Production and purification of Clostridium perfringens type D epsilon toxin and IgY antitoxin.

The aim of this study was to purify Clostridium perfringens type D epsilon toxin and produce and purify anti-epsilon chicken immunoglobulin Y (IgY). A single-step ion exchange chromatography resulted in a high-yield and high-purity toxin, while ion exchange chromatography followed by gel filtration resulted in the highest purity of the toxin, but at a lower yield. Purified and inactivated epsilon toxin were then administered in chickens via four inoculations and IgY was obtained at a high purity and yield, with an antibody titer of 50 IU/mL and high levels of avidity (73.2%). In summary, C. perfringens type D epsilon toxin and chicken anti-epsilon IgY were successfully produced and purified, and may be used for the diagnosis of enterotoxemia caused by the epsilon toxin, as well as in potency tests of existing and future vaccines against enterotoxemia.

Intracellular survival of Clostridium chauvoei in bovine macrophages.

Clostridium chauvoei is the etiological agent of blackleg, a severe disease of domestic ruminants, causing myonecrosis and serious toxemia with high mortality. Despite the known importance of this agent, studies evaluating its pathogenesis of blackleg are scarce, and many are based on an unproven hypothesis that states that macrophages are responsible for carrying C. chauvoei spores from the intestines to muscles in the early stages of blackleg. Therefore, the present study aimed to investigate the survival of C. chauvoei vegetative cells or spores after phagocytosis by a murine macrophage cell line (RAW 264.7) and bovine monocyte-derived macrophages and to profile inflammatory and anti-inflammatory cytokine transcripts of bovine macrophages infected with C. chauvoei vegetative cells or spores. Both vegetative cells and spores of C. chauvoei remain viable after internalization by murine and bovine macrophages. Bovine macrophages infected with vegetative cells showed a pro-inflammatory profile, while those infected with spores displayed an anti-inflammatory profile. Together, these results corroborate the classical hypothesis that macrophages may play a role in the early pathogenesis of blackleg. Moreover, this is the first study to evaluate the infection kinetics and cytokine profile of bovine monocyte-derived macrophages infected with a Clostridium species.

Botulism in ruminants in Brazil / Botulismo em ruminantes no Brasil

ABSTRACT: Botulism is an intoxication caused by the ingestion of neurotoxins secreted by Clostridium botulinum. It affects all warm-blooded animals and some fishes and is characterized by progressive flaccid symmetrical paralysis. Despite the known importance of this disease in ruminants, the Brazilian literature only includes case studies and outbreak descriptions. In addition, the epidemiology of botulism in cattle has markedly changed over the past two decades. The aim of this study is to review these characteristics and to provide recent data on the occurrence of botulism in ruminants in Brazil.

RESUMO: O botulismo é uma intoxicação causada pela ingestão das neurotoxinas secretadas por Clostridium botulinum . A doença afeta todos os animais de sangue quente e alguns tipos de peixes, sendo caracterizada por uma paralisia flácida simétrica progressiva. Apesar da reconhecida importância dessa doença nos ruminantes, a literatura brasileira baseia-se apenas em relatos de casos e descrições de surtos. Além disso, a epidemiologia do botulismo em bovinos apresentou marcantes alterações nas últimas duas décadas. Dessa forma, é essencial uma revisão que elucide as principais características da doença em animais no Brasil. O presente trabalho tem como objetivo revisar tais características, além de fornecer dados recentes sobre a sua ocorrência em ruminantes.

Immunogenicity of a Trivalent Recombinant Vaccine Against Clostridium perfringens Alpha, Beta, and Epsilon Toxins in Farm Ruminants.

Clostridium perfringens is an anaerobic bacterium that produces several toxins. Of these, the alpha, beta, and epsilon toxins are responsible for causing the most severe C. perfringens-related diseases in farm animals. The best way to control these diseases is through vaccination. However, commercially available vaccines are based on inactivated toxins and have many production drawbacks, which can be overcome through the use of recombinant antigens. In this study, we produced recombinant alpha, beta, and epsilon toxins in Escherichia coli to formulate a trivalent vaccine. Its effectiveness was evaluated through a potency test in rabbits, in which the vaccine generated 9.6, 24.4, and 25.0 IU/mL of neutralizing antibodies against the respective toxins. Following this, cattle, sheep, and goats received the same formulation, generating, respectively, 5.19 ± 0.48, 4.34 ± 0.43, and 4.70 ± 0.58 IU/mL against alpha toxin, 13.71 ± 1.17 IU/mL (for all three species) against beta toxin, and 12.74 ± 1.70, 7.66 ± 1.69, and 8.91 ± 2.14 IU/mL against epsilon toxin. These levels were above the minimum recommended by international protocols. As such, our vaccine was effective in generating protective antibodies and, thus, may represent an interesting alternative for the prevention of C. perfringens-related intoxications in farm animals.

Jejunal hemorrhage syndrome in a Zebu cow in Brazil / Síndrome de jejuno hemorrágico em gado Zebu no Brasil

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.

Clostridium perfringens tipo A tem sido considerado agente etiológico da síndrome do jejuno hemorrágico (SJH), que é uma doença que afeta comumente os rebanhos de gado. Embora este microrganismo seja considerado um importante enteropatógeno, a patogênese da SJH ainda não foi elucidada, e não havia sido reportada no Brasil até então. O alvo deste estudo foi descrever pela primeira vez a ocorrência da SJH causada por C. perfringens tipo A, carreador do gene da toxina beta-2, em um zebuíno no Brasil.

Concentração inibitória mínima (CIM) de oito antimicrobianos frente isolados de Streptococcus suis / Mininum inhibitory concentration of eigth antibiotics toward isolates of Streptococcus suis

Avaliou-se a concentração inibitória mínima (CIM) de 75 isolados de Streptococcus suis frente a oito antimicrobianos freqüentemente utilizados no controle da infecção por esse microrganismo. A CIM foi realizada em placas previamente preparadas com ágar sangue, contendo concentrações variando de 0,25 a 256 μg/ml dos seguintes antimicrobianos: amoxicilina, ampicilina, penicilina, ceftiofur sódico, florfenicol, lincomicina, penicilina, sulfametoxazol-trimetoprim e tetraciclina. A amoxicilina nas concentrações de 1 e 2 μg/ml, o florfenicol a 1 μg/ml e o sulfametoxazol-trimetoprim nas concentrações de 2 e 8 μg/ml, foram os antimicrobianos frente aos quais os microrganismos apresentaram menor resistência. Em contraste, a ampicilina, tetraciclina, ceftiofur sódico e lincomicina, foram menos efetivos, apresentando CIM de (64 e 128 μg/ml), (64 e 128 μg/ml), (128 e 256 μg/ml) e (>256 μg/ml), respectivamente. Os resultados deste estudo demonstram que a amoxicilina e o florfenicol são os antibióticos de escolha para o tratamento de infecções pelo S. suis em suínos.

The minimun inhibitory concentration (MIC) was determined toward amoxicilin, ampicilin, penicillin, ceftiofur, florfenicol, lincomycin, trimethoprim-sulfadiazine and tetracycline for 75 strains of Streptococcus suis. The MIC was performed on sheep blood agar plates containing concentrations varying from 0,25 to 256 μg/ml of the antibiotic described above. The amoxicilin in the concentrations of 1 and 2 μg/ml, florfenicol in the concentration of 1 μg/ml and trimethoprim-sulfadiazine in the concentrations of 2 e 8 μg/ml, were the antibiotics that presented minor resistance. In contrast, the ampicilin, tetracycline, ceftiofur and lincomycin, presented MIC of 64 and 128 μg/ml, 64 and 128 μg/ml, 128 and 256 μg/ml and >256 μg/ml, respectively. The results of this study show that the amoxicilin and florfenicol are the antibiotics of choice for the treatment of diseases by S. suis in swine.

Primeiro relato no Brasil de mastite necrótica bovina por Clostridium perfringens tipo A / First report in Brazil of bovine necrotic mastitis due to Clostridium perfringens type A

Relata-se o primeiro caso no Brasil de mastite bovina por Clostridium perfringens tipo A. O quadro clínico caracterizou-se por necrose da papila mamária e porção ventral do quarto afetado. O agente foi isolado em cultura pura e identificado como tipo A por PCR a partir do leite do quarto mamário afetado.

Edema maligno em suíno / Malignant edema in swine

Edema maligno em suíno é uma infecção fatal da musculatura esquelética e tecido subcutâneo de ruminantes e outras espécies animais, causada principalmente por Clostridium septicum, embora possa ocorrer em associação com Clostridium chauvoei, Clostridium sordellii, Clostridium novyi tipo A e Clostridium perfringens tipo A. O diagnóstico efetivo do edema maligno deve levar em consideração, além dos dados clínicos e lesões de necropsia, os achados microscópicos, isolamento do agente, imunofluorescência direta, imunoistoquímica e PCR. Embora haja poucos relatos de edema maligno em suínos na literatura, o C. septicum tem sido associado à maioria dos casos. Este trabalho relata um caso de mionecrose e edema subcutâneo em um leitão causado por C. septicum.