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1
Dias, Liliane Dane. „Avaliação da eficiência de vacinas contra Clostridium septicum“. Universidade Federal de Minas Gerais, 2003. http://hdl.handle.net/1843/BUOS-8C4FGT.
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Twelve commercial vaccines against clostridiosis were evaluated having in their composition Clostridium septicum toxoids and/or bacterial cells, have been evaluated for potency by mice serum neutralization tests using rabbit or bovine sera and challenge test in guinea-pig. The vaccines, coded as T1, T10 and T11, elicited alpha antitoxin titers in rabbits which were superior to the minimum test level of 2.5 lU/mL, as recommended for quality control, and the vaccines T2 and T4 eliciting titers of 2.0 and 2.5 lU/mL. Similar results were obtained as detected in bovine sera against vaccines T1, T2, T4 and T10. The T11 vaccine was not evaluated in bovines. According to the challenge tests in guinea pigs no vaccine met the minimum requeriments for approval. Clostridium septicum vaccines were in general inefficient for stimulating the immune response as evaluated by the recomendaded tests.Foram avaliadas quanto à eficiência 12 vacinas comerciais contra clostridioses, que continham toxóides e/ou bacterinas de Clostridium septicum, pelo teste de soroneutralização em camundongos a partir de soros de coelhos e bovinos vacinados e pelo teste de desafio direto em cobaias. As vacinas codificadas como T1, T 10 e T11 apresentaram, em coelhos, títulos de antitoxina alfa superiores ao nível mínimo de teste de 2,5 Ul/mL recomendado pelo controle deste produto e as vacinas T2 e T4 títulos de 2.0 e 2.5 Ul/mL. Resultados semelhantes foram obtidos nos soros de bovinos em relação às vacinas T1, T2, T4 e T10. A vacina T11 não foi testada em bovinos. Pelo método do desafio direto em cobaias, nenhuma vacina atendeu aos requisitos mínimos. As vacinas contra Clostridium septicum, em sua maioria, foram ineficientes em estimular resposta compatível com níveis de teste.
2
Pinto, Maristela Pimentel. „Produção de conjugados para identificação de Clostridium septicum e Clostridium chauvoei para a técnica de imunofluorescência direta“. Universidade Federal de Minas Gerais, 1992. http://hdl.handle.net/1843/BUOS-8QLMQR.
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The production of conjugates anti-Clostridium septicum and anti-Clostridium chauvoei for direct immunoiluorescense testing is described. The culture medium used, enriched with glucose 0,5% and cysteine 0,1%, propitiated fast and expressive C. septibum and C. chauuoefs growing. Antigens containing 3,2x10 8 to 3,4x10 8 cells/ml were considered satisfactory for immune sera production and aglutination tests. Conjugates prepared from immune sera fractionated by combination of caprylic acid and ammonium sulfate precipitation exhibited eight times higher titres than conjugates prepared by ammonium sulfate precipitation. Heterologous antigens and field materials sent to UFMG's Veterinary School were tested. Fluorescent inibition tests and heterologous and homologous antigens adsorption tests were peiformed.Foram produzidos conjugados anti-C, septicum e anti-C. chauvoei para a técnica dc imunofluorescência direta. O meio de cultura utilizado, enriquecido com 0,5% dc glicose e 0,1% dc cisteína, propiciou crescimento rápido e expressivo dc C. septicum e C. chauvoei. Os antígenos contendo 3,2x10 8 a 3,4x10 8 células/ml foram considerados satisfatórios para produção de soro imune e testes de aglutinação. Os conjugados preparados a partir de soros imunes fracionados com ácido caprílico/sulfato dc amônio obtiveram títulos oito vezes maiores que os precipitados com sulfato de amônio. Para verificação da especificidade dos conjugados foram testados antígenos heterólogos, assim como amostras isoladas a partir de materiais de campo enviados à Escola de Veterinária da UFMG e foram realizados testes de inibição da fluorescência e adsorção com antígenos heterólogos e homólogos.
3
Assis, Ronnie Antunes de. „Padronização da imunohistoquímica para detecção de clostridium chauvoei e clostridium septicum e comparação com a imunofluorescência direta“. Universidade Federal de Minas Gerais, 2001. http://hdl.handle.net/1843/BUOS-8C7F85.
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Standardized the technique of immunohistochemistry using the complex streptavidin-biotin immunoenzymatic marked (LSAB) to detect Clostridium septicum and Clostridium chauvoei and compared it with the technique of direct immunofluorescence (DIF), using different tissues of guinea pigs inoculated with chauvoesi Clostridium, Clostridium septicum, Clostridium sordellii, Clostridium novyi type A and Clostridium chauvoei and anti-Clostridium septcum were produced in rabbits and purified by DEAE-cellulose hill. Portions of imonoglobunas were conjugated with FITC and the remainder was used in the LSAB technique. Conjugates anto-and anti-chauvoei Clostridium Clostridium septicum showed titers of 256 and 512, respectively, while the primary anti-Clostridium chuvoei and anti-Clostridium septicum were standardized at a dilution of 1:2000 / 1:4000 and 15 minutes / 40 minutes of incubation, respectively. There were no cross-reactivity between antibodies evaluated and none of the other species of clostridia used. Chauvoei Clostridium septicum and Clostridium were detected in all sections and tissue impressions by LSAV techniques and IFD. LSAB technique developed in this study was efficient to detect Clostridium septicum and Clostridium chauvoei in tissues of guinea pigs fixed in formalin and embedded in paraffin, and had 100% sensitivity and specificity of DIFPadronizou-se a técnica da imunohistoquimica empregando o complexo de imunoenzimatico streptavidina-biotina marcada (LSAB) para detectar Clostridium chauvoei e Clostridium septicum e comparou-se a mesma com a técnica de imunofluorescencia direta (IFD), usando diferentes tecidos de cobaios inoculados experimentalmente com Clostridium chauvoesi, Clostridium septicum, Clostridium sordellii, Clostridium novyi tipo A e Clostridium chauvoei e anti-Clostridium septcum foram produzidos em coelhos e purificados em colina de DEAE-celulose. Partes das imonoglobunas foram conjugadas com isotiocianato de fluoresceína e o restante foi utilizada na técnica de LSAB. Os conjugados anto-Clostridium chauvoei e anti-Clostridium septicum apresentaram títulos de 256 e 512, respectivamente, enquanto o anticorpo primário anti-Clostridium chuvoei e anti-Clostridium septicum foram padronizadas na diluição de 1:2000/15 minutos e 1:4000/40 minutos de incubação, respectivamente. Não foram observadas reações cruzadas entre os anticorpos avaliados e nenhuma das outras espécies de clostridios utilizadas. Clostridium chauvoei e Clostridium septicum foram detectadas em todas as seções e impressões de tecidos pelas técnicas de LSAV e IFD. A técnica de LSAB desenvolvida neste estudo, mostrou-se eficiente para detectar Clostridium chauvoei e clostridium septicum em tecidos de cobaios fixados pelo formol e incluídos em parafina, e apresentou 100% de sensibilidade e especificidade em relação à IFD
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Wilson, Lynn Margaret. „Physiological studies on swarming and production of virulence determinants in Clostridium septicum“. Thesis, University of Cambridge, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.627209.
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5
Jansen, Katja. „Methodische Untersuchungen zu Eigenschaften, Nachweis, Reinigung und Antigenität des a-Toxins [Alpha-Toxins] von Clostridium septicum“. [S.l.] : [s.n.], 2000. http://deposit.ddb.de/cgi-bin/dokserv?idn=961183098.
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6
Salvarani, Felipe Masiero. „Padronização de um teste de potência de toxóide de Clostridium septicum em linhagem contínua de célula“. Universidade Federal de Minas Gerais, 2007. http://hdl.handle.net/1843/SSLA-7UDPJ9.
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Clostridium septicum is the pathogen that causes the malignat edema. Because of its strong cytotoxic alpha toxin, infections are often lethal. To prevent losses in animal, vaccination with alpha toxóide vaccines is carried out. The potency test of the vaccines have to be done byneutralization test in mice or intradermally into guinea-pig skin. An alternative method to detect neutralizing antibodies based on the use of cell cultures was standardized. Cell culture titrations of C. septicum alpha toxin performed on rabbit at the levels of test prescribed by BritishPharmacopeia have produced consistent results which agree closely with animal models. Cell culture indicators are more sensitive than in vivo methods, permitting detection of substantially lower titers than possible in vivo indicators. It concluded that cell culture titration offers a valid invitro alternative to the use mouse lethal and guinea-pig dermonecrotic indicators for the titration of sera of potency test of C. septicum alpha toxin vaccinesClostridium septicum é o principal patógeno responsável pelo quadro de edema maligno.Devido à ação citotóxica da toxina alfa as infecções geralmente são fatais. Para o controle da doença é utilizado a vacinação com toxóide alfa. O teste de potência das vacinas clostridiais érealizado através da técnica de soroneutralização em camundongos ou teste intradérmico em cobaio, porém com o objetivo de estudar técnicas alternativas in vitro padronizou-se um método para detecção de anticorpos neutralizantes utilizando cultura de células. Os títulosobservados na soroneutralização em cultivo celular, obtidos a partir do soro de coelhos vacinados frente à toxina alfa de C. septicum padronizada nos níveis de teste prescritos pela Farmacopéia Britânica, demonstraram resultados com significativa correlação quando secompara aos modelos animais. O indicador cultura de célula é mais sensível do que os modelos in vivo, permitindo a detecção de títulos de anticorpos substancialmente mais baixos. Conclui-se que a soroneutralização em cultura de célula é uma alternativa ao uso dosindicadores de letalidade em camundongos e dermonecrótico em cobaios na titulação de soros na avaliação da potência de vacinas contra C. septicum
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J?NCK, Fernanda. „A intoxica??o de bovinos por Pteridium (aquilinum) arachnoideum em Santa Catarina e a identifica??o das bact?rias envolvidas nos infartos do quadro agudo“. Universidade Federal Rural do Rio de Janeiro, 2014. https://tede.ufrrj.br/jspui/handle/jspui/2306.
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Pteridium (aquilinum) arachnoideum is a cosmopolitan plant, responsible for heavy losses in cattle for heavy losses in almost all Brazil. This plant causes three clinical pictures: an acute disease, characterized by hemorrhages and fever, and two chronic diseases characterized by bladder tumors and carcinomas of the superior digestive tract. In Santa Catarina, in a retrospective study, the acute poisoning is the form that prevailed. Most of the cases ocurred in the autumn and the main clinical signs observed were fever and faeces with blood. The macroscopic lesions were widespread hemorrhages and infarcts mainly in lung, liver, intestine and lymphnodes, which were characterized by necrosis associated with groups of basophylic bact?ria, sometimes with formation of bubbles of gas. In the bone marrow there was rarefaction or absence of hematopoietic tissue. The experimental reproduction of the disease was realized in four cattle, two vaccinated against clostridioses and two not vaccinated. These cattle received doses of 20, 20, 14 and 10g/kg/day, and they died after 82, 94, 46 and 76 days, when they had ingested 149, 180, 71 and 75% of the plant in relation to their weight. The course of the clinical signs was 5, 4, 1 and 5 days, and at post-mortem examination the lesions found were hemorrhages of varied degrees and locations. Liver infarcts were found in bovines 2, 3 and 4, and in the intestine in all the cattle. The histological lesions were characterized by rarefaction and absence of hematopoietic tissue in the bone marrow, necrosis and bacterial aggregates in the liver, lung, intestine and lymphnodes. Histopathology did not reveal inflammatory reaction and if present the intensity was slight. Samples of organs with infarcts collected at necropsy, in the spontaneous and experimental intoxication, sown in Tarozzi medium, produced gas. Five samples caused death in mice after inoculation of the medium. At necropsy the carcasses had putrid smell, the subcutaneous tissue was red and there was edema and red liquid in the abdominal cavity. Two mice that were sacrificed presented inflammatory reaction in the place of the application, characterized by areas of adherence of the skin to the subcutaneous tissue and presence of abscesso, and four presented putrid smell at necropsy. The impression of of the liver capsula of the mice that died or got sick, revealed small Gram positive rods. Histopathology of the mice that died, revealed in the skeletal musculature of the thighs edema between the fibers with necrosis and eosinofilia of fibers and great amount of small basophylic rods, associated with slight inflammatory mononuclear infiltration and hemorrhage. In the skin also inflammatory filtrate was observed, with edema in the derma and great amount of small basophylics rods. The rest of the mice were sacrificed and no alterations were found. The identification by Chain reaction of Polimerase (PCR) of the liver of the mice that had died, in Tarozzi medium, resulted in Clostridium septicum.
Pteridium (aquilinum) arachnoideum ? planta cosmopolita, respons?vel por perdas vultuosas na cria??o de bovinos em quase todas as regi?es do Brasil. Esta planta ? respons?vel por causar tr?s quadros cl?nicos: um quadro agudo, caracterizado por hemorragias e febre, e dois quadros cr?nicos caracterizados por tumores de bexiga e do trato digest?rio superior. Em Santa Catarina, em estudo retrospectivo, a intoxica??o aguda ? a forma que prevaleceu sobre as demais. A maioria dos casos ocorreu no outono e os principais sinais cl?nicos observados foram febre e fezes com sangue. As les?es macrosc?picas encontradas foram hemorragias generalizadas e infartos principalmente em pulm?o, f?gado, intestino e linfonodo, os quais se caracterizavam por necrose associada a agregados bacterianos bas?filos, em alguns casos com forma??o de bolhas de g?s. Na medula ?ssea havia rarefa??o ou aus?ncia do tecido hematopo?tico. A reprodu??o experimental da doen?a foi realizada em quatro bovinos, dois vacinados contra clostridioses e dois n?o vacinados. Estes receberam doses de 20, 20, 14 e 10g/kg/dia de Pteridium (aquilinum) arachnoideum, e morreram com 82, 94, 46 dias e 76 dias, quando tinham ingerido 149, 180, 71 e 75% de planta em rela??o ao peso vivo. A evolu??o dos sinais cl?nicos foi de 5, 4, 1 e 5 dias, e, ? necropsia, as les?es consistiram de hemorragias em variados graus e localiza??es. Infartos de f?gado foram encontrados nos Bovinos 2, 3 e 4 e no intestino em todos os bovinos. As les?es histol?gicas se caracterizaram por rarefa??o e aus?ncia de tecido hematopo?tico na medula ?ssea, necrose e agregados bacterianos no f?gado, pulm?o, intestino e linfonodo. As les?es histol?gicas n?o revelaram rea??o inflamat?ria e quando presente, a intensidade era leve. Amostras de ?rg?os com infartos coletadas de necropsias nas intoxica??es espont?nea e experimental foram semeadas no meio de cultivo de Tarozzi e produziram g?s. Cinco amostras causaram a morte dos camundongos ap?s inocula??o do meio. ? necropsia desses camundongos verificou-se carca?as com cheiro p?trido, tecido subcut?neo avermelhado e com edema e l?quido avermelhado na cavidade abdominal. Dois camundongos que foram eutanasiados apresentaram rea??o inflamat?ria no local da aplica??o, caracterizada por ?reas de ader?ncia da pele com o tecido subcut?neo e abscessos; quatro exalavam cheiro p?trido na hora da realiza??o da necropsia. A impress?o da c?psula do f?gado dos camundongos que morreram e que ficaram doentes, revelou bastonetes Gram-positivos. ? histologia dos camundongos que morreram verificou-se na musculatura esquel?tica da regi?o da coxa, edema e hemorragia entre as fibras, necrose e eosinofilia de fibras e grande quantidade de bastonetes bas?filos, associado a infiltrado inflamat?rio mononuclear leve e hemorragia. Na pele tamb?m verificou-se, infiltrado inflamat?rio, com edema na derme e grande quantidade de bastonetes bas?filos. Os demais camundongos foram eutanasiados e n?o tiveram altera??es. A identifica??o por Rea??o em Cadeia de Polimerase (PCR) do meio de Tarozzi do f?gado dos camundongos que morreram foi detectado Clostridium septicum.
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Frossard, Christine. „Etude phénotypique du groupe "Clostridium argentinense", "Clostridium hastiforme", "Clostridium subterminale"“. Paris 5, 1991. http://www.theses.fr/1991PA05P154.
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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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Couchman, Edward. „Investigating the Type IV pili of Clostridium difficile and Clostridium sordellii“. Thesis, Imperial College London, 2016. http://hdl.handle.net/10044/1/48055.
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Type IV pili (T4P) are the only type of bacterial pili known to be produced by both Gram-negative and Gram-positive organisms. Though the main pilus shaft consists primarily of only one protein (the major pilin), T4P are unusual in their complexity, requiring multiple (10 or more) different protein components for assembly. Like most types of pili, T4P often function as virulence factors. In particular, T4P frequently operate as adhesins, enabling bacteria on which they are present to stick to each other (to form a biofilm or suchlike) or to adhere directly to host cells. Many T4P systems are able to retract, in which case the T4P may mediate flagella-independent motility. Most research into T4P has historically been performed on Gram-negative organisms, with T4P-encoding genes only being identified in Gram-positive organisms more recently. In particular, all sequenced species of the genus Clostridium are known to encode T4P, but only minimal investigation of these systems has been performed to date. In this study, the T4P of Clostridium difficile were investigated. C. difficile is an important pathogen, being the leading cause of antibiotic-associated diarrhoea in the developed world and thus a considerable burden on Western healthcare systems. By investigating the T4P of this species it was hoped to further elucidate its mechanisms of pathogenicity. Data is presented demonstrating the control of T4P expression by cyclic-di-GMP, and identifying which genes are essential for T4P production in C. difficile. Additionally, a genomic analysis of the related pathogen Clostridium sordellii was performed, using the first high quality genome sequence produced for this species. Genes encoding T4P were identified, analysed and investigated. Furthermore, plasmids carrying the genes encoding the species’ key virulence factors (Lethal Toxin, TcsL, and in some cases haemorrhagic toxin, TcsH) were identified. These plasmids appear to be unstable, a fact with significant implications for diagnosis of C. sordellii disease.
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Hamdi, Cassandra. „Clostridium difficile : Rapid typing Clostridium difficile using MALDI-TOF MS analysis“. Thesis, Högskolan i Skövde, Institutionen för biovetenskap, 2019. http://urn.kb.se/resolve?urn=urn:nbn:se:his:diva-17659.
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Chang, Wei-Lun. „Acetone-Butanol-Ethanol Fermentation by Engineered Clostridium beijerinckii and Clostridium tyrobutyricum“. The Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1282108408.
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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/.
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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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Båverud, Viveca. „Clostridium difficile in horses /“. Uppsala : Dept. of Veterinary Microbiology, Swedish Univ. of Agricultural Sciences ([Institutionen för veterinärmedicinsk mikrobiologi], Sveriges lantbruksuniv.), 2002. http://epsilon.slu.se/avh/2002/91-576-6378-5.pdf.
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Robert, Céline Rabaud Christian. „Bactériémies à Clostridium spp“. [S.l.] : [s.n.], 2007. http://www.scd.uhp-nancy.fr/docnum/SCDMED_T_2007_ROBERT_CELINE.pdf.
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Cairns, Michelle Dawn. „Evolution of Clostridium difficile“. Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10060221/.
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Clostridium difficile continues to be a leading cause of healthcare-associated infections in the developed world. Increased detection of C. difficile infection (CDI) and development of typing schemes to differentiate between strains is primarily due to the recognition of global outbreaks of a single strain, BI/NAP1/027 which is characterised by three common typing techniques; restriction endonuclease analysis (REA), pulsed-field gel electrophoresis (PFGE) and PCR ribotyping. Phylogenetic analysis using multilocus sequence typing (MLST) divides C. difficile into five phylogenetic lineages which align the well-known PCR ribotypes; 027, 023, 017, 078 and a lineage containing diverse PCR ribotypes. MLST data in this thesis confirmed the five phylogenetic lineages were maintained after testing a larger collection of isolates from varied sources with further micro-diversity within the individual lineages. MLST investigation did not identify a lineage exclusive to nonhuman strains or any correlation between sequence type and geographical location. Data in this thesis also supports the notion that PCR ribotyping and REA do not correspond as well as previously considered. This may result in phylogenetically similar strains being designated as a different type or variant. The toxin A-B+ PCR ribotype 017 strain that forms a predominant lineage is little investigated. Through whole genome sequencing (WGS) and single nucleotide polymorphism (SNP) analysis, a historical clone of PCR ribotype 017 was identified from a London hospital ward. Although no phenotype exclusive to the clonal strain was characterised, this is the first report in the UK investigating the phylohistory of isolates from hospitalised patients with CDI due to PCR ribotype 017. Further investigation of PCR ribotype 017 with a larger and global collection of strains revealed two distinct sub-lineages containing multiple independent clonal expansions, antimicrobial resistant SNP determinants, deletions and insertions which were well distributed geographically and temporally. The data suggests transmission between humans and animals and findings support a USA origin with multiple, global transmission events. The key findings of this thesis are that C. difficile as a species is continually evolving with the appearance of divergent sub-lineages. WGS is superior to routine typing methodologies for tracking this evolution and will have significant impacts for outbreak investigation, understanding the phylohistory and phylogeography of C. difficile and other pathogens that are a threat to human health.
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Goya, Odile. „Clostridium et boues thermales“. Bordeaux 2, 1993. http://www.theses.fr/1993BOR2P039.
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Wheeldon, Laura J. „Studies on Clostridium difficile“. Thesis, Aston University, 2008. http://publications.aston.ac.uk/15406/.
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Clostridium difficile Is the major cause of nosocomial diarrhoea in the UK and is associated with high morbidity and mortality rates. There has been a large increase in cases of C. difficile associated disease (CDAD) in the last decade and It is thought that the emergence of the hypervirulent strain (ribotype 027) has contributed towards this rise. A major factor in the control and prevention of the disease is adequate cleaning of the clinical environment and disinfection, usually with chlorine based agents. However, the spores of C. difficile are highly resistant to many disinfectants.
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Caproni, Lisa J. „Antibiotics and Clostridium difficile“. Thesis, University of Edinburgh, 2004. http://hdl.handle.net/1842/24132.
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The aims of this thesis were to investigate the antibiotic susceptibility patterns of C. difficile with relation to the S-type of the isolates over a period of 18 months. Detailed growth curves were performed on strains NCTC 11223, the sequenced strain 630 and an endemic isolate 338a. Toxin A was shown to be produced upon entry to stationary phase in agreement with other studies. OD600 was found to be a good predictor of growth phase and allowed this measurement to be used for subsequent experiments. MICs were performed on 186 random isolates of C. difficile collected during an 18-month epidemiological study to investigate the patterns of sensitivity to six different antibiotics. No evidence of resistance was seen to the two treatment antibiotics and all strains were resistant to cefoxitin (MICs 64-256mg/ml), the antibiotic used in most selective media. Most strains (98.9%) had intermediate resistance to ceftriaxone. The MIC50 and MIC90 of the strains to amoxicillin and clindamycin were very close (8 and 16 for amoxicillin and 16 for clindamycin) but the range of MICs was great. Clindamycin resistance was common with 67% of strains resistant (MICs of > 8mg/ml), 25% with intermediate resistance (MIC > 4mg/ml) and only 8% sensitive (MICs of < 2mg/ml). Twelve isolates from six different patients had very high resistance to clindamycin with MICs > 128mg/ml. Multiple isolates from the same patient, taken at different times, showed changes in susceptibility patterns over time. The only major change in susceptibility over the time period was in clindamycin resistance; some strains appearing to become more resistant while others became less resistant. No differences were apparent in the MIC50 and MIC90 of the different S-types of C. difficile identified, although some S-types were present in very small numbers. No links between antibiotics prescribed and susceptibility patterns were found. Three strains (NCTC 11223, strain 630 and endemic isolate 338a) were cultured in sub-lethal concentrations of the six antibiotics (1/2,1/4 and1/8 of the MIC) over 104 hours and growth and toxin A measured three times a day. The effects varied between strain and antibiotic. The most common effect on the growth of the strains was to increase the initial lag period by ca. 4h compared to the antibiotic-free controls through the clindamycin resistant strain NCTC 11223, (MIC >512mg/ml) showed not lag whatsoever in comparison to the controls when grown in this antibiotic. The most common effect on toxin A production was in the onset of toxin elaboration. Normally toxin began to appear in low levels in early stationary phase before accumulating to high levels by the start of decline.
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Mordaka, Pawel Mateusz. „Reductions using Clostridium sporogenes“. Thesis, University of Nottingham, 2014. http://eprints.nottingham.ac.uk/14165/.
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Clostridium sporogenes was previously shown to be an extraordinary source for unusual reductases. It can catalyze reduction of wide a range of substrates such as nitroalkenes, enoates and nitro compounds, and can be used to generate chiral products. In preliminary studies, the ClosTron gene knock-out system for Clostridia was used to inactivate the fldZ gene assumed to encode the enzyme responsible for reduction of cinnamic acid in the reductive branch of L-phenylalanine fermentation via the Stickland reaction. Biotransformations with the fldZ mutant showed that C. sporogenes possesses multiple enzymatic activities, reducing enoates, β,β- and α,β-disubstituted nitroalkenes with different yields and enantioselectivities. The fldZ reductase was found to be responsible for reduction of cinnamic acid, (E)-1-nitro-2-phenylpropene, (E)-2-nitro-1-phenylpropene and β-nitrostyrene. However, the mutant could still reduce (E)-2-nitro-1-phenylpropene, β-nitrostyrene and cinnamic acid confirming the presence of other C=C double bond reductases in C. sporogenes. The analysis of the C. sporogenes genome sequence allowed identification of two hypothetical genes encoding proteins with homology to flavin-containing C=C double bond reductases, fldZ 2-enoate reductase and OYE-like reductase, which were subsequently cloned, overexpressed in E. coli under anaerobic conditions and tested for reduction of unsaturated compounds. The activity tests showed that fldZ possesses a narrow substrate range and can reduce only aromatic enoates such as cinnamic acid or p-coumaric acid. FldZ also reduced (E)-1-nitro-2-phenylpropene and (E)¬-2-nitro-1-phenylpropene with excellent and poor enantioselectivities (>99% and 16% respectively). On the other hand, the OYE-like reductase did not show activity towards unsaturated substrates in the activity assays and the substrate range of this reductase is unknown. Growth experiments comparing wild type C. sporogenes and the mutant in complex and minimal media showed that the fldZ reductase in not involved in the L-phenylalanine fermentation. Further analysis of the C. sporogenes genome resulted in identification of a novel reductase that might be involved in reduction of cinnamoyl-CoA to 3-phenylpropionyl-CoA in the Stickland reaction. Biocatalytic reduction of aromatic nitro compounds to amines can be used as alternative to chemo-reductive routes in preparation of pharmaceutical and agrochemical products. Protein extracts of C. sporogenes were found to reduce aromatic nitro compounds with different yields depending on the substrate structure and electron donor used in the reaction. The genome of C. sporogenes was screened and that allowed identification of six genes encoding hypothetical nitroreductases, which were subsequently overexpressed in E. coli. However, biotransformations using the recombinant nitroreductases did not show amine product formation. A novel Nylon 6 biosynthesis pathway was designed starting from biorenewable feedstocks. The crucial step in this pathway, reductive cleavage of pipecolic acid to 6-aminocaproic acid was proposed to be catalysed by C. sporogenes D-proline reductase. Thus, the activity of this enzyme was tested towards L- and D-pipecolic acid. Biotransformations showed that pipecolic acid was not accepted as a substrate. In the future, the idea of using D-proline reductase for Nylon biosynthesis may be exploited by improving the reductase activity using protein engineering techniques.
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Anderson, Michael Anthony. „Porcine Enteric Disease Caused by Clostridium difficile and Clostridium perfringens: Epidemiology, Pathogenesis and Immunity“. Diss., The University of Arizona, 2008. http://hdl.handle.net/10150/195681.
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Clostridium difficile and Clostridium perfringens are among the most commonagents of enteric disease in both humans and domestic animals. The former continues to increase in prevalence and diseases caused by the latter persist. Infection with a recently emergedhypervirulent strain (NAP1/027/III) of C. difficile is increasingly common andserious sequelae and fatalities are much more common in these patients. In neonatalpiglets, C. difficile infection (CDI) has become a common occurrence. Historically,isolation of C. perfringens type A from patients with enteric disease has been consideredinconsequential due to its presence in the normal intestine and to the mild nature ofdisease syndromes such as porcine enteritis. However, both CDI and type A diseasecause losses to the swine industry and pigs have been implicated as a possible source ofC. difficile for infection in humans. We investigated the epidemiology and pathogenesisof porcine CDI, and immunity against porcine CDI and type A enteritis. The occurrenceof CDI in integrated swine production facilities was most common in neonatal pigs.Infection in sows was rare, and finisher pigs were culture negative. All C. difficile strainswere ribotype 078. Hypervirulent strain NAP1/027/TTIII was more virulent in neonatalpigs than both a historic human historic human strain and a porcine strain with toxinproducing potential similar to ribotype 027 strains. Inoculation of anti-microbial-treatedadolescent pigs with NAP1/027/III did not cause disease. Ingra-gastric inoculation ofpigs with purified TcdA resulted in severe small intestine damage which isuncharacteristic of natural disease; effects of TcdB were minimal. Passive immunizationof piglets against C. difficile TcdA or C. perfringens type A alpha (CPA) and beta 2(CPB2) toxins did not prevent disease.
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Desvaux, Mickaël Petitdemange Henri. „La fermentation de la cellulose par Clostridium cellulolyticum métabolisme modèle d'un Clostridium cellulolytique mésophile /“. [S.l.] : [s.n.], 2001. http://www.scd.uhp-nancy.fr/docnum/SCD_T_2001_0174_DESVAUX.pdf.
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Desvaux, Mickaël. „La fermentation de la cellulose par Clostridium cellulolyticum : métabolisme modèle d'un Clostridium cellulolytique mésophile“. Nancy 1, 2001. http://docnum.univ-lorraine.fr/public/SCD_T_2001_0174_DESVAUX.pdf.
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Clostridium cellulolyticum est une bactérie cellulolytique anaérobie mésophile, dont le métabolisme carboné fut jusqu'à présent étudié sur cellobiose. En batch sur une cellulose native cette bactérie apparaît sensible à l'acidification tandis qu'à pH régulé l'arrêt de croissance est le résultat d'un flux carboné entrant élevé entraînant l'accumulation de composé(s) intracellulaire(s) inhibiteur(s). L'analyse des flux métaboliques en chémostat sur cellulose en conditions limitantes et/ou saturantes de carbone et/ou d'ammonium a montré que (i) le cellulosome régule le flux carboné entrant (ii) plus la cellodextrine incorporée est longue et plus il y a de glucose 1-phosphate généré (iii) la phosphoglucomutase contrôle l'entrée du flux carboné vers la glycolyse (iv) le glycogène absorbe le surplus de carbone (v) le noeud métabolique du pyruvate est capital dans le contrôle du flux électronique et énergétique (vi) les flux restent globalement inférieurs à ceux obtenus sur cellobioseSo far carbon metabolism of Clostridium cellulolyticum, a mesophilic cellulolytic anaerobic bacteria, has been only investigated with cellobiose. In batch with cellulose, this bacterium appears sensitive to the acidification while in pH-control culture the growth arrest results from a high entering carbon flow leading to the accumulation of intracellular inhibitory compound(s). The metabolic flux analysis in chemostat fed with cellulose under limited or saturated conditions of carbon and/or ammonium showed (i) the carbon entry was regulated by the cellulosome (ii) the longer the cellodextrin incorporated into the cell is, the more the glucose 1-phosphate is generated (iii) the phosphoglucomutase control the orientation of the flux towards glycolyse (iv) the glycogen buffer the carbon surplus (v) the pyruvate metabolic node is of key importance in electronic and energetic fluxes regulation (vi) the metabolic fluxes remain always lower with cellulose than those obtained with cellobiose
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Ferraris, Laurent. „Le microbiote intestinal du nouveau-né prématuré : le genre Clostridium et l'espèce Clostridium butyricum“. Paris 5, 2011. http://www.theses.fr/2011PA05P636.
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Chez le nouveau-né prématuré l’implantation du microbiote intestinal est considérée comme anormale, avec un retard d’implantation des bactéries anaérobies, en particulier des bifidobactéries. Concernant la colonisation intestinale par le genre Clostridium les données sont rares, alors que ce genre est impliqué dans des pathologies digestives, notamment l’entérocolite ulcéro-nécrosante (ECUN), première urgence gastro-intestinale chez le prématuré. Ce travail de thèse a eu deux objectifs : (1) étudier les espèces de Clostridium s’implantant au niveau intestinal chez les nouveau-nés prématurés, et identifier les facteurs périnataux influençant la colonisation ; et (2) réaliser une étude fondamentale sur l’espèce butyricum fréquemment impliquée dans l’ECUN. Nous avons mis en évidence que les espèces C. Perfringens, C. Difficile, C. Butyricum et C. Paraputrificum font parties du microbiote intestinal du nouveau-né prématuré sain. Les souches colonisatrices sont sensibles aux antibiotiques anti-anaérobies (excepté C. Difficile). Les antibiothérapies périnatale et néonatale influencent les niveaux de colonisation des clostridies. Le facteur majeur influençant la colonisation des Clostridium est le centre hospitalier, suggérant l’importance de la colonisation à partir de l’environnement. La comparaison différentielle des profils protéiques cytosoliques de souches de C. Butyricum isolées de prématurés ayant développé ou non une pathologie de type ECUN et de la souche de référence VPI3266 a permis de mettre en évidence des différences au niveau métabolique. L’existence d’un lien entre les différences observées et des caractéristiques de virulence dans le cadre de l’ECUN reste à démontrerIn the preterm neonates, the establishment of the intestinal microbiota is known to be abnormal, with a delay in the strictly anaerobic bacteria establishment, especially for bifidobacteria. As far as clostridia are concerned, data on their intestinal colonization neonates are scarce, although this genus is involved in gastrointestinal diseases, in particular in necrotizing enterocolitis (NEC), the main gastrointestinal emergency in premature neonates. This work aimed at (1) analyzing Clostridium species establishment in the gut of preterm neonates and identifying perinatal factors influencing this colonization, and (2) performing a basic study on the species C. Butyricum commonly involved in NEC. We have shown that the species C. Perfringens, C. Difficile, C. Butyricum and C. Paraputrificum are part of the intestinal microbiota of healthy preterm neonates. Colonizing strains were sensitive to anti-anaerobic antibiotics (except C. Difficile). The perinatal and neonatal antibiotic courses had influence on clostridial colonization levels. The major factor influencing the colonization was the hospital, suggesting the importance of the colonization from the environment. The comparison of the cytosolic protein profiles between C. Butyricum strains isolated from premature neonates who developed or not a NEC and the reference strain VPI3266 highlighted differences in the metabolism. A link between the observed differences and virulence characteristics of the strains in terms of NEC onset remains to be demonstrated
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Camiade, Émilie. „Étude de deux autolysines à activité N-acétylglucosaminidase chez Clostridium perfringens et Clostridium difficile“. Rouen, 2010. http://www.theses.fr/2010ROUES025.
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L'objectif de ce travail était de caractériser et d'étudier le rôle de deux peptidoglycane hydrolases à activité N-acétylglucosaminidase chez Clostridium perfringens et Clostridium difficile. Les peptidoglycane hydrolases possèdent en effet un rôle essentiel dans la croissance bactérienne, peuvent contribuer à la virulence de certaines espèces pathogènes, et peuvent être impliquées dans l'activité bactéricide des antibiotiques inhibiteurs de la synthèse de la paroi. Le lien phylogénétique des genres Staphylococcus et Clostridium nous a permis de caractériser, par analyse génomique, deux gènes de peptidoglycane hydrolases, acp et acd chez C. Perfringens et C. Difficile. Acp et Acd ont une organisation modulaire comparable comprenant un domaine catalytique à activité N-acétylglucosaminidase en C-terminal et un domaine d'ancrage composé de séquences répétées à motifs SH3_3 en position N-terminale. L'étude des fonctions de ces deux peptidoglycane hydrolases a ensuite été réalisée grâce à la construction de leurs mutants respectifs. Acp se révèle être impliquée (i) dans la séparation des cellules filles de C. Perfringens lors de sa croissance et (ii) dans la réponse à la lyse induite par les sels biliaires et la vancomycine. En ce qui concerne Acd de C. Difficile, son activité est vraisemblablement compensée par d'autres peptidoglycane hydrolases actuellement en cours de caractérisationThe objective of this work was to characterize and study the function of two peptidoglycan hydrolases from Clostridium perfringens and Clostridium difficile. Peptidoglycan hydrolases are implicated into the bacterial growth, may contribute to the virulence of certain pathogenic species and can be implicated in the bactericidal effect of cell wall-targeting antibiotics. The phylogenetic link between Staphylococcus and Clostridium allowed us to characterize, by genomic analysis, two genes of peptidoglycan hydrolases, Acp and Acd in C. Perfringens and C. Difficile. Acp and Acd have a modular organization composed of a C-terminal catalytic domain with N-acetylglucasaminidase activity and an anchoring domain composed of SH3_3 repeated sequences. Acp is involved (i) in the daughter cells separation of C. Perfringens during growth and (ii) in response to bile salts and vancomycin induced-lysis. For Acd of C. Difficile, its activity seems to be compensated by other peptidoglycan hydrolases that are actually in characterization
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BALDACINI, OLIVIER. „La cytotoxine de clostridium sordellii. Etude comparee a la toxine b de clostridium difficile“. Université Louis Pasteur (Strasbourg) (1971-2008), 1992. http://www.theses.fr/1992STR13037.
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La mise au point d'un protocole de purification rapide de la cytotoxine ou de la toxine l de clostridium sordellii nous a permis de comparer ses caracteristiques physicochimiques et biologiques et ses modalites d'action avec celles de la cytotoxine ou toxine b de clostridium difficile, egalement purifiee dans notre laboratoire. La toxine l et la toxine b, bien que presentant des homologies sur le plan immunologique, induisent des modifications morphologiques cellulaires distinctes: les deux toxines provoquent la depolymerisation des microfilaments mais les caracteristiques morphometriques et la distribution de l'actine f residuelle different selon que les cellules sont traitees par la toxine l ou la toxine b. Ces deux cytotoxines entrainent egalement des variations de phosphorylation de proteines cellulaires et en particulier de la tropomyosine et de la vimentine, deux composants du cytosquelette. Enfin, nous avons montre qu'un inhibiteur de phosphatases, l'acide okadaique, et la toxine l utilisent des mecanismes d'intoxication en partie communs. L'ensemble des resultats obtenus permet de proposer de nouvelles hypotheses quant aux mecanismes d'action des cytotoxines l et b, lesquelles agiraient en empruntant les voies de la phosphorylation-dephosphorylation cellulaires
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Reeve, Byron William Patrick. „Nitrogen metabolism and butanol production by South African clostridium beijerinckii and clostridium saccharobutylicum strains“. Doctoral thesis, University of Cape Town, 2014. http://hdl.handle.net/11427/12946.
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Includes bibliographical references.The acetone- butanol-ethanol (ABE) fermentation was one of the first fermentation processes to be industrialized on a large scale, and the dominant product, butanol is particularly significant due to its potential as a modern day fuel additive or fuel extender in the petrochemical industry. A collection of 19 solventogenic Clostridium beijerinckii and 11 Clostridium saccharobutylicum strains isolated from the National Chemical Products (NCP) ABE fermentation plant in Germiston, South Africa, were classed according to species by a quick species-specific colony PCR and by rifampicin screening methods respectively. The speciesspecific PCR aims to provide a rapid means of assessing any contamination of an ABE batch fermentation by differentiating between C. saccharobutylicum and C. beijerinckii species. Random Amplification of Polymorphic DNA (RAPD) analysis generated four C. beijerinckii and two C. saccharobutylicum strain groups respectively. Multilocus Sequence Typing (MLST) was developed for a smaller selection of strains and showed a further two strain groups within the NCP C. beijerinckii strains and three groups within the C. saccharobutylicum strains.
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Ulbrik, Teresa Yolanda Lustosa. „Cellulolytic fermentation by clostridium thermocellum“. Thesis, Georgia Institute of Technology, 1991. http://hdl.handle.net/1853/10027.
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Karlsson, Sture. „Toxin production in Clostridium difficile /“. Stockholm : Karolinska institutet, 2004. http://diss.kib.ki.se/2004/91-77349-812-2/.
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Permpoonpattana, Patima. „Clostridium difficile : infection and immunity“. Thesis, Royal Holloway, University of London, 2013. http://repository.royalholloway.ac.uk/items/33009ec4-7815-0803-d39b-f968c8d9cdbb/7/.
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Clostridium difficile is a Gram positive pathogen of significant importance in the UK, Europe and the USA. No vaccine has been developed and current treatments are focused on hospital management and the use of antibiotics. The disease is spread in hospitals in the spore form and the role of spores in C. difficile infecton is poorly understood. In this project spores of C. difficile have been characterised. The proteins from the outermost layers of the spore were identified and the genes cloned. Three of these surface proteins have unique enzymatic properties that maybe important for symptoms of disease. The ability of C. difficile spores to adhere to intestinal cells was found to be far greater than with live cells and through this we have identified that the spore may play an important role in colonisation. The regulation of spore coat gene expression during sporulation was also examined and temporal phases of genes expression identified. A major part of this project was to develop a mucosal vaccine to C. difficile. The approach used was to clone the C-terminus of toxin A onto the surface of Bacillus subtilis spores and use these recombinant spores to immunise mice and hamsters. We found that oral delivery of these spores conferred 75% protection to C. difficile infection in a hamster model of infection. Further, parenteral immunisation of the same antigens (toxin A and B) failed to generate mucosal responses and this showed that mucosal immunisation is critical for good protection. Finally, we found that antibodies to the C-terminus of toxin A were cross reactive to the C-terminus of toxin B. This showed that mucosal delivery of just the C-terminus of toxin A is sufficient to confer protection in an animal model of infection. The outcome of this work is that we have shown the parameters for successful immunisation and vaccination against C. difficile.
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Underwood, Sarah. „Sporulation initiation in Clostridium difficile“. Thesis, University of Leeds, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.505066.
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Clostridium difficile is a leading cause of hospital-acquired diarrhoea, responsible for over 30% of cases of antibiotic-associated colitis, nearly all cases of pseudomembranous colitis and costs the NHS over 200 million per year. This bacterium is able to persist in the hospital environment to cause recurrent infection by the formation of stable spores, refractile to current decontamination procedures. A more comprehensive understanding of the sporulation signal transduction pathway is essential for the design of a decontamination regime effective in removing the spores from the nosocomial environment and the logical design of novel antimicrobial agents. This project aimed to elucidate the mechanism of sporulation initiation . regulation and the role of sporulation-associated proteins in other C. difficile virulence processes, such as toxin production and colonisation. Analysis of sporulation in response to various hospital cleaning agents showed that the combination of a neutral detergent (such as Hospec) with EDTA is a more effective cleaning agent than the chlorine-based agents currently used, as the combination product is uniquely able to both kill vegetative cells and inhibit spore formation. A variety of molecular approaches were used to elucidate information regarding the C. difficile sporulation initiation pathway and the relationship between sporulation and toxin production. Three putative C. difficile sporulation-associated sensor histidine kinases (CD1A, CD2A and CD3B) were identified and shown to be independently involved in sporulation initiation. Furthermore, CD3B has been shown to directly phosphorylate the master response regulator SpoOA, strongly suggesting that this pathway is a two-component system, as opposed to the extended phosphore lay pathway found in B. subtilis. Previous studies on bacteria capable of both toxin production and endospore formation have described links between the two processes. Data presented here indicates SpoOA has a role in indirectly regulating C. difficile toxin A and B production, as the protein is capable of specifically binding promoter regions of the toxin regulatory genes tcdC and tcdD. Inoculation of a triple-stage continuous-culture chemostat that modelled the human gut with C. difficile spoDA- mutant provided further evidence that SpoDA has a key role in both colonisation a!1d toxin production. Overall, this work adds to the growing body of evidence that SpaDA is a master global regulator and has a crucial role in the pathogenicity of C. difficile, making it an excellent target for future novel antimicrobial therapies.
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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.
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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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Cross, Stephen. „The chaperonins of Clostridium thermocellum“. Thesis, University of Kent, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.294324.
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Martins, Luís Filipe Pires. „Clostridium difficile uma ameaça renovada“. Dissertação, Instituto de Ciências Biomédicas Abel Salazar, 2008. http://hdl.handle.net/10216/21105.
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César, Artur Jorge Fernandes. „Clostridium difficile - prevenção e controlo“. Dissertação, Faculdade de Medicina da Universidade do Porto, 2009. http://hdl.handle.net/10216/50362.
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Liew, Fung Min. „Metabolic engineering of Clostridium autoethanogenum“. Thesis, University of Nottingham, 2016. http://eprints.nottingham.ac.uk/32451/.
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Gas fermentation has emerged as a promising technology that converts waste gases containing CO, CO2 and H2 (also known as syngas) into fuels and chemical commodities. Employed by LanzaTech Inc., Clostridium autoethanogenum is an industrial acetogen that converts gases into ethanol, 2,3-butanediol, acetate, and lactate. Metabolic engineering offers unique opportunities to eliminate side-products, synthesize novel, high-value molecules as diversification strategies, and increase productivities of natural products. However, there had been no scientific reports of genetic manipulation of this acetogen so the overall goal of this PhD project was to develop genetic tools for this gas-utilizing microorganism and construct a hyper-ethanol producing strain via metabolic engineering. The formulation of electroporation and conjugation procedures allowed exogenous DNA to be routinely introduced into the bacterial host. ClosTron mutagenesis and Allele-Coupled Exchange (ACE) techniques were fully exemplified in this bacterium during the construction of knockout, in-frame deletion, and overexpression mutants. Carbon monoxide dehydrogenases (cooS1, cooS2 and acsA) were specifically targeted to elucidate their roles in supporting CO oxidation and carbon fixation. In the ethanol formation pathway, inactivation of bi-functional aldehyde/alcohol dehydrogenases (adhE1 and adhE2) impaired growth on pure CO but elevated ethanol titres. Conversely, inactivation of the more highly expressed aldehyde:ferredoxin oxidoreductase (aor1), but not the weakly expressed aor2, significantly reduced ethanol production, highlighting the importance of aor1 in autotrophic ethanol formation. A double KO mutant of aor1 and aor2 was also generated via ClosTron mutagenesis and pyrE-mediated allelic exchange. In an effort to engineer a robust biocatalyst, the native chaperone systems groESL and/or grpE-dnaK-dnaJ were overexpressed in C. autoethanogenum, resulting in enhanced tolerance towards ethanol, heat and salts. In summary, this study demonstrated the genetic tractability of C. autoethanogenum and revealed gene targets for future metabolic engineering of a hyper-ethanol producing acetogen.
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Canard, Bruno. „Organisation genomique de clostridium perfringens“. Paris 7, 1991. http://www.theses.fr/1991PA077145.
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Clostridium perfringens est un bacille a gram positif, pathogene pour l'homme et les animaux. Nous avons construit une carte physique et genetique du chromosome de la souche de reference cpn50, de type a, ainsi que de sept autres isolats appartenant aux serotypes a, b, d et e. Nous avons ainsi pu etudier comparativement l'organisation genomique des genes et loci impliques dans la maintenance des fonctions vitales de la bacterie, dans la virulence, et dans la plasticite genomique. Nous avons caracterise respectivement au niveau moleculaire les operons ribosomiques rrn, le gene nagh codant pour une n-acetyl-beta-d-glucosaminidase, et neuf regions genomiques sujettes a des polymorphismes de restriction. Quatre de ces dernieres sont associees a des facteurs de virulence, et une de ces quatre tolere des variations en taille de l'ordre de 0,5 megabase. L'organisation des genes de virulence et la plasticite genomique sont etroitement imbriquees
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Hecker, Kim Ione. „Bleach-It-Away Clostridium difficile“. ScholarWorks, 2018. https://scholarworks.waldenu.edu/dissertations/5471.
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Hospital-associated infections (HAIs) are infections patients contract as a result of being hospitalized. HAI rates decreased for almost all pathogens in the past few years, with the exception of Clostridium difficile infections (CDIs), which have been steadily climbing, placing hospital-acquired CDI at the top of the HAI list. The Center for Disease Control and Prevention reported in 2010 almost a half a million people were infected with CDIs yearly in the United States, and CDIs claimed the lives of approximately 29,000 people, representing a 4-fold increase from 1993. To address the problem in the local hospital, a quality improvement initiative called Bleach-It-Away was initiated. The initiative involved nurses wiping down the high touch areas in the patient's medical intensive care (MICU) rooms once every shift. The purpose of this quantitative research project was to evaluate the effectiveness of the Bleach-It-Away practice. The project question asked if the Bleach-It-Away practice was effective in reducing CDI rates. Deidentified CDI rates were provided by the clinical practice site covering a period of 12 months prior to implementation and 12 months after implementation of the practice. An independent t-test was used to determine whether there were significant improvements in CDI rates in the MICU. No significant improvement was seen in the postimplementation total CDI rates (p=.07) compared to the preimplementation rates. While the process did not demonstrate a significant improvement, positive social change is possible as hospitals recognize the many factors contributing to CDIs and the need for collaboration from various disciplines to control the problem.
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Martins, Luís Filipe Pires. „Clostridium difficile uma ameaça renovada“. Master's thesis, Instituto de Ciências Biomédicas Abel Salazar, 2008. http://hdl.handle.net/10216/21105.
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César, Artur Jorge Fernandes. „Clostridium difficile - prevenção e controlo“. Master's thesis, Faculdade de Medicina da Universidade do Porto, 2009. http://hdl.handle.net/10216/50362.
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Guttenberg, Gregor [Verfasser], und Manfred [Akademischer Betreuer] Jung. „Clostridiale Glukosylierende Toxine: Untersuchungen zur Autoprozessierung von Clostridium sordellii Letalem Toxin und Clostridium novyi alpha-Toxin sowie funktionelle Charakterisierung von Clostridium perfringens TpeL-Toxin“. Freiburg : Universität, 2012. http://d-nb.info/1123467994/34.
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Hoelzle, Robert Donald. „Genetic improvement of Clostridium tyrobutyricum for butanol production by insertion of adhE from Clostridium acetobutylicum“. Connect to resource, 2010. http://hdl.handle.net/1811/45459.
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Ticchi, Laurence. „Clostridium difficile et ses toxines : prévalence de "Clostridium difficile" et de la toxine A asymptomatique“. Paris 5, 1990. http://www.theses.fr/1990PA05P183.
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Schué, Véronique. „Comparaison des cytotoxines de clostridium difficile et clostridium sordellii : effets cellulaires et subcellulaires, support genetique“. Université Louis Pasteur (Strasbourg) (1971-2008), 1994. http://www.theses.fr/1994STR13229.
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Notre travail a porte essentiellement sur l'etude comparative des cytotoxines b et l de clostridium difficile et clostridium sordellii. Nous avons pu realiser l'analyse des caracteristiques moleculaires et cellulaires des cytotoxines b et l qui presentent des parentes immunologiques. Mais, malgre cette parente, les modifications morphologiques induites par les deux cytotoxines sur modele cellulaire approprie sont fondamentalement distinctes: la cytotoxine b de c. Difficile hyperphosphoryle la calnexine, proteine chaperone ; alors que la cytotoxine l de c. Sordellii phosphoryle la caldesmone, proteine du cytosquelette. En fait, les modifications morphologiques induites par les deux cytotoxines sont la resultante de mecanismes d'action complexes, utilisant notamment les voies de la phosphorylation et de la dephosphorylation. Les etudes plus fines de caracterisation moleculaire a savoir le clonage et le sequencage du gene de la cytotoxine l de c. Sordellii, impliquent une homologie genetique et moleculaire (76%) entre les deux cytotoxines. Ces etudes laissent penser que les deux cytotoxines peuvent decouler d'un support genetique ancestral commun, et apportent un debut de justification en suggerant la creation d'une classe nouvelle de toxines clostridiennes genetiquement proches
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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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Kawaichi, Marisa Emiko. „Efeito da inoculação de esporos de Clostridium estertheticum e Clostridium gasigenes em carne bovina ambalada a vácuo e capacidade de Clostridium estertheticum de formar biofilmes“. [s.n.], 2010. http://repositorio.unicamp.br/jspui/handle/REPOSIP/255466.
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Orientador: Arnaldo Yoshiteru KuayeDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos
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Resumo: Casos de estufamento de carne bovina embalada à vácuo, causado por Clostridium estertheticum e Clostridium gasigenes, mantida sob temperatura de refrigeração vêm sendo observados em diversas regiões do Brasil, principalmente no Centro-Oeste, onde foi detectada a presença destes microrganismos em ambientes de abatedouros-frigoríficos. A complexidade analítica da técnica utilizada em pesquisas com esses microrganismos faz com que sejam escassos os trabalhos que visam o controle desse tipo de deterioração. Diante do exposto e considerando-se a importância da exportação de carne para a economia brasileira, o presente estudo teve como objetivo avaliar o efeito da inoculação de esporos de Clostridium estertheticum e Clostridium gasigenes em carne bovina embalada a vácuo e a eficiência esporicida in vitro do uso de ácidos orgânicos sobre estes microrganismos. Além disso, avaliou-se a capacidade de formação de biofilmes por C. estertheticum em superfície de aço inoxidável, e a eficiência de agentes químicos para sua remoção. Observou-se em carnes embaladas a vácuo, que C. gasigenes produzem maiores quantidades de gases, promovendo o estufamento da embalagem mais rápido em relação ao C. estertheticum. O primeiro indício da formação de bolhas por C. gasigenes foi observado aos 21 dias de incubação à temperatura de 7°C de armazenamento, enquanto que para C. estertheticum observou-se somente aos 35 dias na mesma temperatura. Notou-se uma produção de gás mais intensa à temperatura de armazenamento mais alta. Na carne inoculada com C. gasigenes houve um aumento progressivo no valor do pH, atingindo valores maiores em temperaturas mais elevadas, por volta de 49-77 dias, seguido por uma queda. Por sua vez, a inoculação com C. estertheticum não promoveu a mesma variação de pH, notando-se uma acentuada queda até 21-35 dias, seguido por um progressivo aumento. A utilização, de ácidos orgânicos mostrou reduzido ou nenhum efeito sobre esporos, de C. gasigenes e C. estertheticum, não sendo viável sua aplicação em cortes de carnes bovinas com a finalidade de inibir ou minimizar o blown pack. Observou-se que tanto a cepa de C. estertheticum padrão DSM 8809T quanto a cepa LHCE-13 isolada de equipamento (rolete de retirada do couro) foram capazes de formar biofilmes após 07 dias de contato com a superfície de aço inoxidável e os sanitizantes, ácido peracético (500mg/L) e peróxido de hidrogênio (200 mg/L) em contato por 15 minutos, mostraram-se eficientes no controle destes. São necessários programas de higienização mais rigorosos e efetivos para o controle de C. estertheticum e C. gasigenes nos abatedouros-frigoríficos, pois eles podem estar presentes no ambiente em forma de esporos e também como biofilmes, em locais com grande acúmulo de material orgânico ou em associações com outros microrganismos
Abstract: Occurrence of blown pack in vacuum packaged refrigerated meat caused by Clostridium estertheticum and Clostridium gasigenes have been reported by studies from several Brazilian states where these microorganisms were detected in abattoir environment such as the Midwestern region. Analytical complexity of the researches with C. estertheticum e C. gasigenes makes the studies to prevent or to control this spoilage scarce. For these reasons and it is considering the importance of Brazil¿s beef export to economy, the current study aim to assess the effect of inoculated spores of C. estertheticum and C. gasigenes on vaccum packaged meat and the efficiency in vitro of organic acid on reported microorganisms spores. Furthermore this study tested, for the first time, the ability of C. estertheticum to form biofilm in stainless steel surface and the use of some sanitizers to remove it. Results obtained from behavior analysis of C. estertheticum and C. gasigenes spores inoculated in vacuum packaged meat, showed us differences in growth between these microorganisms. C. gasigenes produced more amount of gas, leading the blown pack faster than C. estertheticum. The first evidence of gas bubble production by C. gasigenes was observed after 21 storage days at 7°C, whereas C. estertheticum started to produce bubbles only after 35 days at the same storage conditions. Gas production was intense in higher storage temperatures. Through the pH measurement it was possible verify that C. gasigenes increases progressively the pH, reaching maximum values after 49-77 days at higher storage temperatures, followed by decreasing. Moreover C. estertheticum do not cause variation in the pH, but it may be seen an accentuated decrease until 21-35 days, followed by an increase of the values, probably because of C. estertheticum ability to revert a falling pH through fast lactate fermentation, started when availability of glucose ends. Regarding to use of organic acids, there is reduced or none effect on C. gasigenes and C. estertheticum spores, it becomes this technique no viable to reduce or control blown pack in vacuum packaged meat. For the first time is demonstrate that C. estertheticum forms biofilm. Strains of C. estertheticum DSM 8809T and C. estertheticum LHCE-13 isolated from abattoir equipment (leather removal drums) are able to colonize surfaces of stainless steel in 07 days. The sanitizers peracetic acid (500mg/L) and hydrogen peroxide (200 mg/L) exposure in 5-25 day-old biofilms at least 15 minutes were effective to control them. Severe and effective hygienization program is needed to control C. estertheticum and C. gasigenes in abattoir environment, mainly because they may be present on equipment surfaces in spore forms and also through biofilm formation in places with high organic material presence
Mestrado
Engenharia de Alimentos
Mestre em Tecnologia de Alimentos
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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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Perelle, Sylvie. „Toxine IOTA de "Clostridium perfringens" et toxines apparentées“. Paris 11, 1996. http://www.theses.fr/1996PA114811.
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Ali, Ali Abdulkareem Ali. „Isolation and characterization of Clostridium perfringens bacteriophages and optimization of electro-transformation parameters for Clostridium difficile“. Thesis, University of Leicester, 2018. http://hdl.handle.net/2381/42289.
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Clostridium perfringens (C. perfringens) is responsible for a variety of diseases in humans and animals. Clostridium difficile (C. difficile) is the leading cause of antibiotic-associated diarrhoea. The available treatments for infections caused by both pathogens are not effective. Bacteriophage (phage) therapy is a promising treatment strategy to tackle and treat any infections or diseases caused by C. perfringens or C. difficile. The aim of this study was to isolate bacteriophages that infect C. perfringens and characterize them, and to optimize electroporation parameters for C. difficile with the end goal being to be able to genetically modify C. difficile phages. Five phages that infect C. perfringens were isolated and characterized from environmental samples. Two phages were sequenced and annotated, one was found to be a Podovirus and the other a Siphovirus. The Podovirus is a strictly lytic phage that does not possess any undesired genes such as the transduction gene, antibiotic resistance gene, and toxin genes. The endolysin of the Podovirus was cloned, expressed, and purified. The muralytic activity of the enzyme was confirmed by a zymogram. This endolysin has the ability to completely lyse 85.7 % of the tested Clostridium perfringens strains. A series of electroporation experiments were carried out using many experimental settings and varying parameters in order to deliver engineered phage and plasmid DNA genome to C. difficile. The electroporation refractory C. difficile R076 was treated with a cysteine protease inhibitor E- 64 and lysostaphin to facilitate electroporation. E-64 was able to reduce the thickness of the C. difficile surface layer proteins. The treatment with lysostaphin resulted in cell lysis. Unfortunately, all the attempts made to optimize the electroporation protocol for C. difficile were unsuccessful as no cells were transformed. However, the experimental observations provide a strong foundation for further work to develop an effective electro-transformation protocol for C. difficile.
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Andre, Gaëlle. „Mécanismes de régulation en réponse à la disponibilité en soufre chez Clostridium acetobutylicum et Clostridium perfingens“. Paris 6, 2009. http://www.theses.fr/2009PA066322.
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Au cours de ce travail, nous avons étudié le métabolisme du soufre chez deux clostridies, Clostridium acetobutylicum, une bactérie non-pathogène d’intérêt industriel, et Clostridium perfringens, une bactérie pathogène pour l’homme. Nous avons identifié un mécanisme original de régulation de l’opéron ubiGmccBA responsable de la conversion de la méthionine en cystéine chez C. Acetobutylicum. Cet opéron est régulé par un mécanisme complexe impliquant un ARN antisens dont la synthèse est réprimée par la méthionine via une boîte S. Cet ARN antisens agirait en cis par interférence transcriptionnelle. Chez C. Perfringens, l’expression de l’opéron ubiGmccBAluxS est régulée par une boîte T spécifique de la cystéine, par le petit ARN régulateur virX et par le système VirR-VirS via le petit ARN régulateur, le VR-RNA. Nous avons également montré qu’une carence en cystéine module l’expression des voies de biosynthèse et d’entrée de la cystéine, de protéines impliquées dans la biogenèse des centres [Fe-S], de certaines voies de fermentation et d’utilisation de sources de carbone et de systèmes redox qui pourraient participer à la résistance aux espèces réactives de l’oxygène. Parmi les gènes induits en carence en cystéine, nous avons identifié le régulateur de la biogenèse des centres [Fe-S], Cpe1786, qui contrôle aussi les voies de fermentation et des enzymes de dégradation de composés de l’hôte.