Escherichia coli in Chicken Carcasses in Southern Brazil: Absence of Shigatoxigenic (STEC) and Isolation of Atypical Enteropathogenic (aEPEC)

The aim of this study was to investigate the presence of shiga toxin-producing Escherichia coli (STEC) and atypical enteropathogenic Escherichia coli (aEPEC) in frozen chicken carcasses sold at stores in southern Brazil. Typical E. coli colonies were enumerated in 246 chicken carcasses, and the presence of stx1, stx2, eae genes was investigated in their rinse liquid and in E. coli strains isolated from those carcasses. Strains of E. coli were also investigated for the presence of bfp gene. A median of 0.6 cfu.g-1(ranging from 0.1 to 242.7 cfu.g-1) of typical E. coli colonies was found in the carcasses. Shiga toxin-encoding genes (stx1 and stx2) were not detected, indicating that the chicken carcasses were negative for STEC. The intimin protein gene (eae) was detected in E.coli isolated from 4.88% of the carcasses; all tested strains were negative for the bfp gene and were classified as aEPEC. Twenty-two aEPEC strains were tested for resistance to ten antimicrobials and subjected to macrorestriction (PFGE). All the tested aEPEC strains were fully susceptible to cephalosporins, ciprofloxacin and colistin. Resistance to sulfonamide (65%), ampicillin (55%), tetracycline (50%) and gentamicin (45%) were the most frequent. The PFGE profile demonstrated a low level of similarity among the resistant strains, indicating that they were epidemiologically unrelated. The results indicate that aEPEC strains can contaminate chicken meat, and their association with strains implicated in human diarrhea needs to be further investigated.(AU)

Escherichia coli in Chicken Carcasses in Southern Brazil: Absence of Shigatoxigenic (STEC) and Isolation of Atypical Enteropathogenic (aEPEC)

The aim of this study was to investigate the presence of shiga toxin-producing Escherichia coli (STEC) and atypical enteropathogenic Escherichia coli (aEPEC) in frozen chicken carcasses sold at stores in southern Brazil. Typical E. coli colonies were enumerated in 246 chicken carcasses, and the presence of stx1, stx2, eae genes was investigated in their rinse liquid and in E. coli strains isolated from those carcasses. Strains of E. coli were also investigated for the presence of bfp gene. A median of 0.6 cfu.g-1(ranging from 0.1 to 242.7 cfu.g-1) of typical E. coli colonies was found in the carcasses. Shiga toxin-encoding genes (stx1 and stx2) were not detected, indicating that the chicken carcasses were negative for STEC. The intimin protein gene (eae) was detected in E.coli isolated from 4.88% of the carcasses; all tested strains were negative for the bfp gene and were classified as aEPEC. Twenty-two aEPEC strains were tested for resistance to ten antimicrobials and subjected to macrorestriction (PFGE). All the tested aEPEC strains were fully susceptible to cephalosporins, ciprofloxacin and colistin. Resistance to sulfonamide (65%), ampicillin (55%), tetracycline (50%) and gentamicin (45%) were the most frequent. The PFGE profile demonstrated a low level of similarity among the resistant strains, indicating that they were epidemiologically unrelated. The results indicate that aEPEC strains can contaminate chicken meat, and their association with strains implicated in human diarrhea needs to be further investigated.

Detection of the mcr-1 gene in Enteropathogenic Escherichia coli (EPEC) and Shigatoxigenic E. coli (STEC) strains isolated from broilers / Detecção do gene mcr-1 em estirpes de Escherichia coli Enteropatogênicas (EPEC) e Shigatoxigênicas (STEC) isoladas de frangos de corte

Enteropathogenic Escherichia coli (EPEC) and Shigatoxigenic E. coli (STEC) strains are among the major pathotypes found in poultry and their products, which are capable of causing human enteric infections. Colistin has been claimed the drug of choice against diseases caused by multidrug-resistant Gram-negative bacteria (MDRGN) in humans. The mcr-1 gene was the first plasmidial gene that has been described to be responsible for colistin resistance and has also been detected in birds and poultry products. Our study aimed to detect the mcr-1 gene in enteropathogenic strains of E. coli in order to evaluate the resistance to colistin in broilers. The material was obtained from 240 cloacal samples and 60 broiler carcasses. The strains were isolated by the conventional bacteriological method and by the virulence genes, which characterize the enteropathogenic strains and resistance, and the samples were detected by polymerase chain reaction (PCR). Of the 213 isolated strains of E. coli, 57 (26.76%) were characterized as atypical EPEC and 35 (16.43%) as STEC. The mcr-1 gene was found in 3.5% (2/57) of the EPEC strains and 5.7% (2/35) of the STEC strains. In this study, it was possible to confirm that the mcr-1 resistance gene is already circulating in the broiler flocks studied and may be associated with the pathogenic strains.(AU)

Escherichia coli Enteropatogênica (EPEC) e Shigatoxigênica (STEC) estão entres os principais patotipos encontrados em aves e produtos avícolas que são capazes de causar doença entérica no homem. A colistina tem sido preconizada como droga de escolha para o tratamento de doenças causadas por bactérias Gram-negativas multirresistentes em humanos. O gene mcr-1 foi o primeiro gene plasmidial a ser descrito como responsável pela resistência a colistina e tem sido descrito em aves e produtos avícolas. Este estudo tem como objetivo a detecção do gene mcr-1 em estirpes de E. coli enteropatogênicas a fim de avaliar a resistência a colistina em frangos de corte. O material foi obtido a partir de 240 amostras cloacais e 60 carcaças de frango de corte. As estirpes foram isoladas pelo método bacteriológico convencional e os genes de virulência, que caracterizam as estirpes enteropatogênicas, e resistência foram detectados pela reação em cadeia pela polimerase (PCR). Das 213 estirpes de E. coli isoladas, 57 (26,76%) foram caracterizadas como EPEC atípica e 35 (16,43%) como STEC. O gene mcr-1 foi encontrado em 3,5% (2/57) das estirpes EPEC e 5,7% (2/35) das estirpes STEC. Neste estudo foi possível confirmar que o gene de resistência mcr-1 já está em circulação nos lotes de frango de corte estudados e pode estar associado às estirpes patogênicas.(AU)

Detection of the mcr-1 gene in Enteropathogenic Escherichia coli (EPEC) and Shigatoxigenic E. coli (STEC) strains isolated from broilers / Detecção do gene mcr-1 em estirpes de Escherichia coli Enteropatogênicas (EPEC) e Shigatoxigênicas (STEC) isoladas de frangos de corte

Enteropathogenic Escherichia coli (EPEC) and Shigatoxigenic E. coli (STEC) strains are among the major pathotypes found in poultry and their products, which are capable of causing human enteric infections. Colistin has been claimed the drug of choice against diseases caused by multidrug-resistant Gram-negative bacteria (MDRGN) in humans. The mcr-1 gene was the first plasmidial gene that has been described to be responsible for colistin resistance and has also been detected in birds and poultry products. Our study aimed to detect the mcr-1 gene in enteropathogenic strains of E. coli in order to evaluate the resistance to colistin in broilers. The material was obtained from 240 cloacal samples and 60 broiler carcasses. The strains were isolated by the conventional bacteriological method and by the virulence genes, which characterize the enteropathogenic strains and resistance, and the samples were detected by polymerase chain reaction (PCR). Of the 213 isolated strains of E. coli, 57 (26.76%) were characterized as atypical EPEC and 35 (16.43%) as STEC. The mcr-1 gene was found in 3.5% (2/57) of the EPEC strains and 5.7% (2/35) of the STEC strains. In this study, it was possible to confirm that the mcr-1 resistance gene is already circulating in the broiler flocks studied and may be associated with the pathogenic strains.(AU)

Escherichia coli Enteropatogênica (EPEC) e Shigatoxigênica (STEC) estão entres os principais patotipos encontrados em aves e produtos avícolas que são capazes de causar doença entérica no homem. A colistina tem sido preconizada como droga de escolha para o tratamento de doenças causadas por bactérias Gram-negativas multirresistentes em humanos. O gene mcr-1 foi o primeiro gene plasmidial a ser descrito como responsável pela resistência a colistina e tem sido descrito em aves e produtos avícolas. Este estudo tem como objetivo a detecção do gene mcr-1 em estirpes de E. coli enteropatogênicas a fim de avaliar a resistência a colistina em frangos de corte. O material foi obtido a partir de 240 amostras cloacais e 60 carcaças de frango de corte. As estirpes foram isoladas pelo método bacteriológico convencional e os genes de virulência, que caracterizam as estirpes enteropatogênicas, e resistência foram detectados pela reação em cadeia pela polimerase (PCR). Das 213 estirpes de E. coli isoladas, 57 (26,76%) foram caracterizadas como EPEC atípica e 35 (16,43%) como STEC. O gene mcr-1 foi encontrado em 3,5% (2/57) das estirpes EPEC e 5,7% (2/35) das estirpes STEC. Neste estudo foi possível confirmar que o gene de resistência mcr-1 já está em circulação nos lotes de frango de corte estudados e pode estar associado às estirpes patogênicas.(AU)

Detection of the mcr-1 gene in Enteropathogenic Escherichia coli (EPEC) and Shigatoxigenic E. coli (STEC) strains isolated from broilers

ABSTRACT: Enteropathogenic Escherichia coli (EPEC) and Shigatoxigenic E. coli (STEC) strains are among the major pathotypes found in poultry and their products, which are capable of causing human enteric infections. Colistin has been claimed the drug of choice against diseases caused by multidrug-resistant Gram-negative bacteria (MDRGN) in humans. The mcr-1 gene was the first plasmidial gene that has been described to be responsible for colistin resistance and has also been detected in birds and poultry products. Our study aimed to detect the mcr-1 gene in enteropathogenic strains of E. coli in order to evaluate the resistance to colistin in broilers. The material was obtained from 240 cloacal samples and 60 broiler carcasses. The strains were isolated by the conventional bacteriological method and by the virulence genes, which characterize the enteropathogenic strains and resistance, and the samples were detected by polymerase chain reaction (PCR). Of the 213 isolated strains of E. coli, 57 (26.76%) were characterized as atypical EPEC and 35 (16.43%) as STEC. The mcr-1 gene was found in 3.5% (2/57) of the EPEC strains and 5.7% (2/35) of the STEC strains. In this study, it was possible to confirm that the mcr-1 resistance gene is already circulating in the broiler flocks studied and may be associated with the pathogenic strains.

RESUMO: Escherichia coli Enteropatogênica (EPEC) e Shigatoxigênica (STEC) estão entres os principais patotipos encontrados em aves e produtos avícolas que são capazes de causar doença entérica no homem. A colistina tem sido preconizada como droga de escolha para o tratamento de doenças causadas por bactérias Gram-negativas multirresistentes em humanos. O gene mcr-1 foi o primeiro gene plasmidial a ser descrito como responsável pela resistência a colistina e tem sido descrito em aves e produtos avícolas. Este estudo tem como objetivo a detecção do gene mcr-1 em estirpes de E. coli enteropatogênicas a fim de avaliar a resistência a colistina em frangos de corte. O material foi obtido a partir de 240 amostras cloacais e 60 carcaças de frango de corte. As estirpes foram isoladas pelo método bacteriológico convencional e os genes de virulência, que caracterizam as estirpes enteropatogênicas, e resistência foram detectados pela reação em cadeia pela polimerase (PCR). Das 213 estirpes de E. coli isoladas, 57 (26,76%) foram caracterizadas como EPEC atípica e 35 (16,43%) como STEC. O gene mcr-1 foi encontrado em 3,5% (2/57) das estirpes EPEC e 5,7% (2/35) das estirpes STEC. Neste estudo foi possível confirmar que o gene de resistência mcr-1 já está em circulação nos lotes de frango de corte estudados e pode estar associado às estirpes patogênicas.

Diarrheagenic Escherichia coli in raw milk, water, and cattle feces in non-technified dairy farms / Escherichia coli diarreiogênica em leite cru, água e fezes bovinas de propriedades leiteiras não tecnificadas

This study focused on detecting diarrheagenic Escherichia coli, enteropathogenic E. coli (EPEC), Shiga-toxin-producing E. coli (STEC), enterohemorrhagic E. coli (EHEC or STEC:EPEC), enterotoxigenic E. coli (ETEC), and enteroaggregative E. coli (EAEC) in raw milk, water, and cattle feces sampled from non-technified dairy farms located in the northeastern São Paulo State, Brazil. Thirty-six water samples were collected at different points, namely, water wells (8 samples), water intended for human consumption (8 samples), water from milking parlor (8 samples), and water intended for animal consumption (7 samples), headwaters (1 sample), rivers (3 samples), and reservoirs (1 sample). Three raw milk samples were taken directly from bulk tanks in each farm, totalizing 24 samples. Feces samples were collected using rectal swabs from 160 bovines (20 animals per farm). E. coli was detected in 128 feces samples (80%), 16 raw milk samples (66.67%), and 20 water samples (55.56%). STEC (26 samples, 16.25%), EPEC (10 samples, 6.25%), STEC: EPEC (5 samples, 3.13%), and STEC: ETEC (1 sample, 0.63%) were the most prevalent strains detected in samples from cattle feces. EPEC, STEC, and STEC: EPEC strains were detected in 4.17% (1 sample), 16.67% (4 samples), and 4.17% (1 sample) of raw milk samples, respectively. STEC strains were detected in water used in the milking parlor, while no EAEC strain was detected. As a conclusion, cattle feces are important contamination sources of pathogenic E. coli in non-technified dairy farms and, consequently, cross-contamination among feces, water, and/or raw milk can occur. The use of quality water and hygienic practices during milking are recommended to avoid contamination since pathogens can be transmitted to humans via raw milk or raw milk cheese ingestion.

Este estudo teve como objetivo realizar a detecção de Escherichia coli diarreiogênica (EPEC, STEC, ETEC e EAEC) em leite, água e fezes bovinas em pequenas propriedades leiteiras localizadas na Região Nordeste do Estado de São Paulo, Brasil. E. coli foi detectada em amostras obtidas de fezes (80%), leite cru (66,67%) e água (55,56%). STEC, EPEC, STEC:EPEC e STEC:ETEC foram as cepas mais prevalentes em amostras de fezes bovinas, respectivamente. Em relação ao leite cru, cepas de EPEC, STEC e STEC:EPEC foram detectadas em 4,17%, 16,67% e 4,17% das amostras, respectivamente. Ainda, detectou-se STEC na amostra de água utilizada na sala de ordenha, enquanto EAEC não foi detectada em nenhuma amostra. Conclui-se que fezes de bovinos é uma importante fonte de contaminação de E. coli patogênicas em propriedades leiteiras e podem consequentemente contaminar o leite cru e água. A importância da qualidade da água e da adoção efetiva de práticas higiênicas durante a obtenção do leite para evitar a contaminação são recomendadas devido à possibilidade de transmissão de microorganismos patogênicos a seres humanos devido a ingestão de leite cru ou queijos produzidos a partir de leite não pasteurizado.

Diarrheagenic Escherichia coli in raw milk, water, and cattle feces in non-technified dairy farms / Escherichia coli diarreiogênica em leite cru, água e fezes bovinas de propriedades leiteiras não tecnificadas

This study focused on detecting diarrheagenic Escherichia coli, enteropathogenic E. coli (EPEC), Shiga-toxin-producing E. coli (STEC), enterohemorrhagic E. coli (EHEC or STEC:EPEC), enterotoxigenic E. coli (ETEC), and enteroaggregative E. coli (EAEC) in raw milk, water, and cattle feces sampled from non-technified dairy farms located in the northeastern São Paulo State, Brazil. Thirty-six water samples were collected at different points, namely, water wells (8 samples), water intended for human consumption (8 samples), water from milking parlor (8 samples), and water intended for animal consumption (7 samples), headwaters (1 sample), rivers (3 samples), and reservoirs (1 sample). Three raw milk samples were taken directly from bulk tanks in each farm, totalizing 24 samples. Feces samples were collected using rectal swabs from 160 bovines (20 animals per farm). E. coli was detected in 128 feces samples (80%), 16 raw milk samples (66.67%), and 20 water samples (55.56%). STEC (26 samples, 16.25%), EPEC (10 samples, 6.25%), STEC: EPEC (5 samples, 3.13%), and STEC: ETEC (1 sample, 0.63%) were the most prevalent strains detected in samples from cattle feces. EPEC, STEC, and STEC: EPEC strains were detected in 4.17% (1 sample), 16.67% (4 samples), and 4.17% (1 sample) of raw milk samples, respectively. STEC strains were detected in water used in the milking parlor, while no EAEC strain was detected. As a conclusion, cattle feces are important contamination sources of pathogenic E. coli in non-technified dairy farms and, consequently, cross-contamination among feces, water, and/or raw milk can occur. The use of quality water and hygienic practices during milking are recommended to avoid contamination since pathogens can be transmitted to humans via raw milk or raw milk cheese ingestion.(AU)

Este estudo teve como objetivo realizar a detecção de Escherichia coli diarreiogênica (EPEC, STEC, ETEC e EAEC) em leite, água e fezes bovinas em pequenas propriedades leiteiras localizadas na Região Nordeste do Estado de São Paulo, Brasil. E. coli foi detectada em amostras obtidas de fezes (80%), leite cru (66,67%) e água (55,56%). STEC, EPEC, STEC:EPEC e STEC:ETEC foram as cepas mais prevalentes em amostras de fezes bovinas, respectivamente. Em relação ao leite cru, cepas de EPEC, STEC e STEC:EPEC foram detectadas em 4,17%, 16,67% e 4,17% das amostras, respectivamente. Ainda, detectou-se STEC na amostra de água utilizada na sala de ordenha, enquanto EAEC não foi detectada em nenhuma amostra. Conclui-se que fezes de bovinos é uma importante fonte de contaminação de E. coli patogênicas em propriedades leiteiras e podem consequentemente contaminar o leite cru e água. A importância da qualidade da água e da adoção efetiva de práticas higiênicas durante a obtenção do leite para evitar a contaminação são recomendadas devido à possibilidade de transmissão de microorganismos patogênicos a seres humanos devido a ingestão de leite cru ou queijos produzidos a partir de leite não pasteurizado.(AU)

Shigatoxigenic and atypical enteropathogenic Escherichia coli in fish for human consumption

Shigatoxigenic and enteropathogenic Escherichia coli with virulence and multidrug resistance profile were isolated from Nile tilapia. This study finding is of great importance to public health because they help understand this pathogen epidemiology in fish and demonstrate how these animals can transmit E. coli related diseases to humans.(AU)

Genes encoding Shiga toxin and the intimin receptor detected in faecal samples collected from wild canids

Background: Enteropathogenic Escherichia coli (EPEC) and Shiga toxin-producing E. coli (STEC) are diarrheagenic E.coli that can cause disease in humans. The pathotype EPEC leads to the attaching and effacing lesion, causing damage tothe microvilli following to diarrhea. STEC pathotypes produces cytotoxins, which in humans are responsible for hemorrhagic colitis or hemolytic uremic syndrome. Animals are the reservoirs of these pathotypes, especially ruminants. However,other animal’s species can be associated as carriers of EPEC and STEC strains. The aim of this study was to analyze wildcanid crab-eating fox (Cerdocyon thous) as potential natural carriers of STEC and EPEC E. coli.Materials, Methods & Results: Seven fecal samples were analyzed from the crab-eating fox of free-living, captured in aperi-urban area. Samples were collected from the rectal ampulla, and the animals were clinic evaluated, being consideredhealthy at the captured moment. The feces were inoculated on medium MacConkey agar, and then the plates were incubated at 37°C for 24 h. All colony forming units (CFU) were collected by plate washing with ultrapure water (2 mL) andposterior freezing at -20°C. The total bacterial DNA from the CFU collected was extracted, followed by PCR assay tosearch for three genes: stx1, stx2 (responsible for the synthesis of the Shiga toxin) and tir, which encodes the translocatedintimin receptor, related to the A/E lesion formation. Three samples were detected as positive, being one animal detected ascarrier of the stx2 gene (STEC strain), while two animals were identified as carrier of the tir gene (EPEC strains).The stx1gene was not identified on the samples. Also, in the samples, only the presence of one gene studied at a time was observed.Therefore, we have found out that the crab-eating fox...

Genes encoding Shiga toxin and the intimin receptor detected in faecal samples collected from wild canids

Background: Enteropathogenic Escherichia coli (EPEC) and Shiga toxin-producing E. coli (STEC) are diarrheagenic E.coli that can cause disease in humans. The pathotype EPEC leads to the attaching and effacing lesion, causing damage tothe microvilli following to diarrhea. STEC pathotypes produces cytotoxins, which in humans are responsible for hemorrhagic colitis or hemolytic uremic syndrome. Animals are the reservoirs of these pathotypes, especially ruminants. However,other animals species can be associated as carriers of EPEC and STEC strains. The aim of this study was to analyze wildcanid crab-eating fox (Cerdocyon thous) as potential natural carriers of STEC and EPEC E. coli.Materials, Methods & Results: Seven fecal samples were analyzed from the crab-eating fox of free-living, captured in aperi-urban area. Samples were collected from the rectal ampulla, and the animals were clinic evaluated, being consideredhealthy at the captured moment. The feces were inoculated on medium MacConkey agar, and then the plates were incubated at 37°C for 24 h. All colony forming units (CFU) were collected by plate washing with ultrapure water (2 mL) andposterior freezing at -20°C. The total bacterial DNA from the CFU collected was extracted, followed by PCR assay tosearch for three genes: stx1, stx2 (responsible for the synthesis of the Shiga toxin) and tir, which encodes the translocatedintimin receptor, related to the A/E lesion formation. Three samples were detected as positive, being one animal detected ascarrier of the stx2 gene (STEC strain), while two animals were identified as carrier of the tir gene (EPEC strains).The stx1gene was not identified on the samples. Also, in the samples, only the presence of one gene studied at a time was observed.Therefore, we have found out that the crab-eating fox...(AU)

Molecular diagnosis of diarrheagenic Escherichia coli isolated from Psittaciformes of illegal wildlife trade / Diagnóstico molecular de Escherichia coli diarreiogênicas isoladas de Psittaciformes do tráfico ilegal de animais silvestres

Diarrheagenic Escherichia coli (DEC) are considered one of the major causes of human diarrhea in developing countries. Some studies have pointed wild birds as important reservoirs for these pathogens. However, scarce species from the Psittaciformes order have been investigated. This study aimed to evaluate the presence of DEC strains in Psittaciformes from illegal wildlife trade. A total of 78 E. coli strains isolated from cloacal swab samples of 167 Psittaciformes in the Ceará State, Brazil, were evaluated regarding the presence of the following DEC virulence genes by polymerase chain reaction (PCR): eaeA and bfpA genes (Enteropathogenic E. coli - EPEC); stx1 and stx2 (Shiga toxin-producing E. coli - STEC); estA and eltB (Enterotoxigenic E. coli - ETEC); ipaH (Enteroinvasive E. coli - EIEC); aatA and aaiC (Enteroaggregative E. coli - EAEC). Positive strains for eaeA and bfpA genes were considered typical EPEC, while strain positive exclusively for the eaeA gene were classified as atypical EPEC. The eaeA gene was identified in 20 E. coli strains and bfpA in 22 isolates. In addition, 11 and 9 belonged to tEPEC and aEPEC, respectively. No strain was positive for stx1 or stx2. A total of 47 (60.3%) strains and a total of 136 birds (81.4%) were negative for the remaining DEC pathotypes investigated. In conclusion, psittacine from illegal wildlife trade in Ceará State, Brazil, presented a relevant prevalence of typical and atypical EPEC, potentially playing a role as reservoirs of DEC strains in the environment. Thus, proper control measures must be adopted to block the spread of these pathogens.(AU)

Escherichia coli diarreiogênicas (DEC) são consideradas uma das causas mais importantes de diarreia em países em desenvolvimento. Alguns estudos têm apontado aves silvestres como importantes reservatórios destes patógenos, entretanto, poucas espécies da ordem Psittaciformes têm sido investigada. O objetivo deste estudo foi analisar a presença de cepas de E. coli diarreiogênicas em Psittaciformes do tráfico de animais silvestres. Um total de 78 amostras de E. coli isoladas de suabes cloacais provenientes de 167 de Psittaciformes do Ceará, Brasil, foram avaliadas quanto a presença dos seguintes genes de virulência DEC por meio de reação em cadeia de polimerase (PCR): eaeA e bfpA (E. coli Enteropatogênica - EPEC); stx1 e stx2 (E. coli produtora de Shiga - STEC); estA e eltB (E. coli Enterotoxigênica - ETEC); ipaH (E. coli Enteroinvasiva - EIEC); aatA e aaiC (E. coli Enteroagregativa - EAEC). As cepas positivas para os genes eaeA e bfpA foram consideradas EPEC típicas, enquanto que as positivas exclusivamente para o gene eaeA foram classificadas como EPEC atípicas. O gene eaeA foi identificado em 20 cepas de E. coli e o gene bfpA em 22 dos isolados. Adicionalmente, 11 e 9 cepas foram classificadas como EPEC típicas e atípicas, respectivamente. Nenhuma cepa foi positiva para os genes stx1 e stx2. Um total de 47 cepas (60,3%) e um total de 136 aves (81,4%) foram negativas para os demais patotipos DEC pesquisados. Em conclusão, psitacídeos provenientes do tráfico de aves silvestres do estado do Ceará, Brasil, apresentaram relevante prevalência de EPEC típicas e atípicas, potencialmente participando como reservatórios de cepas DEC no ambiente. Portanto, medidas de controle devem ser adotadas para inibir a disseminação destes patógenos.(AU)

Molecular diagnosis of diarrheagenic Escherichia coli isolated from Psittaciformes of illegal wildlife trade / Diagnóstico molecular de Escherichia coli diarreiogênicas isoladas de Psittaciformes do tráfico ilegal de animais silvestres

Diarrheagenic Escherichia coli (DEC) are considered one of the major causes of human diarrhea in developing countries. Some studies have pointed wild birds as important reservoirs for these pathogens. However, scarce species from the Psittaciformes order have been investigated. This study aimed to evaluate the presence of DEC strains in Psittaciformes from illegal wildlife trade. A total of 78 E. coli strains isolated from cloacal swab samples of 167 Psittaciformes in the Ceará State, Brazil, were evaluated regarding the presence of the following DEC virulence genes by polymerase chain reaction (PCR): eaeA and bfpA genes (Enteropathogenic E. coli - EPEC); stx1 and stx2 (Shiga toxin-producing E. coli - STEC); estA and eltB (Enterotoxigenic E. coli - ETEC); ipaH (Enteroinvasive E. coli - EIEC); aatA and aaiC (Enteroaggregative E. coli - EAEC). Positive strains for eaeA and bfpA genes were considered typical EPEC, while strain positive exclusively for the eaeA gene were classified as atypical EPEC. The eaeA gene was identified in 20 E. coli strains and bfpA in 22 isolates. In addition, 11 and 9 belonged to tEPEC and aEPEC, respectively. No strain was positive for stx1 or stx2. A total of 47 (60.3%) strains and a total of 136 birds (81.4%) were negative for the remaining DEC pathotypes investigated. In conclusion, psittacine from illegal wildlife trade in Ceará State, Brazil, presented a relevant prevalence of typical and atypical EPEC, potentially playing a role as reservoirs of DEC strains in the environment. Thus, proper control measures must be adopted to block the spread of these pathogens.(AU)

Escherichia coli diarreiogênicas (DEC) são consideradas uma das causas mais importantes de diarreia em países em desenvolvimento. Alguns estudos têm apontado aves silvestres como importantes reservatórios destes patógenos, entretanto, poucas espécies da ordem Psittaciformes têm sido investigada. O objetivo deste estudo foi analisar a presença de cepas de E. coli diarreiogênicas em Psittaciformes do tráfico de animais silvestres. Um total de 78 amostras de E. coli isoladas de suabes cloacais provenientes de 167 de Psittaciformes do Ceará, Brasil, foram avaliadas quanto a presença dos seguintes genes de virulência DEC por meio de reação em cadeia de polimerase (PCR): eaeA e bfpA (E. coli Enteropatogênica - EPEC); stx1 e stx2 (E. coli produtora de Shiga - STEC); estA e eltB (E. coli Enterotoxigênica - ETEC); ipaH (E. coli Enteroinvasiva - EIEC); aatA e aaiC (E. coli Enteroagregativa - EAEC). As cepas positivas para os genes eaeA e bfpA foram consideradas EPEC típicas, enquanto que as positivas exclusivamente para o gene eaeA foram classificadas como EPEC atípicas. O gene eaeA foi identificado em 20 cepas de E. coli e o gene bfpA em 22 dos isolados. Adicionalmente, 11 e 9 cepas foram classificadas como EPEC típicas e atípicas, respectivamente. Nenhuma cepa foi positiva para os genes stx1 e stx2. Um total de 47 cepas (60,3%) e um total de 136 aves (81,4%) foram negativas para os demais patotipos DEC pesquisados. Em conclusão, psitacídeos provenientes do tráfico de aves silvestres do estado do Ceará, Brasil, apresentaram relevante prevalência de EPEC típicas e atípicas, potencialmente participando como reservatórios de cepas DEC no ambiente. Portanto, medidas de controle devem ser adotadas para inibir a disseminação destes patógenos.(AU)

Molecular diagnosis of diarrheagenic Escherichia coli isolated from Psittaciformes of illegal wildlife trade

ABSTRACT: Diarrheagenic Escherichia coli (DEC) are considered one of the major causes of human diarrhea in developing countries. Some studies have pointed wild birds as important reservoirs for these pathogens. However, scarce species from the Psittaciformes order have been investigated. This study aimed to evaluate the presence of DEC strains in Psittaciformes from illegal wildlife trade. A total of 78 E. coli strains isolated from cloacal swab samples of 167 Psittaciformes in the Ceará State, Brazil, were evaluated regarding the presence of the following DEC virulence genes by polymerase chain reaction (PCR): eaeA and bfpA genes (Enteropathogenic E. coli - EPEC); stx1 and stx2 (Shiga toxin-producing E. coli - STEC); estA and eltB (Enterotoxigenic E. coli - ETEC); ipaH (Enteroinvasive E. coli - EIEC); aatA and aaiC (Enteroaggregative E. coli - EAEC). Positive strains for eaeA and bfpA genes were considered typical EPEC, while strain positive exclusively for the eaeA gene were classified as atypical EPEC. The eaeA gene was identified in 20 E. coli strains and bfpA in 22 isolates. In addition, 11 and 9 belonged to tEPEC and aEPEC, respectively. No strain was positive for stx1 or stx2. A total of 47 (60.3%) strains and a total of 136 birds (81.4%) were negative for the remaining DEC pathotypes investigated. In conclusion, psittacine from illegal wildlife trade in Ceará State, Brazil, presented a relevant prevalence of typical and atypical EPEC, potentially playing a role as reservoirs of DEC strains in the environment. Thus, proper control measures must be adopted to block the spread of these pathogens.

RESUMO: Escherichia coli diarreiogênicas (DEC) são consideradas uma das causas mais importantes de diarreia em países em desenvolvimento. Alguns estudos têm apontado aves silvestres como importantes reservatórios destes patógenos, entretanto, poucas espécies da ordem Psittaciformes têm sido investigada. O objetivo deste estudo foi analisar a presença de cepas de E. coli diarreiogênicas em Psittaciformes do tráfico de animais silvestres. Um total de 78 amostras de E. coli isoladas de suabes cloacais provenientes de 167 de Psittaciformes do Ceará, Brasil, foram avaliadas quanto a presença dos seguintes genes de virulência DEC por meio de reação em cadeia de polimerase (PCR): eaeA e bfpA (E. coli Enteropatogênica - EPEC); stx1 e stx2 (E. coli produtora de Shiga - STEC); estA e eltB (E. coli Enterotoxigênica - ETEC); ipaH (E. coli Enteroinvasiva - EIEC); aatA e aaiC (E. coli Enteroagregativa - EAEC). As cepas positivas para os genes eaeA e bfpA foram consideradas EPEC típicas, enquanto que as positivas exclusivamente para o gene eaeA foram classificadas como EPEC atípicas. O gene eaeA foi identificado em 20 cepas de E. coli e o gene bfpA em 22 dos isolados. Adicionalmente, 11 e 9 cepas foram classificadas como EPEC típicas e atípicas, respectivamente. Nenhuma cepa foi positiva para os genes stx1 e stx2. Um total de 47 cepas (60,3%) e um total de 136 aves (81,4%) foram negativas para os demais patotipos DEC pesquisados. Em conclusão, psitacídeos provenientes do tráfico de aves silvestres do estado do Ceará, Brasil, apresentaram relevante prevalência de EPEC típicas e atípicas, potencialmente participando como reservatórios de cepas DEC no ambiente. Portanto, medidas de controle devem ser adotadas para inibir a disseminação destes patógenos.

Captive wild birds as reservoirs of enteropathogenic E. coli (EPEC) and Shiga-toxin producing E. coli (STEC)

ABSTRACT Psittacine birds have been identified as reservoirs of diarrheagenic Escherichia coli, a subset of pathogens associated with mortality of children in tropical countries. The role of other orders of birds as source of infection is unclear. The aim of this study was to perform the molecular diagnosis of infection with diarrheagenic E. coli in 10 different orders of captive wild birds in the state of São Paulo, Brazil. Fecal samples were analyzed from 516 birds belonging to 10 orders: Accipitriformes, Anseriformes, Columbiformes, Falconiformes, Galliformes, Passeriformes, Pelecaniformes, Piciformes, Psittaciformes and Strigiformes. After isolation, 401 E. coli strains were subjected to multiplex PCR system with amplification of genes eae and bfp (EPEC), stx1 and stx2 for STEC. The results of these tests revealed 23/401 (5.74%) positive strains for eae gene, 16/401 positive strains for the bfp gene (3.99%) and 3/401 positive for stx2 gene (0.75%) distributed among the orders of Psittaciformes, Strigiformes and Columbiformes. None of strains were positive for stx1 gene. These data reveal the infection by STEC, typical and atypical EPEC in captive birds. The frequency of these pathotypes is low and restricted to few orders, but the data suggest the potential public health risk that these birds represent as reservoirs of diarrheagenic E. coli.(AU)

Captive wild birds as reservoirs of enteropathogenic E. coli (EPEC) and Shiga-toxin producing E. coli (STEC)

ABSTRACT Psittacine birds have been identified as reservoirs of diarrheagenic Escherichia coli, a subset of pathogens associated with mortality of children in tropical countries. The role of other orders of birds as source of infection is unclear. The aim of this study was to perform the molecular diagnosis of infection with diarrheagenic E. coli in 10 different orders of captive wild birds in the state of São Paulo, Brazil. Fecal samples were analyzed from 516 birds belonging to 10 orders: Accipitriformes, Anseriformes, Columbiformes, Falconiformes, Galliformes, Passeriformes, Pelecaniformes, Piciformes, Psittaciformes and Strigiformes. After isolation, 401 E. coli strains were subjected to multiplex PCR system with amplification of genes eae and bfp (EPEC), stx1 and stx2 for STEC. The results of these tests revealed 23/401 (5.74%) positive strains for eae gene, 16/401 positive strains for the bfp gene (3.99%) and 3/401 positive for stx2 gene (0.75%) distributed among the orders of Psittaciformes, Strigiformes and Columbiformes. None of strains were positive for stx1 gene. These data reveal the infection by STEC, typical and atypical EPEC in captive birds. The frequency of these pathotypes is low and restricted to few orders, but the data suggest the potential public health risk that these birds represent as reservoirs of diarrheagenic E. coli.

Prevalence of Shiga Toxin-Producing and Enteropathogenic Escherichia coli in Wild and Pet Birds in Iran

The aim of this study was to investigate the prevalence of Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) strains and to identify the stx gene types in wild captive and companion birds. In total,657 E. coli isolates from 219 birds belonging to 38 different species were investigated for the presence of STEC and EPEC strains. It was shown that five birds (2.28%) carried strains positive for one or more of the virulence factors investigated. The results indicated that 1.8% (n=4) and 0.45% (n=1) of the birds carried STEC and EPEC strains, respectively. All STEC strains harbored the stx2f and eae genes and this finding reveals the role of other birds, in addition to pigeons, as reservoirs of STEC. The only EPEC strain in this study was isolated from a Myna. Based on our knowledge, this is the first report of Stx2f-producing STEC in Geese, Duck and Lesser kestrel. In conclusion, the results indicate a low frequency of STEC carriage in wild and companion birds, and point out the need of additionally screening for the presence of stx2f in all the eae-harboring strains from birds.

Prevalence of Shiga Toxin-Producing and Enteropathogenic Escherichia coli in Wild and Pet Birds in Iran

The aim of this study was to investigate the prevalence of Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) strains and to identify the stx gene types in wild captive and companion birds. In total,657 E. coli isolates from 219 birds belonging to 38 different species were investigated for the presence of STEC and EPEC strains. It was shown that five birds (2.28%) carried strains positive for one or more of the virulence factors investigated. The results indicated that 1.8% (n=4) and 0.45% (n=1) of the birds carried STEC and EPEC strains, respectively. All STEC strains harbored the stx2f and eae genes and this finding reveals the role of other birds, in addition to pigeons, as reservoirs of STEC. The only EPEC strain in this study was isolated from a Myna. Based on our knowledge, this is the first report of Stx2f-producing STEC in Geese, Duck and Lesser kestrel. In conclusion, the results indicate a low frequency of STEC carriage in wild and companion birds, and point out the need of additionally screening for the presence of stx2f in all the eae-harboring strains from birds.(AU)

PERIGOS MICROBIOLÓGICOS E QUALIDADE HIGIÊNICO-SANITÁRIA DE QUEIJOS MINAS FRESCAL INFORMAIS COMERCIALIZADOS NO NORTE DO TOCANTINS

O queijo é um produto popular que compõe quase que diariamente a dieta da população. É rico em nutrientes e por isso também um excelente meio de multiplicação de microrganismos, inclusive os patogênicos. A contaminação microbiológica desses produtos assume destacada relevância tanto para a indústria, pelas perdas econômicas, como para a saúde pública, pelo risco de causar doenças transmitidas por alimentos. Diante do exposto, o objetivo desta pesquisa foi avaliar a qualidade higiênico-sanitária e a presença de patógenos bacterianos nos queijos Minas Frescal clandestinos comercializado nas feiras livres do município de Araguaína TO. Foram coletadas 21 amostras avaliando-se a presença de coliformes totais (CT) e termotolerantes (CTT), Escherichia coli e os patótipos EPEC, STEC e EHEC, estafilococos coagulase positiva (ECP), Salmonella spp. e Listeria monocytogenes. O resultado demonstrou que 100% das amostras de queijo estavam em desacordo com o padrão máximo de CT e CTT previstos na legislação que regulamenta o controle da qualidade desse alimento. Além desses grupos, 85,71% das amostras encontravam-se acima do limite máximo previsto para ECP, com contaminação satisfatória para a potencial produção de toxinas estafilocócicas em quantidade suficiente para provocar intoxicação alimentar. Na pesquisa de E. coli diarreiogênicas 52,38%, 66,6% e 4,76% das amostras foram positivas para EPEC, STEC e EHEC, respectivamente, indicando contaminação de origem fecal nas amostras e pela potencial presença de outros enteropatógenos. Em nenhuma amostra foi detectada a presença de Salmonella spp. e Listeria monocytogenes. A alta contagem de coliformes totais e termotolerantes encontrados nas amostras demonstram condição sanitária insatisfatória na produção, armazenamento e/ou comercialização deste alimento. A presença de EPEC, STEC e EHEC e estafilococos coagulase positiva em concentrações em que há a produção de enterotoxinas, evidenciam o iminente risco à saúde pelo consumo do queijo Minas Frescal clandestino.

Cheese is a popular product that makes up the daily diet of the population almost daily. It is a product rich in nutrients and therefore also an excellent substract of multiplication of microorganisms, including pathogens. The microbiological contamination of these products assumes prominent relevance to industry, economic losses and public health, due to the risk of causing foodborne diseases. The ain of this research was to evaluate the hygienic-sanitary quality and the presence of bacterial pathogens in the clandestine Minas Frescal cheeses sold in the free markets of Araguaína TO, Brazil. Twenty-one samples were collected evaluating the presence of total (TC) and thermotolerant (CTT) coliforms, Escherichia coli and the pathotypes EPEC, STEC and EHEC, coagulase positive staphylococci (CPS), Salmonella spp. and Listeria monocytogenes. The results showed that 100% of the cheese samples were in disagreement with the maximum standard of CT and CTT in the Brazilian legislation that regulates the quality control of this food. In addition to these groups, 85.71% of the samples were above the maximum limit for, with sufficient contamination for the potential production of enough staphylococcal toxins to cause food poisoning. In the study of diarrheagenic E. coli 52.38%, 66.6% and 4.76% of the samples were positive for EPEC, STEC and EHEC respectively, indicating contamination of fecal origin in samples and potential consumer risk. No Salmonella spp. or Listeria monocytogenes were deteced in those cheese samples. The high count of total and thermotolerant coliforms found in the samples demonstrates unsatisfactory sanitary condition in the production, storage and /or commercialization of this food. The presence of EPEC, STEC and EHEC and coagulase positive staphylococci at concentrations that produce enterotoxin, shows the health risk from the imminent consumption of Minas Frescal cheese.

DISTRIBUIÇÃO MICRORREGIONAL E SUSCEPTIBILIDADE ANTIMICROBIANA DE ESCHERICHIA COLI PATOGÊNICA ISOLADAS DE BOVINOS NO MUNICÍPIO DE ARAGUAÍNA-TO

A resistência bacteriana é um problema de saúde global, onde a Escherichia coli (E. coli) está frequentemente envolvida em situações de multirresistência aos antibióticos de importância clínica para o tratamento em humanos, sendo essa espécie bacteriana utilizada frequentemente como indicador de resistência em produtos de origem animal, por ser essa uma das principais vias de disseminação de resistência a antibióticos para humanos de acordo com a Organização Mundial de Saúde (OMS). Além disso, os bovinos são sabidamente considerados reservatórios assintomáticos e potenciais disseminadores de O157:H7, sorotipo esse de importância mundial por desencadear complicações renais e neurológicas graves em humanos. Da mesma forma o sorogrupo de E. coli enteropatogência destaca-se também por ser uma das principais causas de diarreia infantil em todo o mundo, no entanto, existem poucos dados epidemiológicos acerca da frequência do mesmo em bovinos destinados ao consumo humano. Assim, o objetivo de o presente trabalho foi determinar a susceptibilidade a antibióticos utilizados rotineiramente na atenção à saúde através do teste de sensibilidade microbiana por disco- difusão, bem como identifificação dos soropatotipos O157:H7, EPEC (A, B e C) e EIEC (A e B) e pesquisa do gene eae e rfb O157:H7 em 259 amostras de E. coli isoladas de fezes e conteúdo ruminal de 323 bovinos abatidos sob Inspeção Estadual em Araguaína-To. No total, foram obtidos 259 isolados de E. coli, sendo 186 isolados de fezes e 73 isolados de conteúdo ruminal. Quanto a sorologia, 149 dos isolados foram positivos frente aos anti-soros O157:H7 e EPEC, sendo 22 para O157:H7 e 127 para EPEC (A, B e C), houve uma maior taxa de positivos para o sorogrupo B. foram obtidas 110 amostras negativas frente aos anti-soros. E quanto a pesquisa dos genes, 7 amostras das 259 foram positivas para o gene eae e nenhuma para o gene rfb O157:H7. No geral, a sulfonamida foi o antibiótico com maior índice de resistência, seguida de florfenicol, gentamicina, enrofloxacina e amoxicilina + ácido clavulânico, doxiciclina e cefepime. Ao analisarmos os diferentes materiais biológicos pesquisados, os isolados de fezes foram resistentes a quatro antibióticos e os do conteúdo ruminal a oito, e quando comparado o perfil de multirresistência os isolados de fezes foram resistentes a ciprofloxacina e os do conteúdo ruminal ao cloranfenicol, enrofloxacina, cefepime e doxiciclina. Os isolados positivos frente ao anti-soro O157:H7 apresentaram multirresistência a 9 antibióticos, e os isolados positivos frente ao anti-soro EPEC (A, B e C) foram multirresistentes a 2 antibióticos. Os isolados positivos ao gene eae apresentaram multirresistência a 6 antibióticos. Em nosso estudo, as amostras negativas frente aos anti-soros testados apresentaram perfil de multirresistência a 5 antibióticos. O perfil de multirresistência encontrados nesse trabalho sugerem a necessidade de uma maior cautela no uso de antibióticos para bovinos na região estudada, tendo em vista as altas taxas de resistência obtidas e o uso concomitante de alguns antibióticos de importância clínica em humanos, e a técnica de aglutinação direta utilizada, sugerem bovinos como potencial reservatório da categoria EPEC.

Bacterial resistance is a global health problem, where Escherichia coli (E. coli) is often involved in multidrug resistance to clinically important antibiotics for treatment in humans, and this bacterial species is often used as an indicator of resistance in bacterial products. animal origin, as this is one of the main routes of spread of antibiotic resistance to humans according to the World Health Organization (WHO). In addition, cattle are known to be asymptomatic reservoirs and potential disseminators of O157: H7, a serotype of worldwide importance for triggering severe renal and neurological complications in humans. Similarly, E. coli enteropathogens serogroup also stands out as being a major cause of childhood diarrhea worldwide, however, there is little epidemiological data on its frequency in cattle intended for human consumption. Thus, the aim of the present study was to determine the susceptibility to antibiotics routinely used in health care by means of disc-diffusion microbial susceptibility testing, as well as to identify O157: H7, EPEC (A, B and C) and EIEC seropathotypes. (A and B) and search for the eae and rfb O157: H7 gene in 259 fecal isolated E. coli samples and ruminal content of 323 cattle slaughtered under State Inspection in Araguaína-To. In total, 259 isolates of E. coli were obtained, 186 isolates of feces and 73 isolates of ruminal content. Regarding serology, 149 of the isolates were positive against O157: H7 and EPEC antisera, 22 for O157: H7 and 127 for EPEC (A, B and C), there was a higher positive rate for serogroup B. 110 negative samples were obtained from the antisera. And as for gene research, 7 samples out of 259 were positive for the eae gene and none for the rfb O157: H7 gene. Overall, sulfonamide was the antibiotic with the highest resistance, followed by florfenicol, gentamicin, enrofloxacin and amoxicillin + clavulanic acid, doxycycline and cefepime. By analyzing the different biological materials investigated, stool isolates were resistant to four antibiotics and those of rumen content to eight, and when comparing the multidrug resistance profile stool isolates were resistant to ciprofloxacin and those of rumen content to chloramphenicol, enrofloxacin, cefepime and doxycycline. O157: H7 antiserum positive isolates were multidrug resistant to 9 antibiotics, and EPEC antisera positive isolates (A, B and C) were multidrug resistant to 2 antibiotics. The isolates positive for the eae gene showed multidrug resistance to 6 antibiotics. In our study, negative samples against the antisera tested showed multidrug resistance profile to 5 antibiotics. The multidrug resistance profile found in this study suggests the need for greater caution in the use of antibiotics for cattle in the region studied, given the high resistance rates obtained and the concomitant use of some clinically important antibiotics in humans, and the technique of Direct agglutination used suggest cattle as a potential reservoir of the EPEC category.

Distribution of non-LEE-encoded type 3 secretion system dependent effectors in enteropathogenic Escherichia coli

Enteropathogenic Escherichia coli (EPEC) are important human gastroenteritis agents. The prevalence of six non-LEE genes encoding type 3 translocated effectors was investigated. The nleC, cif and nleB genes were more prevalent in typical than in atypical EPEC, although a higher diversity of genes combinations was observed in atypical EPEC.