Genetic diversity and known virulence genes in Listeria innocua strains isolated from cattle abortions and farm environment.

Listeria innocua is considered as non-pathogenic bacteria living in an environment although several cases of immunocompromised humans and ruminant listeriosis infections have been reported. Previously, L. innocua was identified as a potential pathogen and virulence in association with L. monocytogenes PrfA dependent virulence (LIPI-1) gene cluster was demonstrated in hemolytic L. innocua. L. innocua usually considered non-pathogenic versus pathogenic L. monocytogenes and L. ivanovii because of the main virulence gene loss. There are limited studies and reports available about L. innocua-caused illness in cattle. A total of 18 STs were identified in cattle abortions while 17 STs in the farm environment with majority of STs were present in both abortions and environmental samples. Genome sequencing showed that in one farm identical L. innocua clones were represented in water, feed, soil, and faeces sample groups, suggesting that animals most likely through the faecal shedding may remain as the main source of L. innocua in a farm environment. Out of all L. innocua isolates PrfA-dependent virulence genes were not found in aborted foetuses isolates and environmental L. innocua isolate groups; however, in 20% of isolates a complete LIPI-3 pathogenicity island encoding listeriolysin S was identified. In this study, we demonstrated that genetically diverse L. innocua clones were widely distributed in cattle farm environment and certain isolates had a significant pathogenicity potential for cattle, thus causing adverse health effects, including abortions.

Prevalence, virulence determinants, and genetic diversity in Yersinia enterocolitica isolated from slaughtered pigs and pig carcasses.

Yersinia enterocolitica is an important zoonotic foodborne pathogen that could be transferred from infected pigs to their carcasses at slaughter, with subsequent introduction of the pathogen into the food chain. The aim of the present study was to study the prevalence, virulence characteristics, and genetic diversity of Y. enterocolitica isolates present in slaughtered pig tonsils and carcasses by using the WGS approach. A total of 200 slaughtered pig tonsils from 11 pig farms were collected in 2020-2021 at six slaughterhouses located in Latvia. Out of these samples, n = 190 were obtained from slaughtered pigs raised on Latvian farms while n = 10 were of Lithuanian origin, with the number of farms sampled being 10 and 1, respectively. Additionally, 30 pig carcasses were sampled at five slaughterhouses from pigs originating from five farms in 2021. Samples were investigated microbiologically, Y. enterocolitica isolates were biotyped and serotyped. Y. enterocolitica 4/O:3 was screened for antimicrobial resistance with the EUVSEC test panels. Whole genome sequence analysis (WGS) was performed in order to detect virulence genes and to assess the genetic diversity of Y. enterocolitica isolates. A total of 139 isolates, including one to three isolates from 84 Y. enterocolitica positive slaughtered pig tonsils and 13 pig carcass samples, were subjected to WGS analysis. The prevalence of Y. enterocolitica 4/O:3 in slaughtered pig tonsils and carcasses was 35% (70/200) and 13% (4/30), respectively. Antimicrobial resistance to ampicillin and tetracycline was detected in 97% (72/74) and 1% (1/74) of Y. enterocolitica 4/O:3 isolates. Y. enterocolitica 4/O:3 was represented only by ST18, while Y. enterocolitica 1A by ST3, ST147, ST304, ST307, and ST473. The ST18 isolates harbored the same main chromosomal (ail, inv, myfA, ystA) and majority shared plasmid-borne virulence genes (virF, yadA, yop virulon). The main virulence genes were not identified within the STs of Y. enterocolitica 1A and only minor differences were found between ST3, ST147, ST304, ST307, and ST473. Among ST18 isolates, cgMLST analysis revealed 43 cgMLST genotypes while 16 cgMLST genotypes were found among Y. enterocolitica 1A STs. The present study has shown the distribution of genetically distant cgMLST genotypes in slaughtered pigs from pig farms located in different geographical regions of Latvia, with one to 11 cgMLSTs identified within each sampled farm. The presence of undistinguishable cgMLST genotypes in slaughtered pig tonsils and the respective carcasses supported the link between the slaughter of Y. enterocolitica - positive pigs and carcass contamination with Y. enterocolitica 4/O:3.

Characterization and Genetic Diversity of Listeria monocytogenes Isolated from Cattle Abortions in Latvia, 2013-2018.

Listeria monocytogenes can cause disease in humans and in a wide range of animal species, especially in farm ruminants. The aim of the study was to determine the prevalence and genetic diversity of L. monocytogenes related to 1185 cattle abortion cases in Latvia during 2013-2018. The prevalence of L. monocytogenes among cattle abortions was 16.1% (191/1185). The seasonality of L. monocytogenes abortions was observed with significantly higher occurrence (p < 0.01) in spring (March-May). In 61.0% of the cases, the affected cattle were under four years of age. L. monocytogenes abortions were observed during the third (64.6%) and second (33.3%) trimesters of gestation. Overall, 27 different sequence types (ST) were detected, and four of them, ST29 (clonal complex, CC29), ST37 (CC37), ST451 (CC11) and ST7 (CC7), covered more than half of the L. monocytogenes isolates. Key virulence factors like the prfA-dependent virulence cluster and inlA, inlB were observed in all the analyzed isolates, but lntA, inlF, inlJ, vip were associated with individual sequence types. Our results confirmed that L. monocytogenes is the most important causative agent of cattle abortions in Latvia and more than 20 different STs were observed in L. monocytogenes abortions in cattle.

Prevalence, Genetic Diversity and Factors Associated with Distribution of Listeria monocytogenes and Other Listeria spp. in Cattle Farms in Latvia.

Listeria spp. is a diverse genus of Gram-positive bacteria commonly present in the environment while L. monocytogenes and L. ivanovii are well known human and ruminant pathogens. The aim of the present study was to reveal the prevalence and genetic diversity of L. monocytogenes and other Listeria spp. and to identify the factors related to the abundance of pathogen at cattle farms. A total of 521 animal and environmental samples from 27 meat and dairy cattle farms were investigated and the genetic diversity of L. monocytogenes isolates was studied with WGS. The prevalence of Listeria was 58.9%, while of L. monocytogenes it was -11%. The highest prevalence of L. monocytogenes was found in the environment-soil samples near to manure storage (93%), mixed feed from the feeding trough and hay (29%), water samples from farms drinking trough (28%) and cattle feces (28%). Clonal complexes (CC) of CC37 (30%), CC11 (20%) and CC18 (17%) (all IIa serogroup) were predominant L. monocytogenes clones. CC18, CC37 and CC8 were isolated from case farms and CC37, CC11 and CC18 from farms without listeriosis history. Only one hypervirulent CC4 (1%) was isolated from the case farm. Sequence types (STs) were not associated with the isolation source, except for ST7, which was significantly associated with soil (p < 0.05). The contamination of soil, feeding tables and troughs with L. monocytogenes was associated with an increased prevalence of L. monocytogenes at farms. Our study indicates the importance of hygienic practice in the prevention of the dissemination of L. monocytogenes in the cattle farm environment.

Virulence Determinants and Genetic Diversity of Yersinia Species Isolated from Retail Meat.

Yersinia enterocolitica is an important foodborne pathogen, and the determination of its virulence factors and genetic diversity within the food chain could help understand the epidemiology of yersiniosis. The aim of the present study was to detect the prevalence, and characterize the virulence determinants and genetic diversity, of Yersinia species isolated from meat. A total of 330 samples of retailed beef (n = 150) and pork (n = 180) in Latvia were investigated with culture and molecular methods. Whole genome sequencing (WGS) was applied for the detection of virulence and genetic diversity. The antimicrobial resistance of pathogenic Y. enterocolitica isolates was detected in accordance with EUCAST. Yersinia species were isolated from 24% (79/330) of meats, and the prevalence of Y. enterocolitica in pork (24%, 44/180) was significantly higher (p < 0.05) than in beef (13%, 19/150). Y. enterocolitica pathogenic bioserovars 2/O:9 and 4/O:3 were isolated from pork samples (3%, 6/180). Only resistance to ampicillin was confirmed in Y. enterocolitica 4/O:3 and 2/O:9 isolates, but not in other antimicrobials. Major virulence determinants, including ail, inv, virF, ystA and myfA, were confirmed with WGS in Y. enterocolitica 2/O:9 and 4/O:3. MLST typing revealed 15 STs (sequence types) of Y. enterocolitica with ST12 and ST18, which were associated with pathogenic bioserovars. For Y. enterocolitica 1A, Y. kristensenii, Y. intermedia and Y. frederiksenii, novel STs were registered (ST680-688). The presence of virulence genes and genetic characteristics of certain Y. enterocolitica STs confirm the common knowledge that pork could be an important source of pathogenic Yersinia.