Prevalence and Characterisation of Listeria Monocytogenes Isolated From Chicken and Beef

Listeria monocytogenes is an opportunistic haemolytic pathogen of humans and animals involved in several outbreaks and sporadic cases of listeriosis associated with the consumption of contaminated food. Growing antibiotic resistance demands the constant reassessment of antimicrobial efficacy, particularly in countries with wide antibiotic used in veterinary science, where higher resistance prevalence is often found and it has been reported that antibiotic resistant is associated with plasmid in bacteria. The purpose of this study is to characterize L. monocytogenes isolated from chicken and beef meat by antibiotic resistance test, plasmid DNA profile and arbitrarily primed (AP)- and repetitive sequence (RS)-PCR analysis and to determine whether there is any correlation between the antibiotic resistant and the incidence of plasmid. A total of 112 samples of chicken meat and 101 samples of beef were collected from different wet markets and night markets in Malaysia. Listeria spp. were detected in 39 (34.8%) of the chicken meat and 21 (20.8%) of the beef samples, respectively. L. monocytogenes were detected in 27 (24.1%) and 17 (16.8%) of the chicken meat and beef, respectively. Out of all the Listeria spp., 42 and 20 isolates from the chicken meat and beef, respectively, were confirmed as L. monocytogenes. The antibiotic susceptibility test with sixteen different types of antibiotics revealed that the highest prevalence of resistance among the L. monocytogenes isolates was observed against nalidixic acid (100%). None (0%) of the isolates were resistant to vancomycin. The L. monocytogenes strains showed resistance to at least two or more of the antibiotics tested with their multiple antibiotic resistance index (MARI) ranging between 0.13 to 0.63. Thirty-three different patterns of resistance were observed among all the L. monocytogenes isolates. Plasmid analysis of the L. monocytogenes strains revealed that 24 (38.7%) of the strains carried plasmid DNA ranging in sizes from 1.75 to 104.0 megadalton (MDa). Spearman’s rho correlation analysis was utilised to determine the correlation coefficient between MARI and the incidence of plasmid in the L. monocytogenes isolates. The result shows that there were no significant correlation between the MARI and the incidence of the plasmid (r= 0.143, p= 0.269). The AP- and RS-PCR analyses generated diverse PCR patterns with multiple DNA fragments in sizes ranging between 250 and 3000 bp. Dendrograms generated from the PCR patterns clustered the isolates into several groups and subgroups. PCR analyses with primers GEN1-50-02, GEN1-50-10 and repetitive primer clustered the chicken isolates into 3, 13 and 6 main groups; and the beef isolates into 2, 4 and 5 main groups; respectively. These main groups were further clustered into several subgroups. Isolation of L. monocytogenes from the beef and chicken meat suggests that there is a risk of acquiring listeriosis through these popular meat sources in Malaysia. L. monocytogenes will be killed by cooking and raw or semi raw meat are not consumed in Malaysia. However, L. monocytogenes in raw beef and chicken meat may pose a health hazard in kitchen if contaminating cooked food or other ready to eat food. Considering outbreaks of listeriosis associated with different foods, avoidance of consumption of insufficiently cooked meat by at-risk populations is recommended. Hygiene quality control of meat and its products must be recommended during the slaughtering, transportation carriage, other used devices and stuff carriers. Diligent enforcement of sanitary conditions of food contact surface and handling areas, and personal hygiene practices should reduce the potential contamination of meat products by L. monocytogenes at the retail level. Resistance to two or more antibiotics among these isolates was common. It is suggested that incorrect use of these antimicrobial agents for therapeutical purposes in veterinary science may lead to the development of antibiotic resistance. The high MARI value indicated that the L. monocytogenes strains were originated from high risk sources of contamination in the geographical area. L. monocytogenes can no longer be thought to be uniformly susceptible to antibiotics active against Gram-positive bacteria. Continued surveillance of emerging antimicrobial resistance among L. monocytogenes is important to ensure effective treatment of human listeriosis. The non-significant correlation between the antibiotic resistance and the incidence of plasmid suggest that the plasmids could be cryptic plasmids which have no apparent effect on the phenotype, especially the antibiotic resistance of the host. The presence of hemolysin gene in the beef and chicken meat isolates is of public health concern, as this virulence gene is associated with pathogenicity of the bacteria in human listeriosis. Both AP-PCR and RS-PCR, having high discriminatory power, have revealed the high diversity of food related L. monocytogenes isolates and their suitability to track down the contamination sources. The results suggest how complex the epidemiology of the L. monocytogenes in the study area, as a result of several strains as opposed to the widespread transmission of a single type. The results do not support that certain genetically related strains are better adapted to a particular food source. The genomic heterogeneity of the L. monocytogenes found in this study confirms the usefulness of the AP- and RS-PCR analyses for the rapid differentiation and grouping of this organism.

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