A Multiplex PCR Assay for Detection of Salmonella sp., Listeria monocytogenes, Campylobacter sp., Staphylococcus aureus and Escherichia coli

A multiplex PCR assay was evaluated for diagnosis of diarrheagenic Escherichia coli in stool samples of patients with diarrhoea submitted to a diagnostic microbiology laboratory. Two procedures of DNA template preparationproteinase K buffer method and the boiling method were evaluated to examine isolates of E. coli from 150 selected diarrhoeal cases. By proteinase K buffer method, 119 strains (79.3%) of E. coli were characterized to various categories by their genes that included 55.5% enteroaggregative E. coli (EAEC), 18.5% enterotoxigenic E. coli (ETEC), 1.7% enteropathogenic E. coli (EPEC), and 0.8% Shiga toxin-producing E. coli (STEC). Although boiling method was less time consuming (<24 hrs) and less costly (<8.0 US $/ per test) but was less efficient in typing E. coli compared to proteinase K method (41.3% vs. 79.3% ; p<0.001). The sensitivity and specificity of boiling method compared to proteinase K method was 48.7% and 87.1% while the positive and negative predictive value was 93.5% and 30.7%, respectively. The majority of pathogenic E. coli were detected in children (78.0%) under five years age with 53.3% under one year, and 68.7% of the children were male. Children under 5 years age were frequently infected with EAEC (71.6%) compared to ETEC (24.3%), EPEC (2.7%) and STEC (1.4%). The multiplex PCR assay could be effectively used as a rapid diagnostic tool for characterization of diarrheagenic E. coli using a single reaction tube in the clinical laboratory setting.Bangladesh J Med Microbiol 2007; 01 (02): 38-42

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Detection of the Panton-Valentine Leukocidin Gene in Swedish Isolates of Methicillin-Resistant Staphylococcus aureus using a Multiplex PCR Assay

10.35248/2155-9597.19.10.353 ◽

2019 ◽

Vol 10 (2) ◽

Author(s):

Waleed Abu Al-Soud

Keyword(s):

Staphylococcus Aureus ◽

Multiplex Pcr ◽

Methicillin Resistant Staphylococcus Aureus ◽

Pcr Assay ◽

Methicillin Resistant ◽

Multiplex Pcr Assay ◽

Panton Valentine Leukocidin ◽

Valentine Leukocidin

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Molecular detection of antibiotic resistance genes in multidrug-resistant Staphylococcus aureus isolates from dog dental plaque

BULGARIAN JOURNAL OF VETERINARY MEDICINE ◽

10.15547/bjvm.2099 ◽

2019 ◽

Vol 22 (4) ◽

pp. 419-427

Author(s):

S. Nouri Gharajalar ◽

M. Onsori

Keyword(s):

Staphylococcus Aureus ◽

Antibiotic Resistance ◽

Multiplex Pcr ◽

Resistance Genes ◽

Dental Plaque ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

Diffusion Method ◽

Pcr Assay ◽

Multiplex Pcr Assay

Multidrug resistant Staphylococcus aureus strains are a major health care problem both in humans and animals. In this work we described three multiplex PCR assays for detection of clinically relevant antibiotic resistance genes in S. aureus isolated from dog dental plaques. Thirty dental plaque samples were collected; then cultural, biochemical and molecular tests performed for isolation and identification of S. aureus from samples. The antibiotic susceptibility of the isolates were checked by Kirby Bauer disc diffusion method and the prevalence of antibiotic resistance genes determined using multiplex PCR assay. As a result S. aureus was isolated from 18 dog plaque samples. Fifteen of these isolates were resistant to penicillin. The mecA gene was more prevalent than blaZ among penicillin-resistant bacteria. Ten of the isolates were resistant to tetracycline. The percentage of tetM was higher than tetK among them. Also, 10 of the isolates were resistant to cefazolin among them bla TEM detected in higher rate than blaSHV and blaOXA-1. Hence multiplex PCR assay is a suitable method for detection of antibiotic resistance patterns of S. aureus isolates.

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Development of multiplex PCR assay for rapid detection Listeria monocytogenes in clinical samples

Tạp chí Y học Dự phòng ◽

10.51403/0868-2836/2020/246 ◽

2021 ◽

Vol 30 (4) ◽

pp. 20-26

Author(s):

Le Thanh Huong ◽

Ha Thi Phuong Mai ◽

Hoang Thi Thu Ha ◽

Nguyen Dong Tu ◽

Bui Tien Sy ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Multiplex Pcr ◽

Rapid Detection ◽

Pathogenic Bacteria ◽

Vulnerable Groups ◽

Clinical Samples ◽

Specific Gene ◽

Pcr Assay ◽

Multiplex Pcr Assay ◽

Specificity And Sensitivity

Listeria monocytogenes is widely present in the natural environment. This bacteria can cause infections in both humans and animals. In humans, the most vulnerable groups to be infected with L. monocytogenes are the elderly, people with an impaired immune system and chronically illness, pregnant women, and newborn babies. The aim of this study was to develop a multiplex PCR assay for the rapid detection of L. monocytogenes in mock clinical samples. A pair of primers were designed for detection of L. monocytogenes based on prs, a Listeria genus specific gene, and hly, a hemolysin gene. The specificity of the primers were tested by using different L. monocytogenes strains and other common pathogenic bacteria. The results showed that L. monocytogenes strains were positive in the detection and other tested strains were negative in mock (spiked) clinical samples. The sensitivity of multiplex PCR assay was 102 CFU/ml per reaction. The specificity and sensitivity of multiplex PCR technology for detecting L. monocytogenes in mock (spiked) clinical samples were high, and the assay could be completed within 1.5 hours. Therefore, this established multiplex PCR provides a rapid and reliable method and will be useful for the detection of L. monocytogenes in mock clinical samples.

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Corrigendum to “Enhanced multiplex PCR assay for the typing of staphylococcal cassette chromosome mec types I to V in methicillin-resistant Staphylococcus aureus”

Molecular and Cellular Probes ◽

10.1016/j.mcp.2019.03.004 ◽

2019 ◽

Vol 45 ◽

pp. 68

Author(s):

Kunyan Zhang ◽

Jo-Ann McClure ◽

John M. Conly

Keyword(s):

Staphylococcus Aureus ◽

Multiplex Pcr ◽

Methicillin Resistant Staphylococcus Aureus ◽

Pcr Assay ◽

Methicillin Resistant ◽

Multiplex Pcr Assay ◽

Staphylococcal Cassette Chromosome

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Application of a Multiplex Polymerase Chain Reaction Assay for the Simultaneous Confirmation of Listeria monocytogenes and Other Listeria Species in Turkey Sample Surveillance†

Journal of Food Protection ◽

10.4315/0362-028x-65.5.780 ◽

2002 ◽

Vol 65 (5) ◽

pp. 780-785 ◽

Cited By ~ 30

Author(s):

IRENE V. WESLEY ◽

KAREN M. HARMON ◽

JAMES S. DICKSON ◽

ANN RAMOS SCHWARTZ

Keyword(s):

Polymerase Chain Reaction ◽

Listeria Monocytogenes ◽

Multiplex Pcr ◽

Multiplex Polymerase Chain Reaction ◽

Rrna Gene ◽

Chain Reaction ◽

Pcr Assay ◽

Listeria Species ◽

Multiplex Pcr Assay ◽

Polymerase Chain

A multiplex polymerase chain reaction was developed to simultaneously identify Listeria monocytogenes and species of the genus Listeria. Two sets of primers were used, with the first amplifying a 938-bp region of the 16S rRNA gene that is highly conserved in all Listeria species and the second amplifying a 174-bp region of the listeriolysin (hlyA) gene of L. monocytogenes. Thus, isolates of Listeria spp. yield a single 938-bp product, whereas L. monocytogenes isolates yield both the 938-bp product and a 174-bp product. The specificity of the assay was verified with all six Listeria species and 11 serotypes of L. monocytogenes, as well as nonrelated bacteria. The multiplex PCR assay was used to determine the incidence of Listeria spp., especially L. monocytogenes, in mechanically separated turkey samples (n = 150 samples). L. monocytogenes strains were selected by using the University of Vermont two-step enrichment protocol and plating to selective Palcam agar. The multiplex PCR assay was used for verification of presumptive Listeria colonies. Approximately 38% of mechanically separated turkey samples (57 of 150) yielded L. monocytogenes; an additional 18% of these samples (27 of 150) harbored other Listeria spp. Fifty-one percent (29 of 57) of the L. monocytogenes isolates were of serogroup 1, 44% (25 of 57) were of serogroup 4, and 2% (1 of 57) were assigned to serogroups other than 1 and 4.

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