PMA-LAMP for rapid detection of Escherichia coli and shiga toxins from viable but non-culturable state.
In exposure to outer pressure, microorganisms are capable of entry into the Viable But Non-Culturable (VBNC) state, and thus survive under various elimination processing. The survival microorganisms may yield negative results on culturing, and cause false negative for this golden standard methodology. In this study, a novel PMA-LAMP assay on the detection of Enterohemorrhage E. coli and shiga toxins has been developed and evaluated, with further application on a number of food borne E. coli strains. LAMP primers were designed on the target of rfbe for Enterohemorrhage E. coli and stx1with stx2 for shiga toxins. Via specific penetration through the damaged cell membrane of dead cells and intercalating into DNA, PMA could prevent DNA amplification of dead bacteria from LAMP, which enabled the differentiation of bacteria between VBNC state and dead state. The established PMA-LAMP showed significant advantage in rapidity, sensitivity and specificity, compared with regular PCR assay. The applicability had also been verified, demonstrating the PMA-LAMP was capable of detection on Enterohemorrhage E. coli and shiga toxins.
Department of Haematology, Guangzhou Women and Children's Medical Centre, Guangzhou Medical University, Guangzhou 510623, PR China. Electronic address: robinbobo@qq.com.
Muxia
M
Yan
Department of Haematology, Guangzhou Women and Children's Medical Centre, Guangzhou Medical University, Guangzhou 510623, PR China. Electronic address: 574096897@qq.com.
Ling
L
Xu
Department of Haematology, Guangzhou Women and Children's Medical Centre, Guangzhou Medical University, Guangzhou 510623, PR China. Electronic address: 1006239731@qq.com.
Hua
H
Jiang
Clinical Microbiogy, Guangzhou Women and Children's Medical Centre, Guangzhou Medical University, Guangzhou 510180, PR China. Electronic address: 471122711@qq.com.
Zhenwen
Z
Zhou
School of Bioscience and Bioengineering, South China University of Technology, 382 Zhonghuan Road East, Guangzhou 510006, PR China. Electronic address: zhoushishui2017@163.com.
Shishui
S
Zhou
Department of Haematology, Guangzhou Women and Children's Medical Centre, Guangzhou Medical University, Guangzhou 510623, PR China. Electronic address: sherryhomles@163.com.
Li
L
Zhang
D001431:Bacteriological Techniques / Q000379:methods*; D002238:Carbohydrate Epimerases / Q000235:genetics; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000302:isolation & purification*; D019453:Escherichia coli O157 / Q000235:genetics / Q000302:isolation & purification; D005516:Food Microbiology / Q000379:methods*; D059022:Food Safety / Q000379:methods; D005517:Foodborne Diseases / Q000382:microbiology; D008826:Microbial Sensitivity Tests; D050296:Microbial Viability; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D016133:Polymerase Chain Reaction; D022622:Shiga Toxin 1 / Q000235:genetics; D022641:Shiga Toxin 2 / Q000235:genetics; D022603:Shiga Toxins / Q000235:genetics / Q000302:isolation & purification*; D000637:Transaminases / Q000235:genetics
D017931:DNA Primers; D004269:DNA, Bacterial; D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; D022603:Shiga Toxins; D000637:Transaminases; D002238:Carbohydrate Epimerases; C061272:perosamine synthetase
Enterohemorrhage Escherichia coli; LAMP; PMA; Shiga toxins; VBNC