ABSTRACT
The objective of this study was to detect C. difficileA/B toxins and to isolate strains of C. perfringensand C. difficile from diarrheic and non-diarrheic dogs in Brazil. Stool samples were collected from 57 dogs, 35 of which were apparently healthy, and 22 of which were diarrheic. C. difficileA/B toxins were detected by ELISA, and C. perfringensand C. difficilewere identified by multiplex PCR. C. difficileA/B toxins were detected in 21 samples (36.8%). Of these, 16 (76.2%) were from diarrheic dogs, and five (23.8%) were from non-diarrheic dogs. Twelve C. difficile strains (21.1%) were isolated, of which ten were A+B+and two were A-B-. All non-toxigenic strains were isolated from non-diarrheic animals. The binary toxin gene cdtBwas found in one strain, which was A+B+and was derived from a non-diarrheic dog. C. perfringensstrains were isolated from 40 samples (70.2%). Of these, 18 (45%) were from the diarrheic group, and 22 (55%) belonged to the non-diarrheic group. All isolates were classified as C. perfringenstype A and there was an association between the detection of the cpegene and the presence of diarrhea. Interestingly, ten strains (25%) were positive for the presence of the cpb2gene. The high rate of detection of the A/B toxins in non-diarrheic dogs suggests the occurrence of subclinical disease in dogs or carriage of its toxins without disease. More studies are needed to elucidate the epidemiology of C. difficileand C. perfringensin dogs and to better our understanding of C. difficileas a zoonotic agent. This is the first study to report the binary toxin gene in C. difficilestrains isolated from dogs in Brazil.
Subject(s)
Animals , Dogs , Clinical Laboratory Techniques , Clostridium perfringens , Clostridioides difficile/genetics , Clostridioides difficile/isolation & purification , Diarrhea/genetics , Enterocolitis, Pseudomembranous , Fecal Impaction/genetics , Polymerase Chain Reaction , Bacterial Toxins/genetics , Bacterial Toxins/isolation & purification , Diagnosis , Immunoassay , Methods , Spores, Bacterial , VirulenceABSTRACT
Clostridium difficile has emerged as a major cause of neonatal colitis in piglets, displacing classic bacterial pathogens. However, there is no information regarding the distribution of this microorganism in pig farms in Brazil. In the present study, the presence of toxins A/B and of C. difficile strains in stool samples from 60 diarrheic or non-diarrheic newborn piglets (one to seven days old), from 15 different farms, was studied. The presence of toxins A/B was detected by ELISA and PCR was used to identify toxin A, toxin B and binary toxin gene in each isolated strain. C. difficile A/B toxins were detected in ten samples (16.7 percent). Of these, seven were from diarrheic and three were from non-diarrheic piglets. C. difficile was recovered from 12 out of 60 (20 percent) fecal samples. Of those, three strains were non-toxigenic (A-B-) and nine were toxigenic. Of the nine toxigenic strains, four were A+B+ strains and five were A-B+ strains. The presence of binary toxin observed in the present study was much higher (50 percent) than in previously reported studies. All three non-toxigenic strains were isolated from otherwise healthy piglets. The results suggest the occurrence of neonatal diarrhea by C. difficile in farms in Brazil.
Clostridium difficile tem sido relatado como o principal causador de colite neonatal em suínos. Apesar da crescente importância deste agente, não há dados sobre infecções causadas por C. difficile em suínos no Brasil. O objetivo do presente estudo foi detectar as toxinas A/B e isolar C. difficile a partir de 60 amostras de fezes de leitões diarreicos ou apararentemente saudáveis, com no máximo sete dias de vida, e oriundos de 15 granjas diferentes. As toxinas A/B foram detectadas por ELISA e uma PCR multiplex foi utilizada para detecção dos genes responsáveis pela codificação das toxinas A, B e toxina binária. As toxinas A/B de C. difficile foram detectadas em dez amostras de fezes (16.7 por cento). Dessas, sete eram de animais diarreicos e três de leitões aparentemente saudáveis. Foi possível isolar C. difficile em 12 das 60 (20 por cento) amostras trabalhadas. Dessas, três estirpes eram não-toxigênicas (A-B-) e nove eram toxigênicas, sendo quatro caracterizadas como A+B+ e cinco como A-B+. O gene responsável pela codificação da toxina binária foi encontrado em 50 por cento das estirpes isoladas, proporção superior ao relatado em estudos anteriores. Todas as estirpes não toxigênicas foram isoladas de animais não diarreicos. Os resultados encontrados sugerem a ocorrência de diarreia por C. difficile em granjas no Brasil.