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

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

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

Prevalencia y caracterización del perfil de susceptibilidad antimicrobiana de Campylobacter jejuni y Campylobacter coli en niños con diarrea de la ciudad de Loja, Ecuador / Prevalence and antimicrobial behavior of Campylobacter jejuni and Campylobacter coli in children with diarrhea in Loja city, Ecuador

Resumen Introducción: Campylobacter es un importante agente de diarrea en el ser humano. En Ecuador, la información sobre Campylobacter es escasa y no existen antecedentes de susceptibilidad antimicrobiana. Objetivo: Describir la prevalencia de Campylobacter en niños con diarrea y su comportamiento in vitro frente a cinco antimicrobianos. Método: Se estudiaron 253 niños entre siete meses y 9 años de edad, que consultaron por diarrea en dos hospitales de la ciudad de Loja. Se realizó cultivo de muestras fecales e identificación por pruebas fenotípicas y por RPC múltiple. La susceptibilidad antimicrobiana fue determinada por el método de difusión en disco. Resultados: Campylobacter fue diagnosticado en 16 (6,3%) de las muestras, aislándose C. jejuni en 13 (5,1%) y C. coli en 3 (1,2%). Todas las cepas fueron susceptibles a gentamicina y ampicilina/ ácido clavulánico, con baja resistencia a ampicilina y eritromicina y alta resistencia a ciprofloxacina.

Introduction: Campylobacter is an important agent of diarrhea in humans. In Ecuador, the information on Campylobacter is scarce and there are not antecedents about antimicrobial susceptibility. Objective: To describe Campylobacter prevalence in children with diarrhea and their behavior against five antimicrobials in vitro. Method: We studied 253 children with diarrhea aging 7 months to 9 years who consulted for diarrhea in two hospitals in the city of Loja. Fecal samples were cultured and identification by tests by phenotypic tests and multiplex PCR. Susceptibility to 5 antibiotics was determined by the disc-diffusion method. Results: Campylobacter was found in 16 (6.3%) children, being C. jejuni the most frequent one (5.1%), followed by C. coli (1.2%). All strains were susceptible to gentamicin and ampicillin/clavulanic acid, being found low resistance to ampicillin and erythromycin and high resistance to ciprofloxacin.

Detection of CDT toxin genes in Campylobacter spp. strains isolated from broiler carcasses and vegetables in São Paulo, Brazil

Campylobacteriosis is a worldwide distributed zoonosis. One of the main virulence factors related to Campylobacter spp. in animals and humans is the cytolethal distending toxin (CDT), encoded by three adjacent genes (cdtA, cdtB, cdtC). The occurrence of Campylobacter spp. in samples of vegetables has not been reported in Brazil yet, and has seldom been described in the international literature. The detection of CDT in these strains has not been reported, either. The objectives of the present study were to determine the occurrence of Campylobacter spp. strains carrying virulence factors in samples of poultry and vegetables (lettuce and spinach) from different points of sale, thus verifying if vegetables are as an important vehicle for potentially virulent Campylobacter spp. strains as poultry. Twenty four strains were identified as Campylobacter jejuni by phenotypic and genotypic methods: 22 from broiler carcasses and two from lettuce samples. Three strains were identified as Campylobacter coli: two from broiler carcasses and one from lettuce. The presence of the cdt genes were detected in 20/24 (83.3%) C. jejuni strains, and 3/3 (100%) C. coli strains. The isolation of Campylobacter spp. strains with the cdt gene cluster in lettuce samples points to a new possible source of contamination, which could have an impact in the vegetable production chain and risk to public health. Results show that potentially virulent C. jejuni and C. coli strains remain viable in samples of broiler carcasses and vegetables at the points of sale.

Detection and species identification of Campylobacter in stool samples of children and animals from Vellore, south India.

Campylobacter spp. are an important cause of bacterial gastroenteritis frequently isolated from animal, poultry and environmental samples. In this study, we investigated the zoonotic potential of Campylobacter spp. by comparing prevalence rates and species in 394 children with diarrhoea and 652 animals in Vellore using PCR-based tools. Eighteen children (4.5%) had campylobacteriosis, a majority of whom had co-pathogens (15/18) and most were infected with Campylobacter jejuni (16/18). A few C. coli and mixed infections with both species were also seen. Among the animal samples, 16/25 chicken samples (64%) were positive and all were found to be C. jejuni.

Genotipificación y resistencia antibacteriana de cepas de Campylobacter spp aisladas en niños y en aves de corral / Genotyping and antibacterial resistance of Campylobacter spp strains isolated in children and in free range poultry

Poultry is a main reservoir and source of human infection in campylobacteriosis. Three hundred and forty one stool samples (291 human, 50 avian) were analyzed. In the human group, 220 samples were collected from children with acute diarrheal disease (183 inpatients, 37 outpatients) and 71 from healthy children. Erythromycin and ciprofloxacin agar dilution MIC tests, Penner serotyping and RAPD-PCR genotyping were performed on 23 strains isolated. C. jejuni was reported only in patients with acute diarrhea (5.4 percent inpatients, 2.2 percent outpatients). Campylobacter prevalence in poultry was 34 percent. Cross-resistance to nalidixic acid and ciprofloxacin was found in 33.3 percent of human samples and 11.8 percent of animal samples. Human samples could not be typed using the Penner method. F serotype was the most expressed in poultry. We obtained a total of 14 genotypes (4 / 5 human and 10/15 avian). In conclusion, the predominant species in poultry and humans was C. jejuni, a significant amount of quinolone-resistant human and avian samples were obtained, and avian genotypes and serotypes were not found in human samples. The latter would mean that another source of infection could exist; therefore other reservoirs must be studied.

Las aves de consumo constituyen uno de los principales reservorios y fuente de infección humana de la campilo-bacteriosis. Se analizaron 341 muestras de deposiciones, 291 humanas y 50 aviares. De las muestras, 220 de niños con síndrome diarreico agudo-SDA (183 hospitalizados y 37 consultantes ambulatorios) y 71 niños sanos. A las 23 cepas obtenidas se les realizó CIM por dilución en agar a eritromicina y ciprofloxacina, serotipificación de Penner y genotipiicación por RAPD-PCR. Se encontró Campylobacterjejuni sólo en pacientes con SDA, de ellos 5,4 por ciento ambulatorios y 2,2 por ciento hospitalizados. En aves, la prevalencia de Campylobacter spp., fue de 34 por ciento. Hubo resistencia cruzada a ácido nalidixico y ciprofloxacina en 33,3 por ciento cepas de origen humano y 11,8 por ciento animal. Las cepas humanas fueron no tipiicables por el método de Penner. Predominó entre las aves el serotipo F. Se obtuvo un total de 14 genotipos (4/5 humanos y 10/15 aviares). En conclusión, la especie predominante en aves de corral y en humanos fue C. jejuni, existiendo una alta prevalencia de cepas de origen humano y aviar resistentes a quinolonas. Los genotipos y serotipos aviares no fueron encontrados en cepas de origen humano, lo que indica que podría existir otra fuente de infección, por lo que se requiere estudiar otros reservorios.

Pathological studies on Campylobacter jejuni and C. coli isolated from diarrheic camel - calves

A total of 200 internal organs [intestine, liver, spleen and mesenteric lymph node] were collected from slaughtered camel-calves aged 14-18 months at slaughtered houses in Cairo and Giza; diarrheic camel-calves [150] and apparently healthy camel-calves [50]. All samples were investigated bacteriologically to detected Campylobacter species. The positive samples were examined histopathalogically. The results of bacteriological studies were recovered, 37 [20%] were positive for the infection with C. jejuni and C. coli, while 10 [5%] were positive for the apparent healthy cases. The isolation rate of C. jejuni in the diarrheic calves was [15.33%] and [14%] from apparently healthy cases, while the rates of isolated C. coli were 7[4.66%] and 3[6%] respectively. No isolation can be detected for both microorganisms in the mesenteric lymph node, meanwhile, the higher rate of C. jejuni and C. coli isolate was from the intestine 18[30%], 5[8.33%], then 7[23.33%], 2[6.66%] from the liver of diarrheic and apparently healthy cases. The isolate rate of C. jejuni was 5[16.66%], 2[20%] from the spleen of diarrheic and apparently healthy calves. The histopathalogical examination revealed cryptitis, destruction and degeneration of intestinal covering in the cases which affected with C. coli. Necrosis of intestinal glands, surfac erosion and proliferation of cryptal enterocytes with the presence of bacteria in the apical destructed cells at the tip of epithelial covering were detected from C. jejuni affected cases, while a marked hyperplasia of enterocytes, resulting in distortion of normal architecture of tightly packed proliferating enterocytes were noticed in C. jejuni and C. coli infected cases. The liver revealed hepatic necrosis with inflammatory cell aggregations, also the spleen showed a pronounced follicular depletion with necrosis in the cases which suffered a mixed infection of both bacteria. Finally, this study indicated that camel-calves are reservoir of human infection with Campylobacter infection. This study can provide a basis for the development of specific needs of public health inspection involved in protecting and promoting food safety

Survey of chicken abattoir for the presence of Campylobacter jejuni and Campylobacter coli

The genus Campylobacter is of great importance to public health because it includes several species that may cause diarrhea. These species may be found in water, food and in the intestinal tract of chickens. This study investigated the presence of Campylobacter jejuni and Campylobacter coli in chicken abattoirs in São Paulo State, Brazil. A total of 288 samples of feces, feathers, scald water, evisceration water, chiller water, and the rinse water of eviscerated, not eviscerated and chilled carcasses were collected in six chicken abattoirs. Polymerase Chain Reaction (PCR) was performed in Campylobacter spp.-positive isolates using the gene HIP, specific for hippuricase enzyme from Campylobacter jejuni and aspartokinase gene, specific to detect Campylobacter coli. The percentage of positive isolates of Campylobacter jejuni was 4.9 percent (14/288). Isolation was greater in feces samples (22 percent, 8/36). One sample was positive for the species C. coli. In conclusion, the results indicate that it is necessary to improve quality control for Campylobacter spp. in chicken abattoirs.

O gênero Campylobacter tem grande destaque em saúde pública, principalmente por pertencerem a este gênero várias espécies que podem causar diarréia. Estas espécies podem ser encontradas em amostras de água, alimentos e no trato intestinal das aves. Este estudo investigou a presença de Campylobacter jejuni e Campylobacter coli em abatedouros de aves no Estado de São Paulo. As 288 amostras foram coletadas em seis estabelecimentos e incluíram: fezes; penas; água de escaldamento, de evisceração e de resfriamento; e água de enxaguadura de carcaça não eviscerada, eviscerada e resfriada. Após o isolamento microbiológico das amostras positivas de Campylobacter spp. foi realizada uma Reação em Cadeia da Polimerase (PCR) utilizando o gene HIP, da hipuricase, específico para Campylobacter jejuni e o gene da enzima aspartoquinase, específico para Campylobacter coli. A porcentagem de amostras positivas para Campylobacter spp. foi de 4,9 por cento (14/288), sendo que o isolamento foi maior em amostras de fezes (22 por cento, 8/36). Foi isolada uma amostra positiva para C. coli. Em conclusão, os resultados indicam que há uma necessidade de melhorar a qualidade higiênico-sanitária do controle de Campylobacter em abatedouros de aves.

Molecular characterization of Campylobacter isolated from chickens and humans in northern Thailand.

A study was conducted in northern Thailand to characterize Campylobacter spp isolated from chickens at farms, slaughterhouse, and chicken meat at fresh markets, and stools from chicken farmers as well as neighboring crop farmers who served as controls. The Campylobacter isolates were collected during the rainy months of the years 2000, 2001, and 2002. Standard methods were used for primary isolation and identification of Camplobacter and the resulting isolates were frozen and stored in 30% glycerol with Mueller-Hinton broth at -70 degrees C until used in 2003. A multiplex PCR assay was used for differentiation of the Campylobacter spp. A total of 415 Campylobacter spp were isolated from 849 (48.9%) samples from chickens at the farm, slaughter house and fresh chicken meat market. Campylobacter spp were isolated from 5 of the 129 (3.9%) chicken farmers but none from the 100 neighboring crop farmers. C. jejuni was the most prevalent (42.5%) at the farm, followed by C. coli (39.1%) and other species (8.0%). In contrast, C. coli was the most prevalent at the slaughter house (72.4%) while C. jejuni was only 17.2% and others 3.4%. Similarly, at the fresh chicken market, C. coli was the most prevalent (54.4%) while C. jejuni was 26.5% and others were 13.2%. Campylobacter spp isolated from the chicken farmers were predominantly (75%) C. coli and the rest (25%) were C. jejuni. The results of the study show that both C. coli and C. jejuni are highly prevalent in chickens, along the chicken production system and in chicken farmers in northern Thailand. Critical control points for exposure and contamination of the chicken meat supply should be identified so that methods can be developed to protect human exposure to Campylobacter spp.

Diversity of Campylobacter isolates from three activated sludge systems

Thermophilic campylobacters were isolated from three sewage plants in Rio de Janeiro, RJ, Brazil, and identified. Laboratory analysis of 390 sewage samples showed the presence of 169 thermophilic strains. The results demonstrated that human and animal pathogenic biotypes could be isolated from activated sludge during the initial processing steps. The aeration tank could be considered a barrier to Campylobacter survival. C. jejuni was the prevalent species isolated (40.8 per cent). The most common biotypes were C. jejuni biotype I (21.3 per cent), C. coli biotype I (16 per cent) and C. jejuni biotype II (14.8 per cent).

Characterization of campylobacter spp. using restriction fragment length polymorphism and SDS-polyacrylamide gel electrophoresis

Twenty four Campylobacter jejuni and coli isolates obtained from Egyptian children were characterized using restriction fragment length polymorphism [RFLP] of flagellin genes and polyacrylamide gel electrophoresis of whole cell and glycine-extracted proteins. The isolates were found to fall into nine polymorphism groups, eight of which were reported previously in Egypt but one group displayed by 3 isolates represented a new group that was not reported before. Furthermore, the relative prevalence of polymorphic groups in the population studied is different from that reported previously. Analysis of whole-cell and acid glycine-extracted proteins showed that the profiles of these isolates are typical profiles of Campylobacters isolated from other humans

Actividades enzimáticas de Campylobacter jejuni, C. coli y C. lari / Enzymatic activities in Campylobacter jejuni, C. coli and C. lari

Clásicamente la prueba de sensibilidad al ácido nalidíxico se ha considerado una técnica útil en la identificación de miembros del género Campylobacter. Sin embargo, el progresivo incremento de resistencias adquiridas ha creado la necesidad de incluir en la rutina convencional de laboratorio otras pruebas de identificación. Hemos empleado el sistema Api ZYM (bioMérieux) para determinar la actividad enzimática de 180 cepas de Campylobacter spp, aisladas a partir de muestras clínicas, y evaluar su utilidad en la caracterización de estos microorganismos. De las 19 actividades enzimáticas detectables por el sistema; 13 enzimas (lipasa-C14, valina arilamidasa, cistina arilamidasa, tripsina, alfa-quimiotripsina, alfa-galactosidasa, beta-galactosidasa, beta-glucoridasa, alfa-glucosidasa, beta-glucosidasa, N-acetil-beta-glucosaminidasa, alfa-manosidasa y alfa-fucosidasa) fueron uniformemente negativas en todas las cepas estudiadas y 3 enzimas (fosfatasas alcalina, fosfatasa ácida y naftol-A-S-BI-fosfohidrolasa) estuvieron presentes en el 96,6 o/o. Las 3 reacciones restantes (esterasa-C1, esterasa lipasa-C8 y leucina arilamidasa) presentaron una reactividad variable dependiendo de las cepas estudiadas. Aunque el patrón enzimático detectado a nivel de género Campylobacter fue bastante homogéneo, se observaron diferencias entre las distintas especies. Todas las cepas C. lari evidenciaron un perfil enzimático diferente al observado en C. jejuni y C. coli, las cuales no mostraron diferencias significativas entre ellas. El sistema Api ZYM se ha mostrado como un método sencillo, fácil y complementario en los esquemasj de caracterización de las campilobacterias

Byotipe and lior's serogroup of Campylobacter jejuni and Campylobacter coli isolated in Säo Paulo, Brazil

Foi avaliada a frequência de Campylobacter sp no período 1983 a 1989, quanto à sua distribuiçäo por grupo etário e ao percentual em relaçäo a outros enteropatógenos. As cepas isoladas foram classificadas em biotipos e sorogrupos, segundo as metodologias descritas por Lior. Foram utilizados 46 anti-soros preparados com cepas de referências recebidas do "National Reference Service for Campylobacter" do Canadá. Neste período foram isoladas 285 cepas de Campilobacter sp., correspondendo a 3,72 por cento de C. jejuni e 11,93 por cento de C. coli, das quais 62,81 por cento de C. jejuni por cento foram isoladas de crianças com menos de 2 anos de idade. Em 67 vezes (23,51), C. jejuni e C. coli foram isolados em infecçöes mistas. Houve prevalência do C. jejuni biotipo II com 24,22 por cento e C. coli biotipo I com 9,76 por cento. Foram sorotipadas 84,94 por cento das cepas isoladas, havendo predominância dos sorogrupos; 22 cepas (10,04 por cento) no puderam ser tipadas com os 46 anti-soros utilizados e outras 11 (5,02 por cento) se apresentaram autoaglutinantes

Biotipificación y serotipificación de campylobacter jejuni, acmpylobacter coli y campylobacter lari / Biotyping and sertyping of campylobacter jejuni, campylobacter coli and campylobacter lari

Se serotipificaron 26 cepas de Campylobacter termofílicos por el método de hemaglutinación pasiva y se biotipificaron 62 cepas por el método de Lior. Dos cepas correspondieron al serogrupo 13/50 y otras dos expresaron antígenos del serogrupo 4; el 78 por ciento restante se distribuyó entre diferentes serogrupos; 21,7 por ciento de las cepas fueron no tipificables. Entre los biotipos, 47 (75,8 por ciento) correspondieron a C. jejuni, 44 (71 por ciento) al biotipo I, 3 (4,8 por ciento) al biotipo II; 14 (22,6 por ciento) a C. coli, 11 (17,8 por ciento) al biotipo I, 3 (4,8 por ciento) al biotipo II y 1 (1,6 por ciento) a C. lari. Las cepas de C. jejuni y C. coli resistentes al ácido nalidíxico pueden complicar la identificación

Avaliação do método da passivação do cobre para a obtenção de microaerofilia e do ágar cefoxitina no cultivo de Campylobacter jejuni e Campylobacter coli / Evaluation of cefoxitin mediun and the microaerophilic environment produced by a combination of iron, copper and sodium bicarbonate for the culture of Campylobacter jejuni e Campylobater coli

Foi realizado um estudo sobre o efeito que os meios ágar sangue cefoxitina, ágar Skirrow e ágar Campy-BAP e os métodos de obtenção de microaerofilia que empregam os princípios da passivação do cobre, de envelopes geradores de gases (anaerocult C - Merck) e da evacuação do ar atmosférico e substituição por uma mistura de gases, exercem sobre o crescimento de estirpes de Campylobacter jejuni e Campylobacter coli, utilizando-se culturas puras e suspensões obtidas a partir da água de lavagem de 34 carcaças de frangos. Não houve diferença estatisticamente significativa entre os métodos de obtenção de microaerofilia no cresciemtno das amostras testadas, assim como no isolamento primário de Campylobacter termofílicos a partir de carcaças de frangos. O meio ágar sangue cefoxitina e o método de obtenção de microaerofilia da passivação do cobre apresentaram um bom rendimento, constituindo-se numa alternativa prática e de baixo custo para o isolamento destes microrganismos em laboratórios de médios e pequeno porte.