[Study on rapid detection techniques of PCR-ELISA for Listeria monocytogenes].
OBJECTIVE
To develop rapid PCR-ELISA methods for detecting Listeria monocytogenes, and detect food samples artificially contaminated with Listeria monocytogenes.


METHODS
Specific primers for Listeria monocytogenes pathogenic gene hlyA were selected based on the Genbank data by using molecular biological software DNAman6.0. Digoxigenin-labeled hlyA fragments were obtained by using commercial kit. Specific capture probes were obtained by comparing bacterial pathogenic gene sequences in the Genbank. PCR-ELISA methods were developed and the Listeria monocytogenes isolates with different serotypes were detected. The sensitivity of PCR and PCR-ELISA was determined by artificially inoculating Listeria monocytogenes strains in milk.


RESULTS
It took 6 hours to detect Listeria monocytogenes in food samples by PCR-ELISA. The accordance rate to the bacteriological method was 100%. After 12 h pre-enrichment, the detection limit of PCR-ELISA method was 1 CFU/25 ml milk. The sensitivity of PCR-ELISA method was 10-100 times as PCR.


CONCLUSIONS
The PCR-ELISA method for rapidly detecting Listeria monocytogenes was established. The sensitivity, specificity and reliability of the method proved to be good. It would be valuable to improve the precaution and prediction abilities of food-borne diseases and enhance the chronergy and accuracy of detection method.
Cao
Wei
W
Institute of Nutrition and Food Safety, Chinese Center for Disease Control and Prevention, Beijing.
Wang
Ning
N
Wang
Xiaoying
X
Liu
Hongsheng
H
Guo
Yunchang
Y
D001427:Bacterial Toxins / Q000032:analysis / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D006360:Heat-Shock Proteins / Q000032:analysis / Q000235:genetics*; D006460:Hemolysin Proteins / Q000032:analysis / Q000235:genetics*; D008089:Listeria monocytogenes / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D001427:Bacterial Toxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; C038705:hlyA protein, Listeria monocytogenes