Carlos Augusto Oliveira Junior

Carriage of Clostridium difficile in free-living South American coati (Nasua nasua) in Brazil

The objective of this study was to isolate and characterize Clostridium difficile strains in stool samples from a wild urban mammal, a South American coati (Nasua nasua) in Brazil. Forty-six free-living N. nasua were trapped, and stool samples were collected. C. difficile was isolated from three (6.5%) sampled animals, two strains were toxigenic (A+B+CDT-, PCR ribotype 014/020 and 106) and one was non toxigenic (A-B-CDT-, PCR ribotype 053). The present work confirms that ring-tailed coati (N. nasua) could harbor C. difficile strains, including those PCR ribotypes commonly reported in C. difficile infection in humans.

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.

ISOLATION AND GENOTYPING OF CLOSTRIDIUM PERFRINGENS FROM FREE-LIVING SOUTH AMERICAN COATI (NASUA NASUA).

Abstract:   The importance of Clostridium perfringens for most wild animal species remains unclear. This study aimed to isolate and genotype C. perfringens in stool samples from free-living South American coati (Nasua nasua) in Brazil. Forty-six free-living N. nasua were trapped and stool samples were collected. Two different protocols for C. perfringens isolation were tested: direct plating onto selective agar and pre-enrichment in broth followed by plating in selective agar. Clostridium perfringens type A was isolated from 15 (32.6%) animals by direct plating and 36 (78.3%) animals by broth PE, and the rate of isolation was significantly different between these two methods (P < 0.01). Twelve of the 36 (33.3%) isolated strains by the PE protocol were positive for the β-2 toxin-encoding gene (cpb2) whereas the enterotoxin-encoding gene (cpe) and necrotic enteritis like-B toxin gene (netb) were not found. These results suggest that C. perfringens is commonly part of the microbiota of free-living coatis. Additionally, the use of a PE protocol appears to be essential for studies on C. perfringens in this species.

Genotyping and antimicrobial susceptibility of Clostridium perfringens isolated from Tinamidae, Cracidae and Ramphastidae species in Brazil

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.

Antimicrobial susceptibility of Clostridium difficile isolated from animals and humans in Brazil

The objective of this study was to evaluate antimicrobial susceptibility in Clostridium difficile strains isolated from animals and humans in Brazil. The 54 C. difficile strains used were isolated from stool samples from piglets (n=16), dogs (n=13), humans (n=13), foals (n=8) calves (n=2), an ocelot (n=1) and a maned wolf (n=1). Antimicrobial susceptibility was determined using the serial plate agar dilution method for penicillin, florfenicol, oxytetracycline, erythromycin, vancomycin, metronidazole and tylosin. The C. difficile strains assessed were susceptible to metronidazole and vancomycin. Florfenicol resistance was rarely observed; 52 (96.4%) strains were sensitive to this antimicrobial. Five (9.3%), five (9.3%), 14 (25.9%) and 20 (37.0%) strains were resistant to oxytetracycline, penicillin, tylosin and erythromycin respectively.

Detection of virulence genes and the phylogenetic groups of Escherichia coli isolated from dogs in Brazil

This study identified the virulence genes, pathovars, and phylogenetic groups of Escherichia coli strains obtained from the feces of dogs with and without diarrhea. Virulence genes and phylogenetic group identification were studied using polymerase chain reaction. Thirty-seven E. coli isolates were positive for at least one virulence factor gene. Twenty-one (57.8%) of the positive isolates were isolated from diarrheal feces and sixteen (43.2%) were from the feces of non-diarrheic dogs. Enteropathogenic E. coli (EPEC) were the most frequently (62.2%) detected pathovar in dog feces and were mainly from phylogroup B1 and E. Necrotoxigenic E. coli were detected in 16.2% of the virulence-positive isolates and these contained the cytotoxic necrotizing factor 1 (cnf1) gene and were classified into phylogroups B2 and D. All E. coli strains were negative for the presence of enterotoxigenic E. coli (ETEC) enterotoxin genes, but four strains were positive for ETECrelated fimbriae 987P and F18. Two isolates were Shiga toxin-producing E. coli strains and contained the toxin genesStx2 or Stx2e, both from phylogroup B1. Our data showed that EPEC was the most frequent pathovar and B1 and E were the most common phylogroups detected in E. coli isolated from the feces of diarrheic and non-diarrheic dogs.

Type C botulism in domestic chickens, dogs and black-pencilled marmoset ( Callithrix penicillata ) in Minas Gerais, Brazil

Botulism is a well-known intoxication that affects humans and animals. The disease is endemic in cattle in Brazil and recently emerged as an important disease in commercial laying hens and broiler chickens in Europe. Dogs and other animal species can also be affected. Although antitoxins are commonly administered to humans diagnosed with botulism, in animals this is rarely the case and the treatment of botulism is still based only on support therapy. In the present work, we report an outbreak of type C botulism in Brazil that simultaneously affected domestic chickens, dogs and a black-pencilled marmoset (Callithrix penicillata). The successful use of Clostridium botulinum types C and D antitoxin for the treatment of an affected dog is also described.