Complex class 1 integrons harboring CTX-M-2-encoding genes in clinical Enterobacteriaceae from a hospital in Brazil.

INTRODUCTION: CTX-M enzymes are the most prevalent extended-spectrum beta-lactamases (ESBLs) in Brazil and around the world. The spread of CTX-M lies in their ability to be mobilized by insertion sequences and integrons. This study aimed to identify the mobile genetic structures associated with bla(CTX-M) genes from clinical Enterobacteriaceae strains. METHODOLOGY: Twenty-eight clinical non-clonal Enterobacteriaceae were screened by PCR for the presence of bla(CTX-M) genes and class 1 integrase (int1), and for the association of bla(CTX-M) with class 1 integrons. Plasmid incompatibility groups were assessed by PBRT. Wild-type plasmids were transformed into electrocompetent E. coli, and the S1-PFGE technique was used to verify the presence of high-molecular-weight plasmids in both wild-type strains and E. coli transformants. RESULTS: Sequencing showed that strains carried bla(CTX-M-2) (n = 25) and bla(CTX-M-59) (n = 3) genes inserted into the 3'-end of complex class 1 integrons. Thirteen strains also carried bla(TEM) and bla(SHV) genes. CTX-M-2/59-containing complex class 1 integrons were also present in E. coli transformants. The most frequent Inc groups were IncA/C (n = 10) and IncF (n = 8). Heavy plasmids were observed in both wild-type strains and E. coli transformants. CONCLUSIONS: The presence of the same bla(CTX-M-2-group)-containing genetic structure in seven Enterobacteriaceae species isolated at seven hospital wards shows the great mobility potential of complex class 1 integrons. Also, this is the first report of TEM-15, SHV-45, and SHV-55 in Latin America. The genetic environment of bla(CTX-M-2) accounts for their maintenance and spread among Gram-negative bacteria.

Molecular and biochemical characterization of CTX-M-131, a natural Asp240Gly variant derived from CTX-M-2, produced by a Providencia rettgeri clinical strain in São Paulo, Brazil.

CTX-M-131 is a natural Asp240Gly variant from the CTX-M-2 group detected in a Providencia rettgeri clinical strain from Brazil. Molecular analysis showed that blaCTX-M-131 was inserted in a complex class 1 integron harbored by a 112-kb plasmid, which has not been previously described as a platform for CTX-M-encoding genes with the Asp240Gly mutation. Steady-state kinetic parameters showed that the enzyme has a typical cefotaximase catalytic profile and an enhanced activity against ceftazidime.

An overview of antimicrobial resistance and its public health significance.

Multiple papers have been published regarding the bacterial resistance theme over the last years. A variety of information has reached general and scientific public, daily bringing up data on new resistant microorganisms, new drugs, outbreaks, epidemiological news, resistance gene dissemination, and the lack of information in a particular field has caught our attention: the public health department. Most of researchers, physicians and government employees interpret the public health field as a separate department, not linked to this antibiotic resistance era that we are living nowadays. In this paper we carefully tried to fill in the blanks between public health and the bacteria resistance issue, also considering historical, social, economical and biological problematic that come with this possible pre-antibiotic era.

Resistência a tetraciclinas em isolados clínicos e ambientais de Escherichia coli, Klebsiella pneumoniae e Aeromonas spp: identificação e mapeamento do ambiente genético de genes tet / Tetracycline resistance in clinical and environmental isolates of Escherichia coli, Klebsiella pneumoniae and Aeromonas spp. identification and mapping of tet genes genetic context

A resistência bacteriana a antibióticos é aceita como um dos maiores problemas de saúde pública. As tetraciclinas são antibióticos de amplo espectro, e após seu uso indiscriminado observou-se o surgimento de bactérias resistentes, levando médicos e veterinários a diminuírem seu uso. Objetivos. Verificar o perfil de sensibilidade a tetraciclinas em isolados clínicos e ambientais de Escherichia coli, Klebsiella pneumoniae e Aeromonas spp., bem como pesquisar os principais genes tet associados à resistência a esta classe de antibióticos e determinar a potencial forma de disseminação destes genes através da caracterização de seu ambiente genético. Material e Métodos. Os perfis de sensibilidade à tetraciclina (TET), doxiciclina (DOX), minociclina (MIN) e tigeciclina (TGC) de 572 isolados foram obtidos através das técnicas de Disco-Difusão e Concentração Inibitória Mínima. Os isolados não-sensíveis à tetraciclina foram submetidos a reações de PCR para pesquisa de grupos Inc, genes tet e para a caracterização de seu ambiente genético pela pesquisa das integrases de classes 1, 2, 3 e 4, e dos elementos genéticos móveis Tn1721, IS26, Tn10 e ISAS5. Perfis de similaridade genética dos isolados foram obtidos através das técnicas de ERIC-PCR e PFGE. Após análise destes resultados 33 cepas foram selecionadas para as técnicas de S1-PFGE e transformação. Resultados. A partir dos 572 isolados 18,5 por cento foram resistentes à TET, 13,5 por cento à DOX, 8 por cento à MIN...

The antibiotic resistance is accepted as one of the major problems for public health. Tetracyclines are broad spectrum antibiotics, and its indiscriminate use promoted the emergence of resistant bacteria, leading physicians and veterinarians to decrease its use. Objectives. Verify the susceptibility of clinical and environmental isolates of Escherichia coli, Klebsiella pneumoniae and Aeromonas spp. to tetracyclines, and also search for the main tet genes associated with resistance to these antibiotics and determine the potential mechanism of tet genes dissemination by characterizing their genetic context. Material and Methods. Disk-Diffusion and Minimum Inhibitory Concentration tests were carried out in 572 isolates using tetracycline (TET), doxycycline (DOX), minocycline (MIN) and tigecycline (TGC). PCR was carried out in TET non-susceptible isolates for the detection of Inc groups, tet genes and its genetic context determination through the search of classes 1, 2, 3, and 4 integrases, and Tn1721, Tn10, IS26 and ISAS5 mobile genetic elements. Genetic similarities patterns were determined by ERIC-PCR and PFGE techniques. After analyzing the results 33 strains were selected for the S1-PFGE and transformation experiments. Results. From 572 isolates, 18.5 per cent were TET-resistant, 13.5 per cent DOX-resistant, 8 per cent MIN-resistant and none resistant to TGC. Twenty-two per cent and 16.3 per cent of clinical and environmental isolates were TET-resistant...

An overview of antimicrobial resistance and its public health significance

Multiple papers have been published regarding the bacterial resistance theme over the last years. A variety of information has reached general and scientific public, daily bringing up data on new resistant microorganisms, new drugs, outbreaks, epidemiological news, resistance gene dissemination, and the lack of information in a particular field has caught our attention: the public health department. Most of researchers, physicians and government employees interpret the public health field as a separate department, not linked to this antibiotic resistance era that we are living nowadays. In this paper we carefully tried to fill in the blanks between public health and the bacteria resistance issue, also considering historical, social, economical and biological problematic that come with this possible pre-antibiotic era.

Presence of blaTEM-116 gene in environmental isolates of Aeromonas hydrophila and Aeromonas jandaei from Brazil

It is known that Aeromonas spp. possess different chromosomal â-lactamase genes. Presence and phenotypic expression of blaTEM, blaSHV, and blaCTX-M ESBL-encoding genes were investigated in environmental water isolates of Aeromonas hydrophila and Aeromonas jandaei. Presence of blaSHV and blaCTX-M genes was not observed, and blaTEM gene was verified in 91 percent of the isolates. Sequencing of 10 fragments showed the occurrence of blaTEM-116.

A modified method for detecting Cryptosporidium oocysts using DNA templates extracted from environmental samples / Método modificado para detecção de oocistos de Cryptosporidium utilizando DNA extraído de amostras ambientais

The protozoan parasite Cryptosporidium has emerged as one of the most important water contaminants, causing outbreaks of waterborne diarrhea worldwide. In order to assess the importance for public health of this pathogen’s presence in environmental samples, several methods have been developed to isolate and detect Cryptosporidium oocysts. In the present study, a reliable and reproducible method has been standardized for detecting and identifying Cryptosporidium oocysts in water samples in the State of São Paulo, Brazil as the first step for future genotyping studies. Water samples were concentrated by filtration, and then subjected to ultrasound in Tween 80 0.1%, the obtained sediment was transferred into micro tubes containing 1.0 ml of distilled water and stored at -20ºC. DNA was extracted with the addition of 1% PVP in lysis buffer, the organic extraction was performed in Phase Lock GelHeavy®. There was a 214 bp amplification on the expected fragment in five out of the 11 water samples analyzed. The results of this study demonstrated the application usefulness of the standardized test in epidemiological studies and surveillance programs because the technology allowed to increase significantly the amount of amplified product.

O protozoário parasito Cryptosporidium tem emergido como um dos mais importantes contaminantes da água, causando surtos de diarreia de veiculação hídrica em todo mundo. Para avaliar o significado, para a saúde pública,da presença desse agente patogênico em amostras ambientais, vários métodos têm sido desenvolvidos para isolar e detectar oocistos de Cryptosporidium. No presente estudo foi padronizado um método confiável e reprodutível para detectar e identificar oocistos de Cryptosporidium em amostras de água no Estado de São Paulo, Brasil, comoo primeiro passo para futuros estudos de genotipagem. Amostras de água foram concentradas por filtração,submetidas a ultrasom em solução de Tween 80 a 0.1%; o sedimento obtido foi transferido para microtuboscontendo 1,0 ml de água destilada e conservado a -20ºC. O DNA foi extraído com adição de 1% de PVP no tampão de lise; a extração foi realizada em tubo Phase Lock Gel Heavy®. Houve amplificação do fragmento esperado de 214 bp em cinco das 11 amostras de água analisadas. Os resultados deste estudo demonstraram a utilidade deaplicação do teste padronizado em estudos epidemiológicos e em programas de vigilância, em virtude da técnica ter apresentado sensibilidade para incrementar significativamente a quantidade de produto amplificado.

Presence of bla TEM-116 Gene in Environmental Isolates of Aeromonas Hydrophila and Aeromonas Jandaei from Brazil.

It is known that Aeromonas spp. possess different chromosomal ß-lactamase genes. Presence and phenotypic expression of bla TEM, bla SHV, and bla CTX-M ESBL-encoding genes were investigated in environmental water isolates of Aeromonas hydrophila and Aeromonas jandaei. Presence of bla SHV and bla CTX-M genes was not observed, and bla TEM gene was verified in 91% of the isolates. Sequencing of 10 fragments showed the occurrence of bla TEM-116.

Pesquisa de genes codificadores de enterotoxinas e Beta-lactamases em Aeromonas jandaei e Aeromonas hydrophila provenientes de ambientes aquáticos / Enterotoxins and Beta-lactamases encoding genes investigation in Aeromonas jandaei e Aeromonas hydrophila from aquatic environments

O gênero Aeromonas está amplamente distribuído em ambientes aquáticos, e estudos recentes incluem o gênero no grupo de patógenos emergentes, devido à sua freqüente associação com infecções locais e sistêmicas em humanos. Este trabalho foi realizado com objetivo de pesquisar por meio da PCR e confirmar por meio de sequenciamento, a ocorrência dos genes de virulência act, alt, ast, e resistência cphA, bla(imp), bla(SPM-1), bla(CTX-M), bla(TEM), bla(SHV), verificando também o perfil de resistência a partir de antibiogramas, e a ocorrências de plasmídios nas cepas estudadas. A partir dos resultados observou-se que das 100 cepas selecionadas inicialmente, 87 pertenciam às espécies A. jandaei (46) e A. hydrophila (41). Dentre as quais pôde-se observar a ocorrência de act, alt,e ast, respectivamente em 70, 7 por cento (29), 97,6 por cento (40) e 26,8 por cento (11) das cepas de A. hydrophila, e em 4,4 por cento (2), 0 por cento (0) e 32,6 por cento (15) nas cepas de A. jandaei. Os genes bla(IMP), bla(VIM), bla(SPM-1), blaCTX-M, blaSHV não foram encontrados em nenhuma cepa. O gene cphA foi encontrado em 97,6 por cento (40) e 100 por cento (46) das cepas de A. hydrophila e A. jandaei, respectivamente e o gene bla(TEM) foi encontrado em 97,6 por cento (40) das cepas de A. hydrophila e em 85 por cento (39) das cepas de A. jandaei. Foi verificada a presença de plasmídios em 10/41 (24,4 por cento) das cepas de A. hydrophila e em 16/46 (34,9 por cento) das cepas de A. jandaei.