Tesi: "Pantoea" – Grafiati

1

Brady, Carrie Louise. "Taxonomy of Pantoea associated with bacterial blight of Eucalyptus". Pretoria : [s.n.], 2005. http://upetd.up.ac.za/thesis/available/etd-02092006-110117.

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2

Weller-Stuart, Tania. "Genomic and functional characterization of motility in Pantoea ananatis". Thesis, University of Pretoria, 2015. http://hdl.handle.net/2263/79208.

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Pantoea ananatis causes disease symptoms in a wide range of economically important plants such as Eucalyptus, maize and onions. This study specifically focussed on the interactions of P. ananatis LMG20103 with onion seedlings where it causes symptoms that include water-soaked lesions, wilting and bleaching of the leaves, and maceration of the bulbs. The pathogenicity of P. ananatis is not well understood, however, motility plays an important role in many other well-known phytopathogens such as Ralstonia solanacearum and Pseudomonas syringae. It is therefore hypothesised that P. ananatis uses motility to colonise and infect its hosts. Chapter 1 reviews the literature on how motility aids phytopathogenic bacteria in locating their host, attaching and initiating infection, as well as dissemination. The two dominant forms of motility utilised are swimming and twitching motility. Swimming motility is essentially the rotation of flagella which propels the bacterial cell forward through a fluid environment in response to chemotactic signals. The motor that drives the flagella is made up of several proteins that include the MotAB proteins and its function is dependent on a proton motive force. While twitching motility is not as fast as swimming motility, it is a rapid means of surface colonisation. Bacteria twitch by extending their type IV pili, attaching to the surface, and then retracting, bringing the whole cell closer to the point of contact. This motion is powered by the ATPase PilT. In Chapter 2 the flagellum and type IV pilus biosynthetic gene clusters are compared between strains of P. ananatis and closely related enterobacterial strains. The four fully annotated and sequenced P. ananatis strains used in this study were isolated from various different sources and provided a greater understanding of how P. ananatis exploits its flagella and type IV pili to infect such a wide variety of hosts. While Chapter 3 focuses on the creation of four motility mutants and their respective complements in P. ananatis LMG20103, Chapter 4 consists of an array of tests and assays comparing the mutants to the wild-type strain to elucidate the role of swimming and twitching motility in the colonisation and infection of P. ananatis in onions. Chapter 5 is a published article titled, “Draft genome sequences of the onion centre rot pathogen Pantoea ananatis PA4 and maize brown stalk rot pathogen P. ananatis BD442.” Both strains are South African isolates and were sequenced using the Illumina HiSeq 2500 platform. A greater understanding of how P. ananatis uses motility to target tissues and infect its host plant increases the currently limited body of knowledge available to develop strategies to limit the damage caused by this pathogen to agronomic crops in plantations and nurseries.
Thesis (PhD)--University of Pretoria, 2015.
Microbiology and Plant pathology
PhD
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3

du, Plessis Marike. "Phylo- and comparative genomics of the Pantoea core genome". Diss., University of Pretoria, 2014. http://hdl.handle.net/2263/79232.

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The delineation of bacterial species and genera has always been problematic as a clear definition of these concepts are lacking. In an attempt to classify bacteria into workable groups, operational criteria have been applied to delimitate boundaries for these taxonomic ranks. This approach has unfortunately led to artificial groupings that are often not comparable in terms of diversity in different groups of bacteria. A classification system needs to reflect natural groupings to depict the evolution of bacteria and predict the phenotypic and genetic diversity for these groups. In order to understand the forces that play a role in the evolution of a bacterial genus a review of the current literature was presented in Chapter 1. The major focus was on vertical inheritance and how this process can be used to depict the evolutionary path of members belonging to the same genus. The largest amount of genetic material in any one cell is thought to have been transferred from parent to progeny, supporting the idea that the vertical signal is recoverable and can in fact be the dominant signal present in the genome when looking at conserved genes. The effect of horizontal gene transfer (HGT) on the evolutionary picture obtained by vertical descent was also discussed. The core genome of a genus is defined as the genes conserved between all species of a genus and are thought to mostly include genes that are essential for the survival of members of that particular genus. In Chapter 2, the hypothesis was tested that the boundaries used to delineate genera could be based on an analysis of the shared core genome. For this purpose coherence within the core genome of the genus Pantoea was investigated. The core was characterised in terms of its functional diversity through Clusters of Orthologous Genes (COGs) and compared to the core genomes of other bacterial genera. It was seen that the core genome does give an indication of the coherence of a genus and that shared genome content can be used as a tool to delimitate genera. Previous taxonomic studies have shown that species in the genus Pantoea are well defined but that the phylogenetic relationships between these species are not well elucidated. Generally accepted approaches for phylogenetic inference, like 16S rRNA gene trees and multi-locus sequence analysis (MLSA), does not give sufficient resolution to determine the deeper evolutionary relationships between these species. In Chapter 3, phylogenomic analyses were performed to determine if a robust phylogeny, reflecting the evolutionary history of the genus, can be obtained using the core genome of the genus. The core genome as well as subsets thereof (based on COGs), was used for phylogenetic inference, to obtain a robust phylogeny for the genus.
Dissertation (MSc)--University of Pretoria, 2014.
Microbilogy and Plant pathology
MSc
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4

Galbraith, Matthew Dominic. "Further studies on AGA production by Pantoea agglomerans strain Eh1087". Thesis, University of Canterbury. Biological Sciences, 2004. http://hdl.handle.net/10092/6785.

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Pantoea agglomerans strain Eh1087 produces the phenazine antibiotic Dalanylgriseoluteic acid (AGA). A cluster of 16 genes has previously been shown to be responsible for the production of, and resistance to, AGA. The present study has refined and tested a number of hypotheses arising from the preliminary characterisation of the AGA pathway. The products of the first five genes of the AGA cluster, Group 1, are similar to proteins responsible for phenazine-1-carboxy lie acid by fluorescent pseudomonads. However, Eh1087 appears to be missing a duplication of the ehpA gene, and it was hypothesised that EhpA was responsible for the ability of Eh1087 to produce both phenazine-1- carboxylic acid and phenazine-1,6-dicarboxylic. Comparison of EhpA to related proteins of known structure suggested a catalytic function, and EhpA was found to influence the relative amounts of phenazine-1-carboxylic acid and phenazine-1,6- dicarboxylic acid produced by Group 1. The final step in the AGA pathway is the addition of a D-alanyl residue to griseoluteic acid to form AGA, and is catalysed by the EhpMNO proteins. The previous model for AGA biosynthesis suggested that EhpM was an integral membrane protein and that EhpMNO operated in the periplasm. EhpM and EhpN are similar to components of nonribosomal peptide synthetases, while EhpO is similar to ketosynthases involved in fatty acid and polyketide biosynthesis. Comparison of EhpM with known structures, and preliminary analysis of the subcellular location of EhpMNO suggest that these proteins may be localised to the cytoplasmic side of the inner membrane, rather than the periplasm. An Eh1087 transposon-mutant with an insertion that affected the expression of glnA was isolated. This mutant lacked glutamine synthetase, and was, therefore, not able to incorporate labelled nitrogen from ammonium sulphate, via glutamine, into phenazines. It was concluded that glutamine is the source of the two nitrogen atoms of the phenazine nucleus. In addition, the function of the putative Eh1087 glnA locus was confirmed by complementation of an E.coli glnA mutant and the gene found to encode a type I glutamine synthetase typical of the Enterobacteriaceae.

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5

Sibanda, Siphathele. "Role of quorum sensing in the virulence of Pantoea ananatis". Diss., University of Pretoria, 2012. http://hdl.handle.net/2263/30941.

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Pantoea ananatis, a plant pathogenic bacterium, inflicts significant economic losses to the agricultural and forestry industries. It is ubiquitous and capable of surviving in a diverse range of environmental conditions. The mechanism underlying host infection and colonization by this pathogen is poorly understood. The genome sequence of P. ananatis led to the discovery of putative pathogenicity determinants such as quorum sensing. In this study, a PCR-mediated protocol that makes use of the lambda () red genes was used to knockout the genes for the three quorum sensing systems in P. ananatis LMG 2665T. The mutant strain was named EanΔI/R,RhlΔI/R,ΔLuxS. Growth assays conducted using this mutant and the wild-type strain showed that the mutations did not affect its growth in liquid broth. This mutant was used to determine the role of quorum sensing in the virulence of P. ananatis. Virulence assays conducted showed that quorum sensing is required for virulence in P. ananatis. To elucidate the role of individual quorum sensing systems in the virulence of P. ananatis, mutants lacking one system were constructed following the  Red-mediated PCR protocol. The mutant strains were complemented by cloning the wild-type genes for the respective quorum sensing systems into the broad-host-range plasmid pBR1MCS-5. The mutant strains were named EanΔI/R, RhlΔI/R and ΔLuxS based on the quorum sensing genes that were mutated. The complemented strains were named EanΔI/R::EanI/R, RhlΔI/R::RhlI/R and ΔLuxS::LuxS, respectively. In vitro growth studies showed that the genetically modified P. ananatis strains were not impaired in their growth. The P. ananatis quorum sensing mutant strains and complemented mutant strains were used to determine the functional role of each quorum sensing system in P. ananatis. Characterization of the quorum sensing mutant strains revealed that the three quorum sensing systems are required for virulence of P. ananatis in onion seedlings. The virulence assays conducted showed that the LuxS quorum sensing system is the most crucial system for virulence in P. ananatis. Furthermore, in vitro studies of quorum sensing regulation of specific phenotypes of P. ananatis showed that quorum sensing governs biofilm formation and exopolysaccharide production. The phenotypes that were impaired in the quorum sensing mutant strains were restored to wild-type levels by genetic complementation. This study also showed that swarming and twitching motility, as well as rhamnolipid production are not influenced by quorum sensing in P. ananatis. The dependence of specific phenotypes on quorum sensing indicates the significance of the functional role of quorum sensing genes in the virulence of P. ananatis.
Dissertation (MSc)--University of Pretoria, 2012.
Microbiology and Plant Pathology
MSc
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6

Swart, Lorinda. "Pantoea and Xanthomonas species associated with blight and die-back of Eucalyptus". Diss., University of Pretoria, 2010. http://hdl.handle.net/2263/31420.

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The pulp and paper industry is expanding world-wide to supply the needs and demands of the consumer. Due to this rapid expansion of commercial forests and our ever changing climate including the sporadic increase and decrease in rain and the increasing temperature caused by global warming, previously described and new pathogens are emerging which infect and cause diseases on commercial forest trees and agricultural crops. Research efforts are required to investigate mechanisms of disease control and eradication to prevent the propagation and rapid spread of these pathogens and ensure that there is limited economical loss of forestry and other agriculturally important plants and trees. Since both Xanthomonas and Pantoea species are becoming increasing important as emerging bacterial pathogens, their rapid and accurate identification is crucially important. Little is known about bacterial pathogens on forestry trees since the most prominent diseases of these hosts are caused by fungi. The focus of this study was to investigate and identify the bacterial pathogens associated with Eucalyptus. However, as has been seen in various studies including this one, the identification of these pathogens is not always straightforward and often time consuming. In this study the use of polyphasic identification approach was used which employs a combination of phenotypic and genotypic identification techniques. Both of the genera investigated in this study, namely Pantoea and Xanthomonas, have been found to infect a variety of agriculturally important plant hosts. Pantoea species have previously been isolated from Eucalyptus trees suffering from blight and dieback symptoms. The species isolated have included P. eucalypti from Uruguay, P. vagans isolated from Argentina, Colombia, Uganda and Uruguay and P. deleyi from Uganda. Since the first report of Pantoea on Eucalyptus trees from South Africa in 2002 it has spread locally causing sporadic outbreaks. This pathogen has also been isolated from Eucalyptus trees in other parts of the world including, Argentina, Colombia, Thailand, Uganda and Uruguay. Xanthomonas campestris pv. eucalypti was previously found to cause disease on Eucalyptus trees in Australia. Since then, three other Xanthomonas species have been isolated from Eucalyptus, namely, Xanthomonas spp. from Brazil (Goncalves et al., 2008), X. vasicola from South Africa and X. fuscans from Uruguay as seen in this study.
Dissertation (MSc)--University of Pretoria, 2010.
Microbiology and Plant Pathology
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7

Ramachandran, Revathy. "Investigation of the quorum-sensing regulon in the corn pathogen Pantoea stewartii". Diss., Virginia Tech, 2014. http://hdl.handle.net/10919/56840.

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Pantoea stewartii subsp. stewartii is a bacterium that causes Stewart’s wilt disease in corn plants. The bacteria are transmitted to the plants via an insect vector, the corn flea beetle Chaetocnema pulicaria. Once in the plant, the bacteria migrate to the xylem and grow to high cell densities, forming a biofilm by secreting excess capsular exopolysaccharide, which blocks water transport and causes wilting. The timing of virulence factor synthesis is regulated by the cell-density dependent quorum sensing (QS) system. Such temporal regulation is crucial in establishing infection and is orchestrated by the QS-dependent transcriptional regulator EsaR. EsaR represses expression of capsular exopolysaccharide at low cell densities. At high cell densities, an acylated homoserine lactone (AHL) molecule produced during growth by the cognate AHL-synthase EsaI accumulates. The AHL binds to and inactivates EsaR, causing derepression of capsule production. EsaR is a member of the LuxR family of QS-dependent transcriptional factors. Most LuxR homologs are unstable and/or insoluble in the absence of AHL which has hindered structural studies. Chapter Two describes the changes in the structure of EsaR due to binding of AHL ligand as determined through biochemical methods. EsaR was found to be stable and retain its multimeric state in the absence or presence of AHL, but intra- and inter-domain changes occurred that affect its DNA-binding capacity. Apart from repressing expression of capsule at low cell-densities, EsaR represses its own expression and activates production of a small RNA, EsaS, with unknown function. In Chapter Three a proteomic approach was used to identify an additional 30 QS-controlled proteins. Genes encoding three of these proteins are directly regulated by EsaR and the EsaR binding sites in the respective promoters were defined. In Chapter Four, a high-throughput RNA-Seq method identified even more genes in the QS regulon that the proteomic approach overlooked. RNA-Seq analysis of rRNA-depleted RNA from two strains of P. stewartii was used as a screen to help identify 11 promoters, subsequently shown to be directly regulated by EsaR in vitro. Most of the genes controlled by QS grouped into three major physiological responses, capsule & cell wall production, surface motility & adhesion and stress response. In Chapter Five, the role of two QS regulated genes, dkgA (encoding 2, 5-diketo-D-gluconate) and lrhA (encoding a repressor of chemotaxis, adhesion and motility), in plant virulence were examined. These studies have better characterized the QS regulator EsaR and its interaction with the AHL ligand, and shown that QS has a more global response in P. stewartii than previously recognized. Further characterization of the genes identified in this study could facilitate identification of factors crucial in plant pathogenesis or insect-vector symbiosis and aid in the development of molecular-based approaches for possible disease intervention.
Ph. D.

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BEJI, AMOR. "Individualisation et definition de trois nouvelles especes regroupees anterieurement dans le complexe erwinia herbicola-enterobacter agglomerans". Lille 2, 1989. http://www.theses.fr/1989LIL2P265.

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9

Moreno, González M. del Carmen. "Characterization and mechanism of action of the biological control agent Pantoea agglomerans EPS125". Doctoral thesis, Universitat de Girona, 2006. http://hdl.handle.net/10803/7796.

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La soca EPS125 ha mostrat ser un efectiu agent de control biològic de diferents patògens fúngics de postcollita en diferents fruits. Degut a la seva elevada eficàcia, es va plantejar desenvolupar aquesta soca comercialment i per aquest motiu en el present treball es plantejà complementar la informació necessària pel seu registre.
D'acord amb els resultats obtinguts mitjançant proves fenotípiques i genotípiques, la soca EPS125 queda inclosa dins l'espècie Pantoea agglomerans (Enterobacter agglomerans-Erwinia herbicola). En relació a la utilització de fonts de carboni, en el perfil i contingut d'àcids grassos cel·lulars i en el polimorfisme en la longitud dels fragments de macrorestricció genòmica (MRFLP), la soca EPS125 mostrà trets característics que la diferencien d'altres soques. Els dos marcadors moleculars (125.2 i 125.3) específics per la soca EPS125 dissenyats en el present treball mostraren ser semiespecífics per la seva detecció mitjançant la tècnica PCR i Real Time PCR. Quedant pendent l'anàlisi d'especificitat de l'ús combinat dels dos marcadors moleculars en una reacció PCR multiplex. P. agglomerans EPS125 ha mostrat ser molt efectiva en el control de Penicillium expansum en poma amb una dosi efectiva mitjana de 2.7x105 a 7x105 ufc/ml, i una ratio de 25-101 cèl·lules de la soca EPS125 per inactivar una espora del patogen segons el model de saturació hiperbòlica. Segons les aproximacions fenotípiques i estudis genotípics realitzats, sembla que els mecanismes de biocontrol utilitzats per la soca EPS125 contra P. expansum en poma estan directament relacionats amb la capacitat de formació de biofilm per aquesta soca.
Strain EPS125 has shown effectiveness against a wide range of fungal pathogens in a large variety of fruit. However, to develop this strain as commercial biopesticide an extensive characterization is essential. For this reason, the objective of this PhD thesis was to complete the necessary information for its future registration.
According to morphological and biochemical tests, strain EPS125 pertain to Pantoea agglomerans (Enterobacter agglomerans-Erwinia herbicola) species. This strain showed typical traits different from other bacteria in relation to the ability to use several carbon sources, the fatty acid profiles and the macrorestriction fragment length polymorphism (MRFLP) pattern. The two DNA molecular markers of P. agglomerans EPS125 (125.2 and 125.3) obtained in the present work were semispecific in the detection of strain EPS125 by means of PCR and Real Time PCR. However, the combined use of the two primer sets in a multiplex PCR reaction would be specific. P. agglomerans EPS125 was highly effective against P. expansum in apple fruit having a median effective dose from 2.7x105 to 7x105 cfu/ml and a ratio of 101 and 25 EPS125 cells to inactivate one pathogen spore according to the hyperbolic saturation model.
Biocontrol mechanisms used by P. agglomerans EPS125 against P. expansum in apple fruit may be related with the ability of biofilm formation by this strain as show phenotypic approaches and genotypic studies.

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Kini, Kossi. "Pantoea spp : une nouvelle menace bactérienne pour la production rizicole en Afrique subsaharienne". Thesis, Montpellier, 2018. http://www.theses.fr/2018MONTG015.

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Parmi les 24 espèces de Pantoea décrites à nos jours, cinq ont été signalées jusqu'à 46 fois dans 21 pays comme phytopathogènes d'au moins 31 cultures. En effet, P. ananatis et P. agglomerans ont été signalés comme bactéries phytopathogènes pour au moins dix cultures économiquement importantes, y compris le riz. Récemment, le Centre du riz pour l'Afrique et ses partenaires ont soupçonné la présence d'une bactérie émergente qui provoque le flétrissement bactérienne du riz dans plusieurs pays africains, et l'agent causal a été confirmé comme appartenant au genre Pantoea. Les objectifs de notre projet de thèse étaient (i) d'améliorer la collection d'isolats d’AfricaRice existante par de nouvelles collections (ii) de développer des outils de diagnostic et de caractérisations pour une analyse fine de la diversité génétique, phénotypique et des études d’épidémio-suivaillance. Nos résultats ont montré que les bactéries capables de produire des symptômes de flétrissement bactérien du riz en Afrique forment un complexe d'espèces composé principalement de P. ananatis, P. stewartii et P. agglomerans. Différents types d'outils de diagnostic et de caractérisations ont ensuite été développés et validés. Les résultats de l'utilisation de ces outils ont permis de mettre en évidence la présence de ce complexe bactérien dans plusieurs pays Africains et de fournir des détails sur sa repartition géographique. Ainsi, au total, nous avons diagnostiqué un complexe d'espèces bactériennes, phytopathogènes du riz dans 11 pays africains (Bénin, Burkina Faso, Burundi, Ghana, Côte d'Ivoire, Mali, Niger, Nigeria, Sénégal, Tanzanie, Togo). En outre, l'analyse de trois génomes de P. ananatis Africain isolé du riz, et le développement, l'évaluation et l'application d'outils d'analyse VNTR à locus multiples (MLVA) ont permis de mieux comprendre les relations phylogénétiques et phylogénomiques existant entre les souches de P. ananatis isolées du riz et des souches d' autres sources (plantes, animaux et environnement). En effet, les résultats préliminaires ont montré que plusieurs souches de P. ananatis isolées du riz en Afrique, en Asie et en Europe étaient phylogénétiquement liées et formaient un groupe qui les différenciait de P. ananatis d'autres sources. En conclusion, les résultats de ce projet de thèse fournissent une base solide qui facilitera les futures études de Pantoea spp en Afrique
Among the 24 species of Pantoea described so far, five have been reported up to 46 times in 21 countries as phytopathogens of at least 31 crops. Indeed, P. ananatis and P. agglomerans have been reported as phytopathogenic bacteria for at least ten economically important crops, including rice. Recently, Africa Rice Center and its partners have suspected the presence of an emerging bacterium that causes rice bacterial blight in several African countries, and the causal agent has been confirmed as belonging to the genus Pantoea. The objectives of our thesis project were (i) to improve the collection of existing AfricaRice isolates by new collections (ii) to develop diagnostic and characterization tools for fine analysis of genetic, phenotypic and epidemio-follow-up studies. Our results showed that bacteria capable of producing bacterial blight symptoms of rice in Africa form a species complex composed mainly of P. ananatis, P. stewartii and P. agglomerans. Different types of diagnostic tools and characterizations were then developed and validated. The results from the use of these tools helped to point out the presence of this bacterial complex in several African countries and to provide details on its geographical structure. Thus, in total, we diagnosed a bacterial species complex, phytopathogenic of rice in 11 African countries (Benin, Burkina Faso, Burundi, Ghana, Ivory Coast, Mali, Niger, Nigeria, Senegal, Tanzania, Togo). In addition, analyzes of three genomes of african P. ananatis and the development, evaluation, and application of Multiple Locus VNTR Analysis (MLVA) tools provided insights into the phylogenetic and phylogenomic relationships that exist between P. ananatis strains isolated from rice and strains from other sources (plants, animals and environment). Indeed, preliminary results showed that several strains of P. ananatis isolated from rice in Africa, Asia and Europe were phylogenetically linked and formed a group that differentiated them from P. ananatis from other sources. In conclusion, the results of this thesis project provide a solid foundation that will facilitate future studies of Pantoea spp in Africa

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Prione, Lilian Parra. "Adaptação bacteriana: plasticidade fenotípica de Pantoea ananatis CCT 7673 exposta ao herbicida mesotrione". UNIVERSIDADE ESTADUAL DE PONTA GROSSA, 2014. http://tede2.uepg.br/jspui/handle/prefix/979.

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Made available in DSpace on 2017-07-21T20:00:00Z (GMT). No. of bitstreams: 1 Lilian Prione.pdf: 1165937 bytes, checksum: d73d6916708640fa9af6db139db8b8c2 (MD5) Previous issue date: 2014-02-07
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Herbicides are widely used to increase crop production and account for 35% of all agrochemicals applied annually. After application, the herbicides can remain in the soil as hazardous residues. Mesotrione, (2-[4-methylsulfonyl-2-nitrobenzoyl]1,3-cyclohenanedione), is the active ingredient of Callisto, an herbicide commonly used in corn. Pantoea ananatis CCT 7673, an Enterobacteria isolated from water, has been previously cited as mesotrione-degrading strain. The aim of this study was to evaluate the influence of mesotrione and Callisto as oxidative stress-inducing agents for cellular metabolism of Pantoea ananatis CCT7673 and identify possible mechanisms of tolerance. SOD, CAT and GR activities were evaluated in non-denaturing PAGE and CAT, GR and GST in spectrophotometer. Also, the rates of malonaldehyde (MDA), superoxid and peroxide hydrogen (H2O2) were measured in a spectrophotometer. Minimal medium with no herbicide (MM) was used as control. Lipid peroxidation, superoxide and peroxide hydrogen quantification and SOD, CAT, GR and GST activities were analyzed before and after degradation of mesotrione. The herbicide proved to be the cause of oxidative stress, according to peroxide hydrogen data. Unexpectedly, the rates of lipid peroxidation (MDA) and GR showed to be lower in the presence of the herbicide when compared to the control, with no changes in bacterial growth. The activity of GST was higher in mesotrione treatment in comparison to control and Callisto, during and after degradation. These results suggest that this enzyme may be related to the mesotrione degradation, probably by cometabolism. The rates of lipid peroxidation were shown to be lower in the presence of the herbicide compared to the control, with no changes in growth rates when exposed to herbicide. P. ananatis CCT 7673 showed changes in the saturation of membrane lipids. These changes may interfere with herbicide entry into the cell. These characteristics may be associated with a level of phenotypic plasticity in P. ananatis CCT 7673, making this an interesting bacterium for studies of herbicide tolerance and evolution of microbiota in environments subjected to different degrees of selective pressure model.
Herbicidas são amplamente utilizados para aumentar a produção agrícola e são responsáveis por 35% de todos os agrotóxicos aplicados anualmente. Após a aplicação, os herbicidas podem permanecer no solo como resíduos perigosos. O mesotrione (2 - [ 4 - metilsulfonil - 2 - nitrobenzoil ] 1,3- cyclohenanedione ) é o ingrediente ativo de Callisto, um herbicida utilizado no milho . Pantoea ananatis CCT 7673, é uma enterobactéria isolada de água e foi previamente caracterizada como linhagem degradadora do mesotrione. Os objetivos deste trabalho foram o avaliar a influência do mesotrione e Callisto como agentes indutores de estresse oxidativo para o metabolismo celular de Pantoea ananatis CCT7673, bem como identificar possíveis mecanismos de tolerância a estes herbicidas. As atividades enzimáticas de SOD, CAT e GR foram avaliadas em PAGE não desnaturante e de CAT, GR e GST em espectrofotômetro. Além disto, as taxas de malondialdeído (MDA), radical superóxido e peróxido de hidrogênio (H2O2) foram medidas em espectrofotômetro. Todas as análises foram realizadas antes e após a degradação do mesotrione. Meio mínimo sem herbicida (MM) foi utilizado como controle. O herbicida provou ser agente causal do estresse oxidativo pelos dados de peróxido de hidrogênio. Inesperadamente, as taxas de MDA e GR mostraram-se inferiores nos tratamentos com herbicida em relação ao controle, sem alteração no crecimento bacteriano. A atividade de glutationa-S-transferase foi maior no tratamento com mesotrione em comparação com o controle e Callisto, durante e após a degradação. Estes resultados sugerem que essa enzima pode estar relacionada com o processo de degradação do mesotrione, provavelmente por cometabolismo. As taxas de peroxidação lipídica mostraram ser menores na presença do herbicida em comparação com o controle, sem mudanças na taxa de crescimento Quando exposta ao herbicida, P. ananatis CCT 7673 apresentou alterações na saturação de lipídios de membrana. Estas mudanças podem interferir na entrada do herbicida na célula. Tais características podem estar associadas a um nível de plasticidade fenotípica em P. ananatis CCT 7673, tornando essa bactéria um modelo interessante para estudos de tolerância a herbicidas e evolução de microbiotas em ambientes submetidos a diferentes graus de pressão seletiva.

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Shin, Giyo Yoon. "Functional characterisation of Hfq and identification of Hfq-dependent sRNAs in Pantoea ananatis". Thesis, University of Pretoria, 2020. http://hdl.handle.net/2263/77882.

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The trans-encoded small RNAs (sRNAs) are novel gene expression regulators in bacteria. In response to external stimuli, sRNAs rapidly modulate the expression of genes at the post-transcriptional level to allow bacteria to reprogram its cellular environment for survival and fitness. These regulatory actions of sRNAs are chaperoned by the indispensable RNA-binding protein, Hfq, that stabilizes sRNAs and facilitates the base-pairing between the sRNA and target mRNA. The functional role of this protein in a broad-host-range phytopathogen P. ananatis was determined by constructing an hfq-null mutant strain of P. ananatis LMG 2665T. Overall, deletion of the hfq gene in P. ananatis had a negative effect on motility, biofilm formation, production of quorum sensing autoinducer molecule and pathogenicity of the bacterium, suggesting a collective involvement of Hfq in above-mentioned physiological processes. To identify trans-encoded sRNAs that are dependent on Hfq, strand-specific sRNA sequencing was conducted on total RNA extracted from both the wild-type and hfq mutant strains of P. ananatis at low (OD600 = 0.2) and high (OD600 = 0.6) cell-density conditions. The resulting sRNA transcriptome data were computationally screened for putative sRNAs by applying known sRNA characteristics. The filtering yielded 615 putative sRNAs (302 antisense sRNAs, 249 intergenic sRNAs and 64 gene-overlapping sRNAs). Of these sRNAs, 276 candidate sRNAs were differentially regulated by the loss of hfq (low cell condition = 58, high cell condition = 154 and both cell conditions = 64). Among 276 differentially regulated sRNA candidates, 41 were positive for Rho-independent terminator sequences, which included enterobacteria-conserved Hfq-dependent sRNAs such as ArcZ, FnrS, GlmZ, RprA, RyeB, RyhB, RyhB2, Spot42, SsrA as well as 16 novel P. ananatis sRNAs. Of the 41, the expression profile of nine sRNAs determined by sequencing depth plots and quantitative PCR (qRT-PCR) showed that the abundances of these sRNAs were negatively affected by the absence of the P. ananatis hfq gene, supporting their dependency on Hfq. In addition, computationally predicted target genes of selected sRNAs, whose full-length sequence was determined by 5’ rapid amplification of cDNA ends (5’ RACE) analysis, included those involved in the synthesis of exopolysaccharide, cell wall degrading enzyme and type 6 secretion system. The result suggests possible sRNA-regulation of virulence-related genes. Future works will focus on determining the functional roles of sRNAs and experimentally validating the predicted targets of these sRNAs. Overall, the findings of the current work highlight the importance of Hfq as a global virulence regulator in P. ananatis and identified previously uncharacterized sRNAs that possibly play essential roles in regulating bacterium’s adaptive responses.
Thesis (PhD)--University of Pretoria, 2020.
Microbiology and Plant Pathology
PhD
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13

Pennerman, Kayla Kara. "Development of Methods for Structural Characterization of Pantoea stewartii Quorum-Sensing Regulator EsaR". Thesis, Virginia Tech, 2014. http://hdl.handle.net/10919/25297.

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The LuxR family of proteins serves as quorum-sensing transcriptional regulators in proteobacteria. At high population densities, a small acyl-homoserine lactone (AHL) molecule, produced by a LuxI homologue, accumulates in the environment. The LuxR proteins bind to their respective AHL when the ligand accumulates to sufficient levels. Once bound to AHL, the holoproteins usually become functional as transcriptional activators. However, there is a subset of LuxR homologues, the EsaR subfamily, which is active without the AHL ligand and becomes inactivated once bound to it. EsaR is the best understood member of this subfamily. It controls virulence in the corn pathogen Pantoea stewartii ssp. stewartii. Solubility issues have previously limited structural studies of LuxR homologues as the proteins could not be purified without the AHL ligand. A soluble recombinant EsaR protein, HMGE, is biologically active and can be purified in the absence and presence of AHL, unlike most other LuxR homologues. Using HMGE, amino acid substitutions and Förster resonance energy transfer (FRET), experimental methods were designed for determining the dimerization interface of EsaR and for testing the hypothesis that EsaR undergoes a conformational shift when presented with the AHL ligand. To identify residues of the dimerization interface, heterodimerization assays were designed, involving either coexpression or coincubation of wild-type EsaR and variant HMGE proteins. In this assay, the inability of the proteins to copurify by nickel affinity chromatography would indicate that the modified residue(s) are important for dimerization of EsaR. To determine the conformational change that EsaR undergoes when bound to the AHL ligand, a FRET assay was developed to estimate the distances between amino acid residues in the absence and presence of AHL. Future work will have to include a few modifications to the methods and/or control experiments. This study provides the basis upon which the present methods can be further developed and later used for structural studies of EsaR.
Master of Science

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14

De, Maayer Pieter. "Genome comparisons to identify selected pathogenicity factors of a plant-associated Pantoea ananatis strain". Thesis, University of Pretoria, 2010. http://hdl.handle.net/2263/30849.

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Pantoea ananatis is a ubiquitous organism found in almost every environment on earth. It has been implicated in diseases of a wide range of agronomic crops worldwide, including onion, maize, rice and pineapple, as well as a human disease. In South Africa, P. ananatis causes blight and dieback of Eucalyptus, resulting in severe losses of this important forestry resource. Nevertheless, little is known about the pathogenicity mechanisms utilised by this pathogen to cause disease in this host. The whole genome of a highly virulent Eucalyptus-pathogenic P. ananatis strain, LMG20103, was sequenced. This genome sequence was subsequently mined to identify a vast array of genes encoding putative pathogenicity determinants. Comparative genomics revealed that it has evolved to be able to thrive in a wide range of environments and that this strain carries pathogenicity determinants that may allow it to infect hosts in both the animal and plant Kingdom. Interestingly, no Type II and III secretion systems, which form a major part of the pathogenicity arsenal of many plant pathogenic bacteria are present in P. ananatis. However, three loci on the genome encode three distinct copies of the Type VI secretion system, which has recently been demonstrated to play an important role in diseases caused by many plant- and animal-pathogenic bacteria. In silico analysis of these secretion systems showed that they likely secrete several pathogenicity effectors which may have a role in P. ananatis infection of both plant and animal hosts. Another putative pathogenicity determinant identified from the genome, the exopolysaccharide ananatan, was experimentally demonstrated to play a role in disease expression on both onion seedlings and pineapple fruit. This was done through the production of a library of mutants which encompasses all the genes on the P. ananatis genome. Genome sequencing enabled the identification of all the putative pathogenicity factors of P. ananatis LMG20103 and the use of the mutant library and post-genomic techniques has and will allow the functional characterization of many of these pathogenicity determinants. By this means, the mechanisms underlying the disease caused by P. ananatis on Eucalyptus and other hosts can be better understood. With this information, more directed and effective strategies for the control of this pathogen and its diseases can be developed.
Thesis (PhD)--University of Pretoria, 2010.
Microbiology and Plant Pathology
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15

Gonçalves, Ricardo Marcelo. "Estudos etiológicos da mancha branca do milho e identificação de hospedeiros alternativos de Pantoea ananatis". Universidade Estadual de Londrina. Centro de Ciências Agrárias. Programa de Pós-Graduação em Agronomia, 2012. http://www.bibliotecadigital.uel.br/document/?code=vtls000175423.

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A doença mancha branca do milho (MBM) é alvo de divergências com relação à sua etiologia. Alguns autores descrevem o fungo Phaeospharia maydis (forma anamórfica Phyllosticta sp, sinonímia Phoma sp.) como agente etiológico da doença, outros a bactéria Pantoea ananatis e ainda há autores que relatam a existência de diferentes agentes causais da doença, como Phyllosticta sp., o Phoma sorghina e a Sporormiella sp., e que dependendo do local, um ou outro fungo poderia ser o agente causal da doença. Desta forma, um dos objetivos deste trabalho foi determinar a etiologia da doença e avaliar a possibilidade de duas espécies de gramíneas do gênero Digitaria serem hospedeiros alternativos de P. ananatis. Plantas de milho híbrido HS200 cultivadas em casa de vegetação (CV) foram inoculadas com P. ananatis 45 dias após a semeadura. Sintomas típicos da doença MBM foram observados 10 dias após a inoculação. As plantas permaneceram na CV até as lesões evoluírem para o estádio necrótico quando procedeu-se a coleta das folhas contendo as lesões. Também foram coletadas à campo, folhas contendo lesões necróticas resultantes de infecção natural. Muitas das lesões necróticas, tanto aquelas obtidas por infecção natural, como aquelas obtidas por inoculação em CV apresentaram estruturas fúngicas em seu interior. As folhas foram lavadas e segmentos foliares apresentando lesões com estruturas fúngicas foram acondicionados em câmara úmida. Os fungos foram isolados, purificados e a técnica da PCR foi usada para amplificar o espaço interno transcrito (ITS) do rDNA. Os amplicons foram sequenciados e a identificação dos isolados foi realizada pelo grau de similaridade das sequências com àquelas depositadas no GenBank. Os fungos isolados tanto de lesões necróticas obtidas pela inoculação de P. ananatis em CV como aquelas coletadas à campo foram identificados como pertencentes às espécies Epicoccum nigrum, Leptosphaerulina chartarum, Fusarium chlamydosporum, Alternaria alternata, Alternaria ricini, Fusarium equiseti, Gibberella moniliformis (Fusarium moniliforme), Curvularia sp., Phoma sp. (sinonímia de várias espécies de Phyllosticta sp.) e Phoma sorghina. Para fornecer maior subsídio com relação à etiologia da MBM, também foi realizado o monitoramento das lesões em diferentes estádios de desenvolvimento. Para tanto, lesões de MBM obtidas por meio de infecção natural foram coletadas a campo, classificadas em quatro estádios diferentes de desenvolvimento, de acordo com o progresso da lesão, e o DNA total extraído de um pool de lesões para cada estádio. Após extração, o DNA foi amplificado por PCR com primers específicos para P. ananatis (ANAF/ANAR) e primers universais para fungos (ITS). P. ananatis foi identificada em todos os estádios de desenvolvimento das lesões, enquanto que fungos foram detectados somente nos tecidos necróticos das lesões. Os experimentos comprovaram que a bactéria P. ananatis é o agente causal da MBM, e que os fungos instalam-se nas lesões preestabelecidas pela bactéria. A ocorrência de hospedeiros alternativos de P. ananatis foi analisada por meio de inoculações cruzadas realizadas em casa de vegetação entre os isolados: WT-2 (P. ananatis controle positivo, isolado da MBM); Pa2DH (isolado obtido de lesões de MBM, o qual foi inoculado e reisolado a partir de lesões de Digitaria horizontalis) e Pa1DI (isolado de Digitaria insularis). A partir dos sintomas obtidos, os reisolados foram empregados em sucessivas inoculações. Sintomas da doença foram observados em casa de vegetação 10 e 30 dias após as inoculações em plantas de Digitaria spp e milho, respectivamente. A bactéria foi reisolada após os testes de patogenicidade e todos os reisolados foram identificados como P. ananatis por meio do sequenciamento parcial do gene 16S rDNA e dos amplicons com os primers específicos ANAF/ANAR.
The maize white spot (MWS) disease is largely studed due to the divergence related to its etiology. Some researchers describe the fungus Phaeospharia maydis (anamorphic form of Phyllosticta sp., synonymy Phoma sp.) as the causal agent, and some the bacteria Pantoea ananatis and yet another researchers who report the existence of different causative agents of disease, Phyllosticta sp., Phoma sorghina and Sporormiella sp., that depending on location, either fungus could be the causal agent of disease. By this mean one of the goals of this work was to confirm the etiology of MWS. Another goal of this work was to evaluate the possibility of two grass species from Digitaria genus to be optional hosts for P. ananatis. Hybrid maize plants HS200 were grown in a greenhouse and inoculated with P. ananatis 45 days after planting. Characteristic symptons were observed 10 days after the inoculation. The plants were held at the greenhouse until the injuries have developed to the necrotic stage, when the leaves with the injuries were collected. Leaves were also collected at field condition, containing necrotic injuries, caused by natural infection. Many of the necrotic injuries, no matter if obtained by natural infection or by the inoculation at the green house, have shown fungi structures in its interior. The leaves were washed and sections showing injuries with central fungi structures were packed into a moist chaimber. The fungi were then isolates, purified and the PCR technique was used to amplify the internal transcribed spacer (ITS) of rDNA. The amplicons were sequenced and the identification of the fungi isolates was performed by the similarity degree of the sequencies with those deposited at GenBank. The fungi isolates, both from the necrotic injuries acquired by the inoculation of P. ananatis in greenhouse and those collected at field conditions were identified as belonging to the species Epicoccum nigrum, Leptosphaerulina chartarum, Fusarium chlamydosporum, Alternaria alternata, Alternaria ricini, Fusarium equiseti, Gibberella moniliformis (Fusarium moniliforme), Curvularia sp., Phoma sp (synonymy of many species of Phyllosticta sp.) and Phoma sorghina. To provide more subsidy relationed to the etiology of MWS, also the monitoring of the injuries in diferente degrees of development. For this, injuries of MWS were obtained at field condition by natural infection, classified in four different development stages, according to the injury development, and the total DNA was extracted from an injury pool for each stage. After the extraction, the DNA was amplified by a PCR with specific primers for P. ananatis (ANAF/ANAR) and the universal primers for fungi (ITS). P. ananatis is present in every stage of the injury development, when the fungi were detected after the injury tissue becomes necrotic. So these experiments proved to be the bacterium P. ananatis the causal agent of MWS, and fungi install itself in the injures pre-established by the P. ananatis bacteria. The incidence of optional hosts for P. ananatis was analised through crossed inoculations, taken in a greenhouse between the isolates: WT-2 (P. ananatis positive control, isolated from MWS), Pa2DH (isolate taken from MWS injuries, which was inoculated and reisolated from Digitaria horizontalis injuries) and Pa1DI (isolated from Digitaria insularis). From the gotten symptons, the reisolate were used in subsequent inoculations. Disease symptons were observed in greenhouse condition between 10 and 30 days after inoculation in Digitaria and maize plants, respectively. The bacteria was reisolated after pathogenicity tests and all of the reisolated were identified as P. ananatis by partial sequencing of gene 16S rDNA and from the amplicons with specific primes ANAF/ANAR.

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16

Schu, Daniel Joseph. "Structure/Function Analysis of the Quorum-sensing Regulator EsaR from the Plant Pathogen Pantoea stewartii". Diss., Virginia Tech, 2009. http://hdl.handle.net/10919/38807.

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Pantoea stewartii subsp. stewarti is the causative agent of Stewartâ s wilt disease in maize. Disease symptoms develop after the bacteria grow to high cell densities in the plant xylem and secrete an abundance of exopolysaccharide (EPS). EPS production is regulated by quorum sensing. Two regulatory proteins are key to the process of quorum sensing, the LuxI and LuxR homologues EsaI and EsaR. Most LuxR homologues function as activators of transcription in the presence of their cognate acylated homoserine lactone signal (AHL). EsaR utilizes an AHL-response opposite of the majority of the LuxR homologues. EsaR represses EPS production at low cell densities. However, at high cell densities when high concentrations of AHL are present, EsaR is inactivated and derepression of EPS production occurs. The mechanism that enables EsaR to respond to AHL in a manner opposite to that of most LuxR homologues remains elusive. A comparative study of EsaR and the well characterized quorum-sensing regulators LuxR from Vibrio fischeri and TraR from Agrobacterium tumefaciens was initiated. Previous studies demonstrated that in the absence of AHL, EsaR retains the ability to function as a weak activator of the lux operon in recombinant Escherichia coli. This thesis research further characterized the role of EsaR as an activator. Variant forms of EsaR with deletions or single residue substitutions were generated and their ability to regulate transcription was examined in vivo. Furthermore, a native EsaR-activated promoter has been identified, which controls expression of a putative regulatory sRNA in P. stewartii. It is apparent that EsaR functions as a transcription factor at low concentrations of AHL as demonstrated by its ability to inhibit EPS production. At high concentrations, the AHL appears to bind and cause a conformational shift in the protein leading to its inactivation. The second goal of this study was to further elucidate the mechanism by which AHL regulates EsaR. Pulse-chase experiments demonstrated that EsaR is resistant to proteases with or without AHL in vivo. Limited proteolytic digestions in vitro suggest that the protein does undergo conformational changes in response to AHL. Gel filtration chromatography, sucrose gradient ultracentrifugation, and cross-linking experiments proved that this conformational change does not impact the multimeric state of EsaR. To better understand the mechanism of regulation by AHL, the final goal of this project was to examine the interactions which result in EsaR-responsiveness to AHL. Several individual amino acid substitutions were identified that cause EsaR to function in an AHL-independent manner, by which variants retain the ability to bind and block gene expression in the presence of AHL. These residues have been mapped onto a homology model of EsaR and their role has been examined in vitro. The ability of these EsaR* variants to bind AHL and an analysis of the effects individual mutations have on the overall conformation of the protein was performed. Overall this study has revealed several unique aspects of the quorum-sensing system in P. stewartii whereby gene expression is regulated at both low and high cell density. Studies were also initiated to examine the mechanism of AHL-responsiveness of EsaR. The mechanism by which AHL modulates most LuxR homologues remains elusive. The ability to purify EsaR +/- its cognate AHL may prove critical in elucidating this mechanism.
Ph. D.

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17

Burke, Alison Kernell. "Analysis of the Quorum Sensing Regulons of Vibrio parahaemolyticus BB22 and Pantoea stewartii subspecies stewartii". Diss., Virginia Tech, 2015. http://hdl.handle.net/10919/77872.

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Quorum sensing is utilized by many different proteobacteria, including the two studied for this dissertation work, Vibrio parahaemolyticus and Pantoea stewartii subsp. stewartii. V. parahaemolyticus causes acute gastroenteritis in people who eat contaminated raw or undercooked shellfish. It is found in warmer marine waters and in rare cases, causes systemic infections when bacteria enter the body through open wounds. P. stewartii, on the other hand, is a phytopathogen that causes Stewart's wilt in maize. It is found in soil or the mid-gut of the corn flea beetle, its insect vector. Both V. parahaemolyticus and P. stewartii utilize quorum sensing to control their pathogenicity. Quorum sensing enables coordinate gene expression across a bacterial population. The V. parahaemolyticus quorum-sensing system utilizes the master regulator OpaR, which is homologous to the V. harveyii LuxRVh and the P. stewartii system contains EsaR which is homologous to the V. fischeri LuxRVf regulator. While the two systems differ in the molecular details of their mechanistic control, they are both forms of cell density dependent regulation that are either directly or indirectly controlled by small signaling molecules. Three different signaling molecules are found in V. parahaemolyticus, and only one signal is used in P. stewartii. The focus of this dissertation has been on understanding the downstream targets of OpaR and EsaR in their respective quorum-sensing systems. Prior to this work, it was known that when OpaR is not present or is nonfunctional V. parahaemolyticus changes from an opaque to a translucent colony morphology phenotype and the cells also become swarm proficient and more pathogenic. The complete genome of the V. parahaemolyticus BB22OP strain was assembled and annotated (Chapter 2). RNA-Seq was then used to analyze the transcriptomes of OpaR-active and OpaR-deficient strains of V. parahaemolyticus and identify genes that were regulated via quorum sensing (Chapter 3). Similarly, P. stewartii was also analyzed using RNA-Seq to identify genes controlled by EsaR in the transcriptome that had not been detected through prior proteomic studies. The initial RNA-Seq work confirmed the control of some previously identified direct targets of EsaR and newly identified ten other genes also directly controlled by EsaR (Chapter 4). Two direct targets of EsaR, rcsA and lrhA, became the focus of additional studies to further define the hierarchy of gene control downstream of the quorum-sensing regulator EsaR. RcsA controls capsule production, while LrhA controls motility and adhesion in P. stewartii. The regulons of rcsA and lrhA were defined by RNA-Seq, which also revealed multi-level control of rcsA gene expression (Chapter 5). Tight coordinated and temporal control of virulence factors is important for successful disease progression by pathogens. This dissertation work aims to enable a better understanding of the quorum-sensing hierarchy of genetic control in V. parahaemolyticus and P. stewartii.
Ph. D.

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18

Duong, An Duy. "Investigation of Pantoea stewartii Quorum-Sensing Controlled Regulators and Genes Important for Infection of Corn". Diss., Virginia Tech, 2018. http://hdl.handle.net/10919/93208.

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Bacteria interact with their eukaryotic hosts using a variety of mechanisms that range from being beneficial to detrimental. This dissertation focuses on Pantoea stewartii subspecies stewartii (P. stewartii), an endosymbiont in the corn flea beetle gut that causes Stewart's wilt disease in corn. Gaining insights into the interactions occurring between this bacterial pathogen and its plant host may lead to informed intervention strategies. This phytopathogen uses quorum sensing (QS) to coordinate cell density-dependent gene expression and successfully colonize corn leading to wilt disease. Prior to the research presented in this dissertation, the QS master regulator EsaR was shown to regulate two major virulence factors of P. stewartii, capsule production and surface motility. However, the function and integration of EsaR downstream targets in P. stewartii were still largely undefined. Moreover, only a draft genome of a reference strain of P. stewartii was publicly available for researchers, limiting bioinformatics and genome-scale genetic approaches with the organism. The work described in this dissertation has now addressed these important issues. The function of two EsaR direct targets, LrhA and RcsA, was explored (Chapter Two) and the existence of integration in the regulation between them was discovered (Chapters Two and Four). RcsA and LrhA are transcription factors controlling capsule production and surface motility in P. stewartii, respectively. In Chapter Two, the RcsA and LrhA regulons were investigated using RNA-Seq. This led to the discovery of a potential regulatory interaction between them that was confirmed by qRT-PCR and transcriptional gene fusion assays. The involvement of LrhA in surface motility and virulence was also established in this project. A direct interaction between LrhA and promoter of rcsA was defined in Chapter Four. Additional direct regulatory targets of LrhA were also identified. A project to generate a complete assembly of the P. stewartii genome (Chapter Three) enabled more thorough genome-wide analysis and revealed the existence of a previous unknown 66-kb region in the P. stewartii genome believed to contain genes important for motility and virulence. In addition, completion of the genome sequence permitted genes for two distinctive Type III secretion systems, used for interactions with corn or the corn flea beetle, to be placed on two mega-plasmids. Furthermore, the complete genome sequence facilitated a Tn-Seq approach (Chapter Five). Tn-Seq is a potent tool used to identify bacterial genes required for certain environmental test conditions. This project is a pioneering utilization of a Tn-Seq analysis in planta to investigate genes important for colonization and survival of P. stewartii within its corn host. It was discovered that OmpC and Lon are important to in planta growth and OmpA plays a role in plant virulence. In conclusion, these studies have broadened our understanding about the role of the QS regulon and other genes important for the pathogenesis of this phytopathogen. This knowledge may now be applied toward the development of future disease intervention strategies against P. stewartii and other wilt-disease causing plant pathogens.
PHD

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19

Gentzel, Irene Nichole. "Water-Soaked Symptoms in Maize as a Response to the Pathogen Pantoea stewartii ". The Ohio State University, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=osu1565453865973106.

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20

Panta, Utsab R., James A. Joslyn e Rupal D. Shah. "Pantoea agglomerans bacteremia: A rare case of spontaneous human infection by a plant pathogen in an immunocompromised host". Digital Commons @ East Tennessee State University, 2018. https://dc.etsu.edu/asrf/2018/schedule/170.

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Introduction: Pantoea agglomerans is a Gram negative ubiquitous bacteria commonly isolated from plant surfaces, seeds, fruits and animal/human feces usually introduced to human by ingestion of infected fruits/vegetables, thorn pricks and gastrointestinal translocation in lack of stomach acidity. However, the pathogen can also cause opportunistic human infection especially when the immune system is impaired. The aim of this case report is to investigate clinical features in a patient with P. agglomerans bacteremia and bring attention the opportunistic infection by this rare bacteria. Case presentation: We present a case of 57 year old caucasian lady with past medical history of Chronic Obstructive Pulmonary Disease, Atrial fibrillation, Immunoglobulin (IgG) deficiency, recurrent pneumonia, urine infection, oral/vaginal candidiasis, Gastro-esophageal reflux disease who presents with one week history of increased shortness of breath, chest tightness and productive cough without fever/chills. She also had high INR of 4.7 (target 2-3) despite taking normal dose of warfarin. She denies plant exposure. Her vitals were stable, saturation maintained with oxygen supplementation. Chest exam revealed very poor air entry bilaterally suggesting exacerbation of COPD. Oral thrush was present. Recent IgG level within last 6 months was low. Blood culture grew Pantoea agglomerans, pan-sensitive to most of the antibiotics. Chest X ray, CT scan abdomen and urine studies could not localize the source of infection. She was treated with Ceftriaxone, INR normalized to therapeutic range and she improved to baseline after 10 days of treatment. Discussion and conclusion: P. agglomerans is a rare cause of bacteremia which usually presents as fever, chills and general toxicity, however could also present as a cause of exacerbation of chronic diseases. Spontaneous infection can occur in a immunocompromised host, however the pathogen is of low virulence. The link between upper GI symptoms along with antacid receipt and spontaneous P. agglomerans infection could be possible, however needs further study. Hence, P. agglomerans should be considered one of the possible cause of spontaneous bacteremia in a immunocompromised host.

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21

Koziski, Jessica Marie. "Genetic Analysis of the Quorum Sensing Regulator EsaR". Thesis, Virginia Tech, 2008. http://hdl.handle.net/10919/34229.

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Pantoea stewartii subsp. stewartii is the causative agent of Stewartâ s wilt disease in maize plants. The bacteria are injected into the plant by corn flea beetles during feeding. They colonize the xylem and overproduce a capsular exopolysaccharide (EPS) at high cell densities. The production of EPS is regulated by an EsaI/EsaR quorum sensing mechanism, homologous to the LuxI/R system. Although activation of the EPS encoding genes by EsaR occurs after it complexes to the AHL (3-oxo-C6-HSL), unlike the LuxI/R system, this activation occurs by a different mechanism. At low cell densities, dimerized EsaR acts as a repressor. At a high cell population, derepression of the EPS genes occurs via an unknown mechanism once the AHL complexes to EsaR. Hence, a random mutagenesis genetic approach to isolate EsaR* variants that are immune to the effects of AHL has been utilized. Error-prone PCR and site-directed mutagenesis were used to generate desired mutants, which were subsequently screened for their ability to repress transcription in the presence of AHL. Several individual amino acids playing a critical role in the AHL-insensitive phenotype have been identified and mapped onto a homology model of EsaR. A separate study attempted to localize the dimerization region and analyze the stability of the N-terminal domain of EsaR. Truncations of EsaR at amino acids 169 and 178, without and with the extended linker region respectively, were generated using PCR. Dimerization assays similar to those by Choi and Greenberg in 1991 were performed but proved to be unsuccessful. However, the N-terminal domain is stable as determined by western blotting, which may facilitate its future structural analysis. Together, these efforts have contributed to the molecular understanding of AHL-dependent derepression of EsaR.
Master of Science

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22

Heerman, Matthew C. "Bacterial infection, immune responses, and autophagy in lutzomyia longipalpis sand flies". Diss., Kansas State University, 2016. http://hdl.handle.net/2097/32902.

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Doctor of Philosophy
Department of Entomology
Marcelo Ramalho-Ortigao
Kun Yan Zhu
Microbial communities residing within the midgut of insect vectors play a critical role in the response to various zoonotic and human pathogens, and can directly alter the development and survival of the insects. Sand flies are the primary vector of Leishmania, the causative pathogen of leishmaniasis, a neglected tropical disease. Sand flies acquire many microbes from the soil where immature stages develop until emergence as adults. Gram-negative Pantoea agglomerans and gram-positive Bacillus subtilis are two bacteria commonly associated with sand fly populations. Here, I demonstrated that an EGFP- and a GFP-expressing version of these two bacteria localize to different compartments of the midgut; a phenomenon that is achieved, in part, to pH differences found across the length of the gut. Additionally, P. agglomerans is able to selectively induce midgut epithelial apoptosis while B. subtilis does not. This is accompanied by differential immune and homeostasis responses to both bacteria highlighted by immune pathway suppression via the Poor Immune Response upon Knock-in (Pirk) gene. These effects may actually be representative of a broader type of response to bacterial infection that might be present across several insect species. Finally, I demonstrated that during metamorphosis the sand fly relies, at least in part, upon the activation of multiple genes from the autophagy pathway to aid in generating adult tissues. More specifically, I demonstrate, using microscopy, the presence of ATG6 in the cytoplasm of developing midgut epithelial cells of the sand fly pupae.

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23

Lanza, Fabrício Eustáquio. "Mancha-branca do milho: etiologia e resistência de genótipos". Universidade Federal de Viçosa, 2009. http://locus.ufv.br/handle/123456789/4378.

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Conselho Nacional de Desenvolvimento Científico e Tecnológico
The main objectives of this work were to identify the causal agent of the maize White Spot, to obtain preliminary information on the disease dispersal in the field and to characterize the reaction of maize hybrids and inbreds to the disease. For the etiological studies, isolations were performed from white spot lesions the anasarca phase, resulting in the development of bacterial colonies. Maize hybrids BRS2022, BRS1010, 1D2195, BRS1040, BRS1035, BRS1031, BRS3025, BRS1030, 2B710 e P30F35 and inbred lines L3, L228-3, 521274, 521236 e 262841-1-4-1 were evaluated under natural epidemic in a randomized block design with three replications. Cultivars were planted in single row plots, separated by two rows of the resistant hybrid BRS1010. Spreader rows were formed by planting the susceptible genotype DAS657 0,5 m apart and in front of each block. Disease severity was evaluated at a weekly internal starting 60 days after planting, through a 1 to 9 scale of disease severity where 1= no disease and 9= 100% of leaf area affected. Ratings were taken at three different locations within each plot: 1, 2, 3, 4, 5 and 6 meters inoculum source. Data were used for the calculation of the area under disease progress curve (AUDPC), disease severity at 50% of epidemic development (Y50), disease severity at the end of the epidemic, and the rate of disease progress. Inoculations on the susceptible hybrid DAS657, in the greenhouse, reproduced the typical symptoms of the disease. Re-isolations from theses lesions confirmed the presence of the same bacteria isolated from the field, which identified as Pantoea ananatis, confirming previous reports on the involvement of this bacteria in the initial lesions of this disease. No disease gradient was observed based on the disease severity observed in each point of evaluation within each plot. A better distinction between the level of resistance of maize genotypes was obtained through AUDPC and Ymáx values. Maize hybrids BRS1030, BRS1035 and BRS1010 and inbreds L3, and L228-3 were the most resistant genotypes. These inbred lines may be useful in breeding programs for resistance to maize white spot.
Este trabalho objetivou confrimar o agente causal da mancha-branca do milho, obter informações preliminares sobre a dispersão do patógeno e caracterizar a reação de genótipos de milho a doença. Para o estudo etiológico, isolados foram obtidos de lesões de manchabranca em fase de anasarca, resultando em desenvolvimento de colônias bacterianas. Híbridos de milho, BRS2022, BRS1010, 1D2195, BRS1040, BRS1035, BRS1031, BRS3025, BRS1030, 2B710 e P30F35 e as linhagens L3, L228-3, 521274, 521236 e 262841-1-4-1 foram avaliados sob epidemia natural em delineamento de blocos ao acaso e três repetições. Os cultivares foram plantados em fileiras de cinco metros, separadas por uma linha do híbrido resistente BRS1010. A cortina suscetível (fonte de inóculo) formada pelo híbrido DAS657, foi plantada na parte frontal de cada bloco, afastada 0,5 m. A severidade da doença foi avaliada em intervalos semanais a partir dos 60 dias do plantio, utilizando uma escala de 1 a 9, onde: 1= sem doença e 9= 100% de área foliar afetada. As avaliações foram realizadas em 6 pontos dentro da parcela afastados 1, 2, 3, 4, 5 e 6 metros da fonte de inóculo. Os dados de severidade foram usados para o calculo da área abaixo da curva de progresso da doença (AACPD), severidade da doença na metade da epidemia (Y50), severidade da doença no final da epidemia (Ymáx), e taxa de progresso da doença. Com inoculações em híbrido suscetível DAS657, em casa de vegetação, foi possível reproduzir os sintomas típicos da doença. Reisolamento a partir dessas lesões confirmou a presença da mesma bactéria isolada do campo, identificada como Pantoea ananatis, corroborando relatos do envolvimento desta bactéria nos sintomas iniciais da doença. Não foi observada a formação de um gradiente de dispersão baseado na severidade da doença observada em cada ponto de avaliação dentro da parcela. A melhor distinção entre os níveis de resistência de genótipos de milho foi obtida pelos valores de AACPD e Ymáx. Os híbridos de milho BRS1030, BRS1035 e BRS1010 e as linhagens L3, e L228-3 foram os genótipos mais resistentes. Essas linhagens podem ser usadas em programas de melhoramento visando resistência a mancha-branca.

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Pandolfi, Valesca. "Análise transcricional do fitopatógeno Fusarium graminearum Schwabe na interação antagonista com a bactéria Pantoea agglomerans Gavini". Universidade de São Paulo, 2006. http://www.teses.usp.br/teses/disponiveis/64/64133/tde-04052007-085012/.

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Gramíneas cultivadas, como trigo, cevada e milho são produtos agrícolas de fundamental importância no Brasil. Entre os fatores causadores de perdas na produção de grãos dessas espécies estão os estresses causados por fitopatógenos como Fusarium graminearum Schwabe (teleomorfo Gibberella zeae Schw.), agente causador da fusariose e de difícil controle químico, biológico ou mesmo genético. Uma estratégia que tem se mostrado eficiente no controle de doenças é a utilização de microrganismos antagonistas a diferentes fitopatógenos, dentre os quais destaca-se a bactéria P. agglomerans. O presente trabalho teve como objetivo identificar genes diferencialmente expressos em interações fungo fitopatogênico-microrganismo antagonista, considerando como modelo o sistema F. graminearum-P. agglomerans. A construção de uma biblioteca de cDNA de F. graminearum cultivado in vitro proporcionou a geração de 1.983 seqüências válidas, resultando em 1.283 unigenes. As categorias de maior representatividade desta biblioteca foram aquelas constituídas por proteínas envolvidas em vias da informação genética - DNA-RNA-proteína (26 %); proteínas hipotéticas (24 %) e proteínas do metabolismo (16 %). Tanto a categoria de proteínas envolvidas nos processos de desenvolvimento como as envolvidas na percepção a estímulos externos constituíram 10 % dos unigenes. Dentre os genes presumivelmente anotados, foram identificados aqueles codificadores de enzimas de importantes rotas metabólicas como gliceraldeído-3-fosfato-desidrogenase, fosfoglicerato quinases e fosfoenolpiruvato carboxilases, como também componentes produzidos pelo metabolismo secundário como micotoxinas e outras proteínas associadas a estresse e patogenicidade de fungos. Neste trabalho também foi verificado o potencial de antagonismo in vitro da bactéria P. agglomerans frente a três fitopatógenos de trigo: Drechslera tritici-repentis (Died.) Shoem e Bipolaris sorokiniana (Sacc. in Sorok.) e F. graminearum. Foi verificado que a inibição do crescimento destes fungos está associada à liberação de compostos solúveis e voláteis pela bactéria, que foram responsáveis por cerca de 50 % e 40 % de inibição, respectivamente. O perfil da expressão gênica de F. graminearum na interação com a bactéria P. agglomerans foi avaliado via macroarranjo. Dos 1.014 genes avaliados, 29 genes de F. graminearum foram diferencialmente expressos (p < 0,05) durante a interação com a bactéria antagonista, sendo 19 genes induzidos e 10 genes reprimidos. Entre os transcritos induzidos foram identificadas proteínas envolvidas nos processos de defesa e/ou virulência de fungos, cuja expressão foi induzida em resposta a estresses tanto abióticos como bióticos. Dos genes que foram reprimidos, destacaram-se: um transcrito com similaridade a uma proteína com um domínio do tipo dedo de zinco ?zinc finger? que é um fator de transcrição importante no processo de divisão celular, bem como proteínas envolvidas na cadeia respiratória, na modulação protéica e sinalização celular. Os dados do macroarranjo foram validados via transcrição reversa seguida de PCR quantitativo em tempo real (RT-PCRq), metodologia que se mostrou adequada para complementar a análise transcricional obtida por macroarranjo. As informações geradas na análise de antagonismo in vitro, bem como a análise e seqüenciamento dos transcritos, juntamente com a quantificação do nível de expressão na interação, foram fundamentais para compreender o padrão de resposta do fungo F. graminearum na interação com a bactéria P. agglomerans.
Cultivated grasses such as wheat, barley and maize are agricultural products of fundamental economic and social importance in Brazil. Among causing factors of important grain production losses in these species are diseases caused by phytopathogenic fungi such as Fusarium graminearum Schwabe (teleomorfo Gibberella zeae Schw.), the causal agent of fusariosis, a disease of difficult chemical, biological or even genetic control. An efficient and promising strategy to be adopted in order to protect cultivated plants against such diseases is the selection of antagonist microorganisms, amongst them the bacteria Pantoea agglomerans. This microbiota might have an important impact in scab control, isolated or in an integrated management program with chemical treatment. The present work aimed at identifying differentially expressed sequences in pathogenic fungi-antagonistic microorganisms interactions, considering the F. graminearum ? P. agglomerans model. The construction of a cDNA library for F. graminearum grown in PDA medium generated 1,983 valid sequences and provided 1,283 unigenes. The most representative categories in this library were proteins involved in genetic information pathways, DNA-RNA-protein (26 %); hypothetical proteins (24 %); and proteins involved in metabolism (16 %). The protein category involved in developmental processes as well as those related to external stimuli perception comprised 10 % of the obtained unigenes. Among putatively annotated genes, some coding for enzymes of important metabolic routes were identified, such as glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase and phophoenolpyruvate carboxylase. Also secondary metabolism compounds, specially micotoxins and proteins related to fungi stresses and pathogenicity were identified. In the present work, the control of three wheat phytopathogens, Drechslera tritici-repentis (Died.) Shoem, Bipolaris sorokiniana (Sacc.in Sorok.) and F. graminearum, using specific isolates of P. agglomerans was demonstrated. It was observed that the 50 % and 40 % growth inhibition of these fungi is associated to the bacteria release of soluble and volatile compounds, respectively. The gene expression profile of F. graminearum during interaction with the bacteria P. agglomerans was evaluated via macroarray. Among the 1,014 analysed genes, 29 F. graminearum genes were differentially expressed (p < 0,05) during its interaction with the antagonist bacteria: 19 genes were induced while 10 genes were repressed. Among the induced transcripts, proteins involved in fungi defense and/or virulence processes were identified, whose expression was induced in reponse to abiotic or biotic stresses. Among the identified repressed genes, a transcript similar to a protein containing a zinc finger-type domain, a transcription factor relevant in cell division, deserves special attention, as well as proteins involved in respiratory chain, in protein modulation and in cell signaling. Additionally, the macroarray data were validated by reverse transcription followed by real-time quantitative PCR (RT-PCRq), a suitable method for complementing transcriptional analysis through macroarray. Finally, the information generated in in vitro pathogenic fungi-antagonistic microorganisms interactions analysis, as well as in the analysis and sequencing of the obtained transcripts, together with the determination of the level of expression during the evaluated interactions were essential for better understanding the response pattern of the fungus F. graminearum in interaction with the bacteria P. agglomerans

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Sauer, Aline Vanessa. "Monitoramento e caracterização da população epifítica de Pantoea ananatis, agente causal da mancha branca do milho". Universidade Estadual de Londrina. Centro de Ciências Agrárias. Programa de Pós-Graduação em Agronomia, 2010. http://www.bibliotecadigital.uel.br/document/?code=vtls000155148.

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A bactéria Pantoea ananatis, foi descrita recentemente como sendo agente causal da Mancha Branca do Milho. Os sintomas caracterizam-se inicialmente por lesões iniciais cloróticas aquosas, que se tornam necróticas de coloração palha. Este trabalho teve como objetivo, monitorar a população epifítica da bactéria, sob condições naturais de infestação e caracterizar essas bactérias comparando-as com aquelas presentes nas lesões. O antibiótico eritromicina foi usado como marcador de resistência, permitindo o crescimento de P. ananatis e inibição das demais bactérias da folha. Três híbridos, HS200, DAS657, 2B710 foram utilizados, e a partir de 50 dias pós-semeadura (DAS), coletas semanais foram realizadas na safra verão 2008/2009 e quinzenais realizadas na safrinha/2009 até o surgimento das lesões. As folhas foram segmentadas (2,5g de massa fresca) e submetidas a dois processos: 1) Safra verão 2008/2009: agitação por duas horas a 60 RPM à 28ºC em 100mL de tampão fosfato (pH 7,0) + 0,1(g/v) peptona bacteriológica; Safrinha/2009: agitação por duas horas a 140 RPM à 30oC em 100mL de tampão fosfato (pH 7,0) + 0,1(g/v) peptona bacteriológica e 2) maceração em 25mL de tampão fosfato (pH 7,0). Cerca de 0,1 mL de cada tratamento foi plaqueado em meio TSA acrescido de eritromicina (1mg/mL) e em TSA sem antibiótico (controle). As placas foram incubadas a 30ºC por 48 horas até a avaliação das unidades formadoras de colônias. Os isolados utilizados na caracterização para nucleação de gelo foram cultivados em meio TSB por 24 horas, à 30ºC e à 60RPM. 0,1mL da suspensão bacteriana foi acrescida à 1mL de água com temperatura externa abaixo de -1oC e observada a formação de gelo instantânea. Os isolados WT 2 e WT11 foram examinados em microscopia eletrônica de transmissão. Os resultados permitiram observar uma grande variação no tamanho da população epifítica da bactéria durante o desenvolvimento da planta, indicando que P. ananatis possa ficar acumulada em diferentes regiões da planta, sem uma distribuição homogênea. A análise de similaridade revelou que os isolados se agruparam em apenas um grupo e através de testes bioquímicos revelou-se que os mesmos eram pertencentes à espécie P. ananatis. Não foram observadas diferenças entre os isolados de lesão e da superfície epifítica para a atividade de nucleação de gelo. Através de microscopia eletrônica de transmissão, demonstrou-se a presença de estruturas semelhantes à vesículas protéicas na membrana externa, sendo provavelmente as responsáveis pela liberação de núcleos de gelo. No teste de patogenicidade os isolados foram capazes de causar sintomas em casa de vegetação e em câmaras de microumidade.
Pantoea ananatis, was recently described as the causative agent of Maize White Spot. The symptoms are characterized initially by initial chlorotic lesions aqueous, which become necrotic colored straw. This study aimed to monitor the population of epiphytic bacteria under natural conditions of infestation and characterize these bacteria by comparing them with those present in the lesions. The antibiotic erythromycin was used as a marker of resistance, allowing the growth of P. ananatis and inhibition of other bacteria of the leaf. Three hybrids, HS200, DAS657, 2B710 were used, and from 50 days after sowing (DAS), weekly collections were made in the 2008/2009 summer harvest and biweekly in little harvest/2009 until the appearance of lesions. The leaves were targeted (2.5 g fresh weight) and subjected to two processes: 1) Harvest summer 2008/2009: agitation for two hours at 60 RPM 28c in 100 mL of phosphate buffer (pH 7.0) + 0.1 (g/v) bacteriological peptone; little haverst/2009: agitation for two hours at 140 RPM at 30°C in 100 mL of phosphate buffer (pH 7.0) + 0.1 (g / v) bacteriological peptone and 2) in 25 mL of maceration phosphate buffer (pH 7.0). About 0.1 mL of each treatment was plated on TSA medium plus erythromycin (1mg/mL) and TSA without antibiotic (control). The plates were incubated at 30°C for 48 hours until the evaluation of colony forming units. The isolates used for the characterization of ice nucleation were grown in TSB medium for 24 hours, at 30°C and 60RPM. 0.1 mL of bacterial suspension was added to 1 mL of water with outside temperature below 1oC, and observed the formation of ice instantly. Isolates WT 2 and WT11 were examined in transmission electron microscopy. The results showed a wide variation in epiphytic size population of bacteria during plant development, indicating that P. ananatis can be accumulated in different parts of the plant, without a homogeneous distribution. The similarity analysis revealed that the isolates clustered in one group and by biochemical tests revealed that they belonged to the species P. ananatis. No differences were observed among isolates of injury and epiphytic surface for the activity of ice nucleation. Through transmission electron microscopy, showed the presence of structures similar to protein vesicles in the outer membrane, is probably responsible for the release of ice nuclei. In pathogenicity test, the isolates were able to cause symptoms in the greenhouse and chambers of microumidade.

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Geissinger, Jared Scott. "Structure-Function Analysis of the EsaR N-terminal Domain". Thesis, Virginia Tech, 2011. http://hdl.handle.net/10919/46190.

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The LuxR protein family is a class of quorum-sensing regulated bacterial transcription factors that alter gene expression as a function of ligand detection. This coincides with a high population density and/or a low rate of signal ligand diffusion. The majority of LuxR proteins are activated only in the presence of the signal ligand, an acyl-homoserine lactone (AHL). EsaR, from the corn pathogen Pantoea stewartii, represents a subset of LuxR homologues that are active in the absence of AHL and deactivated by its presence. The mechanism by which EsaR responds to AHL in a manner opposite to that of the majority of LuxR homologues remains elusive. Unlike the majority of LuxR homologues, which require AHL for purification, EsaR can be purified and biochemically investigated in the absence and presence of AHL. This work sought to answer questions regarding the structure-function relationship of the LuxR homologue, EsaR. Fluorescence anisotropy was used to determine the relative DNA-binding affinity of wild type EsaR and three AHL-independent EsaR variants in the presence and absence of AHL. This enabled for quantitative analysis of the relative binding affinities of these AHL-independent variants for the EsaR binding site, the esa box. The results demonstrate that one AHL-independent EsaR variant has a slightly higher affinity for the esa box in the presence, rather than the absence of AHL. The affinity of the other two for the DNA is not impacted by AHL, potentially due to an inability to transduce the signal of ligand detection to the DNA binding domain. Constructs containing only the EsaR N-terminal domain (NTD) were also developed. These constructs circumvented solubility issues associated with the full-length protein, allowing for additional biochemical analysis. It was determined that the EsaR NTD alone is sufficient for multimerization and ligand binding. Additionally, preliminary X-ray crystallography efforts have established some of the early parameters required to solve the crystal structure of the EsaR ligand binding domain in both the presence and absence of AHL. If pursued, these structures would be the first solved of a LuxR homologue ligand binding domain in both the presence and absence of the native AHL, potentially demonstrating the conformational change that occurs as a result of ligand binding. Collectively, these findings have established some of the groundwork required to resolve the question of what sort of conformational changes occur in EsaR as a result of ligand binding.
Master of Science

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Soto, Muñoz Lourdes. "Implementación de técnicas moleculares para la detección y cuantificación del agente de biocontrol Pantoea agglomerans CPA-2". Doctoral thesis, Universitat de Lleida, 2014. http://hdl.handle.net/10803/285968.

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Pantoea agglomerans CPA-2 es un agente de biocontrol (ACB) eficaz en el control de enfermedades de postcosecha en fruta de pepita y cítricos. No obstante, para implementar y registrar su uso como estrategia práctica de control en Europa es importante evaluar la capacidad del ACB para colonizar, persistir y propagarse en condiciones habituales de aplicación con un método de detección que permita diferenciar al antagonista del resto de la microbiota. La presente tesis doctoral tuvo como objetivo fundamental el desarrollo de técnicas moleculares para la detección y cuantificación específica de CPA-2 en su proceso de formulación y en su entorno de aplicación (pre- y postcosecha). Para cumplir este objetivo, en primer lugar se desarrolló y validó la técnica de qPCR, pero se observó la limitación de esta técnica al no discriminar entre el ADN de las células viables y las no viables de la superficie de manzanas conservadas a medio y largo plazo. Esta limitación se superó con un pre-tratamiento de las muestras con propidio de monoazida (PMA) en combinación con la qPCR (PMA-qPCR). Posteriormente, la técnica desarrollada del PMA-qPCR se aplicó para determinar la supervivencia del ACB después de su formulación por liofilización, lecho fluido y atomización pero también para evaluar su dinámica poblacional sobre la superficie de naranjas tratadas en pre- y postcosecha, siendo el primer estudio donde se aplica a un ACB. Finalmente, se estudió la persistencia de CPA-2 en tratamientos de pre- y postcosecha, confirmando su presencia mediante PCR convencional. Los resultados de esta tesis demuestran la versatilidad de las técnicas moleculares basadas en la PCR (qPCR, PMA-qPCR y PCR convencional) para detectar y cuantificar la población de CPA-2 durante su formulación y en las condiciones ambientales de aplicación. Así como la limitada dispersión y baja persistencia de CPA-2 en su entorno de aplicación.
Pantoea agglomerans CPA-2 és un agent de biocontrol (ACB) eficaç en el control de malalties de postcollita en fruita de llavor i cítrics. No obstant, per implementar i registrar el seu ús com estratègia pràctica de control a Europa és important avaluar la capacitat de l’ACB per colonitzar, persistir i propagar-se en condicions habituals d’aplicació amb un mètode de detecció que permeti diferenciar a l’antagonista de la resta de la microbiota. La present tesis doctoral va tenir com objectiu fonamental el desenvolupament de tècniques moleculars per a la detecció i quantificació específica de CPA-2 en el seu procés de formulació i en el seu entorn d’aplicació (pre- i postcollita). Per complir aquest objectiu, en primer lloc es va desenvolupar i validar la tècnica de qPCR, però es va observar la limitació d’aquesta tècnica al no discriminar entre el ADN de les cèl·lules viables i les no viables de la superfície de pomes conservades a mig i llarg termini. Aquesta limitació es va superar amb un pretractament de les mostres amb propidi de monoàcida (PMA) en combinació amb la qPCR (PMA-qPCR). Posteriorment, la tècnica desenvolupada del PMA-qPCR es va aplicar per determinar la supervivència de l’ACB després de la seva formulació per liofilització, llit fluït i atomització però també per avaluar la seva dinàmica poblacional sobre la superfície de taronges tractades en pre- i postcollita, sent el primer estudi on s’aplica a un ACB. Finalment, es va estudiar la persistència de CPA-2 en tractaments de pre i postcollita, confirmant la seva presència mitjançant PCR convencional. Els resultats d’aquesta tesis demostren la versatilitat de las tècniques moleculars basades en la PCR (qPCR, PMA-qPCR i PCR convencional) per detectar i quantificar la població de CPA-2 durant la seva formulació i en les condicions ambientals d’aplicació. Així com la limitada dispersió i baixa persistència de CPA-2 en el seu entorn d’aplicació.
Pantoea agglomerans CPA‑2 is a biocontrol agent (ACB) effective in the control of postharvest diseases in pome fruit and citrus. However, in order to implement and register the use of this ACB as a practical control strategy in Europe, it is first necessary to evaluate the ability of the antagonist to colonize, persist and spread in its normal operating conditions with a screening method to identify and quantify the ACB of the rest of the microbiota. The main goal of the present thesis was the development of molecular techniques for the identification and quantification of CPA‑2 strain in its formulation and application in the environment (pre- and postharvest). To achieve this objective, the development and validation of qPCR technique was due at first, however this technique was not able to discriminate between the DNA of viable and non‑viable cells of apple surface stored at middle and long time. This limitation was overcome by treating the samples with the DNA intercalator, propidium of monoazide (PMA), combined with qPCR (PMA‑qPCR). Furthermore, PMA‑qPCR technique was applied to determine the survival of CPA‑2 after its formulation by freeze drying, spray drying and fluidized bed drying; and to assess the population dynamics of the antagonist on the surface of pre‑treated and postharvest oranges, it was the first study where this technique was applied in ACB. Finally, CPA‑2 persistence in pre‑ and postharvest treatment was studied and its presence was confirmed by conventional PCR. The results presented in this thesis demonstrate the versatility of molecular techniques based on PCR (qPCR, PMA‑qPCR and conventional PCR) to detect and quantify P. agglomerans CPA‑2 in normal operating conditions.

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Otero, Bravo Alejandro. "Genome Evolution During Development of Symbiosis in Extracellular Mutualists of Stink Bugs (Pentatomidae)". The Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1586638345682906.

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Williamson, J. J. "Investigation into the biotechnological applications of Pantoea agglomerans for the production of high value terpenoids such as taxadiene". Thesis, University of Nottingham, 2017. http://eprints.nottingham.ac.uk/43555/.

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Plant derived terpenoids represent a diverse class of chemicals with important roles such as fragrances, flavours and pharmaceuticals as well as high value medicinal compounds such as the antimalarial Artemisinin and the anti-cancer drug Paclitaxel (Taxol). High volume production of these products can be difficult due to reliance on plant production or economic difficulties in synthetic production. These high value terpenoids represent an interesting challenge to metabolic engineering and synthetic biology, not only the requirement to produce these chemicals at scale, but also because novel products can be developed via the intermediates of these pathways. Research into methods of microbial production of Taxol and its derivatives have focused on the engineering of Escherichia coli and the precursor pathways leading to geranylgeranyl diphosphate (GGPP), since GGPP is the last non-dedicated precursor to Taxol. The work reported here has taken a different approach and has instead attempted to utilise natural diversity in search of a better host for production of these high value terpenoids, by selecting an organism that already produces a large amount of GGPP. As well as being a precursor to Taxol, GGPP can also be converted into carotenoid pigments such as lycopene, β-carotene and zeaxanthin. Therefore production of carotenoids can be used as a surrogate measure for predicting how well a species will perform in the production of heterologous isoprenoids. Genes for carotenoid biosynthesis from Pantoea species have been used in many heterologous expression experiments, however Pantoea have not been considered as a possible chassis for the production of isoprenoids. As a relatively close phylogenetic relative to E.coli, these Gram negative bacteria are easily transformable, genetically tractable and many tools which have been developed for E. coli function in Pantoea. The work reported here addresses the developments of tools and techniques to assess the ability of Pantoea agglomerans to produce terpenoids including lycopene and taxadiene. The tools developed included: 1) a range of promoters, which were characterised in E. coli and P. agglomerans for their regulation of lycopene biosynthesis, 2) methods for the editing of the genome of P. agglomerans and 3) a range of vectors for the expression of the first 3 dedicated steps in Taxol biosynthesis, taxadiene synthase (txs), taxadiene 5α hydroxylase and taxadiene-5α-ol-O-acetyl transferase (5α) and taxane 10β-hydroxylase (10β). These genes initially could not be cloned in an operon together due to apparent toxicity. However by designing measures to minimise the burden imposed it was possible to express all four enzymes, of the three step process in P. agglomerans. This work discusses the difficulties associated with working with a less well characterised organism (such as P. agglomerans), how we overcame these issues and the unexpected phenotypes exhibited by this genus.

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Miller, Amanda Mota. "Variabilidade genética e nucleação de gelo em isolados de Pantoea ananatis, agente causal da mancha branca do milho". Universidade Estadual de Londrina. Centro de Ciências Agrárias. Programa de Pós-Graduação em Agronomia, 2014. http://www.bibliotecadigital.uel.br/document/?code=vtls000190611.

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Medidas de controle da doença Mancha Branca do Milho (MBM), causada por Pantoea ananatis (Pa), são baseadas preferencialmente no desenvolvimento de cultivares resistentes, porém o desconhecimento e a falta de informações sobre a variabilidade genética do patógeno dificultam o estabelecimento de estratégias de manejo mais estáveis no sistema. Assim, os objetivos do trabalho foram investigar a variabilidade genética de isolados de Pa obtidos de diferentes regiões do Brasil e caracterizá-los quanto à presença e expressão fenotípica do gene inaA, responsável pelo surgimento inicial dos sintomas da doença. Para os estudos, foram utilizados isolados pertencentes ao banco de linhagens do Laboratório de Genética de Microrganismos da Universidade Estadual de Londrina, os quais tiveram seu DNA extraído e identidade confirmada por PCR utilizando os primers espécie-específicos ANAF/ANAR. A investigação da variabilidade genética foi conduzida com marcadores moleculares AFLP (Amplified Fragment Lenght Polymorphism). Na seleção das combinações com maior quantidade de locus polimórficos, foram selecionados para Pa os primers: EcoRI-ACG/MseI-CT, EcoRI-ACG/MseI-CAC e EcoRI-ACG/MseI-CAG. Com o intuito de encontrar possíveis variações no número de plasmídios na espécie, foi a realizada a extração do mesmo por lise alcalina. A caracterização dos isolados quanto à presença do gene inaA foi realizada por PCR utilizando os primers upper INA A/ lower INA A. A expressão do fenótipo INA+ foi avaliada adicionando-se 0,1mL de cultura bacteriana em água ultra-pura com temperatura de -10 oC. Dados de similaridade genética permitiram separar os isolados em dois grupos, porém sem correlação entre o local de origem do hospedeiro com a composição dos mesmos. A porcentagem de polimorfismo de Pa variou de 24,64% a 92,46% e a diversidade gênica de 0,07 a 0,09. A análise de variância molecular mostrou que 99,18% da variabilidade genética encontra-se dentro das populações. Os resultados obtidos para variabilidade genética apontam para a ação de forças evolutivas sobre as populações estudadas. Este é o primeiro relato da descrição de plasmídio de Pa provenientes de lesões de MBM. P. ananatis apresenta pelo menos um plasmídio, com tamanho estimado com base na literatura entre 280-352 kb. Correlação positiva entre a detecção do gene inaA e a atividade de nucleação de gelo foi obtida em Pa. Dos 90 isolados em estudo, apenas três não amplificaram para o primer, e cerca de 20% dos isolados, embora portadores do gene, não expressaram o fenótipo nas condições avaliadas. Concluiu-se que a expressão de gene inaA é dependente de características de cada individuo, pois sua presença no genoma não implica na manifestação fenotípica.
Measures of control the disease Maize White Spot (MWS), caused by Pantoea ananatis (Pa), are based preferably on the development of resistant cultivars, but the lack of information about the genetic variability of the pathogen, hampers to establish management strategies more stable in the system. The objectives of the study were to investigate the genetic variability of isolates of Pa obtained from different regions of Brazil and characterize them for the presence and phenotypic expression of inaA gene, responsible for the initial appearance of disease symptoms. For the studies, was used isolates belonging to the stock strains of the Laboratory of Genetics of Microorganisms, State University of Londrina, that had their DNA extracted and identity confirmed by PCR using species-specific primers ANAF/ANAR. The investigation of the genetic variability was conducted with molecular markers AFLP (Amplified Fragment Lenght Polymorphism). In selecting the combinations with the highest number of polymorphic loci were selected for Pa the EcoRI-ACG/MseI-CT, EcoRI-ACG/MseI-CAC and EcoRI-ACG/MseI-CAG primers. In order to find possible variations in the number of plasmids in the species, the extraction was performed by the alkaline lysis. The characterization of isolates for the presence of inaA gene was performed by PCR using the primers upper INA A/lower INA A. The expression of INA+ phenotype was assessed by adding 0.1 mL of bacterial culture in ultra- pure water to a temperature of -10 °C. Database of genetic similarity allowed to separate the isolates into two groups, but no correlation between the location of origin of the host with the composition. The percentage of polymorphism Pa ranged from 24.64% to 92.46 % and gene diversity from 0.07 to 0.09. The analysis of molecular variance showed that 99.18 % of genetic variation is found within populations. The results obtained for genetic variability point to the action of evolutionary forces on the populations studied. This is the first report describing Pa plasmid from injuries MBM. P. ananatis has at least one plasmid, with size estimated based on the literature between 280-352 kb. Positive correlation between detection of gene inaA and ice nucleation activity was obtained in Pa. Of the 90 isolates studied, only three non-amplified for the primer, and about 20% of the isolates, although gene carriers, did not express the phenotype in the conditions evaluated. It was concluded that the inaA gene expression is dependent on characteristics of each individual, because their presence in the genome does not imply the phenotypic manifestation.

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Pomini, Armando Mateus. "Acil-homosserina lactonas produzidas pelas bacterias fitopatogenicas Pantoea ananatis e Methylobacterium mesophilicum e defesa quimica no opilião Hoplobunus mexicanus". [s.n.], 2009. http://repositorio.unicamp.br/jspui/handle/REPOSIP/249296.

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Orientador: Anita Jocelyne Marsaioli
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica
Made available in DSpace on 2018-08-13T13:06:45Z (GMT). No. of bitstreams: 1 Pomini_ArmandoMateus_D.pdf: 4501241 bytes, checksum: 249f7514962993874b22ddcad9831cc8 (MD5) Previous issue date: 2009
As bactérias Gram-positivas e Gram-negativas possuem um mecanismo de comunicação química intra-específico conhecido como ¿quorum-sensing¿, regulando a expressão de uma vasta gama de atividades biológicas. As bactérias Gram-negativas utilizam acil-homosserina lactonas (acil-HSLs) como principais substâncias sinalizadoras. Na presente tese, relatamos a determinação da configuração absoluta do raro metabólito (S)-(-)-N-heptanoil-HSL produzida pela bactéria fitopatogênica Pantoea ananatis. A configuração absoluta desta substância foi determinada através da técnica de cromatografia gasosa com detecção por ionização em chama com coluna quiral, através de comparações de tempo de retenção e co-injeção com padrões sintetizados. Avaliou-se também a importância da configuração absoluta para a atividade antimicrobiana de acil-HSLs contra bactérias Gram-positivas (Bacillus subtilis, Bacillus cereus e Staphylococcus aureus). Curiosamente, o enantiômero não natural (R)-N-3-oxo-octanoil-HSL foi tão ativo quanto o produto natural (S). Estudou-se também as interações da (S)-N-3-oxo-octanoil-HSL com células de Agrobacterium tumefaciens NTL4(pZLR4) através da técnica de ressonância magnética nuclear de hidrogênio por diferença de transferência de saturação (STD-RMN), revelando que o primeiro evento de interação da substância com a célula ocorre com a região lipídica da membrana celular externa. Finalmente, realizou-se o estudo químico das substâncias sinalizadoras produzidas pela bactéria Methylobacterium mesophilicum, que ocorre simbioticamente com a bactéria Xylella fastidiosa nos vasos condutores de laranjeiras atacadas pela clorose variegada dos citros. Entre os vários resultados inéditos, reportamos a caracterização e síntese do produto natural inédito (S)-N-(2E)-dodecenoil-HSL e a primeira síntese do metabólito (S)-N-(2E, 7Z)-tetradecadienil-HSL. Outrossim, reportamos a primeira caracterização da configuração absoluta de cinco acil-HSLs naturais de cadeia longa. Realizou-se também estudos relacionados aos efeitos das acil-HSLs sintéticas contra bactérias Gram-positivas endofíticas da laranjeira. Adicionalmente, caracterizou-se a secreção de defesa do opilião Hoplobunus mexicanus. O repertório de defesa deste animal é composto por dois componentes voláteis de alta irritabilidade (2,5-dimetil-fenol e 2-metil-5-etil-fenol), além da tanatose e emissão de sons, uma característica inédita em opiliões
Gram-positive and Gram-negative bacteria use quorum sensing communication circuits to regulate a diverse array of physiological activities. In general, Gram-negative bacteria use acylated homoserine lactones (acyl-HSLs) as autoinducers, and Gram-positive bacteria use processed oligo-peptides. In the present work, we relate the absolute configuration determination of the rare metabolite (S)-(-)-N-heptanoyl-HSL produced by the phytopathogen Pantoea ananatis. The absolute configuration was determined by gas chromatography coupled to flame ionization detection with chiral column, through retention time comparison and co-injections with synthetic products. The importance of the absolute configuration for the antimicrobial activity of acyl-HSL against Gram-positive bacteria (Bacillus subtilis, Bacillus cereus and Staphylococcus aureus) was assessed. Curiously, non-natural (R)-N-3-oxo-octanoyl-HSL was as active as the natural product with (S) absolute configuration. The interaction of (S)-N-3-oxo-octanoil-HSL with Agrobacterium tumefaciens NTL4(pZLR4) cells was further studied using hydrogen nuclear magnetic resonance experiments with saturation transfer difference (STD-NMR) revealing that the first binding event is the diffusion through the lipidic part of the outer membrane. Finally, we have investigated the chemical study of the signaling substances produced by Methylobacterium mesophilicum, which co-occurs with Xylella fastidiosa in orange trees affected by the citrus variegated chlorosis disease. Among several results, we report herein the characterization and synthesis of a new natural product [(S)-N-(2E)-dodecenoyl-HSL], the first synthetic procedure for the rare (S)-N-(2E,7Z)-tetradecadienyl-HSL and the occurrence of a rare long, odd chain representative (N-tridecanoyl-HSL) in trace amounts. We report the first absolute configuration determination for five natural acyl-HSLs. We have also studied the effects of synthetic acyl-HSLs on Gram positive bacteria isolated from orange tissues. Additionally, the defensive secretion produced by the harvestman Hoplobunus mexicanus was characterized. The defensive repertory of this arachnid includes two irritating and volatile components (2,5-dimethyl-phenol and 2-methyl-5-ethyl-phenol), besides thanatosis and sound emission, a new behavioral artifice in opilionids
Doutorado
Quimica Organica
Doutor em Ciências

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Bartholomew, Holly Packard. "In planta studies of the corn pathogen Pantoea stewartii subsp. stewartii and applications of a corn-based industrial byproduct". Diss., Virginia Tech, 2020. http://hdl.handle.net/10919/99356.

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Corn is a valuable agricultural commodity in the United States and in the world. The causal agent of Stewart's wilt disease in corn, Pantoea stewartii subsp. stewartii, is a bacterial phytopathogen that is vectored into the plant by the corn flea beetle, Chaetocnema pulicaria. After entering the apoplast of the leaf, the bacteria cause water soaking symptoms before traveling to the plant xylem to form a dense biofilm, thereby blocking water transport and inducing necrosis and wilt. This results in reduced crop yield and may even lead to death of the corn plant. To better understand the in planta requirements of this pathogen, a whole transcriptome study was performed via RNA-Seq to determine genes differentially expressed in the bacteria while inside the corn. It was found that nutrient transporters and stress response genes were upregulated specifically when the bacteria are in their host plant, suggesting a response to nutrient availability and host defense in the xylem. Further elucidation of the genes required for the P. stewartii in planta lifestyle was performed via a reverse genetics approach where in-frame gene deletions and the corresponding complementation strains were constructed for genes that had shown a fitness defect in corn based on a previously published Tn-Seq study: genes encoding seven transcription factors, nsrR, iscR, lrp, nac, DSJ_00125, DSJ_03645, and DSJ_18135, as well as a hypothetical protein DSJ_21690. Investigation of the physiological role of these genes was performed using in planta virulence and competition assays for all strains. An in planta qRT-PCR analysis of bacterial gene transcription was also completed for the strains with deletions in nsrR and iscR. In vitro assays were performed on all strains to determine their capsule production and motility phenotypes. Taken together, it was seen that iscR is important for colonization capabilities in planta, both NsrR and IscR act as regulators, and lrp is important for full disease capabilities, perhaps due to reduced capsule and motility phenotypes. These findings lay the groundwork for finding potential disease intervention strategies not only against P. stewartii, but also other xylem-dwelling bacterial phytopathogens. In addition to exploring ways to enhance crop yield, an additional research area was on repurposing a byproduct of corn ethanol production, syrup. It was hypothesized that this corn-based syrup could be utilized as a carbon source to grown bacteria. In turn, the resulting bacterial biomass could then be added as a fish feed supplement in aquaculture. Syrup was tested as a growth medium for individual soil bacterial isolates as well as a full mixed bacterial community consortium to determine which bacteria could grow most efficiently, both in rate and yield. It was found that the highest growth rate and yield was from Bacillus species, some of which may have probiotic benefits to fish. Ultimately, the collective outcomes from these projects in basic research about a bacterial corn pathogen and applied research about beneficial microbes grown on a corn-based substrate are expected to improve scientific endeavors as well as agricultural practices.
Doctor of Philosophy
Corn is a top agricultural commodity in the United States, as a food for human consumption, a primary nutrient source used in animal feed, and a substrate consumed during biofuel production. These various corn-based industries are impacted by bacteria in multiple ways; in some cases, bacteria may cause disease that reduces crop yield, but other bacteria serve beneficial roles that enhance health. This dissertation research describes studies about the bacterium that causes Stewart's wilt disease in corn, Panteoa stewartii subsp. stewartii. In an initial experiment, the genes that P. stewartii expresses at the highest levels when it grows inside the corn plant were identified. These genes were deduced to be important for the ability of the bacterium to live successfully in this environment. This work was followed up with a more specific approach that examined the role of certain genes that were predicted to be master regulators of the expression of other genes in the ability of the P. stewartii to colonize the plant and/or cause disease. By identifying key bacterial genes, disease intervention strategies to combat Stewart's wilt and other similar bacterial plant pathogen diseases might become possible. Protecting corn yields is important for ethanol production. The final study of this dissertation examined the ability of bacteria to grow on a byproduct of ethanol production called syrup. The goal was to then use the biomass of these beneficial microbes as a food source for animals being produced in aquaculture facilities. Among the species tested, the highest growth rate and yield was from Bacillus subtilis, a safe-to-eat bacterium that has known beneficial health properties when consumed by fish. Overall, the research studies that were completed for this dissertation have the potential to improve agricultural practices by decreasing corn disease leading to increased corn yield and developing new downstream corn-based animal feed products.

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Han, David Youngsun. "Identification an characterization of the systemic resistance-inducing biological control agent Pantoea Agglomerans E278A from compost-amended potting mixes /". The Ohio State University, 1999. http://rave.ohiolink.edu/etdc/view?acc_num=osu1488187049539837.

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Fudge, James R. "Selection and Use of Pantoea dispersa strain JFS as a Non-Pathogenic Surrogate for Salmonella Typhimurium Phage Type 42 in Flour". BYU ScholarsArchive, 2015. https://scholarsarchive.byu.edu/etd/5972.

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Salmonella, a common food pathogen, costs more than any other pathogen in the United States in terms of health care costs and loss of work due to the illnesses it causes. Low-moisture foods, especially flour, are susceptible to being contaminated by Salmonella. Food producers want flour to be pathogen-free but to also retain the same functionality of non-treated flour. Heat treatment is the most common method employed for lowering the concentration of pathogens in food. However, heating can result in the loss of the flour’s functionality. Pantoea dispersa strain JFS has been isolated from flour as a nonpathogenic bacterial surrogate that closely matches the D-value of Salmonella in flour. Flour samples were subjected to dry heat (70, 75, and 80°C) and heat tolerance was determined by plating out at least four different time points for each temperature. The death rate of P. dispersa strain JFS was similar to (p<0.05) Salmonella. This strain of P. dispersa was then used as a surrogate for Salmonella in a continuous and batch heat treatment processes to determine the amount of kill achieved by each. The continuous process was conducted using varying levels of four independent variables: temperature, residence time, use of steam, and manipulation of initial water content. All 15 runs resulted in a reduction of at least 1.5 logs of the surrogate, with the greatest reduction being 2.5 logs. The batch process was conducted using one independent variable, temperature. All runs for the batch process resulted in a reduction of at least 2.5 logs of the surrogate, with the greatest reduction being 4.3 logs at 170°F. Both processes could be used to reduce any Salmonella present in flour.

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Sharma, Ruchira. "Isolation, Characterization, and Genomic Comparison of Bacteriophages of Enterobacteriales Order". BYU ScholarsArchive, 2019. https://scholarsarchive.byu.edu/etd/8577.

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According to CDC, every year at least 2 million people are affected and 23,000 dies as a result of antibiotic resistance in U.S. It is considered one of the biggest threats to global health. More and more bacterial infections are becoming harder to treat. One such infection is fire blight, one of the most destructive disease of apple and pear trees. It is caused by bacteria Erwinia amylovora and its outbreaks have been known to destroy entire orchards in a single season. The conventional method of treatments includes use of antibiotics like streptomycin and oxytetracycline but the incidences like presence of multi-drug resistant bacteria in the mammals grazing in the fields have raised concerns. Phage therapy is considered one of the few ways available to combat bacterial resistance and prevent fire blight. In this method, a cocktail of highly lytic bacteriophages is prepared and sprayed on the trees at different time intervals. Bacteriophages are an “intelligent” drug. They multiply at the site of the infection until there are no more bacteria and then they are excreted back into the nature. These phenomena make them more efficient than an antibiotic, which kills all kind of bacteria including good bacteria and can be maintained in the environment for long periods of time. These qualities of bacteriophage have resulted in many commercially available phage therapies. The initial part of this research focuses on isolation, characterization and genomic comparison of bacteriophages that infect a plant pathogen E.amylovora of Erwiniaceae family of Enterobacteriales order. In this study, 28 novel bacteriophages were isolated, fully sequenced, characterized and grouped into seven families based on phage homology. To take this further, we characterized a novel jumbo family of bacteriophages that has a small burst size of 4.6-4.9 and are most similar to bacteriophages that infect Pseudomonas and Ralstonia rather than Enterobacteriales bacteria by protein similarity. These bacteriophages are shown to infect Erwinia and Pantoea bacterial strains, but no infection of 9 other bacterial strains tested, was seen, under laboratory conditions. The results of this work provide an insight on special characteristics that makes bacteriophage so unique and adaptable. The final part of this research explores the enormous diversity of bacteriophages. In 2014 Grose and Casjens grouped 337 fully sequenced tailed phages into 56 diverse clusters (32 lytic and 24 temperate). We further expanded our current understanding of these clusters by performing the comprehensive analysis of genomes and proteomes of 1037 tailed bacteriophages, posted on GenBank. The results of this work provide insights into diversity and relatedness of bacteriophages and the data is posted on GenBank.

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Merighi, Massimo. "Molecular biology and biochemistry of regulation of Hrp/type III secretion genes in the corn pathogen Pantoea stewartii pv. stewartii". Connect to this title online, 2004. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1069854564.

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Thesis (Ph. D.)--Ohio State University, 2004.
Document formatted into pages; contains xxxiii, 421 p. Includes bibliographical references. Abstract available online via OhioLINK's ETD Center; full text release delayed at author's request until 2005 Dec. 2.

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Mittapelly, Priyanka. "Molecular interactions of brown marmorated stink bug, Halyomorpha halys with its bacterial endosymbiont, Pantoea carbekii and their role in nutrient provisioning". The Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1543246781490262.

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Delétoile, Alexis. "Phylogénie, espèces et populations bactériennes : Séquençage de gènes ubiquistes chez les Enterobacteriaceae et Bifidobacterium". Paris 7, 2009. http://www.theses.fr/2009PA077047.

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La diversité phylogénétique bactérienne est d'une richesse considérable, tant en nombre d'espèces que par la diversité clonale en leur sein. La connaissance de cette biodiversité est nécessaire pour comprendre l'évolution des propriétés biologiques, les mécanismes de spéciation et pour le suivi épidémiologique. L'emploi de gènes multiples codant pour des protéines apporte une précision phylogénétique nettement supérieure à l'ARN 16S et minimise les distorsions causées par la recombinaison. Les objectifs de cette thèse étaient d'appliquer le séquençage de gènes de ménage multiples à large échelle en utilisant des gènes universellement conservés, de déterminer le pouvoir résolutif de cette approche à différents niveaux phylogénétiques (de la famille au clone) et d'étudier la diversité et l'évolution de deux groupes importants. La famille des Enterobacteriaceae a été choisie en raison de sa grande diversité écologique et de son importance médicale et agronomique. Une phylogénie précise et robuste (171 taxa, 6 gènes) a permis de retracer l'histoire de sa diversification écologique. Le genre Bifidobacterium comprend des espèces importantes de la flore intestinale et dans l'industrie agroalimentaire. Le séquençage de sept gènes des quatre espèces B. Animalis, B. Bifidum, B. Brève et B. Longum (130 souches) a permis de préciser leurs frontières phylogénétiques. Chez ces deux groupes, la recombinaison homologue inter-espèces est rare, mais peut perturber la phylogénie de gènes individuels. L'emploi du même jeu de gène a permis de déterminer la diversité clonale des espèces d'enterobactéries Pantoea agglomerans, Plesiomonas shigelloides et Enterobacter cloacae et des quatre espèces de Bifidobacterium. Ce travail démontre l'efficacité de l'utilisation de gènes ubiquistes à la fois pour la phylogénie, la délimitation des espèces, la génétique des populations et le typage épidémiologique
The phylogenetic diversity of bacteria is impressive, both by the number of species and by the clonal diversity within them. Knowledge of this biodiversity is needed to understand the evolution of biological properties, the mechanisms of speciation and for epidemiological tracking. The use of multiple protein-coding genes provides a better phylogenetic precision compared to 16S RNA and minimizes the distortions caused by recombination. The aims of this thesis were to apply the sequencing of multiple housekeeping genes on a large scale by using universally' conserved genes, to determine the discrimination power of this approach at different phylogenetic levels (from family to qlone) and to study diversity and the evolution of two major groups. The Enterobacteriaceae family was chosen because of its large ecological diversity and its medical and agricultural importance. A precise and robust phylogeny was established (171 taxa, 6 genes) and revealed the history of ecological diversification of this family. The genus Bifidobacterium includes species that are important members of the intestinal flora and for the food industry. The sequencing of seven genes in the four species B. Animalis, B. Bifidum, B. Breve and B. Longum (130 strains) has clarified the phylogenetic borders between them. In both groups, inter-species homologous recombination is rare, but can disturb the phylogeny of individual genes. The use of the same gene set allowed determining the clonal diversity of the enterobacterial species Pantoea agglomerans, Plesiomonas shigelloides and Enterobacter cloacae and of the four species of Bifidobacterium. This work demonstrates the effectiveness of using ubiquitous genes for phylogeny, for species circumscription and for population genetics and epidemiological typing

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Ji, Yuxia. "BIOLOGICAL SELENIUM CONTROL: SELENIUM REDUCTION BY SHIGELLA FERGUSONII STRAIN TB42616 AND PANTOEA VAGANS STRAIN EWB32213-2 IN BIOREACTOR SYSTEMS". UKnowledge, 2019. https://uknowledge.uky.edu/ce_etds/91.

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Se(VI) and Se(IV), as the two major species of selenium in water, are toxic to aquatic lives and may cause adverse health effects to humans at high levels. Biological reduction of Se(VI) is a two-stage process first from Se(VI) to Se(IV) and then from Se(IV) to Se(0) with potential accumulation of the more toxic Se(IV) due to the slower rate of the second stage. Selenium reduction was first evaluated with batch cultures of Shigella fergusonii strain TB42616 (TB) and Pantoea vagans strain EWB32213-2 (EWB) isolated in our laboratory from sludge and coal slurry sediment samples, respectively. In order to facilitate Se(VI) reduction and reduce Se(IV) accumulation, the Se(VI)-reducing strain TB was co-cultured with a Se(IV)-reducing strain EWB. Although Se(VI) reduction rate was not affected, Se(IV) reduction was significantly enhanced with low Se(IV) accumulation in the defined co-culture. Effects of culture composition as well as nitrate and arsenate on Se(VI) reduction were also investigated. A co-culture composition of 10:1 (EWB:TB) ratio was observed to achieve the best total selenium reduction. In addition, nitrate at 50 mg/L was observed to inhibit Se(IV) reduction but not Se(VI) reduction, while arsenate at 200 mg/L exhibited slight inhibition on both Se(VI) and Se(IV) reduction. Biokinetic parameters were optimized with a Monod-type kinetic model using batch pure culture data through the Robust Global Optimization Algorithm embedded in a computer package. Se(VI) reduction by the defined co-culture was then simulated and verified over a range of culture compositions and initial Se(VI) concentrations, respectively. An inter-species inhibition term was incorporated into the model to illustrate the competition for Se(IV) during Se(VI) reduction in the co-culture. The model showed a significant increase of Se(IV) accumulation with higher initial Se(VI) concentration. However, Se(IV) accumulation can be reduced with increasing population ratio of EWB to TB in the defined co-culture. The relatively high correlation coefficients suggested that the model was robust and applicable in simulating Se(VI) reduction by the defined co-culture. Since activated alumina was reported to be more effective for Se(IV) adsorption than Se(VI), the effect of biological activities on selenium removal was investigated using continuous-flow reactors packed with alum-impregnated activated alumina (AIAA) and cultured with a Se(VI)-reducing strain TB under various influent Se(VI) concentrations and hydraulic retention times (HRTs). A selenium removal efficiency of 92% was achieved in a bioreactor with initial biomass of 2.2×106 cells/g-AIAA after a 70-day operation period. Little improvement was observed by lowering the influent Se(VI) concentration from 50 to 10 mg/L while the removal efficiency was significantly enhanced by either extending the hydraulic retention time from 3.2 to 5.0 days or increasing the attached biomass during the startup. An increase in mass ratios of Se(VI) reduction by immobilized cells to adsorption by AIAA was also observed with increasing cell mass during the operation. Se(VI) reduction using continuous-flow reactors packed with strain TB immobilized Ca2+-alginate beads was investigated under various hydraulic retention times (HRT) and influent Se(VI) concentrations. A high removal efficiency up to 98.7% was achieved under an HRT of 5 days and an influent Se(VI) concentration of 400 mg/L. The results showed that the overall selenium removal was positively correlated to the bed height of the reactor and the HRT but not related to the influent Se(VI) concentration. The steady state was analyzed using a mathematical model based on Monod-type equations with four biokinetic parameters optimized including the half-velocity constants and maximum specific reduction rates. The relatively high correlation coefficients indicate that the model is robust and valid to simulate Se(VI) reduction in the gel-beads-packed continuous-flow system.

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Taylor, Christopher Michael. "Understanding the relationship between the brown marmorated stink bug, Halyomorpha halys (Stal), and its symbiont, Pantoea carbekii, with implications for stink bug management". Thesis, University of Maryland, College Park, 2016. http://pqdtopen.proquest.com/#viewpdf?dispub=10128715.

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Symbiotic relationships between insects and beneficial microbes are very common in nature, especially within the Hemiptera. The brown marmorated stink bug, Halyomorpha halys Stål, harbors a symbiont, Pantoea carbekii, within the fourth region of the midgut in specialized crypts. In this dissertation, I explored this insect-microbe relationship. I determined that the brown marmorated stink bug is heavily reliant on its symbiont, and that experimental removal of the symbiont from the egg mass surface prior to nymphal acquisition led to lower survival, longer development, lower fecundity, and aberrant nymphal behavior. Additionally, I determined that even when the symbiont is acquired and housed in the midgut crypts, it is susceptible to stressors. Stink bugs reared at a higher temperature showed lower survival, longer development, and a cease in egg mass production, and when bugs were screened for their symbiont, fewer had successfully retained it while under heat stress. Finally, with the knowledge that the stink bug suffers decreases in fitness when its symbiont is missing or stressed, I wanted to determine if targeting the symbiont was a possible management technique for the stink bug. I tested the efficacy of a number of different insecticidal and antimicrobial products to determine whether prevention of symbiont acquisition from the egg mass was possible, and results indicated that transmission of the symbiont from the egg mass to the newly hatched nymph was negatively impacted when certain products were applied (namely surfactants or products containing surfactants). Additionally, direct effects on hatch rate and survival were reported for certain products, namely the insect growth regulator azadirachtin, which suggests that nymphs can pick up residues from the egg mass surface while probing for the symbiont. I conclude that P. carbekii plays a critically important role in the survival of its host, the brown marmorated stink bug, and its presence on the egg mass surface before nymphal hatch makes it targetable as a potential management technique.

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41

Correa, Valdir Ribeiro Mr. "FUNCTIONAL GENOMICS OF PANTOEA STEWARTII SUBSP. STEWARTII AND PARTIAL GENOME SEQUENCE OF THE MAIZE STOLBUR PHYTOPLASMA SOLANI, TWO INSECT-TRANSMITTED BACTERIAL PATHOGENS OF MAIZE". The Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1291166530.

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42

Arruda, Andrelisse, e 69-99901-2574. "Identificação de microrganismos cultiváveis associados ao intestino de Anopheles darlingi (Diptera: Culicidae) com potencial à paratransgênese para o controle da malária". Universidade Federal do Amazonas - Universidade Federal de Rondônia, 2017. https://tede.ufam.edu.br/handle/tede/6375.

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Microrganisms living in insects’ midgut have been isolated and identified for developing biotechnological tools to fight vector-born diseases. In this context, the mosquitoes Anopheles from different regions around the world have been studied about their midgut microbiota focused on paratransgenesis. However, information about microrganisms living in neotropical mosquitoes midgut are scarce. And, specially about Anopheles darlingi, the main malaria vector in the Brazilian Amazon, still this study, there were not any report about culturable microrganisms associated to this insect. The first step for paratransgenesis is to isolate culturable microrganisms naturally associated to the insect vector, and thus amenable to experimentation in laboratory. The objectives of this work were to isolate and to identify culturable bacteria and yeasts isolated from feces of Anopheles darlingi, the main vector of malaria in Brazil; to estimate the species richness and frequency distribution of the sampled bacteria and yeasts and to characterize and to select the sampled bacteria and yeasts isolated from feces of An. darlingi with potential for paratransgenesis. The female mosquitoes of An.darlingi were captured in two rural places of Porto Velho, Rondônia, Brasil. For improving the bacterial growth, mosquito feces were collected on LB agar medium and cultivated at 37 ºC for 24 hours, and for improving the yeast growth, mosquito feces were collected on YPD agar medium with Chloramphenicol and cultivated at 30 ºC for 48 hours. Sixty pure bacteria colonies and sixty pure yeast colonies were sampled. The isolates were preserved in -80 ºC freezer. PCR reactions with genomic DNA from each isolate were perfomed using the primers of 16S rRNA genes for bacteria and 26S and ITS for yeasts. From 60 bacterial isolates, 55 samples were identified. From 60 yeast isolates, 27 samples were identified. The fragments were sequenced with the Sanger method and the sequences with similarities above of 97% with sequences in reference database were deposited in Genbank (NCBI). For bacteria, MALDI-TOF, VITEK®2 and BBL Crystal were also used as a complementar protocols to identify the isolates. The identified bacteria fall into 8 genera, Enterobacter, Klebsiella, Cedecea, Pantoea, Serratia, Acinetobacter, Burkholderia and Staphylococcus. The identified yeast fall into 7 genera, Pseudozyma, Papiliotrema (Cryptococcus), Meyerozyma (=Pichia), Rhodotorula, Candida, Hanseniaspora and Metschnikowia. As candidates to paratransgenesis to control of malaria in An. darlingi are those bacteria belonging to the genera Pantoea and Serratia and the yeasts belonging to the genera Meyerozyma (=Pichia), Pseudozyma, Hanseniaspora and Metschnikowia.
Microrganismos contidos no trato digestório de insetos vem sendo isolados e identificados com o intuito de desenvolver ferramentas biotecnológicas para o controle de doenças transmitidas por insetos. Nesse contexto, mosquitos Anopheles de diferentes partes do globo têm sua microbiota investigada com foco em paratransgênese. No entanto, a informação sobre microrganismos associados aos anofelinos neotropicais é escassa. Tratando-se de Anopheles darlingi, o principal vetor de malária no Brasil, até o presente trabalho, não havia informações sobre microrganismos cultiváveis associados a esse vetor. Os objetivos deste trabalho foram isolar e identificar bactérias e leveduras cultiváveis isoladas das fezes de An. darlingi, o principal vetor da malária no Brasil; estimar riqueza e distribuição de frequência das bactérias e leveduras amostradas, e caracterizar e selecionar bactérias e leveduras isoladas das fezes de An. darlingi com potencial para paratransgênese. Os mosquitos An. darlingi fêmeas foram coletados em duas localidades rurais de Porto Velho, Rondônia, Brasil. Para favorecer o crescimento de bactérias, fezes dos mosquitos foram coletadas em meio LB ágar e cultivadas à 37°C por 24 horas, e para propiciar o crescimento de leveduras, fezes dos mosquitos foram coletadas em meio YPD ágar com cloranfenicol e cultivadas à 30°C por 48 horas. Sessenta colônias bacterianas e 60 colônias leveduriformes foram amostradas. Os isolados foram preservados em freezer -80 °C. Foram realizadas PCR utilizado DNA genômico dos isolados com iniciadores para a região do DNA ribossomal 16S para bactérias e 26S e ITS para leveduras. Todas as 60 bactérias isoladas foram identificadas. Das 60 leveduras isoladas, 27 foram identificadas. Os fragmentos foram sequenciados pelo método Sanger e as sequências com similaridades superiores a 97% frente a sequências disponíveis em bancos de dados foram depositadas no GenBank. Para bactérias, MALDI-TOF, VITEK®2 e BBL Crystal foram utilizados como métodos complementares para identificação dos isolados. As bactérias identificadas pertencem a 8 gêneros: Staphylococcus, Burkholderia, Cedecea, Enterobacter, Klebsiella, Pantoea, Serratia e Acinetobacter. As leveduras identificadas pertencem a 7 gêneros: Candida, Meyerozyma (=Pichia), Metschnikowia, Hanseniaspora, Rhodotorula, Papiliotrema (=Cryptococcus) e Pseudozyma. São candidatas à paratransgênese para o controle da malária em An. darlingi as bactérias do gênero Pantoea e Serratia e as leveduras dos gêneros Meyerozyma (Pichia), Metschkowia, Hanseniaspora e Pseudozyma.

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43

Jin, Lin. "The Bacterial AvrE-Family Type-III Effector Proteins Modulate Plant Immunity via Targeting Plant Protein Phosphatase 2A Complexes". The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1458339056.

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44

Rafael, Mallaupoma Zeny C. "Determinación y diferenciación de Pantoea agglomerans y patógenos por medio de perfiles de ácidos grasos a partir de aislamientos de 5 variedades de semillas de Allium cepa L". Master's thesis, Universidad Nacional Mayor de San Marcos, 2007. https://hdl.handle.net/20.500.12672/826.

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Por medio de metil acido esteres (FAMES), fueron analizados 382 aislamientos pertenecientes a cinco variedades de semillas de Allium cepa L. (S1, S2,S3,S4,S5). Se determino que el 20% afecta a cultivos de interés económico, el 36% pertenecen a patógenos que afectan humanos, el 4% afectan a animales y el 40% son del ambiente. A los aislamientos se les identifico por medio de perfiles de ácidos grasos usando un cromatógrafo HP 5890 serie II y una columna capilar agilent ultra 2, software MIDI , Newark , Del. Se encontró principalmente los géneros Erwinia, Enterobacter, Pantoea y Pseudomonas las cuales afectan cultivos de interés económico (Allium cepa L. Carica papaya, Zingiber ofíciale, Eucalyptus spp, Cucumis melón, Ananas comosus, Sorghum). De las variedades analizadas, S2 reportó mayor diversidad de Pantoea agglomerans seguida por S3 que además presenta mayor concentración de Enterobacter cloacae; en la variedad S1 ambos patógenos se presentaron en proporción semejante, a diferencia de S5 que presentó Pantoea ananas, Pantoea ananatis, Enterobacter cloacae y S4 solo presento Pantoea agglomerans.
--- By means of acid methyl esters (FAMES), 382 isolates were analyzed from five varieties of seed Allium cepa L. (S1, S2, S3, S4, S5). It was determined that 20% affect crops of economic interest, 36% belong to pathogens that affect humans, 4% affect animals and 40% are from the environment. The isolates were identified by means of fatty acid profile chromatography using an HP 5890 series II, a capillary column Agilent Ultra 2 and MIDI software, Newark, Del. Findings included mainly the genera Erwinia, Enterobacter, Pantoea and Pseudomonas; all of them are described affecting crops of interest. (Allium cepa L. Carica papaya, Zingiber ofíciale, Eucalyptus spp, Cucumis melón, Ananas comosus, Sorghum). Of the varieties tested, S2 reported the greatest diversity of Pantoea agglomerans S3 followed by the largest concentration of Enterobacter cloacae; In the variety S1, both pathogens were presented in similar proportion, unlike S5 which submitted Pantoea ananas, Pantoea ananatis, and Enterobacter cloacae. S4 only presented Pantoea agglomerans.
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45

Amellal, Najat. "Rôle de bactéries productrices d'exopolysaccharides dans la rhizosphère du blé dur". Nancy 1, 1996. http://www.theses.fr/1996NAN10093.

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Deux souches bactériennes taxonomiquement éloignées produisant des exopolysaccharides (EPS) de nature très différente, ont été isolées de la rhizosphère du blé dur cultive sur un vertisol marocain. L’une de ces deux souches appartient à l'espèce Agrobacterium radiobacter et produit un succinoglycane. L’identification phénotypique de l'autre souche (microplaques Biolog et Biotype 100) a été complétée par une cartographie du gène codant pour l'ARNr 16S. Tout indique que cette souche appartient à une espèce très proche de Pantoea agglomerans. La composition de son exopolysaccharide ne semble pas classique. L’inoculation de ces deux souches A. Radiobacter NAT200 et Pantoea sp. NAS206 a des semences de blé et dans le sol, préalablement stérilisé ou non, a permis en référence a des échantillons témoins non inoculés de montrer leur impact sur la structuration du sol adhérant aux racines. En colonisant fortement la rhizosphère du blé, ces bactéries productrices d'EPS ont augmenté d'une manière importante la masse de sol adhèrent par unité de biomasse racinaire (SA/RA). Cet effet était plus net dans le cas de la souche NAS206 qui non seulement permettait au sol de mieux adhérer aux racines de blé, mais également stimulait la macroporosité de ce sol (diamètre des pores compris entre 10 et 13 µm). L'évaluation de l'effet de l'inoculation de la souche la plus efficiente (NAS206) à différentes humidités du sol, a montré que le maximum de colonisation de la rhizosphère du blé a été atteint dans des conditions de stress hydrique (16% d'humidité du sol), mais que l'activité agrégeante de cette bactérie productrice d'EPS n'était significative qu’à des humidités du sol supérieures à 19%.

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Cramer, Patricia Catherine. "Modeling Florida panther movements to predict conservation strategies in north Florida". Connect to this title online, 1999. http://purl.fcla.edu/fcla/etd/amj9950.

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Smith, Logan Cutler. "The Panther Dancer". Cleveland State University / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=csu1462981865.

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du, Preez Byron Dennis. "The impact of intraguild competition with lion (Panthera leo) on leopard (Panthera pardus) behavioural ecology". Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:6c17014e-2c58-40e5-866e-d1ce88fe0e89.

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Single-species research dominates the field of ecology; however there is a growing appreciation of the importance of a multi-species approach to holistic conservation. Carnivores exert a top-down control on other species, and are vital components of stable ecosystem functioning. Physiologically adapted for predation upon other animals, competition between carnivores can be particularly aggressive; frequently resulting in mortality, and even population suppression. Big cat research has historically focused on those species that are most easily observable; in particular the lion Panthera leo. The majority of the Felidae however are secretive and elusive, and receive relatively little scientific attention. In particular, there are few data available that measure the effect of direct intraguild interactions between carnivores. Using leopards Panthera pardus as a model species, this research aimed to investigate the impact of lions on the behavioural ecology of a socially subordinate carnivore. Leopards are the most abundant large carnivore in Africa, and have the largest global range of all felids; their ecological niche overlapping with that of both lions and tigers. The knowledge gained from examining their competitive interactions is therefore widely relevant, and may be applicable to other subordinate carnivore species that remain unstudied. Biotelemetry and camera-trap data were modelled using novel algorithms to show that lions impact on leopard population density, demographics and spatial ecology. Faecal analyses suggest that dietary niche segregation may facilitate sympatry. These results indicate the level of impact that large carnivores can exert over smaller species, and the potential for a focus on single-species conservation to undermine holistic conservation. The manifestation of intraguild competition has a significant influence on an animal’s ecology; leopards are generalist species that cope with persecution by adapting their behaviour and niche. Ecological specialists may not fare as well under competitive pressure, and proactive conservation initiatives may be required for endangered species.

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Richter, Thomas. "Untersuchungen zur den lokalen Panthea Süd- und Mittelbabyloniens in altbabylonischer Zeit /". Münster : Ugarit Verlag, 2004. http://catalogue.bnf.fr/ark:/12148/cb40038437g.

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Trinkel, M., P. Funston, M. Hofmeyr, S. Dell, C. Packer e R. Slotow. "Inbreeding and density-dependent population growth in a small, isolated lion population". The Zoological Society of London, 2010. http://encore.tut.ac.za/iii/cpro/DigitalItemViewPage.external?sp=1001446.

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Abstract In South Africa, more than 30 small, enclosed game reserves have reintroduced lions over the last two decades, which now house more than 500 individuals. There is a high risk of inbreeding in these fragmented, fenced and isolated populations, which may be compounded by a lack of management guidelines. A population of 11 founder lions Panthera leo was reintroduced to Madikwe Game Reserve in 1995, and this population has in turn become a source for reestablishing other populations. Only four lineages were reintroduced, founder males were related to founder females, and since 1997, only one male lineage maintained tenure for 49 years, resulting in breeding with direct relatives. Interventionist management to limit lion population growth and inbreeding in Madikwe has taken the form of translocating, trophy hunting and culling of mainly sub adult lions. Despite this management, inbreeding started 5 years after reintroduction. Reproductive performance and thus population growth in Madikwe were dependent on the overall lion population density. When lion density was low, females first gave birth at a significantly younger age and produced larger litters, resulting in a high population growth rate, which decreased significantly when lion density in the park reached carrying capacity, that is, 61 lions. This might have profound consequences for future reestablishment of lion populations when restocking new reserves: our study illustrates the need for founder populations of reintroduced endangered predator species to be as large and genetically diverse as possible, and thereafter new genetic material should be supplemented. The development of such management guidelines is becoming very important as large predator populations become increasingly fragmented and managed as metapopulations.

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