| Listeria monocytogenes(L.monocytogenes), an zoonotic disease pathogens, can cause gastroenteritis, encephalitis, meningitis, septicaemia and abortion in humans and animals with a mortality of up to 20～30%。ith the rapid increase in the consumption of chilled and frozen food in recent years,the potential risk of L.monocytogenes in food is also on the rise. Experts and scholars have studied the biological characteristics, detection technology, pathogenicity gene, virulence factors and epidemiological of L.monocytogenes. L.monocytogenes encompasses a diversity of strains with varying pathogenic potential. While many L.monocytogenes strains could be high pathogenieity, others are less virulent or even avirulent and produce little harm in the host. Therefore, accurate and rapid evaluation of L.monocytogenes isolates virulence has become one of the most widely researched topics.This paper isolated and identified L.monocytogenes from 1645 samples.1645 random samples from 8 sorts of meat products, bean product, fruit, quick-frozen food, vegetables, aquatic products, raw meat and raw milk were collected from supermarkets and farm produce markets.49 strains of L.monocytogenes isolated were subjected to PCR assay for seven virulence-associated genes (hly, plcB, inlA, inlB, inlC, inlJ, prfA) and toxicity test in mice, to analyze the relationship between virulence-associated genes and L. monocytogenes virulence, to set up a method of evaluating the virulence of L.m isolates by PCR amplification.The results:The total detectable rate of L.monocytogenes was 3.0%. The results showed that the detectable rate of L.monocytogenes was lower than other areas. The study on the distributing status of L. monocytogenes indicated that the able rate of L.monocytogenes in raw meat samples was the highest (5.8%). The next was meat products (3.3%) and aquatic products (2.9%). The PCR assay revealed that 19 of 49 strains isolated possessed all the seven virulence-associated genes,7 strains were hly-,4 strains were plcB-,4 strains were prfA-,5 strains were inlA-, inlB-,13 strains were inlC-and 12 strains were inlJ-,1 strain was negative for inlA,inlB,inlC and inlJ. Four isolates were defined virulent strains, whose virulence was equivalent to that of the standard strain, ATCC 1961E with the mice LD50 between 1.2×10 and 6.0×10 CFU/mL. The LD50 of screened low virulent strain 09-132 was 1.7×1011 CFU/mL.The conclusions are as follows:1. In terms of detection rate and probability, ALOA selective medium is better and more suitable for lab use; However, L. monocytogenes can not be completely detected when using one media alone in broth.2. The population distribution of Listeria is not the same in different kinds of food samples. Quick-frozen food, raw meat, meat products and aquatic products were polluted by two and above Listeria, and one of them was L.monocytogenes.The results show that there is potential risk of Listeria food poisoning in Chongqing City.3. The serotype of isolates was irrelevant to virulence. The positive rate of the internalization gene (inlA, inlB, inlC, inlJ) from raw poultry meat isolates was higher than that from the raw meat isolates. The internalization gene (inlA, inlB, inlC, inU) loss may lead to virulation declination, while the prfA and plcB genes are less relevant to the virulence isolates. inlC could serve as markers for PCR assessment of L.monocytogenes virulence.Suggestions:The knock-out mutants of virulence-associated genes are needed to better define the relationship between virulence-associated genes and L. monocytogenes virulence. And studies of those kinds are likely to form the basis for the supervision of L.monocytogenes, the examination of explosion and spread and the tracking of polluter. |
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