O presente trabalho objetivou pesquisar e caracterizar Escherichia coli patogênica (E. coli produtora de toxina Shiga STEC; E. coli aviária patogênica - APEC) em fragatas (Fregata magnificens) da costa do Estado de São Paulo e, desta forma, contribuir para a compreensão de aspectos epidemiológicos deste importante grupo de enfermidades zoonóticas bacterianas, as colibaciloses. Para tanto, foram realizadas três expedições científicas aos dois sítios de nidificação de fragatas no estado de São Paulo, Ilha Principal do Arquipélago de Alcatrazes (24°06´S 45°41´W) e Ilha de Castilho (25°16´S 47°57´W), nas quais foi colhido um total de 42 amostras de \"swabs\", sendo 21 cloacais e 21 de coanas, oriundos de 18 filhotes de sexo indeterminado, 2 fêmeas adultas e 1 macho adulto de fragatas. Das 42 amostras clinicas estudadas, foram identificadas 18 com crescimento de E.coli. Destas, foram isoladas 67 cepas, que foram então submetidas à pesquisa de genes de virulência e caracterização de grupos filogenéticos através da técnica de PCR. Em seguida, as cepas que exibiram fatores de virulência foram analisadas através de sorotipagem. Para investigar os padrões de resistência e sensibilidade a antimicrobianos, foram realizados antibiogramas para 18 tipos distintos de antimicrobianos para uma cepa de cada amostra selecionada de E. coli. Os resultados alcançados mostraram que foi possível identificar 15 diferentes perfis de virulência, com positividade para os seguintes genes: fimH (98,3%), malX (52,4%), traT (31,1%), cvaC (22,9%), fyuA (19,6%), ibeA (19,6%), aer (13,1%) e papC (13,1%). A maioria dos isolados de fragatas foi caracterizado entre os grupos filogenéticos D e B2, e os principais sorotipos encontrados foram O1:H6, O2:H7, O25:H4, e O102:H10. Em relação à resistência a antimicrobianos, 60,1%% dos isolados foram sensível aos 18 diferentes antimicrobianos testados; por outro lado, o antimicrobiano que apresentou maior resistência foi a Tetraciclina, que se revelou inefetiva em 20,1% das amostras pesquisados. Até o momento não foram identificadas cepas de STEC nos indivíduos pesquisados; por outro lado, os isolados albergam genes que caracterizam as ExPEC. Tais resultados mostram que a maioria das cepas isoladas é potencialmente patogênica para as aves, podendo representar significativo risco para sanidade das aves marinhas, além de se configurar como agentes zoonóticos relevantes, enfatizando a importância de se investigar a eventual participação das aves selvagens, em especial as marinhas, na cadeia epidemiológica de doenças ocasionadas por E. coli. Tais informações poderão nortear as pesquisas de doenças selecionadas nas populações das fragatas, assim como embasar a adoção de eventuais ações em relação a aspectos de saúde animal e humana, que tenham as fragatas como um dos elos. / The present study is to investigate and characterize pathogenic Escherichia coli (E. coli Shiga toxin-producing - STEC avian pathogenic E. coli - APEC) on frigates (Fregata magnificens) from the coast of São Paulo and contribute to the understanding of epidemiological aspects in this important group of zoonotic bacterial diseases, the colibacillosis. To this, there were three scientific expeditions to two nesting sites of frigates in the state of Sao Paulo, the main island of Alcatraz (24 ° 06\'S - 45 ° 41\'W) and Castilho´s island (25 ° 16\'S - 47 ° 57\'W), in which it was collected a total of 42 samples of swabs, and 21 cloacal and choanal 21, coming from 18 offspring of indeterminate sex, 2 adult females and 1 adult male frigate. Of the 42 clinical samples studied, 18 were identified with growth of E.coli. Of these, 67 strains were isolated, which were then tested for virulence genes and characterization of phylogenetic groups by PCR technique. Then the strains that exhibited virulence factors were analyzed by serotyping. To investigate the patterns of resistance and sensitivity to antimicrobial susceptibility tests were performed for 18 different types of antimicrobials for one strain of each selected sample of E. coli. The results showed that it was possible to identify 15 different profiles of virulence, tested positive for the following genes: fimH (98.3%), malX (52.4%) traT (31.1%), cvaC (22.9 %), fyuA (19.6%), ibeA (19.6%), aer (13.1%) and papC (13.1%). Most isolates of frigates was characterized between phylogenetic groups D and B2, and the main serotypes were O1:H6, O2:H7, O25:H4, and O102:H10. For resistance to antibiotics, 60.1%% of the isolates were sensitive to 18 different antimicrobial agents tested on the other hand, the antimicrobial with the highest resistance was to tetracycline, which has proved ineffective in 20.1% of the samples studied. So far not been identified in STEC strains studied subjects on the other hand, isolates harbor genes that characterize ExPEC. These results show that most of the strains are potentially pathogenic to birds and may represent significant risk to health of sea birds, including how to configure agents relevant, emphasizing the importance of investigating the possible involvement of wild birds, especially navies in the epidemiological chain of diseases caused by E.coli. Such information could guide the research of selected diseases in populations of the frigates, as well as basing the adoption of any action in relation to aspects of animal and human health, which have the frigates as a link.

Pombos (Columba livia) são aves exóticas, sinantrópicas, reservatórios de algumas doenças de caráter zoonótico. O presente trabalho teve por objetivo caracterizar genotipicamente isolados de Salmonella spp. e Escherichia coli diarreiogênicas de pombos domésticos. Foram coletados suabes de 118 pombos provenientes de dois zoológicos de São Paulo. Para a identificação de Salmonella spp, as amostras foram submetidas ao pré-enriquecimento em água peptonada 0,1% e incubadas a 37º C por 24-48 horas, seguida do enriquecimento seletivo em caldo de tetrationato de sódio, com incubação a 37º C por 24-48h e plaqueamento em meio seletivo XLT4, a 37º C por 24-48h. Para o isolamento de E. coli, as amostras foram semeadas em ágar MacConkey, com incubação a 37º C por 24 h. As colônias suspeitas foram identificadas por série bioquímica. A técnica de PCR foi utilizada para a confirmação do gênero Salmonella spp, identificação dos sorovares de S. Enteritidis e S. Typhimurium e para a determinação dos fatores de virulência de Escherichia coli. Dentre os 118 animais analisados, 4,23% (5/118) foram positivos para Salmonella spp, sendo todos do sorovar S. Typhimurium. Foram identificadas estirpes diarreiogênicas de E. coli, positivas para os fatores de virulência eae (8/118), bfp (4/118) e stx2f (3/118). Na análise filogenética pela técnica de AFLP, as estirpes de E. coli diarreiogênicas apresentaram uma diversidade genética, mas as estirpes de S. Typhimurium foram consideradas clonais. Os dados deste trabalho destacam o risco sanitário relacionado ao forrageamento da espécie Columba livia em zoológicos, por meio da confirmação da presença de Salmonella Typhimurium, EPEC típica, EPEC atípica e STEC nas excretas destas aves / Pigeons (Columba livia) are exotic birds, synanthropic and reservoirs of some zoonotic diseases. The aim of this study was to characterize genotypically isolates of Salmonella spp. and diarrheagenic strains of Escherichia coli isolated from domestic pigeons. Fecal swabs of 118 pigeons were collected from two zoos in São Paulo. The samples were subjected to a pre-enrichment step being placed in peptone water solution of 0.1% and incubated for 24-48 h at 37º C. Thereafter it was performed selective enrichment step where the samples were transferred in sodium tetrathionate broth with incubation for 24-48 h at 37º C, followed by plating on selective medium XLT4 incubation for 24-48h at 37º C for isolation and identification of Salmonella spp. The samples were also placed in BHI broth for a period of 24 h and then plated on MacConkey medium with incubation for 24h at 37º C for isolation of E. coli. Subsequent microbiological step, the isolates were subjected to DNA extraction using the methodology described by Boom for PCR. Polymerase chain reaction (PCR) was employed for identification of the Salmonella serovar Enteritidis and Typhimurium and virulence factors of E. coli. Of the 118 animals tested, 4.23% (5/118) were positive for Salmonella spp., all characterized as being serovar S. Typhimurium. Some E. coli strains (8/118) were positive for virulence factors: eae, 3,38% (4/118), bfp and 2.54% (3/118) to stx2f. In phylogenetic analysis by AFLP, strains of diarrheagenic E. coli showed a genetic diversity, but S. Typhimurium presented a clonal relationship. Data from this study highlight the health risk related to the foraging of Columba livia in zoos, because they can be reservoirs of Salmonella Typhimurium, typical and atypical EPEC and STEC

Les Escherichia coli producteurs de Shiga-toxines (STEC) sont responsables d'infections humaines, allant d'une diarrhée aqueuse bénigne pouvant se compliquer en syndrome hémolytique et urémique (SHU), parfois mortel. La transmission de STEC à l'Homme s'effectue principalement par l'ingestion d'aliments contaminés. Le principal facteur de virulence des STEC est le gène stx (codant la Shiga-toxine), localisé dans le génome d'un prophage. La thèse a été centrée sur les STEC O26:H11, deuxième sérotype à l'origine de SHU dans le monde, et premier retrouvé dans les fromages au lait cru.Le premier objectif était de caractériser génétiquement les STEC O26:H11 et leurs phages Stx (variants du gène stx et sites d'insertion des phages Stx) afin d'identifier d'éventuelles différences entre ces souches selon leur origine. La majorité des souches alimentaires et bovines étudiées possèdent le variant stx1a et leurs phages Stx sont intégrés dans wrbA et yehV. Les souches humaines possèdent les variants stx1a et stx2a en proportions équivalentes. Leurs phages Stx sont aussi intégrés dans wrbA et yehV mais à la différence des souches alimentaires et bovines, le site yecE a été identifié comme site d'insertion. Toutes les souches humaines qui possédaient un phage Stx2a intégré dans wrbA et yecE ont causé des SHU ; ce qui pourrait être un indicateur de haute virulence. Des études ont montré que des souches E. coli Attachant/Effaçant (AEEC) O26:H11 sont isolées à partir d'aliments identifiés comme « stx+ » par PCR. En dehors de l'absence du gène stx, ces souches AEEC sont similaires aux STEC. Leur caractérisation a montré ici que la majorité d'entre elles ont leurs sites d'insertion intacts, caractéristique compatible avec une perte de phage Stx par excision spontanée. La stabilité des phages Stx a donc été évaluée chez les STEC O26:H11. La présence/absence de phages Stx a été quantifiée par PCRq pour chaque souche au niveau de la population bactérienne entière montrant que les STEC ont la capacité de perdre leurs phages Stx. En revanche, cette instabilité n'est pas liée aux sites d'insertion. Plusieurs essais visant à introduire des phages Stx dans les souches AEEC pour les convertir en STEC ont été réalisés mais les résultats ont montré qu'il était difficile d'infecter ces souches. L'étude du taux d'induction des phages Stx in vitro chez les STEC O26:H11 a montré, qu'en présence de mitomycine C, les phages Stx2 étaient plus inductibles que les phages Stx1. En revanche, il n'a pas été mis en évidence de différences en fonction de l'origine des souches testées et du site d'intégration des phages Stx. L'analyse morphologique de quelques phages Stx a montré que le type Stx1 ou Stx2 n'était pas lié à une forme spécifique de phage. L'étude des stress relatifs à la technologie fromagère a montré que le stress salin et le stress oxydatif, lié à la libération potentielle d'H2O2 par d'autres bactéries, entrainaient l'induction des phages Stx. Comme des souches AEEC sont fréquemment isolées à partir d'aliments qualifiés de « stx+» par PCR, le processus analytique d'isolement des STEC a aussi été étudié. La production de phages Stx lors de la phase d'enrichissement semble possible à partir d'un aliment contaminé. En revanche, aucun composant de cette méthode, testé individuellement, n'a pu être identifié comme inducteur des phages Stx. Ces travaux ont permis d'acquérir des connaissances sur la diversité des phages Stx issus de STEC O26:H11 isolées chez l'Homme et dans la filière laitière. Des différences au niveau des variants du gène stx mais aussi des sites d'insertion des phages Stx ont pu être observées en fonction de l'origine des souches. De plus les niveaux d'induction des phages Stx diffèrent selon le variant du gène stx. Ces différences pourraient refléter l'existence de clones distincts, aux potentiels de virulence différents, circulant dans les aliments, chez les bovins et les patients / Shiga toxin-producing Escherichia coli (STEC) are responsible for human infections, ranging from mild diarrhea to hemolytic-uremic syndrome (HUS), sometimes with fatal outcome. Transmission of STEC to humans occurs mainly through the ingestion of contaminated food. The main virulence factor of STEC is the stx gene (encodes Shiga-toxin) located in the genome of a prophage. The PhD thesis was focused on STEC O26:H11, which is the second serotype causing HUS in the world, and the first one found in raw milk cheeses. The first objective was to characterize genetically STEC O26:H11 strains and their Stx phages (stx gene subtypes and insertion sites of Stx phages) in order to identify any differences between these strains according to their origin. The majority of the investigated food and bovine strains possessed stx1a subtype and their Stx phages were integrated into wrbA and yehV. Human strains possessed the stx1a and stx2a subtypes in equivalent proportions. Their Stx phages were also integrated into wrbA and yehV but unlike food and bovine strains, yecE site were identified as insertion site. All the human strains carrying an Stx2a phage integrated into wrbA and yecE caused HUS, which could indicate a high virulence. Studies showed that Attaching/Effacing E. coli (AEEC) O26:H11 strains were isolated from foods identified as "stx +" by PCR. Except for the absence of stx gene, these AEEC strains are similar to STEC. Their characterization showed, in this study, that the majority of them had intact insertion sites, in agreement with a possible loss of Stx phage by spontaneous excision. The stability of Stx phages was evaluated in STEC O26:H11. Presence/absence of Stx phages was quantified for each strain in the total bacterial population, showing that STEC were capable of losing their Stx phages. However, this instability was not related to the insertion sites. Several attempts to introduce Stx phages in AEEC strains in order to convert them into STEC were conducted but the results showed that it was difficult to infect these strains. The induction rate of Stx phages in vitro in STEC O26:H11 showed that, with mitomycin C, the Stx2 phages were more inducible than Stx1 phages. However no difference was found with the origin of the strains tested and the Stx phage integration site used. The morphological analysis of some Stx phages showed that Stx1 or Stx2 type was not related to a specific phage shape. Study of various stress related to the cheese-making process showed that the osmotic and oxidative stress related to the potential release of H2O2 by other bacteria, led to the induction of Stx phages. Because AEEC strains are frequently isolated from food qualified as "stx+" by PCR, the analytical STEC isolation procedure was studied for its ability to induce Stx phages. Production of Stx phages during the enrichment phase seemed possible from contaminated food. However, none of the components of this method, tested individually, could be identified as an inducer of Stx phages. This work highlighted the diversity of Stx phages from STEC O26:H11 isolated from humans and dairy sector. Differences in stx subtypes and Stx phages insertion sites present among the STEC O26:H11 strains were observed depending on the origin of the strains. Moreover the induction levels of Stx phages differed according to the stx subtypes. These differences might reflect the existence of distinct clones, with varying virulence potential, circulating in foods, cattle and patients

O presente trabalho objetivou pesquisar e caracterizar Escherichia coli patogênica (E. coli produtora de toxina Shiga STEC; E. coli aviária patogênica - APEC) em fragatas (Fregata magnificens) da costa do Estado de São Paulo e, desta forma, contribuir para a compreensão de aspectos epidemiológicos deste importante grupo de enfermidades zoonóticas bacterianas, as colibaciloses. Para tanto, foram realizadas três expedições científicas aos dois sítios de nidificação de fragatas no estado de São Paulo, Ilha Principal do Arquipélago de Alcatrazes (24°06´S 45°41´W) e Ilha de Castilho (25°16´S 47°57´W), nas quais foi colhido um total de 42 amostras de \"swabs\", sendo 21 cloacais e 21 de coanas, oriundos de 18 filhotes de sexo indeterminado, 2 fêmeas adultas e 1 macho adulto de fragatas. Das 42 amostras clinicas estudadas, foram identificadas 18 com crescimento de E.coli. Destas, foram isoladas 67 cepas, que foram então submetidas à pesquisa de genes de virulência e caracterização de grupos filogenéticos através da técnica de PCR. Em seguida, as cepas que exibiram fatores de virulência foram analisadas através de sorotipagem. Para investigar os padrões de resistência e sensibilidade a antimicrobianos, foram realizados antibiogramas para 18 tipos distintos de antimicrobianos para uma cepa de cada amostra selecionada de E. coli. Os resultados alcançados mostraram que foi possível identificar 15 diferentes perfis de virulência, com positividade para os seguintes genes: fimH (98,3%), malX (52,4%), traT (31,1%), cvaC (22,9%), fyuA (19,6%), ibeA (19,6%), aer (13,1%) e papC (13,1%). A maioria dos isolados de fragatas foi caracterizado entre os grupos filogenéticos D e B2, e os principais sorotipos encontrados foram O1:H6, O2:H7, O25:H4, e O102:H10. Em relação à resistência a antimicrobianos, 60,1%% dos isolados foram sensível aos 18 diferentes antimicrobianos testados; por outro lado, o antimicrobiano que apresentou maior resistência foi a Tetraciclina, que se revelou inefetiva em 20,1% das amostras pesquisados. Até o momento não foram identificadas cepas de STEC nos indivíduos pesquisados; por outro lado, os isolados albergam genes que caracterizam as ExPEC. Tais resultados mostram que a maioria das cepas isoladas é potencialmente patogênica para as aves, podendo representar significativo risco para sanidade das aves marinhas, além de se configurar como agentes zoonóticos relevantes, enfatizando a importância de se investigar a eventual participação das aves selvagens, em especial as marinhas, na cadeia epidemiológica de doenças ocasionadas por E. coli. Tais informações poderão nortear as pesquisas de doenças selecionadas nas populações das fragatas, assim como embasar a adoção de eventuais ações em relação a aspectos de saúde animal e humana, que tenham as fragatas como um dos elos. / The present study is to investigate and characterize pathogenic Escherichia coli (E. coli Shiga toxin-producing - STEC avian pathogenic E. coli - APEC) on frigates (Fregata magnificens) from the coast of São Paulo and contribute to the understanding of epidemiological aspects in this important group of zoonotic bacterial diseases, the colibacillosis. To this, there were three scientific expeditions to two nesting sites of frigates in the state of Sao Paulo, the main island of Alcatraz (24 ° 06\'S - 45 ° 41\'W) and Castilho´s island (25 ° 16\'S - 47 ° 57\'W), in which it was collected a total of 42 samples of swabs, and 21 cloacal and choanal 21, coming from 18 offspring of indeterminate sex, 2 adult females and 1 adult male frigate. Of the 42 clinical samples studied, 18 were identified with growth of E.coli. Of these, 67 strains were isolated, which were then tested for virulence genes and characterization of phylogenetic groups by PCR technique. Then the strains that exhibited virulence factors were analyzed by serotyping. To investigate the patterns of resistance and sensitivity to antimicrobial susceptibility tests were performed for 18 different types of antimicrobials for one strain of each selected sample of E. coli. The results showed that it was possible to identify 15 different profiles of virulence, tested positive for the following genes: fimH (98.3%), malX (52.4%) traT (31.1%), cvaC (22.9 %), fyuA (19.6%), ibeA (19.6%), aer (13.1%) and papC (13.1%). Most isolates of frigates was characterized between phylogenetic groups D and B2, and the main serotypes were O1:H6, O2:H7, O25:H4, and O102:H10. For resistance to antibiotics, 60.1%% of the isolates were sensitive to 18 different antimicrobial agents tested on the other hand, the antimicrobial with the highest resistance was to tetracycline, which has proved ineffective in 20.1% of the samples studied. So far not been identified in STEC strains studied subjects on the other hand, isolates harbor genes that characterize ExPEC. These results show that most of the strains are potentially pathogenic to birds and may represent significant risk to health of sea birds, including how to configure agents relevant, emphasizing the importance of investigating the possible involvement of wild birds, especially navies in the epidemiological chain of diseases caused by E.coli. Such information could guide the research of selected diseases in populations of the frigates, as well as basing the adoption of any action in relation to aspects of animal and human health, which have the frigates as a link.

Pombos (Columba livia) são aves exóticas, sinantrópicas, reservatórios de algumas doenças de caráter zoonótico. O presente trabalho teve por objetivo caracterizar genotipicamente isolados de Salmonella spp. e Escherichia coli diarreiogênicas de pombos domésticos. Foram coletados suabes de 118 pombos provenientes de dois zoológicos de São Paulo. Para a identificação de Salmonella spp, as amostras foram submetidas ao pré-enriquecimento em água peptonada 0,1% e incubadas a 37º C por 24-48 horas, seguida do enriquecimento seletivo em caldo de tetrationato de sódio, com incubação a 37º C por 24-48h e plaqueamento em meio seletivo XLT4, a 37º C por 24-48h. Para o isolamento de E. coli, as amostras foram semeadas em ágar MacConkey, com incubação a 37º C por 24 h. As colônias suspeitas foram identificadas por série bioquímica. A técnica de PCR foi utilizada para a confirmação do gênero Salmonella spp, identificação dos sorovares de S. Enteritidis e S. Typhimurium e para a determinação dos fatores de virulência de Escherichia coli. Dentre os 118 animais analisados, 4,23% (5/118) foram positivos para Salmonella spp, sendo todos do sorovar S. Typhimurium. Foram identificadas estirpes diarreiogênicas de E. coli, positivas para os fatores de virulência eae (8/118), bfp (4/118) e stx2f (3/118). Na análise filogenética pela técnica de AFLP, as estirpes de E. coli diarreiogênicas apresentaram uma diversidade genética, mas as estirpes de S. Typhimurium foram consideradas clonais. Os dados deste trabalho destacam o risco sanitário relacionado ao forrageamento da espécie Columba livia em zoológicos, por meio da confirmação da presença de Salmonella Typhimurium, EPEC típica, EPEC atípica e STEC nas excretas destas aves / Pigeons (Columba livia) are exotic birds, synanthropic and reservoirs of some zoonotic diseases. The aim of this study was to characterize genotypically isolates of Salmonella spp. and diarrheagenic strains of Escherichia coli isolated from domestic pigeons. Fecal swabs of 118 pigeons were collected from two zoos in São Paulo. The samples were subjected to a pre-enrichment step being placed in peptone water solution of 0.1% and incubated for 24-48 h at 37º C. Thereafter it was performed selective enrichment step where the samples were transferred in sodium tetrathionate broth with incubation for 24-48 h at 37º C, followed by plating on selective medium XLT4 incubation for 24-48h at 37º C for isolation and identification of Salmonella spp. The samples were also placed in BHI broth for a period of 24 h and then plated on MacConkey medium with incubation for 24h at 37º C for isolation of E. coli. Subsequent microbiological step, the isolates were subjected to DNA extraction using the methodology described by Boom for PCR. Polymerase chain reaction (PCR) was employed for identification of the Salmonella serovar Enteritidis and Typhimurium and virulence factors of E. coli. Of the 118 animals tested, 4.23% (5/118) were positive for Salmonella spp., all characterized as being serovar S. Typhimurium. Some E. coli strains (8/118) were positive for virulence factors: eae, 3,38% (4/118), bfp and 2.54% (3/118) to stx2f. In phylogenetic analysis by AFLP, strains of diarrheagenic E. coli showed a genetic diversity, but S. Typhimurium presented a clonal relationship. Data from this study highlight the health risk related to the foraging of Columba livia in zoos, because they can be reservoirs of Salmonella Typhimurium, typical and atypical EPEC and STEC

Dentre os psitacídeos criados como pets, destacam-se as calopsitas (Nymphicus hollandicus), os periquitos-australianos (Melopsittacus undulatus) e agapornis (Agapornis spp.). Os benefícios decorrentes da presença destas aves em ambiente doméstico são reconhecidos, porém pouco se conhece sobre os riscos de transmissão de zoonoses. E. coli pode ser isolada em diversos sítios do corpo humano e animal. Alguns sorotipos são considerados patogênicos e, nestes casos, a infecção pode estar associada à ocorrência de diversas manifestações clínicas. O presente estudo tem como objetivo a caracterização fenotípica e genotípica de Escherichia coli diarreiogênicas isoladas de pequenos psitacídeos mantidos como pets. 171 amostras de fezes de aves domiciliadas na cidade de São Paulo foram colhidas com auxílio de suabes estéreis, sendo 67 de calopsitas, 59 de periquitos australianos e 45 agapornis. Das 171 amostras, foram identificadas 42 colônias de Escherichia coli (22 de calopsitas, 10 de periquitos e 10 de agapornis), submetidas à extração de DNA e pesquisa dos genes eae, bfpA, stx1, stx2, LT, ST e aggR ; aaiA ; aatA ; aaiG pela Reac?a?o em Cadeia da Polimerase (PCR). Os resultados apontaram positividade em dez isolados: eae+, bfpA+, Stx2f+ de três calopsitas e cinco periquitos australianos (8/10); ST de uma calopsita (1/10) e eae+ de um periquito australiano (1/10). As oito estirpes de STEC/tEPEC foram classificadas como O137:HNM e nenhuma das estirpes exibiu um perfil de multirresistência. Apenas um isolado foi positivo para formação de biofilme (DO de 0,064), sendo classificado como fracamente aderente. A análise filogenética dos 4 grupos de Clermont et al. (2013) classificou três estirpes como B2, duas como F, e uma como Clado II. Duas estirpes foram classificadas como \"não tipável\". Os resultados para o AFLP foram: 7/8 estirpes (três de calopsitas e quatro de periquitos) foram agrupadas em um mesmo cluster, apresentando 90-100% de similaridade. Os resultados do sequenciamento de fragmentos internos de genes house-keeping - Multi-locus Sequence Typing (MLST) revelou que as oito estirpes pertenciam ao ST 3072. Neste estudo, nenhuma das estirpes apresentou atividade hemolítica, mas produziram toxinas in vitro. Aderiram in vitro em células HeLa e causaram lesões A/E em modelo de alça intestinal de coelho. As estirpes induziram processo inflamatório e aumento de muco intestinal. Os dados deste trabalho revelaram o potencial zoonótico dos híbridos STEC/tEPEC sorogrupo O137:HNM; ST 3072 isolados de psitacídeos de pequeno porte criados como \"pet\". / Among the parrots (psittacidae family) kept as pets, the cockatiels (Nymphicus hollandicus), budgerigars (Melopsittacus undulatus) and lovebirds (Lovebirds spp.) stand out. Although the benefits from the presence of these birds in a household environment are acknowledged, little is known about the risks of transmission of zoonoses. E. coli can be isolated in various sites of the human and animal body. Some serotypes are considered as pathogenic, and in such cases, the infection may be associated with the occurrence of several clinical manifestations. The aim of the present study is to characterize, phenotypically and genotypically, diarrheagenic Escherichia coli, isolated from the small parrots kept as pets. One hundred and seventy-one fecal samples of birds living in households in the city of Sa?o Paulo were collected with the aid of sterile swabs, with 67 being from cockatiels, 59 from budgerigars, and 45 from lovebirds. From the 171 samples, 42 (22 from cockatiels, 10 from budgerigars and 10 from lovebirds) Escherichia coli colonies were identified, which had their DNA extracted, and the eae, bfpA, stx1, stx2, LT, ST and aggR; aaiA; aatA; aaiG genes were investigated, through Polymerase Chain Reaction (PCR). The results indicated positivity in ten isolates: eae+, bfpA+, Stx2f+ from three cockatiels and five budgerigars (8/10), ST from one cockatiel (1/10), and eae+ from one budgerigar (1/10). The eight STEC/tEPEC lineages were classified as O137:HNM, and none of the lineages showed a multiresistance antibiotic profile. Only one isolate was positive for biofilm formation (DO, 0.064), which was classified as weakly adherent. The phylogenetic analysis of the 4 groups by Clermont et al. (2013) classified three lineages as B2, two as F, and one as Clade II. Two lineages were classified as \"non-typeable.\" The results for AFLP were: 7/8 lineages (three of cockatiels and four of budgerigars) were clustered together, with 90-100% of similarity being observed. The results of the internal fragment sequencing of house- keeping genes - Multi-locus Sequence Typing (MLST) revealed that the eight lineages belonged in to ST 3072. In this study, although none of the lineages showed hemolytic activity, they produced toxins in vitro. They adhered, in vitro, to HeLa cells and caused A/E lesions in an intestinal loop model of rabbits. The lineages induced an inflammatory process and an increase of intestinal mucus. This study data revealed the zoonotic potential of STEC/tEPEC hybrids, O137:HNM; ST 3072 serogroup, isolated from small-sized parrots kept as pets

Ovinos são considerados reservatórios de Escherichia coli produtora da toxina Shiga (STEC). Este estudo pesquisou amostras de 100 carneiros no Estado de São Paulo. PCR foi empregada para detecção de stx1, stx2, eae, ehx, e intiminas. RFLP-PCR foi realizado para identificação das variantes de stx1/stx2 e fliC. Cinco isolados eae+stx- de sorotipos O128:H2, O145:H2, O153:H7 e O178:H7 apresentaram intiminas b, g e e. Cinqüenta e seis STEC foram obtidos e os genótipos foram stx1cstx2d-O118 (56,1%), stx1c (33,3%), stx2d-O118 (7,6%), e stx1cstx2dOX3a (3%). Os sorotipos mais freqüentes foram ONT:H8, ONT:H-, O75:H-, O174:H8 e O5:H-. Gene fliC codificando para H2, H8, H14, H16, H19, H21 ou H28 foi identificado em 25/33 isolados. ehx foi detectado em três/cinco EPEC Atípicas e em 38 (57,6%) STEC. Sensibilidade aos antimicrobianos foi observada em 77,2% dos isolados STEC. A análise da similaridade genética por PFGE revelou 24 perfis entre 40 isolados STEC e EPEC atípicas. O estudo demonstrou que ovinos saudáveis podem se comportar como reservatórios de STEC e EPEC atípica. / Lambs are reported as carriers of Shiga toxin-producing Escherichia coli (STEC) worldwide. This study surveyed 100 sheep in São Paulo, Brazil. PCR was used for detection of stx1, stx2, eae, ehx, and intimins. RFLP-PCR investigated the stx1/stx2 variants and flagellar antigen (fliC). Five isolates were eae+/stx-, belonging to serotypes O128:H2, O145:H2, O153:H7 and O178:H7, harboring b, g and e intimins. Fifty-six STEC isolates were recovered, and stx genotypes were stx1cstx2d-O118 (56.1%), stx1c (33.3%), stx2d-O118 (7.6%), and stx1cstx2dOX3a (3%). A diversity of STEC serotypes was observed, but most frequent were ONT:H8, ONT:H-, O75:H-, O174:H8 and O5:H-. fliC gene coding for H2, H8, H14, H16, H19, H21 or H28 was identified in 25/33 isolates. ehx gene was detected in three Atypical EPEC and in 38 (57,6%) STEC. Fifty-one (77.2%) STEC were susceptible to antimicrobials tested. Pulsed field gel electrophoresis (PFGE) revealed 24/40 profiles. The study showed that healthy ovine can be carrier of STEC and atypical EPEC in Brazil.

Dentre os psitacídeos criados como pets, destacam-se as calopsitas (Nymphicus hollandicus), os periquitos-australianos (Melopsittacus undulatus) e agapornis (Agapornis spp.). Os benefícios decorrentes da presença destas aves em ambiente doméstico são reconhecidos, porém pouco se conhece sobre os riscos de transmissão de zoonoses. E. coli pode ser isolada em diversos sítios do corpo humano e animal. Alguns sorotipos são considerados patogênicos e, nestes casos, a infecção pode estar associada à ocorrência de diversas manifestações clínicas. O presente estudo tem como objetivo a caracterização fenotípica e genotípica de Escherichia coli diarreiogênicas isoladas de pequenos psitacídeos mantidos como pets. 171 amostras de fezes de aves domiciliadas na cidade de São Paulo foram colhidas com auxílio de suabes estéreis, sendo 67 de calopsitas, 59 de periquitos australianos e 45 agapornis. Das 171 amostras, foram identificadas 42 colônias de Escherichia coli (22 de calopsitas, 10 de periquitos e 10 de agapornis), submetidas à extração de DNA e pesquisa dos genes eae, bfpA, stx1, stx2, LT, ST e aggR ; aaiA ; aatA ; aaiG pela Reac?a?o em Cadeia da Polimerase (PCR). Os resultados apontaram positividade em dez isolados: eae+, bfpA+, Stx2f+ de três calopsitas e cinco periquitos australianos (8/10); ST de uma calopsita (1/10) e eae+ de um periquito australiano (1/10). As oito estirpes de STEC/tEPEC foram classificadas como O137:HNM e nenhuma das estirpes exibiu um perfil de multirresistência. Apenas um isolado foi positivo para formação de biofilme (DO de 0,064), sendo classificado como fracamente aderente. A análise filogenética dos 4 grupos de Clermont et al. (2013) classificou três estirpes como B2, duas como F, e uma como Clado II. Duas estirpes foram classificadas como \"não tipável\". Os resultados para o AFLP foram: 7/8 estirpes (três de calopsitas e quatro de periquitos) foram agrupadas em um mesmo cluster, apresentando 90-100% de similaridade. Os resultados do sequenciamento de fragmentos internos de genes house-keeping - Multi-locus Sequence Typing (MLST) revelou que as oito estirpes pertenciam ao ST 3072. Neste estudo, nenhuma das estirpes apresentou atividade hemolítica, mas produziram toxinas in vitro. Aderiram in vitro em células HeLa e causaram lesões A/E em modelo de alça intestinal de coelho. As estirpes induziram processo inflamatório e aumento de muco intestinal. Os dados deste trabalho revelaram o potencial zoonótico dos híbridos STEC/tEPEC sorogrupo O137:HNM; ST 3072 isolados de psitacídeos de pequeno porte criados como \"pet\". / Among the parrots (psittacidae family) kept as pets, the cockatiels (Nymphicus hollandicus), budgerigars (Melopsittacus undulatus) and lovebirds (Lovebirds spp.) stand out. Although the benefits from the presence of these birds in a household environment are acknowledged, little is known about the risks of transmission of zoonoses. E. coli can be isolated in various sites of the human and animal body. Some serotypes are considered as pathogenic, and in such cases, the infection may be associated with the occurrence of several clinical manifestations. The aim of the present study is to characterize, phenotypically and genotypically, diarrheagenic Escherichia coli, isolated from the small parrots kept as pets. One hundred and seventy-one fecal samples of birds living in households in the city of Sa?o Paulo were collected with the aid of sterile swabs, with 67 being from cockatiels, 59 from budgerigars, and 45 from lovebirds. From the 171 samples, 42 (22 from cockatiels, 10 from budgerigars and 10 from lovebirds) Escherichia coli colonies were identified, which had their DNA extracted, and the eae, bfpA, stx1, stx2, LT, ST and aggR; aaiA; aatA; aaiG genes were investigated, through Polymerase Chain Reaction (PCR). The results indicated positivity in ten isolates: eae+, bfpA+, Stx2f+ from three cockatiels and five budgerigars (8/10), ST from one cockatiel (1/10), and eae+ from one budgerigar (1/10). The eight STEC/tEPEC lineages were classified as O137:HNM, and none of the lineages showed a multiresistance antibiotic profile. Only one isolate was positive for biofilm formation (DO, 0.064), which was classified as weakly adherent. The phylogenetic analysis of the 4 groups by Clermont et al. (2013) classified three lineages as B2, two as F, and one as Clade II. Two lineages were classified as \"non-typeable.\" The results for AFLP were: 7/8 lineages (three of cockatiels and four of budgerigars) were clustered together, with 90-100% of similarity being observed. The results of the internal fragment sequencing of house- keeping genes - Multi-locus Sequence Typing (MLST) revealed that the eight lineages belonged in to ST 3072. In this study, although none of the lineages showed hemolytic activity, they produced toxins in vitro. They adhered, in vitro, to HeLa cells and caused A/E lesions in an intestinal loop model of rabbits. The lineages induced an inflammatory process and an increase of intestinal mucus. This study data revealed the zoonotic potential of STEC/tEPEC hybrids, O137:HNM; ST 3072 serogroup, isolated from small-sized parrots kept as pets

Ovinos são considerados reservatórios de Escherichia coli produtora da toxina Shiga (STEC). Este estudo pesquisou amostras de 100 carneiros no Estado de São Paulo. PCR foi empregada para detecção de stx1, stx2, eae, ehx, e intiminas. RFLP-PCR foi realizado para identificação das variantes de stx1/stx2 e fliC. Cinco isolados eae+stx- de sorotipos O128:H2, O145:H2, O153:H7 e O178:H7 apresentaram intiminas b, g e e. Cinqüenta e seis STEC foram obtidos e os genótipos foram stx1cstx2d-O118 (56,1%), stx1c (33,3%), stx2d-O118 (7,6%), e stx1cstx2dOX3a (3%). Os sorotipos mais freqüentes foram ONT:H8, ONT:H-, O75:H-, O174:H8 e O5:H-. Gene fliC codificando para H2, H8, H14, H16, H19, H21 ou H28 foi identificado em 25/33 isolados. ehx foi detectado em três/cinco EPEC Atípicas e em 38 (57,6%) STEC. Sensibilidade aos antimicrobianos foi observada em 77,2% dos isolados STEC. A análise da similaridade genética por PFGE revelou 24 perfis entre 40 isolados STEC e EPEC atípicas. O estudo demonstrou que ovinos saudáveis podem se comportar como reservatórios de STEC e EPEC atípica. / Lambs are reported as carriers of Shiga toxin-producing Escherichia coli (STEC) worldwide. This study surveyed 100 sheep in São Paulo, Brazil. PCR was used for detection of stx1, stx2, eae, ehx, and intimins. RFLP-PCR investigated the stx1/stx2 variants and flagellar antigen (fliC). Five isolates were eae+/stx-, belonging to serotypes O128:H2, O145:H2, O153:H7 and O178:H7, harboring b, g and e intimins. Fifty-six STEC isolates were recovered, and stx genotypes were stx1cstx2d-O118 (56.1%), stx1c (33.3%), stx2d-O118 (7.6%), and stx1cstx2dOX3a (3%). A diversity of STEC serotypes was observed, but most frequent were ONT:H8, ONT:H-, O75:H-, O174:H8 and O5:H-. fliC gene coding for H2, H8, H14, H16, H19, H21 or H28 was identified in 25/33 isolates. ehx gene was detected in three Atypical EPEC and in 38 (57,6%) STEC. Fifty-one (77.2%) STEC were susceptible to antimicrobials tested. Pulsed field gel electrophoresis (PFGE) revealed 24/40 profiles. The study showed that healthy ovine can be carrier of STEC and atypical EPEC in Brazil.

La contamination des zones littorales par des bactéries entériques potentiellement pathogènes pour l’Homme constitue un problème majeur pour la pérennité de certains usages tel que la conchyliculture. Ces bactéries provenant de rejets agricoles ou urbains peuvent atteindre les zones conchylicoles et être impliquées dans des toxi-infections alimentaires collectives (TIAC). Actuellement, très peu de données sont disponibles sur la présence et la diversité des bactéries entériques telles que les Escherichia coli (E. coli) pathogènes de type E. coli producteurs de Shiga-toxines (STEC) et E.coli entéropathogènes (EPEC) dans les coquillages en France.La présence de ces E. coli pathogènes a été recherchée pendant deux ans, dans trois zones conchylicoles françaises et leurs bassins versants. Un total de 28 souches STEC et 89 souches EPEC différentes ont été isolées dans des coquillages, des eaux aux exutoires, le sédiment et l’eau de mer, représentant 1% de la totalité des souches E. coli isolées (n=12016). Ces souches isolées présentaient néanmoins une grande diversité. Elles étaient réparties au sein de 73 profils de virulence différents dont une souche STEC de sérotype O26 :H11 présentant 47 gènes de virulence isolée dans un lot de moule. Soixante-quinze pourcents des souches EPEC présentaient des gènes de virulence associés à des îlots de pathogénicités caractéristiques de souches pathogènes responsables d’infection grave chez l’Homme, révélant le potentiel pouvoir pathogène de certaines souches. Enfin, l’étude de la cinétique de contamination, décontamination d’huîtres au contact de souches STEC, n’a pas montré de différence comparée à un E. coli non STEC.Les travaux réalisés au cours de cette thèse sont à notre connaissance les premiers de ce genre. Ils ont permis de mettre en évidence la faible présence de STEC et de EPEC au niveau des zones conchylicoles françaises étudiées ainsi que le potentiel pouvoir pathogène de certaines souches. La faible prévalence des souches sur ces sites de catégorie B ou C (purification des coquillages avant commercialisation) reste néanmoins plutôt en faveur d’un risque faible de contamination dans les zones sélectionnées. Les résultats acquis au cours de cette thèse sont des éléments importants pour mieux appréhender le risque sanitaire potentiel lié aux STEC et aux EPEC en zone littorale. / The contamination of coastal areas by potentially pathogenic enteric bacteria is of concern for the sustainability of some uses, such as shellfish farming, recreational shellfish harvesting and bathing. The contamination of these environments may occur through the land-spreading of livestock wastes, animal feces deposited on pastures, wastewaters from slaughterhouses. The presence of these bacteria in coastal environment may present a potential risk to human health. In fact, shellfish-borne outbreaks may occur by the consumption of shellfish from contaminated areas. To date, few studies focusing on the presence and the diversity of enteric bacteria, such as pathogenic Escherichia coli (E. coli) more precisely, Shiga-toxin-producing E. coli (STEC) and enteropathogenic E. coli (EPEC) in coastal environments and shellfish have been reported.For this purpose, during a 2-year study, shellfish batches, freshwater, seawater, and surface sediment samples from three French selected shellfish-harvesting areas and their upstream catchments, were analyzed to evaluate the presence of STEC and EPEC strains. Twenty-eight STEC and 89 EPEC strains were isolated representing 1% of the total E. coli(n=12 016). The isolated STEC and EPEC strains belonged to a high diversity. One STEC strain isolated from a mussel batch, belonging to the serotype O26:H11 displayed 47 additional virulence genes. Seventy-five percent of EPEC strains displayed several virulence genes associated with pathogenicity islands specific to pathogenic E. coli involved in human infections.No difference in the kinetics of the contamination and depuration of oysters by STEC and non-STEC E. coli was found.To our knowledge, this study is the first to focus on the diversity of STEC and EPEC strains isolated from coastal environments. This study highlights the weak presence of STECs and EPECs in the French shellfish-harvesting areas studied and a potential pathogenicity of some strains. The low prevalence of STEC and EPEC strains in shellfish fromB- and C-categories (depuration of shellfish before commercialization), is rather in favor of a limited risk of contamination of shellfish in the studied areas. The results obtained during this study are important to better understand the health risk associated with STEC and EPEC in coastal areas.