Articles de revues : « Beta-2 toxin. Clostridium perfringens »

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TSIOURIS (Β.Σ. ΤΣΙΟΥΡΗΣ), V. S., I. GEORGOPOULOU (ΓΕΩΡΓΟΠΟΥΛΟΥ) et E. PETRIDOU (Ε. ΠΕΤΡΙΔΟΥ). « Update on the toxins of Clostridium perfringens and their actions ». Journal of the Hellenic Veterinary Medical Society 61, n^o 3 (17 novembre 2017) : 241. http://dx.doi.org/10.12681/jhvms.14892.

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Clostridia appeared as a distinct class, approximately 2.7 billion years ago, before the initial formation of oxygen. Clostridium perfringens is widely distributed throughout the environment due to its ability to form spores. Furthermore, it is a member of intestinal microbiota in animals and human. In 2002, the complete genome of C perfringens strain 13 was published. Genomic analysis has revealed that C. perfringens lacks the genetic machinery to produce 13 essential amino acids and it obtains these in vivo via the action of its toxins. Toxins of C perfringens can be divided into major, minor and enterotoxin. C perfringens strains are classified into five toxinotypes (A, B, C, D and E), based on the production of four major toxins. Alpha toxin is the best and most studied major toxin of C perfringens and it was the first bacterial toxin established to possess enzymatic activity. It has haemolytic, necrotic and cytolytic activity, it can lyse platelets and leukocytes and it can damage fibroblasts and muscle cell membranes. Expression of epa gene, which is responsible for the production of alpha toxin by C perfringens, is down-regulated in the normal healthy gut, but it is upregulated to initiate enteric disease in response to an environmental signal. C perfringens appears to be regulated in a quorum sensing manner, using oligopeptides, AI-2 or both, to regulate expression of the epa gene, and thus the synthesis of alpha toxin. Beta toxin is recognized as an important agent in necrotic enteritis of humans and it is the second most lethal C. perfringens toxin following epsilon toxin. Beta toxin is a membrane spanning protein that oligomerizes to form channels in susceptible cells or it primarily acts as a neurotoxin. Epsilon toxin is the most potent of the C. perfringens toxins and the third most potent neurotoxin from the Clostridium spp., following botulinum and tetanus toxins. Epsilon toxin of C perfringens type D causes enterotoxaemia and pulpy kidneys disease of lambs. Iota toxin causes disruption of the actin cytoskeleton and cell barrier integrity and it is the less toxic of the major toxins of C perfringens. Although C perfringens enterotoxin is not classified as one of the major toxins of C perfringens, it is the third most common cause of food poisoning in industrialized nations. It is not secreted by the cells of growing bacteria, but it is released only with the sporulation of C perfringens. Not all strains of C perfringens carry the epe gene, which is responsible for the production of enterotoxin. Theta toxin is a pore-forming cytolysin that can lyse red blood cells. It is produced by all types of C perfringens. Together with alpha-toxin, theta-toxin modulates the host inflammatory response. ß2 toxin is a pore forming toxin which is involved in necrotic enteritis of swine and horse, in haemorragic enteritis of bovine in diarrhea cases of dogs and along with enterotoxin in diarrhea cases of humans. Recently, -NetB, a novel toxin that is associated with broiler necrotic enteritis, has been described. The mechanism of its action seems to involve the formation of small hydrophilic pores. Other toxins of C. perfringens include λ-toxin, ô-toxin, μ-toxin, v-toxin, κ-toxin, a-clostripain like protease and neuraminidase/sialidase. These toxins can act as enzymes, while many of them can act synergically or supplementally with major pore forming toxins. Potentially, C. perfringens might produce more toxins, which have not been identified. Finally, the actions of C. perfringens toxins, major or minor, in some diseases have not been figured out.

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Almeida, J. C., R. O. S. Silva, F. C. F. Lobato et R. A. Mota. « Isolation of Clostridium perfringens and C. difficile in crab-eating fox ( Cerdocyon thous - Linnaeus 1776) from Northeastern Brazil ». Arquivo Brasileiro de Medicina Veterinária e Zootecnia 70, n^o 6 (décembre 2018) : 1709–13. http://dx.doi.org/10.1590/1678-4162-9895.

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ABSTRACT The aim of the present study was to isolate Clostridium perfringens and C. difficile in crab-eating fox (Cerdocyon thous) from Northeastern Brazil. Stool samples of 18 captive crab-eating foxes from four states of Northeastern Brazil (Alagoas, Bahia, Paraíba e Pernambuco) were collected and subjected to C. perfringens and C. difficile isolation. Suggestive colonies of C. perfringens were then analyzed for genes encoding the major C. perfringens toxins (alpha, beta, epsilon and iota), beta-2 toxin (cpb2), enterotoxin (cpe), and NetB- (netB) and NetF- (netF) encoding genes. C. difficile strains were analyzed by multiplex-PCR for a housekeeping gene (tpi), toxins A (tcdA) and B (tcdB) and a binary toxin gene (cdtB). Unthawed aliquots of stool samples positive for toxigenic C. difficile were subjected to a commercial ELISA to evaluate the presence of A/B toxins. Clostridium perfringens (type A) was isolated from five (27%) samples, and only one sample was positive for beta-2 enconding gene (cpb2). Two (11%) stool samples were positive for C. difficile, but negative for A/B toxins. These two wild canids were also positive for C. perfringens type A. This is the first report of C. difficile in crab-eating fox.

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Songer, J. Glenn, et Francisco A. Uzal. « Clostridial Enteric Infections in Pigs ». Journal of Veterinary Diagnostic Investigation 17, n^o 6 (novembre 2005) : 528–36. http://dx.doi.org/10.1177/104063870501700602.

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Clostridium perfringens types A and C and Clostridium difficile are the principal enteric clostridial pathogens of swine. History, clinical signs of disease, and gross and microscopic findings form the basis for a presumptive diagnosis of C. perfringens type-C enteritis. Confirmation is based on isolation of large numbers of type-C C. perfringens and/or detection of beta toxin in intestinal contents. Diagnosis of C. perfringens type-A infection, however, remains controversial, mostly because the condition has not been well defined and because type-A organisms and their most important major (alpha) toxin can be found in intestinal contents of healthy and diseased pigs. Isolation of large numbers of C. perfringens type A from intestinal contents, in the absence of other enteric pathogens, is the most reliable criterion on which to base a diagnosis. Recently, beta2 (CPB2) toxin-producing C. perfringens type A has been linked to disease in piglets and other animals. However, implication of CPB2 in pathogenesis of porcine infections is based principally on isolation of C. perfringens carrying cpb2, the gene encoding CPB2, and the specific role of CPB2 in enteric disease of pigs remains to be fully defined. Clostridium difficile can also be a normal inhabitant of the intestine of healthy pigs, and diagnosis of enteric infection with this microorganism is based on detection of its toxins in feces or intestinal contents.

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Gurjar, Abhijit, Jihong Li et Bruce A. McClane. « Characterization of Toxin Plasmids in Clostridium perfringens Type C Isolates ». Infection and Immunity 78, n^o 11 (7 septembre 2010) : 4860–69. http://dx.doi.org/10.1128/iai.00715-10.

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ABSTRACT Clostridium perfringens type C isolates cause enteritis necroticans in humans or necrotizing enteritis and enterotoxemia in domestic animals. Type C isolates always produce alpha toxin and beta toxin but often produce additional toxins, e.g., beta2 toxin or enterotoxin. Since plasmid carriage of toxin-encoding genes has not been systematically investigated for type C isolates, the current study used Southern blot hybridization of pulsed-field gels to test whether several toxin genes are plasmid borne among a collection of type C isolates. Those analyses revealed that the surveyed type C isolates carry their beta toxin-encoding gene (cpb) on plasmids ranging in size from ∼65 to ∼110 kb. When present in these type C isolates, the beta2 toxin gene localized to plasmids distinct from the cpb plasmid. However, some enterotoxin-positive type C isolates appeared to carry their enterotoxin-encoding cpe gene on a cpb plasmid. The tpeL gene encoding the large clostridial cytotoxin was localized to the cpb plasmids of some cpe-negative type C isolates. The cpb plasmids in most surveyed isolates were found to carry both IS1151 sequences and the tcp genes, which can mediate conjugative C. perfringens plasmid transfer. A dcm gene, which is often present near C. perfringens plasmid-borne toxin genes, was identified upstream of the cpb gene in many type C isolates. Overlapping PCR analyses suggested that the toxin-encoding plasmids of the surveyed type C isolates differ from the cpe plasmids of type A isolates. These findings provide new insight into plasmids of proven or potential importance for type C virulence.

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Nagahama, Masahiro, Sadayuki Ochi, Masataka Oda, Kazuaki Miyamoto, Masaya Takehara et Keiko Kobayashi. « Recent Insights into Clostridium perfringens Beta-Toxin ». Toxins 7, n^o 2 (3 février 2015) : 396–406. http://dx.doi.org/10.3390/toxins7020396.

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Sakurai, Jun, et Masahiro Nagahama. « CLOSTRIDIUM PERFRINGENS BETA-TOXIN : CHARACTERIZATION AND ACTION ». Toxin Reviews 25, n^o 1 (janvier 2006) : 89–108. http://dx.doi.org/10.1080/15569540500320979.

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Greco, Grazia, Anna Madio, Vito Martella, Marco Campolo, Marialaura Corrente, Domenico Buonavoglia et Canio Buonavoglia. « Enterotoxemia Associated with Beta2 Toxin–Producing Clostridium Perfringens Type A in Two Asiatic Black Bears (Selenarctos Thibetanus) ». Journal of Veterinary Diagnostic Investigation 17, n^o 2 (mars 2005) : 186–89. http://dx.doi.org/10.1177/104063870501700216.

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Beta2 (β2) toxin–producing Clostridium perfringens type A strains were found to be associated with necrotic and hemorrhagic intestinal lesions in 2 Asiatic black bears ( Selenarctos thibetanus) that died suddenly. Ten isolates were obtained from the liver, lungs, heart, and small and large intestine of the animals and were examined by multiplex polymerase chain reaction for the genes encoding the 4 lethal toxins (alpha, beta, epsilon, and iota) for classification into toxin types as well as for the genes encoding enterotoxin and the novel β2-toxin for subclassification. In addition, the cpb2 sequence of the 10 isolates was different from the published sequence of cpb2 of pig type C isolate CWC245, whereas it was highly similar to the cpb2 sequence of the C. perfringens type A strain 13. This finding suggests the existence of 2 cpb2 subtypes. This is the first report of enterotoxemia associated with the presence of C. perfringens producing β2-toxin in the tissues and intestinal content of Asiatic black bears.

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Diab, S. S., H. Kinde, J. Moore, M. F. Shahriar, J. Odani, L. Anthenill, G. Songer et F. A. Uzal. « Pathology of Clostridium perfringens Type C Enterotoxemia in Horses ». Veterinary Pathology 49, n^o 2 (18 avril 2011) : 255–63. http://dx.doi.org/10.1177/0300985811404710.

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Clostridium perfringens type C is an important cause of enteritis and enterocolitis in foals and occasionally in adult horses. The disease is a classic enterotoxemia, and the enteric lesions and systemic effects are caused primarily by beta toxin, 1 of 2 major toxins produced by C. perfringens type C. Until now, only sporadic cases of C. perfringens type C equine enterotoxemia have been reported. We present a comprehensive description of the lesions in 8 confirmed cases of type C enterotoxemia in foals and adult horses. Grossly, multifocal to segmental hemorrhage and thickening of the intestinal wall were most common in the small intestine, although the colon and cecum were also frequently affected. All horses had variable amounts of fluid, often hemorrhagic intestinal contents. The most characteristic microscopic lesion was necrotizing or necrohemorrhagic enteritis, with mucosal and/or submucosal thrombosis. Numerous gram-positive rods were occasionally seen in affected mucosa. A definitive diagnosis of C. perfringens type C enterotoxemia in all 8 cases was based on the clinical history, gross and histologic lesions, and detection of the beta toxin in intestinal contents.

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Ferreira, Thais Sebastiana Porfida, Andrea Micke Moreno, Renata Rodrigues de Almeida, Cleise Ribeiro Gomes, Debora Dirani Sena de Gobbi, Pedro Henrique Nogueira de Lima Filsner et Marina Moreno. « Molecular typing of Clostridium perfringens isolated from swine in slaughterhouses from São Paulo State, Brazil ». Ciência Rural 42, n^o 8 (août 2012) : 1450–56. http://dx.doi.org/10.1590/s0103-84782012000800020.

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Clostridium perfringens is an anaerobic Gram-positive bacterium known as common pathogen for humans, for domestic and wildlife animals. Although infections caused by C. perfringens type C and A in swine are well studied, just a few reports describe the genetic relationship among strains in the epidemiological chain of swine clostridioses, as well as the presence of the microorganism in the slaughterhouses. The aim of the present study was to isolate C. perfringens from feces and carcasses from swine slaughterhouses, characterize the strains in relation to the presence of enterotoxin, alpha, beta, epsilon, iota and beta-2 toxins genes, using polymerase chain reaction (PCR) and comparing strains by means of Pulsed field gel electrophoresis (PFGE). Clostridium perfringens isolation frequencies in carcasses and finishing pig intestines were of 58.8% in both types of samples. According to the polymerase chain reaction assay, only alfa toxin was detected, being all isolates also negative to enterotoxin and beta2 toxin. Through PFGE technique, the strains were characterized in 35 pulsotypes. In only one pulsotype, the isolate from carcass sample was grouped with fecal isolate of the same animal, suggesting that the risk of cross-contamination was low. Despite the high prevalence of C. perfringens in swine carcasses from the slaughterhouses assessed, the risk of food poisoning to Brazilian pork consumers is low, since all strains were negative to cpe-gene, codifying enterotoxin.

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Fisher, Derek J., Mariano E. Fernandez-Miyakawa, Sameera Sayeed, Rachael Poon, Victoria Adams, Julian I. Rood, Francisco A. Uzal et Bruce A. McClane. « Dissecting the Contributions of Clostridium perfringens Type C Toxins to Lethality in the Mouse Intravenous Injection Model ». Infection and Immunity 74, n^o 9 (septembre 2006) : 5200–5210. http://dx.doi.org/10.1128/iai.00534-06.

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ABSTRACT The gram-positive anaerobe Clostridium perfringens produces a large arsenal of toxins that are responsible for histotoxic and enteric infections, including enterotoxemias, in humans and domestic animals. C. perfringens type C isolates, which cause rapidly fatal diseases in domestic animals and enteritis necroticans in humans, contain the genes for alpha toxin (plc), perfringolysin O (pfoA), beta toxin (cpb), and sometimes beta2 toxin (cpb2) and/or enterotoxin (cpe). Due to the economic impact of type C-induced diseases, domestic animals are commonly vaccinated with crude type C toxoid (prepared from inactivated culture supernatants) or bacterin/toxoid vaccines, and it is not clear which toxin(s) present in these vaccines actually elicits the protective immune response. To improve type C vaccines, it would be helpful to assess the contribution of each toxin present in type C supernatants to lethality. To address this issue, we surveyed a large collection of type C isolates to determine their toxin-producing abilities. When late-log-phase vegetative culture supernatants were analyzed by quantitative Western blotting or activity assays, most type C isolates produced at least three lethal toxins, alpha toxin, beta toxin, and perfringolysin O, and several isolates also produced beta2 toxin. In the mouse intravenous injection model, beta toxin was identified as the main lethal factor present in type C late-log-phase culture supernatants. This conclusion was based on monoclonal antibody neutralization studies and regression analyses in which the levels of alpha toxin, beta toxin, perfringolysin O, and beta2 toxin production were compared with lethality. Collectively, our results highlight the importance of beta toxin for type C-induced toxemia.

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El-sify, Ahmed. « A review on Clostridium perfringens toxins with special reference to Beta 2 toxin ». Journal of Current Veterinary Research 9, n^o 1 (1 septembre 2015) : 85–100. http://dx.doi.org/10.21608/jcvr.2015.37191.

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Sakurai, Jun, et Yoshio Fujii. « Purification and characterization of Clostridium perfringens beta toxin ». Toxicon 25, n^o 12 (janvier 1987) : 1301–10. http://dx.doi.org/10.1016/0041-0101(87)90008-0.

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Duracova, Miloslava, Jana Klimentova, Alena Myslivcova Fucikova, Lenka Zidkova, Valeria Sheshko, Helena Rehulkova, Jiri Dresler et Zuzana Krocova. « Targeted Mass Spectrometry Analysis of Clostridium perfringens Toxins ». Toxins 11, n^o 3 (23 mars 2019) : 177. http://dx.doi.org/10.3390/toxins11030177.

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Targeted proteomics recently proved to be a technique for the detection and absolute quantification of proteins not easily accessible to classical bottom-up approaches. Due to this, it has been considered as a high fidelity tool to detect potential warfare agents in wide spread kinds of biological and environmental matrices. Clostridium perfringens toxins are considered to be potential biological weapons, especially the epsilon toxin which belongs to a group of the most powerful bacterial toxins. Here, the development of a target mass spectrometry method for the detection of C. perfringens protein toxins (alpha, beta, beta2, epsilon, iota) is described. A high-resolution mass spectrometer with a quadrupole-Orbitrap system operating in target acquisition mode (parallel reaction monitoring) was utilized. Because of the lack of commercial protein toxin standards recombinant toxins were prepared within Escherichia coli. The analysis was performed using proteotypic peptides as the target compounds together with their isotopically labeled synthetic analogues as internal standards. Calibration curves were calculated for each peptide in concentrations ranging from 0.635 to 1101 fmol/μL. Limits of detection and quantification were determined for each peptide in blank matrices.

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Shatursky, Oleg, Robert Bayles, Marianne Rogers, B. Helen Jost, J. Glenn Songer et Rodney K. Tweten. « Clostridium perfringens Beta-Toxin Forms Potential-Dependent, Cation-Selective Channels in Lipid Bilayers ». Infection and Immunity 68, n^o 10 (1 octobre 2000) : 5546–51. http://dx.doi.org/10.1128/iai.68.10.5546-5551.2000.

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ABSTRACT Recombinant beta-toxin from Clostridium perfringenstype C was found to increase the conductance of bilayer lipid membranes (BLMs) by inducing channel activity. The channels exhibited a distribution of conductances within the range of 10 to 380 pS, with the majority of the channels falling into two categories of conductance at 110 and 60 pS. The radii of beta-toxin pores found for the conductance states of 110 and 60 pS were 12.7 and 11.1 Å, respectively. The single channels and the steady-state currents induced by beta-toxin across the BLMs exhibited ideal monovalent cation selectivity. Addition of divalent cations (Zn2+, Cd2+, or Mg2+) at a concentration of 2 mM increased the rate of beta-toxin insertion into BLMs and the single-channel conductance, while application of 5 mM Zn2+ to a beta-toxin-induced steady-state current decreased the inward current by approximately 45%. The mutation of arginine 212 of beta-toxin to aspartate, previously shown to increase the 50% lethal dose of beta-toxin for mice nearly 13-fold, significantly reduced the ability of beta-toxin to form channels. These data support the hypothesis that the lethal action of beta-toxin is based on the formation of cation-selective pores in susceptible cells.

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Matsuda, Asami, Meiji Soe Aung, Noriko Urushibara, Mitsuyo Kawaguchiya, Ayako Sumi, Mayumi Nakamura, Yuka Horino, Masahiko Ito, Satoshi Habadera et Nobumichi Kobayashi. « Prevalence and Genetic Diversity of Toxin Genes in Clinical Isolates of Clostridium perfringens : Coexistence of Alpha-Toxin Variant and Binary Enterotoxin Genes (bec/cpile) ». Toxins 11, n^o 6 (6 juin 2019) : 326. http://dx.doi.org/10.3390/toxins11060326.

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Clostridium perfringens (C. perfringens) is responsible for food-borne gastroenteritis and other infectious diseases, and toxins produced by this bacterium play a key role in pathogenesis. Although various toxins have been described for C. perfringens isolates from humans and animals, prevalence of individual toxins among clinical isolates has not yet been well explored. In the present study, a total of 798 C. perfringens clinical isolates were investigated for prevalence of eight toxin genes and their genetic diversity by PCR, nucleotide sequencing, and phylogenetic analysis. Besides the alpha-toxin gene (plc) present in all the isolates, the most common toxin gene was cpe (enterotoxin) (34.2%), followed by cpb2 (beta2 toxin) (1.4%), netB (NetB) (0.3%), and bec/cpile (binary enterotoxin BEC/CPILE) (0.1%), while beta-, epsilon-, and iota-toxin genes were not detected. Genetic analysis of toxin genes indicated a high level of conservation of plc, cpe, and netB. In contrast, cpb2 was revealed to be considerably divergent, containing at least two lineages. Alpha-toxin among 46 isolates was classified into ten sequence types, among which common types were distinct from those reported for avian isolates. A single isolate with bec/cpile harbored a plc variant containing an insertion of 834-bp sequence, suggesting its putative origin from chickens.

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Sayeed, Sameera, Jihong Li et Bruce A. McClane. « Characterization of Virulence Plasmid Diversity among Clostridium perfringens Type B Isolates ». Infection and Immunity 78, n^o 1 (26 octobre 2009) : 495–504. http://dx.doi.org/10.1128/iai.00838-09.

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ABSTRACT The important veterinary pathogen Clostridium perfringens type B is unique for producing the two most lethal C. perfringens toxins, i.e., epsilon-toxin and beta-toxin. Our recent study (K. Miyamoto, J. Li, S. Sayeed, S. Akimoto, and B. A. McClane, J. Bacteriol. 190:7178-7188, 2008) showed that most, if not all, type B isolates carry a 65-kb epsilon-toxin-encoding plasmid. However, this epsilon-toxin plasmid did not possess the cpb gene encoding beta-toxin, suggesting that type B isolates carry at least one additional virulence plasmid. Therefore, the current study used Southern blotting of pulsed-field gels to localize the cpb gene to ∼90-kb plasmids in most type B isolates, although a few isolates carried a ∼65-kb cpb plasmid distinct from their etx plasmid. Overlapping PCR analysis then showed that the gene encoding the recently discovered TpeL toxin is located ∼3 kb downstream of the plasmid-borne cpb gene. As shown earlier for their epsilon-toxin-encoding plasmids, the beta-toxin-encoding plasmids of type B isolates were found to carry a tcp locus, suggesting that they are conjugative. Additionally, IS1151-like sequences were identified upstream of the cpb gene in type B isolates. These IS1151-like sequences may mobilize the cpb gene based upon detection of possible cpb-containing circular transposition intermediates. Most type B isolates also possessed a third virulence plasmid that carries genes encoding urease and lambda-toxin. Collectively, these findings suggest that type B isolates are among the most plasmid dependent of all C. perfringens isolates for virulence, as they usually carry three potential virulence plasmids.

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Nagahama, Masahiro, Masahiro Shibutani, Soshi Seike, Mami Yonezaki, Teruhisa Takagishi, Masataka Oda, Keiko Kobayashi et Jun Sakurai. « The p38 MAPK and JNK Pathways Protect Host Cells against Clostridium perfringens Beta-Toxin ». Infection and Immunity 81, n^o 10 (22 juillet 2013) : 3703–8. http://dx.doi.org/10.1128/iai.00579-13.

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ABSTRACTClostridium perfringensbeta-toxin is an important agent of necrotic enteritis and enterotoxemia. Beta-toxin is a pore-forming toxin (PFT) that causes cytotoxicity. Two mitogen-activated protein kinase (MAPK) pathways (p38 and c-Jun N-terminal kinase [JNK]-like) provide cellular defense against various stresses. To investigate the role of the MAPK pathways in the toxic effect of beta-toxin, we examined cytotoxicity in five cell lines. Beta-toxin induced cytotoxicity in cells in the following order: THP-1 = U937 > HL-60 > BALL-1 = MOLT-4. In THP-1 cells, beta-toxin formed oligomers on lipid rafts in membranes and induced the efflux of K+from THP-1 cells in a dose- and time-dependent manner. The phosphorylation of p38 MAPK and JNK occurred in response to an attack by beta-toxin. p38 MAPK (SB203580) and JNK (SP600125) inhibitors enhanced toxin-induced cell death. Incubation in K+-free medium intensified p38 MAPK activation and cell death induced by the toxin, while incubation in K+-high medium prevented those effects. While streptolysin O (SLO) reportedly activates p38 MAPK via reactive oxygen species (ROS), we showed that this pathway did not play a major role in p38 phosphorylation in beta-toxin-treated cells. Therefore, we propose that beta-toxin induces activation of the MAPK pathway to promote host cell survival.

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Sakurai, Jun, Yoshio Fujii et Masahiro Nagahama. « Effect of p-chloromercuribenzoate on Clostridium perfringens beta toxin ». Toxicon 30, n^o 3 (mars 1992) : 323–30. http://dx.doi.org/10.1016/0041-0101(92)90872-3.

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Dinh, Phat X. « Detecting toxin genes of Clostridium perfringens isolated from diarrhea piglets using multiplex PCR ». Journal of Agriculture and Development 17, n^o 06 (31 décembre 2018) : 24–30. http://dx.doi.org/10.52997/jad.4.06.2018.

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Clostridium perfringens is currently classified into five types (A, B, C, D, E) based on the different toxins produced. Type A and C are known as the causative agent of enteritis and enterotoxaemia in newborn and young piglets with severe intestinal lesions including edema, hemorrhage and necrosis. A multiplex PCR (mPCR) was developed in order to quickly and early determine the presence of genotypes of C. perfringens based on their genes of cpa, cpb, cpb2 and cpe encoding alpha toxin, beta toxin, beta2 toxin and enterotoxin with predicted products of 324 bp, 196 bp, 107 bp and 257 bp respectively. The detection limit of the mPCR assay was 1 × 103 copies/reaction for each gene. Sequencing of mPCR products performed with clinical samples collected from C. perfringens suspected pigs showed that the mPCR test functioned specifically. In conclusion, the developed mPCR test successfully detected the presence of genes cpa, cpb, cpb2 and cpe in the examined samples. Analysis of the bacteria isolated from field samples of diarrheal piglets collected in this study indicated that C. perfringens carrying gene cpa counted for 96.66% and 3.33% was identified as C. perfringens carrying genes cpa and cpb concurrently. Gene cpe was not found in this study, while gene cpb2 was detected coincidently in 73.33% of the samples with cpa gene. The results indicate that the prevalence of these four toxin genes is cpa, cpb2, cpb and cpe in decending order.

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Omer, Sawsan A., Ebtesam M. Al-Olayan, Salah Eldin H. Babiker, Mohammed Z. Aljulaifi, Abdulaziz N. Alagaili et Osama B. Mohammed. « Genotyping of Clostridium perfringens Isolates from Domestic Livestock in Saudi Arabia ». BioMed Research International 2020 (24 mars 2020) : 1–9. http://dx.doi.org/10.1155/2020/9035341.

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The present study was undertaken to confirm the genetic identity of Clostridium perfringens isolates from domestic livestock in Saudi Arabia and to characterize the genes encoding to alpha, beta, epsilon, and iota (α-, β-, ε-, and ι-) toxins. C. perfringens were confirmed in 104 out of 136 isolates on multiplex polymerase chain reaction using specific primers amplifying genes related to toxins produced by C. perfringens. Genes encoding α-toxins were detected in 104 samples. Of the isolates, 80.8% were diagnosed as type A, 15.4% as type D, 2.9% as type C, and 0.96% as type B. None of the isolates has genes encoding iota (ι-) toxin. All isolates investigated yielded enterotoxin (cpe) products and none yielded β2 (cpb2-toxin) or NetB products. PLC gene sequences encoding α-toxin showed >96.7% similarity. Isolates which had α-toxins as well as enterotoxin (cpe) are regarded as type F. Phylogenetic analysis using maximum likelihood analysis yielded two clades, and the majority of the isolates were in one group while only two isolates clustered on the second clade. Within the Kingdom of Saudi Arabia strains, 54 variable positions and 23 polymorphic amino acids were noticed. Isolates with ε- and β-toxins were variable and were found to be close to those published for C. perfringens. ETX gene sequences encoding ε-toxins were found to be related to CPE sequences.

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Mehdizadeh Gohari, Iman, Stefan Unterer, Ashley E. Whitehead et John F. Prescott. « NetF-producing Clostridium perfringens and its associated diseases in dogs and foals ». Journal of Veterinary Diagnostic Investigation 32, n^o 2 (21 février 2020) : 230–38. http://dx.doi.org/10.1177/1040638720904714.

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The role of type A Clostridium perfringens in canine acute hemorrhagic diarrhea syndrome and foal necrotizing enteritis is poorly characterized. However, a highly significant association between the presence of novel toxigenic C. perfringens and these specific enteric diseases has been described. These novel toxigenic strains produce 3 novel putative toxins, which have been designated NetE, NetF, and NetG. Although not conclusively demonstrated, current evidence suggests that NetF is likely the major virulence factor in strains responsible for canine acute hemorrhagic diarrhea syndrome and foal necrotizing enteritis. NetF is a beta–pore-forming toxin that belongs to the same toxin superfamily as CPB and NetB toxins produced by C. perfringens. The netF gene is encoded on a conjugative plasmid that, in the case of netF, also carries another putative toxin gene, netE. In addition, these strains consistently also carry a cpe tcp-conjugative plasmid, and a proportion also carry a separate netG tcp-conjugative plasmid. The netF and netG genes form part of a locus with all the features of the pathogenicity loci of tcp-conjugative plasmids. The netF-positive isolates are clonal in origin and fall into 2 clades. Disease in dogs or foals can be associated with either clade. Thus, these are strains with unique virulence-associated characteristics associated with serious and sometimes fatal cases of important enteric diseases in 2 animal species.

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Silva, Rodrigo Otávio Silveira, Francisco Carlos Ferreira Junior, Marcus Vinícius Romero Marques, Carlos Augusto Oliveira Junior, Nelson Rodrigo da Silva Martins et Francisco Carlos Faria Lobato. « Genotyping and antimicrobial susceptibility of Clostridium perfringens isolated from Tinamidae, Cracidae and Ramphastidae species in Brazil ». Ciência Rural 44, n^o 3 (mars 2014) : 486–91. http://dx.doi.org/10.1590/s0103-84782014000300016.

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The aim of this study was to isolate, genotype and evaluate the antimicrobial susceptibility of Clostridium perfringens found in species Tinamidae, Cracidae and Ramphastidae in Brazil. C. perfringens was isolated in 13 (5%) out of 260 swab samples and five (8.3%) out of 60 stool samples. All strains were classified as C. perfringens type A, and nine (50%) were positive for the beta-2 toxin-encoding gene. No strains were positive for the necrotic enteritis toxin B-like (NetB)-encoding gene. All isolates were susceptible to penicillin, metronidazole and vancomycin, whereas four (22.2%), five (27.8%) and 13 (72.2%) strains were considered resistant to erythromycin, oxytetracycline and lincomycin, respectively.

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Freitas, Nayra F. Q. R., Denis Y. Otaka, Cleideanny C. Galvão, Dayane M. de Almeida, Marcos R. A. Ferreira, Clóvis Moreira Júnior, Marina M. M. H. Hidalgo, Fabricio R. Conceição et Felipe M. Salvarani. « Humoral Immune Response Evaluation in Horses Vaccinated with Recombinant Clostridium perfringens Toxoids Alpha and Beta for 12 Months ». Toxins 13, n^o 8 (13 août 2021) : 566. http://dx.doi.org/10.3390/toxins13080566.

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In horses, Clostridium perfringens is associated with acute and fatal enterocolitis, which is caused by a beta toxin (CPB), and myonecrosis, which is caused by an alpha toxin (CPA). Although the most effective way to prevent these diseases is through vaccination, specific clostridial vaccines for horses against C. perfringens are not widely available. The aim of this study was to pioneer the immunization of horses with three different concentrations (100, 200 and 400 µg) of C. perfringens recombinant alpha (rCPA) and beta (rCPB) proteins, as well as to evaluate the humoral immune response over 360 days. Recombinant toxoids were developed and applied to 50 horses on days 0 and 30. Those vaccines attempted to stimulate the production of alpha antitoxin (anti-CPA) and beta antitoxin (anti-CPB), in addition to becoming innocuous, stable and sterile. There was a reduction in the level of neutralizing anti-CPA and anti-CPB antibodies following the 60th day; therefore, the concentrations of 200 and 400 µg capable of inducing a detectable humoral immune response were not determined until day 180. In practical terms, 200 µg is possibly the ideal concentration for use in the veterinary industry’s production of vaccines against the action of C. perfringens in equine species.

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Gurtner, Corinne, Francesca Popescu, Marianne Wyder, Esther Sutter, Friederike Zeeh, Joachim Frey, Conrad von Schubert et Horst Posthaus. « Rapid Cytopathic Effects of Clostridium perfringens Beta-Toxin on Porcine Endothelial Cells ». Infection and Immunity 78, n^o 7 (19 avril 2010) : 2966–73. http://dx.doi.org/10.1128/iai.01284-09.

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ABSTRACT Clostridium perfringens type C isolates cause fatal, segmental necro-hemorrhagic enteritis in animals and humans. Typically, acute intestinal lesions result from extensive mucosal necrosis and hemorrhage in the proximal jejunum. These lesions are frequently accompanied by microvascular thrombosis in affected intestinal segments. In previous studies we demonstrated that there is endothelial localization of C. perfringens type C β-toxin (CPB) in acute lesions of necrotizing enteritis. This led us to hypothesize that CPB contributes to vascular necrosis by directly damaging endothelial cells. By performing additional immunohistochemical studies using spontaneously diseased piglets, we confirmed that CPB binds to the endothelial lining of vessels showing early signs of thrombosis. To investigate whether CPB can disrupt the endothelium, we exposed primary porcine aortic endothelial cells to C. perfringens type C culture supernatants and recombinant CPB. Both treatments rapidly induced disruption of the actin cytoskeleton, cell border retraction, and cell shrinkage, leading to destruction of the endothelial monolayer in vitro. These effects were followed by cell death. Cytopathic and cytotoxic effects were inhibited by neutralization of CPB. Taken together, our results suggest that CPB-induced disruption of endothelial cells may contribute to the pathogenesis of C. perfringens type C enteritis.

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Finzel, J., V. Florian, H. Schoepe, G. Woitow, H. J. Selbitz et S. Springer. « Vorkommen und Bekämpfung des Clostridium-perfringens-Typ-A-assoziierten Durchfalls der Saugferkel unter besonderer Berücksichtigung der Immunprophylaxe ». Tierärztliche Praxis Ausgabe G : Großtiere / Nutztiere 40, n^o 06 (2012) : 375–82. http://dx.doi.org/10.1055/s-0038-1623143.

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Zusammenfassung Gegenstand und Ziel: Clostridium perfringens Typ A wird häufig in Verbindung mit Durchfällen von Saugferkeln isoliert. Die Bedeutung des Alpha-(α-) und Beta(β)2-Toxins für die Pathogenese der Erkrankung ist nicht abschließend geklärt. Zur Bekämpfung der Erkrankung wurden in der Vergangenheit insbesondere stallspezifische Impfstoffe eingesetzt. Ziel der Untersuchungen war die Typisierung und quantitative Bestimmung des α- und β2-Toxins von C.-perfringens-Stämmen, die im Rahmen der Herstellung bestandsspezifischer Impfstoffe isoliert wurden. Mithilfe eines Intoxikationsmodells wurde weiterhin die Wirksamkeit eines kommerziell erhältlichen Impfstoffs gegen den C.-perfringens-Typ-A-assoziierten Durchfall der Saugferkel geprüft. Material und Methoden: Im ersten Teil der Untersuchungen wurden 1434 C.-perfringens-Stämme, die von an Durchfall erkrankten Ferkeln stammten, mithilfe einer Multiplex-PCR typisiert. Gleichzeitig wurde das α- und β2-Toxin-Bildungsvermögen der Stämme mittels ELISA-Tests untersucht. Zur Prüfung der α- und β2-toxoidhaltigen C.-perfringens-Typ-A-Vakzine für Schweine (Clostriporc A, IDT Biologika GmbH) wurden im zweiten Teil der Untersuchungen 18 Jungsauen im letzten Drittel der Trächtigkeit zweimal im Abstand von 3 Wochen immunisiert. Ergebnisse: 87,9% der Stämme gehörten zur Toxovar A (cpa, cpb2), 6,3% zur Toxovar A (cpa) und 5,8% zur Toxovar C (cpa, cpb, cpb2). Bei der quantitativen Untersuchung des Toxinbildungsvermögens der C.-perfringens-Typ-A-Stämme (cpa, cpb2) zeichneten sich Stämme, die geringe oder mittlere Gehalte des α-Toxins bildeten, häufig durch eine starke Expression des β2-Toxins aus. Die geimpften Sauen bildeten Antikörper gegen das α- und β2-Toxin, die über das Kolostrum auf die Nachzucht übertragen wurden und neugeborene Ferkel signifikant (p < 0,05) vor einer Intoxikation mit einem α- und β2-toxinhaltigen Überstand eines heterologen C.-perfringens-Typ-A-Stammes schützten. Schlussfolgerung und klinische Relevanz: Die Ergebnisse unterstreichen die Bedeutung von α- und β2-toxinbildenden C.-perfringens-Typ-A-Stämmen im Durchfallgeschehen der Saugferkel. Im Intoxikationsmodell ließ sich eine deutliche Schutzwirkung der eingesetzten Vakzine vor dem α- und β2-Toxin nachweisen.

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Hauer, P. J. « Purification and characterization of Clostridium perfringens type C beta toxin ». Veterinary Microbiology 37, n^o 3-4 (novembre 1993) : 410. http://dx.doi.org/10.1016/0378-1135(93)90041-5.

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Tweten, Rodney K. « Clostridium perfringens beta toxin and Clostridium septicum alpha toxin : their mechanisms and possible role in pathogenesis ». Veterinary Microbiology 82, n^o 1 (septembre 2001) : 1–9. http://dx.doi.org/10.1016/s0378-1135(01)00372-8.

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Pires, Prhiscylla Sadanã, Jose Azael Zambrano Uribe, Antônio Último de Carvalho, Rodrigo Otávio Silveira Silva, Felipe Masiero Salvarani, Rodrigo Melo Meneses, Luiza Bossi Leite et al. « Jejunal hemorrhage syndrome in a Zebu cow in Brazil ». Ciência Rural 45, n^o 8 (12 mai 2015) : 1476–79. http://dx.doi.org/10.1590/0103-8478cr20141304.

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Clostridium perfringens type A has been incriminated as the etiologic agent in jejunal hemorrhage syndrome (JHS), which is a disease that affects dairy cattle. Although this microorganism is considered an important enteropathogen the pathogenesis of JHS is still not clear, and there have been no reports of its occurrence in Brazil so far. The aim of this study was to describe the occurrence of JHS by infection with a C. perfringens type A strain carrying the beta-2 toxin gene in a zebu cow in Brazil, for the first time.

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Svobodová, I., I. Steinhauserová et M. Nebola. « Incidence of Clostridium perfringens in Broiler Chickens in the Czech Republic ». Acta Veterinaria Brno 76, n^o 8 (2007) : S25—S30. http://dx.doi.org/10.2754/avb200776s8s025.

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Clostridium perfringens is a causative agent of human and animal foodborne diseases. It is known as a normal inhabitant of the intestinal tract of chickens as well as a potential pathogen causing necrotic enteritis. The aim of the present study was to detect the incidence of C. perfringens in healthy broiler chickens. From May 2005 to September 2006, 609 samples of caecal content from broilers from 23 intensive poultry farms were analyzed. The samples were cultivated on TSC and blood agar, typical colonies were identified and biochemically confirmed. the total number of positive samples was 112 (18.39%). the samples were processed by the multiplex polymerase chain reaction method (PCR) for toxin genotyping. The presence of alpha, beta, beta2 and enterotoxin gene was detected. All C. perfringens isolates were classified as type A, four isolates had the cpb2 gene. In conclusion the prevalence of C. perfringens-positive farms is approximately 74% and the amount ranges about 104 cfu/g of caecal content.

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Carvalho, A. V. A., L. G. D. Heneine, R. A. Assis, V. L. V. Abreu, L. A. Gonçalves et F. C. F. Lobato. « Production and purification of beta-toxin from Clostridium perfringens type C ». Arquivo Brasileiro de Medicina Veterinária e Zootecnia 58, n^o 2 (avril 2006) : 276–78. http://dx.doi.org/10.1590/s0102-09352006000200018.

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Hamza, Dalia, Sohad Dorgham et Ashraf Hakim. « Toxinotyping and antimicrobial resistance of Clostridium perfringens isolated from processed chicken meat products ». Journal of Veterinary Research 61, n^o 1 (1 mars 2017) : 53–58. http://dx.doi.org/10.1515/jvetres-2017-0007.

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Abstract Introduction: The toxinotyping and antimicrobial susceptibility of Clostridium perfringens strains isolated from processed chicken meat were determined. Material and Methods: Two hundred processed chicken meat samples from luncheon meats, nuggets, burgers, and sausages were screened for Clostridium perfringens by multiplex PCR assay for the presence of alpha (cpa), beta (cpb), epsilon (etx), iota (ia), and enterotoxin toxin (cpe) genes. The C. perfringens isolates were examined in vitro against eight antibiotics (streptomycin, amoxicillin, ampicillin, ciprofloxacin, lincomycin, cefotaxime, rifampicin, and trimethoprim-sulfamethoxazole) Results: An overall of 32 C. perfringens strains (16%) were isolated from 200 processed chicken meat samples tested. The prevalence of C. perfringens was significantly dependent on the type of toxin genes detected (P = 0.0), being the highest in sausages (32%), followed by luncheon meats (24%), burgers (6%), and nuggets (2%). C. perfringens type A was the most frequently present toxinotype (24/32; 75%), followed by type D (21.9 %) and type E (3.1%). Of the 32 C. perfringens strains tested, only 9 (28%) were enterotoxin gene carriers, with most representing type A (n = 6). C. perfringens strains differed in their resistance/susceptibility to commonly used antibiotics. Most of the strains tested were sensitive to ampicillin (97%) and amoxicillin (94%), with 100% of the strains being resistant to streptomycin and lincomycin. It is noteworthy that the nine isolates with enterotoxigenic potential had a higher resistance than the non-enterotoxigenic ones. Conclusion: The considerably high C. perfringens isolation rates from processed chicken meat samples and resistance to some of the commonly used antibiotics indicate a potential public health risk. Recent information about the isolation of enterotoxigenic C. perfringens type E from chicken sausage has been reported.

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Saadh, Mohamed J., Issam J. Sa'adeh, Moeen F. Dababneh, Ammar M. Almaaytah et Mohammad F. Bayan. « Production, immunogenicity, stability, and safety of a vaccine against Clostridium perfringens beta toxins ». August-2020 13, n^o 8 (2020) : 1517–23. http://dx.doi.org/10.14202/vetworld.2020.1517-1523.

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Background and Aim: The beta toxin is causing the most severe Clostridium perfringens-related diseases. This work was dedicated to developing a vaccine against beta toxin using C. perfringens type C (NCTC 3180). Materials and Methods: The crude toxoid harvest contained 710 limits of flocculation (Lf)/mL. The vaccine was formulated. Each 1 mL of the final vaccine product contained at least 50 Lf/mL of beta toxoids, 0.2 mL 3% aluminum hydroxide gel (equivalent to 5.18 mg of aluminum), <0.001% W/V thiomersal, formaldehyde <0.05% W/V, and ∼0.7 mL phosphate-buffered saline (pH 7.2). The efficacy of the vaccine was evaluated by potency, stability, and safety tests. Results: The vaccine demonstrated 24.36 IU/mL (standard deviation, ±0.56) and 14.74 IU/mL (±0.36) of neutralizing antibodies in rabbits and cattle, respectively. Indeed, these levels were above the minimum recommended by international protocols since the obtained antibody levels had 2.43- and 1.47-fold increase in both rabbits and cattle, respectively, over the minimum antitoxin level suggested by the United States Department of Agriculture. Interestingly, our formulation was capable of inducing 1.65-fold higher immune responses in rabbits than that stimulated in cattle (65% increase) with a significant difference (p<0.0001). The vaccine was stable up to 30 months. The vaccinated rabbits were suffered from a temporarily slight increase in temperatures in the first 10 h without any significant difference (p>0.05). Conclusion: The research showed a procedure for the manufacturing process of the vaccine against C. perfringens beta toxins with a feasible quantity and the vaccine described here showed to be effective in eliciting levels of neutralizing antibodies higher than required by international standards. In addition, The vaccine was stable up to 30 months. Thus, it may represent an effective and safe for preventing C. perfringens-related diseases in rabbits and cattle, although further studies to prove its efficacy in the field on other farm animals are still needed.

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Rood, J. « Regulation of extracellular toxin production in Clostridium perfringens ». Trends in Microbiology 3, n^o 5 (mai 1995) : 192–96. http://dx.doi.org/10.1016/s0966-842x(00)88919-2.

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Petit, L., et M. R. Popoff. « Clostridium perfringens epsilon toxin cytotoxicity on MDCK cells ». Toxicon 35, n^o 12 (décembre 1997) : 1670–71. http://dx.doi.org/10.1016/s0041-0101(97)90111-2.

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Alsaab, Fehaid, Ali Wahdan et Elhassan M. A. Saeed. « Phenotypic detection and genotyping of Clostridium perfringens associated with enterotoxemia in sheep in the Qassim Region of Saudi Arabia ». March-2021 14, n^o 3 (6 mars 2021) : 578–84. http://dx.doi.org/10.14202/vetworld.2021.578-584.

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Background and Aim: Enterotoxemia caused by Clostridium perfringens toxinotypes is an often fatal disease of sheep of all ages, with a substantial economic loss to the sheep industry. This study was conducted to isolate C. perfringens from suspected cases of enterotoxemia in sheep in the central part of the Qassim Region, Saudi Arabia, and to determine the prevalent toxinotype by detecting alpha (cpA), beta (cpB), and epsilon (etX) toxin genes, which might help control this disease locally. Materials and Methods: A total of 93 rectal swabs and intestinal content samples were collected from diseased and animals suspected of having died of enterotoxemia in early 2020. Samples were subjected to bacteriological examination, biochemical analysis of isolates by VITEK 2, and molecular toxinotyping of isolates by LightCycler® real-time polymerase chain reaction (RT-PCR). Results: Our results revealed that only 14 isolates were confirmed by VITEK 2 as being C. perfringens, with excellent identification (probability of 95% and 97%). According to the toxinotyping of isolates by RT-PCR, all 14 isolates possessed both the cpA and etX toxin genes, while the cpB toxin gene was not detected in any of the isolates. Conclusion: Our findings demonstrated that C. perfringens type D was the only toxinotype found in the central part of the Qassim Region in 2020; moreover, according to the culture method, only 15% (14/93) of the suspected cases of enterotoxemia were confirmed to be caused by C. perfringens infection, which highlighted the importance of clinical and laboratory differential diagnosis of enterotoxemia in sheep.

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HSIEH, H. « Measurement of Clostridium perfringens $beta;-toxin production by surface plasmon resonance immunoassay ». Vaccine 16, n^o 9-10 (mai 1998) : 997–1003. http://dx.doi.org/10.1016/s0264-410x(97)00282-x.

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Miclard, J., M. Jäggi, E. Sutter, M. Wyder, B. Grabscheid et H. Posthaus. « Clostridium perfringens beta-toxin targets endothelial cells in necrotizing enteritis in piglets ». Veterinary Microbiology 137, n^o 3-4 (juin 2009) : 320–25. http://dx.doi.org/10.1016/j.vetmic.2009.01.025.

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Nagahama, Masahiro, Shinsuke Morimitsu, Atsushi Kihara, Masahiko Akita, Koujun Setsu et Jun Sakurai. « Involvement of tachykinin receptors in Clostridium perfringens beta-toxin-induced plasma extravasation ». British Journal of Pharmacology 138, n^o 1 (janvier 2003) : 23–30. http://dx.doi.org/10.1038/sj.bjp.0705022.

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Steinthorsdottir, Valgerdur, Haraldur Halldórsson et OÂlafur S. Andrésson. « Clostridium perfringens beta-toxin forms multimeric transmembrane pores in human endothelial cells ». Microbial Pathogenesis 28, n^o 1 (janvier 2000) : 45–50. http://dx.doi.org/10.1006/mpat.1999.0323.

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Roos, S., M. Wyder, N. Regenscheit, A. Candi, C. Nathues, F. Van Immerseel et H. Posthaus. « Effect OF Clostridium perfringens Beta Toxin on the Porcine Small Intestinal Mucosa ». Journal of Comparative Pathology 152, n^o 1 (janvier 2015) : 56. http://dx.doi.org/10.1016/j.jcpa.2014.10.062.

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Seike, Soshi, Masaya Takehara, Keiko Kobayashi et Masahiro Nagahama. « Role of pannexin 1 in Clostridium perfringens beta-toxin-caused cell death ». Biochimica et Biophysica Acta (BBA) - Biomembranes 1858, n^o 12 (décembre 2016) : 3150–56. http://dx.doi.org/10.1016/j.bbamem.2016.10.003.

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Nagahama, Masahiro, Soshi Seike, Hidenori Shirai, Teruhisa Takagishi, Keiko Kobayashi, Masaya Takehara et Jun Sakurai. « Role of P2X7 receptor in Clostridium perfringens beta-toxin-mediated cellular injury ». Biochimica et Biophysica Acta (BBA) - General Subjects 1850, n^o 11 (novembre 2015) : 2159–67. http://dx.doi.org/10.1016/j.bbagen.2015.08.011.

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Nagahama, Masahiro, Mariko Umezaki, Masataka Oda, Keiko Kobayashi, Shigenobu Tone, Taiji Suda, Kazumi Ishidoh et Jun Sakurai. « Clostridium perfringens Iota-Toxin b Induces Rapid Cell Necrosis ». Infection and Immunity 79, n^o 11 (12 septembre 2011) : 4353–60. http://dx.doi.org/10.1128/iai.05677-11.

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ABSTRACTClostridium perfringensiota-toxin is a binary toxin composed of an enzyme component (Ia) and a binding component (Ib). Each component alone lacks toxic activity, but together they produce cytotoxic effects. We examined the cytotoxicity of iota-toxin Ib in eight cell lines. A431 and A549 cells were susceptible to Ib, but MDCK, Vero, CHO, Caco-2, HT-29, and DLD-1 cells were not. Ib bound and formed oligomers in the membranes of A431 and MDCK cells. However, Ib entered MDCK cells but not A431 cells, suggesting that uptake is essential for cellular survival. Ib also induced cell swelling and the rapid depletion of cellular ATP in A431 and A549 cells but not the insensitive cell lines. In A431 cells, Ib binds and oligomerizes mainly in nonlipid rafts in the membranes. Disruption of lipid rafts by methyl-β-cyclodextrin did not impair ATP depletion or cell death caused by Ib. Ib induced permeabilization by propidium iodide without DNA fragmentation in A431 cells. Ultrastructural studies revealed that A431 cells undergo necrosis after treatment with Ib. Ib caused a disruption of mitochondrial permeability and the release of cytochromec. Staining with active-form-specific antibodies showed that the proapoptotic Bcl-2-family proteins Bax and Bak were activated and colocalized with mitochondria in Ib-treated A431 cells. We demonstrate that Ib by itself produces cytotoxic activity through necrosis.

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Zafar Khan, Muhammad Umar. « Genetic Diversity of Clostridium perfringens Strains Isolated from Broiler Chickens Revealed by PFGE Analysis in China and Pakistan ». Pakistan Veterinary Journal 41, n^o 01 (1 mars 2021) : 85–91. http://dx.doi.org/10.29261/pakvetj/2020.087.

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Clostridium perfringens (C. perfringens) is widely distributed in broiler chickens causing clinical and subclinical enteritis and is especially known for causing necrotic enteritis (NE). There are numerous reports of NE outbreaks in Pakistan as well as China but there is a lack of information related to PFGE profile from both the countries. To close this gap, we designed this study and obtained samples from broiler chicken farms located in 3 different regions of Pakistan and 4 different regions of China. A total of 79 fecal swabs (Pakistan=29; China=50) were collected and grown on FTA media. Further, isolates were grown on TSE agar and black colonies were selected for DNA extraction. All 79 isolates were tested for toxin profiles by PCR (α-gene; beta-2; netB gene) and PFGE profiling (pulsotypes analysis). Toxinotyping results revealed that all the isolates (n=50) from China were type A (α-toxin positive) while 23 and 6 isolates (n=29) from Pakistan were type A (α-toxin positive) and type G (α-toxin, NetB positive), respectively. Toxinotyping revealed α-toxin is highly prevalent in both the countries while from Pakistani isolates, NetB toxin was also detected. PFGE discriminated 79 isolates into 45 different PFGE patterns (pulsotypes). The analysis further showed different pulsotypes originating from China and Pakistan and isolates were subtyped by SmaI. The results showed high genetic polymorphism in C. perfringens even within the same strain. These preliminary findings of genetic variations will further help to design control strategies

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LIU, ZONGLIN L., et HANS P. BLASCHEK. « Monoclonal Antibody-Based ELISA for Detection of Clostridium perfringens Alpha-Toxin ». Journal of Food Protection 59, n^o 6 (1 juin 1996) : 621–25. http://dx.doi.org/10.4315/0362-028x-59.6.621.

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A monoclonal antibody-based ELISA was developed to detect alpha-toxin present in Clostridium perfringens bacterial cell lysates and cell-free culture supernatants, Monoclonal antibodies against C. perfringens alpha-toxin were produced in hybridoma tissue culture supernatants and in BALB/c mice ascites fluid, The monoclonal antibodies obtained from hybridoma culture supernatant and ascites fluid showed identical antigen specificity, but the latter showed a higher titer, with a 50% endpoint at 1/4,000. The monoclonal antibodies were specific for phospholipase C produced by C. perfringens, but not by Bacillus cereus. The lower limit of phospholipase C detection was 16 ng/ml. The dose-dependent relationship between absorption at 490 nm and concentration of phospholipase C diluted in HEPES (N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid) or Trypticase glucose yeast broth fit a four-parameter and a quadratic model, respectively. The monoclonal antibody-based ELISA developed is a rapid, sensitive and specific detection method and can be used for quantitative characterization of C. perfringens alpha-toxin.

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Miyamoto, Kazuaki, Jihong Li, Sameera Sayeed, Shigeru Akimoto et Bruce A. McClane. « Sequencing and Diversity Analyses Reveal Extensive Similarities between Some Epsilon-Toxin-Encoding Plasmids and the pCPF5603 Clostridium perfringens Enterotoxin Plasmid ». Journal of Bacteriology 190, n^o 21 (5 septembre 2008) : 7178–88. http://dx.doi.org/10.1128/jb.00939-08.

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ABSTRACT Clostridium perfringens type B and D isolates produce epsilon-toxin, the third most potent clostridial toxin. The epsilon-toxin gene (etx) is plasmid borne in type D isolates, but etx genetics have been poorly studied in type B isolates. This study reports the first sequencing of any etx plasmid, i.e., pCP8533etx, from type B strain NCTC8533. This etx plasmid is 64.7 kb, carries tcp conjugative transfer genes, and encodes additional potential virulence factors including beta2-toxin, sortase, and collagen adhesin but not beta-toxin. Interestingly, nearly 80% of pCP8533etx open reading frames (ORFs) are also present on pCPF5603, an enterotoxin-encoding plasmid from type A isolate F5603. Pulsed-field gel electrophoresis and overlapping PCR indicated that a pCP8533etx-like etx plasmid is also present in most, if not all, other type B isolates and some beta2-toxin-positive, cpe-negative type D isolates, while other type D isolates carry different etx plasmids. Sequences upstream of the etx gene vary between type B isolates and some type D isolates that do not carry a pCP8533etx-like etx plasmid. However, nearly all type B and D isolates have an etx locus with an upstream IS1151, and those etx loci typically reside near a dcm ORF. These results suggest that pCPF5603 and pCP8533etx evolved from insertion of mobile genetic elements carrying enterotoxin or etx genes, respectively, onto a common progenitor plasmid.

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Theoret, James R., Francisco A. Uzal et Bruce A. McClane. « Identification and Characterization of Clostridium perfringens Beta Toxin Variants with Differing Trypsin Sensitivity andIn VitroCytotoxicity Activity ». Infection and Immunity 83, n^o 4 (2 février 2015) : 1477–86. http://dx.doi.org/10.1128/iai.02864-14.

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By producing toxins,Clostridium perfringenscauses devastating diseases of both humans and animals.C. perfringensbeta toxin (CPB) is the major virulence determinant for type C infections and is also implicated in type B infections, but little is known about the CPB structure-function relationship. Amino acid sequence comparisons of the CPBs made by 8 randomly selected isolates identified two natural variant toxins with four conserved amino acid changes, including a switch of E to K at position 168 (E168K) that introduces a potential trypsin cleavage site into the CPB protein of strain JGS1076. To investigate whether this potential trypsin cleavage site affects sensitivity to trypsin, a primary host defense against this toxin, the two CPB variants were assayed for their trypsin sensitivity. The results demonstrated a significant difference in trypsin sensitivity, which was linked to the E168K switch by using site-directed recombinant CPB (rCPB) mutants. The natural CPB variants also displayed significant differences in their cytotoxicity to human endothelial cells. This cytotoxicity difference was mainly attributable to increased host cell binding rather than the ability to oligomerize or form functional pores. Using rCPB site-directed mutants, differences in cytotoxicity and host cell binding were linked to an A300V amino acid substitution in the strain JGS1076 CPB variant that possessed more cytotoxic activity. Mapping of sequence variations on a CPB structure modeled using related toxins suggests that the E168K substitution is surface localized and so can interact with trypsin and that the A300V substitution is located in a putative binding domain of the CPB toxin.

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Profeta, Francesca, Cristina E. Di Francesco, Andrea Di Provvido, Massimo Scacchia, Alessandra Alessiani, Elisabetta Di Giannatale, Giuseppe Marruchella, Massimiliano Orsini, Tonino Toscani et Fulvio Marsilio. « Prevalence of netB-positive Clostridium perfringens in Italian poultry flocks by environmental sampling ». Journal of Veterinary Diagnostic Investigation 32, n^o 2 (25 octobre 2019) : 252–58. http://dx.doi.org/10.1177/1040638719885841.

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Clostridium perfringens type G is one of the pathogens involved in enteric diseases in poultry. NetB, a pore-forming toxin, is considered the main virulence factor responsible for necrotic enteritis during C. perfringens infection. We carried out a field study involving 14 farms to evaluate the occurrence of netB-positive C. perfringens and the impact of infection in Italian poultry flocks. Environmental samples ( n = 117) and 50 carcasses were screened by microbiologic and molecular methods. Microbiologic investigations yielded 82 C. perfringens isolates. DNA was extracted from all samples and screened for α-toxin and NetB encoding genes by real-time PCR. The C. perfringens α-toxin gene was detected in 151 of 167 extracts (90.4%), and 31 of 151 (20.5%) were netB gene positive also. Sixteen isolates from a turkey flock with mild enteric disorders were also netB positive, demonstrating their occurrence not only in broiler but also in turkey flocks. A pulsed-field gel electrophoresis protocol was optimized to evaluate the diversity among isolates and revealed high genetic heterogeneity. The complete NetB toxin-coding gene of 2 C. perfringens isolates from turkey and broiler flocks were analyzed and showed very high relatedness with analogous sequences worldwide.

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VAHJEN, WILFRIED, K. GOLLNISCH, O. SIMON et E. SCHULZ. « Development of a semiquantitative PCR assay for the detection of the Clostridium perfringens type C beta toxin gene in purified nucleic acid extracts from the intestinal tract of pigs ». Journal of Agricultural Science 134, n^o 1 (janvier 2000) : 77–87. http://dx.doi.org/10.1017/s002185969900725x.

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Total DNA was extracted and purified from luminal digesta samples (1 g) of 14-week-old (c. 43 kg bodyweight) pigs. Digoxigenin (DIG) labelled PCR products of the beta toxin gene of Clostridium perfringens type C were generated with DIG-11-UTP and chemoluminescent signals of DIG-labelled PCR products were detected on slot blots and recorded with a sensitive charge coupled densitometric camera. Detection limit and correlation of chemoluminescent signals to initial target DNA concentration were evaluated. Detection of the beta toxin gene could be achieved with 100 ag C. perfringens DNA in the presence of 1 μg unspecific DNA. The semiquantitative DIG-PCR assay was used to evaluate the relative abundance of the beta toxin gene of C. perfringens type C in purified DNA extracts from intestinal samples of pigs, which were fed a wheat/barley diet (controls), supplemented with the antibiotic avilamycin (40 mg/kg feed), a xylanase (4000 U/kg feed) or a combination of both feed additives. The antibiotic treatment significantly reduced the amount of PCR product in ileal and colon DNA extracts. Xylanase supplementation significantly reduced the amount of PCR product in colon samples, but led to an increase in jejunal samples. From the results it is concluded, that the antibiotic inhibited growth of C. perfringens type C in the ileum and colon of pigs. The semiquantitative DIG-PCR assay can be used to sensitively monitor the relative abundance of specific pathogens in the intestinal tract.

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Si, Weiduo, Joshua Gong, Yanming Han, Hai Yu, John Brennan, Huaijun Zhou et Shu Chen. « Quantification of Cell Proliferation and Alpha-Toxin Gene Expression of Clostridium perfringens in the Development of Necrotic Enteritis in Broiler Chickens ». Applied and Environmental Microbiology 73, n^o 21 (7 septembre 2007) : 7110–13. http://dx.doi.org/10.1128/aem.01108-07.

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ABSTRACT Cell proliferation and alpha-toxin gene expression of Clostridium perfringens in relation to the development of necrotic enteritis (NE) were investigated. Unlike bacitracin-treated chickens, non-bacitracin-treated birds exhibited typical NE symptoms and reduced growth performance. They also demonstrated increased C. perfringens proliferation and alpha-toxin gene expression that were positively correlated and progressed according to the regression model y = b 0 + b 1 X − b 2 X 2. The average C. perfringens count of 5 log10 CFU/g in the ileal digesta appears to be a threshold for developing NE with a lesion score of 2.

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