Detection of shigella in lettuce by the use of a rapid molecular assay with increased sensitivity

• Published in: Brazilian journal of microbiology Vol. 41; no. 4; pp. 993 - 1000
• Main Authors: Jiménez, Kenia Barrantes(Universidad de Costa Rica Instituto de Investigaciones en Salud Infection-Nutrition Section), McCoy, Clyde B.(University of Miami Department of Epidemiology and Public Health), Achí, Rosario(Universidad de Costa Rica Instituto de Investigaciones en Salud Infection-Nutrition Section)
• Format: Journal Article
• Language: English
• Published: Brazil Sociedade Brasileira de Microbiologia 01-10-2010; Springer Nature B.V
• Subjects: Bacteria; food; Food contamination & poisoning; Lettuce; MICROBIOLOGY; PCR; Polymerase chain reaction; rapid method; Shigella; rapid method; lettuce; PCR; Shigella; food
• ISSN: 1517-8382; 1678-4405; 1678-4405
• DOI: 10.1590/S1517-83822010000400018

A Multiplex Polymerase Chain Reaction (PCR) assay to be used as an alternative to the conventional culture method in detecting Shigella and enteroinvasive Escherichia coli (EIEC) virulence genes ipaH and ial in lettuce was developed. Efficacy and rapidity of the molecular method were determined as compared to the conventional culture. Lettuce samples were inoculated with different Shigella flexneri concentrations (from 10 CFU/ml to 10(7) CFU/ml). DNA was extracted directly from lettuce after inoculation (direct-PCR) and after an enrichment step (enrichment PCR). Multiplex PCR detection limit was 10(4) CFU/ml, diagnostic sensitivity and specificity were 100% accurate. An internal amplification control (IAC) of 100 bp was used in order to avoid false negative results. This method produced results in 1 to 2 days while the conventional culture method required 5 to 6 days. Also, the culture method detection limit was 10(6) CFU/ml, diagnostic sensitivity was 53% and diagnostic specificity was 100%. In this study a Multiplex PCR method for detection of virulence genes in Shigella and EIEC was shown to be effective in terms of diagnostic sensitivity, detection limit and amount of time as compared to Shigella conventional culture.