BIOCHEMICAL CHARACTERIZATION, MOLECULAR CONFIRMATION AND ANTIMICROBIAL SUSCEPTIBILITY TEST OF Listeria monocytogenes FROM RAW BOVINE MILK COLLECTED FROM HAWASSA CITY, DALE AND ARSI NEGELE DISTRICTS OF SOUTHERN ETHIOPIA

Abstract

Listeria monocytogenes is one of the most important foodborne pathogens and causes zoonotic disease due to its abundance in the environment. It is responsible for Listeriosis in humans and animals with low incidence but high fatality rate. Hence, this study was conducted to assess the prevalence, antibiotic susceptibility patterns, and molecular characterization of Listeria monocytogenes from raw cow milk. A cross-sectional study design was employed from December 2019 to September 2020. A total of 240 raw bovine milk samples were collected from dairy farms and smallholder dairy producers using a simple random sampling technique. Milk samples were collected in two season. Listeria monocytogenes isolation was performed using standard cultural procedures including Twin pack Listeria selective broth and agar medium as well as confirmatory broths like rhamnose, xylose and mannitol. Heamolysis and CAMP tests were also conducted. Molecular detection was performed by multiplex PCR by targeting five virulence-associated genes (prfA, plcA, actA, hlyA, and iapA) and by 16S rRNA genes sequence. The antimicrobial susceptibility profile of L. monocytogenes was also assessed by using the standard disk diffusion method. Out of 240 raw bovine milk samples tested, 69 (28.75%) were found positive for Listeria spp. of which 17 (7.08%) isolates were identified as Listeria monocytogenes after morphological and biochemical confirmation. The prevalence of Listeria monocytogenes in the wet season was 9.32%, but 4.92% in the dry season. Antimicrobial resistance profiling showed that all the Listeria monocytogenes isolates were completely resistant to Nalidixic acid followed by erythromycin (88.24%), ampicillin (23.53%), chloramphenicol (17.65%), streptomycin (11.76%) and 5.88% isolate showed resistance to cefotaxime. However, all the L. monocytogenes isolates were sensitive to vancomycin, gentamicin, and sulfamethoxazole. Gene of 16S rRNA was found in all the 17 tested isolates, but hlyA and iapA were found only in the 11 tested isolates and no isolate was found to contain prfA, actA, and plcA genes. The study indicates that raw bovine milk is a significant source of L. monocytogenes and shows a potential health risk for listeriosis.