Tesis sobre el tema "Clostridium botulinum"
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Sharma, Davinder Kumar. "Toxin production by Clostridium botulinum". Thesis, University of East Anglia, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.301991.
Texto completoResumen
The endopeptidase activity assay developed for measurement of purified botulinum neurotoxin type A (BoNT/A) in clinical therapeutic preparations has been adopted to provide a specific measure of BoNT/A activity in culture supernatants of proteolytic C. botulinum type A. Electrophoretic studies and inhibition of BoNT/A activity by anti-A antibody confirmed the specificity of the assay. The minimum detection limit was 0.2 MLD50/ml indicating the assay as more sensitive than the standard mouse bioassay or any other in vitro assay available to date. Whilst the assay did not exhibit any cross reactions with non-proteolytic (saccharolytic) clostridia, proteolytic C. botulinum types B and F and C. sporogenes showed some cross reactions. The endopeptidase assay was used to investigate physiological aspects of BoNT/A production by proteolytic C. botulinum type A strain NCTC 7272. Growth studies at 15°C, 25°C and 37°C with strain NCTC 7272 demonstrated that the first appearance of BoNT/A (0.1-1.0 MLD50 ml) occurred during mid-late exponential or early stationary phase of growth. Extracellular BoNT/A formation was not proportional to viable count. Slightly more BoNT/A was detected at 25°C than 37° or 15°C. The results of BoNT/A formation by one of the growth curves at 25°C measured by the endopeptidase assay and mouse bioassays were very similar confirming the specificity of the assay. A simple method was developed to lyre the cells so that BoNT/A formation could be subsequently measured in the endopeptidase assay. The data obtained following lysis of cells and measurement of intracellular BoNT/A showed that both intracellular BoNT/A and total BoNT/A formation is not constitutive but are more closely proportional to viable count than extracellular BoNT/A. Release of BoNT/A from cells was not associated with autolysis. The conversion of BoNT/A from the single-chain to dichain form during growth has been measured. The use of the endopeptidase assay has been also exploited to study BoNT/A formation by this strain within the population of cells. There was only a four-fold difference in BoNT/A production by cells of strain NCTC 7272, and further work in this area is warranted. Attempts were made to use MAPs for the production of monoclonal antibodies to SNAP-25 following cleavage by BoNT/E. Whilst the outcome was unsuccessful, the soundness of the principle was demonstrated
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Raffestin, Stéphanie. "Régulation de la toxinogenèse chez Clostridium botulinum et Clostridium tetani". Paris 7, 2005. http://www.theses.fr/2005PA077044.
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Davis, Tom Owen. "Regulation of botulinum toxin complex formation in Clostridium botulinum : type A NCTC 2916". Thesis, Open University, 1998. http://oro.open.ac.uk/57744/.
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Genomic DNA fragments encoding the silent type B neurotoxin gene from Clostridium botulinum NCTC 2916 have been cloned and the complete nucleotide sequence determined. The translated sequence revealed that the gene encoded a neurotoxin which was closely related to type B neurotoxin genes from Group I Clostridium botulinum. However among the nucleotide sequence differences, aG to T transition has interrupted the coding sequence with the formation of a stop codon. In addition the deletion of an adenine residue has resulted in a frame-shift mutation. Analysis of the DNA sequence contiguous with the silent type B neurotoxin gene revealed the presence of a gene encoding a Nontoxic-Nonhaemagglutinin protein which appears to share a bicistronic mRNA transcript with the type B neurotoxin gene. In the reverse orientation, the partial sequence of a gene encoding a haemagglutinin protein was found, typical of type A and B botulinal neurotoxin complexes. Separating the genes encoding the 'components of the neurotoxin complex was a gene of 178 amino acids which possessed features commonly associated with transcriptional factors. To facilitate the in vivo study of botulinal neurotoxin complex regulation, a gene transfer system using clostridial components has been developed. The minimal replicon of the cryptic plasmid pCB 102 from Clostridium butyricum NCIB 7423 was located to 1.6 kb DNA fragment by deletion analysis, enabling the identification of hitherto undiscovered putative ORFs and secondary structures, consistent with a replicative function. The replicon has been incorporated in to a number of Escherichia coli vectors resulting in a versatile series of shuttle vectors which have demonstrated high structural and segregational stabilities in a heterologous host Clostridium beyerinckii NCI NIB 8052. Gene transfer of a Group I Clostridium botulinum type A strain was demonstrated with a representative pCB 102-derived shuttle vector, pMTL540E. In addition, a 5.9 kb plasmid indigenous to C. hotulimun NCTC 2916 was cloned and the complete nucleotide sequence determined. Eight putative ORFs have been identified, including a putative replication protein and recombinase.
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Cooksley, Clare Marie. "Characterisation of a putative agr system in Clostridium botulinum and Clostridium sporogenes". Thesis, University of Nottingham, 2008. http://eprints.nottingham.ac.uk/11463/.
Texto completoResumen
Botulinum neurotoxin induces a potentially fatal paralytic condition in humans and various animal species collectively known as 'botulism'. It consequently poses a major problem to the food industry, due to the ability of its spores to survive in cooked foods. The incidence of wound botulism has also suffered a recent increase in the UK. The genome sequence of the C botulinum Group I strain ATCC 3502 has recently been determined. In silico analysis has revealed the presence of two distinct loci capable of encoding proteins with homology to AgrB and AgrD of the Staphylococcus aureus agr quorum sensing system. The functional characterisation of these genes has been carried out in order to determine whether they play a role in quorum sensing. To simplify laboratory procedures, C. sporogenes, the non-toxic counterpart of C. botulinum, was initially focused on. The agr regions in C. sporogenes were sequenced and their proteins compared with those of C. botulinum and other Gram-positive bacteria. Regions of conservation were observed amongst the clostridia and, to a lesser extent, between clostridia and staphylococci. Transcriptional linkage assays showed some of the genes of the C sporogenes agr regions to be co-expressed, and Real-Time RT-PCR demonstrated the maximal expression of these genes during late exponential growth. Modulation of the expression of the identified agr genes is a prerequisite to determining their function. Due to an initial lack of an effective gene knockout tool, antisense RNA expression was used for this purpose in C sporogenes, and showed that down regulation of the agrB genes affects sporulation. The development of an integrative vector system for gene inactivation in C sporogenes was also attempted. Knockout mutants in C botulinum and C sporogenes were later constructed using the newly-developed ClosTron system. These mutants were used to demonstrate that AgrDl, AgrD2 and an orphan sensor kinase protein all play a role in the control of sporulation in C. botulinum and C. sporogenes.
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Hielm, Sebastian. "Molecular detection, typing and epidemiology og Clostridium botulinum". Helsinki : University of Helsinki, 1999. http://ethesis.helsinki.fi/julkaisut/ela/elint/vk/hielm/.
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Connan, Chloé. "Neurotoxinogénèse et Passage des neurotoxines botuliques à travers la barrière intestinale". Thesis, Paris 11, 2013. http://www.theses.fr/2013PA114830/document.
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Les neurotoxines botuliques (BoNTs), produites par C. botulinum, sont responsables du botulisme humain et animal. Dans sa forme naturelle, le botulisme résulte le plus souvent d’une absorption des toxines botuliques à partir du tube digestif après ingestion d’aliments contaminés par la toxine et C. botulinum. L’intoxination peut être divisée en 4 grandes étapes : production de toxine par la bactérie, ingestion d’aliments contenant la toxine préformée, passage de la neurotoxine à travers la barrière intestinale et action protéolytique aux terminaisons nerveuses. La régulation de la production des toxines et le passage des neurotoxines botuliques à travers la barrière intestinale sont mal compris. BoNT s’associe à des protéines non toxiques (NAPs) pour former des complexes de différentes tailles. Les gènes codant les BoNTs et NAPs sont regroupés sur le locus botulique et leur expression est contrôlée positivement par le facteur sigma alternatif BoTR/A. La toxinogénèse chez C. botulinum est contrôlée par un réseau complexe de régulateurs incluant au moins 3 systèmes à deux composants (TCS), identifiés pas la méthode d’ARN antisens, qui régulent positivement la production de complexe botulique indépendamment de BoTR/A. D’autre part, l’entrée de BoNT/B dans la barrière intestinale a été suivie à l’aide du fragment HcB marqué en fluorescence dans une anse intestinale ligaturée de souris. Des analyses en microscopie à fluorescence, immunohistochimie et microscopie électronique ont permis de mettre en évidence que HcB transcytose à travers les entérocytes par une voie d’endocytose dépendante de la dynamine. HcB cible les terminaisons nerveuses acétylcholinergiques de la lamina propia des villosités et gagne les neurones acétylcholinergiques et sérotoninergiques de la sous-muqueuse et de la musculeuse en seulement 10 minutes. Une étude in vitro réalisée sur cellules intestinales (m-ICcl2) montre que l’entrée de HcB est dépendante de récepteurs gangliosidiques GD1b/GT1b présents à la surface des cellules mais pas de la synaptotagmine II qui est requise pour l’entrée de BoNT/B dans les cellules neuronalesBotulinum neurotoxins (BoNTs), produced by C. botulinum, are responsible for animal and human botulism. In its natural form, botulism is mostly acquired after absorption of BoNTs in the digestive tract after ingestion of food contaminated with C. botulinum and its toxins. The intoxination can be divided in 4 major steps: toxin production, ingestion of food contaminated with BoNTs, passage of BoNTs through the intestinal barrier, and proteolytic activity on nerve endings. Regulation of toxin production and passage of BoNTs through the intestinal barrier are poorly understood. BoNT associates with non toxic protein (NAPs) to form complexes of various sizes. The BoNTs and NAPs genes are clustered in the botulinum locus and are positively regulated by an alternative sigma factor BotR/A. Toxinogenesis in C. botulinum is regulated by a complex regulatory network containing at least 3 two components systems (TCS), identified by antisens RNA strategy, which regulate the production of botulinum complex independently of BotR/A. On the other hand, BoNT/B entry was monitored with fluorescent HcB fragment in ligatureted mouse intestinal loop. Fluorescent imaging analysis, immunohistochemistry and electron microscopy, have evidence that HcB is transcytosed through enterocytes cells by an endocytosis dynamin dependant. HcB targets acetylcholinergic nerves localized in lamina propria of villi then reaches serotoninergic and acetylcholinergic nerve endings in the submucosa and musculosa within 10 minutes. In vitro experiments performed on intestinal cell line (m-ICcl2) shows that the endocytosis of HcB is dependent on the GD1b/GT1b gangliosidic receptors on the cell surface but not on the synaptotagmine II protein which is recquiered HcB entry in neuronal cells
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Masuyer, Geoffrey. "Structure and activity of Clostridium botulinum neurotoxin functional fragments". Thesis, University of Bath, 2012. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.557825.
Texto completoResumen
Botulinum neurotoxins (BoNTs) cause flaccid paralysis by inhibiting neurotransmission at cholinergic nerve terminals. BoNTs consist of three essential domains for toxicity: the cell binding domain (Hc), the translocation domain (Hn) and the catalytic domain (LC). The binding function of the Hc domain is essential for BoNTs to bind the neuronal cell membrane, therefore removal of the Hc domain results in a product that retains the endopeptidase activity of the LC but is non-toxic. Functional derivatives (LHn) of the parent neurotoxin composed of Hn and LC domains have been recombinantly produced and characterised. The crystallographic structures of LHn from serotypes A and B are reported here and demonstrate the stability of the LHn fragment in comparison to the full length toxins. The activity of LHn has been assessed on recombinant substrates and on cultured neuronal cells. LHn retains the capacity to internalise and cleave its intracellular SNARE substrate when applied to the cells at high concentration. These activities demonstrate the utility of engineered botulinum neurotoxin fragments as analytical tools to study the mechanisms of action of BoNT neurotoxins and of SNARE proteins. Targeted secretion inhibitors (TSI) are a new class of engineered biopharmaceutical molecules derived from the botulinum neurotoxins. These functional derivatives are expressed as single-chain proteins and require post-translational activation into di-chain molecules for function. A range of BoNT derivatives are presented and demonstrate the successful use of engineered SNARE substrate peptides at the LC-Hn interface to give these molecules self-activating capabilities while retaining the functions of LHn. Several novel molecules with therapeutic potential have been produced and their crystallisation for structural investigation is reported. These results provide an understanding of the structural implications and challenges of engineering therapeutic molecules that combine functional properties of the LHn fragment from BoNTs with specific ligand partners to target different cell types.
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Woudstra, Cedric. "Clostridium botulinum, du génotypage de la toxine en passant par les flagellines jusqu'au séquençage de génomes : un aperçu de la diversité génétique des Clostridies associés au botulisme animal et humain". Thesis, Paris Est, 2016. http://www.theses.fr/2016PESC1020/document.
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Le botulisme est une maladie nerveuse, commune à l’homme et aux animaux, due à l’action de la toxine botulique produite par Clostridium botulinum. Il existe 8 types de toxines dénommées A à H. Les bactéries capables de produire cette toxine se différencient en six groupe sur la base de leurs caractéristiques phénotypiques et biologiques. Les souches de C. botulinum responsables du botulisme humain appartiennent aux groupes I et II selon qu’elles soient protéolytiques ou non. Elles produisent les toxines A, B, E et F, ainsi que le nouveau type H récemment découvert. C. butyricum et C. baratii sont également capables de produire les toxines botuliques de type F et E et appartiennent au groupe V et VI. C. argentinense appartient au groupe IV et est capable de synthétiser la toxine de type G. Elle a été soupçonnée d’être impliquée dans des cas de botulisme infantile en Argentine. Les souches de C. botulinum responsables du botulisme animal appartiennent au groupe III (C. novyi sensu lato) et produisent les toxines C, D et leurs formes mosaïques C/D et D/C. La toxine botulique est le poison le plus puissant connu à ce jour. La dose létale nécessaire pour tuer une personne en bonne santé par intoxication alimentaire est de 70 µg seulement. C’est pourquoi cette toxine a fait l’objet d’études particulièrement approfondies, notamment celles impliquées dans des cas de botulisme humain. Elle peut également être utilisée pour le traitement de certaine pathologie ou la chirurgie esthétique (Botox). Malheureusement, elle peut également être utilisée à mauvais escient, en tant qu’arme de guerre ou à des fins de bioterrorisme. C’est pourquoi l’emploi de la toxine botulique ou de sa bactérie productrice fait l’objet d’une législation particulièrement stricte. Mon projet de doctorat s’est organisé autour de plusieurs projets de recherche visant à développer des méthodes de détection et de typage de du germe et de sa toxine (projets Européens BIOTRACER et AniBioThreat ; projets NRBC-bio ; LNR botulisme aviaire en France). Lors de mes recherches j’ai concentré mon travail sur le développement de méthodes capable de suivre et remonter à la source d’une contamination, qu’elle soit délibérée, accidentelle ou naturelle. Afin d’y parvenir j’ai investigué les gènes des flagellines de C. botulinum groupe I à III, responsables du botulisme humain et animal. L’analyse des gènes flaA et flaB a mis en évidence 5 groupes majeurs et 15 sous-groupes, certain étant spécifiques de régions géographiques. FlaB s’est montré spécifique de C. botulinum type E. Les gènes flagellines fliC, spécifiques à C. botulinum du groupe III, se divisent 5 groupes, avec fliC-I et fliC-IV associés aux types mosaïques C/D et D/C. J’ai étudié la prévalence des souches productrices de toxine de type mosaïques chez les volailles et les bovins. Les résultats montrent que les types C/D et D/C sont majoritaires en Europe. Enfin, j’ai séquencé 17 génomes provenant de souches responsables de botulisme animal en France (14 types C/D et 3 types D/C). Leur analyse montre que ces souches sont très proche génétiquement, entre elles et avec les souches Européennes. Grâce à ces données j’ai mis en évidence un large contenu extra chromosomique dans les souches C/D, qui peut être utilisé pour créer une carte d’identité génétique. D’autre part, l’étude des séquences Crisps à des fins de typage ne s’est pas avérée suffisamment résolutive, du fait de système Crispr-Cas déficient chez les souches C/D. Enfin, un très haut degré de discrimination a été atteint par typage SNP, qui a permis de distinguer jusqu’à l’origine de chaque souche. L’ensemble de ces résultats est développé dans le présent manuscritClostridium botulinum is the etiologic agent of botulism, a deadly paralytic disease that can affects both human and animals. Different bacteria, producing neurotoxins type A to H, are responsible for the disease. They are separated into different groups (I to VI) on the basis of their phenotypical and biological characteristics. Human botulism is mainly due to Groups I and II producing neurotoxins A, B, E and F, with type H recently discovered. Also C. butyricum and C. baratii species (Groups V and VI), producing toxins type F and E respectively, are scarcely reported. C. argentinense Group IV, producing toxin type G, which has been suspected to be associated with infant botulism in Argentina. Animal botulism is mainly due to Group III, which is constituted by C. novyi sensu lato species. They produce toxin types C, D and their mosaic variants. Botulinum neurotoxins are the most powerful toxin known to date with as little as 70 µg enough to kill a person by food poisoning. Therefore, it received a great deal of attention. Botulinum neurotoxins have been deeply studied, especially human related toxins compared to animal. The toxins found to be useful for medical or cosmetic (Botox) treatments, but it was also used as a biological warfare agent, and for bioterrorism. Its extreme potency is equal to its dangerousness. Therefore, governments show concerns of its potential misuse as a bioterrorism weapon; research programs are funded to study and raise awareness about both the toxins and the producing organisms. My PhD work was structured by the different projects I was involved in, which were related to C. botulinum detection and typing, like BIOTRACER and AniBioThreat European projects, the French national CBRN program, or the NRL for avian botulism. The main transversal objective I followed lead me to develop new methods to trace back the origin of C. botulinum contamination, in case of a deliberate, accidental or naturally occurring botulism outbreak. I investigated flagellin genes as potential genetic targets for typing C. botulinum Group I-II and III, responsible for human and animal botulism respectively. Flagellin genes flaA and flaB showed the investigated C. botulinum Group I and II strains to cluster into 5 major groups and up to 15 subgroups, some being specific for certain geographical areas, and flaB being specific to C. botulinum type E. Flagellin fliC gene investigated in C. botulinum Group III showed to cluster into five groups, with fliC-I and fliC-IV associated to type C/D and D/C respectively, being not discriminative enough to differentiate highly genetically related strains. I also studied the prevalence of mosaic toxin genes in C. botulinum Group III in animal botulism, mainly in poultry and bovine. The results brought out the mosaic toxin types C/D and D/C to be predominant in the samples investigated throughout Europe. Finally, I explored the full genome sequences of 14 types C/D and 3 types D/C C. botulinum Group III strains, mainly originating from French avian and bovine botulism outbreaks. Analyses of their genome sequences showed them to be closely related to other European strains from Group III. While studying their genetic content, I was able to point out that the extrachromosomal elements of strains type C/D could be used to generate a genetic ID card. Investigation of Crispr typing method showed to be irrelevant for type C/D, due to a deficient Crispr-Cas mechanism, but deserve more investigation for type D/C. The highest level of discrimination was achieved while using SNP core phylogeny, which allowed distinguishing up to the strain level. Here are the results I’m going to develop in this manuscript
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Bradbury, Mark. "Genomic and flow cytometric studies of Clostridium sporogenes, a non-toxigenic surrogate for Clostridium botulinum". Thesis, Federation University Australia, 2014. http://researchonline.federation.edu.au/vital/access/HandleResolver/1959.17/97216.
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Clostridium sporogenes and Group I Clostridium botulinum are two bacterial species belonging to the same phylogenetic group, primarily differentiated by the expression of botulinum neurotoxin. Both organisms are of significant commercial importance in regards to the spoilage of and/or disease in thermally processed food products due to their ability to form heat resistant spores. As such, these species are often used as the target organism for the design of thermal inactivation processes, particularly in regards to thermal sterilisation. Two specific aspects pertaining to these organisms were investigated to further enhance knowledge with respect to their use in thermal processing studies: the genetic relationship between C. sporogenes and Group I C. botulinum; and the mechanism of heat resistance in spores of C. sporogenes. The first part of this thesis describes the assembly, annotation and analysis of the draft genome of C. sporogenes PA 3679 (the most widely used surrogate for Group I C. botulinum). These data allowed identification of unique variants genes related to spore germination, analogous toxin regions and mobile elements between species. MLST analysis revealed that phylogeny was an ineffective indicator of toxigenicity in this group and thus prompted a pan-genomic analysis. The pan-genome of C. sporogenes/Group I C. botulinum was found to consist of 8799 coding sequences (CDS’s) and a core genome consisting of 1590 CDS’s. Analysis of this pan-genome revealed the significant role that mobile genetic elements have played in genetic diversity within this group of organisms. The second part of this thesis investigated the heat inactivation of C. sporogenes PA 3679 spores in regards to structural variation and population heterogeneity. A novel flow cytometric approach was developed and utilised to investigate isothermal spore inactivation; and implications of the impact of NaCl on the intrinsic variability throughout this process and during a subsequent recovery period. Based on these approaches a possible mechanistic description for the thermal inactivation of spores was developed. Together, these studies present significant evidence supporting the continued suitability of C. sporogenes as a surrogate for Group I C. botulinum, whilst also enhancing the understanding of clostridial spore inactivation during a moist heat process.Doctor of Philosophy
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Bucknavage, Martin M. "Growth and survival of Clostridium botulinum type E in pasturized oysters". Thesis, This resource online, 1988. http://scholar.lib.vt.edu/theses/available/etd-04122010-083636/.
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LA, TORRE ANGELA. "Studi molecolari del processo di germinazione in Clostridium sporogenes, modello non-patogeno di Clostridium botulinum". Doctoral thesis, Università Cattolica del Sacro Cuore, 2016. http://hdl.handle.net/10280/10793.
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Quando le condizioni sono sfavorevoli alla crescita, membri dei generi Bacillus e Clostridia (incluso Clostridium botulinum, l’agente eziologico del botulismo) formano endospore, forme cellulari estremamente resistenti, metabolicamente dormienti e difficili da distruggere. Tuttavia, le spore attraverso il processo di germinazione, riattivano il ciclo vegetativo non appena le condizioni tornano favorevoli. Questa capacità di “riattivazione” delle spore è causa di “food spoilage” e di intossicazioni alimentari.
Considerando che le specie Clostridium botulinum e Clostridium sporogenes sono filogeneticamente correlate, in questo lavoro, il ceppo Clostridium sporogenes UC9000, isolato da latte crudo, è stato utilizzato come modello non-patogeno di Clostridium botulinum per studiare la germinazione.
Studi fisiologici hanno rivelato che le spore del ceppo UC9000 germinano in presenza di L-alanina/ L-cisteina in combinazione con L-lattato, mentre un analisi in silico ha permesso di identificare omologhi dei recettori coinvolti nella risposta all’L-alanina in Bacillus. Attraverso l’analisi del genoma sono stati identificati gli enzimi SleB, CwlJ e SleL, responsabili della degradazione del cortex. CwlJ è stato localizzato nel coat della spora grazie ad uno studio di proteomica, è stato espresso in forma solubile in E. coli ed un test di attività in vitro ha evidenziato la sua capacità di indurre la germinazione di spore “decoated”When environmental conditions are unfavorable to the growth, Bacillus and Clostridium bacteria (including Clostridium botulinum, the causative agent of foodborne botulism) form endospores, metabolically dormant cell types resistant to several adverse conditions and difficult to kill. However, under suitable conditions, spores resume the vegetative life by triggering the germination process. Thus, spores are dangerous agents of human foodborne disease and food spoilage. In this work, the strain Clostridium sporogenes UC9000, isolated from raw milk, was used like not-pathogenic model of Clostridium botulinum to better understand the mechanisms underpinning the Clostridium germination. Clostridium sporogenes is a species phylogenetically related to Clostridium botulinum and often used like its surrogate. Physiological studies revealed that UC9000 spores germinate in presence of L-alanine/L-cysteine in combination with L-lactate, while in silico analyses allowed the identification of homologues of the Bacillus germinant receptors responsive to L-alanine. The genome screening also detected genes coding for SleB, CwlJ and SleL, enzymes participating to the cortex degradation. CwlJ was found resident in the spore coat by performing a proteomic analysis, it was expressed in soluble form in E. coli and an in vitro assay of activity revealed its capability to induce germination when added exogenously to decoated spores
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LA, TORRE ANGELA. "Studi molecolari del processo di germinazione in Clostridium sporogenes, modello non-patogeno di Clostridium botulinum". Doctoral thesis, Università Cattolica del Sacro Cuore, 2016. http://hdl.handle.net/10280/10793.
Texto completoResumen
Quando le condizioni sono sfavorevoli alla crescita, membri dei generi Bacillus e Clostridia (incluso Clostridium botulinum, l’agente eziologico del botulismo) formano endospore, forme cellulari estremamente resistenti, metabolicamente dormienti e difficili da distruggere. Tuttavia, le spore attraverso il processo di germinazione, riattivano il ciclo vegetativo non appena le condizioni tornano favorevoli. Questa capacità di “riattivazione” delle spore è causa di “food spoilage” e di intossicazioni alimentari.
Considerando che le specie Clostridium botulinum e Clostridium sporogenes sono filogeneticamente correlate, in questo lavoro, il ceppo Clostridium sporogenes UC9000, isolato da latte crudo, è stato utilizzato come modello non-patogeno di Clostridium botulinum per studiare la germinazione.
Studi fisiologici hanno rivelato che le spore del ceppo UC9000 germinano in presenza di L-alanina/ L-cisteina in combinazione con L-lattato, mentre un analisi in silico ha permesso di identificare omologhi dei recettori coinvolti nella risposta all’L-alanina in Bacillus. Attraverso l’analisi del genoma sono stati identificati gli enzimi SleB, CwlJ e SleL, responsabili della degradazione del cortex. CwlJ è stato localizzato nel coat della spora grazie ad uno studio di proteomica, è stato espresso in forma solubile in E. coli ed un test di attività in vitro ha evidenziato la sua capacità di indurre la germinazione di spore “decoated”When environmental conditions are unfavorable to the growth, Bacillus and Clostridium bacteria (including Clostridium botulinum, the causative agent of foodborne botulism) form endospores, metabolically dormant cell types resistant to several adverse conditions and difficult to kill. However, under suitable conditions, spores resume the vegetative life by triggering the germination process. Thus, spores are dangerous agents of human foodborne disease and food spoilage. In this work, the strain Clostridium sporogenes UC9000, isolated from raw milk, was used like not-pathogenic model of Clostridium botulinum to better understand the mechanisms underpinning the Clostridium germination. Clostridium sporogenes is a species phylogenetically related to Clostridium botulinum and often used like its surrogate. Physiological studies revealed that UC9000 spores germinate in presence of L-alanine/L-cysteine in combination with L-lactate, while in silico analyses allowed the identification of homologues of the Bacillus germinant receptors responsive to L-alanine. The genome screening also detected genes coding for SleB, CwlJ and SleL, enzymes participating to the cortex degradation. CwlJ was found resident in the spore coat by performing a proteomic analysis, it was expressed in soluble form in E. coli and an in vitro assay of activity revealed its capability to induce germination when added exogenously to decoated spores
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Preising, Claudia. "Literaturstudie zum Vermehrungs- und Toxinbildungs- vermögen von Clostridium botulinum, zu den Eigenschaften des Botulinumtoxins sowie zum Vorkommen und zur Tenazität der Clostridium botulinum-Sporen". Doctoral thesis, Universitätsbibliothek Leipzig, 2006. http://nbn-resolving.de/urn:nbn:de:swb:15-20070118-093729-8.
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1) Einleitung 2) Pathogenese und Klinik des Botulismus 3) Vorkommen von C. botulinum und des Botulismus 4) Beeinflussung von Spore, vegetativer Zelle und BoNT 5) Diagnostik, Therapie und Prophylaxe 6) Diskussion
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Bruhne, Lars. "Untersuchungen zur Beziehung zwischen positivem Clostridium botulinum Antikörper-Nachweis, ausgewählten Stoffwechselparametern, Akute-Phase-Proteinen und Erkrankungshäufigkeiten, Herdengröße sowie Herdenmilchleistung von Milchrindern". Doctoral thesis, Universitätsbibliothek Leipzig, 2015. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-171930.
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Brown, Robert Christopher. "Development of a novel expression system in Clostridium perfringens". Thesis, Open University, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.321561.
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Marvaud, Jean-Christophe. "Contribution a l'etude des proteines associees aux neurotoxines clostridiennes et vectorisation de proteines dans les cellules (doctorat : microbiologie)". Paris 11, 1998. http://www.theses.fr/1998PA114851.
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Cadieux, Brigitte. "Development of a novel, rapid, in vitro assay for the detection of Clostridium botulinum neurotoxin type E". Thesis, McGill University, 2001. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=32836.
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Botulism is a foodborne intoxication caused by ingestion of Clostridium botulinum neurotoxin (BoNT). Preliminary studies focussed on the production of polyclonal antisera against BoNT/E by immunizing a rabbit with botulinal toxoid type E. The antiserum was subsequently used to detect BoNT/E using the slot blot immunoassay where samples were applied to a slot blot filtration manifold and drawn by vacuum through a membrane. The membrane was then immunologically processed before chemiluminescent detection. However, the antisera lacked specificity and cross-reacted with closely related clostridia strains.The specificity of the antisera was increased by adsorbing cross-reactive antibodies from whole antisera with affinity columns made with total proteins from culture supernatants of closely related clostridia. Alternatively, specific antibodies were isolated with an affinity column prepared with C. botulinum type E toxoid.
Different methods of concentrating BoNT/E in each sample prior to testing them were evaluated to increase the sensitivity of the assay.
The slot blot immunoassay was then evaluated for detection of BoNT/E in mixed cultures and in food samples. (Abstract shortened by UMI.)
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Alberto, François. "Etude physiologique et moléculaire de la germination des spores clostridium botulinum". Aix-Marseille 3, 2003. http://www.theses.fr/2003AIX30009.
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L'objectif de cette thèse est de contribuer à la compréhension des mécanismes physiologiques et moléculaires de la germination des spores de Clostridium botulinum. Cette bactérie anaérobie stricte est capable de produire des spores, formes de dormance, quand les conditions environnementales deviennent défavorables. Ces spores peuvent survivre très longtemps et sont résistantes à de nombreux traitements physico-chimiques. La germination est définie comme le phénomène permettant à une spore en dormance de donner une cellule végétative métaboliquement active. Les spores bactériennes germent en réponse à de petites molécules appelés germinants. La première étape du travail a consisté à établir les principales conditions de germination de différentes souches de C. Botulinum mésophiles. Ainsi ces bactéries germent, à 30ʿC, en présence de L-alanine et autres acides aminés hydrophobes. Cette réponse est, dans la majorité des cas, améliorée par l'ajout de L-lactate. La température d'incubation des spores influence fortement la germination. La recherche du support moléculaire des récepteurs putatifs des germinants a permis de mettre en évidence, chez C. Botulinum Beans B, un opéron de 3 gènes homologues aux gènes de l'opéron gerA de germination codant pour le récepteur de la L-alanine chez Bacillus subtilis. Il a été mis en évidence une expression pendant les phases tardives de la sporulation sous contrôle du facteur sG. Des expériences de localisation de la protéine GerAB, dans les différentes structures de la spore, ont permis de mettre en évidence la présence de la protéine au niveau de la membrane interne de la spore. Les résultats obtenus chez C. Botulinum ainsi que les données connues chez d'autres espèces sporulées suggèrent que le mécanisme des phases précoces de germination est relativement conservé chez ces bactériesThe aim of this work is to elucidate some of the physiological and molecular mechanisms implicated in Clostridium botulinum spore germination. This strictly anaerobic bacterium is able to produce dormant spores when the environmental conditions become unfavourable. Spores can survive for a long time and are resistant to a wide range of physical and chemical treatments. Germination is defined as the conversion of a dormant spore into a metabolic active vegetative cell. Bacterial spores germinate in response to small molecules (amino acids, sugars, ions) called germinants. The first step of this work was to determine the main conditions of germination for different strains of mesophylic C. Botulinum. It was demonstrated that C. Botulinum spores germinate at 30ʿC in response to an induction by L-alanine at various germination rates according to the strain. This response is mainly enhanced by addition of L-lactate as a co-germinant. Only hydrophobic amino acids are able to trigger C. Botulinum spore germination. Spore germination is influenced dramatically by temperature. At 15ʿC, few spores germinate in L-alanine/L-lactate. The molecular analysis of the putative germinant receptors has revealed the presence of a three-gene operon in C. Botulinum Beans B, homologous to genes gerAA, gerAB and gerAC of the Bacillus subtilis gerA operon. Analysis of the gene transcription and mRNA mapping showed that expression is very high during late sporulation and controlled by a sG factor. Western-blot analysis have revealed that GerAB protein is located in inner membrane of the spore. The results obtained with C. Botulinum and the previous data on other sporulating species suggest that the first step of spore germination may be the same in all spore-forming bacteria
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Blount, Benjamin Andrew. "Analysis of the roles and regulation of flagella in Clostridium botulinum". Thesis, University of Nottingham, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.588079.
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Flagella are complex structures with important roles in bacterial motility, virulence and protein secretion. As of yet, little work has been undertaken to characterise the function and regulation of flagella in Clostridium botulinum, an organism which produces an extremely potent neurotoxin which is the causative agent of botulism and is utilised for numerous pharmaceutical purposes. Genes encoding structural components of the flagellar basal body, hook and filament were inactivated using the ClosTron insertional mutation method, resulting in reduced swarming motility and reduced neurotoxin release from the cell into culture supernatant as determined by an ELISA method. To determine regulatory factors affecting flagellar assembly and secretion, sigD, the gene encoding δD, an alternative sigma factor that has been shown to be responsible for the expression of late flagellar genes in various bacterial species, was inactivated. The mutant strain was analysed by microarray and quantitative real-time reverse-transcription PCR methods. It was shown that δD positively regulates the expression of multiple genes including the late flagellar genes, the clostridiolysin S genes and an operon of secreted proteases. Phenotypic assays were performed on the sigD mutant and confirmed that it was null for motility and that proteolysis by secreted proteases was reduced. This reduced proteolysis phenotype was complemented back towards wild-type levels by the introduction of a plasmid containing the sigD gene from Clostridium sporogenes under the control of its native Pft9B promoter. To aid future work, a method for inducing and selecting for double homologous recombination events in C. botulinum was developed for the purpose of creating a strain in which the chromosomal botA gene is converted to a toxoid encoding sequence by allelic exchange. This work has therefore implicated flagella in the secretion of the neurotoxin, characterised the regulatory roles of δD and developed a novel method for allelic exchange in clostridia.
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Ashton, Philip Matthew. "A genomic and proteomic investigation into the Clostridium botulinum neurotoxin complex". Thesis, Open University, 2013. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.664471.
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Clostridium botulinum and some strains of C. baratii and C. butyricum produce one of the most potent toxins known to man, botulinum neurotoxin, and are responsible for the disease botulism. This severe neuroparalytic disease IS the result of botulinum neurotoxin negotiating a complex path to the cholinergic nerve endings. There, it interferes with the release of excitatory neurotransmitters resulting in flaccid paralysis and if untreated, death. The neurotoxin, itself a multi-faceted protein, does not act alone but is produced as part of a large, hetero-multimeric complex with the associated non-toxic proteins. This complex, known to protect the toxin from acids and proteases in the gut, has recently been suggested to play a more active role in toxicity. Here, the proteins from the lesser-studied toxin' complex type (the OrfX type) are shown for the first time to share sequence similarity and syriteny with clusters of proteins that are co-localised with various putative toxin genes in diverse other species. The extracellular supernatant proteome of C. botulinum is characterised and mined for potential novel virulence factors, with metabolic cost of extracellular protein being highlighted as a potential marker of virulence associated extracellular proteins. The supernatant of 22 clinical strains of C. botulinum were investigated for the presence of the toxin complex; all toxin complex components were jdentified in the majority of strains indicating the importance of these proteins in the causation of botulism. A relationship between ' OrfX-encoding strains and infant botulism was also uncovered in clinical strains from the UK. Hypotheses to explain this association are explored. Finally, the transcriptome of C. botulinum was investigated using RNA-sequencing. This uncovered a complex and diverse picture of transcription in C. botulinum and raised questions as to the role of the alternative sigma factor BotR in the regulation of bont.
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Fohler, Svenja [Verfasser]. "Occurrence of Clostridium botulinum neurotoxin genes and toxin-genotypes of Clostridium perfringens in dairy cattle / Svenja Fohler". Hannover : Bibliothek der Tierärztlichen Hochschule Hannover, 2016. http://d-nb.info/1104403897/34.
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Morineaux, Valérie. "Identification et Quantification des Sous-Types de la Neurotoxine Botulique de Type A par Spectrométrie de Masse". Thesis, Paris 11, 2015. http://www.theses.fr/2015PA114826.
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Les toxines botuliques (BoNTs) sont les substances les plus toxiques connues. Elles sont responsables du botulisme, une maladie rare mais le plus souvent mortelle sans prise en charge médicale. Cependant, les applications médicales des BoNTs sont de plus en plus nombreuses du fait de leurs propriétés paralysantes. Leur toxicité par voie inhalée en fait un des 6 principaux agents du risque intentionnel. Les BoNTs, produites par Clostridium botulinum, se répartissent en 7 types sérologiques qui se déclinent en sous-types. Cette biodiversité rend difficile leur identification par les méthodes classiques utilisées pour les toxines protéiques (approches immunologiques). Jusqu’à présent, seule l’analyse génétique permettait de distinguer les différents sous-types entre eux. Dans ce travail a été développée une méthode d’analyse en LC-QqQ-MS/MS en mode MRM pour identifier les différents sous-types de la BoNT/A dans des matrices complexes à partir de peptides communs et spécifiques à ces sous-types. Un traitement d’échantillon par immunocapture sur billes magnétiques couplées à des anticorps anti-peptides a été développé pour isoler la toxine de l’échantillon avant analyse. Des surnageants de culture des sous-types A1 à A3, A5, A7 à A8 ont été utilisés pour valider la méthode. La limite de détection de la méthode est compatible avec les taux de toxine retrouvés habituellement dans les échantillons naturellement contaminés. Cette méthode de spectrométrie de masse a ensuite été utilisée pour quantifier les différents sous-types de la BoNT/A dans une matrice complexe (surnageants de culture de C. botulinum). Une technique de quantification, utilisant un isotope stable de la chaine légère de type A1, ([13C6]K et [13C6]R), a été retenu comme étalon interne. Les différents sous-types de BoNT/A ont été quantifiés dans les surnageants et la quantité de BoNT correspondante à une dose létale minimale de 100% a été déterminée pour chaque sous-typeBotulinum neurotoxins (BoNTs) are the most poisonous substances known. They are responsible for human botulism, a rare but potentially fatal disease if not quickly treated. However, BoNTs were approved for the treatment of numerous medical applications due to their temporary paralysis effects. BoNTs are among the six agents with the highest risk of potential use as bio-weapons because of their high toxicity in aerosol form. BoNTs, produced by Clostridium botulinum, are divised into seven toxinotypes and each toxinotype contains several subtypes. This biodiversity makes more difficult their identification with classical methods by immunological ways. Until now, only molecular genetical methods could differenciate subtypes among them. The aim of this work was to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) in MRM mode to efficiently discrimate the distinct subtypes from specific and common peptides. Immunocapture sample preparation with antipeptides antibodies was used and allowed the isolation of the toxin from the sample. Subtyping was performed with crude supernatants (BoNT/A1 to /A3, /A5, /A7 and /A8) in order to validate the method. Limit of detection (LOD) of the proposed method is in the range of minimal toxin concentration found in naturally contamined samples. In a second part of this work, this mass spectrometry method was used to quantify the neurotoxin in complex matrices (supernatants of Clostridium botulinum cultures). Isotope labeled light chain (13C6]K et [13C6]R) from botulinum A1 neurotoxin was produced and used as internal standart. Subtypes were quantified in supernatants and the quantity of neurotoxin for one minimal lethal dose 100% was determined for each subtype
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Taylor, Reed H. "Conditions Associated with Clostridium sporogenes Growth as a Surrogate for Clostridium botulinum in Non-thermally Processed Canned Butter". BYU ScholarsArchive, 2010. https://scholarsarchive.byu.edu/etd/2447.
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Shelf-stable canned butter is currently available in retail stores, and many home-preservationists promote home-canning of butter. Non-cultured butter is a low-acid canned food, which would presumably require thermal processing. The lack of a thermal process step in canned butter products raises questions of potential safety, because they are hermetically sealed and generally exhibit anaerobic growth conditions, which are optimal for Clostridium botulinum growth. Without thermal processing, low-acid canned foods (LACF) must have inhibitory factors present to prevent C. botulinum growth. Some potential intrinsic inhibitory factors, or "hurdles", within butter include: reduced water activity (aw), acidity (pH) in cultured products, elevated salt content, and the micro-droplet nature of the aqueous phase in the butter emulsion. It was hypothesized that a normal intact butter emulsion would have sufficient "hurdles" to prevent C. botulinum growth, while a broken butter emulsion would result in a larger aqueous phase that would allow for growth. Butter was prepared using a batch churn method with either inoculated or uninoculated cream. Butter samples with four different salt amounts (0, 0.8, 1.6, & 2.4% added NaCl) were prepared and placed in coated aluminum cans for storage. Samples were stored for 1 or 2 week periods at either 22°C or 41°C and then plated for C. sporogenes growth. Samples stored at 41°C showed a significant increase over those stored at 22°C. This growth increase occurred due to incubation near the optimal growth temperature for C. sporogenes and damage to emulsion structure. Furthermore, sodium chloride (NaCl) addition was found to have a significant effect on C. sporogenes growth, with 0.8 % NaCl promoting more growth than 0%, but with decreases in growth beyond 0.8%. Uninoculated control plates were also found to have bacterial growth. This growth was attributed to other anaerobic bacteria present within the cream. It was concluded that removal of the butter structure "hurdle" could result in C. botulinum growth even at elevated salt levels and therefore home preparation of canned butter is not advisable. It is also possible that commercially canned butter, if heat abused, could potentially allow for C. botulinum growth and therefore consumption is not recommended.
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Ruth, Vieira de Lemos Vasconcelos Maria. "Efeito da acidificação com diferentes ácidos sobre as características sensoriais e inibição do Clostridium botulinum no palmito de pupunha em conserva". Universidade Federal de Pernambuco, 2004. https://repositorio.ufpe.br/handle/123456789/9010.
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Previous issue date: 2004A porção comestível do tronco de algumas palmeiras, utilizadas para a produção de palmito é um produto de importância para o Brasil como principal produtor e exportador na forma em coonserva, detendo 85% da produção mundial. A região Nordeste tem se destacado como produtora da variedade pupunha (Bactris gasipaes), nativa da Amazônia, que apresenta vantagens ecológicas por se tratar de palmeira cultivada, com características de perfilhamento, precocidade de produção e qualidade satisfatória. A ocorrência de toxinfecções por Clostridium botulinum provocadas pelo consumo de palmito industrializado levou o Ministério da Saúde a estabelecer a obrigatoriedade do uso de ácido associado à salmoura para produção de palmito com o objetivo de baixar o pH a limites inferiores a 4,5, desfavorecendo o desenvolvimento do Clostridium botulinum, assegurando portanto a saúde do consumidor. Normalmente o ácido cítrico é utilizado para atender as exigências, no entanto admite-se que outros ácidos podem ser utilizados com vantagem quanto aos aspectos sensoriais. Este trabalho teve por objetivo avaliar o efeito da acidificação do palmito de pupunha, cultivado em Pernambuco, com diferentes ácidos sobre as características do produto em conserva. Foram testados os ácidos: cítrico, málico, láctico e tartárico na estabilidade do produto embalado em frascos de vidro com capacidade para 600 ml e quanto às características organolépticas. Os resultados obtidos demonstraram a viabilidade da acidificação do palmito de pupunha com esses diferentes ácidos sem prejuíso nas características físicas, químicas, organolépticas e microbiológicas
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Vogelsgesang, Martin. "Rho-ADP-ribosylierende Exoenzyme von Clostridium botulinum und Clostridium limosum : Untersuchungen zur Substratspezifität und Interaktion mit Ral-GTPasen /". [S.l. : s.n.], 2005. http://www.gbv.de/dms/bs/toc/513916806.pdf.
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Sharkey, Freddie. "Toxin gene expression in Clostridium botulinum type E under different growth conditions". Thesis, University of Ulster, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.274025.
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McGrath, Susan. "The development of an RNA assay for Clostridium botulinum toxin gene expression". Thesis, University of Ulster, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.326309.
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Gil, Luciana Aquini Fernandes. "Produção e caracterização de quimeras recombinantes C e D de Clostridium botulinum". Universidade Federal de Pelotas, 2012. http://repositorio.ufpel.edu.br/handle/ri/2486.
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Previous issue date: 2012-08-08Bovine Botulism is a lethal intoxication caused by the ingestion of the neurotoxins produced by Clostridium botulinum types C and D that inhibit the release of acetylcholine at the neuromuscular junction leading to death by flaccid paralysis. It produces important economic losses, being a major cause of casualties in cattle in several regions of Brazil. The control of the disease depends on the presence of neutralizing antibodies against botulinum neurotoxins (BONTs) in immunized cattle. Immunization is obtained inoculating toxoids produced from cultures of selected strains of C. botulinum types C and D, whose industrial production has limitations concerning efficiency and productivity. An alternative to the use of these toxoids is the production of recombinant antigens with high levels of purity and antigenicity. The C-terminal fraction of the heavy chain of botulinum neurotoxins has been the main target in the development of recombinant vaccines with promising results. In this work, two recombinant bivalent chimeras for the control of bovine botulism consisting of the neuronal receptor binding domains (NRBDs) of botulinum C and D toxins were efficiently produced in Escherichia coli. They were characterized and evaluated in mice, with promising results. Both the recombinant chimeras rLTB-C-D and rC-D were produced by cloning and expressing a synthetic gene encoding the C-terminal portion of both BONTs. The former also included the preferred codons of the E. coli heat labile enterotoxin B subunit (LTB), a potent humoral immune adjuvant. The levels of expression of the recombinant antigens were satisfactory, yielding approximately 100 mg of each recombinant antigen per liter of culture. An ELISA performed to assess the antigenicity of the molecules showed that both were recognized by sera of immunized mice suggesting the preservation of epitopes with the properties of native BONTs. Both chimeras induced high levels of neutralizing antibodies without undesirable effects. The level of neutralizing antibodies of the groups inoculated with equimolar concentrations of rLTB-C-D and rC-D containing Aluminum Hydroxide as adjuvant were similar, confirming the adjuvant properties of LTB. These results demonstrated that the recombinant chimeras were immunogenic. Sera from mice inoculated with commercial vaccines were also analyzed by ELISA using as antigens rC and rD, corroborating the neutralization.
O botulismo bovino é uma intoxicação letal causada pela ingestão da neurotoxina produzida pelo Clostridium botulinum principalmente dos tipos C e D que atua inibindo a liberação de acetilcolina na junção neuromuscular levando à morte por paralisia flácida, com grande importância econômica e sanitária, sendo uma das principais causas de morte em bovinos adultos no Brasil. O controle imunológico do botulismo bovino depende da presença de anticorpos neutralizantes contra as neurotoxinas botulínicas (NBOTs) no momento da ingestão da toxina pré-formada, por meio de imunização dos animais. Atualmente, a imunização é realizada com toxóides obtidos da detoxificação do extrato de cultivos de cepas selecionadas de C. botulinum dos tipos C e D que apresentam limitações quanto à eficiência e produção. Uma alternativa ao uso dos toxóides clássicos é a produção de vacinas recombinantes usando antígenos específicos de alta pureza e imunogenicidade. A fração C-terminal da cadeia pesada da neurotoxina botulínica tem sido o alvo principal no desenvolvimento de alternativas recombinantes a serem utilizadas como vacinas. Neste trabalho, duas quimeras recombinantes bivalentes compostas pelos domínios de ligação ao receptor neuronal (DLRNs) foram produzidas em Escherichia coli, caracterizadas e avaliadas em camundongos. As quimeras recombinantes rLTB-C-D e rC-D foram produzidas a partir da clonagem e expressão de um gene sintético que codifica a porção C-terminal das NBOTs construído com os códons preferenciais de E. coli e a subunidade B da enterotoxina termolábil de E. coli (LTB), um potente adjuvante da resposta imune humoral. O nível de expressão dos antígenos foi de aproximadamente 100mg de cada antígeno recombinante por litro de cultura. Um ELISA realizado para avaliar a antigenicidade das moléculas mostrou que ambas foram reconhecidas pelos soros padrões, sugerindo conservação de epitopos semelhantes aos dos DLRNs nativos. Ambas as quimeras foram inócuas para os camundongos, os quais não apresentaram lesões no local da inoculação bem como alteração de comportamento. Nos soros dos camundongos inoculados com as quimeras recombinantes foi possível detectar níveis de anticorpos neutralizantes. O grupo inoculado com a rLTB-C-D apresentou nível de anticorpos neutralizantes semelhante ao do grupo rC-D + hidróxido de alumínio confirmando o potencial adjuvante da LTB. As quantidades de antígenos utilizados foram equimolares. Esses resultados demonstram que as quimeras recombinantes foram imunogênicas. Os soros dos camundongos inoculados com as diferentes vacinas também foram analisados por ELISA indireto utilizando rC e rD como antígenos. Os dados obtidos neste ELISA corroboram os dados da soroneutralização.
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OUAKNINE, ROGER. "La conservation des aliments par ionisation ; les problemes poses par clostridium botulinum". Strasbourg 1, 1987. http://www.theses.fr/1987STR10748.
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Rheinhart, Courtney Elizabeth. "Clostridium botulinum toxin development in refrigerated reduced oxygen packaged Atlantic croaker (Micropogonias undulatus)". Thesis, Virginia Tech, 2007. http://hdl.handle.net/10919/32440.
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The purpose of this study was to determine the effects of storage temperature and film oxygen transmission rate (OTR) on toxin development by Clostridium botulinum in refrigerated raw vacuum packaged croaker fillets, and to determine if toxin development precedes microbiological and/or organoleptic spoilage. Raw croaker fillets were vacuum packaged in oxygen-permeable films (OTR of 10,000 cc/m2/24hr or 3,000 cc/m2/24hr) and stored at either 4ºC or 10ºC. Type 83F, 17 Type B, Beluga, Minnesota, and Alaska nonproteolytic strains of C. botulinum were used to inoculate fish prior to vacuum packaging. At both temperatures, microbial spoilage preceded toxin production in fillets vacuum packaged in both film types. At 4ºC microbial spoilage occurred after approximately 7 days for fillets vacuum packaged in the 10,000 cc/m2/24hr OTR film and after 8 days for fillets vacuum packaged in the 3,000 cc/m2/24hr OTR film. However, toxin was not detected until day 8. At 10ºC microbial spoilage occurred after approximately 3 days for fillets vacuum packaged in the 10,000 cc/m2/24hr OTR film, while toxin production occurred on day 5. For fillets vacuum packaged in the 3,000 cc/m2/24hr OTR film microbial spoilage occurred after 4 days. However toxin production did not occur until day 6. In contrast, at both temperatures toxin production preceded or coincided with organoleptic spoilage in fillets vacuum packaged in both film types. At 4ºC organoleptic spoilage occurred after 10 days for fillets packaged in the 10,000 cc/m2/24hr OTR film and after 9 days in the 3,000 cc/m2/24hr OTR film, while toxin production occurred on day 8. At 10ºC organoleptic spoilage occurred after 6 days for fillets packaged in the 10,000 cc/m2/24hr OTR film, and toxin was detected on day 5. For fillets packaged in the 3,000 cc/m2/24hr OTR film and stored at 10ºC, organoleptic spoilage occurred after 6 days, while toxin production occurred on day 6. Although toxin production preceded or coincided with organoleptic spoilage in both film types, this may have been because samples were presented on ice, which could have masked potential odors. This study shows that there are not significant differences between these film types when it comes to microbial and organoleptic spoilage. Therefore lower OTR films, such as 3,000 cc/m2/24hr film, may be used to vacuum package Atlantic croaker.Master of Science in Life Sciences
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Cleverley, Stephen. "Exploring the role of the small GTPase Rho in T-lymphocyte biology". Thesis, University College London (University of London), 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.311966.
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Arritt, Fletcher M. III. "The Effects of Modified Atmosphere Packaging on Toxin Production by Clostridium botulinum in Raw Aquacultured Flounder Fillets and Fully Cooked Breaded and Battered Pollock Portions". Diss., Virginia Tech, 2004. http://hdl.handle.net/10919/28790.
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Fish products under vacuum (VAC) and/or modified atmosphere packaging (MAP) conditions can have a significantly extended shelf life. Prevention of toxin production by Clostridium botulinum is essential for processors of VAC and MAP refrigerated fishery products. The objective of this study was to determine if C. botulinum toxin development precedes microbiological spoilage and sensory rejection in fully cooked breaded and battered Alaskan Pollock or raw aquacultured flounder fillets.
Aquacultured summer flounder (Paralichthys dentatus) fillets and fully cooked breaded and battered Alaskan pollock (Theragra chalcogramma) were either aerobically packed (Oxygen Transmission Rate (OTR) of 3,000 cc/m2/24h@70°F for flounder and 6,000 cc/m2/24h@70°F for Pollock), vacuum packed or MAP packaged in a 100% CO2 atmosphere (OTR of 7.3 cc/m2/24h@70°F). Flounder fillets were stored at either 4 or 10°C while pollock portions were stored at 8 and 12°C. Based on the time to spoilage (counts >107 CFU/g), additional samples were inoculated with five strains of nonproteolytic C. botulinum and analyzed qualitatively for botulinum toxin using a mouse bioassay.
For flounder at 4°C, toxin formation did not occur after 35 days in aerobically packed fillets. Vacuum packed and 100% CO2 fillets produced toxin before spoilage at days 20 and 25, respectively. In the aerobic packages at 10°C, toxin production occurred after spoilage at day 8, but before spoilage in the vacuum and 100% CO2 packages at day 9. Sensory evaluation of toxic vacuum and 100% CO2 packages at 4°C revealed toxin production proceeded spoilage and absolute sensory rejection. However, at 10°C toxin production was evident only after absolute sensory rejection and microbiological spoilage for aerobically packed fillets. Vacuum packages and 100% CO2 packages were toxic during spoilage but before absolute sensory rejection.
Toxin was not present in the aerobically and 100% CO2 packed pollock samples at 8°C and the 100% CO2 packed samples at 12°C after 35 days. Aerobically packed portions stored at 12°C first produced toxin at day 25; toxicity occurred after absolute sensory rejection and before spoilage. The vacuum packed portions first formed toxin at day 25 for 8 and 12°C storage before spoilage and absolute sensory rejection.Ph. D.
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Fach, Patrick. "Évaluation des techniques de biologie moléculaire pour la détection et le typage des Clostridium pathogènes en agro-alimentaire : application à Clostridium perfringens et Clostridium botulinum". Compiègne, 1994. http://www.theses.fr/1994COMPD689.
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Les affections à C. Perfringens et C. Botulinum sont observées dans le monde entier. C. Botulinum est l'agent responsable du botulisme. Cette maladie grave, qui est fort heureusement peu répandue, atteint l'homme et les animaux. Elle est due à l'action d'une neurotoxine qui est le composant toxique le plus actif que l'on connaisse et qui agit sur les neurones périphériques en bloquant la libération de neuromédiateur. On distingue 7 toxinotypes (A à G), les toxinotypes E et F sont également élaborés par certains Clostridium autres que C. Botulinum. Les infections à C. Perfringens sont plus répandues. Les souches entérotoxinogènes de type A sont responsables de toxi-infections alimentaires chez l'homme. Elles sont au troisième rang après les toxi-infections dues à Salmonella et Staphylococcus. Les souches de type C sont responsables d'entérites nécrosantes graves (Pig-bel et Darmbrand) mais exceptionnelles. Chez l'animal, différents toxinotypes (A à E) peuvent être en cause. Ils sont à l’ origine d'entérotoxémies qui peuvent provoquer de lourdes pertes sur le plan économique. Notre travail a porté sur la mise en évidence et la caractérisation, par des techniques de biologie moléculaire, des gènes codant pour les neurotoxines de C. Botulinum et les toxines majeures ainsi que l'entérotoxine de C. Perfringens. Un test d'amplification génique (PCR) à l'aide d'oligonucléotides partiellement dégénérés a permis l'amplification d'un fragment d'ADN de 270 bp dans chacun des 5 gènes codant pour les neurotoxines botuliques A, B, E, F et G. Les produits d'amplification ont été identifiés à l'aide de sondes internes spécifiques de type. Le modèle a été développé pour identifier et typer rapidement C. Botulinum A, B et E dans les aliments. Le test a montré une sensibilité comparable au test de référence, de létalité sur souris. Les gènes des neurotoxines botuliques C et D ont été identifiés par un test de bi-amplification. La technique a permis la détection et le typage de C. Botulinum C et D au sein des échantillons pathologiques analysés. Nous avons également pu caractériser le gène de l'entérotoxine de C. Perfringens par une technique d'amplification génique et d'hybridation ADN-ADN. Ceci nous a permis d'adapter un test de diagnostic par PCR duplex dans les aliments destinés à l'homme et à l'animal. Outre son utilisation en agro-alimentaire, cet outil a aussi été utilisé dans les matières fécales d'enfants atteints de toxi-infections à C. Perfringens. Par ailleurs, les gènes des quatre toxines majeures : alpha, bêta, epsilon et iota de C. Perfringens ont pu être caractérisés à l'aide de plusieurs oligonucléotides spécifiques. L'étude du typage moléculaire de C. Perfringens par PCR a contribué à montrer l'existence, chez certaines souches de C. Perfringens de type C, d'un gène codant pour la toxine bêta différent de celui décrit dans la littérature.
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Ragazani, Adriana Valim Ferreira [UNESP]. "Ocorrência de clostrídios patogênicos em solo de pastagem da micro-região de Jaboticabal, SP". Universidade Estadual Paulista (UNESP), 2007. http://hdl.handle.net/11449/103894.
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Entre as espécies de Clostridium de importância em patologia animal, destaca-se o Clostridium perfringens, o Clostridium botulinum, o Clostridium chauvoei. O objetivo desta pesquisa foi verificar a presença de bactérias anaeróbias esporuladas (Clostridium sp), assim como, identificar as espécies de Clostridium patogênicos para a saúde animal, principalmente de bovinos, no solo de pastagem da micro-região de Jaboticabal, SP. Foram coletadas 250 amostras de solo e realizadas contagens de bactérias esporuladas do gênero Clostridium e identificação das espécies patogênicas presentes. Os resultados permitiram demonstrar a contagem de UFC de Closrtidium sp com média em log 10 igual a 2,79, sendo que os valores mínimo e máximo obtido foram 2,15 e 3,68 respectivamente. Para caracterização e identificação, os resultados permitiram identificar a presença de bactérias anaeróbias esporuladas em 233 amostras (93,2%), entre estas 180 eram do gênero Clostridium...
The species of Clostridium of major importance to animal pathology are Clostridium perfringens, Clostridium botulinum, and C. chauvoei. Considering this, the objective of this research was to verify the presence of anareobic sporulate bacteria (Clostridium sp), and also identify the species of pathogenic Clostridium for the animal health, mostly to bovine, in pasture soil of Jaboticabal-SP. A total of 250 samples were collected and used to determine the number of sporulated bacteria from Clostridium genderand identify the pathogenic species present. The results demonstrated that the average number of CFU in log 10 of Closrtidium sp was 2,79, and the minimum and maximum values obtained were 2,15 and 3,68 respectively. After characterization and isolation and identification, the results showed the presence of 233 samples (93,2%) of sporulated bacteria, of these 180 were of Clostridium gender. The biochemical tests were identified in 42 samples, being 23 samples (9,2%) of Clostridium perfringens, 13 samples (5,2%) of Clostridium botulinum and 6 samples (2,4%) of C. chauvoei ...(Complete abstract, click electronic access below)
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Ferreira, Rosa Maria Moraes. "Contaminação ambiental pelo Clostridium Botulinum tipos C e D de valas de captação hidrica e cultivo do microrganismo em um sistema experimental". [s.n.], 2002. http://repositorio.unicamp.br/jspui/handle/REPOSIP/316869.
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Orientador : Iveraldo dos Santos DutraDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Mestrado
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Hunter, Leonie C. "The role of Clostridium botulinum type C in the pathogenesis of equine grass sickness". Thesis, University of Edinburgh, 2001. http://hdl.handle.net/1842/24726.
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This thesis investigates the hypothesis that EGS is caused by a toxicoinfection with C. botulinum type C, where the organism grows and produces toxin in the gastrointestinal tract (GI). The study has taken three main directions: 1) detection of the botulinum type C neurotoxin (BoNT/C) in the GI tract of horses with and without EGS. 2) isolation and characterisation of organisms phenotypically resembling C. botulinum type C, and 3) detection of antibodies to the type C neurotoxin and surface antigens in the serum, GI tract, colostrum and milk. BoNT/C was detected by ELISA both directly in GI contents and after enrichment of the sample in order to determine the presence of both preformed toxin-producing organisms. BoNT/C was detected by direct detection and/or enrichment in 74% of horses with acute grass sickness, 67% of horses with subacute grass sickness and 67% of horses with chronic grass sickness compared to 10% of controls. BoNT/C was also detected directly and/or after enrichment in the GI tract of 81% cats with feline dysautonomia. These results support the hypothesis that toxicoinfection with C. botulinum type C is involved in the aetiology of EGS and possibility other similar dysautonomias, such as feline dysautonomia. C. botulinum type C is phenotypically indistinguishable from C. botulinum type D and Clostridium novyi type A. These three organisms are grouped together as "Group III" botulinum strains and are differentiated on the basis of the major toxins they produce. However, the type C and D neurotoxins and the C. novyi alpha toxin are encoded on separate pseudolysogenic bacteriophages that can be readily lost. Loss of these phages results in loss of toxigenicity and non-toxigenic isolates are essentially indistinguishable from each other. Sixteen isolates phenotypically resembling Group III botulinum were isolated from the GI tract of nine horses, one cat and one hare. The animals from which these organisms were isolated all had some association with dysautonomia: five horses, one cat and one hare had dysautonomia at the time of sampling; two horses had recovered from EGS; two horses had been in recent contact with EGS.
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Lux, Romy [Verfasser]. "Glutamat-Aufnahme und -Freisetzung durch Astrozyten nach Clostridium botulinum C3-Protein-Behandlung / Romy Lux". Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2009. http://d-nb.info/1023783649/34.
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Piepgras, Johannes [Verfasser]. "Der Einfluss von Clostridium botulinum C3 Proteinen auf die neuronale Glutamat-Aufnahme / Johannes Piepgras". Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2019. http://d-nb.info/1201650356/34.
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Campbell, Kathryn Deirdre. "Molecular inter-relationships of psychrotrophic Clostridium botulinum based on 23S rRNA and BoNT genes". Thesis, University of Reading, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.262610.
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Dukart, Irene [Verfasser]. "Charakterisierung der Interaktion des C2-Toxins von Clostridium botulinum mit Cyclophilin A / Irene Dukart". Ulm : Universität Ulm. Medizinische Fakultät, 2014. http://d-nb.info/1049238400/34.
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Zeiller, Matthias [Verfasser] y Anton [Akademischer Betreuer] Hartmann. "Systemische Kolonisierung von Weißklee (Trifolium repens) durch Clostridium botulinum / Matthias Zeiller. Betreuer: Anton Hartmann". München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2016. http://d-nb.info/1110749392/34.
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Ismaiel, Adnan A. "The inhibition of Clostridium botulinum growth and toxin production by essential oils of spices". Diss., Virginia Polytechnic Institute and State University, 1987. http://hdl.handle.net/10919/77803.
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The essential oils of clove, thyme, black pepper, pimenta, origanum, garlic, onion, and cinnamon were evaluated for their effect on germination, outgrowth, growth, and toxin production of C. botulinum strains in microbiological media. The oils of clove, thyme, origanum, and cinnamon were studied for their mechanism of inhibition of C. botulinum 67B. The most effective oil, in combination with sodium nitrite at different levels was further tested against the growth and toxin production of C. botulinum (mixed types) in a meat model system.
Among all the spice oils, origanum and pimenta were the most effective in inhibiting six strains of types A, B, and E of C. botulinum in prereduced PY medium. These oils at a concentration of 200 ppm completely inhibited C. botulinum growth. Garlic, onion and black pepper exhibited the lowest inhibitory activity towards the growth of C. botulinum strains. Strains of type A were more sensitive to the inhibitory action of the oils than those of types B and E.
The inhibition of germination of C. botulinum by the eight spice oils indicated that garlic oil was the most potent inhibitor. Oils of pimenta, and clove were the least effective in inhibiting germination. The inhibitory effect of the oils was shown to be reversible. The oils appeared to have no significant effect on the outgrowth of the germinated spores. Nevertheless, the oils were highly active in inhibiting vegetative growth (cell division). Black pepper, clove, cinnamon, and origanum were the strongest inhibitors of vegetative growth. Yet, the oils had no direct effect on toxin production. The delay in toxin production caused by the oils was attributed to the effect of the oils on growth rather than on toxin production. Origanum oil acted synergistically with sodium nitrite in inhibiting the growth. of C. botulinum in a microbiological medium. In vacuum-packaged comminuted pork, origanum oil at 400 ppm, in combination with 50-100 ppm of sodium nitrite, significantly delayed the growth and toxin production of C.botulinum (mixed types). The probability of growth and toxin production of C. botulinum (p) in the vacuum packaged comminuted pork was calculated with Hauschild 's formula. The results showed that sodium nitrite significantly lowered p values, whereas origanum oil had very low effect on p values.Ph. D.
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Ragazani, Adriana Valim Ferreira. "Ocorrência de clostrídios patogênicos em solo de pastagem da micro-região de Jaboticabal, SP /". Jaboticabal : [s.n.], 2007. http://hdl.handle.net/11449/103894.
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Orientador: Ruben Pablo Schocken-IturrinoBanca: Maria Luiza Poiatti
Banca: Alessandra Aparecida Medeiros
Banca: Hélio José Montassier
Banca: Antônio Carlos Monteiro
Resumo: Entre as espécies de Clostridium de importância em patologia animal, destaca-se o Clostridium perfringens, o Clostridium botulinum, o Clostridium chauvoei. O objetivo desta pesquisa foi verificar a presença de bactérias anaeróbias esporuladas (Clostridium sp), assim como, identificar as espécies de Clostridium patogênicos para a saúde animal, principalmente de bovinos, no solo de pastagem da micro-região de Jaboticabal, SP. Foram coletadas 250 amostras de solo e realizadas contagens de bactérias esporuladas do gênero Clostridium e identificação das espécies patogênicas presentes. Os resultados permitiram demonstrar a contagem de UFC de Closrtidium sp com média em log 10 igual a 2,79, sendo que os valores mínimo e máximo obtido foram 2,15 e 3,68 respectivamente. Para caracterização e identificação, os resultados permitiram identificar a presença de bactérias anaeróbias esporuladas em 233 amostras (93,2%), entre estas 180 eram do gênero Clostridium...(Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The species of Clostridium of major importance to animal pathology are Clostridium perfringens, Clostridium botulinum, and C. chauvoei. Considering this, the objective of this research was to verify the presence of anareobic sporulate bacteria (Clostridium sp), and also identify the species of pathogenic Clostridium for the animal health, mostly to bovine, in pasture soil of Jaboticabal-SP. A total of 250 samples were collected and used to determine the number of sporulated bacteria from Clostridium genderand identify the pathogenic species present. The results demonstrated that the average number of CFU in log 10 of Closrtidium sp was 2,79, and the minimum and maximum values obtained were 2,15 and 3,68 respectively. After characterization and isolation and identification, the results showed the presence of 233 samples (93,2%) of sporulated bacteria, of these 180 were of Clostridium gender. The biochemical tests were identified in 42 samples, being 23 samples (9,2%) of Clostridium perfringens, 13 samples (5,2%) of Clostridium botulinum and 6 samples (2,4%) of C. chauvoei ...(Complete abstract, click electronic access below)
Doutor
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Carpusca, Irina. "The ADP-ribosylating mosquitocidal toxin (MTX) from bacillus Sphaericus : Role of its ricin-like domain in autoinhibition and translocation". Université Louis Pasteur (Strasbourg) (1971-2008), 2005. http://www.theses.fr/2005STR13003.
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Junqueira, Valeria Christina Amstalden. "Avaliação da incidencia de Clostridium botulinum e da produção de toxina em mortadela e presunto". [s.n.], 1994. http://repositorio.unicamp.br/jspui/handle/REPOSIP/254645.
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Orientador: Antonio de Melo SerranoDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos
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Resumo: Na presente pesquisa foi realizada uma avaliação do risco de transmissão de botulismo por mortadela e presunto. Numa primeira fase, foram examinadas amostras de mortadela e' presunto cozido, coletadas ao acaso no varejo do município de Campinas - SP. A partir de 25 amostras de cada produto, em quadruplicata de 75g, num total de 100 subamostras, foi calculado o Número Mais Provável (NMP) de C. botulinum por kg de mortadela e presunto. Nesta fase, ainda foram determinados o tempo e a temperatura de estocagem na revenda, bem como o pH, atividade de água, concentração de cloreto de sódio e de nitrito de sódio e umidade desses produtos. Na segunda fase, mortadela e presunto elaborados por nós e artificialmente contaminados com 104 esporos de C. botulinum tipos A e 8 por amostra foram avaliados quanto à possibilidade de toxigênese do microrganismo, quando submetidos à estocagem inadequada, à temperatura de 30°C. Os resultados obtidos permitiram estimar a incidência de Clostridium botulinum na ordem de O,27NMP/kg de mortadela e inferior a O, 13NMP/kg de presunto. A formação de toxina botulínica foi observada nas amostras de presunto artificialmente contaminadas, após 12 dias de estocagem à temperatura de 30°C. Ao fim de 28 dias de estocagem, à mesma temperatura, não foi detectada toxina nas amostras de mortadela
Abstract: The risk of transmitting botulism via mortadella and ham was evaluated in this research. In the first phase, samples of mortadella and cooked ham were acquired at random from shops in the region of Campinas - SP. Twenty five samples of each product were acquired and examined in quadruplicate (75g each) giving a total of 100 sub-samples. The Most Probable Number (MPN) for C. botulinum per kg of mortadella or ham, was calculated. The storage times and temperatures at the distributers were also determined plus the pH, water activity, sodium chloride and sodium nitrite concentrations and water content. In the second phase, mortadella and ham were produced in the pil0t plant and contamined with 104 C. botulinum types A and B spores per sample. These samples were evaluated to determine the possibility of toxin formation by the microorganism when the products were submitted to inadequate storage at a temperature of 30°C. The results obtained allowed for the estimation of an incidence of C. botulinum at a level of 0,27 MPN/kg in mortadella and below 0,13 MPN/kg in ham. The formation of botulinum toxin was observed after 12 days of storage at 30°C in the artificially contamined ham samples. After 28 days of storage at the same temperature, no toxin was detected in the samples of mortadella
Mestrado
Mestre em Tecnologia de Alimentos
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Reddy, Divya Shree A. "Iron Requirement of Clostridiiyum Botulinum Type A and Characterization of Iron-Sulfur Proteins in Nitrite Treated and Untreated Botulinal Cells". DigitalCommons@USU, 1985. https://digitalcommons.usu.edu/etd/5303.
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The effect of added iron on the growth of Clostridium botulinum type A in a chemically defined medium was studied. Growth of C. botulinum was supported by an iron level of 0.05 ug/ml with maximum growth observed at a level of 3 ug iron/ml.
Electron paramagnetic resonance (EPR) studies were conducted to detect the presence of iron-sulfur centers and iron-nitric oxide complexes in untreated and nitrite treated cell-free extracts of C. botulinum type A. Untreated extracts of C. botulinum exhibited EPR signals in the oxidized and reduced states characteristic of a "HiPiP-type" iron-sulfur center (g=2.02) in the oxidized state and a reduced signal at g=l.94, characteristic of a reduced iron-sulfur center. Extracts of C. botulinum treated with nitrite exhibited an EPR signal at g=2.035, characteristic of iron-nitrosyl complexes, with the simultaneous disappearance of the the signal at g=l.94. This indicates that nitrite reacts with the iron-sulfur centers in botulinal cells to form iron-nitrosyl complexes. Addition of ascorbate with nitrite intensified the EPR signal at g=2.035, probably by enhancing the reduction of nitrite to nitric oxide.
A cytochrome c reduction method was used for the determination of ferredoxin activity in untreated and nitrite treated cells of C. botulinum type A from which ferredoxin had been partially purified. Untreated extracts of C. botulinum reduced cytochrome c which demonstrates ferredoxin activity within the cells. Treatment of the cells with nitrite at a level of 1000 ppm for 45 min was found to inhibit ferredoxin activity by 90%. Boiling the partially purified ferredoxin from the untreated cells for 5 min inactivated the protein.
Pyruvate-ferredoxin oxidoreductase activity in partially purified extracts of nitrite treated and untreated cells of C. botulinum was determined by assaying for FAD reduction and acylhydroxamate formation. Nitrite treated cells exhibited an inhibition of 70% of FAD reducing activity and 80% inhibition of acylhydroxamate formation when compared to the untreated cells. Boiling inhibited the activity of partially purified oxidoreductase activity by more than 90% in both the assays.
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Phillips, Daphne 1956. "Safety studies with proteolytic Clostridium botulinum in high-moisture bakery products packaged under modified atmospheres". Thesis, McGill University, 2002. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=84412.
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Initial challenge studies with spores of proteolytic Clostridium botulinum types A and B (~104 spores/g) showed that while air- and gas-packaged English-style crumpets (aw 0.990) and pizza crust (aw 0.960) were toxic after 42-days storage at ambient temperature (25°C), no neurotoxin was detected in bagels (a w 0.944). Further challenge studies with similarly packaged crumpets inoculated with C. botulinum (~102 spores/g), pre- or post-baking, demonstrated that all crumpets were toxic within 4 to 6 days at 25°C and that toxigenesis preceded spoilage. Furthermore, reformulating crumpets to pH 8.3 and packaging in 100% CO2 had little effect in delaying the growth of C. botulinum compared to crumpets formulated to pH 6.5 and packaged in 60% CO2.Subsequent studies were directed at determining the levels of additional barriers that could be used to ensure the safety of high-moisture MAP crumpets. While ethanol vapour proved to be an effective additional barrier in crumpets (100-g, [aw 0.990, pH 6.5]) challenged with ~102 spores/g of C. botulinum, spoilage preceded toxigenesis due to absorption of ethanol from the package headspace by crumpets. Modelling studies in Trypticase Peptone Glucose Yeast (TPGY) broth confirmed the anti-botulinal nature of ethanol and showed that a level of ~4% (vol/vol) could be used for complete inhibition of this pathogen, depending on the aw and pH of the growth medium. However, while ethanol vapour could be used to inhibit the growth of C. botulinum in high-moisture crumpets, its anti-botulinal efficacy was influenced by the method of crumpet leavening (yeast v chemical).
Preliminary studies were also done to assess the potential of mastic oil, a novel inhibitor, against C. botulinum. While direct and indirect application of ethanolic extracts of mastic oil inhibited the growth of C. botulinum in vivo, they failed to do so in crumpets.
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Yusof, Farida Zuraina Md. "The distribution of toxin genes among isolates of Clostridium botulinum Group II from different environments". Thesis, University of Ulster, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.274549.
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Pospiech, Janina Marta Lucia. "Untersuchung von Gärresten und Gärsubstraten aus landwirtschaftlichen Biogasanlagen des Freistaates Sachsen: Auswahl und Etablierung von bakteriologischen und molekularbiologischen Verfahren zum Nachweis ausgewählter Indikatorkeime". Doctoral thesis, Universitätsbibliothek Leipzig, 2015. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-189760.
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Die im Biogasprozess anfallenden Gärreste werden oftmals als Wirtschaftsdünger verwendet. Krankheitserreger, die sich in den Gärresten befinden können über die Düngung in die Lebensmittelkette gelangen. Die Möglichkeit einer Vermehrung von Bakterien in den Biogasanlagen sowie deren Ausbreitung schürt die Bedenken der Öffentlichkeit. Das Ziel dieser Arbeit war es, Nachweismethoden für die Untersuchung von Proben aus Biogasanlagen zu etablieren, die Praxistauglichkeit dieser anhand von Proben aus Biogasanlagen zu überprüfen und die mikrobielle Belastung dieser Proben hinsichtlich ausgewählter Indikatorkeime zu erfassen. Bei den Indikatorkeimen handelte es sich um Clostridium perfringens, Clostridium botulinum, Enterokokken, Escherichia coli, ESBL-bildende Enterobaceriaceae und Salmonellen.
Für die Etablierung der bakteriologischen Nachweismethoden wurde autoklavierter Gärrest mit einer definierten Keimmenge beimpft und auf verschiedene Nährmedien aufgebracht. Diese wurden bebrütet, ausgezählt und die KbE/ml berechnet. Mittels Probitanalyse wurde für jedes Medium die untere Grenze für den Nachweis aus beimpftem Gärrest bestimmt. Bei den Nährmedien handelte es sich um Brilliance™ Salmonella Agar, XLT4 Agar und XLD Agar für den Nachweis von Salmonella spp. Für E. coli wurden Tergitol 7 Lactose TCC Agar und Brilliance™ E. coli/Coliform Selektiv Agar verwendet. Der Nachweis von Enterokokken erfolgte mittels Slanetz Bartley Agar und Enterococcus Selektivagar. Für die ESBL-bildenden Enterobacteriaceae wurde der Brilliance™ ESBL Agar eingesetzt. Die getesteten Nährmedien zum Nachweis von C. perfringens waren Membran Clostridium Perfringens (mCP) Selektivnährboden sowie Tryptose Sulphite Cycloserine (TSC) Agar überschichtet mit TSC Agarbasis. Für C. botulinum erfolgte der Nachweis auf Eigelb Laktose Agar. Darüber hinaus wurde eine PCR zur C. perfringens Toxintyp-Bestimmung nach dem Protokoll von VAN ASTEN et al. (2009) etabliert. Zum Nachweis von C. botulinum wurde die PCR nach dem Protokoll von HILL et al. (2010) eingesetzt. Bei der Untersuchung der Praxistauglichkeit wurden Proben aus zehn Biogasanlagen des Freistaates Sachsen entnommen und untersucht. Hierbei handelte es sich um Proben aus Abschnitten vor, während und nach der Fermentation. Anhand der ermittelten Nachweisgrenze sowie der Handhabung wurden die folgenden Nährmedien für die Untersuchung der Biogasanlagen-Proben ausgewählt: Brilliance™ Salmonella Agar, XLT4 Agar, Brilliance™ E. coli/Coliform Selektiv Agar, Slanetz Bartley Agar, Brilliance™ ESBL Agar, TSC Agar überschichtet mit TSC Agarbasis und Eigelb Laktose Agar. Für die Anzucht anaerober Bakterien wurden die Proben vor der Beimpfung der Agarplatten erhitzt. Zudem erfolgte eine Anreicherung des zuvor erhitzten Probenmaterials in TPYG Bouillon. Diese wurde genutzt, um daraus aufgereinigte DNA mittels PCR auf C. botulinum und C. perfringens zu untersuchen. Die verwendeten Nährmedien wurden im Praxistest positiv evaluiert. Die Ergebnisse für die Proben aus den Biogasanlagen zeigten, dass, mit Ausnahme von C. perfringens, alle Indikatororganismen während des Biogasprozesses einer Reduktion unterlagen. Die durchschnittliche anaerobe Lebendkeimzahl belief sich auf 107 bis 108 KbE/g Probe. E. coli erfuhr eine Reduktion um bis zu vier Zehnerpotenzen. Enterokokken wurden um 1 bis 2 log10 Stufen reduziert. ESBL-bildende Enterobacteriaceae konnten in sechs der zehn Biogasanlagen nachgewiesen werden. Hierbei handelte es sich überwiegend um E. coli und Klebsiella spp. In keiner der Proben konnten Salmonellen oder C. botulinum nachgewiesen werden. Typ A war der am häufigsten nachgewiesene C. perfringens-Toxintyp. Das β2-Toxin-Gen wurde in 20 Fällen nachgewiesen. Einmal konnte C. perfringens Typ C, β2-Toxin-Gen-positiv detektiert werden.
Der hygienische Status der Gärreste entsprach in etwa dem hygienischen Status von Gülle. In Abhängigkeit vom Indikatorkeim war eine Verbesserung des Status durch eine Reduktion der Keimzahl festzustellen.
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Haug, Gerd. "Clostridium-botulinum-C2-Toxin Interaktion der Toxinkomponenten und Translokation der Enzymkomponente in das Zytosol von Zielzellen /". [S.l.] : [s.n.], 2005. http://deposit.ddb.de/cgi-bin/dokserv?idn=974136484.
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