Phenotypic and molecular characterization of erythromycin resistance in Campylobacter jejuni and Campylobacter coli strains isolated from swine and broiler chickens / Caracterização fenotípica e molecular da resistência à eritromicina em cepas de Campylobacter jejuni e Campylobacter coli isoladas de suínos e frangos de corte

Campylobacter spp. is a bacterial agent that causes gastroenteritis in humans and may trigger Guillain-Barré Syndrome (GBS) and is also considered one of the main foodborne diseases in developed countries. Poultry and pigs are considered reservoirs of these microorganisms, as well as raw or undercooked by-products are often incriminated as a source of human infection. Treatment in human cases is with macrolide, such erythromycin, that inhibits the protein synthesis of the microorganism. This study aimed to isolate Campylobacter jejuni and Campylobacter coli from intestinal content samples of broiler chickens (n=20) and swine (n=30) to characterize the erythromycin resistance profile of the strains and to detect molecular mechanisms involved in this resistance. The minimum inhibitory concentration was determined by agar dilution. The Mismatch Amplification Mutation Assay-Polymerase Chain Reaction (MAMA-PCR) was performed to detect mutations at positions 2074 and 2075 of 23S rRNA region, in addition to PCR test to detect the erm(B) gene. From the intestinal content of broiler chickens, 18 strains of C. jejuni and two strains of C. coli were isolated, whereas, from swine samples, no C. jejuni strain and 14 strains of C. coli were isolated. All C. coli strains were resistant, and three C. jejuni strains from broilers chickens were characterized with intermediate resistance to erythromycin. The MIC of the strains ranged from ≤0.5mg/μL to ≥128mg/μL. All resistant strains had the A2075G mutation, and one strain with intermediate resistance had the A2075G mutation. However, the A2074C mutation and the erm(B) gene were not detected. High resistance levels were detected in C. coli strains isolated from swine. The MAMA-PCR is a practical tool for detecting the erythromycin resistance in Campylobacter strains.(AU)

Campylobacter spp. é um agente bacteriano causador de gastroenterite em humanos e associado à síndrome de Guillain-Barré, sendo a campilobacteriose considerada uma das principais enfermidades de origem alimentar. Aves e suínos são importantes reservatórios desses microrganismos e seus produtos derivados crus ou mal cozidos são muitas vezes incriminados como fonte de infecção humana. A primeira escolha para o tratamento em casos humanos são os antimicrobianos da classe dos macrolídeos como à eritromicina. Dentro desse contexto, o objetivo deste estudo foi isolar Campylobacter jejuni e C. coli a partir de 20 amostras de conteúdo intestinal de frangos de corte e de 30 de suínos ao abate e investigar a resistência à eritromicina das estirpes obtidas e os possíveis mecanismos moleculares envolvidos nesta resistência. A concentração inibitória mínima foi determinada pela diluição em ágar e a técnica MAMA-PCR foi utilizada para detecção de mutações nas posições 2074 e 2075 da região 23s rRNA, foi pesquisado também a presença do gene erm(B) pela PCR. A partir do conteúdo intestinal de frangos de corte foram isoladas 18 estirpes de C. jejuni e duas de C. coli, enquanto de suínos foram obtidas 14 estirpes de C. coli e nenhuma estirpe de C. jejuni. Todas as estirpes de C. coli de suínos foram identificadas como resistentes e três estirpes de C. jejuni de frangos foram caracterizadas com resistência intermediária. A CIM das estirpes variou de ≤0,5mg/μL a ≥128mg/μL. Todas as estirpes resistentes tinham a mutação A2075G e uma cepa com resistência intermediária também apresentou a mutação A2075G. Não foi detectada a mutação A2074C ou a presença do gene erm(B) em nenhuma das estirpes obtidas. Os resultados revelam um alto nível de resistência em estirpes de C. coli isoladas de suínos frente a eritromicina. A técnica MAMA PCR utilizada se constitui em uma ferramenta prática para detecção da resistência à eritromicina em estirpes de C. jejuni e C. coli.(AU)

Phenotypic and molecular characterization of erythromycin resistance in Campylobacter jejuni and Campylobacter coli strains isolated from swine and broiler chickens / Caracterização fenotípica e molecular da resistência à eritromicina em cepas de Campylobacter jejuni e Campylobacter coli isoladas de suínos e frangos de corte

Campylobacter spp. is a bacterial agent that causes gastroenteritis in humans and may trigger Guillain-Barré Syndrome (GBS) and is also considered one of the main foodborne diseases in developed countries. Poultry and pigs are considered reservoirs of these microorganisms, as well as raw or undercooked by-products are often incriminated as a source of human infection. Treatment in human cases is with macrolide, such erythromycin, that inhibits the protein synthesis of the microorganism. This study aimed to isolate Campylobacter jejuni and Campylobacter coli from intestinal content samples of broiler chickens (n=20) and swine (n=30) to characterize the erythromycin resistance profile of the strains and to detect molecular mechanisms involved in this resistance. The minimum inhibitory concentration was determined by agar dilution. The Mismatch Amplification Mutation Assay-Polymerase Chain Reaction (MAMA-PCR) was performed to detect mutations at positions 2074 and 2075 of 23S rRNA region, in addition to PCR test to detect the erm(B) gene. From the intestinal content of broiler chickens, 18 strains of C. jejuni and two strains of C. coli were isolated, whereas, from swine samples, no C. jejuni strain and 14 strains of C. coli were isolated. All C. coli strains were resistant, and three C. jejuni strains from broilers chickens were characterized with intermediate resistance to erythromycin. The MIC of the strains ranged from ≤0.5mg/μL to ≥128mg/μL. All resistant strains had the A2075G mutation, and one strain with intermediate resistance had the A2075G mutation. However, the A2074C mutation and the erm(B) gene were not detected. High resistance levels were detected in C. coli strains isolated from swine. The MAMA-PCR is a practical tool for detecting the erythromycin resistance in Campylobacter strains.(AU)

Campylobacter spp. é um agente bacteriano causador de gastroenterite em humanos e associado à síndrome de Guillain-Barré, sendo a campilobacteriose considerada uma das principais enfermidades de origem alimentar. Aves e suínos são importantes reservatórios desses microrganismos e seus produtos derivados crus ou mal cozidos são muitas vezes incriminados como fonte de infecção humana. A primeira escolha para o tratamento em casos humanos são os antimicrobianos da classe dos macrolídeos como à eritromicina. Dentro desse contexto, o objetivo deste estudo foi isolar Campylobacter jejuni e C. coli a partir de 20 amostras de conteúdo intestinal de frangos de corte e de 30 de suínos ao abate e investigar a resistência à eritromicina das estirpes obtidas e os possíveis mecanismos moleculares envolvidos nesta resistência. A concentração inibitória mínima foi determinada pela diluição em ágar e a técnica MAMA-PCR foi utilizada para detecção de mutações nas posições 2074 e 2075 da região 23s rRNA, foi pesquisado também a presença do gene erm(B) pela PCR. A partir do conteúdo intestinal de frangos de corte foram isoladas 18 estirpes de C. jejuni e duas de C. coli, enquanto de suínos foram obtidas 14 estirpes de C. coli e nenhuma estirpe de C. jejuni. Todas as estirpes de C. coli de suínos foram identificadas como resistentes e três estirpes de C. jejuni de frangos foram caracterizadas com resistência intermediária. A CIM das estirpes variou de ≤0,5mg/μL a ≥128mg/μL. Todas as estirpes resistentes tinham a mutação A2075G e uma cepa com resistência intermediária também apresentou a mutação A2075G. Não foi detectada a mutação A2074C ou a presença do gene erm(B) em nenhuma das estirpes obtidas. Os resultados revelam um alto nível de resistência em estirpes de C. coli isoladas de suínos frente a eritromicina. A técnica MAMA PCR utilizada se constitui em uma ferramenta prática para detecção da resistência à eritromicina em estirpes de C. jejuni e C. coli.(AU)

Isolation and antimicrobial resistance of Campylobacter jejuni and Campylobacter coli found in chilled chicken carcasses in the Federal District Region and surrounding areas / Isolamento e resistência antimicrobiana de Campylobacter jejuni e Campylobacter coli encontradas em carcaças de frango resfriadas na região do Distrito Federal e entorno

This study aimed to isolate Campylobacter jejuni and Campylobacter coli from chilled chicken carcasses marketed in the Federal District Region and surrounding areas, as well as to detect the occurrence of antimicrobial resistance and genes responsible for the same. A total of 105 chilled chicken carcasses were collected, of which 7 (6.67%) were positive for C. jejuni and 4 (3.81%) were positive for C. coli. These results were obtained using both the conventional microbiological isolation method and polymerase chain reaction assays. All of the positive strains were subjected to antimicrobial susceptibility testing for seven antimicrobials. The resistance incidences found in the C. jejuni strains were as follows: 71.43% for tetracycline and nalidixic acid, 42.86% for streptomycin and gentamicin, 57.14% for ciprofloxacin and erythromycin, and 28.57% for chloramphenicol. Among the C. coli strains, 100% were resistant to tetracycline and streptomycin, 75% were resistant to erythromycin, 50% were resistant to ciprofloxacin, gentamicin, and nalidixic acid, and no strains were resistant to chloramphenicol. While analyzing the presence of antimicrobial resistance genes in the isolated C. jejuni strains, the aph3-1 (resistance to aminoglycosides), aadE (resistance to streptomycin), and tet(O) (resistance to tetracycline) genes were identified, with occurrence rates of 57.14%, 28.57%, and 42.86%, respectively, whereas in the C. coli strains, there was a 25% occurrence rate for both the aph3-1 and tet(O) genes. The aadE gene was not found in the C. coli isolates. The results of this study demonstrated the presence of C. jejuni and C. coli in chilled chicken carcasses marketed in the Federal District Region and surrounding areas, as well as the antimicrobial resistance and the presence of resistance genes in these bacteria, which may pose threats to public health.

O objetivo deste trabalho foi isolar Campylobacter jejuni e Campylobacter coli de carcaças de frango resfriadas comercializadas no Distrito Federal e entorno, bem como detectar a ocorrência de resistência antimicrobiana e genes responsáveis pela resistência antimicrobiana. Foram coletadas um total de 105 carcaças de frango resfriadas, das quais 7 (6,67%) foram positivas para C. jejuni e 4 (3,81%) para C. coli. Estes resultados foram obtidos usando tanto o método convencional de isolamento microbiológico quanto os ensaios de reação em cadeia da polymerase (PCR). Todas as cepas positivas foram submetidas ao teste de susceptibilidade antimicrobiana para sete antimicrobianos. As incidências de resistência encontradas nas cepas de C. jejuni foram as seguintes: 71,43% para tetraciclina e ácido nalidíxico, 42,86% para estreptomicina e gentamicina, 57,14% para ciprofloxacina e eritromicina e 28,57% para cloranfenicol. Entre as cepas de C. coli, 100% foram resistentes à tetraciclina e à estreptomicina, 75% eram resistentes à eritromicina, 50% eram resistentes à ciprofloxacina, gentamicina e ácido nalidíxico, e nenhuma cepa eram resistentes ao cloranfenicol. Ao analisar a presença de genes de resistência antimicrobiana nas cepas isoladas de C. jejuni, foram identificados os genes aph3-1 (resistência a aminoglicosídeos), aadE (resistência à estreptomicina) e tet (O) (resistência à tetraciclina) genes foram identificados, com taxas de ocorrência de 57,14%, 28,57% e 42,86%, respectivamente, enquanto que nas cepas de C. coli, houve uma taxa de ocorrência de 25% para os genes aph3-1 e tet (O). O gene aadE não foi encontrado nos isolados de C. coli. Os resultados deste estudo demonstraram a presença de C. jejuni e C. coli em carcaças de frango resfriadas comercializadas na Região do Distrito Federal e entorno, além da resistência antimicrobiana e a presença de genes de resistência nestas cepas, que podem se tornar uma possível ameaça a saúde pública.

Isolation and antimicrobial resistance of Campylobacter jejuni and Campylobacter coli found in chilled chicken carcasses in the Federal District Region and surrounding areas / Isolamento e resistência antimicrobiana de Campylobacter jejuni e Campylobacter coli encontradas em carcaças de frango resfriadas na região do Distrito Federal e entorno

This study aimed to isolate Campylobacter jejuni and Campylobacter coli from chilled chicken carcasses marketed in the Federal District Region and surrounding areas, as well as to detect the occurrence of antimicrobial resistance and genes responsible for the same. A total of 105 chilled chicken carcasses were collected, of which 7 (6.67%) were positive for C. jejuni and 4 (3.81%) were positive for C. coli. These results were obtained using both the conventional microbiological isolation method and polymerase chain reaction assays. All of the positive strains were subjected to antimicrobial susceptibility testing for seven antimicrobials. The resistance incidences found in the C. jejuni strains were as follows: 71.43% for tetracycline and nalidixic acid, 42.86% for streptomycin and gentamicin, 57.14% for ciprofloxacin and erythromycin, and 28.57% for chloramphenicol. Among the C. coli strains, 100% were resistant to tetracycline and streptomycin, 75% were resistant to erythromycin, 50% were resistant to ciprofloxacin, gentamicin, and nalidixic acid, and no strains were resistant to chloramphenicol. While analyzing the presence of antimicrobial resistance genes in the isolated C. jejuni strains, the aph3-1 (resistance to aminoglycosides), aadE (resistance to streptomycin), and tet(O) (resistance to tetracycline) genes were identified, with occurrence rates of 57.14%, 28.57%, and 42.86%, respectively, whereas in the C. coli strains, there was a 25% occurrence rate for both the aph3-1 and tet(O) genes. The aadE gene was not found in the C. coli isolates. The results of this study demonstrated the presence of C. jejuni and C. coli in chilled chicken carcasses marketed in the Federal District Region and surrounding areas, as well as the antimicrobial resistance and the presence of resistance genes in these bacteria, which may pose threats to public health.(AU)

O objetivo deste trabalho foi isolar Campylobacter jejuni e Campylobacter coli de carcaças de frango resfriadas comercializadas no Distrito Federal e entorno, bem como detectar a ocorrência de resistência antimicrobiana e genes responsáveis pela resistência antimicrobiana. Foram coletadas um total de 105 carcaças de frango resfriadas, das quais 7 (6,67%) foram positivas para C. jejuni e 4 (3,81%) para C. coli. Estes resultados foram obtidos usando tanto o método convencional de isolamento microbiológico quanto os ensaios de reação em cadeia da polymerase (PCR). Todas as cepas positivas foram submetidas ao teste de susceptibilidade antimicrobiana para sete antimicrobianos. As incidências de resistência encontradas nas cepas de C. jejuni foram as seguintes: 71,43% para tetraciclina e ácido nalidíxico, 42,86% para estreptomicina e gentamicina, 57,14% para ciprofloxacina e eritromicina e 28,57% para cloranfenicol. Entre as cepas de C. coli, 100% foram resistentes à tetraciclina e à estreptomicina, 75% eram resistentes à eritromicina, 50% eram resistentes à ciprofloxacina, gentamicina e ácido nalidíxico, e nenhuma cepa eram resistentes ao cloranfenicol. Ao analisar a presença de genes de resistência antimicrobiana nas cepas isoladas de C. jejuni, foram identificados os genes aph3-1 (resistência a aminoglicosídeos), aadE (resistência à estreptomicina) e tet (O) (resistência à tetraciclina) genes foram identificados, com taxas de ocorrência de 57,14%, 28,57% e 42,86%, respectivamente, enquanto que nas cepas de C. coli, houve uma taxa de ocorrência de 25% para os genes aph3-1 e tet (O). O gene aadE não foi encontrado nos isolados de C. coli. Os resultados deste estudo demonstraram a presença de C. jejuni e C. coli em carcaças de frango resfriadas comercializadas na Região do Distrito Federal e entorno, além da resistência antimicrobiana e a presença de genes de resistência nestas cepas, que podem se tornar uma possível ameaça a saúde pública.(AU)

Campylobacter jejuni and Campylobacter coli originated from chicken carcasses modulate their transcriptome to translate virulence genes in human cells / Campylobacter jejuni e Campylobacter coli originadas de carcaças de frango modulam seu transcriptoma para traduzir genes de virulência em células humanas

The aim was to determine the spread of genetically similar profiles of Campylobacter in chicken carcasses and evaluate their ability to produce transcripts for ciaB, dnaJ, p19 and sodB genes, before and after cultivation in Caco-2 cells. The strains used were isolated from 420 samples of chicken carcasses chilled and frozen ready for marketing. The species were identified by PCR-multiplex, the phylogeny was determined by RAPD-PCR and the presence of transcripts was performed by RT-PCR. We identified 74 (17.6%) of Campylobacter strains, being 55 (74.3%) C. jejuni and 19 (25.7%) C. coli. The phylogenetic relationship demonstrated heterogeneity between isolates of the same species, with absence of clones, indicating the high level of diversity of circulating genotypes. The gene transcription showed conflicting results before and after the culture in Caco-2 cell, so that before cultivation isolates showed greater capacity to transcribe genes related to survival and after the interaction with human cells, the strains showed higher potential to transcribe genes associated with virulence. The result of this study contributes to the understanding of how these seemingly fragile microorganisms are the most prevalent bacterial agents in human gastroenteritis.(AU)

O objetivo foi determinar a disseminação de perfis geneticamente semelhantes de Campylobacter em carcaças de frango e avaliar sua capacidade de produzir transcritos para os genes ciaB, dnaJ, p19 e sodB, antes e após o cultivo em células Caco-2. As cepas utilizadas foram isoladas de 420 amostras de carcaças de frango resfriadas e congeladas prontas para comercialização. As espécies foram identificadas por PCR-multiplex, a filogenia foi determinada por RAPD-PCR e a presença de transcritos foi realizada por RT-PCR. Identificamos 74 (17,6%) das cepas de Campylobacter, sendo 55 (74,3%) C. jejuni e 19 (25,7%) C. coli. A relação filogenética demonstrou heterogeneidade entre isolados da mesma espécie, com ausência de clones, indicando o alto nível de diversidade dos genótipos circulantes. A transcrição gênica mostrou resultados conflitantes antes e após a cultura em células Caco-2, de modo que, antes do cultivo, os isolados apresentaram maior capacidade de transcrever genes relacionados à sobrevivência e após a interação com células humanas, as linhagens apresentaram maior potencial para transcrever genes associados à virulência. O resultado deste estudo contribui para a compreensão de como esses microrganismos aparentemente frágeis são os agentes bacterianos mais prevalentes na gastroenterite humana.(AU)

Campylobacter jejuni and Campylobacter coli originated from chicken carcasses modulate their transcriptome to translate virulence genes in human cells / Campylobacter jejuni e Campylobacter coli originadas de carcaças de frango modulam seu transcriptoma para traduzir genes de virulência em células humanas

The aim was to determine the spread of genetically similar profiles of Campylobacter in chicken carcasses and evaluate their ability to produce transcripts for ciaB, dnaJ, p19 and sodB genes, before and after cultivation in Caco-2 cells. The strains used were isolated from 420 samples of chicken carcasses chilled and frozen ready for marketing. The species were identified by PCR-multiplex, the phylogeny was determined by RAPD-PCR and the presence of transcripts was performed by RT-PCR. We identified 74 (17.6%) of Campylobacter strains, being 55 (74.3%) C. jejuni and 19 (25.7%) C. coli. The phylogenetic relationship demonstrated heterogeneity between isolates of the same species, with absence of clones, indicating the high level of diversity of circulating genotypes. The gene transcription showed conflicting results before and after the culture in Caco-2 cell, so that before cultivation isolates showed greater capacity to transcribe genes related to survival and after the interaction with human cells, the strains showed higher potential to transcribe genes associated with virulence. The result of this study contributes to the understanding of how these seemingly fragile microorganisms are the most prevalent bacterial agents in human gastroenteritis.(AU)

O objetivo foi determinar a disseminação de perfis geneticamente semelhantes de Campylobacter em carcaças de frango e avaliar sua capacidade de produzir transcritos para os genes ciaB, dnaJ, p19 e sodB, antes e após o cultivo em células Caco-2. As cepas utilizadas foram isoladas de 420 amostras de carcaças de frango resfriadas e congeladas prontas para comercialização. As espécies foram identificadas por PCR-multiplex, a filogenia foi determinada por RAPD-PCR e a presença de transcritos foi realizada por RT-PCR. Identificamos 74 (17,6%) das cepas de Campylobacter, sendo 55 (74,3%) C. jejuni e 19 (25,7%) C. coli. A relação filogenética demonstrou heterogeneidade entre isolados da mesma espécie, com ausência de clones, indicando o alto nível de diversidade dos genótipos circulantes. A transcrição gênica mostrou resultados conflitantes antes e após a cultura em células Caco-2, de modo que, antes do cultivo, os isolados apresentaram maior capacidade de transcrever genes relacionados à sobrevivência e após a interação com células humanas, as linhagens apresentaram maior potencial para transcrever genes associados à virulência. O resultado deste estudo contribui para a compreensão de como esses microrganismos aparentemente frágeis são os agentes bacterianos mais prevalentes na gastroenterite humana.(AU)

Campylobacter jejuni and Campylobacter coli originated from chicken carcasses modulate their transcriptome to translate virulence genes in human cells

ABSTRACT: The aim was to determine the spread of genetically similar profiles of Campylobacter in chicken carcasses and evaluate their ability to produce transcripts for ciaB, dnaJ, p19 and sodB genes, before and after cultivation in Caco-2 cells. The strains used were isolated from 420 samples of chicken carcasses chilled and frozen ready for marketing. The species were identified by PCR-multiplex, the phylogeny was determined by RAPD-PCR and the presence of transcripts was performed by RT-PCR. We identified 74 (17.6%) of Campylobacter strains, being 55 (74.3%) C. jejuni and 19 (25.7%) C. coli. The phylogenetic relationship demonstrated heterogeneity between isolates of the same species, with absence of clones, indicating the high level of diversity of circulating genotypes. The gene transcription showed conflicting results before and after the culture in Caco-2 cell, so that before cultivation isolates showed greater capacity to transcribe genes related to survival and after the interaction with human cells, the strains showed higher potential to transcribe genes associated with virulence. The result of this study contributes to the understanding of how these seemingly fragile microorganisms are the most prevalent bacterial agents in human gastroenteritis.

RESUMO: O objetivo foi determinar a disseminação de perfis geneticamente semelhantes de Campylobacter em carcaças de frango e avaliar sua capacidade de produzir transcritos para os genes ciaB, dnaJ, p19 e sodB, antes e após o cultivo em células Caco-2. As cepas utilizadas foram isoladas de 420 amostras de carcaças de frango resfriadas e congeladas prontas para comercialização. As espécies foram identificadas por PCR-multiplex, a filogenia foi determinada por RAPD-PCR e a presença de transcritos foi realizada por RT-PCR. Identificamos 74 (17,6%) das cepas de Campylobacter, sendo 55 (74,3%) C. jejuni e 19 (25,7%) C. coli. A relação filogenética demonstrou heterogeneidade entre isolados da mesma espécie, com ausência de clones, indicando o alto nível de diversidade dos genótipos circulantes. A transcrição gênica mostrou resultados conflitantes antes e após a cultura em células Caco-2, de modo que, antes do cultivo, os isolados apresentaram maior capacidade de transcrever genes relacionados à sobrevivência e após a interação com células humanas, as linhagens apresentaram maior potencial para transcrever genes associados à virulência. O resultado deste estudo contribui para a compreensão de como esses microrganismos aparentemente frágeis são os agentes bacterianos mais prevalentes na gastroenterite humana.

Caracterização de estirpes de Campylobacter coli isoladas de carcaças de ovinos e de efluentes de abatedouro do estado de São Paulo / Characterization of Campylobacter coli strains isolated from the carcasses of sheep and of wastewater of abattoir in the state of São Paulo

Doença bacteriana zoonótica, a campilobacteriose é responsável mundialmente por frequentes casos de gastroenterite humana. Campylobacter spp. apresenta fator de virulência associado à diarreia, denominado toxina citoletal distensiva (CDT), sendo codificado pelos genes do complexo cdt. Os objetivos do presente estudo foram: 1) isolar e identificar estirpes de Campylobacter spp. de 102 suabes de carcaças e 102 suabes retais de ovinos (Ovis aries) e de sete amostras de água dos efluentes, antes e depois do tratamento de desinfecção de abatedouro localizado no estado de São Paulo; e 2) detectar, pela técnica de multiplex-PCR, a presença do complexo de genes cdt. Foram isoladas e identificadas, por métodos fenotípicos e genotípicos, sete estirpes de Campylobacter coli provenientes de 4/102 (3,92%) das amostras de suabes retais, 1/102 (0,98%) de suabes de carcaças e 2/7 (28,5%) das águas dos efluentes. Dos isolados de suabes retais, em 2/7 (28,6%) estirpes foi detectada a presença dos genes cdt. Trata-se do primeiro relato de isolamento de estirpes de Campylobacter coli provenientes de abatedouro de ovinos e das estirpes portadoras do complexo de genes cdt nessa espécie animal no Brasil.(AU)

A zoonosis and bacterial disease, campylobacteriosis is responsible for frequent cases of human gastroenteritis worldwide. Campylobacter spp. presents the virulence factor called cytolethal distensive toxine (CDT), responsible for diarrhea and codified by the cdt gene. The aims of this study were: 1) to isolate and identify Campylobacter spp. strains from 102 carcass swabs and 102 rectal swabs of sheep (Ovis aries) and seven samples of wastewater, before and after the disinfection treatment, collected from the abattoir of the state of São Paulo; and 2) to detect the presence of cdt gene complex by Multiplex-PCR in strains of Campylobacter spp. Seven strains of Campylobacter coli were isolated and identified by phenotypic and genotypic methods: 4/102 (3.92%) from rectal swabs, 1/102 (0.98%) from carcass swabs and 2/7 (28.5%) from wastewater. From the rectal swab samples 2/7 (28.6%) strains were detected with the cdt gene. This is the first report on the isolation of Campylobacter coli from sheep abattoir, and of strains carrying the cdt gene complex in this animal species in Brazil.(AU)

Campylobacter isolation from the feces of sheep with a history of reproductive disorders bred in the state of São Paulo, Brazil / Isolamento de Campylobacter em fezes de ovinos com distúrbios reprodutivos pertencentes a criatórios do Estado de São Paulo, Brasil

Campylobacter species are a significant cause of sheep abortion in most sheep-raising countries. The relationship between the presence of Campylobacter spp. in fecal samples and reproductive disorders was investigated in 274 sheep from 28 properties in the state of São Paulo, Brazil. Biological samples from 16 aborted fetuses, one uterus, six placentas, five uterine secretion samples, five vaginal swabs, 17 semen samples, and three preputial swabs were also subjected to bacterial isolation. The bacteria were isolated from fecal samples of 14.9% (5/28) of the properties, affecting 3.65% (10/274) of the sheep, 3.5% (9/255) of females and 5.3% (1/19) of males. Campylobacter jejuni was the most prevalent species, present in 66.67% (7) of the positive samples, followed by Campylobacter coli, present in 22.22% (2), and one strain was identified as Campylobacter spp. The birth of weak lambs (p=0.06, OR=6.83 and CI=1.73 to 27.05) and neonatal death (p=0.087, OR=3.5 and CI=0.83 to 14.72) were associated with the fecal isolation of Campylobacter spp. Diarrhea was also associated with the bacteria (p=0.003, OR=9.83 and CI=2.19 to 44.18). The dissemination of Campylobacter spp. in Brazilian sheep is low and that, at present, the existing strains are not responsible for significant economic losses in sheep production, especially in adult animals.(AU)

Espécies de Campylobacter são uma importânte causa de abortos em ovinos de muitos paíse. No intuito do isolamento de Campylobacter spp. em ovinos, foram colhidas 274 amostras fecais de ovinos do Estado de São Paulo, Brasil que possuiam histórico de distúrbios reprodutivos. O isolamento bacteriano ocorreu em 3,65% (10/274) das amostras, correspondendo 3,5% (9/255) das fêmeas e 5,3% (19/01) dos machos. Campylobacter jejuni foi a mais prevalente, responsável por 66,67% (7) das amostras positivas e o Campylobacter coli responsáveis por 22,22% (2) e uma cepa foi identificada como Campylobacter spp. Os parâmetros analisados que apresentaram valor de p 0,1 foram o nascimento de cordeiros "fracos" (p=0,06, OR=6,83 e IC=1,73 a 27,05) e morte neonatal (p=0,087, OR=3,5 e IC=0,83 a 14,72). Foi observado que os animais infectados apresentam maior risco de quadros de diarreia (p=0,003, OR=9,83 e IC=2,19 a 44,18). Campylobacter jejuni e Campylobacter coli estão presentes nos rebanhos paulistas e observou-se associação de ovinos com isolamento bacteriano fecal com quadro de nascimento de cordeiros fracos e consequente morte neonatal. Além de sua associação com quadros de diarreia que predispões à infecção oral de cordeiros a bactéria. A disseminação de Campylobacter spp. nos rebanhos ovinos Brasileiros é baixa e que, por enquanto, as cepas existentes não são responsáveis por grandes prejuízos na ovinocultura, principalmente em animais adultos.(AU)

Campylobacter isolation from the feces of sheep with a history of reproductive disorders bred in the state of São Paulo, Brazil / Isolamento de Campylobacter em fezes de ovinos com distúrbios reprodutivos pertencentes a criatórios do Estado de São Paulo, Brasil

Campylobacter species are a significant cause of sheep abortion in most sheep-raising countries. The relationship between the presence of Campylobacter spp. in fecal samples and reproductive disorders was investigated in 274 sheep from 28 properties in the state of São Paulo, Brazil. Biological samples from 16 aborted fetuses, one uterus, six placentas, five uterine secretion samples, five vaginal swabs, 17 semen samples, and three preputial swabs were also subjected to bacterial isolation. The bacteria were isolated from fecal samples of 14.9% (5/28) of the properties, affecting 3.65% (10/274) of the sheep, 3.5% (9/255) of females and 5.3% (1/19) of males. Campylobacter jejuni was the most prevalent species, present in 66.67% (7) of the positive samples, followed by Campylobacter coli, present in 22.22% (2), and one strain was identified as Campylobacter spp. The birth of weak lambs (p=0.06, OR=6.83 and CI=1.73 to 27.05) and neonatal death (p=0.087, OR=3.5 and CI=0.83 to 14.72) were associated with the fecal isolation of Campylobacter spp. Diarrhea was also associated with the bacteria (p=0.003, OR=9.83 and CI=2.19 to 44.18). The dissemination of Campylobacter spp. in Brazilian sheep is low and that, at present, the existing strains are not responsible for significant economic losses in sheep production, especially in adult animals.

Espécies de Campylobacter são uma importânte causa de abortos em ovinos de muitos paíse. No intuito do isolamento de Campylobacter spp. em ovinos, foram colhidas 274 amostras fecais de ovinos do Estado de São Paulo, Brasil que possuiam histórico de distúrbios reprodutivos. O isolamento bacteriano ocorreu em 3,65% (10/274) das amostras, correspondendo 3,5% (9/255) das fêmeas e 5,3% (19/01) dos machos. Campylobacter jejuni foi a mais prevalente, responsável por 66,67% (7) das amostras positivas e o Campylobacter coli responsáveis por 22,22% (2) e uma cepa foi identificada como Campylobacter spp. Os parâmetros analisados que apresentaram valor de p 0,1 foram o nascimento de cordeiros "fracos" (p=0,06, OR=6,83 e IC=1,73 a 27,05) e morte neonatal (p=0,087, OR=3,5 e IC=0,83 a 14,72). Foi observado que os animais infectados apresentam maior risco de quadros de diarreia (p=0,003, OR=9,83 e IC=2,19 a 44,18). Campylobacter jejuni e Campylobacter coli estão presentes nos rebanhos paulistas e observou-se associação de ovinos com isolamento bacteriano fecal com quadro de nascimento de cordeiros fracos e consequente morte neonatal. Além de sua associação com quadros de diarreia que predispões à infecção oral de cordeiros a bactéria. A disseminação de Campylobacter spp. nos rebanhos ovinos Brasileiros é baixa e que, por enquanto, as cepas existentes não são responsáveis por grandes prejuízos na ovinocultura, principalmente em animais adultos.

Occurrence of Campylobacter jejuni and Campylobacter coli biotypes and antimicrobial susceptibility in healthy dogs in Southern Chile

Background: The thermotolerant species of Campylobacter, mainly C. jejuni and C. coli, are as important agents of human gastroenteritis worldwide, being a serious public health problem. They are widely distributed in the environment and the animal kingdom having as reservoirs a wide variety of animals, including dogs, which in turn can act as a source of infection for humans. The Campylobacter isolation rates found in dogs are heterogeneous with few available data in Latin American countries. However Campylobacter diarrhea in humans is a self-limited clinical process being antimicrobial treatment not always necessary, it is necessary to know their antimicrobial susceptibility/resistance profiles in clinical and reservoirs isolates, especially in C. jejuni and C. coli strains isolated from animals close to man as dogs are. The aims of this work were to determine the isolation frequency of C. jejuni and C. coli and their biotypes in healthy dogs and the susceptibility/resistance profiles of the isolated strains to six antimicrobial drugs. Materials, Methods & Results: Fecal samples from 141 healthy dogs were obtained, seeded into the TEC transport medium. In the laboratory, each sample was plated out on modified Skirrow medium and incubated at 42°C for 48 h under microaerobic conditions. The isolated strains were identified to species and biotype levels through their phenotypic characteristics using the API Campy® procedure (bioMérieux, Marcy/Etoile, France) and the method described by Lior, respectively. The antimicrobial susceptibility patterns to ampicillin, ciprofloxacin, chloramphenicol, erythromycin, gentamicin and tetracycline were determined by means of the E-test method, considering as the minimal inhibitory concentrations (MICs) the lowest antibiotic concentrations yielding no growth. Ampicillin resistant strains were tested for ß-lactamase production with the chromogenic cephalosporin test and the disc diffusion susceptibility test for ampicillin-sulbactam. The recovery rate of Campylobacter was 31.2% (C. jejuni 22.7%, C. coli 8.5%). Three of the four biotypes described for C. jejuni and the two described for C. coli were found being C. jejuni biotype I the most frequent one (53.1%). None of the strains showed resistance to chloramphenicol, erythromycin, gentamicin and tetracycline. Resistant strains were found for ciprofloxacin (6 strains, 13.9%) and ampicillin (4 strains, 9.1%). All the ampicillin resistant strains were found to be ß-lactamase producers. Discussion: The Campylobacter isolation rate (31.2%) found is not negligible. It may provide an epidemiological insight about the risk of zoonotic infection for individuals maintaining contact with dogs, especially for children. Direct contact with pets is a risk factor for acquiring campylobacteriosis, which was also observed in Latin-American countries. Ampicillin resistance is related to ß-lactamase production and was reported in different countries, including Chile. Campylobacter resistance to quinolones has been increasingly reported in strains of human and animal origin, especially from animals related to husbandry. The ciprofloxacin resistant strains found in this study probably could be a reflection of their spread to human pets. Because dogs have been identified as a potential source of human Campylobacter infections, exposure to animals carrying fluoroquinolone-resistant microorganisms could be a risk factor for acquiring this kind of strains.

Occurrence of Campylobacter jejuni and Campylobacter coli biotypes and antimicrobial susceptibility in healthy dogs in Southern Chile / Occurrence of Campylobacter jejuni and Campylobacter coli biotypes and antimicrobial susceptibility in healthy dogs in Southern Chile

Background: The thermotolerant species of Campylobacter, mainly C. jejuni and C. coli, are as important agents of human gastroenteritis worldwide, being a serious public health problem. They are widely distributed in the environment and the animal kingdom having as reservoirs a wide variety of animals, including dogs, which in turn can act as a source of infection for humans. The Campylobacter isolation rates found in dogs are heterogeneous with few available data in Latin American countries. However Campylobacter diarrhea in humans is a self-limited clinical process being antimicrobial treatment not always necessary, it is necessary to know their antimicrobial susceptibility/resistance pro- fi les in clinical and reservoirs isolates, especially in C. jejuni and C. coli strains isolated from animals close to man as dogs are. The aims of this work were to determine the isolation frequency of C. jejuni and C. coli and their biotypes in healthy dogs and the susceptibility/resistance profiles of the isolated strains to six antimicrobial drugs. Materials, Methods & Results: Fecal samples from 141 healthy dogs were obtained, seeded into the TEC transport medium. In the laboratory, each sample was plated out on modified Skirrow medium and incubated at 42C for 48 h under microaerobic conditions. The isolated strains were identified to species and biotype levels through their pheno

Background: The thermotolerant species of Campylobacter, mainly C. jejuni and C. coli, are as important agents of human gastroenteritis worldwide, being a serious public health problem. They are widely distributed in the environment and the animal kingdom having as reservoirs a wide variety of animals, including dogs, which in turn can act as a source of infection for humans. The Campylobacter isolation rates found in dogs are heterogeneous with few available data in Latin American countries. However Campylobacter diarrhea in humans is a self-limited clinical process being antimicrobial treatment not always necessary, it is necessary to know their antimicrobial susceptibility/resistance pro- fi les in clinical and reservoirs isolates, especially in C. jejuni and C. coli strains isolated from animals close to man as dogs are. The aims of this work were to determine the isolation frequency of C. jejuni and C. coli and their biotypes in healthy dogs and the susceptibility/resistance profiles of the isolated strains to six antimicrobial drugs. Materials, Methods & Results: Fecal samples from 141 healthy dogs were obtained, seeded into the TEC transport medium. In the laboratory, each sample was plated out on modified Skirrow medium and incubated at 42C for 48 h under microaerobic conditions. The isolated strains were identified to species and biotype levels through their pheno

ISOLAMENTO E CARACTERIZAÇÃO FENOTÍPICA DE Campylobacter spp. EM MARRECOS DE PEQUIM (Anasbochas) ABATIDOS E COMERCIALIZADOS NO ESTADO DO RIO DE JANEIRO

As doenças oriundas de agentes etiológicos presentes nos alimentos são consideradas um grande problema de saúde pública mundial, e entre os microrganismos que podem ser veiculados destacam-se as bactérias do gênero Campylobacter spp.Há menos relatos sobre a ocorrência de Campylobacter em marrecos de Pequim em comparação ao frango de corte, o que induziu ao desenvolvimento desta pesquisa; como objetivo de isolar, identificar e avaliar a ocorrência de Campylobacter spp. nos marrecos de Pequim (Anas bochas), em amostras de pele, carne, miúdo e ceco de carcaças provenientes de abatedouro avícola submetido ao Serviço de Inspeção Estadual (SIE) e abatedouro clandestino de vendas a varejo, e comercializadas no Estado do Rio de Janeiro. Foram analisadas por cultura, isolamento, caracterização morfotintorial e bioquímica um total de 360 amostras (135 amostras de ceco, 75 de pele, 75 de carne e 75 de miúdos). Dentre as amostras analisadas pela metodologia de cultura e isolamento, 66 (18,33%) foram positivas para Campylobacter spp., sendo 62 provenientes de amostras de ceco e quatro de amostras de pele. Pela caracterização bioquímica, 36 (55%) foram positivas para Campylobacter jejuni e 30 (45%) para Campylobacter coli, dois dos agentes etiológicos mais comuns de gastroenterite ocasionada pela ingestão de alimentos contaminados em vários países do mundo, embora no Brasil a campilobacteriose seja subdiagnosticada e os dados subestimados. Observou-se que alguns isolados sugestivos por apresentarem positividade em características morfotintoriais, não sobreviveram nos meios de cultura de manutenção para posterior realização de provas bioquímicas e, portanto, foram considerados negativos. A presença de Campylobacter spp. nas amostras avaliadas é indicativa da importância do microrganismo e da necessidade de se colocar em prática os programas de Boas Práticas de Fabricação (BPF), Procedimentos Padrão de Higiene Operacional (PPHO) e Análise de Perigos e Pontos Críticos de Controle (APPCC) nos estabelecimentos industrializadores e comercializadores de alimentos. Sugere-se a inclusão do agente etiológico noPrograma Nacional de Controle de Patógenos (PNCP), o que possibilitaria a identificação de sua prevalência em produtos de origem animal e a implantação de medidas específicas de controle.

The diseases caused by etiological agents present in food are considered a major public health problem worldwide, and among the microorganisms that can be transmitted are the bacteria of the genus Campylobacter spp. There are fewer reports on the prevalence of Campylobacter in Pekin ducks(Anas boschas) compared to broiler chicken, which led to the development of this research; in order to isolate, identify and evaluate the incidence of Campylobacter spp.in Pekin ducks(Anas boschas), in samples of skin, meat, kid and cecum ofcarcasses from a poultry slaughterhouse submitted to the State Inspection Service (SIE) and a clandestine slaughterhouse of retail sales, and marketed in the State of Rio de Janeiro. A total of 360 samples (135 samples of cecum, 75 of skin, 75 of meat and 75 of kids) were analyzed by culture, isolation, morphotintorial and biochemical characterization. Among the samples analyzed by the culture and isolation methodology, 66 (18.33%) presented positive results for Campylobacter spp., of which 62 came from cecum samples and four from skin samples. By the biochemical characterization, 36 (55%) were positive for Campylobacter jejuni and 30 (45%) for Campylobacter coli, two of the most common etiological agents of foodborne gastroenteritis in several countries of the world, although in Brazil the campylobacteriosis are underdiagnosed and underestimated data. It was observed that some isolates suggestive of morphotintorial characteristics did not survive in the maintenance culture media for subsequent biochemical tests and, therefore, were considered negative. The presence of Campylobacter spp. in the samples evaluated is indicative of the need to put into practice the Good Manufacturing Practice (GMP), Sanitation Standard Operating Procedures (SSOP) and Hazard Analysis and Critical Control Points (HACCP) programs in food processing and marketing establishments. It is suggested the inclusion of the etiological agent in the National Pathogen Control Program (NPCP), which would allow the identification of its prevalence in products of animal origin and the implementation of specific control measures.

Transmission of Campylobacter coli in chicken embryos

Campylobacter coli is an important species involved in human cases of enteritis, and chickens are carriers of the pathogen mainly in developing country. The current study aimed to evaluate the transmission of C. coli and its pathogenic effects in chicken embryos. Breeder hens were inoculated intra-esophageally with C. coli isolated from chickens, and their eggs and embryos were analyzed for the presence of bacteria using real-time PCR and plate culture. The viability of embryos was verified. In parallel, SPF eggs were inoculated with C. coli in the air sac; after incubation, the embryos were submitted to the same analysis as the embryos from breeder hens. In embryos and fertile eggs from breeder hens, the bacterium was only identified by molecular methods; in the SPF eggs, however, the bacterium was detected by both techniques. The results showed no relationship between embryo mortality and positivity for C. coli in the embryos from breeder hens. However, the presence of bacteria is a cause of precocious mortality for SPF embryos. This study revealed that although the vertical transmission is a possible event, the bacteria can not grow in embryonic field samples.

Detecção de fatores de virulência em estirpes de Campylobacter spp. isoladas de carcaças de suínos abatidos em frigoríficos / Virulence factors in strains of Campylobacter spp. isolated from the carcasses of swine slaughtered in abattoirs

Isolaram-se estirpes de Campylobacter spp. em amostras de carcaças (n=65), fezes (n=65) e linfonodos mesentéricos (n=65) de suínos abatidos em frigoríficos do estado de São Paulo e detectaram, pela técnica da Multiplex-PCR, a presença do complexo de genes cdt, responsáveis pela expressão do fator de virulência da toxina CDT. Do total de 195 amostras de origem suína, Campylobacter spp. foi isolado de 31 (15,9%), sendo 29 (93,6%) de amostras de suabe retal, 1/65 (3,2%) de suabe de carcaça e um (3,2%) de linfonodo. Vinte e oito estirpes de C. coli foram positivas para a detecção dos genes cdt, e três estirpes de C. jejuni foram negativas para a detecção desses genes. Foi detectada, pela primeira vez no estado de São Paulo, a presença dos genes cdt em 100% das estirpes de Campylobacter coli provenientes de suínos abatidos em frigoríficos.(AU)

The purposes of this study were to isolate and identify Campylobacter spp. strains from the carcasses (n=65), feces (n=65) and mesenteric lymph nodes (n=65) of swine slaughtered in abattoirs in the State of Sao Paulo and to detect the presence of the cdt gene complex - responsible for the expression of the virulence factor cytolethal distensive toxin - in these Campylobacter spp. strains through Multiplex-PCR. From 195 samples analyzed, Campylobacter spp. was isolated in 31 (15.9%): 29 (93,6%) samples of rectal swab, 1 (3.2%) carcass swab and 1 (3.2%) lymph node sample. The 28 strains of isolated C. coli were positive for CDT toxin genes and the three strains of isolated C. jejuni were negative for these genes. It was also the first time that the cdt gene cluster was detected in strains isolated from swine in the state of São Paulo. These findings indicate swine as a potential spreading source of virulent strains of Campylobacter coli, either for slaughterhouse staff or consumers of carcasses and sub products.(AU)

Occurrence of Campylobacter jejuni and Campylobacter coli biotypes and antimicrobial susceptibility in healthy dogs in Southern Chile / Occurrence of Campylobacter jejuni and Campylobacter coli biotypes and antimicrobial susceptibility in healthy dogs in Southern Chile

Background: The thermotolerant species of Campylobacter, mainly C. jejuni and C. coli, are as important agents of human gastroenteritis worldwide, being a serious public health problem. They are widely distributed in the environment and the animal kingdom having as reservoirs a wide variety of animals, including dogs, which in turn can act as a source of infection for humans. The Campylobacter isolation rates found in dogs are heterogeneous with few available data in Latin American countries. However Campylobacter diarrhea in humans is a self-limited clinical process being antimicrobial treatment not always necessary, it is necessary to know their antimicrobial susceptibility/resistance pro- fi les in clinical and reservoirs isolates, especially in C. jejuni and C. coli strains isolated from animals close to man as dogs are. The aims of this work were to determine the isolation frequency of C. jejuni and C. coli and their biotypes in healthy dogs and the susceptibility/resistance profiles of the isolated strains to six antimicrobial drugs. Materials, Methods & Results: Fecal samples from 141 healthy dogs were obtained, seeded into the TEC transport medium. In the laboratory, each sample was plated out on modified Skirrow medium and incubated at 42C for 48 h under microaerobic conditions. The isolated strains were identified to species and biotype levels through their pheno

Background: The thermotolerant species of Campylobacter, mainly C. jejuni and C. coli, are as important agents of human gastroenteritis worldwide, being a serious public health problem. They are widely distributed in the environment and the animal kingdom having as reservoirs a wide variety of animals, including dogs, which in turn can act as a source of infection for humans. The Campylobacter isolation rates found in dogs are heterogeneous with few available data in Latin American countries. However Campylobacter diarrhea in humans is a self-limited clinical process being antimicrobial treatment not always necessary, it is necessary to know their antimicrobial susceptibility/resistance pro- fi les in clinical and reservoirs isolates, especially in C. jejuni and C. coli strains isolated from animals close to man as dogs are. The aims of this work were to determine the isolation frequency of C. jejuni and C. coli and their biotypes in healthy dogs and the susceptibility/resistance profiles of the isolated strains to six antimicrobial drugs. Materials, Methods & Results: Fecal samples from 141 healthy dogs were obtained, seeded into the TEC transport medium. In the laboratory, each sample was plated out on modified Skirrow medium and incubated at 42C for 48 h under microaerobic conditions. The isolated strains were identified to species and biotype levels through their pheno

Neonatal Campylobacter coli hemorrhagic enteritis and bacteraemia

A case of neonatal campylobacteriosis with hemorrhagic enteritis and bacteraemia due to Campylobacter coli is presented. The mother, from a rural area, had three febrile self-limited diarrheic episodes during pregnancy. The neonate probably acquired the infection during labor. The newborn's serum showed high levels of specific immunoglobulins which could explain the scarce symptoms in this newborn, despite the delay in establishing the gentamicin therapy.

Um caso de campylobacteriose neonatal com enterite hemorrágica e bacteremia produzido por Campylobacter coli é apresentado. A mãe, proveniente de uma região rural, apresentou durante a gravidez, três episódios de diarréia autolimitada. A infecção no recém nascido provavelmente foi adqüirida durante o parto. Os altos níveis séricos de immunoglobulinas específicas poderiam explicar a escassa sintomatologia, apesar da demorada prescrição do tratamento com gentamicina.