Pathogenicity, genotyping and antibacterial susceptibility of the Listeria spp. recovered from stray dogs

Abstract

The present study aimed to determine the prevalence of Listeria spp. in stray dogs in the Kayseri province of Turkey. In addition, serotyping, genotyping and virulence gene analysis of the isolated Listeria spp. were performed and their pathogenicity and antibacterial susceptibility were investigated.The study included 80 rectal swaps taken from 80 stray dogs of different ages and gender that were sheltered in the Kayseri Municipal Dog Shelter. Listeria selective broth and Listeria selective agar were used for the isolation of Listeria spp. and the isolates were identified using a Microbact 12L (Oxoid, England) identification test kit. 16S rDNA sequencing and species-specific polymerase chain reaction (PCR) were performed for molecular identification of the isolates, multiplex PCR and a serological test were performed for serotyping, and PCR was used for virulence gene analysis. For determining the pathogenicity of L. monocytogenes and L. innocua isolates, a total of 100 mice (50 pregnant and 50 non-pregnant) were used. The mice were infected intraperitoneally; the inoculation dose was 1 × 109 CFU/mL and 0.2 mL was used for each animal. Tissue samples obtained from infected mice were processed for the re-isolation of the Listeria spp. and then stained with hematoxylin eosin and Brown-Brenn Gram stain. The antibiotic susceptibilities of the isolates were determined by the disk diffusion method.Listeria spp. were isolated from 5 (6.25%) of the 80 fecal samples. While 1 of the isolates was identified as L. monocytogenes, 4 of them were identified as L. innocua. Serotyping by serological and molecular methods revealed the isolate of L. monocytogenes to be serotype 1/2a. According to the phylogenetic trees, L. innocua and L. monocytogenes strains were clustered in different groups. The L. monocytogenes isolate was positive for all virulence genes tested. All L. innocua isolates were positive for the plcB gene. While all L. innocua isolates were negative for the lin1068 gene, 3 L. innocua isolates were found to be positive for the lin0558 gene. In mice infected with L. monocytogenes, pathological findings were observed in the uterus, intestines, pancreas, and heart. In mice infected with L. innocua, pathological findings were observed in the stomach, intestines and spleen. L. monocytogenes- or L. innocua-related infections or other inflammatory reactions were not observed in the brains of infected animals. On histopathological examination with Gram stain, an abundance of Listeria spp. was observed in the lesions of the liver, spleen, uterus, and kidney. Moreover, while abortion was observed in all animals infected with L. monocytogenes, it was not observed in any of the animals infected with L. innocua. Antibiotic susceptibility testing revealed that all 5 isolates were sensitive to ampicillin, amoxicillin/clavulanic acid, erythromycin, gentamicin, penicillin G, and trimethoprim-sulfamethoxazole and were resistant to nalidixic acid, streptomycin, and cefuroxime sodium.Considering also the pathogenicity of the isolated microorganisms, it can be suggested that stray dogs as carriers of Listeria spp. are a significant risk to public health. As L. innocua isolates, which are considered apathogenic, led to the occurrence of lesions similar to those caused by L. monocytogenes, detailed studies on the pathogenesis of L. innocua infections caused by L. innocua isolates recovered from various sources are required.