Tesis sobre el tema "Insect vectors"

The aim of the research was to identify the phytoplasmas detected in insects that were found on various phytoplasma-infected plants, and to reveal phytoplasma insect-vectors as well as phytogenetical relationships of identified phytoplasmas. From previous research, we already know a few mostly widespread phytoplasma groups, subgroups, and many of their host plants in Lithuania. The data on potential vectors of these bacteria are very scarce in Lithuania. The identification and research of insect vectors will help to create more effective strategies and systems to fight with phytoplasmal infections. Identification of phytoplasmas and their vectors will provide important data for research of ecology, distribution, origin, epidemiology, and ways of spreading of these pathogens. Such information is beneficial for plant protection institutions and plant growers in Lithuania and neighbouring countries. It will help to ascertain possible invasive insect species and phytoplasma strains in Lithuania. During this research for the first time in Lithuania, we determined possible phytoplasma insect vectors using molecular biology methods. Most of the detected phytoplasma subgroups were found in the identified insect species for the first time in Lithuania and worldwide. Our data on new potential insect vector species extend the spectrum of phytoplasma vectors in our region. Phytoplasmas were detected for the first time in five plant species in Lithuania. We identified in this work one... [to full text]
Disertacijos darbo tikslas – aptikti ir identifikuoti Lietuvoje paplitusias fitoplazmas vabzdžiuose, surinktuose nuo įvairių augalų su fitoplazminiais simptomais ir nustatyti fitoplazmų vabzdžius pernešėjus bei atskleisti identifikuotų ir kitų fitoplazmų filogenetinius giminingumus. Lietuvoje jau žinomos keletas labiausiai paplitusių fitoplazmų grupių bei pogrupių, taip pat aptikta nemažai jų augalų-šeimininkų. Duomenų apie galimus šių bakterijų pernešėjus Lietuvoje beveik nėra. Pernešėjų identifikavimas ir tyrimas padės kurti veiksmingesnes strategijas bei sistemas kovai su fitoplazminėmis infekcijomis. Fitoplazmų ir jų pernešėjų identifikavimas suteiks svarbių duomenų tiriant šių patogenų ekologiją, paplitimą, kilmę, epidemiologiją, plitimo kelius. Informacija bus naudinga Lietuvos ir kaimyninių šalių augalų apsaugai. Taip pat galės padėti nustatant galimų invazinių vabzdžių rūšių bei fitoplazmų kamienų atsiradimą Lietuvoje dėl klimato kaitos. Šio darbo metu pirmą kartą Lietuvoje molekuliniais metodais buvo išaiškinti fitoplazmų vabzdžiai pernešėjai. Daugelis aptiktų fitoplazmų pogrupių nustatytos identifikotuose vabzdžiuose pirmą kartą, kaip Lietuvoje taip ir pasaulyje. Penkiose augalų rūšyse fitoplazmos aptiktos pirmą kartą Lietuvoje. Darbo metu nustatytas vienas visiškai naujas Lietuvai ir pasauliui ir vienas naujas Lietuvai fitoplazmų pogrupiai bei jų augalai šeimininkai, kas prisideda prie Lietuvoje bei pasaulyje aptinkamų fitoplazmų paplitimo ir bioįvairovės tyrimo... [toliau žr. visą tekstą]

Les vecteurs dérivés du virus adéno-associé (AAVr) constituent des outils de choix pour le transfert de gène in vivo. Leur innocuité a notamment contribué à leur attractivité et leur utilisation dans des essais cliniques de thérapie génique. Afin d'étendre le champ de leur application au traitement de maladies systémiques, un défi majeur reste à relever : leur production à grande échelle. Le système d'infection de cellules d'insecte par des baculovirus peut répondre à ce challenge, pourtant la biologie de l'AAV dans ces cellules reste méconnue. Ceci se répercute par la présence de particules vides ou par une perte d'infectiosité des vecteurs viraux produits. Le travail présenté dans ce manuscrit a pour objectifs de 1) déterminer l'efficacité et la spécificité d'encapsidation du gène d'intérêt dans les capsides d'AAVr 2) étudier le lien entre ces paramètres et l'expression des protéines Rep et 3) définir le rôle de la protéine AAP (assembly-activating protein) en cellules d'insecte. De façon inédite, nous avons montré que moins de 30% des particules générées contenaient le transgène et que l'ADN baculoviral représentait jusqu'à 2,1% du contenu des capsides d'AAV, avec une prédominance pour les séquences proches des ITR (inverted terminal repeats). Enfin, nous avons démontré que l'AAP était essentielle pour l'assemblage des particules d'AAV2 dans les cellules Sf9. Ce projet participe non seulement à l'élucidation des mécanismes· d'encapsidation des AAV dans les cellules d'insecte mais répond également aux exigences des organismes réglementaires en proposant une technique d' avant-garde d'évaluation des contaminants ADN présents dans les stocks de vecteurs AAV
Due to their efficiency and safety, recombinant adenoassociated virus (rAAV) vectors have been widely used for gene therapy. ln the past few years, there have been a large number of positive clinical outputs using AAVbased products spanning broad therapeutic areas. However, the generation of rAAV at sufficient quantity and quality appears as a bottleneck on the path to commercialization. The baculovirus-infected insect cell platform has proven to tackle this challenge, yet, surprisingly, the biology of rAAV in insect cells remains largely unknown. As a result, current vectors suffer from quality problems such as generation of empty particles or reduced infectivity. The objectives of the present work are 1) to determine the rAAV packaging efficiency and specificity in insect cells 2) to investigate the link between packaging and Rep proteins expression, and 3) to decipher the role of the assembly-activating protein (AAP). First, we showed that less than 30% of rAAV particles contained the gene of interest in S19 cells cleared lysate. Second, we found that baculoviral DNA contamination is below 2.1% of encapsidated DNA, with a higher representativity for sequences close to the inverted terminal repeats. Finally, we demonstrated that functional AAP is strictly required for rAAV2 particles assembly in insect cells. Altogether, our data provide novel insights into the biological mechanism of rAAV genome packaging in insect cells and suggest that there is still room for improvement in order to increase vector quality. From a safety perspective, this project has allowed the development of an accurate quality control method to assess DNA contamination in viral vector stocks

Bois Noir (BN) is a disease of the grapevine yellows (GY) complex associated with ‘Candidatus Phytoplasma solani’ (CaPsol) strains, which causes economic crop losses in viticulture worldwide. The epidemiology of BN is very complex due to the involvement of different herbaceous plants and several insect vectors that transmit CaPsol to grapevine. Therefore, the BN containment is very difficult and require massive efforts for possible spread reduction. The heavy application of chemical insecticides was not successful to control the insect vector presence within the vineyard. The thesis work was framed considering the directives provided by the European council 2009/128/EC regarding the promotion of low use of pesticides in sustainable management approaches. In the present thesis dissertation, CaPsol insect vectors and diseased grapevines were the main targets prioritized for successful containment of BN in organically cultivated vineyards in northern Italy. Since H. obsoletus is the widely distributed insect vector in Europe, the management of the leafhopper population was carefully considered. The use of Vitex agnus-castus as trap plant for H. obsoletus as an indirect control strategy was evaluated. Vitex agnus-castus tended to be a preferred host plant for H. obsoletus, but transmission trials demonstrated its ability to harbor CaPsol and indicated the impossibility of using this plant to avoid BN spread. In addition, the efficacy of different entomopathogenic nematodes and fungi as direct control strategy were evaluated against H. obsoletus nymphs and adults. Their application in a laboratory and semi-field conditions showed a promising killing effect that can be implemented for insect vector control in open field. Due to the very low density of H. obsoletus population in heavily BN infected vineyards questions were raised to figure out the other possible presence of alternative insect vectors. Surveys on Auchenorrhyncha coupled with molecular analyses revealed the presence of numerous putative vectors. Some of them, selected on the basis of their abundance, CaPsol-infection rate and CaPsol strains harbored, went through transmission trials. Eight insects were found able to transmit CaPsol to grapevines. Characterization of the bacterial microbiota associated with H. obsoletus and the alternative insect vectors indicated an interesting perspective regarding the microbial signatures associated with xylem- and phloem-feeding insects, and determinants that could be relevant to establish whether an insect species can be a vector or not, opening up new avenues for developing microbial resource management-based approaches. Moreover, grafting of materials collected from recovered grapevines was conducted in field trials with the aim to evaluate its preventive and curative potentials against BN. Results of symptom observation and CaPsol molecular detection on grafted and non-grafted grapevines showed that grafting of recovered shoots can have a curative effect, increasing the natural recovery. Results obtained in this PhD thesis opened new perspectives to develop integrated sustainable strategies for BN management.

Understanding emerging viruses is critical for disease monitoring and prediction; however, surveys of novel viruses are hindered by the lack of a universal assay for viruses. Viral metagenomics, consisting of viral particle purification and shotgun sequencing, is a powerful technique for discovering viruses in a wide variety of sample types. However, current protocols are not effective on tissue samples (e.g., lungs, livers and tumors), where they are hindered by the high amount of host nucleic acids which limits the percentage of sequences that originate from viruses. In this dissertation, a modified viral metagenomics protocol was developed and utilized to effectively purify viruses from tissues, enabling the sequencing of novel viruses from animals, plants, and insect vectors. Viral metagenomics performed directly on tissue samples enabled the discovery of novel vertebrate, plant, insect and bacterial viruses. From a sea turtle fibropapilloma, viral metagenomics revealed a novel tornovirus STTV1, which is only the second single-stranded DNA virus known in reptiles and is extremely different from any previously described viruses. Similarly, from the lung of a sea lion involved in a mortality event, viral metagenomics identified a novel sea lion anellovirus (ZcAV), which is the first anellovirus characterized from a marine animal. The STTV1 and ZcAV genomes were highly divergent from known viruses, to a degree that they could not have been detected by degenerate PCR assays or microarrays, demonstrating viral metagenomics as an effective method for characterizing novel viruses. In addition to discovery of viruses in individual diseased animals, this dissertation pioneered a technique called vector-enabled metagenomics (VEM) to examine viruses present in insect vectors. VEM combines the power of metagenomics to sequence novel viruses with the ability of insect vectors to integrate viral diversity over space, time, and many host individuals and species. VEM allows for the investigation of viral diversity among the broad range of hosts that the insects feed on, providing an unprecedented snapshot of the viral diversity in natural reservoirs. This dissertation describes the first viral metagenome performed on mosquitoes and whiteflies, providing significant insights to the viral diversity in animal and plant reservoirs. Both animal and plant viruses were represented in the mosquito viromes, which likely originate from animal blood and plant nectar that the mosquitoes feed on. Mosquito viromes contained a diverse range of viruses, including vertebrate, insect, plant, and bacterial viruses, and almost all the viral sequences were novel, suggesting the pan-animal virome is largely uncharacterized. In contrast, only plant viruses were observed in the whitefly viromes because whiteflies feed solely on plants. Whitefly viromes contained known and novel viral sequences infecting crops, novel viral sequences infecting native plants, as well as novel satellites that were the first viral satellites to be documented in North America. Distinct viromes were found amongst the three mosquito samples as well as between the two whitefly samples, demonstrating the diverse and dynamic nature of the viruses in plant and animal reservoirs. By enabling the discovery of virus in diseased organisms and in insect vectors, viral metagenomics is a powerful technique that will significantly enhance our fundamental scientific understanding of the diversity, transmission, biogeography, and emergence of viruses. The viral metagenomic approach described here has implications for surveillance of emerging viruses, prediction of viral epidemics, and proactive control of diseases.

Measles virus is an animal enveloped virus that is a member of the genus morbillivirus in the paramyxoviridae family. Its envelope contains two surface glycoproteins H and F which are required for viral attachment and entry respectively. Virus penetration occurs via a process which involves fusion of the viral membrane with the plasma membrane at the cell surface. Replication of the virus thus follows and leads to giant cell (syncytia) formation.
The infectivity of measles virus is dependent upon a host proteolytic cleavage of the F$ sb0$ glycoprotein into two active subunits F$ sb1$ and F$ sb2$. This cleavage was later shown to expose a hydrophobic sequence at a NH$ sb2$ terminal of the F$ sb1$ which is directly involved in cell fusion and virus penetration.
In order to increase our knowledge concerning cell mediated fusion events we have expressed the fusion glycoprotein of measles virus in insect and human cells by using recombinant baculo- and adenoviruses respectively. Analysis by SDS-PAGE demonstrated that our protein was first synthesized as a 60 Kd protein and cleaved subsequently into its two respective subunits F$ sb1$ and F$ sb2$ of 40 Kd and 20 Kd respectively. Hemolysis assays confirmed the biological activity of this protein in both systems. However, the fusion protein was unable to fuse insect cells.

The objective of this project was to determine the extent and impact of infection with Trypanosoma evansi in livestock in Mindanao, Philippines, evaluate economic benefits of control options and determine its vectors. The project was undertaken because of insufficient knowledge on the dynamics and impact of surra in livestock in the island and because sporadic serious epidemics have occurred in recent years despite the implementation of control measures. Data from cross-sectional surveys conducted in 2002-6 involving more than 2,000 animals were utilized to estimate the impact of T. evansi infection in buffalo populations. A bio-economic infectious disease model was also developed using these data and data from follow up surveys to evaluate economic losses and benefits of control of T. evansi in different animal hosts. Trypanosoma evansi infection caused significant negative impact on buffalo populations with high mortality and reproductive losses. The estimated financial losses from T. evansi infection are high. However, targeted treatment of all sick animals throughout the year using a highly effective drug would have substantial benefits. The estimated annual total financial net benefit from an effective surra control for a typical village in a moderate/high-surra risk area in Mindanao was US $158,000. The value added to buffaloes, cattle, horses, goats/sheep and pigs as a result of this control was US $88, $84, $151, $7, $114 per animal per year, respectively. Follow up surveys were conducted in 2007-8 to determine the prevalence of T. evansi infection in 2,383 buffaloes and other animals (290 goats, 226 cattle, 151 pigs and 35 horses) from 73 villages in Mindanao, investigate associations between T. evansi and other pathogens (Neospora caninum and Brucella abortus) with reproductive failure and calf mortality in buffalo cows, and to confirm the presence of RoTat 1.2 gene in 168 local isolates of T. evansi. Trypanosoma evansi was detected using MHCT, MIT, PCR and CATT in livestock in a number of high-surra risk areas with 59%, 41%, 41%, 35% and 25% seroprevalence in buffaloes, cattle, horses, goats and pigs, respectively. Trypanosoma evansi was associated with reproductive failure and early calf mortality in buffalo cows. The RoTat 1.2 gene was detected in all 168 local isolates of T. evansi tested but was probably not expressed in all cases. The seroprevalence and impact of combined infections of T. evansi and F. gigantica were determined in 1,163 buffaloes from 32 villages in high- and low-surra risk areas in Mindanao. Fasciola gigantica infection was highly prevalent in buffaloes in both areas and combined infections of T. evansi and F. gigantica were highly prevalent in high-surra risk villages. Buffaloes that were seropositive to T. evansi infection were more likely to be seropositive with F. gigantica than uninfected buffaloes and combined infections were associated with poor body conditions and low PCV. Trapping of tabanids was conducted in 2007-8 in selected villages in high-and low-surra risk provinces to determine the local tabanid fauna and their abundance, detect trypanosomes in tabanids and determine the hosts of the flies using genetic markers. All five species of trapped tabanids were more abundant in low- than high-altitude areas and abundance was significantly associated with high rainfall. Trypanosoma evansi and T. theileri were detected from at least one fly of every tabanid species caught. Buffaloes, pigs, goats, humans and chickens were identified as hosts of tabanids in Mindanao. There is a need to identify tabanid fauna in other areas in Mindanao and confirm their active role in the transmission of T. evansi in livestock. Results support the conclusions that: (a) Trypanosoma evansi infection causes significant economic losses in livestock in Mindanao but its effective control would provide substantial financial benefits; (b) Trypanosoma evansi infection is highly prevalent in livestock in Mindanao which is highly associated with poor reproduction performance in buffaloes; (c) RoTat 1.2 based tests (PCR and CATT) are applicable in the diagnosis of surra in Mindanao but the value of the CATT still requires further evaluation; (d) Fasciolosis needs to be included in the control strategy for surra in high risk areas; and, (e) Tabanids identified in Mindanao are potential transmitters of T. evansi and their control should be explored. There is a need, therefore, to sustain surveillance and implement an integrated and more effective control programme against T. evansi infection in livestock in Mindanao.

Insects can cause economic and environmental damages by transmitting bacterial pathogens to plants, making the use of control and prevention measures imperative. On the other hand, methods aimed at preventing crop pests’ outbreaks, such as tilling and insecticide applications, pose a threat to human and ecosystems health. Innovative and more sustainable control methods are therefore urgently needed to address farmers’ needs and preserve ecosystem services provided by biodiversity. Biotremology is an emerging science that studies the substrate-borne vibrational signals emitted by insects to communicate and interact with their others in the environment. As many insect pests rely on these signals to communicate, their behaviour can be manipulated by transmitting vibrations to the substrate where they thrive. Vibrational signals can, for instance, interfere with target’s behaviours such as mating, feeding, and residence time on the plant, and can therefore support integrated pest management strategies alternative to insecticides. In this work, I investigated the mating behaviour and vibrational communication of two insect vectors having an important economic impact: Philaenus spumarius (Hemiptera: Aphrophoridae) and Bactericera cockerelli (Hemiptera: Triozidae). I also provided novel insights regarding the vibrational manipulation of their behaviour. I characterised the sexual behaviour and associated vibrational signals of the meadow spittlebug P. spumarius, and I described other intraspecific interactions. I evaluated the correlation between the female signalling activity and development of the reproductive organs, suggesting that a vibrational mating disruption could be effective only if applied from the onset of egg maturation, which triggers the female proceptivity (from August onwards). Besides gathering behavioural data, I used occupancy models to estimate P. spumarius presence and detection probability in Trentino (Northern Italy), a rarely deployed approach to study insect distribution. Site covariates impacted P. spumarius detectability, suggesting that spittlebug presence can be overlooked if detection is not considered. As plant composition influences the presence and abundance of the spittlebug, I studied the association between P. spumarius and its host plants. Such information can support monitoring programs and control strategies aimed at reducing P. spumarius population in crops of interest by shaping the plant composition of the ground cover. After investigating the behaviours and the ecology of the spittlebug, I attempted to disrupt its feeding activity using vibrations, opening the gates for future research. In this regard, I combined for the first time biotremology with the Electrical Penetration Graph (EPG) technique. The tomato potato psyllid B. cockerelli is an important threat to New Zealand crops. Besides the characterisation of the pair formation process and associated vibrational signals, I used vibrations to disrupt mate finding by transmitting vibrations to the host plant. Given that the stimulus attracted male psyllids at small scale, this strategy is a feasible approach that could be implemented for use in fields and greenhouses. In this way, an innovative trapping and mating disruption device could join the farmers’ toolbox for monitoring and controlling B. cockerelli. In conclusion, I provided new information regarding the ethology of two insect vectors and suggested that there is room for applied biotremology to support sustainable management strategies.

Insects can cause economic and environmental damages by transmitting bacterial pathogens to plants, making the use of control and prevention measures imperative. On the other hand, methods aimed at preventing crop pests’ outbreaks, such as tilling and insecticide applications, pose a threat to human and ecosystems health. Innovative and more sustainable control methods are therefore urgently needed to address farmers’ needs and preserve ecosystem services provided by biodiversity. Biotremology is an emerging science that studies the substrate-borne vibrational signals emitted by insects to communicate and interact with their others in the environment. As many insect pests rely on these signals to communicate, their behaviour can be manipulated by transmitting vibrations to the substrate where they thrive. Vibrational signals can, for instance, interfere with target’s behaviours such as mating, feeding, and residence time on the plant, and can therefore support integrated pest management strategies alternative to insecticides. In this work, I investigated the mating behaviour and vibrational communication of two insect vectors having an important economic impact: Philaenus spumarius (Hemiptera: Aphrophoridae) and Bactericera cockerelli (Hemiptera: Triozidae). I also provided novel insights regarding the vibrational manipulation of their behaviour. I characterised the sexual behaviour and associated vibrational signals of the meadow spittlebug P. spumarius, and I described other intraspecific interactions. I evaluated the correlation between the female signalling activity and development of the reproductive organs, suggesting that a vibrational mating disruption could be effective only if applied from the onset of egg maturation, which triggers the female proceptivity (from August onwards). Besides gathering behavioural data, I used occupancy models to estimate P. spumarius presence and detection probability in Trentino (Northern Italy), a rarely deployed approach to study insect distribution. Site covariates impacted P. spumarius detectability, suggesting that spittlebug presence can be overlooked if detection is not considered. As plant composition influences the presence and abundance of the spittlebug, I studied the association between P. spumarius and its host plants. Such information can support monitoring programs and control strategies aimed at reducing P. spumarius population in crops of interest by shaping the plant composition of the ground cover. After investigating the behaviours and the ecology of the spittlebug, I attempted to disrupt its feeding activity using vibrations, opening the gates for future research. In this regard, I combined for the first time biotremology with the Electrical Penetration Graph (EPG) technique. The tomato potato psyllid B. cockerelli is an important threat to New Zealand crops. Besides the characterisation of the pair formation process and associated vibrational signals, I used vibrations to disrupt mate finding by transmitting vibrations to the host plant. Given that the stimulus attracted male psyllids at small scale, this strategy is a feasible approach that could be implemented for use in fields and greenhouses. In this way, an innovative trapping and mating disruption device could join the farmers’ toolbox for monitoring and controlling B. cockerelli. In conclusion, I provided new information regarding the ethology of two insect vectors and suggested that there is room for applied biotremology to support sustainable management strategies.

I. Ricci, C. Damiani, P. Rossi, A. Capone, P. Scuppa, A. Cappelli, U. Ulissi, M. Mosca, M. Valzano, S. Epis, E. Crotti, D. Daffonchio, A. Alma, L. Sacchi, M. Mandrioli, C. Bandi and G. Favia, Mosquito symbioses: from basic research to the paratransgenic control of mosquito-borne diseases, J. Appl. Entomol. 135 (2011) 487493 Guido Favia, Irene Ricci, Patrizia Scuppa, Claudia Damiani, Paolo Rossi, Aida Capone, Chenoa De Freece, Matteo Valzano, Alessia Cappelli, Michela Mosca and Ulisse Ulissi, Facing Malaria Parasite with Mosquito Symbionts Irene Ricci, Matteo Valzano, Ulisse Ulissi, Sara Epis, Alessia Cappelli, Guido Favia, Symbiotic control of mosquito borne disease M. Valzano, G. Achille, F. Burzacca, I. Ricci, C. Damiani, P. Scuppa, G. Favia, Deciphering microbiota associated to Rhynchophorus ferrugineus in Italian samples: a preliminary study, Journal of Entomological and Acarological Research 2012; volume 44:e16

Orientador: Cláudio Casanova
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
Made available in DSpace on 2018-08-26T11:03:53Z (GMT). No. of bitstreams: 1 CollaJacques_FernandaElisa_D.pdf: 17373999 bytes, checksum: 18adb4b65dc2639da145a80ed8b6f4e2 (MD5) Previous issue date: 2014
Resumo: Flebotomíneos (Diptera: Psychodidae) são insetos de importância médico-veterinária pois participam do ciclo de doenças tais como leishmanioses, que acometem milhares de pessoas ao redor do mundo. O conhecimento sobre a ecologia e biologia desses dípteros é de fundamental importância para determinar a capacidade e competência vetorial de suas populações, e portanto, relevantes do ponto de vista da saúde pública. Este trabalho buscou esclarecer alguns pontos com informações deficientes sobre a biologia e ecologia das espécies vetoras de Leishmania (Leishmania) infantum chagasi e Leishmania (Viannia) braziliensis, respectivamente, Lutzomyia longipalpis e Nyssomyia neivai, no estado de São Paulo. Para a leishmaniose visceral americana (LVA), objetivou-se esclarecer questões a respeito de criadouros urbanos preferenciais de Lu. longipalpis e sobre sua expansão geográfica no estado de São Paulo. Em referências às lacunas no conhecimento da ecoepidemiologia da leishmaniose tegumentar americana, a competência vetorial de Nyssomyia neivai para L. (V.) braziliensis e o estudo da composição da comunidade flebotomínica em área endêmica no nordeste do estado, bem como sua variação mensal. Criadouros de Lu. longipalpis foram encontrados predominantemente em ambientes sombreados com grande quantidade de matéria orgânica, com maior concentração de larvas associadas a galinheiros. Quanto à sua dispersão, esta parece ter seguido uma rota a partir de Mato Grosso do Sul, seguindo em direção oeste-leste e também para norte e para sul a partir dessa linha central. A ocorrência de casos de LVA em humanos frequentemente é precedida pela detecção de Lu. longipalpis e casos caninos. Nyssomyia neivai, bastante frequente em áreas endêmicas para LTA no vale do Rio Mogi Guaçu, mostrou-se competente para adquirir, manter e transmitir o parasito em condições laboratoriais, cumprindo assim mais uma etapa necessária para a sua incriminação como espécie-vetora. Ainda no vale do rio Mogi Guaçu, a comunidade de flebotomíneos mostrou-se bastante diversa, com 17 espécies, das quais cinco são importantes para o ciclo da LTA. A fauna flebotomínica encontrada era composta majoritariamente por Nyssomyia whitmani, Migonemyia migonei e Expapillata firmatoi e, mais uma vez, abrigos para animais, tais como galinheiros, foi o local preferido para estes insetos, onde a sua maioria foi capturada. Desta forma, este trabalho apresenta dados relevantes para o conhecimento a respeito desses insetos vetores, que podem ser usados como base para futuros projetos ou no desenvolvimento de novas estratégias de controle dessas doenças
Abstract: Phlebotomine sandflies (Diptera: Psychodidae) are insects of medical and veterinary importance because they take part in the cicle of diseases, such as leishmaniasis which affect thousands of people around the world. Learning about these dipteras¿s ecology and biology is fundamentaly important to define the vectorial capacity and competence of their populations, and therefore, relevant for the public health perspective. This study aimed at adressing some issues with deficiente information on the biology and ecology of vector species of Leishmania (Leishmania) infantum chagasi and Leishmania (Viannia) braziliensis, respectively, Lutzomyia longipalpis and Nyssomyia neivai, in São Paulo State. For the American visceral leishmaniasis (AVL), this paper tried to answer questions on the preferencial breeding site of Lu. longipalpis and its geographical expantion in São Paulo State. In relation to the knowledge of the ecoepidemiology of American cutaneous leishmaniasis (ACL), the vectorial competence of Nyssomyia neivai to L. (V.) braziliensis and the analysis of the sandfly community in and endemic area in the Northeast of the state were studied. Lutzomyia longipalpis breeding sites were found predominantly in shaded environments with great amount of organic matter, and the greatest concentration of larvae associated to chicken sheds. Considering its dispersion, Lu. longipalpis seemed to follow a route from Mato Grosso do Sul to São Paulo city, in a west to east direction and from that central line, an expansion to the north and south. The occurance of human AVL is generally preceded by the detection of Lu. longipalpis and canine cases. Nyssomyia neivai, frequently found in endemic áreas for ACL in the Mogi Gauçu River Valley, was shown to be competente to aquire, host and transmit the parasite under laboratory conditions, thus fulfilling another requirement to be recognized as a vector species. Still in Mogi Guaçu River Valley, the sandfly community was greatly diverse, comprised of 17 species, of which five are considered important for the ACL cicle. The sandfly fauna was mainly comprised of Nyssomyia whitmani, Migonemyia migonei and Expapillata firmatoi and, once again, animal shelters, such as chicken sheds, were the preferred site for these insects, where the greatest amount was collected. Therefore, this work presents relevant data for the knowledge on these vector insects, which can be used as basis for future projects or in the development of new control strategies against these diseases
Doutorado
Relações Antrópicas, Meio Ambiente e Parasitologia
Doutora em Biologia Animal

Submitted by Jean Medeiros (jeanletras@uepb.edu.br) on 2016-05-11T14:27:17Z No. of bitstreams: 1 PDF - Noiana de Paula Noia.pdf: 1156392 bytes, checksum: a6a9764178b9ec1cd59f8dadd9cb363e (MD5)
Approved for entry into archive by Secta BC (secta.csu.bc@uepb.edu.br) on 2016-07-22T13:24:22Z (GMT) No. of bitstreams: 1 PDF - Noiana de Paula Noia.pdf: 1156392 bytes, checksum: a6a9764178b9ec1cd59f8dadd9cb363e (MD5)
Made available in DSpace on 2016-07-22T13:24:22Z (GMT). No. of bitstreams: 1 PDF - Noiana de Paula Noia.pdf: 1156392 bytes, checksum: a6a9764178b9ec1cd59f8dadd9cb363e (MD5) Previous issue date: 2013-12-16
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
The Aedes genus is of great epidemiological importance, highlighting the Aedes aegypti Linnaeus, 1762 and Aedes albopictus Skuse, 1894, the main vectors of dengue, yellow fever and other arboviruses. How are spread across the planet, have become a public health risk factors. Aimed to evaluate the effects of temperature on competition and in the life cycle, and the source of blood meal in fecundity and longevity of females of A. aegypti and A. albopictus. The collections of eggs were performed using traps ovitraps type. The experiments were performed at the Laboratory of Entomology, University of Paraíba State - UEPB in Campina Grande, PB. Competition between A. aegypti and A. albopictus was examined at temperatures of 20, 24, 28, 32 and 36 ° C in the proportion of 10:0, 7:3, 5:5, 3:7 and 10:0 (A. aegypti : A. albopictus). The cycle life was examined at the same temperatures. Feeding two treatments with each meal in quail and the other in swiss mice were used. For all experiments we used 12h light and 70% humidity. In interspecific relationship, there was overlap of A. aegypti on A. albopictus from the first generation (F1), independent of temperature and the proportion of species. The long life cycle did not differ significantly among species independent of temperature, but decreased with increasing temperature. The base temperature (Tb) and thermal constant (K) and hence the number of generations was greater for A. albopictus than for A. aegypti. The repast in mice resulted in greater number of eggs and female longevity for both A. aegypti as for A. albopictus. The substrate used for biting that was more appropriate for the two species, the mouse was, since we obtained the highest female fertility and longevity. The temperature and the substrate used for blood feeding interfere with competition, development, longevity and fecundity of the species studied. The number of generations of insects in the laboratory is higher for A. albopictus than for A. aegypti.
O grande importância epidemiológica, destacando o Aedes aegypti Linnaeus, 1762 e Aedes albopictus Skuse, 1894, principais vetores da dengue, febre amarela e outras arboviroses. Como estão distribuídos por todo o planeta, se tornaram fatores de risco à saúde pública. Objetivou-se avaliar os efeitos da temperatura na competição e no ciclo de vida, e da fonte de repasto sanguíneo na fecundidade e longevidade de fêmeas de A. aegypti e A. albopictus. As coletas dos ovos foram realizadas utilizando armadilhas do tipo ovitraps. Os experimentos foram realizados no Laboratório de Entomologia da Universidade Estadual da Paraíba – UEPB, em Campina Grande, PB. A competição entre A. aegypti e A. albopictus foi analisada nas temperaturas de 20, 24, 28, 32 e 36ºC nas proporções de 10:0, 7:3, 5:5, 3:7 e 10:0 (A. aegypti: A. albopictus). O ciclo de vida foi analisado nas mesmas temperaturas. Para nutrição foram utilizados dois tratamentos um com o repasto em codorna e o outro em camundongo swiss. Para todos os experimento utilizou-se fotofase de 12h e umidade de 70%. Na relação interespecífica, constatou-se a sobreposição de A. aegypti sobre A. albopictus a partir da primeira geração (F1), independente da temperatura e da proporção das espécies. O tempo do ciclo de vida não diferiu significativamente entre as espécies, independente da temperatura, porem diminuiu em função do aumento da temperatura. A temperatura base (Tb) e a constante térmica (K) e consequentemente o número de gerações anuais foi maior para A. albopictus do que para A. aegypti. O repasto em camundongos resultou em maior número de ovos e longevidade das fêmeas tanto para A. aegypti como para A. albopictus. O substrato utilizado para hematofagia que se mostrou mais adequado para as duas espécies, foi o camundongo, visto que obteve-se a maior fertilidade e longevidade da fêmea. A temperatura e o substrato utilizado para o repasto sanguíneo interferem na competição, desenvolvimento, longevidade e fecundidade das espécies estudadas. O número de gerações anuais dos insetos em laboratório é maior para A. albopictus do que para A. aegypti.

I fitoplasmi sono procarioti pleomorfi, privi di parete cellulare, simili a batteri ed appartenenti alla classe Mollicutes e sono caratterizzati da un genoma molto ridotto. L’assenza di vie metaboliche essenziali li rende parassiti obbligati di piante ed insetti. Nelle prime, la presenza di questi patogeni è limitata ai tessuti floematici e causa sintomi associati a profondi disordini nel normale bilancio ormonale. Negli insetti, gli effetti dei fitoplasmi sulla fitness sono vari e, a seconda delle relazioni evolutive, possono essere deleteri o benefici. I fitoplasmi sono associati a centinaia di patologie di specie vegetali distribuite in tutto il mondo, comprese colture di elevate importanza economica, fruttiferi e anche piante ornamentali. Il principale mezzo di trasmissione di questi patogeni è costituito da insetti vettori e le interazioni tra fitoplasmi ed insetti sono, in alcuni casi, molto specifiche e coinvolgono una complessa catena di eventi. Solo fitomizi floematici, come cicadomorfi, fulgoromorfi e psille hanno il potenziale per acquisire e trasmettere questi parassiti obbligati. ‘Candidatus Phytoplasma mali’ è l’agente eziologico di Apple proliferation (AP), una malattina che rappresenta uno dei più gravi problemi nei meleti europei. In Trentino, una delle principali regioni produttrici di mele in Italia, AP è una minaccia per la produzione. I sintomi più evidenti sono la presenza di scopazzi sui rami, di stipole ingrandite nelle foglie e di arrossamenti anticipati delle chiome in autunno. I frutti degli alberi infetti, che hanno dimensioni inferiori e proprietà organolettiche alterate, non possono essere commercializzati. Finora due psille, Cacospylla picta e Cacopsylla melanoneura (Homoptera: Psyllidae), sono state confermate vettori della malattia, ma il loro reale ruolo nell’epidemiologia di AP è ancora oggetto di discussione, dal momento che studi condotti in aree geografiche diverse mostrano efficienze di trasmissione differenti. Questa ricerca si inserisce all’interno di un progetto avviato in Trentino dopo che un’esplosione della fitoplasmosi è stata segnalata in Valsugana (Trentino sud-orientale) nel 2011. I principali obiettivi di questa ricerca hanno riguardato lo studio di alcuni aspetti dell’epidemiologia di AP alcune caratteristiche biologiche degli insetti vettori. Il primo scopo è stato quello di monitorare l’avanzamento della malattia e le dinamiche di popolazione dei vettori, valutando le percentuali di infezione naturale delle psille e alcuni parametri relativi al processo di trasmissione, come la capacità di acquisizione, l’efficienza di trasmissione e la possibilità di trasmissione transovarica del fitoplasma. La distribuzione delle piante infette è stata mappata lungo la Valsugana e le popolazioni delle psille vettrici sono state monitorate tra il 2014 e il 2016. Dopo tre anni di rilievi, si è ottenuta una drastica diminuzione della percentuale di piante sintomatiche. Riguardo gli insetti vettori, C. melanoneura si è rivelata essere la specie con densità di popolazione più elevate sia in frutteti coltivati, sia in frutteti abbandonati, ma la specie con percentuali di infettività maggiori è risultata essere C. picta. I parametri di trasmissione per entrambe le specie di psilla sono stati valutati attraverso prove di acquisizione e trasmissione e prove specifiche sono state condotte per verificare se vi è trasmissione transovarica del fitoplasma in C. picta. Gli esperimenti condotti in condizioni di semi-campo e serra, durante la primavera e l’estate del 2015 e del 2016, hanno riguardato adulti svernanti di entrambe le specie (campionati in Valsugana) e ninfe e adulti di nuova generazione allevati su piante di melo infette. Alla fine di ogni esperimento, piante e insetti sono stati analizzati tramite real-time PCR per verificare la presenza di fitoplasma. I risultati confermano che C. picta è un vettore molto più efficiente in Trentino, ma suggeriscono la possibilità di acquisizione e trasmissione del fitoplamsa anche in C. melanoneura, anche se con una bassa efficienza. Per entrambe le specie, gli adulti svernanti sono stati in grado di trasmettere il fitoplasma solo dopo aver trascorso un periodo di acquisizione su piante infette e la capacità di acquisizione degli stadi che si sono sviluppati su piante infette è stata molto elevata. In più, è stata dimostrato per la prima volta che individui infetti di C. picta sono capaci di trasmettere ‘Ca. P. mali’ alla progenie. Uno degli obiettivi di questa ricerca era lo studio del rapporto tritrofico tra fitoplasma e i suoi ospiti, attraverso indagini sulla diversità genetica del fitoplasma in piante e insetti vettori e sulla loro distribuzione geografica. Quindi il fitoplasma è stato caratterizzato geneticamente e gli isolati ottenuti da piante di melo e insetti raccolti in differenti aree geografiche sono stati analizzati attraverso un metodo di multilocus sequencing typing basato su quattro geni del fitoplasma. I risultati ottenuti sono solo parziali, ma i differenti genotipi di ‘Ca. P. mali’ ottenuti finora indicano la presenza di un’elevata variabilità genetica negli insetti rispetto alle piante, che suggerisce l’ipotesi dell’esistenza di relazioni specifiche tra genotipi di fitoplasma e insetti vettori. Nuove informazioni sono state ottenute sull’etologia delle psille, in particolare per quanto riguarda il comportamento riproduttivo. Per questo, sono state condotte osservazioni etologiche e registrazioni dei segnali vibrazionali emessi durante il corteggiamento attraverso l’uso di un laser vibrometro. Il segnale sembra essere specie-specifico, ma non sembra essere un prerequisito per il corteggiamento e l’accoppiamento. Inoltre, come già visto in altri psillidi, indagini al microscopio elettronico a scansione hanno evidenziato la presenza di strutture stridulatorie su torace e ali di entrambe le specie. Infine, poiché è noto che altre specie di omotteri sono vettori di fitoplasmosi, un obiettivo di questo studio è stato la ricerca di potenziali nuovi vettori di ‘Ca. P. mali’, caratterizzando allo stesso tempo le comunità di cicadomorfi e fulgoromorfi dei meleti della Valsugana e valutando l’effetto dei paesaggi circostanti sulla loro distribuzione. I campionamento sono stati condotti in meleti circondati da diversi paesaggi e anche all’interno di diversi habitat nello stesso meleto. I risultati di questo studio indicano che i paesaggi influenzano la ricchezza in specie e che, riguardo gli habitat, il cotico erboso del frutteto è visitato da numeri più elevati di specie e individui in tutti i paesaggi considerati. Tutti gli individui raccolti sono stati analizzati tramite real-time PCR e i risultati indicano che almeno due specie di cicaline sono state capaci di acquisire ‘Ca. P. mali’. Le ricerche condotte in questa tesi hanno permesso di tracciare un quadro diffusione di AP in Valsugana. I risultati ottenuti nei diversi ambiti di ricerca descritti sopra, dagli studi epidemiologici e di variabilità genetica, passando attraverso il comportamento sessuale dei vettori e la biodiversità nell’agroecosistema, rappresentano le basi teoriche per aiutare consulenti tecnici e agricoltori ad ottimizzare l’attuale gestione della malattia e i ricercatori a sviluppare strategie di controllo innovative e sostenibili.
Phytoplasmas are pleomorphic wall-less prokaryotes related to bacteria belonging to the Mollicutes class and are characterized by a very small genome. The lack of essential biosynthetic pathways makes them obligate parasites of plants and insect vectors. In plants, these pathogens are restricted to the phloem tissue and cause symptoms suggesting profound disorders in the normal balance of hormones. In insects, their effects on fitness can range from detrimental to beneficial, depending on the evolutionary relationships. Phytoplasmas are associated with diseases of hundreds of plant species worldwide, including many economically important crops, fruit trees, and ornamental plants. Insect vectors represent the principal means of phytoplasma transmission and interactions between phytoplasmas and insect hosts are, in some cases, very specific and involve a complex sequence of events. Only phloem-feeding insects like leafhoppers, planthoppers, and psyllids can potentially acquire and transmit these obligate parasites. ‘Candidatus Phytoplasma mali’ is the etiological agent of Apple proliferation (AP) disease, which represents one of the most severe problems in European apple orchards. In Trentino, one of the main apple producing regions in Italy, AP is the major threat for the production. The main symptoms are witches’ brooms, enlarged stipules, and early leaf reddening in autumn. The fruits of infected trees, which are smaller and have altered organoleptic properties, cannot be commercialized. So far, two psyllid species, Cacospylla picta and Cacopsylla melanoneura (Homoptera: Psyllidae), are confirmed vectors, but their actual role in the epidemiology of AP is still debated, as studies conducted in different geographical regions show different transmission efficiencies. This research is part of a project launched in Trentino after a serious outbreak of the disease reported in Valsugana (southeastern Trentino) in 2011. The main objectives of this work regarded the study of epidemiological traits of the disease and biological features of the insect vectors. The first aim was monitoring the disease spread and vector populations’ dynamics, evaluating the natural infection level of psyllids and some transmission parameters, such as acquisition capacity, transmission efficiency, and the possibility of a transovarial phytoplasma transmission. The distribution of infected plants was mapped along Valsugana and the populations of the psyllid vectors were monitored in the period 2014-2016. After a three-year survey, the percentage of symptomatic apple plants drastically decreased. Regarding the psyllid vectors, C. melanoneura showed higher population levels compared to C. picta in both conventional and untreated orchards, but the percentages of infected individuals were higher in the latter species. The transmission parameters were evaluated in psyllids during acquisition and transmission trials carried out with C. picta and C. melanoneura. Specific trials were conducted with C. picta to assess the vertical transmission of the phytoplasma. Experiments, conducted under semi-field and greenhouse conditions in spring and summer 2015 and 2016, involved overwintered adults of both species collected in Valsugana and nymphs and emigrant adults reared on infected apple plants. After each experiment, insects and test plants were analyzed by real-time PCR to assess the phytoplasma presence. Results confirm C. picta as a more competent ‘Ca. P. mali’ vector in Trentino, but suggest the possibility of acquiring and transmitting the phytoplasma also for C. melanoneura, even though with a low efficiency. For both species, overwintered adults were able to transmit the disease only after an acquisition period spent on infected plants and the acquiring capacity of the stages developed on infected apple plants was very high. Moreover, for the first time was demonstrated that infected C. picta individuals are able to transmit ‘Ca. P. mali’ to the progeny. Another goal of this research was to unravel the tri-trophic relationship involving phytoplasma and its hosts by the study of the genetic diversity of phytoplasma isolates in plants and insect vectors and their geographical distribution. So, a genetic characterization of phytoplasma was conducted, and isolates obtained from insects and apple plants collected in different geographical areas were analyzed by a multilocus sequence typing method based on four phytoplasma genes. The results obtained are only partial, but the different ‘Ca. P. mali’ genotypes observed so far indicate a higher genetic diversity in insects compared to host plants and suggest the hypothesis of specific relationships between phytoplasma genotypes and insect vectors. New insights on psyllid ethology were achieved. In particular, a research was conducted to investigate the vibrational communication involved in reproductive behavior. For the two vector species, ethological observations and laser vibrometer recordings of the vibrational signals emitted during courtship were carried out. Signals appeared to be species-specific, but they did not seem to be a prerequisite for courtship and mating. Moreover, as already seen in other psyllid species, a scanning electron microscopy investigation showed the presence of a stridulatory mechanism on thorax and wings of both species. Finally, as other homopteran species are known to be phytoplasma vectors, a goal of this research was to look for potential new vectors of AP phytoplasma, characterizing the leafhoppers’ and planthoppers’ communities in apple orchards of Valsugana and studying the effect of surrounding landscapes on their distribution. Samplings were conducted in apple orchards surrounded by different landscapes and in different habitats inside the orchard. The results of this study indicate that landscapes influence the species richness and, regarding habitats, that grasses are visited by higher numbers of species and individuals in all landscapes considered. All insect collected were tested by real-time PCR and results indicate that three samples belonging to three different species tested positive to ‘Ca. P. mali’. Research conducted in this thesis drew a picture of AP disease spread in Valsugana. Results obtained in all the topics described above, from epidemiological studies to phytoplasma genetic variability, passing through vectors’ courtship behavior and agroecosystem biodiversity, represent the theoretical background that helps advisors and growers to optimize the current disease management, and researchers to develop innovative and sustainable control strategies.

Made available in DSpace on 2014-06-11T19:34:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-11-23Bitstream added on 2014-06-13T19:05:00Z : No. of bitstreams: 1 macedo_maa_dr_botfca.pdf: 1148372 bytes, checksum: 9b6be36e41f3f7e9a3ddc6047c6ba682 (MD5)
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
O objetivo do presente trabalho foi estudar diferentes estratégias de controle de insetospragas nas culturas do tomateiro e do pimentão, procurando associar amostragens, níveis de controle, resistência de plantas a insetos, controle químico, produtos naturais, irradiação e controle cultural. Foram amostrados o número de B. tabaci, F. schultzei, M. persicae, a porcentagem de plantas com virose, a produção de frutos e a porcentagem do custo dos produtos aplicados em relação a produção. Os dados obtidos foram submetidos à análise de variância pelo teste de Fisher, e quando significativo às médias foram comparadas pelo teste de Tukey, a 5% de probabilidade. Pelos resultados obtidos conclui-se que: a) cultura do tomateiro: os cultivares menos infestados por T. absoluta foram AP 529 e AP 533 na fase de frutificação; a estratégia convencional foi a mais eficiente no controle dos insetos vetores, porcentagem de plantas com virose e da traça, e, promoveu maior produção de frutos; as estratégias MIP, doses de 75,100 e 125 Gy + MIP-nim a 0,5% e MIP-nim a 0,5% e MIP-nim a 0,5% com barreiras de plantas de milho, foram promissores no controle de insetos vetores e traças na fase vegetativa, quando em baixa pressão de populações da traça; o número de pulverizações quando é adotado o MIP é menor que no convencional, b) cultura do pimentão: os cultivares menos infestados por mosca branca, tripes, pulgões e vetores totais foram Priscila e AF 3159, porém não muitos produtivos; os cultivares mais infestados por insetos vetores foram Magali R e Laser, sendo os mais produtivos; a estratégia convencional foi mais eficiente no controle dos insetos vetores, promovendo maior produção de frutos e menor porcentagem de custo dos produtos aplicados; as estratégias óleo...
This work was conducted to evaluate insect-pest control strategies in tomato and green pepper. Sampling, action thresholds, genetic resistance, chemical control, natural insecticides, irradiation and cultural control were studied. The number of B. tabaci, F. schultzei and M. persicae were sampled, and the rate of virus-infected plants, pepper yield and the costs of the products in relation to the production were determined. Data were subjected to an ANOVA through Fisher s test, and means were compared by Tukey test at 5% of probability. Our results showed that: a) In tomato: the cultivars AP 529 and AP 533 were less infested by T. absoluta in the first and second experiments, respectively, during the fruiting stage; the conventional strategy was the most efficient in controlling insect vectors, virus infection and the leafminer, and allowed higher fruit yield. The strategies IPM, radiation of 75, 100 and 125 Gy + IPM - 0.5% Neem, IPM with 0.5% Neem and IPM with 0.5% Neem combined with maize barrier, were promising in controlling insect vectors and leafminers at the vegetative stage, when there is low leafminer population pressure; the number of sprayings in IPM is lower than in conventional control, b) In the pepper experiment, the cultivars Priscila and AF 3159 were the least infested by white flies, thrips, aphids and total vectors, but did not show high pepper yield. The most infested cultivars were Magali R and Laser, which were also the most pruductive. The conventional strategy was the most efficient in controlling insect vectors, allowing the highest yields and the lowest relative control costs. The control with 0.5 and 0.75% Neem oil, radiation of 75, 100 and 125 Gy combined with 0.5% Neem oil and 0.5 and 1.0% Neem oil, were promising to control insect vectors, because they allowed high pepper yield. A linear, negative correlation between the rate... (Complete abstract, click electronic access below)

A espécie Aedes scapularis é um dos culicídeos de grande importância médica. Está distribuída nas Américas e tem grande competência vetora para diversos arbovírus. No estado de São Paulo, há ocorrência de Ae. scapularis em vários municípios, como em Pariquera-Açu e São Paulo. O objetivo desse trabalho foi testar se há diferenciação genética - morfológica entre essas populações, podendo diagnosticar existência de espécies crípticas em Aedes scapularis. As populações estudadas foram Pariquera-Açu (PAR), Parque Ecológico do Tietê em São Paulo (PET) e Butantã (BUT). Os parâmetros utilizados foram: Morfometria geométrica da asa (forma e tamanho), estudo do gene mitocondrial COI, análise dos espaçadores internos transcritos ITS2 e análise morfológica de ovos. Com os resultados obtidos, podemos concluir que a divergência populacional é atestada por padrões geográficos de forma alar, gene mitocondrial COI e razão comprimento e largura dos ovos, mas extensões de estudos em outras áreas precisam ser feitos para poder atestar espécies crípticas em Aedes scapularis.
The species Aedes scapularis is a culicidae of medical importance. It is distributed in the Americas and has a high vector competence for many arboviruses. In state of São Paulo, have occurrence of Ae. scapularis in many cities, such as Pariquera-Acu and the city of São Paulo. The aim of this study was to determine differentiation genetic- morphology among these populations, being able to diagnose the existence of cryptic species in Aedes scapularis. The populations studied were Pariquera-Acu (PAR), the Tietê Ecological Park in Sao Paulo (PET) and Butantã (BUT). The parameters used were: wing geometric morphometry (shape and size), study of mitochondrial gene COI, analysis of internal transcribed spacers ITS2 and morphological analysis of eggs. With these results, we conclude that divergence population is attested by the geographical patterns of wing shape, and COI mitochondrial gene length and width ratio of eggs, but extensions of studies in other areas need to be made in order to attest cryptic species in Aedes scapularis.

Resumo: O objetivo do presente trabalho foi estudar diferentes estratégias de controle de insetospragas nas culturas do tomateiro e do pimentão, procurando associar amostragens, níveis de controle, resistência de plantas a insetos, controle químico, produtos naturais, irradiação e controle cultural. Foram amostrados o número de B. tabaci, F. schultzei, M. persicae, a porcentagem de plantas com virose, a produção de frutos e a porcentagem do custo dos produtos aplicados em relação a produção. Os dados obtidos foram submetidos à análise de variância pelo teste de Fisher, e quando significativo às médias foram comparadas pelo teste de Tukey, a 5% de probabilidade. Pelos resultados obtidos conclui-se que: a) cultura do tomateiro: os cultivares menos infestados por T. absoluta foram AP 529 e AP 533 na fase de frutificação; a estratégia convencional foi a mais eficiente no controle dos insetos vetores, porcentagem de plantas com virose e da traça, e, promoveu maior produção de frutos; as estratégias MIP, doses de 75,100 e 125 Gy + MIP-nim a 0,5% e MIP-nim a 0,5% e MIP-nim a 0,5% com barreiras de plantas de milho, foram promissores no controle de insetos vetores e traças na fase vegetativa, quando em baixa pressão de populações da traça; o número de pulverizações quando é adotado o MIP é menor que no convencional, b) cultura do pimentão: os cultivares menos infestados por mosca branca, tripes, pulgões e vetores totais foram Priscila e AF 3159, porém não muitos produtivos; os cultivares mais infestados por insetos vetores foram Magali R e Laser, sendo os mais produtivos; a estratégia convencional foi mais eficiente no controle dos insetos vetores, promovendo maior produção de frutos e menor porcentagem de custo dos produtos aplicados; as estratégias óleo... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: This work was conducted to evaluate insect-pest control strategies in tomato and green pepper. Sampling, action thresholds, genetic resistance, chemical control, natural insecticides, irradiation and cultural control were studied. The number of B. tabaci, F. schultzei and M. persicae were sampled, and the rate of virus-infected plants, pepper yield and the costs of the products in relation to the production were determined. Data were subjected to an ANOVA through Fisher’s test, and means were compared by Tukey test at 5% of probability. Our results showed that: a) In tomato: the cultivars AP 529 and AP 533 were less infested by T. absoluta in the first and second experiments, respectively, during the fruiting stage; the conventional strategy was the most efficient in controlling insect vectors, virus infection and the leafminer, and allowed higher fruit yield. The strategies IPM, radiation of 75, 100 and 125 Gy + IPM - 0.5% Neem, IPM with 0.5% Neem and IPM with 0.5% Neem combined with maize barrier, were promising in controlling insect vectors and leafminers at the vegetative stage, when there is low leafminer population pressure; the number of sprayings in IPM is lower than in conventional control, b) In the pepper experiment, the cultivars Priscila and AF 3159 were the least infested by white flies, thrips, aphids and total vectors, but did not show high pepper yield. The most infested cultivars were Magali R and Laser, which were also the most pruductive. The conventional strategy was the most efficient in controlling insect vectors, allowing the highest yields and the lowest relative control costs. The control with 0.5 and 0.75% Neem oil, radiation of 75, 100 and 125 Gy combined with 0.5% Neem oil and 0.5 and 1.0% Neem oil, were promising to control insect vectors, because they allowed high pepper yield. A linear, negative correlation between the rate... (Complete abstract, click electronic access below)
Orientador: Arlindo Leal Boiça Junior
Coorientador: Carlos Frederico Wilcken
Banca: Wilson Badiali Crocomo
Banca: José Roberto Scarpellini
Banca: Antonio Carlos Busoli
Banca: Valter Arthur
Doutor

The overall objective of this study was to test the hypothesis that insects can vector F. tumidum conidia to infect gorse plants with the aim of developing an alternative approach to mycoherbicide delivery to control weeds. Four potential insect species (Apion ulicis, Cydia ulicetana, Epiphyas postvittana and Sericothrips staphylinus) were assessed for their ability to vector F. tumidum conidia. To achieve this, the external microflora (bacteria and fungi) and the size and location of fungal spores on the cuticle of these insect species were determined. In addition, the ability of the insects to pick up and deposit F. tumidum conidia on agar was studied. Based on the results from these experiments, E. postvittana was selected for more detailed experiments to determine transmission of F. tumidum to infect potted gorse plants. The factors promoting pathogenicity of F. tumidum against gorse and the pathogen loading required to infect and kill the weed were also determined. The external microflora of the four insect species were recovered by washing and plating techniques and identified by morphology and polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) and sequencing of internally transcribed spacer (ITS) and 16S rDNA. A culture-independent technique (direct PCR) was also used to assess fungal diversity by direct amplification of ITS sequences from the washings of the insects. All insect species carried Alternaria, Cladosporium, Nectria, Penicillium, Phoma, Pseudozyma spp. and entomopathogens. Ninety four per cent of the 178 cloned amplicons had ITS sequences similarity to Nectria mauritiicola. E. postvittana carried the largest fungal spores (mean surface area of 125.9 ìm2) and the most fungal CFU/insect. About 70% of the fungi isolated from the insects were also present on the host plant (gorse) and the understorey grass. The mean size of fungal spores recovered from the insect species correlated strongly with their body length (R² = 85%). Methylobacterium aquaticum and Pseudomonas lutea were common on all four insect species. Pseudomonas fluorescens was the most abundant bacterial species. In the pathogenicity trials, the effectiveness of F. tumidum in reducing root and shoot biomass of 16 and 8 wk old gorse plants was significantly increased with wounding of the plants. Older plants (32 wk old) which were wounded and inoculated were significantly shorter, more infected and developed more tip dieback (80%) than plants which were not wounded (32%). This indicates that damage caused by phytophagous insect species present on gorse through feeding and oviposition may enhance infection by F. tumidum. Wounding may release nutrients (e.g. Mg and Zn) essential for conidia germination and germ tube elongation and also provide easier access for germ tube penetration. Conidial germination and germ tube length were increased by 50 and 877%, respectively when incubated in 0.2% of gorse extract solution for 24 h compared with incubation in water. Inoculum suspensions amended with 0.2% of gorse extract caused more infection and significantly reduced biomass production of 24 wk old gorse plants than suspensions without gorse extract. A minimum number of about 900 viable conidia/infection site of F. tumidum were required to infect gorse leaves. However, incorporation of amendments (which can injure the leaf cuticle) or provision of nutrients (i.e. gorse extract or glucose) in the formulation might decrease the number of conidia required for lesion formation. Scanning electron micrographs showed that germ tube penetration of gorse tissue was limited to open stomata which partly explain the large number of conidia required for infection. The flowers and leaves were more susceptible to F. tumidum infection than the spines, stems and pods. An experiment to determine the number of infection sites required to cause plant mortality showed that the entire plant needs to be inoculated in order for the pathogen to kill 10 wk old plants as F. tumidum is a non systemic pathogen. The number of infection sites correlated strongly with disease severity (R² = 99.3%). At least 50% of the plant was required to be inoculated to cause a significant reduction in shoot dry weight. F. tumidum, applied as soil inoculant using inoculated wheat grains in three separate experiments, significantly suppressed gorse seedling emergence and biomass production. In experiments to determine the loading capacity of the insect species, E. postvittana, the largest insect species studied, carried significantly more (68) and deposited significantly more (29) F. tumidum conidia than the other species. Each E. postvittana, loaded with 5,000 conidia of F. tumidum, transmitted approximately 310 conidia onto gorse plants but this did not cause any infection or affect plant growth as determined by shoot fresh weight and shoot height. E. postvittana on its own did not cause any significant damage to gorse and did not enhance F. tumidum infection. It also failed to spread the pathogen from infected plants to the healthy ones. There was no evidence of synergism between the two agents and damage caused by the combination of both E. postvittana and F. tumidum was equivalent to that caused by F. tumidum alone. This study has shown that E. postvittana has the greatest capacity to vector F. tumidum since it naturally carried the largest and the most fungal spores (429 CFU/insect). Moreover, it naturally carried Fusarium spp. such as F. lateritium, F. tricinctum and Gibberella pulicaris (anamorph Fusarium sambucinum) and was capable of carrying and depositing most F. tumidum conidia on agar. Coupled with the availability of pheromone for attracting the male insects, E. postvittana may be a suitable insect vector for delivering F. tumidum conidia on gorse using this novel biocontrol strategy. Although it is a polyphagous insect, and may visit non-target plants, F. tumidum is a very specific pathogen of gorse, broom and a few closely related plant species. Hence, using this insect species to vector F. tumidum in a biological control programme, should not pose a significant threat to plants of economic importance. However, successful control of gorse using this "lure-load-infect" concept would depend, to a large extent on the virulence of the pathogen as insects, due to the large size of F. tumidum macroconidia, can carry only a small number of it.

Doenças causadas por virus do gênero Begomovirus (família Geminiviridae) são incomuns em espécies de passifloras. Nos últimos dez anos, entretanto, foram encontrados no Brasil begomovirus infectando passifloras em pomares nos municipios de São Fidelis (RJ), Paragominas (PA), Patos de Minas (MG) e Araguari (MG). Estes isolados foram transmitidos mecanicamente para plantas de Nicotiana benthamiana, que apresentaram sintomas de mosaico e deformação foliar. Também foi possível a transmissão para plantas de Sida rhombifolia através de inoculação por biobalística com o DNA amplificado do isolado de São Fidelis. Estas plantas demonstraram sintomas de mosaico amarelo e deformação foliar. O DNA total extraído de plantas infectadas foi amplificado por RCA, sendo que o componente A (DNA-A) dos isolados de Paragominas e Patos de Minas foram sequenciados diretamente por \"primer walking\". O DNA-A dos isolados de São Fidelis e Araguari foram clonados e sequenciados. As sequências de nucleotideos dos isolados de Paragominas e de São Fidelis apresentaram 90% de similaridade ao Sida mottle virus (SiMoV), enquanto a sequência de nucleotídeos do isolado de Araguari apresentou 96% de similaridade ao Sida micrantha mosaic virus (SimMV). Assim esses isolados encontrados em maracujazeiro podem ser considerados estirpes do SiMoV e SimMV, respectivamente. Não foi possivel a transmissão desses isolados através de Bemisia tabaci biótipo B, no entanto, os insetos foram capazes de adquirir o vírus. O isolado de São Fidelis foi detectado separadamente na região onde se encontra a glandula salivar (cabeça e protórax) e na região posterior do inseto, indicando que o vírus transpos a barreira do mesenteron e circulou pela hemolinfa do inseto. Alves (2008) obteve uma forma atenuada do isolado de São Fidelis através de inoculações mecanicas em plantas de N. benthamiana. O DNA-A desta forma atenuada foi sequênciado e apresentou 90% de identidade ao isolado do qual se originou. A forma atenuada do begomovirus foi capaz de proteger plantas de maracujazeiro contra a estirpe severa do vírus.
Diseases caused by begomoviruses (family Geminiviridae) are hardly found in Passiflora species. In the last years, however, begomovirus infected passionflowers were found in orchards in the counties of São Fidelis (state of Rio de Janeiro), Paragominas (Pará), Araguari and Patos de Minas (Minas Gerais). These isolates were mechanically transmitted to Nicotiana benthamiana plants, which showed variable symptoms of mosaic and leaf distortion. Another susceptible host is Sida rhombifolia, which was biolistic inoculated with amplified DNA of São Fidelis isolate, and showed symptoms of yellow mosaic and leaf distortion. Total DNA extracted from field infected passiflora was amplified by RCA, and the DNA-A of Paragominas and Patos de Minas isolates were directly sequenced by primer walking. The A component of São Fidelis and Araguari isolates were cloned and also completely sequenced. The complete nucleotide sequence of DNA-A of Araguari isolate shared 96% identity with that of Sida micrantha mosaic virus (SimMV), whereas the DNA-A of Paragominas and São Fidelis isolates shared 90% identity with that of Sida mottle virus (SiMoV). These viruses may be consider as strains of SiMoV and SimMV, respectively. It was not possible to transmit these isolates by Bemisia tabaci biótipo B, although the insects were able to acquire the virus. São Fidelis isolate could be detected separately at salivary gland region and posterior region of the insect, indicating that the virus could cross the digestive tract and circulate in the hemolymph. Alves (2008) obtained a mild strain of São Fidelis isolate by mechanical inoculation in N. benthamiana plants. The mild isolate was able to protect passionflower against the severe isoalte of this begomovirus.

Xylella fastidiosa é uma bactéria gram-negativa que coloniza os vasos de xilema das plantas, podendo ser patogênica para algumas culturas de importância econômica. Esta bactéria é transmitida por cigarrinhas (Hemiptera: Auchenorrhyncha) da superfamília Cercopoidea (Aphrophoridae, Cercopidae e Clastopteridae) e família Cicadellidae (subfamília Cicadellinae). Na Itália este fitopatógeno esta causando uma nova doença em plantas de oliveira, denominada síndrome do declínio rápido da oliveira. Recentemente também foi detectada a ocorrência desta bactéria associada a sintomas de requeima foliar na Argentina e no Brasil. Sendo assim, surgiu a necessidade de realizar levantamentos de espécies de cigarrinhas que ocorrem nas regiões onde estão localizados os pomares de oliveira na Serra da Mantiqueira, nos estados de São Paulo e Minas Gerais, na faixa de 800 a 1800 m de altitude. O objetivo deste trabalho foi realizar a análise faunística das espécies de cigarrinhas, como também mensurar a similaridade da comunidade de espécies encontradas entre os pomares, e indicar possíveis vetores de Xylella fastidiosa para a cultura da oliveira. Foram selecionadas sete pomares de oliveira localizados na Serra da Mantiqueira, seguindo um gradiente altitudinal, sendo estes: Wenceslau Braz (MG-1780 m), São Bento do Sapucaí (Entre Vilas) (SP-1602 m), São Bento do Sapucaí (São José) (SP-1512 m), Maria da Fé (MG), com três pomares (área do Lago-1329 m, área da Suíça-1318 m, área da Atemoia-1310 m) e Cabreúva (SP-883 m). Para a captura de cigarrinhas, foram instalados em cada pomar nove cartões adesivos amarelos, em duas alturas, 0,8 m e 1,6 m do nível do solo, totalizando 18 cartões por pomar, que foram trocados a cada 15 dias no período de junho de 2015 a maio de 2017. Coletaram-se 40.324 indivíduos de 270 espécies incluídos em Auchenorrhyncha, distribuídos em 12 famílias, com maior frequência de indivíduos do grupo Cicadellidae (68%), e Cicadellinae (43% de todos os cicadelídeos coletados). A análise faunística indicou 44 espécies predominantes, sendo 20 pertencentes a grupos de potenciais vetores. Dentre elas, Clastoptera sp. 1 (Clastopteridae), Macugonalia cavifrons e Scopogonalia sp. 1 (Cicadellinae) foram predominantes em todos os pomares, exceto no pomar de Cabreúva, sendo que a primeira coloniza a oliveira. Os maiores índices de diversidade, riqueza e equitabiliade foram obtidos para os pomares de Wenceslau Braz, São Bento do Sapucaí (São José) e São Bento do Sapucaí (Entre Vilas) (0,87), respectivamente. A maior similaridade de Jaccard foi encontrada entre os pomares de Maria da Fé (55%). A maior similaridade de Morisita foi encontrada entre os pomares de São Bento do Sapucaí (São José) e Maria da Fé (área da Suíça) (91%) e a menor entre os pomares de Cabreúva e São Bento do Sapucaí (Entre Vilas) (6%). Para algumas espécies predominantes de Cicadellinae e Cercopoidea, houve diferença na eficiência de captura por armadilhas adesivas posicionadas em alturas distintas na copa das oliveiras.
Xylella fastidiosa is a gram-negative bacterium that colonizes the xylem vessels of plants and may be pathogenic to some crops. This bacterium is transmitted by leafhoppers (Hemiptera: Auchenorrhyncha) from the superfamily Cercopoidea (Aphrophoridae, Cercopidae and Clastopteridae) and family Cicadellidae (subfamily Cicadellinae). In Italy this phytopathogen is causing a new disease in olive trees, named \'olive quick decline syndrome\'. Recently, the occurrence of this bacterium associated to a \'leaf scorch\' was also detected in Argentina and Brazil. Thus, it is necessary to carry out surveys of Auchenorrhyncha species in olive orchards in the region of the Mantiqueira Mountain Range, where the olives are grown in the states of São Paulo and Minas Gerais, at 800 to 1800 m altitude range. The objectives of this work were to do faunistic analysis of Auchenorrhyncha species, to measure species similarity of the community of Auchenorrhyncha among orchards, and to indicate possible vectors of Xylella fastidiosa in olives. Seven olive orchards located in the Mantiqueira Mountain Range were selected, following an altitudinal gradient: Wenceslau Braz (MG-1780 m), São Bento do Sapuccaí (EntreVilas) (SP-1602 m), São Bento do Sapucaí (São José) (SP-1512 m), Maria da Fé (MG), with three orchards (area of Lago-1329 m, area of Suíça-1318 m, area of Atemoia-1310 m) e Cabreúva (SP-883 m). Nine yellow stick traps were installed in each orchard at two heights, 0.8 m and 1.6 m above soil level, totaling 18 cards per orchard, which were replaced fortnightly from June/2015 to May/2017. A total of 40,324 individuals from 270 species included in Auchenorrhyncha were collected, distributed in 12 families, with the highest frequency of individuals in the group Cicadellidae (68%) and group Cicadellinae (43% of all Cicadellidae). The faunistic analyses classified 44 species as predominant, 20 belonging to groups of possible vectors of X. fastidiosa. Among them, the species Clastoptera sp. 1 (family Clastopteridae), Macugonalia cavifrons and Scopogonalia sp. 1 (subfamily Cicadellinae) are predominant in all orchards, except in the Cabreúva orchard. Nymphs of Clastoptera sp. 1 were observed developing on young olive branches. Higher diversity, richness and equitability indices were observed in Wenceslau Braz, São Bento do Sapucaí (São José) and São Bento do Sapucaí (Entre Vilas) orchards, respectively. Jaccard\'s greatest similarity was found between Maria da Fé orchards (55%). The greatest similarity of Morisita was found between the orchards of São Bento do Sapucaí (São José) and Maria da Fé (91%) and the lowest one between orchards of Cabreúva and São Bento do Sapucaí (Entre Vilas) (6%). For some predominant species of Cicadellinae and Cercopoidea, capture efficiency was influenced by trap height on the olive tree canopy.

Disertacijos darbo tikslas – aptikti ir identifikuoti Lietuvoje paplitusias fitoplazmas vabzdžiuose, surinktuose nuo įvairių augalų su fitoplazminiais simptomais ir nustatyti fitoplazmų vabzdžius pernešėjus bei atskleisti identifikuotų ir kitų fitoplazmų filogenetinius giminingumus. Lietuvoje jau žinomos keletas labiausiai paplitusių fitoplazmų grupių bei pogrupių, taip pat aptikta nemažai jų augalų-šeimininkų. Duomenų apie galimus šių bakterijų pernešėjus Lietuvoje beveik nėra. Pernešėjų identifikavimas ir tyrimas padės kurti veiksmingesnes strategijas bei sistemas kovai su fitoplazminėmis infekcijomis. Fitoplazmų ir jų pernešėjų identifikavimas suteiks svarbių duomenų tiriant šių patogenų ekologiją, paplitimą, kilmę, epidemiologiją, plitimo kelius. Informacija bus naudinga Lietuvos ir kaimyninių šalių augalų apsaugai. Taip pat galės padėti nustatant galimų invazinių vabzdžių rūšių bei fitoplazmų kamienų atsiradimą Lietuvoje dėl klimato kaitos. Šio darbo metu pirmą kartą Lietuvoje molekuliniais metodais buvo išaiškinti fitoplazmų vabzdžiai pernešėjai. Daugelis aptiktų fitoplazmų pogrupių nustatytos identifikotuose vabzdžiuose pirmą kartą, kaip Lietuvoje taip ir pasaulyje. Penkiose augalų rūšyse fitoplazmos aptiktos pirmą kartą Lietuvoje. Darbo metu nustatytas vienas visiškai naujas Lietuvai ir pasauliui ir vienas naujas Lietuvai fitoplazmų pogrupiai bei jų augalai šeimininkai, kas prisideda prie Lietuvoje bei pasaulyje aptinkamų fitoplazmų paplitimo ir bioįvairovės tyrimo... [toliau žr. visą tekstą]
The aim of the research was to identify the phytoplasmas detected in insects that were found on various phytoplasma-infected plants, and to reveal phytoplasma insect-vectors as well as phytogenetical relationships of identified phytoplasmas. From previous research, we already know a few mostly widespread phytoplasma groups, subgroups, and many of their host plants in Lithuania. The data on potential vectors of these bacteria are very scarce in Lithuania. The identification and research of insect vectors will help to create more effective strategies and systems to fight with phytoplasmal infections. Identification of phytoplasmas and their vectors will provide important data for research of ecology, distribution, origin, epidemiology, and ways of spreading of these pathogens. Such information is beneficial for plant protection institutions and plant growers in Lithuania and neighbouring countries. It will help to ascertain possible invasive insect species and phytoplasma strains in Lithuania. During this research for the first time in Lithuania, we determined possible phytoplasma insect vectors using molecular biology methods. Most of the detected phytoplasma subgroups were found in the identified insect species for the first time in Lithuania and worldwide. Our data on new potential insect vector species extend the spectrum of phytoplasma vectors in our region. Phytoplasmas were detected for the first time in five plant species in Lithuania. We identified in this work one... [to full text]

Sabe-se que a grande maioria dos fitopatógenos depende quase que exclusivamente de vetores para disseminarem-se para novos hospedeiros, porém pouco foi estudado no que se refere aos efeitos dos micro-organismos sobre seus insetos vetores. Sendo Bemisia tabaci uma praga de elevada importância e vetora de inúmeros vírus para plantas cultiváveis, é de extrema relevância estudar os efeitos provocados pelos vírus sobre seu desempenho biológico. Assim, esta pesquisa objetivou: a) avaliar os efeitos direto e indireto dos begomovírus Tomato severe rugose vírus (ToSRV) e Tomato yellow leaf curl virus (TYLCV) sobre parâmetros biológicos de B. tabaci biótipo B e Q respectivamente, dos quais: duração e viabilidade do período ninfal; razão sexual; fecundidade; fertilidade e longevidade. Os resultados encontrados variam de positivos, neutros a negativos, dependendo do parâmetro, efeito e espécie do vírus estudado. Sendo assim, observou-se que há efeito direto de ambos os vírus na duração do período ninfal, sendo menor em insetos virulíferos que em sadios. Ademais, há um incremento no número médio de ovos depositados por insetos virulíferos com ToSRV (225,2 ovos/fêmea) quando comparado com insetos sadios (180,1 ovos/fêmea). Já TYLCV afetou diretamente a longevidade de machos os quais tiveram a duração da fase adulta incrementada quando virulíferos (30 dias) e 24 dias quando sadios. Há um efeito indireto negativo de ToSRV sobre a viabilidade ninfal de seu vetor, sendo de 52% quando estes são mantidos em plantas infectadas e 86% em plantas sadias de tomate. A razão sexual também foi afetada por este vírus, favorecendo as fêmeas, sendo de 2:1 a proporção entre fêmeas e machos em plantas infectadas. Ademais, a longevidade de machos foi reduzida drasticamente quando em plantas infectadas com ToSRV. Também foi detectado um efeito indireto positivo do TYLCV sobre a fecundidade de fêmeas de B. tabaci biótipo Q, as quais depositaram em média maior quantidade de ovos em plantas infectadas que em plantas sadias de tomate, sendo 52,8 e 33,2 ovos respectivamente. Tais resultados permitem concluir que, nas condições em que os ensaios foram realizados, ToSRV afeta diretamente de forma positiva seu vetor, enquanto possui efeitos indiretos principalmente negativos sobre parâmetros biológicos de B. tabaci biótipo B. Igualmente TYLCV possui efeitos diretos positivos sobre o biótipo Q da espécie de mosca-branca. Já indiretamente este vírus, diferentemente de ToSRV, afeta positivamente a biologia de seu vetor B. tabaci biótipo Q, favorecendo a fecundidade dos indivíduos que se desenvolveram em plantas infectadas de tomate.
It is known that the vast majority of pathogens relies almost exclusively vector for spreading to new hosts, but little has been studied regarding the effects of micro-organisms on its insect vectors. Bemisia tabaci is a pest of high importance vector of numerous virus to cultivated plants, it is extremely important to study the effects caused by viruses on its biological performance. Thus, this study aimed to: a) evaluate the direct and indirect effects of the begomovirus Tomato severe rugose virus (ToSRV) and Tomato yellow leaf curl virus (TYLCV) on biological parameters of B. tabaci biotype B and Q respectively, of which duration and viability of nymphal development; sex ratio; fecundity; fertility and longevity. The results range from positive, neutral to negative, depending on the parameter, effect and species of the virus studied. Thus, it was observed that there is a direct effect of the both virus in the duration of nymphal development, being less than viruliferous insects in healthy individuals. Moreover, there is an increase in the average number of eggs laid by viruliferous insects with ToSRV (225,2 eggs / female) when compared with healthy insects (180,1 eggs / female). Already TYLCV directly affected the longevity of males which lasted adulthood increased when viruliferous (30 days and 24 when healthy). There is an indirect negative effect on the viability of ToSRV nymphal of its vector, and 52% when they are kept in infected plants and 86% in healthy tomato plants. The sex ratio was also affected by this virus, favoring females, with a ratio of 2:1 between females and males in infected plants. Furthermore, the longevity of males was reduced dramatically when plants infected with ToSRV. We also detected a positive indirect effect of TYLCV on fertility of female B. tabaci biotype Q, which placed greater average number of eggs in infected plants than on healthy plants of tomato, 52,8 and 33,2 eggs respectively. These results indicate that, under conditions in which the tests were performed, ToSRV directly affects positively its vector, while indirect effects has mostly negative on biological parameters of B. tabaci biotype B. TYLCV also has positive direct effects on the Q biotype of the whitefly species. Already indirectly this virus, unlike ToSRV, positively affects the biology of its vector B. tabaci biotype Q, favoring the fecundity of individuals that developed in infected tomato plants.

O brócolis (Brassica oleraceae var. italica) é uma das hortaliças mais importantes do país, cujo volume de comercialização na CEAGESP é de aproximadamente 13 mil toneladas por ano. Recentemente, uma nova doença tem causado perdas relevantes para as culturas instaladas na maior região produtora do Estado de São Paulo. Os sintomas característicos da doença são expressos pelo enfezamento da planta e necrose dos vasos de floema. Devido ao fato destes sintomas indicarem a presença de fitoplasmas nas culturas de repolho e couve-flor, localizadas na mesma região geográfica onde foi observada esta nova doença, levantou-se a suspeita de que estes mesmos agentes patogênicos pudessem estar associados com as plantas doentes de brócolis. Assim, o DNA total de plantas de brócolis sintomáticas foi analisado por PCR com primers específicos para a região 16S rDNA de fitoplasmas. Os resultados revelaram que estes patógenos estavam associados com as plantas doentes. Através das técnicas de RFLP do sequenciamento de nucleotídeos desta mesma região genômica, os fitoplasmas foram identificados como pertencentes aos grupos 16SrI, 16SrIII e 16SrXIII. Através de análise de RFLP, fitoplasmas também foram identificados em diversas espécies de plantas daninhas e em cigarrinhas da família Cicadellidae coletadas em áreas adjacentes a campos de produção de brócolis. Fitoplasmas do grupo 16SrIII foram identificados em plantas daninhas das espécies Agetarum conyzoides (mentrasto), Crotalaria lanceolata (crotalária), Lepidium virginicum (mentruz), Nicandra physalodes (juá-de-capote), Paulicourea marcgravii (erva-de-rato), Ricinus communis (mamona), Sida rhombifolia (guanxuma), Sonchus oleraceae (serralha amarela), Bidens pilosa (picão preto), Erigeron bonariensis (buva), Emilia sonchifolia (falsa serralha), Leonorus sibiricus (rubim), enquanto que fitoplasmas do grupo 16SrVII foram encontrados as últimas quatro espécies citadas. Com relação aos insetos, fitoplasmas foram detectados em indivíduos das subfamílias Deltocephalinae, Agalliinae e Typhlocybinae. Dentro da subfamília Deltocephalinae, a cigarrinha Balclutha hebe portava fitoplasma do grupo 16SrI, enquanto que cigarrinhas das espécies Atanus nitidus, Planicephalus flavicosta e Schapytopius fuliginosus abrigavam fitoplasmas do grupo 16SrIII. Nos tecidos de duas cigarrinhas da subfamília Agalliinae e uma da Typhlocybinae, as quais não foram identificadas quanto a espécie, foram encontrados fitoplasmas do grupo 16SrIII. As análises epidemiológicas revelaram um padrão espacial agregado de plantas doentes e a ocorrência de um maior progresso da doença nos bordos dos campos de cultivo de brócolis, que estão localizados nas proximidades de áreas com a presença de plantas daninhas.
Broccoli (Brassica oleraceae var. italica) is one of the most important vegetables in Brazil, whose trading volume in CEAGESP is approximately 13 000 tons per year. Recently, a new disease has caused significant losses in this crop cultivated in the largest producing region of the São Paulo State. The characteristic symptoms of the disease are expressed by plant stunting and necrosis of phloem vessels. Because these symptoms indicate the presence of phytoplasmas in cabbage and cauliflower crops, grown in the same geographical region, it was suspected that the same pathogens could be associated with the affected broccoli plants. Therefore, the total DNA from symptomatic plants of broccoli was analyzed by PCR with specific primers for the 16S rDNA of phytoplasmas. Through the techniques of RFLP and nucleotide sequencing of the same genomic region, the phytoplasmas were identified as belonging to the groups 16SrI, 16SrIII and 16SrXIII. Through RFLP analysis, phytoplasmas were also identified in several species of weeds and leafhoppers in the family Cicadellidae collected in adjacent areas of broccoli fields. Phytoplasmas belonging of the 16SrIII group were identified in the weeds belonging to the species Agetarum conyzoides, Crotalaria lanceolata, Lepidium virginicum, Nicandra physalodes, Paulicourea marcgravii, Ricinus communis, Sida rhombifolia, Sonchus oleraceae, Bidens pilosa, Erigeron bonariensis, Emilia sonchifolia, Leonorus sibiricus, while phytoplasmas of the 16SrVII group were found in the last four mentioned species. In respect to insects, phytoplasmas were detected in individuals from subfamilies Deltocephalinae, Agalliinae and Typhlocybinae. Within the subfamily Deltocephalinae, the leafhopper Balclutha hebe carried phytoplasmas of the 16SrI group, while that of the species Atanus nitidus, Planicephalus flavicosta e Schapytopius fuliginosus harbored phytoplasmas of the 16SrIII group. In the tissues of two leafhoppers of the subfamily Agalliinae and one of the Typhlocybinae, which were not identified as specie, were found phytoplasmas of the 16SrIII group. The epidemiological analysis revelead an aggregated pattern of the diseased plants and a higher progress of the diseased in the border of the broccoli fields, whitch were located nearby areas where the presence of weeds was abundant.

La Fièvre catarrhale ovine (FCO) est une arbovirose émergente en Europe depuis la fin desannées 90. Elle affecte principalement les ruminants par la piqûre de petits moucheronshématophages, les Culicoides (Diptera : Ceratopogonidae). Pendant l’été 2006, l’introductiondu sérotype 8 de la FCO, dans la région de Maastricht (Pays-Bas) a rapidement diffusé dansles Ardennes, générant de lourdes pertes pour les éleveurs de bovins et d’ovins. Cesévènements interrogent sur la capacité des Culicoides de la région paléarctique à transmettrela FCO. Ils révèlent la nécessité de mieux connaître la biologie de ces diptères.Nous avons développé successivement dans ce travail, trois axes de recherche qui se sontappuyés sur un travail de terrain réalisé principalement au sein de deux élevages situés dansles Ardennes françaises.Dans un premier temps, nous avons réalisé une expérimentation de gorgement de Culicoidesde captures et d’émergences, provenant des Ardennes, sur petits ruminants virémiques pour leBTV8. A l’issue des expérimentations, une femelle gorgée de l’espèce Culicoides obsoletus apondu et a été retrouvée faiblement positive lors de la recherche du génome du virus de laFCO. Les résultats obtenus ainsi que les difficultés rencontrées lors de la réalisation de cetype d’expérimentation sont discutés.Le deuxième travail exposé s’est intéressé au comportement trophique des Culicoides parl’étude de l’origine du repas sanguin de femelles de Culicoides piégées dans des biotopesvariés. A cette fin, nous avons utilisé des marqueurs moléculaires pour amplifier l’ADN devertébré présent dans les estomacs de femelles gorgées. Ces analyses ont permis de mettre enévidence que des espèces appartenant aux complexes Obsoletus, Pulicaris, ou encore,Culicoides dewulfi, avaient un spectre d’hôte large. Certaines d’entre elles peuvent se gorger àla fois sur les ruminants domestiques et sur la faune sauvage. De plus, ce type d’étuderenseigne sur l’écologie des différentes espèces de Culicoides.Enfin, nous présentons les résultats d’une étude faunistique fondée sur des captures avec despièges lumineux, mais aussi, des prélèvements de boue pour la recherche des gîtes larvaires.Les résultats de piégeages entre les deux exploitations ont été comparés, notamment en termesde biodiversité, et sont discutés en regard des différences de pratiques d’élevage entre lesdeux exploitations choisies d’une part, et la mise en évidence des gîtes larvaires d’autre part.De nombreuses espèces de Culicoides ont émergé au laboratoire à partir des prélèvements deboues, qui ont été caractérisés macroscopiquement. Les gîtes larvaires de C. obsoletus, peuconnus jusqu’alors, ont été mis en évidence dans les deux fermes. Ils ont fait l’objet d’un suivisur plusieurs mois.L’ensemble de ces études contribue à la meilleure connaissance des Culicoides présents dansles Ardennes et de leur biologie, elles permettent de rendre compte des espèces qui semblenttrès inféodées à l’élevage de bovins, et celles qui sont plus ubiquistes. Certains travauxprésentés pourraient être poursuivis pour mettre en évidence les espèces ou populations deCulicoides plutôt sylvatiques, et pour mettre en place de nouvelles expérimentations sur lacompétence et la capacité vectorielle des Culicoides
Since the late 90’s, Bluetongue disease (BT) can be considered as an emerging arbovirose inEurope. This disease is mainly transmitted to ruminants by the bites of minute size midges,the Culicoides (Diptera: Ceratopogonidae), also known as biting midges. An outbreak of BTserotype 8 occurred during summer 2006, in the region of Maastricht (Netherlands) andspread quickly to the Ardennes region. The epizooty lead to severe losses in cattle and sheepholdings. These events highlighted the lack of knowledge on the vectorial capacity ofpaleartic Culicoides species, and more generally on their biology.Three approaches are successively treated in this document. They are all based on field workconducted mainly in two holdings located in the Ardennes region.First, an experiment to assess oral susceptibility of Culicoides to Bluetongue virus (BTV) 8was undertaken. Field collected and emerging Culicoides coming from the Ardennes wereengorged on viremic small ruminants. At the end of the experiments, one Culicoides obsoletusfemale was found bloodfed and laid eggs. She was tested for BTV and was found weaklypositive for BTV genome. This result and the difficulties met during the experiment havebeen discussed.The second study focused on the bloodmeal origin of engorged females of Culicoides. Thesewere collected by light traps set in different kinds of environment. Molecular markers wereused in order to amplify the DNA of vertebrates present in the stomach of bloodfed females.Some of the species processed belonging to the Obsoletus or the Pulicaris complex, andCulicoides dewulfi fed on a wide variety of hosts, including domestic ruminants and wildanimals. Moreover, this kind of study brings information on the ecology of different speciesof Culicoides.Finally, a faunistic survey is presented. It was achieved through light trap collections ofmidges and also thanks to the sampling of potential breeding sites. Biodiversity in thecollection of midges captured by light traps between the two holdings were compared.Differences observed are discussed taking into account the differences in breeding practicesbetween the two holdings and the breeding sites investigations. Numerous species ofCulicoides emerged in the laboratory from soil samples which were macroscopicallydescribed. Breeding sites of C. obsoletus, which were not well documentated in the literature,were found in both farms. These were monitored over some months.This work contributes to a better knowledge of the Culicoides present in the Ardennes andtheir biology. It highlights the species which are closely related to the cattle holdingenvironment, and those which are ubiquist. Some of these studies could be continued in orderto highlight the species more related to the forested areas, and to set new experiments onvectorial competence and capacity

A Escaldadura das Folhas da Ameixeira é o principal problema fitossanitário para a produção de ameixas no Brasil. A doença é causada pela bactéria Xylella fastidiosa, transmitida por cigarrinhas (Hemiptera: Cicadellidae, Cicadellinae). Neste trabalho foi realizado o levantamento, análise faunística e a flutuação populacional de Cicadellinae em pomares de ameixeira localizados nos Estados do Rio Grande do Sul (RS) e São Paulo (SP). A captura das cigarrinhas foi realizada com cartões adesivos amarelos (8,5 x 11,5 cm) em dois pomares de ameixeira de cada estado, localizados nos municípios de Paranapanema (SP) e Bento Gonçalves e Farroupilha (RS). Em cada pomar, foram instalados 20 cartões, distribuídos em 10 pontos espaçados 35 x 35 m, com duas alturas de amostragem (0,5 e 1,7m acima do nível do solo). Os cartões adesivos foram trocados quinzenalmente no período de setembro de 2006 a setembro de 2008. Com base na análise faunística, as espécies de cigarrinhas mais abundantes, constantes, freqüentes e dominantes foram analisadas para se conhecer a sua flutuação populacional. Nos dois pomares do RS foram coletados 1103 indivíduos de Cicadellinae distribuídos em 27 espécies. Destas, Erythrogonia dorsalis (Signoret, 1853), Sibovia sagata (Signoret,1854), Spinagonalia rubrovittata Cavichioli, 2008, Macugonalia cavifrons Stål, 1862, Dilobopterus dispar (Germar, 1821), Bucephalogonia xanthophis (Berg, 1879) e Molomea lineiceps Young, 1968 foram predominantes. Em SP, foram coletados 293 cicadelíneos, distribuídos em 10 espécies, sendo Oncometopia facialis (Signoret, 1854) e Molomea consolida Schröder, 1959 predominantes. Verificou-se uma menor diversidade e abundância de espécies de Cicadellinae nos pomares localizados em SP quando comparados com os pomares do RS. A distribuição populacional das espécies predominantes nos pomares do RS concentrou-se nos meses de janeiro a março, e em SP, de outubro a janeiro, correspondendo aos períodos de colheita e póscolheita em ambas as regiões. Os menores índices de captura de cicadelíneos foram obtidos nos meses de entressafra em todos os pomares amostrados.
The plum leaf scald (PLF) is a major threat for plum production in Brazil. PLF is caused by the bacteria Xylella fastidiosa which is transmitted by sharpshooter leafhoppers (Hemiptera: Cicadellidae, Cicadellinae). In this investigation, we surveyed the Cicadellinae species associated with plum orchards located in Rio Grande do Sul (RS) and São Paulo (SP) states, Brazil, in order to identify potential vectors of X. fastidiosa in these regions. We also studied the seasonal fluctuation of predominant species determined by faunistic analysis. In each area, adult sharpshooters were sampled using yellow sticky cards (8,5 x 11,5 cm) in two orchards in Paranapanema (SP) and one in Bento Gonçalves and Farroupilha (RS), respectivelly. In all orchards, traps were spaced 35 m apart in 10 sample units. Vertical distribution was also evaluated by placing cards at 0.5 and 1.7 m above soil level in each sample unit. The survey was conducted during two years from September 2006 to September 2008, by removing sticky cards fortnightly. In plum orchards located in RS state, we collected a total of 1103 specimens of 27 sharpshooter species. Erythrogonia dorsalis (Signoret, 1853), Sibovia sagata (Signoret,1854), Spinagonalia rubrovittata Cavichioli, 2008, Macugonalia cavifrons Stål, 1862, Dilobopterus dispar (Germar, 1821), Bucephalogonia xanthophis (Berg, 1879) and Molomea lineiceps Young, 1968 were predominant in these orchards. We collected 293 individuals of 10 sharpshooter species in SP state, where Oncometopia facialis (Signoret, 1854) and Molomea consolida Schröder, 1959 were predominant. We found a lower diversity and abundance of Cicadellinae species in plum orchards located in SP when compared to RS state. The predominant species were more abundant from January to March in RS and, from October to January in SP, corresponding to plum harvest and pos-harvest periods in both regions. The lowest population of sharpshooters in plum orchards was observed during the plant dormancy period.

As brássicas compreendem diversas espécies de grande relevância comercial dentre as demais espécies olerícolas cultivadas no Brasil. A região localizada próxima à cidade de São Paulo (SP) tem se destacado no cultivo de brássicas, especialmente do repolho, da couve-flor e do brócolis. Em campos de cultivo destas espécies, tem sido observadas plantas exibindo redução de tamanho, inflorescências mal formadas, avermelhamento de folhas e necrose dos vasos condutores. Investigações tem mostrado que estas anormalidades estão associadas aos fitoplasmas e a doença tem sido denominada enfezamento. Ainda, estudos anteriores têm sugerido a ocorrência de algumas espécies de cigarrinhas potencialmente vetoras destes fitoplasmas. Considerando estas informações, o presente trabalho teve por objetivo identificar espécies transmissoras de fitoplasmas para plantas de brócolis, buscando aumentar os conhecimentos sobre os vetores de fitoplasmas envolvidos com o enfezamento desta cultura. Para isto, foram coletados insetos no interior e áreas marginais de campos cultivados. Estes insetos foram separados em grupos, identificados taxonomicamente e confinados em plantas sadias de brócolis. A avaliação da transmissão foi feita com base na detecção de fitoplasmas nos tecidos dos insetos e das plantas, usando-se a técnica de PCR duplo, com primers específicos para identificação de fitoplasmas do grupo 16SrIII. A sobrevivência dos insetos nas plantas de brócolis foi pouco duradoura, não excedendo 48 horas. A transmissão experimental foi constatada em 30% das plantas inoculadas. Dentre as 8 potenciais espécies de vetores que foram testadas, as espécies Atanus nitidus, Balclutha hebe, Agalliana sticticollis e Agallia albidula transmitiram fitoplasma para plantas de brócolis. Os resultados deste estudo confirmaram aqueles obtidos nas investigações anteriores, as quais sugeriam a ocorrência de potenciais espécies vetoras de fitoplasmas dentre aquelas presentes nos campos de cultivo. No entanto, o conhecimento de detalhes sobre a transmissão necessita de estudos com populações sadias e infectivas das espécies vetoras, sob condições controladas. Apesar desta necessidade, uma etapa importante foi cumprida no presente trabalho, o qual se constitui numa contribuição relevante tanto para o conhecimento de aspectos epidemiológicos relacionados à disseminação do agente causal do enfezamento do brócolis como para a área de conhecimento relacionada à transmissão de patógenos por insetos vetores, nas condições brasileiras.
Cole crops include several species of commercial importance among the vegetable crops cultivated in Brazil. The region located near the city of São Paulo (SP) has excelled in the cultivation of brassica, especially cabbage, cauliflower and broccoli. In cultivated fields with these species have been observed plants showing reduction of size, malformed inflorescences, reddening of leaves and necrosis of region of vessels. Previous investigations have shown that these abnormalities are associated with phytoplasmas and the disease has been called stunt. In addition, previous studies have suggested the occurrence of some species of leafhoppers potentially vectors of phytoplasmas. Considering this information, the present study aimed to identify species that transmit phytoplasmas to plants of broccoli, seeking to increase knowledge about vectors of phytoplasmas involved with this culture. Thus, insects were collected within and in marginal areas of cultivated fields. These insects were separated into groups, taxonomically identified and confined in healthy plants of broccoli. The evaluation of transmission was based on detection of phytoplasmas in the tissues of plants and insects using the technique of nested PCR with specific primers for identification of phytoplasmas group 16SrIII. The survival of insects on plants of broccoli was short-lived, not exceeding 48 hours. The experimental transmission was observed in 30% of inoculated plants. Among the 8 potential vector species that were tested, the species Atanus nitidus, Balclutha hebe, Agalliana sticticollis and Agallia albidula transmitted phytoplasma to plants of broccoli. The results of the present study confirmed those obtained in previous research, which suggested the occurrence of potential vector species of phytoplasmas among those present in crop fields. However, details about these species as vectors require the creation of healthy populations of these species and infective for broadcast demonstration in controlled conditions. Despite this need, an important step has been accomplished in this work, which constitutes a significant contribution both to the knowledge of epidemiological aspects related to the spread of causal agent of broccoli stunt and the area of knowledge related to the transmission of pathogens by insects vectors, the Brazilian conditions.

Este trabalho foi realizado com o objetivo de identificar cigarrinhas vetoras ou potenciais vetoras de Xylella fastidiosa Wells et al. em pomares cítricos do litoral norte da Bahia, determinando-se as espécies predominantes com ênfase na subfamília Cicadellinae. Os levantamentos foram conduzidos no período de março/2002 a fevereiro/2003, em cinco propriedades do litoral norte do Estado da Bahia, nos municípios de Alagoinhas, Inhambupe e Rio Real, em pomares de laranja doce [Citrus sinensis (L.) Osbeck] cv. Pêra, enxertada sobre limão cravo (Citrus limonia Osbeck) com idade de 7 a 9 anos. Em cada pomar, foram instalados 15 cartões adesivos amarelos de 8,5 X 11,5 cm, dispostos em um espaçamento de 40x40 m. Os cartões foram amarrados a 1,5 m de altura em ramos da face norte, na periferia da copa, sendo substituídos periodicamente. Também foram realizadas avaliações periódicas na vegetação rasteira presente nas entrelinhas dos pomares através de rede de varredura, amostrando-se, em cada avaliação, cinco pontos escolhidos aleatoriamente e efetuando-se 30 redadas por ponto. Os dados de coleta foram submetidos a uma análise faunística, calculando-se os índices de constância, freqüência, abundância, dominância, diversidade, eqüitabilidade e similaridade. Coletaram-se 1860 espécimes pertencentes a 54 espécies de sete famílias da subordem Auchenorrhyncha, sendo a maioria deles das famílias Cicadellidae (84,3%) e Membracidae (14,2%). Um maior número de espécies de Auchenorrhynca foi observado em pomar vizinho a mata nativa pouco degradada. A subfamília Cicadellinae, que inclui os vetores de X. fastidiosa, foi a mais representativa em número de espécies (15) e de indivíduos coletados (63,92%). Observou-se variação na composição de espécies de Cicadellinae dependendo do método e local de amostragem, evidenciando-se a existência de dois grupos de potenciais vetores habitando a copa das laranjeiras e a vegetação rasteira. Acrogonia flagellata Young, A. citrina Marucci & Cavichioli, Homalodica sp. e Cicadellini sp.1 predominaram nos cartões adesivos amarelos (árvores cítricas), enquanto que Hortensia similis (Walker) e Erythrogonia dubia (Medler) foram dominantes na rede de varredura (vegetação rasteira). Entre as espécies de Cicadellinae já conhecidas como vetoras de X. fastidiosa em citros, apenas A. citrina, Bucephalogonia xanthophis (Berg) e Ferrariana trivittata (Signoret) foram observadas, sendo que as duas últimas foram de ocorrência acidental na vegetação rasteira. Entre os novos potenciais vetores, A. flagellata, Homalodisca sp. e Cicadellini sp.1 são espécies predominantes nos pomares cítricos do litoral norte da Bahia, devendo ser investigadas quanto à capacidade de transmissão de X. fastidiosa para citros.
This study was carried out to identify vectors or potential vectors of Xylella fastidiosa Wells et al. in citrus groves of the north coast of Bahia State, Brazil, based on a faunistic survey and determination of predominant sharpshooter leafhoppers (subfamily Cicadellinae). The survey was carried out from March/2002 to February/2003 in five old sweet orange [Citrus sinensis (L.) Osbeck, cv. Pêra, grafted on rangpur lime (Citrus limonia Osbeck); 7-9 years old] groves of northeastern Bahia State, located in Alagoinhas, Inhambupe and Rio Real. Fifteen yellow sticky cards (8,5 X 11,5 cm) were installed 40x40 m apart in each grove, hanged at the height of 1.5 m on the upper north side of citrus canopies. Cards were replaced monthly or fortnightly. A sweep net was periodically used to sample leafhoppers in the herbaceous weeds inside the groves, by selecting five points at random and performing 30 sweeps in each point (sample unit). Data was uses to calculate indices of constancy, frequency, abundance, dominance, diversity, equitability and similarity. A total of 1860 specimens of 54 species belonging to 7 families of suborder Auchenorrhyncha were collected, most of them in the families Cicadellidae (84,3%) and Membracidae (14,2%). A larger number of species was observed in a grove located nearby a non-degraded native wood. The subfamily Cicadellinae, which includes the sharphooter vectors of X. fastidiosa, showed the largest number of species (15) and specimens (63,92%). There was a variation in Cicadellinae species composition depending upon the sampling method, which indicates that two distinct groups of potential sharpshooter vectors are inhabiting the citrus canopy and the weedy vegetation. Acrogonia flagellata Young, A. citrina Marucci & Cavichioli, Homalodica sp. e Cicadellini sp.1 predominate in the yellow sticky cards hanged on the citrus canopy, whereas Hortensia similis (Walker) e Erythrogonia dubia (Medler) were dominant species on weeds. Among the Cicadellinae species already known as vectors of X. fastidiosa in citrus, only A. citrina, Bucephalogonia xanthophis (Berg) e Ferrariana trivittata (Signoret) were found; the two latter species were accidentally trapped by sweep net in the weedy vegetation. Among the new potential vectors, A. flagellata, Homalodisca sp. e Cicadellini sp.1 are predominant species in citrus groves of the north coast of Bahia and should be tested with respect to their ability to transmit X. fastidiosa to citrus.

Triatoma sordida é considerada de importância secundária no ciclo da Doença de Chagas humana, uma vez que vem ocupando o lugar de Triatoma infestans no peri-domicílio das casas. Triatoma costalimai é considerada uma espécie silvestre e endêmica do cerrado brasileiro. Recentemente tem ocorrido um aumento do número de invasões domiciliares por T. sordida e T. costalimai, devido ao impacto causado pelo homem no meio ambiente. Ambas as espécies já foram encontradas naturalmente infectadas por Trypanosoma cruzi e, portanto, contribuem para o ciclo antropozoótico da doença. O presente trabalho teve como objetivo verificar a variabilidade genética e morfológica dessas duas espécies, por morfometria geométrica alar e da cabeça e marcadores moleculares mitocondriais sendo T. sordida coletado nos estados de Mato Grosso do Sul (5 populações), Goiás (4 populações) e Minas Gerais (3 populações); e T. costalimai coletados nos estados da Bahia (1 população), Goiás (2 populações) e Minas Gerais (1 população). A hipótese Os resultados mostram que as populações de T. sordida encontram-se altamente estruturadas geneticamente, e que a morfologia alar apresenta uma heterogeneidade, o que permite concluir que mesmo estruturadas geneticamente, não há um processo de especiação ocorrendo para essa espécie. As populações de T. costalimai apresentam alta variabilidade morfológica do conexivo, embora as asas e cabeças apresentam certa similaridade entre as populações estudadas. Os marcadores genéticos indicam distinção entre espécimes que apresentam uma linha laranja continua no conexivo daqueles que apresentam manchas laranjas triangulares. As diferenças encontras sugerem que T. costalimai compreende duas subespécies, com diferenças morfológicas e cromáticas.
Triatoma sordida is considered of secondary importance in the cycle of Human Chagas Disease, since it has occupied the place of Triatoma infestans in the peri-domicile of the houses. Triatoma costalimai is a wild and endemic species of Brazilian cerrado. Recently there has been an increase in the number of home invasions by these species, due to the impact caused by man in the environment. Both species have already been found naturally infected by Trypanosoma cruzi and, therefore, contribute to the antropozootic cycle of the disease. The present work had as objective to verify the genetic and morphological variability of these two species, through the geometric morphometry of the head and mitochondrial molecular markers. The results show that the populations of T. sordida are highly structured genetically, and that the wing morphology shows heterogeneity in the wing shape, which allows to conclude that even if genetically structured, there is no speciation process occurring for this species. The populations of T. costalimai have high morphological variability of the connexivum, although the wings and heads present some similarity between the populations studied. Genetic markers indicate a distinction between specimens with a continuous orange line in the connexivum of those with triangular orange spots. The differences found suggest that T. costalimai comprises two subspecies, with morphological and chromatic differences.

Orientador: Alessandra Alves de Souza
Texto em português e inglês
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
Made available in DSpace on 2018-08-25T17:07:39Z (GMT). No. of bitstreams: 1 Niza_Barbara_M.pdf: 2368175 bytes, checksum: 2c97ffad74e7a3e1c33ae99c78818b99 (MD5) Previous issue date: 2014
Resumo: A citricultura brasileira é um importante setor para a economia do país, contribuindo com superávits comerciais e geração de empregos, entretanto, o setor passa por uma grave crise econômica em decorrência do alto custo de produção e do baixo valor pago pela caixa de laranja no Brasil. O principal motivo do alto custo de produção é a alta incidência de pragas e doenças que atingem essa cultura. Dentre as doenças, a Clorose Variegada do Citros (CVC) causada pela bactéria Xylella fastidiosa e transmitida a seus hospedeiros por cigarrinhas vetoras, é a que até hoje causou mais danos à citricultura brasileira. O mecanismo de patogenicidade da X. fastidiosa permanece não conclusivo porém a hipótese mais aceita está relacionada à facilidade da bactéria em colonizar o hospedeiro, ou seja, em se movimentar e multiplicar dentro dos vasos do xilema da planta infectada, seguido da formação do biofilme. O conhecimento da doença bem como das interações planta-patógeno e vetor-patógeno estão muito avançados para a doença de Pierce (PD), doença causada pela X. fastidiosa em videiras nos Estados Unidos. Esse avanço no conhecimento para PD ocorreu principalmente devido à obtenção de estirpes geneticamente modificadas da bactéria, permitindo a execução de estudos funcionais e de colonização. A implementação da aquisição de X. fastidiosa em sistema de dieta artificial para estudos com o vetor também foi de grande contribuição para esse avanço, uma vez que esse sistema elimina a utilização de plantas fonte para aquisição da bactéria. Em citros, sabe-se que existem fontes de resistência natural à CVC como tangerinas, tangors, limas e limões, entretanto, todas as variedades de laranja doce plantadas no Brasil são suscetíveis a essa doença. Sabe-se também que há uma resposta genética diferente entre um genótipo resistente e o suscetível quando inoculados com X. fastidiosa, porém, não se conhece como se dá a colonização in planta, e se existe uma correlação entre a resposta genética da planta e o comportamento da bactéria. Buscando melhorar o entendimento dos fatores envolvidos no patossistema CVC este trabalho teve como objetivo a obtenção de uma estirpe patogênica de X. fastidiosa de citros transformada com a proteína verde fluorescente (GFP) afim de avaliar sua colonização in planta em genótipos parentais e híbridos de citros, resistentes e suscetíveis, além da implementação da aquisição de X. fastidiosa por cigarrinhas vetores por meio do sistema de dieta artificial. A obtenção do transformante de X. fastidiosa expressando GFP permitiu o acompanhamento da colonização da bactéria nos vasos do xilema de plantas suscetíveis e resistentes e a avaliação mostrou uma colonização diferenciada entre caule e pecíolo. Também foi verificado um padrão diferencial de colonização dos caules de genótipos suscetíveis em relação aos resistentes, no qual a bactéria parece não capaz de se mover em genótipos resistentes, permanecendo aprisionada no xilema primário dessas plantas, sugerindo um possível mecanismo de resistência. A implementação da aquisição de células bacterianas em sistema de dieta artificial foi estabelecida com sucesso para vetor e estirpe de X. fastidiosa em citros, abrindo perspectivas para vários estudos na área de interação vetor-patógeno e transmissão da CVC
Abstract: The citrus agribusiness is an important segment for the country economy, contributing to employment and trade surpluses, however, it is passing through an economic crisis on behalf of the high cost of production and the low price paid by the orange box in Brazil. The main reason for the high cost of production is the high incidence of pests and diseases affecting this crop. Among the diseases, the Citrus Variegated Chlorosis (CVC) is caused by the bacterium Xylella fastidiosa and transmitted to its hosts by sharpshooters, and it is the disease that more damage have been causing to the citrus agribusiness in Brazil. The X. fastidiosa pathogenicity mechanism still not clear but the currently accept hypothesis is related to its facility to colonize the host, in other words, on move and multiply within the xylem vessels of an infected host, followed by the biofilm formation. The knowledge about the disease and the interactions between plant-pathogen and vector-pathogen are advanced for the Pierce disease, which is caused by X. fastidiosa on grapes in the United States. This occurs mainly on behalf of bacterial mutant¿s obtainment, allowing functional and colonization studies, besides the artificial diet system establishment for insect vectors studies, once this system eliminates the use of source plant acquisition, a limiting factor for X. fastidiosa acquisition since its colonization in host plants is random. In citrus, is known that natural resistant sources against CVC exists like tangerine, tangors, limes and lemons, while all the sweet orange varieties grown in Brazil are susceptible. Also, was verified a different genetic response between a resistant and a susceptible genotype when inoculated with X. fastidiosa, although, the colonization process in planta still unkown as well as if there is a correlation between the plant genetic response and the bacterial behavior. In order to better understand the factors involved in the CVC pathosystem, this work had the goal of obtain a pathogenic X. fastidiosa strain from citrus transformed with the green fluorescent protein (GFP) to evaluate its colonization in planta on parent and hybrid citrus genotypes, resistant and susceptible, besides de X. fastidiosa artificial acquisition by the insect vector using the artificial diet system. Was obtained a X. fastidiosa strain transformed with the GFP which allowed the bacterial colonization monitoring within the xylem vessels of resistant and susceptible plants and this evaluation showed a differential colonization of stems and petioles. Was also verified a different stem colonization pattern between resistant and susceptible genotypes on which the bacteria seems to be not able to move in resistant ones, staying contained into the primary xylem of these plants, suggesting a possible mechanism of resistance. The bacterial cells acquisition on artificial diet system was successfully established using a citrus insect vector and bacterial strain, opening perspectives for various studies on vector-pathogen interaction and transmission of CVC
Mestrado
Genetica de Microorganismos
Mestra em Genética e Biologia Molecular

A cultura do tomateiro (Solanum lycopersicum L.) é importante mundialmente devido ao alto consumo de seus frutos. Nos últimos anos surgiram nesta cultura no Brasil alguns vírus emergentes com altas taxas de disseminação, como begomovírus e crinivírus, transmitidos pela Bemisia tabaci biótipo B, que podem causar danos à produção do tomateiro. A espécie de begomovírus atualmente mais encontrada no Brasil, em plantios de tomateiro, é o Tomato severe rugose virus (ToSRV). De 2002 a 2004, pesquisadores relataram incidências desse vírus em mais da metade das amostras com sintomas de geminiviroses coletadas em vários estados brasileiros e sua presença continua sendo verificada frequentemente. No ano de 2006, um crinivírus, o Tomato chlorosis virus (ToCV), foi relatado no Brasil, infectando plantas de tomate no Estado de São Paulo e atualmente encontra-se presente em diveros estados brasileiros. Os objetivos desse trabalho foram: determinar os períodos mínimos de acesso à aquisição e à inoculação do ToSRV e do ToCV pela B. tabaci biótipo B; identificar o período de retenção do ToSRV no inseto e a interação do ToSRV e do ToCV na aquisição e na transmissão por esse aleirodídeo. Também foi avaliada a eficiência do inseticida cloridrato de cartape no controle da disseminação primária e secundária do ToSRV pela B. tabaci biótipo B em tomateiros em gaiolas em casa de vegetação. Finalmente avaliou-se a eficiência do aleirodídeo Trialeurodes vaporariorum na transmissão de um isolado brasileiro do ToCV. Os períodos mínimos de acesso à aquisição e à inoculação de ambos os vírus pela B. tabaci biótipo B foram de cinco minutos. O tempo de retenção do ToSRV em B. tabaci biótipo B foi de 25 dias. A eficiência de um único adulto de B. tabaci na transmissão simultânea do ToSRV e do ToCV para tomateiros foi de 44,7%, similar àquela da transmissão isolada do ToRSV (47,4%) e do ToCV (44,7%). A eficiência de T. vaporariorum na transmissão do ToCV foi inferior à da B. tabaci biótipo B. Usando 40 insetos por vaso com duas plantas as eficiências de transmissão foram 57,7% e 100%, respectivamente. O inseticida cloridrato de cartape reduziu a infecção secundária do ToSRV pela B. tabaci biótipo B, mas não foi eficiente para reduzir a infecção primária em tomateiros.
Tomato (Solanum lycopersicum) is one of the leading vegetables grown and consumed in Brazil and in the world, after potato. The importance of tomato is related to its high consumption worldwide and also its nutritive value. Presently the most important virus diseases responsible for yield losses on tomato crops in Brazil are those caused by begomovirus and crinivirus, both transmitted by Bemisia tabaci biotype B. At the moment the prevalent species of begomovirus is Tomato severe rugose virus (ToSRV). From 2002 to 2004, researchers reported incidence of this virus in more than half of the symptomatic tomato samples collected in several Brazilian states. In 2006, a crinivirus, Tomato chlorosis virus (ToCV), was reported for the first time in Brazil, infecting tomato plants in the State of São Paulo and at present the virus occurs in several Brazilian states. The objectives of this study were to determine the minimum acquisition and inoculation access periods of ToSRV and ToCV by B. tabaci biotype B; identify the retention period of ToSRV in the insect; and the interaction of ToSRV and ToCV on the transmission by this aleyrodidae. It was also evaluated the effectiveness of the insecticide cartap hydrochloride in controlling the primary and secondary spread of ToSRV by B. tabaci biotype B on tomato plants in a greenhouse. Finally, it was evaluated the efficiency of Trialeurodes vaporariorum in the transmission of a Brazilian isolate of ToCV. The minimum acquisition and inoculation access periods for both viruses by B. tabaci biotype B were five minutes. The maximum retention time of ToSRV in B. tabaci biotype B was 25 days. The efficiency of a single adult of B. tabaci to simultaneously transmit ToSRV and ToCV to tomato plants was 44.7%, similar to the transmission of ToRSV (47.4%), and ToCV (44.7%) separately. T. vaporariorum was less efficient than B. tabaci on the transmission of ToCV. Using 40 insects per pot with two plants, transmission efficiencies were 57.7% and 100%, respectively. The insecticide cartap hydrochloride reduced secondary infection of ToSRV transmitted by B. tabaci biotype B, but was not effective in reducing the primary infection in tomato.

Huanglongbing (HLB), considerada uma das mais graves doenças dos citros, está associada a Candidatus Liberibacter spp., bactérias endógenas restritas ao floema e de difícil cultivo em meio de cultura. Diferentemente de outras doenças que afetam plantas cítricas, ainda não foram encontradas dentro do gênero Citrus espécies resistentes ao HLB. Genes de interesse agronômico têm sido empregados na transformação genética de citros visando a resistência a doenças. Dentre estes, destacam-se os que conferem resistência a bactérias incluindo attA, que codificam os peptídeos antibacterianos atacinas, e hrpN, que ativam a produção de proteínas harpinas, relacionadas ao sistema de defesa. O objetivo deste trabalho foi avaliar a resistência de plantas de laranja doce contendo o gene da atacina A (attA) ou o gene da harpina (hrpN) à Candidatus Liberibacter asiaticus (CLas) utilizando duas formas de inoculação: borbulhões infectados e o inseto vetor da bactéria, Diaphorina citri. Para as plantas contendo o gene hrpN, apenas o segundo método foi utilizado. Os principais sintomas do HLB foram observados aos quatro e oito meses após a inoculação por borbulhões. A infecção das plantas foi confirmada com a detecção de CLas por PCR (quatro e oito meses) e Rt-qPCR (oito meses). Em plantas inoculadas por D. citri, os sintomas foram observados oito e doze meses após a inoculação, assim como a detecção da bactéria por PCR. Após 15, 17 e 18 meses, foi realizada uma nova avaliação por Rt-PCR a partir de spots (imprints em membrana) de folhas. Rt-PCR foi empregado também em spots dos psilídeos utilizados na inoculação. Não foi possível avaliar a resistência ao HLB em plantas contendo o gene attA ou hrpN a partir da inoculação por D. citri. Os resultados de detecção da bactéria nas plantas e nos psilídeos utilizados para inoculação indicam que, possivelmente, não ocorreu a inoculação devido ao baixo percentual de psilídeos que continham CLas utilizados. Dentre as plantas transgênicas contendo o gene attA inoculadas por borbulhões infectivos, oito eventos (cinco de laranja Pera, dois de laranja Hamlin e um de laranja Valência) apresentaram menores títulos bacterianos e algumas também demonstraram redução dos sintomas do HLB quando comparadas com plantas não transgênicas, oito meses após a inoculação, indicando uma possível ação do peptídeo atacina A contra o agente causal do HLB.
Haunglongbing (HLB), considered one of the most serious diseases of citrus, is associated to Candidatus Liberibacter spp., endogenous and phloem-inhabiting bacteria not easily grown in culture medium No species within the genus Citrus is known to resist this bacterial infection. The use of genes of agronomic interest for genetic transformation aiming disease resistance in citrus has been reported. Among these genes, attA that codes for the antibacterial peptides attacin, and hrpN, that codes for proteins harpin that activate the plant defense system may have potential in searching for HLB resistance. The objective of this study was to evaluate the resistance of sweet orange containing attA or hrpN to Candidatus Liberibacter asiaticus (CLas) inoculated through infected budstick grafting or the insect vector, Diaphorina citri. For the plants containing hrpN, only the second method was used. The most obvious HLB symptoms were observed four and eight months after inoculation by infected budstick when CLas also was detected by PCR (four months) and RT-qPCR (eight months). For those inoculated with D. citri, symptoms were observed and bacteria detected eight and twelve months after inoculation. Fifteen, 17 and 18 months after inoculation, a new attempt was made for CLas detection, now through Rt-PCR from leaf and psyllids imprinting spots on membrane. It was not possible to evaluate the HLB resistance in plants containing attA or hrpN gene from D. citri inoculation. The results of CLas detection in plants and psyllids indicate that possibly there was no inoculation due the low rate of psyllids contained CLas used. Among the plants containing attA, five, two and one event of, respectively, Pera, Hamlin, and Valencia sweet orange had lower bacterial titers than those non transgenic plants and some also showed milder HLB symptoms, eight months after inoculation, suggesting a possible effect of attacin A against the causal agent of HLB.

O psilídeo Diaphorina citri é vetor das bactérias Candidatus Liberibacter asiaticus e Ca. L. americanus, associadas ao Huanglongbing (HLB) dos citros. Apesar da grande importância desta doença no mundo, sabe-se pouco sobre a atividade alimentar deste vetor, o que seria fundamental para entender o processo de transmissão desses patógenos e aprimorar estratégias de manejo da doença. Assim, esta pesquisa teve por objetivos: a) examinar aspectos morfológicos do aparato bucal de D. citri; b) caracterizar o seu comportamento alimentar em mudas de laranja doce (Citrus sinensis) através da técnica de Electrical Penetration Graph (EPG)-sistema DC; e c) avaliar o efeito da idade de folhas cítricas na penetração estiletar deste inseto e na eficiência de aquisição de Ca. L. asiaticus. Adultos de D. citri apresentam um rostro que se projeta logo após o par de coxas protoráxicas. Verificou-se que na extremidade distal desta estrutura há quatro pares de sensilas em simetria bilateral. O rostro abriga um feixe de quatro estiletes (2 mandíbulas e 2 maxilas) com comprimento médio de 512 µm. Os estiletes maxilares se acoplam formando os canais alimentar e salivar com diâmetros de 0,90 µm e 0,47 µm, respectivamente; esses dois canais se fundem próximo à extremidade distal de uma das maxilas para formar um canal comum, que se estende por, aproximadamente, 4,1 µm. No estudo de EPG, as formas de onda foram descritas quanto à amplitude, frequência, nível de voltagem e origem elétrica, sendo correlacionadas com atividades estiletares baseando-se em análises histológicas da posição da extremidade distal da bainha salivar no tecido vegetal e semelhanças com outras formas de onda descritas para afídeos. Cinco formas de onda foram descritas: (C) penetração intercelular no parênquima; (D) contato com o floema; (E1) salivação no floema; (E2) ingestão no floema; e (G) ingestão no xilema. Com um tempo médio de 206,1 min em 8 h de registro, a ingestão no floema (E2) é a principal atividade observada em D. citri. O tempo médio para atingir o floema desde a primeira prova é de 154 min, após 20,3 provas. D. citri mostra nítida preferência por folhas novas da porção superior da planta (71,3% dos indivíduos), e pela nervura principal na face abaxial das folhas (87,1%). Cortes histológicos revelam que folhas maduras apresentam lignificação nas paredes de células que compõem a camada de fibras, tecido que antecede o feixe floemático; já as folhas novas não apresentam esse espessamento, favorecendo o caminhamento estiletar rumo ao floema. Adultos de D. citri adquirem Ca. L. asiaticus com maior eficiência em folhas novas (assintomáticas) do que em folhas maduras (sintomáticas) de plantas infectadas. Aplicando-se a técnica de EPG, verificou-se que as folhas novas favorecem a alimentação floemática de D. citri; quase que a totalidade dos indivíduos avaliados sobre folhas novas assintomáticas encontraram o floema e permaneceram por mais tempo neste tecido, o que não foi constatado sobre folhas maduras sintomáticas. Esta pesquisa mostra que o comportamento alimentar de D. citri é influenciado pela idade das folhas cítricas, com implicações sobre a eficiência de aquisição da bactéria.
The Asian citrus psyllid, Diaphorina citri, is the vector of the phloem-limited bacteria Candidatus Liberibacter asiaticus and Ca. L. americanus, which are associated to the serious disease known as Citrus Huanglongbing (HLB). Despite the importance of this disease worldwide, little is known about the feeding activities of the vector, which is essential to understand transmission mechanisms and establish disease management strategies. Thus, the goals of this research were to examine the morphology of D. citri mouth apparatus, to characterize its probing behavior on sweet orange (Citrus sinensis) seedlings by using the Electrical Penetration Graph (EPG)-DC technique, and to assess the effect of citrus leaf age on the stylet penetration and efficiency of acquisition of Ca. L. asiaticus. D. citri shows a coneshaped rostrum that extends itself to behind the prothoracic pair of thighs. By electron microscopy, we found four pairs of sensilla symmetrically distributed on the distal end of the rostrum. The rostrum houses a stylet bundle comprised of two mandibulae and two maxillae with mean length of 512 µm. The two maxillary stylets are hold together and have internal grooves that form the food and salivary duct, with mean diameters of 0.9 and 0.47 µm, respectively. These two canals merge at a distance of 4.1 µm from the distal tip of the stylets, forming a common duct. In the EPG study, waveforms were described according to characteristics of amplitude, frequency, voltage level and electrical origin, and correlated with stylet activities based on histological analysis of salivary sheath termini in the plant tissue and similarities with EPG waveforms previously described for aphids. Five distinct waveform types were described: (C) intercellular stylet pathway through parenchyma, (D) phloem contact, (E1) salivation in the phloem sieve tubes, (E2) phloem sap ingestion, and (G) in xylem sap ingestion. Phloem sap ingestion (E2) is the main activity observed for D. citri, with an average time of 206.1 min in 8 h of recordings. Mean time to reach sieve tubes from the first probe is 154 min, after 20.3 probes. This insect shows a clear preference for young leaves of the upper portion of the plant (71.3% of the individuals), and main vein on the abaxial leaf surface (87.1%). Mature leaves show lignification in cell walls of the fiber layer adjacent to the phloem. In contrast, young leaves do not show cell wall thickening in this fiber layer, favoring stylet pathway towards the phloem. Adults of D. citri acquired L. Ca asiaticus with greater efficiency on young (asymptomatic) leaves than in mature (symptomatic) leaves from infected plants. Applying the EPG technique, it was found that young asymptomatic leaves favor phloem feeding by D. citri, since most individuals reached phloem sieve tubes and remained longer in this tissue; this was not the case for individuals confined on mature symptomatic leaves. This study shows that feeding behavior of D. citri is influenced by citrus leaf age, with implication on efficiency of bacterial acquisition from infected plants.

Made available in DSpace on 2015-03-26T13:30:51Z (GMT). No. of bitstreams: 1 texto completo.pdf: 408527 bytes, checksum: 3f5ff39d24998982838ce196768d85c4 (MD5) Previous issue date: 2011-02-18
Conselho Nacional de Desenvolvimento Científico e Tecnológico
Predators capture, kíll and consume theír prey. Thís ís the only ap- proach consídered ín rnost studíes Of predator-prey ínteractíons. However, sorne recent studíes have been concerned Wíth the índírect effects Of pre- datíorL When threatened, prey can change theír behavíor, thus reducing theír chances Of death. Frorn thís perspectíve, experíments performed wíth rnosquítoes specíes ín aquatíc envíronments shows that, ín several ínstances, preys are able to ídentífy the presence Of predators. Thís perceptíon leads to behavíoral alteratíons such as fernale ovíposítíon repellency and reduced foragíng 1arvae. However, the type Of sígnal perceíved by the prey, the írn- portance Of índuced responses to the survíval Of índívíduals, and the reasons for these evolve ín sorne species and not others, remain under díscussíon. The objectíve Was to deterrníne Whether chemícal cues of predator larva Toxorhyn- chites theobaldi (Díptera: Culícídae) are capable Of ínñuencíng the behavíor Of fernales and ímmatures ín three prey species Of the sarne farníly, although wíth Very dístínct behavíor (Culea: molli8, Limatus durhamii and Aedes al- bopictu5). We found that, the ínfusíon Where T. theobaldi larvae Was shel- tered, probably releasíng chernícal sígnals, had a lower proportíon of larvae (5.4 %) of C. molli5, cornpared Wíth control (95.6 %), índícatíng ovíposítíon repellency. Meanwhíle, the other two specíes ovíposíted índíscrímínately on ínfusíon treated and controL We also observed that the presence of traces of the sarne predator, C. mollis larvae spend rnost of theír tíme ín rest (82.7 compared Wíth no sígns of T. theobaldi (49.3 %). However. L. durhamii and A. albopictus larvae do not show behavíoral changes ín the presence of the sarne sígnals. Yet, C. mollis larvae ís preyed upon ín a smaller proportíon than the others, suggestíng that these changes ínñuence ín species Vulnera- bílíty. Frorn these results, ít ís argued that the líkelíhood of the development of defensive responses índuced by predatíon rísk ís related to several factors such as hístoríc of contact Wíth the predator, the ovíposítíon strategy of the females and the ñxed and ñexíble behavíor standard and posítíoníng of 1ar- Vae. Also ís díscussed the ímportance of natíve predators to curb the ínvasíon of exotíc species and díseases spread by these Vectors.
Predadores capturam, abatem e consomem suas presas. Esta é a única abordagem considerada na maior parte dos estudos da ínteração predador- presa. Entretanto, alguns trabalhos recentes têrn se preocupado com os efeitos indiretos da predação. Quando ameaçadas, presas podem alterar seu comportamento, reduzindo suas chances de morte. Sob essa perspectiva, experimentos realizados com espécies de mosquitos em ambientes aquáticos demonstram que, em diversas situações, presas são capazes de identificar a presença de predadores. Tal percepção induz alterações comportamentais como a repelência de ovíposição das fêmeas e redução no forrageamento das 1arvas. Entretanto, o tipo de sinal percebído pelas presas, a importância das respostas induzidas para a sobrevivência dos indivíduos, e as razões destas evoluírem ern algumas espécies, e não em outras, permanecem em discussão. O objectivo do trabalho foi verificar se pistas químicas da presença larva predadora Toxorhynchites theobaldi (Díptera: Culícídae) influenciam o comportamento tanto de fêmeas ovípositantes quanto de imaturos de três espécies de presas da mesma famílía, entretanto com comportamentos bastante distintos, sendo elas Culex molli5, Limatus durhamii e Aedes albopictus (Díptera: Culícídae). Verificamos que, na infusão onde larvas de T. theobaldi permaneceram abrigadas previamente, liberando possíveis sinais químicos, foi encontrada uma menor proporção de larvas (5,4%) de C. molli5, quando comparado com o controle (95,6%) , indicando repelência na ovíposição Enquanto isso, as outras duas espécies ovíposítaram indiscriminadamente na infusão tratada e na controle. Observamos tarnbérn que, na presença de indícios desse mesmo predador, imaturos de C. mollis passaram maior parte do tempo ern repouso (82,7%), quando compadrado com a ausência de sinais de T. theobaldi (49,3%). Entretanto, ímaturos de L. durhamii e A. albopic- tus não apresentam alterações comportamentais na presença dos mesmos sinais. Ainda, C. mollis é predado em uma menor proporção que as demais, sugerindo que essas alterações influenciam na Vulnerabilidade das espécies. A partir desses resultados, é discutido que a probabilidade da evolução de respostas defensivas induzidas pelo risco de predação está relacionada a diversos fatores como o histórico de contato com o predador, a estratégia de ovíposição das fêmeas e o padrão fixo e flexicível de comportamento e posícionarnento das 1arvas. É abordada também a importância dos predadores nativos em conter a invasão de espécies exóticas e a disseminação de doenças por elas Veiculadas.

O Greening ou Huanglongbing (HLB) é a doença mais devastadora dos citros na atualidade. Levantamentos mostram que a enfermidade está presente em cerca de 60% dos talhões do parque citrícola paulista, demostrando sua grande capacidade de disseminação. O psilídeo-asiático-dos-citros, Diaphorina citri Kuwayama (Hemiptera: Psylidae) é um importante vetor dos agentes causais associados ao HLB. Sua capacidade de dispersão está intimamente relacionada ao progresso da doença no campo. Não se tem conhecimento dos principais fatores que possam interferir no deslocamento do inseto até plantas cítricas. Sendo assim, a presente pesquisa teve como objetivo investigar a influência de alguns fatores ambientais sobre a dispersão do psilídeo D. citri, discutindo-os em relação à epidemiologia do HLB dos citros. Inicialmente foram realizados testes em laboratório para adaptação da metodologia de marcação, liberação e recaptura usando pó fluorescente. Grupos de psilídeos adultos foram marcados e sua sobrevivência avaliada, verificando a influência de quatro cores distintas de pó fluorescente (amarelo, azul, rosa e verde). Posteriormente realizaram-se estudos de campo com quatro liberações de psilídeos, com marcadores distintos, em quatro áreas experimentais circulares e adjacentes, sendo formadas por círculos concêntricos localizados a 18, 24 e 30 m de cada centro de liberação. Os insetos foram monitorados visualmente e por armadilhas adesivas amarelas. Nas 1ª e 2ª liberações, foi verificado o efeito das coberturas vegetais milho (Zea mays L.) e capim-braquiária (Brachiaria decumbens Stapf.) na dispersão de D. citri. Já nas 3ª e 4ª liberações, foi estudado o efeito de brotações no deslocamento do inseto. A influência dos ventos foi avaliada em ambas as situações. A análise de sobrevivência indicou que não houve interferência das quatro cores de pó fluorescente em D. citri, sendo a marcação visível no corpo dos insetos durante os 48 dias de avaliação. De acordo com as análises fatoriais, não foi observada qualquer interferência da cobertura vegetal milho, medindo 2 m de altura, na dispersão do inseto. Na presença de brotações, adultos de D. citri dispersaram menos em busca de novos hospedeiros, acumulando-se nas primeiras linhas de plantas cítricas. Após 6 horas da liberação, a capacidade de dispersão máxima de D. citri foi três vezes maior na condição de ausência de brotações. A dispersão média e a área total abrangida variaram assimptoticamente com o tempo, ou seja, o inseto possui inicialmente grande velocidade de dispersão, porém esta tende a se estabilizar com o incremento do tempo. Esses resultados deixam claro que D. citri realiza movimentos dependendo das condições fenológicas de seus hospedeiros, ressaltando a grande importância de se intensificar métodos de controle regionais em épocas em que o fluxo vegetativo das plantas cítricas não é tão intenso.
Greening or Huanglongbing (HLB) is the most devastating disease of citrus today. Surveys show that the disease is present in about 60% of the park stands citrus Sao Paulo, demonstrating its great ability to spread. The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psylidae) is an important vector of the causative agents associated with HLB. Its ability to spread is closely related to disease progression in the field. There is no knowledge of the main factors that can affect the movement of the insect to citrus plants. Thus, the present study aimed to investigate the influence of some environmental factors on the dispersal of psyllid D. citri, discussing them in relation to the epidemiology of HLB of citrus. Initially tests were performed in the laboratory to adapt the methodology of marking, release and recapture using fluorescent powder. Groups of adult psyllids were marked and their survival assessed by checking the influence of four different colors of fluorescent powder (yellow, blue, pink and green). Subsequent studies were conducted with four field releases psyllids, with distinct markers on four experimental areas adjacent circular and being formed by concentric circles located at 18, 24 and 30 m for each center release. The insects were monitored visually and by yellow sticky traps. In the 1st and 2nd releases, was to determine the effect of cover crops maize (Zea mays L.) and grass (Brachiaria decumbens Stapf.) in the dispersal of D. citri. In the 3rd and 4th releases, were studied the effect of shootings in insect movement. The influence of the wind has been evaluated in both situations. Survival analysis indicated that there was no interference of the four colors of fluorescent powder on D. citri, and marking was visible on the body of insects during the 48 days of evaluation. According to the factor analysis, no interference of vegetation corn (measuring 2 m in height) was observed in the dispersal of the insect. In the presence of shoots, adult D. citri less dispersed looking for new hosts and it accumulate in the first lines of citrus plants. After 6 hours of release, the D. citri maximum dispersal was three times higher in the condition of absence of shoots. The dispersal medium and the total area covered varied asymptotically with time, ie the insect has initially high speed dispersal, but this tends to stabilize with increasing time. These results clearly show that D. citri performs movements depending on the phenology of its hosts, emphasizing the importance of intensifying regional control methods at times when the flow of plant citrus is not intense.

Bacillus thuringiensis (Bt) é uma bactéria entomopatogênica utilizada como biopesticida contra insetos, principalmente Lepidoptera, Coleoptera e Diptera, e na produção de organismos geneticamente modificados. Com a descoberta da capacidade de Bt se movimentar sistemicamente em plantas, potencialmente atingindo insetos sugadores, surge uma nova possibilidade para o controle de Diaphorina citri Kuwayama, que transmite bactérias associadas ao Huanglongbing, uma séria doença da citricultura. O objetivo do trabalho foi desenvolver sistemas de aquisição de Bt por D. citri, in vitro e in planta, para estudos de patogenicidade. Uma estirpe de Bt transformada com o gene green fluorescent protein (Btk-gfp), cultivada em meio NYSM, foi usada como marcador de aquisição pelo inseto e movimentação nas plantas. Para o sistema de aquisição in vitro, selecionaram-se dietas com base na sobrevivência e atividade alimentar de D. citri. Btk-gfp foi adicionada à dieta selecionada, composta por uma solução de sacarose a 30% em água mineral com corantes alimentícios (verde 0,1% e amarelo 0,4%). A dieta foi acondicionada em um sachê formado por duas membranas de Parafilm®, sobre a parte inferior de uma placa de Petri de 40 mm de diâmetro (gaiola de alimentação). Dez insetos foram confinados em cada gaiola para períodos de acesso à aquisição (PAA) de até 48 h, estabelecendo-se 10 gaiolas por tratamento; como controle, utilizou-se a dieta sem Btk-gfp. Para testes de aquisição in planta, suspensões de Btk-gfp foram inoculadas em ramos novos cortados e no solo, próximo à haste de seedlings de laranja doce [Citrus sinensis (L.) Osbeck] e de murta [Murraya paniculata (L.) Jack], e em diferentes concentrações, avaliando-se a movimentação da bactéria após um período de 48 h. Posteriormente, adultos e ninfas de D. citri foram confinados nestas plantas para avaliar a aquisição de Btk-gfp, usando-se plantas inoculadas apenas com água como controle. Para os bioensaios de patogenicidade a D. citri, 21 estirpes de Bt foram testadas no sistema de aquisição em ramos cortados de murta e 5 estirpes em ramos cortados de laranja para ninfas de 3º. ínstar. In vitro, testaram-se 3 estirpes contra ninfas de 3º. ínstar e 9 contra adultos. Btgfp foi absorvido pelas raízes e ramos novos cortados de seedlings de laranja doce e de murta, e translocado até as folhas, mostrando movimentação sistêmica. Esta estirpe foi adquirida por adultos e ninfas de D. citri que se alimentaram nos ramos novos cortados, mantidos em suspensão bacteriana. O sistema de alimentação in vitro permitiu a aquisição de Bt-gfp (pellet ressuspendido na dieta de sacarose com corantes) por adultos de D. citri com apenas 12 h de PAA, mas 30 h é um período ótimo para exposição do inseto ao sistema. Nenhuma das nove estirpes testadas no sistema in vitro foi patogênica aos adultos de D. citri. Das 21 estirpes testadas contra ninfas de 3º. ínstar em ramos novos de murta, cinco causaram mortalidade de 24 a 45%, em 5 dias. Essas cinco estirpes também foram testadas contra ninfas dem ramos de citros cortados, causando mortalidade de 35 a 75% em 48 h.
Bacillus thuringiensis (Bt) is an entomopathogenic bacterium widely used as a biopesticide against insect pests, mainly Coleoptera, Lepidoptera and Diptera, or for engineering genetically-modified plants. The recent finding that Bt is able to move systemically within plants, potentially targeting piercing-sucking insects, suggests that the bacterium may be effective for microbial control of Diaphorina citri Kuwayama, the vector of Huanglongbing-associated bacteria, a serious citrus disease. The goal of this research was to develop in vitro and in planta acquisition systems of Bt by D. citri for pathogenicity assays. A transformed Bt strain with the green fluorescent protein gene (Btk-gfp), grown in NYSM medium, was used as a marker to demonstrate bacterial acquisition by the insect and movement within the plants. Artificial diets were selected for the in vitro acquisition system based on insect survival and feeding activity. Btk-gfp was added to the selected diet, a 30% sucrose solution in mineral water with green (0.1%) and yellow (0.4%) food coloring. The diet was placed inside a Parafilm® membrane sachet, covering the opening of the lower half of a 40-mm diameter culture plate, and forming the feeding cage. Ten D. citri adults or nymphs were introduced in each cage and allowed acquisition access periods (AAP) of up to 48 h on the diet; as a control, a diet without Btk-gfp was used. For testing the in planta acquisition system, Btk-gfp suspensions were inoculated in young stem cuttings or in the soil, near the stem of Citrus sinensis (L.) Osbeck and Murraya paniculata (L.) Jack seedlings, as well as in different concentrations, and bacterial movement was assessed after 48 h; plants inoculated with water were used as controls. D. citri adults and nymphs were confined on inoculated plants to verify Btk-gfp acquisition. In pathogenicity assays, 21 Bt strains were tested against 3rd instars of D. citri using the acquisition system with stem cuttings of M. paniculata and 5 strains were tested using citrus stem cuttings. The in vitro acquisition system was used to test pathogenicity of 3 and 9 Bt strains against 3rd-instar nymphs and adults, respectively. Bt-gfp absorbed by roots or young stem cuttings of inoculated C. sinensis and M. paniculata was detected in the leaves, showing systemic movement. Bt-gfp was isolated from groups of nymphs and adults that were fed on inoculated stem cuttings or on artificial diets with bacterial suspension, showing that both in planta and in vitro acquisition take place. D. citri adults can acquire Bt-gfp within 12 h of AAP to the artificial diet with bacterium inoculum, but 30 h is the optimum AAP. None of the nine Bt strains assayed in vitro were pathogenic to D. citri adults. Of 21 strains tested for pathogenicity against nymphs using inoculated stem cuttings of M. paniculata, five caused mortality rates varying from 24 to 45% mortality within 5 days. These five Bt strains were also tested against D. citri nymphs using young stem cuttings of C. sinensis, causing mortality rates of 35 to 75% within 48 h.

A malária é uma doença grave, cujos vetores são mosquitos do gênero Anopheles. Esse gênero é composto por cerca de 500 espécies e aproximadamente 45 são responsáveis pela transmissão do parasito em todo o mundo. Alguns destes vetores são componentes de complexo de espécies crípticas como Complexo Albitarsis e Complexo Triannulatus. O Complexo Albitarsis é formado por quatro espécies: An. albitarsis s.s, An. albitarsis B, An. marajoara e An. deaneorum. O Complexo Triannulatus é composto por três espécies: An. triannulatus, An. halophylus e An. triannulatus C. Algumas técnicas moleculares são utilizadas para auxiliar na distinção destas espécies crípticas. Dessa forma, DNA de An. albitarsis s.l e An. triannulatus s.l., capturados na Usina de Porto Primavera, foram amplificados para ITS2 e para o gene ND4 do DNAmt. Todas as amostras foram submetidas à técnica de RFLP. Algumas amostras foram seqüenciadas diretamente ou clonadas para posterior seqüenciamento a fim de se confirmar a espécie. Das amostras do Complexo Albitarsis, 62,85% foram identificadas como An. deaneorum para ITS2-RFLP. As restantes foram amplificadas para ND4-RFLP e foram identificadas como An. albitarsis s.s. Observou-se, no entanto, que as amostras do Complexo Albitarsis não exibiram similaridade total com as depositadas no GeneBank sendo que para ITS2-PCR estas foram identificadas como An. albitarsis s.s. e An. deaneorum, enquanto que para o gene ND4 todas foram identificadas como An. albitarsis B. O ITS2-RFLP para o An. triannulatus s.l demonstrou polimorfismos entre as espécies. No seqüenciamento, estas amostras foram similares àquelas depositadas no GeneBank. O emprego de marcadores moleculares podem, em parte, auxiliar na distinção de espécies crípticas, porém outros marcadores devem ser avaliados a fim de se elucidar a identificação do Complexo Albitarsis.
Malaria is a severe disease whose vectors are mosquitoes belonging to the genus Anopheles. This genus contains 500 species of which approximately 45 are responsible for the worldwide transmission of these parasites. Some of these vectors belong to cryptic species such as those of the Albitarsis and Triannulatus Complex. The Albitarsis Complex is composed of four species: An. albitarsis s.s., An. albitarsis B, An. marajoara and An. deaneorum. The Triannulatus Complex contains three species: An. triannulatus, An. halophylus and An. triannulatus C. We used molecular techniques to differentiate these cryptic species. Thus, DNA of An. albitarsis s.l and An. triannulatus s.l, captured at the Porto Primavera Dam had their ITS2 and their mtDNA, ND4 genes amplified. All samples were analyzed by the RFLP technique. Some samples were directly sequenced while others were cloned for subsequent sequencing for species confirmation. Within the Albitarsis Complex, 62.85% were identified as An. deaneorum by RFLP-ITS2. The remaining were amplified by RFLP-ND4 and identified as An. albitarsis s.s. However the results of sequencing of the samples of the Albitarsis Complex did not overlap entirely with those deposited in the GeneBank since those amplified by RFLP-ITS2 were identified as An. albitarsis s.s. and An. deaneorum while those obtained by RFLP-ND4 were identified as An. albitarsis B. Also RFLP-ITS2 of A. triannulatus s.l. contained polymorphic regions among different species. By sequencing, these samples were similar to those deposited in the GeneBank. The use of molecular markers did, in some instances help to distinguish species within cryptic complexes, however other markers need to be evaluated to elucidate further identification of the Albitarsis Complex.

O Huanglongbing (HLB) é uma das mais temidas doenças da citricultura mundial, associada a bactérias do gênero Candidatus Liberibacter, que foram detectadas no Brasil em 2004. Em 2008, detectou-se outra bactéria associada a sintomas de HLB no Estado de São Paulo, que foi caracterizada como sendo um fitoplasma do grupo 16 SrIX. Fitoplasmas são molicutes fitopatógenos restritos ao floema de plantas, disseminados por insetos vetores. O presente trabalho buscou identificar cigarrinhas potencialmente vetoras do fitoplasma associado ao HLB e plantas hospedeiras desses insetos, bem como desenvolver uma técnica de quantificação de DNA desse patógeno em insetos e plantas. Amostragens de cigarrinhas foram realizadas quinzenalmente por 12 meses em dois pomares de laranja com histórico de ocorrência do fitoplasma 16SrIX na região norte do Estado de São Paulo, usando-se armadilhas adesivas amarelas em duas alturas (0,3 e 1,5 m) da copa de árvores cítricas, e rede de varredura na vegetação espontânea. Dados faunísticos identificaram uma espécie de Agalliinae (Agallia albidula Uhler) e três de Deltocephalinae, [Balclutha hebe (Kirkaldy), Planicephalus flavicosta (Stål) e Scaphytopius (Convelinus) marginelineatus (Stål)], como os cicadelídeos (Hemiptera: Cicadellidae) mais abundantes e frequentes nas áreas estudadas. Essas espécies predominaram na amostragem com rede de varredura e na menor altura de coleta com armadilhas adesivas, indicando comportamento de alimentação em vegetação rasteira. Com observações visuais, verificou-se associação das espécies com certas plantas invasoras, e influência da composição florística da vegetação rasteira sobre a abundância das cigarrinhas. S. marginelineatus e P. flavicosta ocorreram com maior frequência em Sida rhombifolia L. e Althernantera tenella Colla, respectivamente, enquanto que A. albidula foi predominante em Conyza bonariensis (L.) Cronq., e B. hebe ocorreu exclusivamente em gramíneas, principalmente Panicum maximum Jacq.. Plantas invasoras amostradas nas áreas foram testadas para a presença do fitoplasma 16SrIX, porém sem resultados positivos. No entanto, amostras de campo da cigarrinha S. marginelineatus foram positivas por PCR e sequenciamento para o referido fitoplasma. Indivíduos de S. marginelineatus criados em laboratório e mantidos por um período de acesso à aquisição de 72 h em citros infectado com o fitoplasma 16SrIX, foram capazes de transmití-lo para citros, após 21 dias de latência, porém com baixa eficiência (0,5%). Por meio de PCR quantitativo desenvolvido para esse fitoplasma, verificou-se baixo título do patógeno tanto em S. marginelineatus, quanto em plantas cítricas infectadas, o que pode explicar, pelo menos em parte, a baixa eficiência de transmissão pelo inseto tendo citros como fonte. Isto sugere a existência de outros hospedeiros mais adequados como fontes de inóculo para aquisição do fitoplasma por S. marginelineatus ou outro vetor ainda desconhecido.
Huanglongbing (HLB) is a severe citrus disease associated to phloemlimited bacteria in the genus Candidatus Liberibacter, which were detected in Brazil in 2004. In 2008, another bacterium was found in association with HLB symptom in the São Paulo State, and characterized as a phytoplasma belonging to 16SrIX group. Phytoplasmas are vector-borne phytopathogenic mollicutes that inhabit plant sieve elements. The goals of this study were to identify potential leafhopper vectors of the HLB-associated phytoplasma and their host plants, as well as to establish a real-time PCR procedure for pathogen quantification in vectors and plants. Leafhoppers were sampled fortnightly for 12 months by yellow sticky cards placed at two heights (0.3 and 1.5 m) on the citrus tree canopy and by sweep net in the ground vegetation of two sweet orange groves with history of infection by the 16SrIX phytoplasma, in the northern region of São Paulo State. Faunistic analyses indicated 1 Agalliinae (Agallia albidula Uhler) and 3 Deltocephalinae [Balclutha hebe (Kirkaldy), Planicephalus flavicosta (Stål) e Scaphytopius (Convelinus) marginelineatus (Stål)] species that were the most abundant and frequent leafhoppers (Hemiptera: Cicadellidae) in the experimental areas. These species predominated in sweep net and in sticky traps catches at 0.3 m above soil, showing that they inhabit the ground vegetation. Visual observations indicated a strong association of leafhopper species with some weeds and the influence of weed species composition on leafhopper abundance in the ground vegetation. S. marginelineatus and P. flavicosta were more frequent on Sida rhombifolia L. and Althernantera tenella Colla, respectively, while A. albidula was observed more often on Conyza bonariensis (L.) Cronq., and B. hebe occurred solely on grasses, more abundantly on Panicum maximum Jacq. Fourteen weed species sampled in the area were PCR tested for infection by the 16SrIX phytoplasma, but none was found infected. Nevertheless, 3 out of 30 field-collected samples (10 adults per sample) of S. marginelineatus tested positive for this phytoplasma by PCR and sequencing. Healthy lab-reared adults of S. marginelineatus were able to transmit inefficiently (0,5%) the 16SrIX phytoplasma to healthy citrus after a 72-h acquisition access period on infected citrus plants followed by a 21-day latent period on S. rhombifolia plants. By using the qPCR method developed for this phytoplasma, a very low pathogen titer was found both in S. marginelineatus and in infected citrus plants, which may explain, at least partially, the low transmission efficiency by this vector using citrus as a source plant. It also suggests that existence of alternative hosts that might be more adequate as inoculum sources for phytoplasma acquisition and spread by S. marginelineatus or another vector yet to be discovered.

The early region of polyomavirus encodes three proteins capable of causing tumors in vivo as well as cellular transformation in vitro. Polyomavirus middle T antigen has an associated tyrosine kinase activity due to its interaction with the cellular tyrosine kinase pp60$ sp{ rm c-src}$. To more closely characterize the interaction of these two molecules I have sought to express polyomavirus middle T antigen and pp60$ sp{ rm c-src}$ in insect cells.
A variety of transfer vectors have been designed to allow the introduction of foreign genes into the genome of baculoviruses. cDNAs encoding polyomavirus middle T antigen and pp60$ sp{ rm c-src}$ have been cloned into two such vectors that differ in their non-coding upstream sequences. Putative recombinant viruses were selected and examined for the expression of these two gene products. Although permanent recombinant viral stocks were not established, it was demonstrated that pp60$ sp{ rm c-src}$ can be produced in insect cells following transient transfection. Furthermore, this protein was shown to be biologically active as measured in the in vitro immune complex kinase assay. pp60$ sp{ rm c-src}$ produced in insect cells is capable to phosphorylating an exogenous substrate, enolase, in addition to undergoing autophosphorylation.

La bactérie Xylella fastidiosa est l'organisme de quarantaine avec une incidence socio-économique et environnementale potentielle considérée comme la plus grave en Europe. Elle est transmise par des insectes vecteurs polyphages. La compréhension des réseaux d'interactions écologiques est un prérequis indispensable à la mise en place de mesures de prophylaxie et de lutte. Cette thèse propose une approche transdisciplinaire avec l'étude des insectes vecteurs et une analyse des enjeux autour de la mobilisation de ces connaissances pour la préparation à une éventuelle crise. En écologie, l'étude des insectes vecteurs est menée à la fois en zone contaminée (Corse) et en zone indemne (principalement la Nouvelle-Aquitaine). En zone contaminée, les travaux de recherche permettent de mettre en évidence la bactérie dans les insectes vecteurs, avec une détection de Xylella fastidiosa à l'échelle du territoire Corse. En zone indemne, les insectes vecteurs potentiels et avérés de la bactérie et les interactions trophiques plantes-insectes dans les parcelles cultivées (principalement la vigne) et les milieux semi-naturels adjacents sont étudiés. Enfin, la mobilisation d'outils sociologiques (observation participante et entretiens semi-directifs) dans le champ des STS apportera un éclairage sur la production des connaissances scientifiques et leur mobilisation ou non dans des dispositifs réglementaires et de surveillance de la maladie. L'étude de la bactérie en zone contaminée a conduit à la mise au point d'une méthode à haut débit pour détecter Xylella fastidiosa dans ses insectes vecteurs et son application sur des insectes collectés en Corse entre 2016 et 2020 permet de mettre en évidence que la bactérie est présente sur l'ensemble de l'île. De plus, les résultats de modèles statistiques montrent l'influence de la température sur les populations de bactérie avec une prévalence plus importante lorsque les températures hivernales sont plus douces. Dans une perspective de changements globaux, des projections climatiques jusqu'en 2100 ont été utilisées et ont montré que les aires favorables à la bactérie et son principal insecte vecteur resteraient importantes en Corse et largement superposées avec des déplacements possibles du pathosystème en altitude. Les études en zones indemnes ont montré que Philaenus spumarius, Neophilaenus capestris, Neophilaenus lineatus et Cicadella viridis, sont les quatre espèces d'insectes vecteurs principalement retrouvées sur le continent en France. Ces espèces sont majoritairement collectées en strates herbacées de prairies et en bordures de parcelles cultivées (vigne) par rapport aux inter-rangs. Ces observations mettent en évidence que les milieux semi-naturels et les bordures de parcelles sont plus favorables à la propagation de la bactérie et le risque de transmission de la bactérie aux plantes cultivées semble plus modéré car peu d'insectes ont été collectés dans les parcelles. Les résultats issus de la recherche académique sont mobilisés dans d'autres contextes dont le contexte réglementaire. Dans le cas de Xylella fastidiosa, les résultats montrent que les recherches mondiales sont partagées par l'ensemble de la communauté scientifique bien qu'elles soient regroupées selon la discipline et la maladie étudiées. En France, ces recherches sont dissociées selon les disciplines et les instituts et sont également impactées par la médiatisation de cette bactérie de quarantaine à lutte obligatoire. Enfin, le dispositif d'épidémio-surveillance concernant Xylella fastidiosa en France et appuyé sur la plateforme d'épidémio-surveillance en santé du végétal résulte d'arrangements entre la recherche et le gestionnaire du risque. Ainsi, cette thèse propose une démarche transdisciplinaire afin d'éclairer la gestion intégrée des organismes pathogènes dans le cadre de la preparedness avec la production de résultats en écologie et l'analyse du système d'épidémio-surveillance de la bactérie en France
The bacterium Xylella fastidiosa is the quarantine pest with potential socio-economic and environmental impact considered to be the most serious in Europe. It is transmitted by polyphagous insect vectors. Understanding host plant - insect vector - bacterium interaction networks is an essential prerequisite to implement prophylactic and control measures. This thesis proposes a transdisciplinary approach with the study of insect vectors in ecology and an analysis of the issues around the mobilization of this knowledge for the preparation for a possible crisis. In ecology, the study of insect vectors of the bacterium is carried out both in contaminated areas (Corsica) and in unscathed areas (mainly New Aquitaine). In contaminated areas, research work will make it possible to highlight the bacterium in insect vectors, with the detection of Xylella fastidiosa across the Corsican territory. In free areas, the objective is to study potential and proven insect vectors of the bacterium and plant-insect trophic interactions in crops (mainly vines) and adjacent semi-natural environments. Finally, the mobilization of sociological tools (participant observation and semi-structured interviews) in the field of the study of science and methods will shed light on the production of scientific knowledge and its mobilization or not in regulatory and monitoring systems of the disease. The study of the bacterium in contaminated areas has led to the development of a high-throughput method to detect Xylella fastidiosa in its insect vectors. The application of this method on insects collected in Corsica between 2016 and 2020 makes it possible to demonstrate that the bacterium is present throughout the island. In addition, the results of statistical models show the influence of temperature on bacterial populations with a significant prevalence when winter temperatures are milder. From the perspective of global changes, climate projections up to 2100 were used and showed that the areas favorable to the bacterium and its main insect vector would remain significant in Corsica and largely superimposed with possible movements of the pathosystem at altitude and therefore that the risk of an epidemic would remain present. Studies in free areas have shown that Philaenus spumarius, Neophilaenus capestris, Neophilaenus lineatus and Cicadella viridis are the four vector insect species mainly found on the mainland in France. These species are mainly collected in herbaceous strata of meadows and on the edges of crops (vineyards) in relation to the inter-rows. These observations show that semi-natural environments and the edges of plots are more favorable to the spread of the bacterium and the risk of transmission of the bacterium to cultivated plants seems more moderate because few insects were collected in the plots. The results from academic research are used in other contexts, including the regulatory context. In the case of the bacterium Xylella fastidiosa, the results show that global research is shared by the entire scientific community although it is grouped according to the discipline and the disease studied. In France, this research is dissociated according to the disciplines and the institutes and it is also impacted by the media coverage of this quarantine bacterium which must be controlled. Finally, the epidemio-surveillance system concerning Xylella fastidiosa in France and supported by the plant health epidemio-surveillance platform results from arrangements between research and the risk manager. Thus, this thesis proposes a transdisciplinary approach in order to shed light on the integrated management of pathogenic organisms within the framework of preparedness with the production of results in ecology and the analysis of the epidemio-surveillance system of the bacterium in France