Prevalence and genotype diversity of Campylobacter jejuni in hunted reared pheasants (Phasianus colchicus) in Finland.

Campylobacter spp., especially C. jejuni, is the most common zoonotic pathogen in humans worldwide. In Nordic countries, the prevalence of C. jejuni in broilers, which is an important reservoir of human infections, is generally low. Thus, other sources of domestically acquired infections besides chicken meat need to be considered. Game birds are known to carry a variety of zoonotic agents, including Campylobacter. The aim of this study was to investigate Campylobacter spp. carriage in a flock of reared pheasants at hunting in two successive samplings to better understand the dynamics of Campylobacter infections in pheasants. Overall, 72% of the intestinal samples were positive for Campylobacter spp. by direct culture on mCCDA. C. jejuni was the only species identified. The isolates were genotyped using whole genome sequencing (WGS), multilocus sequence typing (MLST), and ad hoc whole genome MLST (wgMLST). Two distinct C. jejuni clones were identified among the 18 isolates studied, representing MLST sequence types (STs) ST-45 and ST-699. The ST-45 isolates were closely related to previous human clinical isolates using core genome MLST (cgMLST). In contrast, the ST-699 isolates forming the dominant clone in the latter sampling were quite distinct from previously described cgMLST profiles from different hosts and sources worldwide. In conclusion, the intestine of reared pheasants is commonly colonized by C. jejuni and may carry genotypes relevant to infections in livestock and humans. Hygienic measures are needed to limit the spread of infection in reared flocks. Especially farmers and hunters having direct contact with pheasant offal need to be aware of the associated zoonosis risk to protect themselves and their working dogs alike. Biosecurity measures to improve the safety and reduce the zoonosis risk associated with pheasant farming should be further investigated.

Microbiological quality and safety of vacuum-packaged white-tailed deer meat stored at 4 °C.

Vacuum packaging is widely used for extending the shelf life of commercial fresh meat products. It also ensures product hygiene during distribution and storage. However, very little information exists concerning the effects of vacuum packaging on the shelf life of deer meat. One of our aims was to evaluate how storage under vacuum at 4 °C affects the microbial quality and safety of white-tailed deer (Odocoileus virginianus) meat cuts. This was assessed in a longitudinal study based on sensory analyses and measurements of (1) mesophilic aerobic bacteria (MAB), (2) lactic acid bacteria (LAB), (3) enterobacteria (EB), (4) and Escherichia coli (EC) counts, and the presence of foodborne pathogens (Campylobacter, Salmonella, stx-harbouring E. coli (STEC), Yersinia and Listeria). Microbiomes were additionally investigated by 16S rRNA gene amplicon sequencing at the time of spoilage. In total, 50 vacuum-packaged meat cuts from the carcasses of 10 wild white-tailed deer harvested in southern Finland in December 2018 were analysed. A significant (p < 0.001) drop in the odour and appearance scores and a significant increase in MAB (p < 0.001) and LAB (p = 0.001) counts of the vacuum-packaged meat cuts were observed after 3 weeks of storage at 4 °C. A very strong correlation (rs = 0.9444, p < 0.001) between the MAB and LAB counts were found during the 5-week sampling period. Clear spoilage changes, manifested as sour off-odours (odour scores ≤2) and pale colour, were detected in the meat cuts spoilt after 3-week storage. High (≥8 log10 cfu/g) MAB and LAB counts were also detected. According to the 16S rRNA gene amplicon analyses, Lactobacillus was the dominant bacterial genus in these samples, demonstrating that LAB can cause rapid spoilage of vacuum-packaged deer meat cuts stored at 4 °C. The rest of the samples were spoilt after four or five weeks of storage, and a vast number of bacterial genera were identified in them. Listeria and STEC were detected by PCR in 50 % and 18 % of the meat cut samples, respectively, which may indicate a public health problem. Our results demonstrate that it is very challenging to ensure the quality and safety of vacuum-packaged deer meat stored at 4 °C, and freezing is therefore recommended to prolong the shelf life.

Characterisation of Listeria monocytogenes Isolates from Hunted Game and Game Meat from Finland.

Listeria monocytogenes is an important foodborne zoonotic bacterium. It is a heterogeneous species that can be classified into lineages, serogroups, clonal complexes, and sequence types. Only scarce information exists on the properties of L. monocytogenes from game and game meat. We characterised 75 L. monocytogenes isolates from various game sources found in Finland between 2012 and 2020. The genetic diversity, presence of virulence and antimicrobial genes were studied with whole genome sequencing. Most (89%) of the isolates belonged to phylogenetic lineage (Lin) II and serogroup (SG) IIa. SGs IVb (8%) and IIb (3%) of Lin I were sporadically identified. In total, 18 clonal complexes and 21 sequence types (STs) were obtained. The most frequent STs were ST451 (21%), ST585 (12%) and ST37 (11%) found in different sample types between 2012 and 2020. We observed 10 clusters, formed by closely related isolates with 0-10 allelic differences. Most (79%) of the virulence genes were found in all of the L. monocytogenes isolates. Only fosX and lin were found out of 46 antimicrobial resistance genes. Our results demonstrate that potentially virulent and antimicrobial-sensitive L. monocytogenes isolates associated with human listeriosis are commonly found in hunted game and game meat in Finland.

Hunted game birds - Carriers of foodborne pathogens.

Game birds may carry zoonotic bacteria in their intestines and transmit them to hunters through bird handling or through the handling and consumption of contaminated meat. In this study, the prevalence of foodborne bacteria was screened from game bird faeces and mallard breast meat using PCR. The sampling occurred in southern Finland from August to December during the hunting season. Isolates were characterized by multi-locus sequence typing. Mesophilic aerobic bacteria and Escherichia coli counts were used to assess the microbial contamination of mallard meat. In total, 100 woodpigeon (Columba palumbus), 101 pheasants (Phasianus colchicus), 110 mallards (Anas platyrhynchos), and 30 teals (Anas crecca) were screened during the hunting season. Additionally, 100 mallard breast meat samples were collected. Campylobacter and Listeria were commonly detected in the faeces and Listeria on mallard meat. L. monocytogenes of sequence types associated with human listeriosis were frequently found in game bird faeces and on mallard meat. Good hygiene during game bird handling, storing the game bird meat frozen, and proper heat treatment are important measures to minimize the health risk for hunters and consumers.

Population Genetics and Characterization of Campylobacter jejuni Isolates from Western Jackdaws and Game Birds in Finland.

Poultry are considered a major reservoir and source of human campylobacteriosis, but the roles of environmental reservoirs, including wild birds, have not been assessed in depth. In this study, we isolated and characterized Campylobacter jejuni from western jackdaws (n = 91, 43%), mallard ducks (n = 82, 76%), and pheasants (n = 9, 9%). Most of the western jackdaw and mallard duck C. jejuni isolates represented multilocus sequence typing (MLST) sequence types (STs) that diverged from those previously isolated from human patients and various animal species, whereas all pheasant isolates represented ST-19, a common ST among human patients and other hosts worldwide. Whole-genome MLST revealed that mallard duck ST-2314 and pheasant ST-19 isolates represented bacterial clones that were genetically highly similar to human isolates detected previously. Further analyses revealed that in addition to a divergent ClonalFrame genealogy, certain genomic characteristics of the western jackdaw C. jejuni isolates, e.g., a novel cdtABC gene cluster and the type VI secretion system (T6SS), may affect their host specificity and virulence. Game birds may thus pose a risk for acquiring campylobacteriosis; therefore, hygienic measures during slaughter and meat handling warrant special attention.IMPORTANCE The roles of environmental reservoirs, including wild birds, in the molecular epidemiology of Campylobacter jejuni have not been assessed in depth. Our results showed that game birds may pose a risk for acquiring campylobacteriosis, because they had C. jejuni genomotypes highly similar to human isolates detected previously. Therefore, hygienic measures during slaughter and meat handling warrant special attention. On the contrary, a unique phylogeny was revealed for the western jackdaw isolates, and certain genomic characteristics identified among these isolates are hypothesized to affect their host specificity and virulence. Comparative genomics within sequence types (STs), using whole-genome multilocus sequence typing (wgMLST), and phylogenomics are efficient methods to analyze the genomic relationships of C. jejuni isolates.

Microbial contamination of moose (Alces alces) and white-tailed deer (Odocoileus virginianus) carcasses harvested by hunters.

Hunting is currently a very popular activity, and interest in game meat is increasing. However, only limited research is available on the bacterial quality and safety of moose (Alces alces) and white-tailed deer (Odocoileus virginianus) harvested by hunters. Poor hunting hygiene can spread bacteria onto the carcasses, and inadequate chilling of the carcasses may increase the bacterial load on the carcass surface. We studied the bacterial contamination level on carcasses of 100 moose and 100 white-tailed deer shot in southern Finland. Hunters eviscerated carcasses in the field and skinned them in small slaughter facilities. During the sampling, same person visited 25 facilities located in 12 municipalities of four provinces. Moose carcasses had mean mesophilic aerobic bacteria (MAB), Enterobacteriaceae (EB) and Escherichia coli (EC) values of 4.2, 2.6 and 1.2 log10 cfu/cm2, respectively, while deer carcass values were 4.5, 1.5 and 0.7 log10 cfu/cm2, respectively. Moose carcasses were significantly more contaminated with EB and EC than deer carcasses. High bacterial counts (MAB>5.0 log10 cfu/cm2 and EB > 2.5 log10 cfu/cm2) on the carcasses were associated with the smallest facilities having only one room. The outdoor temperature and days between hunting and sampling affected the bacterial counts. High EB counts on the carcasses indicated a gut hit. Male gender was significantly more contaminated by EC and meat-borne pathogenic bacteria: Campylobacter spp., Salmonella spp., enteropathogenic Yersinia spp., stx-harbouring EC (STEC) and Listeria monocytogenes. STEC (28/200) and L. monocytogenes (20/200) were the most commonly detected bacteria by PCR. L. monocytogenes isolates of different sequence types (ST7, 18, 29, 37, 249, 412, 451 and 611) belonged to serotypes 1/2a (seven isolates) and 4b (three isolates). The virulence gene ail was detected in four Yersinia enterocolitica biotype 1A isolates and one Yersinia kristensenii isolate. The bacterial counts on the moose and deer carcasses varied highly, and more attention should be paid to hunting hygiene and training of hunters. Game meat may be a source of meat-borne pathogens, and close attention should therefore be paid when handling and preparing game.