1. Investigación

Listeria monocytogenes is a major human and animal foodborne patho-gen. However, data from environmental reservoirs remain scarce. Here, we usedwhole-genome sequencing to characterize Listeria species isolates recovered over 1year from wild animals in their natural habitats in Spain. Three different Listeria spp.(L. monocytogenes [n = 19], Listeria ivanovii subsp. londoniensis [n = 4], and Listeriainnocua [n = 3]) were detected in 23 animal tonsils (9 deer, 14 wild boars) and 2feeding troughs. No Listeria species was detected in feces. L. monocytogenes wasdetected in tonsils of 44.4% (8 out of 18) of deer and 40.7% (11 out of 27) of wildboars. L. monocytogenes isolates belonged to 3 different core genome multilocussequence typing (cgMLST) types (CTs) of 3 distinct sublineages (SL1, SL387, andSL155) from lineages I and II. While cgMLST type L1-SL1-ST1-CT5279 (IVb; clonalcomplex 1 [CC1]) occurred only in one animal, types L1-SL387-ST388-CT5239 (IVb;CC388) and L2-SL155-ST155-CT1170 (IIa; CC155) were retrieved from multiple ani-mals. In addition, L1-SL387-ST388-CT5239 (IVb; CC388) isolates were collected 1 yearapart, revealing their long-term occurrence within the animal population and/orenvironmental reservoir. The presence of identical L. monocytogenes strains in deerand wild boars suggests contamination from a common food or environmentalsource, although interhost transmission cannot be excluded. Pathogenicity islandsLIPI-1, LIPI-3, and LIPI-4 were present in 100%, 5%, and 79% of the L. monocytogenesisolates, respectively, and all L. monocytogenes lineage II isolates (n = 3) carried SSI-1stress islands. This study highlights the need for monitoring L. monocytogenes envi-ronmental contamination and the importance of tonsils as a possible L. monocyto-genes intrahost reservoir.

Vultures are nature's most successful scavengers, feeding on the carcasses of dead animals present in the field. Availability of domestic carrion has been unstable due to rapidly changing agro-grazing economies and increasing sanitary regulations that may require burial or burning of livestock carcasses. Thus, several griffon vulture (Gyps fulvus) recoveries are based on European legislation that guarantees the animals' welfare, avoids intense persecution of the vultures and allows the feeding of threatened wildlife in supplementary feeding stations (SFS). However, in recent years, many studies have speculated on the likelihood that avian scavengers may be infected by feeding on pig carcasses at SFS from intensive livestock. In this context, the present study evaluated whether free-living griffon vultures and pig farms share zoonotic Salmonella strains to test the hypothesis that vulture are infected during consumption of carcasses provided at SFS. Here, the occurrence, serotypes and genomic DNA fingerprinting (phage typing and pulsed-field gel electrophoresis) of isolated strains were carried out in griffon vultures and pig farms authorised to provided carcasses at SFS in Castellón province (eastern Spain). The bacteriological analyses revealed that 21.1% of vultures and 14.5% for pig farms samples tested were Salmonella-positive. Monophasic S. typhimurium 1,4,[5],12:i:- was the most frequently isolated serovar. Comparison of Salmonella strains isolated from vultures and pig farms revealed that monophasic S. typhimurium 1,4,[5],12:i:-, S. Derby and S. Rissen strains were highly genetically homogeneous (similar DNA fingerprint). In conclusion, the current study indicates that free-living griffon vultures and pig farms that provide the carcasses at SFS share several zoonotic Salmonella strains. On this basis, and although transmission could be bidirectional, our result seems to corroborate the pig carcasses-to-vulture transmission and cross-infection at SFS. As an immediate Salmonella control strategy in wild avian scavengers, we suggest the implementation of a programme to guarantee that solely pig carcasses from Salmonella-free farms arrive at SFS.

Staphylococcus aureus is an opportunistic multi-host pathogen that threatens both human and animal health. Animals can act as a reservoir of S. aureus for humans, but very little is known about wild animals’ epidemiological role. Therefore, in this study, we performed a genomic characterization of S. aureus isolates from wildlife, hunters, and their auxiliary hunting animals of Eastern Spain. Of 20 different species, 242 wild animals were examined, of which 28.1% were S. aureus carriers. The common genet, the Iberian ibex, and the European hedgehog were the species with the highest S. aureus carriage. We identified 30 different sequence types (STs), including lineages associated with wild animals such as ST49 and ST581, multispecies lineages such as ST130, ST398, and ST425, and lineages commonly isolated from humans, including ST1 and ST5. The hunters and the single positive ferret shared ST5, ST398, or ST425 with wild animals. In wildlife isolates, the highest resistance levels were found for penicillin (32.8%). For virulence factors, 26.2% of them carried superantigens, while 14.8% harbored the immune evasion cluster (IEC), which indicates probable human origin. Our findings suggest that wild animals are a reservoir of clinically relevant genes and lineages that could have the potential to be transmitted to humans. These data support the notion that wildlife surveillance is necessary to better understand the epidemiology of S. aureus as a pathogen that circulates among humans, animals, and the environment.

The Iberian hare (Lepus granatensis) is a popular small game species in the Iberian Peninsula, and it has never been reported to be affected by sarcoptic mange. An adult female Iberian hare with overt skin lesions on forelimbs and ventral thorax, suggestive of sarcoptic mange, was culled in Quart de les Valls municipality in the Valencian Community, Spain, in 2019. Skin scrapings were digested in 10% KOH solutions to confirm the presence of mites. Ten Sarcoptes microsatellite markers were used to characterize the genetic structure of mites obtained from the hare, and from sympatric and allopatric wild rabbits (Oryctolagus cuniculus) and red foxes (Vulpes vulpes). A total of 56 alleles were counted across the 10 microsatellite loci. Six private alleles were found at four loci (Sarms 33, 38, 41, 45). The multivariate analysis characterized three main clusters, corresponding to mites collected on foxes originating from Catalonia, foxes from Valencia and the hare plus wild rabbits. To our knowledge, this is the first reported case of sarcoptic mange in the Iberian hare. The origin was molecularly traced back to contacts with endemically infected wild rabbits. We encourage further investigations on cross-transmissibility of S. scabiei between wild rabbits and the diverse representatives of Lepus spp.