Quantification and characterization of Salmonella spp. isolates in sewage sludge with potential usage in agriculture

Abstract\ud \ud Background\ud This study aims to scrutinize Salmonella spp. and its serotypes in sewage sludge samples from wastewater treatment plants, and assesses the presence of virulence genes and antibiotics resistant to the profile. Samples (n = 54) were collected and analyzed in accordance with the EPA Method 1682/2006. For positive serological reaction, 40 strains were selected for PCR analyses and detection of spvC, invA and sseL virulence genes, plasmid presence and resistance to antibiotics.\ud \ud \ud Results\ud \ud Salmonella spp. was detected in 38.9% of the samples collected (<0.006473 to 12.19 MPN/gTS). The most prevalent serotype was Salmonella Infantis. All Salmonella spp. (n = 35) presented at least one of the three virulence genes mentioned above and 40% harboured plasmids. Salmonella Typhimurium strains were isolated harbouring at least one of the following virulence genes: spvC, invA or sseL. Four Salmonella spp. isolates were resistant to tetracycline; three were resistant to trimethoprim-sulfamethoxazole, and one isolate was resistant to ciprofloxacin. Two Salmonella spp. strains presented multi resistance to antimicrobial agents.\ud \ud \ud Conclusions\ud The results obtained demonstrated that Salmonella spp. have been found in sewage sludge, thus it is essential to set measures to mitigate human health risks when it is intended to be applied on agricultural soils.We would like to give thanks to the Coordination of Improvement of Higher\ud Education– CAPES– for providing us with scholarships and also to Fundação\ud de Amparo à Pesquisa do Estado de São Paulo– FAPESP– for its financial\ud support (2010/05664-6)

Molecular epidemiology of Listeria monocytogenes isolated from different sources in Brazil

Listeria monocytogenes is an important foodborne pathogen that primarily affects pregnant women, neonates, the elderly and immune-compromised individuals, and it may cause abortion, septicemia, and meningitis. From the 13 capsular groups described, serotypes 4b, 1/2b and 1/2a are most closely related to human infection. For this reason, serotyping has limited value as an epidemiological tool; thus, improved discriminatory typing methods are required to enhance knowledge of L. monocytogenes contamination and infection. The aim of this study was to characterize the genetic diversity of L. monocytogenes isolates in the pork processing industry in Sao Paulo, Brazil and human infection isolates by ERICPCR and single enzyme AFLP. Serotypes 1/2c and 4b were frequent among isolates from pork and slaughterhouse/market environments, whereas serotypes 4b and 1/2a were observed among human isolates. ERIC-PCR and AFLP revealed 34 and 31 distinct profiles, respectively, which had tendencies of separation according to serogroup and isolate origin. The genetic profiles from slaughterhouse and market environments suggest the possibility of different sources of Listeria contamination in the environment, although in certain cases, continuous contamination caused by the persistence of clonal strains is also a possibility.Listeria monocytogenes é um importante patógeno de origem alimentar que afeta principalmente grávidas, neonatos, idosos e indivíduos imunocomprometidos, e pode causar abortamento, septicemia e meningite. Dos 13 grupos capsulares descritos, os sorotipos 4b, 1/2b e 1/2a são os mais relacionados à infecção humana. Por esta razão, a sorotipagem possui valor limitado como ferramenta epidemiológica e, dessa forma, métodos mais discriminatórios são necessários para melhorar o conhecimento sobre a contaminação e a infecção por L. monocytogenes. O objetivo deste estudo foi caracterizar a diversidade genética de isolados de L. monocytogenes da indústria de processamento de carne suína no Estado de São Paulo, Brasil, e compará-los a isolados de casos de infecção humana através do ERIC-PCR e AFLP com uma única enzima. Os sorotipos 1/2c e 4b foram frequentes em carne suína e ambientes de abatedouros e mercados, enquanto os sorotipos 4b e 1/2a foram observados nos isolados de humanos. ERIC-PCR e AFLP resultaram em 34 e 31 perfis distintos, respectivamente, com uma tendência a separar de acordo com o sorogrupo e a origem do isolado. Os perfis genéticos de ambiente dos abatedouros e mercados sugerem a possibilidade de diferentes origens de contaminação por Listeria nos ambientes estudados, porém, em alguns casos, é possível que ocorra a persistência de cepas clonais causando contaminação contínua

Characterization of antibiotic resistance in Listeria spp. isolated from slaughterhouse environments, pork and human infections

Introduction: Listeria species are susceptible to most antibiotics. However, over the last decade, increasing reports of multidrug-resistant Listeria spp. from various sources have prompted public health concerns. The objective of this study was to characterize the antibiotic susceptibility of Listeria spp. and the genetic mechanisms that confer resistance. Methodology: Forty-six Listeria spp. isolates were studied, and their minimal inhibitory concentrations of antibiotics were determined by microdilution using Sensititre standard susceptibility MIC plates. The isolates were screened for the presence of gyrA, parC, lde, lsa(A), lnu(A), and mprF by PCR, and the amplified genes were sequenced. Results: All isolates were susceptible to penicillin, ampicillin, tetracycline, erythromycin, and carbapenems. Resistance to clindamycin, daptomycin, and oxacillin was found among L. monocytogenes and L. innocua, and all species possessed at least intermediate resistance to fluoroquinolones. GyrA, parC, and mprF were detected in all isolates; however, mutations were found only in gyrA sequences. A high daptomycin MIC, as reported previously, was observed, suggesting an intrinsic resistance of Listeria spp. to daptomycin. Conclusions: These results are consistent with reports of emerging resistance in Listeria spp. and emphasize the need for further genotypic characterization of antibiotic resistance in this genusFAPESP, 2010/19005-4CAPE

Mucopolysaccharidosis I, II, and VI: Brief review and guidelines for treatment

Mucopolysaccharidoses (MPS) are rare genetic diseases caused by the deficiency of one of the lysosomal enzymes involved in the glycosaminoglycan (GAG) breakdown pathway. This metabolic block leads to the accumulation of GAG in various organs and tissues of the affected patients, resulting in a multisystemic clinical picture, sometimes including cognitive impairment. Until the beginning of the XXI century, treatment was mainly supportive. Bone marrow transplantation improved the natural course of the disease in some types of MPS, but the morbidity and mortality restricted its use to selected cases. The identification of the genes involved, the new molecular biology tools and the availability of animal models made it possible to develop specific enzyme replacement therapies (ERT) for these diseases. At present, a great number of Brazilian medical centers from all regions of the country have experience with ERT for MPS I, II, and VI, acquired not only through patient treatment but also in clinical trials. Taking the three types of MPS together, over 200 patients have been treated with ERT in our country. This document summarizes the experience of the professionals involved, along with the data available in the international literature, bringing together and harmonizing the information available on the management of these severe and progressive diseases, thus disclosing new prospects for Brazilian patients affected by these conditions

Not available

No presente estudo 64 amostras de água de superficie e 24 amostras de sedimento provenientes da Represa de Guarapiranga, SP, foram submetidas à técnica dos tubos múltiplos para a pesquisa de Aeromonas spp. As amostras eram submetidas a enriquecimento com água peptonada alcalina (APA), adicionado de 1 por cento de cloreto de sódio, pH 8,6 e isoladas em agar sangue contendo 10 mg/1 de ampicilina. As colônias que se apresentavam hemolíticas ou não, eram submetidas a triagem pelos testes de oxidase e agar ferro de Kligler (Difco) e confirmadas através de provas bioquímicas complementares. Foram isoladas A. hydrophila em 18,7 por cento das amostras, A. caviae em 31,2 por cento, A. sobria em 43,7 por cento e A. jandaei em 76,6 por cento. Não foi observada a correlação das espécies isoladas com temperatura da água, pH da amostra ou contagens de indicadores de contaminação fecal. As maiores contagens de Aeromonas spp foram encontradas em amostras de sedimento. Do total de cepas isoladas, 15 de A. hydrophila, 15 de Aromonas caviae, 15 de A. sobria e 15 de A. jandaei foram submetidas ao teste de alça ligada em coelhos e teste de hemólise em ágar sangue de coelho e humano. A pesquisa de fatores de virulência revelou que 80 por cento das cepas de A. hydrophila, 73,3 por cento de A. caviae, 46,7 por cento de A. sobria e 86,6 por cento de A. jandaei causaram acúmulo de fluido em alça ligada de coelho e que 86,6 por cento de A. hydrophila, 33,3 por cento de A. caviae, 60 por cento de A. sobria e 100 por cento de A. jandaei foram produtoras de beta-hemólise em ágar sangue, confirmando o elevado potencial patogênico desses microrganismos. Considerando os resultados obtidos pode-se concluir que cepas de Aeromonas isoladas de ambiente aquático são capazes de produzir fatores de virulência que constituem um risco à saúde da população.In this work, 64 superfitial water samples and 24 of sedment, were colected from Guarapiranga dam in the State of São Paulo, Brazil, and were analised to the presence of Aeromonas spp. Enrichment of samples were made in Alcaline Peptone Water (APW) with sodium chloride 1 per cent pH 8,6. Blood Agar Ampicilin (10/gl) were used as isolation medium. Colonies that were hemolytic or not, were submetted to oxidase test and Kligler lron Agar (difco), and were confirmed through complementar biochemical tests. A. hydrophila were isolated in 18, 7 per cent of the samples, A. caviae in 31,2 per cent A. sobria in 43,7 per cent and A. jandaei in 76,6 per cent. No correlation was observed between Aeromonas sapecies neither with temperature or pH, nor with fecal contaminantion indicators. The highests values of Aeromonas spp, were observed in sedment samples. From the total of strains that were isolated, 15 of A. hydrophila, 15 A. caviae, 15 A. sobria and 15 of A. jandaei. were submetted to the rabbit ileal loop assay and to the hemolysis prodution on Blood agar with human and rabbit erytrocits. The virulence factors reseach revealed that 80 per cent of A. hydrophila, 73,3 per cent of A. caviae, 46,7 of A. sobria and 86,8 per cent of the A. jandaei were able to cause fluid accumulation in rabbit ileal loop, and, 86,6 per cent of A. hydrophila strains, 33,3 per cent of A. caviae, 60 per cent of A. sobria and 100 per cent of A. jandaei produced beta-hemolisis in blood agar. these results confirms the pathogenic potential of these microorganisms. Considering the given results, it was concluded that Aeromonas strains, isolated from this environment are able to produce virulence factors that are considered a risk to public health

Not available

No presente estudo 64 amostras de água de superficie e 24 amostras de sedimento provenientes da Represa de Guarapiranga, SP, foram submetidas à técnica dos tubos múltiplos para a pesquisa de Aeromonas spp. As amostras eram submetidas a enriquecimento com água peptonada alcalina (APA), adicionado de 1 por cento de cloreto de sódio, pH 8,6 e isoladas em agar sangue contendo 10 mg/1 de ampicilina. As colônias que se apresentavam hemolíticas ou não, eram submetidas a triagem pelos testes de oxidase e agar ferro de Kligler (Difco) e confirmadas através de provas bioquímicas complementares. Foram isoladas A. hydrophila em 18,7 por cento das amostras, A. caviae em 31,2 por cento, A. sobria em 43,7 por cento e A. jandaei em 76,6 por cento. Não foi observada a correlação das espécies isoladas com temperatura da água, pH da amostra ou contagens de indicadores de contaminação fecal. As maiores contagens de Aeromonas spp foram encontradas em amostras de sedimento. Do total de cepas isoladas, 15 de A. hydrophila, 15 de Aromonas caviae, 15 de A. sobria e 15 de A. jandaei foram submetidas ao teste de alça ligada em coelhos e teste de hemólise em ágar sangue de coelho e humano. A pesquisa de fatores de virulência revelou que 80 por cento das cepas de A. hydrophila, 73,3 por cento de A. caviae, 46,7 por cento de A. sobria e 86,6 por cento de A. jandaei causaram acúmulo de fluido em alça ligada de coelho e que 86,6 por cento de A. hydrophila, 33,3 por cento de A. caviae, 60 por cento de A. sobria e 100 por cento de A. jandaei foram produtoras de beta-hemólise em ágar sangue, confirmando o elevado potencial patogênico desses microrganismos. Considerando os resultados obtidos pode-se concluir que cepas de Aeromonas isoladas de ambiente aquático são capazes de produzir fatores de virulência que constituem um risco à saúde da população.In this work, 64 superfitial water samples and 24 of sedment, were colected from Guarapiranga dam in the State of São Paulo, Brazil, and were analised to the presence of Aeromonas spp. Enrichment of samples were made in Alcaline Peptone Water (APW) with sodium chloride 1 per cent pH 8,6. Blood Agar Ampicilin (10/gl) were used as isolation medium. Colonies that were hemolytic or not, were submetted to oxidase test and Kligler lron Agar (difco), and were confirmed through complementar biochemical tests. A. hydrophila were isolated in 18, 7 per cent of the samples, A. caviae in 31,2 per cent A. sobria in 43,7 per cent and A. jandaei in 76,6 per cent. No correlation was observed between Aeromonas sapecies neither with temperature or pH, nor with fecal contaminantion indicators. The highests values of Aeromonas spp, were observed in sedment samples. From the total of strains that were isolated, 15 of A. hydrophila, 15 A. caviae, 15 A. sobria and 15 of A. jandaei. were submetted to the rabbit ileal loop assay and to the hemolysis prodution on Blood agar with human and rabbit erytrocits. The virulence factors reseach revealed that 80 per cent of A. hydrophila, 73,3 per cent of A. caviae, 46,7 of A. sobria and 86,8 per cent of the A. jandaei were able to cause fluid accumulation in rabbit ileal loop, and, 86,6 per cent of A. hydrophila strains, 33,3 per cent of A. caviae, 60 per cent of A. sobria and 100 per cent of A. jandaei produced beta-hemolisis in blood agar. these results confirms the pathogenic potential of these microorganisms. Considering the given results, it was concluded that Aeromonas strains, isolated from this environment are able to produce virulence factors that are considered a risk to public health

Ribotyping of Aeromonas hydrophila, Aeromonas caviae, Aeromonas sobria, and Aeromonas jandaei, potentially pathogenic, isolated from water samples Guarapiranga Reservoir, São Paulo

Neste estudo 60 cepas de Aeromonas, 15 A. hydrophila, 15 A. caviae, 15 A. sobria e 15 A. jandaei, isoladas de 5 diferentes pontos do reservatório de Guarapiranga, São Paulo, e previamente testada quanto à produção de fatores de virulência, acúmulo de fluído em alça ligada e hemólise em ágar sangue, foram submetidas a ribotipagem e a análise do perfil plasmidial. Cada cepa apresentou um perfil de ribotipagem diferente tendo-se observado para as espécies A. hydrophila e A. caviae a diferenciação em 3 agrupamentos, e A. sobria e A. jandaei dois agrupamentos cada. A análise do perfil plasmidial demonstrou que 13,4 por cento das A. hydrophila apresentaram um ou no máximo 2 plasmídios, enquanto 33,3 por cento das A. sobria e 53,3 por cento das A. jandaei apresentaram de 1 a 6 plasmídios para cada espécie; A. caviae não apresentou cepas contendo plasmídios. Não foi observada correlação entre a presença de plasmídios e a produção de fatores de virulência pelas cepas estudadas. A ribotipagem demonstrou haver um polimorfismo genômico dentro de uma mesma espécie de Aeromollas e, ainda, diferenciou cepas isoladas de um mesmo ponto de amostragem. Estas metodologias, ribotipagem e análise do perfil plasmidial, apresentam em geral características que são complementares, demonstrando ser ferramentas importantes a serem empregadas, tanto em estudos epidemiológicos como ecológicos.In this work 60 Aeromonas strains, 15 A. hydrophila, 15 A. caviae, 15 A. sobria and 15 A. jandaei isolated from 5 different points of Guarapiranga Dam, São Paulo, and previously tested for virulence factors production (ileal loop assay and hemolysis on blood agar) were submitted to ribotyping and plasmidial profiles analysis. Each strain showed a different ribopattern and there were observed that for A. hydrophila and A. caviae each specie were grouped in 3 ribotypes, A. sobria and A. jandaei in 2 ribotype each. Plasmidial profiles analysis demonstrated that 13,4 per cent af A. hydrophila had at least one but no more than 2 plasmids, 33,3 per cent of A. sobria and 53,3 per cent af A. jandaei had from one to 6 plasmids each, and A. caviae didn\'t show to have any plasmids. There were not observed correlation between presence of plasmids and virulence factor production. Ribotyping showed that there are genomic polymorphism within the same Aeromonas specie and differentiate strains that were isolated from the same sample point, indicating that those methodologies have in general characteristics that are complementary and are important tools to be used either in epidemiological or ecological studies

Ribotyping of Aeromonas hydrophila, Aeromonas caviae, Aeromonas sobria, and Aeromonas jandaei, potentially pathogenic, isolated from water samples Guarapiranga Reservoir, São Paulo

Neste estudo 60 cepas de Aeromonas, 15 A. hydrophila, 15 A. caviae, 15 A. sobria e 15 A. jandaei, isoladas de 5 diferentes pontos do reservatório de Guarapiranga, São Paulo, e previamente testada quanto à produção de fatores de virulência, acúmulo de fluído em alça ligada e hemólise em ágar sangue, foram submetidas a ribotipagem e a análise do perfil plasmidial. Cada cepa apresentou um perfil de ribotipagem diferente tendo-se observado para as espécies A. hydrophila e A. caviae a diferenciação em 3 agrupamentos, e A. sobria e A. jandaei dois agrupamentos cada. A análise do perfil plasmidial demonstrou que 13,4 por cento das A. hydrophila apresentaram um ou no máximo 2 plasmídios, enquanto 33,3 por cento das A. sobria e 53,3 por cento das A. jandaei apresentaram de 1 a 6 plasmídios para cada espécie; A. caviae não apresentou cepas contendo plasmídios. Não foi observada correlação entre a presença de plasmídios e a produção de fatores de virulência pelas cepas estudadas. A ribotipagem demonstrou haver um polimorfismo genômico dentro de uma mesma espécie de Aeromollas e, ainda, diferenciou cepas isoladas de um mesmo ponto de amostragem. Estas metodologias, ribotipagem e análise do perfil plasmidial, apresentam em geral características que são complementares, demonstrando ser ferramentas importantes a serem empregadas, tanto em estudos epidemiológicos como ecológicos.In this work 60 Aeromonas strains, 15 A. hydrophila, 15 A. caviae, 15 A. sobria and 15 A. jandaei isolated from 5 different points of Guarapiranga Dam, São Paulo, and previously tested for virulence factors production (ileal loop assay and hemolysis on blood agar) were submitted to ribotyping and plasmidial profiles analysis. Each strain showed a different ribopattern and there were observed that for A. hydrophila and A. caviae each specie were grouped in 3 ribotypes, A. sobria and A. jandaei in 2 ribotype each. Plasmidial profiles analysis demonstrated that 13,4 per cent af A. hydrophila had at least one but no more than 2 plasmids, 33,3 per cent of A. sobria and 53,3 per cent af A. jandaei had from one to 6 plasmids each, and A. caviae didn\'t show to have any plasmids. There were not observed correlation between presence of plasmids and virulence factor production. Ribotyping showed that there are genomic polymorphism within the same Aeromonas specie and differentiate strains that were isolated from the same sample point, indicating that those methodologies have in general characteristics that are complementary and are important tools to be used either in epidemiological or ecological studies

OCCURRENCE OF PATHOGENIC MICROORGANISMS IN FISH SOLD IN SAO PAULO, BRAZIL

This study investigated the presence of potentially human pathogenic strains of Vibrio spp., Aeromonas spp., Escherichia coli, Salmonella spp. and Staphylococcus aureus in fish commercialized in street markets of Sao Paulo city, Brazil. Twenty fish of different species were analyzed for foodborne pathogens using conventional methods. High levels of fecal contamination were detected in 25% of samples. S. aureus was isolated from 10% of samples. All were negative for Salmonella. Vibrio species, including Vibrio cholerae non-O1/non-O139, were observed in 85% of samples although Vibrio parahaemolyticus was not found in this study. Aeromonas spp., including A. hydrophila, was isolated from 50% of fish samples. The occurrence of these pathogens suggests that the fish commercialized in Sao Paulo may represent a health risk to the consumers.CECOVISA/USP - Centro Colaborador em Vigilancia Sanitaria/FSP[CA 06/99-44-ANVS/MS