title: A real-time loop-mediated isothermal amplification assay for rapid detection of Shigella species.
abstract: Shigellosis is a foodborne illness caused by the genus Shigella and is an important global health issue. The development of effective techniques for rapid detection of this pathogen is essential for breaking the chain of transmission. Therefore, we have developed a novel loop-mediated isothermal amplification (LAMP) assay targeting the invasion plasmid antigen H (ipaH) gene to rapidly detect Shigella species. This assay could be performed in 90 min at an optimal temperature of 64ºC, with endpoint results visualized directly. Notably, the method was found to be more sensitive than conventional PCR. Indeed, the detection limit for the LAMP assay on pure bacterial cultures was 5.9 x 10(5) CFU/ml, while PCR displayed a limit of 5.9 x 10(7) CFU/ml. In spiked lettuce samples, the sensitivity of the LAMP assay was 3.6 x 10(4) CFU/g, whereas PCR was 3.6 x 10(5) CFU/g. Overall, the assay accurately identified 32 Shigella spp. with one enteroinvasive Escherichia coli displaying positive reaction while the remaining 32 non-Shigella strains tested were negative.
authors.affiliation: Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: P S
authors.initials: PS
authors.lastname: Liew
authors.affiliation: Laboratory of Biomedical Science and Molecular Microbiology, Institute of Graduate Studies, University of Malaya.
authors.forename: C S J
authors.initials: CS
authors.lastname: Teh
authors.affiliation: Department of Parasitology, Faculty of Medicine, University of Malaya.
authors.forename: Y L
authors.initials: YL
authors.lastname: Lau
authors.affiliation: Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: K L
authors.initials: KL
authors.lastname: Thong
mesh_terms: D000942:Antigens, Bacterial / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D004405:Dysentery, Bacillary / Q000175:diagnosis* / Q000382:microbiology; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D012760:Shigella / Q000235:genetics / Q000302:isolation & purification*; D013696:Temperature; D013997:Time Factors
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C063500:ipaH protein, Shigella flexneri