Comparison of culture, PCR and immunoassays for detecting Escherichia coli O157 following enrichment culture and immunomagnetic separation performed on naturally contaminated raw meat products

SUMMARYVibrio parahaemolyticus is a marine bacterium, responsible for gastroenteritis in humans. Most of the clinical isolates produce thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH) encoded by tdh and trh genes respectively. In this study, twenty-three V. parahaemolyticus, previously isolated from oysters and mussels were analyzed by PCR using specific primers for the 16S rRNA and virulence genes (tdh, trh and tlh) and for resistance to different classes of antibiotics and PFGE. Nineteen isolates were confirmed by PCR as V. parahaemolyticus. The tlh gene was present in 100% of isolates, the tdh gene was identified in two (10.5%) isolates, whereas the gene trh was not detected. Each isolate was resistant to at least one of the nine antimicrobials tested. Additionally, all isolates possessed the bla TEM-116 gene. The presence of this gene in V. parahaemolyticus indicates the possibility of spreading this gene in the environment. Atypical strains of V. parahaemolyticus were also detected in this study.

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“…V. harveyi was used as a negative control. DNA extraction was carried out for all samples using the heat-shock technique 7 . Kanagawa Test.…”

Section: Methodsmentioning

confidence: 99%

Characterization of Vibrio Parahaemolyticus isolated from oysters and mussels in São Paulo, Brazil

Rojas

Matté

Dropa

et al. 2011

Rev. Inst. Med. trop. S. Paulo

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“…Total chromosomal DNA from Aeromonas strains was extracted using the heat-shock method according to Chapman et al (2001) with modifications. In brief, the strains were grown overnight in 2 ml of LB (Luria Broth) 1% NaCl and centrifuged for 5 min at 10,000 £ g. The pellet was resuspended in 1 ml sterile MilliQ w water and centrifuged for 3 min at 10,000 £ g. The pellet was resuspended in 200 ml of sterile MilliQ w water and the 1.5 ml microtubes were boiled for 10 min at 958C and then immediately frozen for 30 min at 2208C.…”

Section: Preparation Of Dnamentioning

confidence: 99%

Molecular detection of enterotoxins in environmental strains of Aeromonas hydrophila and Aeromonas jandaei

Balsalobre

Dropa

Matté

et al. 2009

Journal of Water and Health

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Aeromonas species are widely distributed in aquatic environments and recent studies include the genus in the emergent pathogens group because of its frequent association with local and systemic infections in immunocompetent humans. Aiming to search for virulence genes in environmental strains of Aeromonas hydrophila and Aeromonas jandaei, we designed specific primers to detect act/hly A/aer complex and alt genes. Primers described elsewhere were used to detect ast. Eighty-seven strains previously identified using phenotypic and genotypic tests as A. hydrophila (41) and A. jandaei (46) were analysed for the presence of the virulence genes using PCR. DNA fragments of expected size were purified and directly sequenced. Among the 41 strains of A. hydrophila 70.7% (29), 97.6% (40) and 26.8% (11) possessed act/hly A/aer complex, ast and alt genes, respectively. Among the 46 strains of A. jandaei, 4.4% (2), 0% (0) and 32.6% (15) were positive for act/hly A/aer complex, ast and alt genes, respectively. Sequencing allowed for the confirmation of amplified products using BLAST. The present work proposes a specific and rapid diagnostic method to detect the main virulence determinants of Aeromonas, a genus potentially pathogenic to humans.

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“…Isolates were responsible for community-or hospital-acquired infections, and for colonization. Detection of bla genes: Total DNA extraction was carried out for all samples using the heat-shock technique (CHAPMAN et al 2001). Isolates were tested by PCR for the β-lactamase groups SHV, TEM and CTX-M, with previously described oligonucleotides and reaction conditions (CAO et al 2002).…”

Section: Settingmentioning

confidence: 99%

Extended-spectrum beta-lactamases among Enterobacteriaceae isolated in a public hospital in Brazil

Dropa

Balsalobre

Lincopán

et al. 2009

Rev. Inst. Med. trop. S. Paulo

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SUMMARYExtended-spectrum β-lactamases (ESBL) in enterobacteria are recognized worldwide as a great hospital problem. In this study, 127 ESBL-producing Enterobacteriaceae isolated in one year from inpatients and outpatients at a public teaching hospital at São Paulo, Brazil, were submitted to analysis by PCR with specific primers for bla SHV , bla TEM and bla CTX-M genes. From the 127 isolates, 96 (75.6%) Klebsiella pneumoniae, 12 (9.3%) Escherichia coli, 8 (6.2%) Morganella morganii, 3 (2.3%) Proteus mirabilis, 2 (1.6%) Klebsiella oxytoca, 2 (1.6%) Providencia rettgeri, 2 (1.6%) Providencia stuartti, 1 (0.8%) Enterobacter aerogenes and 1 (0.8%) Enterobacter cloacae were identified as ESBL producers. Bla SHV , bla TEM and bla CTX-M were detected in 63%, 17.3% and 33.9% strains, respectively. Pulsed field gel eletrophoresis genotyping of K. pneumoniae revealed four main molecular patterns and 29 unrelated profiles. PCR results showed a high variety of ESBL groups among strains, in nine different species. The results suggest the spread of resistance genes among genetically different strains of ESBL-producing K. pneumoniae in some hospital wards, and also that some strongly related strains were identified in different hospital wards, suggesting clonal spread in the institutional environment.

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Comparison of culture, PCR and immunoassays for detecting Escherichia coli O157 following enrichment culture and immunomagnetic separation performed on naturally contaminated raw meat products

Cited by 61 publications

References 39 publications

Characterization of Vibrio Parahaemolyticus isolated from oysters and mussels in São Paulo, Brazil

Characterization of Vibrio Parahaemolyticus isolated from oysters and mussels in São Paulo, Brazil

Molecular detection of enterotoxins in environmental strains of Aeromonas hydrophila and Aeromonas jandaei

Extended-spectrum beta-lactamases among Enterobacteriaceae isolated in a public hospital in Brazil

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