Pathological and virological insights from an outbreak of European brown hare syndrome in the Italian hare (Lepus corsicanus).

European brown hare syndrome (EBHS) is a highly contagious and fatal viral disease, mainly affecting European brown hares (Lepus europaeus). The etiological agent, EBHS virus (EBHSV), belongs to the Lagovirus genus within the Caliciviridae family. The Italian hare (Lepus corsicanus) is endemic to Central-Southern Italy and Sicily and is classified as a vulnerable species. L. corsicanus is known to be susceptible to EBHS, but virological data available is scarce due to the few cases detected so far. In this study, we describe the occurrence of EBHS in two free-ranging L. corsicanus, found dead in a protected area of Central Italy. The two hares were identified as L. corsicanus using phenotypic criteria and confirmed through mitochondrial DNA analysis. Distinctive EBHS gross lesions were observed at necropsy and confirmed by subsequent histological examination. EBHSV was detected in the livers of the two animals initially using an antigen detection ELISA, followed by an EBHSV-specific reverse transcription-PCR, thus confirming the viral infection as the probable cause of death. The EBHS viruses detected in the two hares were identical, as based on blast analysis performed for the VP60 sequences and showed 98.86% nucleotide identity and 100% amino acid identity with strain EBHSV/GER-BY/EI97.L03477/2019, isolated in Germany in 2019. Phylogenetic analysis places our virus in group B, which includes strains that emerged after the mid-1980s. This study supports previous reports of EBHS in L. corsicanus and further expands the knowledge of the pathological and virological characteristics of the etiological agent. The ability of EBHSV to cause a fatal disease in the Italian hare represents a serious threat to the conservation of this vulnerable species, especially in populations kept in enclosed protected areas.

Brucella ceti Infection in Striped Dolphins from Italian Seas: Associated Lesions and Epidemiological Data.

Brucella ceti infections have been increasingly reported in cetaceans. In this study, we analyzed all cases of B. ceti infection detected in striped dolphins stranded along the Italian coastline between 2012 and 2021 (N = 24). We focused on the pathogenic role of B. ceti through detailed pathological studies, and ad hoc microbiological, biomolecular, and serological investigations, coupled with a comparative genomic analysis of the strains. Neurobrucellosis was observed in 20 animals. The primary histopathologic features included non-suppurative meningoencephalitis (N = 9), meningitis (N = 6), and meningoencephalomyelitis (N = 5), which was also associated with typical lesions in other tissues (N = 8). Co-infections were detected in more than half of the cases, mostly involving Cetacean Morbillivirus (CeMV). The 24 B. ceti isolates were assigned primarily to sequence type 26 (ST26) (N = 21) and, in a few cases, ST49 (N = 3). The multilocus sequence typing (cgMLST) based on whole genome sequencing (WGS) data showed that strains from Italy clustered into four genetically distinct clades. Plotting these clades onto a geographic map suggests a link between their phylogeny and the topographical distribution. These results support the role of B. ceti as a primary neurotropic pathogen for striped dolphins and highlight the utility of WGS data in understanding the evolution of this emerging pathogen.

Genomics Insight into cfr-Mediated Linezolid-Resistant LA-MRSA in Italian Pig Holdings.

The cfr genes encode for a 23S rRNA methyltransferase, conferring a multiresistance phenotype to phenicol, lincosamide, oxazolidinone, pleuromutilin, and streptogramin A antibiotics. These genes have been described in staphylococci, including methicillin-resistant Staphylococcus aureus (MRSA). In this study, we retrospectively performed an in-depth genomic characterisation of three cfr-positive, multidrug-resistant (MDR) livestock-associated (LA) MRSA clonal complexes (CCs) 1 and 398 detected in different Italian pig holdings (2008-2011) during population studies on Italian livestock (2008-2014). We used a combined Illumina and Oxford Nanopore Technologies (ONT) whole genome sequencing (WGS) approach on two isolates (the 2008 CC1 and the 2010 CC398 isolates, but not the 2011 CC1 isolate). Interestingly, the three isolates presented different cfr variants, with only one displaying a linezolid-resistant phenotype. In isolate 2008 CC1, the cfr gene was identified within a Tn558 composite transposon-like structure flanked by IS elements located on a novel 44,826 bp plasmid. This represents the first report of CC1 LA-MRSA harbouring the cfr gene in its functional variant. Differently, cfr was chromosomally located in isolate 2010 CC398. Our findings have significant public health implications, confirm the need for the continuous genomic surveillance of cfr-positive zoonotic LA-MRSA, and backdate cfr presence in LA-MRSA from Italian pigs to at least 2008.

Evidence of structural rearrangements in ESBL-positive pESI(like) megaplasmids of S.Infantis.

The increasing prevalence of pESI(like)-positive, multidrug-resistant (MDR) S. Infantis in Europe is a cause of major concern. As previously demonstrated, the pESI(like) megaplasmid is not only a carrier of antimicrobial resistant (AMR) genes (at least tet, dfr, and sul genes), but also harbours several virulence and fitness genes, and toxin/antitoxin systems that enhance its persistence in the S. Infantis host. In this study, five prototype pESI(like) plasmids, of either CTX-M-1 or CTX-M-65 ESBL-producing strains, were long-read sequenced using Oxford Nanopore Technology (ONT), and their complete sequences were resolved. Comparison of the structure and gene content of the five sequenced plasmids, and further comparison with previously published pESI(like) sequences, indicated that although the sequence of such pESI(like) 'mosaic' plasmids remains almost identical, their structures appear different and composed of regions inserted or transposed after different events. The results obtained in this study are essential to better understand the plasticity and the evolution of the pESI(like) megaplasmid, and therefore to better address risk management options and policy decisions to fight against AMR and MDR in Salmonella and other food-borne pathogens. Graphical representation of the pESI-like plasmid complete sequence (ID 12037823/11). Block colours indicate the function of the genes: red: repB gene; pink: class I integrons (IntI); yellow; mobile elements; blue: resistance genes; green: toxin/anti-toxin systems; grey: mer operon; light green: genes involve in conjugation.

Genomics insights into a Mycobacterium pinnipedii isolate causing tuberculosis in a captive South American sea lion (Otaria flavescens) from Italy.

Tuberculosis (TB) affects humans and other animals, and it is caused by bacteria within the Mycobacterium tuberculosis complex (MTBC). In this study, we report the characterisation of Mycobacterium pinnipedii that caused a TB case in a sea lion (Otaria flavescens) kept in an Italian zoo. The animal died due to severe, progressive disorders involving the respiratory and gastro-enteric systems and the skin. At necropsy, typical gross lesions referable to a TB generalised form were found. In particular, nodular granulomatous lesions were detected in the lungs and several lymph nodes, and colonies referable to Mycobacterium spp. were isolated from lung, mesenteric, and mediastinal lymph nodes. The isolate was identified by PCR as a MTBC, had a spoligotype SB 1480 ("seal lineage"), and was characterised and characterised by whole-genome sequencing analysis confirming that the MTBC involved was M. pinnipedii. The analysis of the resistome and virulome indicated the presence of macrolide and aminoglycoside resistance genes intrinsic in M. tuberculosis [erm-37 and aac(2')-Ic] and confirmed the presence of the region of difference 1 (RD1), harbouring the esxA and esxB virulence genes, differently from its closest taxon, M. microti. As for other MTCB members, M. pinnipedii infection can spill over into non-pinniped mammalian species; therefore, zoological gardens, veterinary practitioners, and public health officers should be aware of the hazard posed by tuberculosis from marine mammals. Since the isolate under study, as well as all available genomes of M. pinnipedii investigated in this study retains almost all the M. tuberculosis virulence genes, it could indeed cause infection, lesions, and disease in other animal species, including humans.

The hazard of carbapenemase (OXA-181)-producing Escherichia coli spreading in pig and veal calf holdings in Italy in the genomics era: Risk of spill over and spill back between humans and animals.

Carbapenemase-producing Enterobacterales (CPE) are considered a major public health issue. In the frame of the EU Harmonized AMR Monitoring program conducted in Italy in 2021, 21 epidemiological units of fattening pigs (6.98%; 95% CI 4.37-10.47%; 21/301) and four epidemiological units of bovines <12 months (1.29%; 95% CI 0.35-3.27%, 4/310) resulted positive to OXA-48-like-producing E. coli (n = 24 OXA-181, n = 1 OXA-48). Whole Genome Sequencing (WGS) for in-depth characterization, genomics and cluster analysis of OXA-181-(and one OXA-48) producing E. coli isolated, was performed. Tracing-back activities at: (a) the fattening holding of origin of one positive slaughter batch, (b) the breeding holding, and (c) one epidemiologically related dairy cattle holding, allowed detection of OXA-48-like-producing E. coli in different units and comparison of further human isolates from fecal samples of farm workers. The OXA-181-producing isolates were multidrug resistant (MDR), belonged to different Sequence Types (STs), harbored the IncX and IncF plasmid replicons and multiple virulence genes. Bioinformatics analysis of combined Oxford Nanopore Technologies (ONT) long reads and Illumina short reads identified bla OXA-181 as part of a transposon in IncX1, IncX3, and IncFII fully resolved plasmids from 16 selected E. coli, mostly belonging to ST5229, isolated during the survey at slaughter and tracing-back activities. Although human source could be the most likely cause for the introduction of the bla OXA-181-carrying IncX1 plasmid in the breeding holding, concerns arise from carbapenemase OXA-48-like-producing E. coli spreading in 2021 in Italian fattening pigs and, to a lesser extent, in veal calf holdings.

Technical specifications for a baseline survey on the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in pigs.

The European Commission requested scientific and technical assistance in the preparation of a EU-wide baseline survey protocol for a European Union (EU) coordinated monitoring programme on the prevalence of methicillin-resistant Staphylococcus Aureus (MRSA) in pigs. The objective of the survey is to estimate the MRSA prevalence in batches of fattening pigs at slaughter at both European and national levels, with a 95% level of confidence and a level of precision of 10% considering an expected prevalence of 50%. The survey protocol defines the target population, the sample size for the survey, sample collection requirements, the analytical methods (for isolation, identification, phenotypic susceptibility testing and further genotypic testing of MRSA isolates), the data reporting requirements and the plan of analysis. The samples are to be analysed according to the laboratory protocols available on the European Union Reference Laboratory (EURL-AR) website. Generalised linear models will be used to estimate proportion (with 95% confidence intervals) of batches of slaughter pigs tested positive to MRSA. The necessary data to be reported by the EU Member States to support this baseline survey are presented in three data models. The results of the survey should be reported using the EFSA data collection framework.

First report of the zoonotic nematode Baylisascaris procyonis in non-native raccoons (Procyon lotor) from Italy.

Baylisascaris procyonis is a nematode parasite of the raccoon (Procyon lotor), and it can be responsible for a severe form of larva migrans in humans. This parasite has been reported from many countries all over the world, after translocation of its natural host outside its native geographic range, North America. In the period between January and August 2021, 21 raccoons were cage-trapped and euthanized in Tuscany (Central Italy), in the context of a plan aimed at eradicating a reproductive population of this non-native species. All the animals were submitted for necroscopic examination. Adult ascariids were found in the small intestine of seven raccoons (prevalence 33.3%). Parasites have been identified as B. procyonis based on both morphometric and molecular approaches. The aim of the present article is to report the first finding of this zoonotic parasite from Italy, highlighting the sanitary risks linked to the introduction of alien vertebrate species in new areas.

Plasmodium matutinum Causing Avian Malaria in Lovebirds (Agapornis roseicollis) Hosted in an Italian Zoo.

Avian malaria is a worldwide distributed, vector-born disease of birds caused by parasites of the order Haemosporida. There is a lack of knowledge about the presence and pathogenetic role of Haemosporida in Psittacidae. Here we report a case of avian malaria infection in lovebirds (Agapornis roseicollis), with the genetic characterization of the Plasmodium species involved. The birds were hosted in a zoo located in Italy, where avian malaria cases in African penguins (Spheniscus demersus) were already reported. Animals (n = 11) were submitted for necropsy after sudden death and were subjected to further analyses including histopathology, bacteriology, and PCR for the research of haemosporidians. Clinical history, gross lesions and histopathological observation of schizonts, together with positive PCR results for Plasmodium spp., demonstrated that avian malaria was the cause of death for one animal and the possible cause of death for the other nine. The sequences obtained were compared using BLAST and analyzed for similarity to sequences available at the MalAvi database. Genetic analyses demonstrated a 100% nucleotide identity to Plasmodium matutinum LINN1 for all the obtained sequences. To our knowledge, this is the first report describing avian malaria in lovebirds.

Plasmodium matutinum Transmitted by Culex pipiens as a Cause of Avian Malaria in Captive African Penguins (Spheniscus demersus) in Italy.

Avian malaria is a parasitic disease of birds caused by protozoa belonging to the genus Plasmodium, within the order Haemosporida. Penguins are considered particularly susceptible, and outbreaks in captive populations can lead to high mortality. We used a multidisciplinary approach to investigate the death due to avian malaria, occurred between 2015 and 2019, in eight African penguins (Spheniscus demersus) kept in two Italian zoos located in central Italy, and situated about 30 km apart. We also provided information about the presence and circulation of Plasmodium spp. in mosquitoes in central Italy by sampling mosquitoes in both zoos where penguin mortalities occurred. In the eight dead penguins, gross and histopathological lesions were consistent with those previously observed by other authors in avian malaria outbreaks. Organs from dead penguins and mosquitoes collected in both zoos were tested for avian malaria parasites by using a PCR assay targeting the partial mitochondrial conserved region of the cytochrome b gene. Identification at species level was performed by sequencing analysis. Plasmodium matutinum was detected in both dead penguins and in mosquitoes (Culex pipiens), while Plasmodium vaughani in Culex pipiens only. Parasites were not found in any of the PCR tested Aedes albopictus samples. Based on our phylogenetic analysis, we detected three previously characterized lineages: Plasmodium matutinum LINN1 and AFTRU5, P. vaughani SYAT05. In Culex pipiens we also identified two novel lineages, CXPIP32 (inferred morphospecies Plasmodium matutinum) and CXPIP33 (inferred morphospecies P. vaughani). Significantly, LINN1 and AFTRU5 were found to be associated to penguin deaths, although only LINN1 was detected both in penguins (along the years of the study) and in Culex pipiens, while AFTRU5 was detected in a single penguin dead in 2017. In conclusion, in our study Plasmodium matutinum was found to cause avian malaria in captive penguins kept in Europe, with Culex pipiens being its most probable vector. Our results are in agreement with previous studies suggesting that Culex pipiens is one of the main vectors of Plasmodium spp. in Europe and the Northern Hemisphere. Zoos maintaining captive penguins in temperate areas where Culex pipiens is abundant should be well aware of the risks of avian malaria, and should put every effort to prevent outbreaks, in particular during the periods when the number of vectors is higher.

Livestock-Associated Methicillin-Resistant Staphylococcus aureus and Related Risk Factors in Holdings of Veal Calves in Northwest Italy.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) have emerged worldwide as zoonotic pathogens. Data on LA-MRSA in veal calf production in Italy are lacking; the aim of this survey was to fill current knowledge gaps in its prevalence and characteristics. Between February 2012 and January 2013 nasal swabs were taken from 1650 three- to six-month-old veal calves on 55 farms in Piedmont (northwest Italy), including gathering-related epidemiological data. S. aureus were screened for methicillin resistance by phenotypic and molecular (mecA gene detection) methods. MRSA were further genotyped by multilocus sequence typing. About 30% of the herds tested positive for MRSA: three different clonal complexes (CC398, CC97, and CC1) and staphylococcal cassette chromosome mec types (IVa, IVb, and V) were detected. Multilevel logistic regression model indicated poor cleaning, importation from Austria, and animal age as risk factors and coagulase-negative staphylococci colonization as a predictive factor for the occurrence of MRSA. The detection of CCs circulating in pigs and dairy cattle in Italy underscores the ability of the LA-MRSA clones to spread among animal production systems. In addition to maintaining preventive control measures for human health, better cleaning procedures need to be implemented, especially after new calves have been introduced into the herd.

Molecular epidemiology of methicillin-resistant Staphylococcus aureus from dairy farms in North-eastern Italy.

Transmission of Staphylococcus aureus along the dairy production chain is an emerging public health problem with human, veterinary, and food safety issues. The prevalence of multidrug-resistant, particularly methicillin-resistant S. aureus (MRSA), has steadily increased in several European countries. In this study, the prevalence of S. aureus in raw cow milk and farm workers was investigated, and the trajectories of MRSA transmission at the primary stage of the dairy chain were assessed. To this purpose, a longitudinal survey was conducted in 618 dairy farms in two contiguous regions with high livestock density in North-eastern Italy. S. aureus contamination of bulk tank milk (BTM) was observed in more than 80% of farms, while MRSA prevalence was 3.6% and 15.9% in BTM and farm workers, respectively. The majority of MRSA isolates from both BTM and farm workers were assigned to ST398, and showed a worrisome multidrug-resistant phenotype. Enterotoxin and Panton-Valentine leukocidin genes were detected in 11.5% and 4.9% of MRSA isolates from both sources. Nearly all MRSA isolates from workers belonged to the same epidemiological type as BTM isolates from the corresponding farm, denoting a bidirectional MRSA transmission pattern. A focus on the ST398 spa type t899 MRSA lineage in the Italian livestock system highlighted the presence of two major clusters whose dissemination was likely facilitated by the selective pressure imposed by antimicrobial use in animal farming. Our findings emphasize the need for continuous monitoring of MRSA along the dairy production chain, not only to avoid transmission between animals and exposed workers, but also to contain the risk of raw milk and dairy product contamination by multidrug resistant and toxigenic strain.

Novel IncFII plasmid harbouring blaNDM-4 in a carbapenem-resistant Escherichia coli of pig origin, Italy.

OBJECTIVES: To the best of our knowledge, we describe the first evidence in Europe of an MDR, blaNDM-4-positive Escherichia coli isolated from a food-producing animal, harboured by a novel IncFII plasmid of which we report the complete sequence. METHODS: One blaNDM-4-positive E. coli isolated in 2019 from the caecal contents of a fattening pig in Italy was in-depth characterized by combined bioinformatic analysis of Oxford Nanopore long reads and Illumina short reads, for in silico typing, determination of the blaNDM-4 genetic context and full reconstruction of the blaNDM-4-carrying plasmid. RESULTS: The isolate belonged to ST641 and to the genoserotype O108:H23 and tested positive for different virulence genes and plasmid replicons. The MDR phenotype of resistance to all ß-lactams, carbapenems, sulfamethoxazole and trimethoprim was mediated by blaTEM-1B, blaNDM-4, sul1/sul3 and dfrA12, respectively. The blaNDM-4 gene was harboured by a novel 53 043 bp IncFII plasmid (pMOL412_FII) composed of four main genetic regions, including an MDR region (MRR-NDM-4) of 16 kb carrying blaNDM-4 and several antimicrobial resistance genes located in a class 1 integron. pMOL412_FII was closely related to another ∼90.3 kb plasmid (pM109_FII) harbouring blaNDM-4 in an E. coli isolated from a human patient in Myanmar. CONCLUSIONS: To the best of our knowledge, we have identified for the first time in Europe an NDM-producing Enterobacterales in livestock and resolved the complete sequence of the novel pMOL412_FII plasmid harbouring blaNDM-4 in an MRR. A global One Health approach, comparing genomic data from different sources and geographical areas, may help to trace back and control possible plasmid-borne carbapenemase gene transmission between animals and humans and along the food chain at international level.

Molecular epidemiology of Salmonella Infantis in Europe: insights into the success of the bacterial host and its parasitic pESI-like megaplasmid.

Salmonella Infantis is one of the five serovars most frequently causing human salmonellosis in Europe, mainly associated with poultry. A clone harbouring a conjugative plasmid of emerging S. Infantis (pESI)-like megaplasmid, carrying multidrug resistant (MDR) and extended-spectrum beta-lactamases (ESBL) genes, has spread in the Italian broiler chicken industry also causing human illness. This work is aimed at elucidating the molecular epidemiology of S. Infantis and pESI-like in Europe using whole-genome sequencing and bioinformatics analysis, and to investigate the genetic relatedness of S. Infantis clones and pESI-like from animals, meat, feed and humans provided by institutions of nine European countries. Two genotyping approaches were used: chromosome or plasmid SNP-based analysis and the minimum spanning tree (MST) algorithm based on core-genome multilocus sequence typing (cgMLST). The European S. Infantis population appeared heterogeneous, with different genetic clusters defined at core-genome level. However, pESI-like variants present in 64.1 % of the isolates were more genetically homogeneous and capable of infecting different clonal lineages in most of the countries. Two different pESI-like with ESBL genes (n=82) were observed: blaCTX-M-1-positive in European isolates and blaCTX-M-65-positive in American isolates (study outgroup). Both variants had toxin-antitoxin systems, resistance genes towards tetracyclines, trimethoprim, sulphonamides and aminoglycosides, heavy metals (merA) and disinfectants (qacEΔ). Worryingly, 66 % of the total isolates studied presented different gyrA chromosomal point mutations associated with (fluoro)quinolone resistance (MIC range 0.125-0.5 mg/L), while 18 % displayed transferable macrolide resistance mediated by mph, mef and erm(B) genes. Proper intervention strategies are needed to prevent further dissemination/transmission of MDR S. Infantis and pESI-like along the food chain in Europe.

Antimicrobial Usage and Resistance in Companion Animals: A Cross-Sectional Study in Three European Countries.

Companion animals have been described as potential reservoirs of antimicrobial resistance (AMR), however data remain scarce. Therefore, the objectives were to describe antimicrobial usage (AMU) in dogs and cats in three European countries (Belgium, Italy, and The Netherlands) and to investigate phenotypic AMR. A questionnaire and one fecal sample per animal (n = 303) were collected over one year and AMU was quantified using treatment incidence (TI). Phenotypic resistance profiles of 282 Escherichia coli isolates were determined. Nineteen percent of the animals received at least one antimicrobial treatment six months preceding sampling. On average, cats and dogs were treated with a standard daily dose of antimicrobials for 1.8 and 3.3 days over one year, respectively. The most frequently used antimicrobial was amoxicillin-clavulanate (27%). Broad-spectrum antimicrobials and critically important antimicrobials for human medicine represented 83% and 71% of the total number of treatments, respectively. Resistance of E. coli to at least one antimicrobial agent was found in 27% of the isolates. The most common resistance was to ampicillin (18%). Thirteen percent was identified as multidrug resistant isolates. No association between AMU and AMR was found in the investigated samples. The issue to address, regarding AMU in companion animal, lies within the quality of use, not the quantity. Especially from a One-Health perspective, companion animals might be a source of transmission of resistance genes and/or resistant bacteria to humans.

Unexpected human cases of cutaneous anthrax in Latium region, Italy, August 2017: integrated human-animal investigation of epidemiological, clinical, microbiological and ecological factors.

On 31 August, a veterinarian and a farmworker were hospitalised for skin lesions. Both had been exposed to a dead cow on 19 August on a farm near Rome, where eight further cattle died of confirmed anthrax later the same month. At admission, the first case showed a black depressed eschar and another smaller lesion on one hand. The second case presented deep infection of the skin, with involvement of both arms. Anthrax diagnosis was confirmed by detection of B. anthracis DNA in eschar fragments from both patients. T-cell specific immunity was studied by flow cytometry and Elispot assay after stimulation with B. anthracis secretome in blood samples collected from Case 1. Immunoglobulin production was detected by complement fixation assay. In Case 1, specific CD4+ T-cell activation was detected, without antibody production. Specific antibodies were detected only in the second patient with severe cutaneous illness. Both patients recovered. The two human anthrax cases were epidemiologically linked, but anthrax was not suspected at admission in either case. The veterinarian had initially unrecognised professional exposure and the exposed farmworker did initially not report exposure to affected animals. A One Health strategy integrating human and animal investigations was essential to confirm the diagnosis.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) spa type t127, Sequence Type (ST)1, quickly spreads and persists among young pigs.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) clones other than Clonal Complex (CC)398, as CC1, have been isolated in pigs in some countries, and appeared to be prevalent in Italy. The aim of this study was to evaluate the capability of Sequence Type (ST)1, CC1, LA-MRSA clone to colonize and to be transmitted among piglets. Eighteen caesarean-derived/colostrum-deprived piglets of 35 days of age were assigned randomly to three groups: four seeder piglets were contaminated with a spa type t127, ST1, SCCmec V, MRSA (Group A), 10 MRSA-negative piglets were exposed to Group A after 2 days post-contamination, dpc (Group B) and 4 piglets were used as control group (Group C). Piglets were evaluated until 44 dpc (Group A) or at 42 days post-exposure, dpe (Group B) and then euthanized and necropsied. All nasal and skin cultures of Group A resulted MRSA-positive throughout the experiment starting from two dpc, while Group C tested always MRSA-negative. At first sampling, all Group B piglets became positive and remained positive throughout the experiment. This is the first colonization/transmission study with a CC1 LA-MRSA in pigs. The results add further knowledge on the ability of CC1 LA-MRSA to colonize pigs, and on colonization/transmission patterns, both suggesting good host adaptation.

Systemic tuberculosis by MYCOBACTERIUM BOVIS in a free-ranging MARSICAN brown bear (URSUS ARCTOS MARSICANUS): a Case report.

BACKGROUND: Mycobacterium bovis is known to have a wide host range and has been isolated from numerous free-ranging wildlife species, carnivores included. In bears, M. bovis has been previously reported only from a culture of pooled lymph nodes of a black bear (Ursus americanus) in the absence of lesions. The aims of this study were to describe gross and microscopic pathological findings of M. bovis tuberculosis in a deceased Marsican brown bear (Ursus arctos marsicanus). CASE PRESENTATION: In March 2014, an adult female Marsican brown bear was found in the Abruzzo, Lazio and Molise National Park (Italy) showing severe non-specific clinical signs. The animal died soon after its discovery and the carcass was submitted to post-mortem examination to identify the cause of death. The bear was diagnosed with a severe Mycobacterium bovis infection, with both pathological and microbiological aspects suggesting ongoing generalization. A presumptive diagnosis of mycobacterial infection was initially made based on gross findings. Histopathology showed the presence of acid-fast bacilli in all sampled tissues along with poorly organized granulomatous lesions. Slow-growing Mycobacterium sp. was isolated from multiple organs (intestine, mesenteric lymph nodes, liver, spleen, lung and kidneys). The PCR and sequencing algorithm identified the Mycobacterium sp. isolate as M. bovis. Spoligotyping demonstrated that the M. bovis isolate belonged to spoligotype SB0120. CONCLUSIONS: This is the first report of lethal M. bovis tuberculosis infection in a free-ranging brown bear. This pathogen could have serious adverse effects in an endangered relic population such as the Marsican brown bear. Stricter application of health regulations in force, surveillance of M. bovis infections in wild ungulates and carnivore scavengers, along with dismissal of supplementary feeding points intended for cattle or wildlife, are warranted to control the presence of bovine tuberculosis in wild and domestic animals in protected areas.

Detection and isolation of Shiga Toxin-producing Escherichia coli (STEC) strains in caecal samples from pigs at slaughter in Italy.

Shiga toxin-producing Escherichia coli (STEC) strains are food-borne pathogens of public health concern. Despite ruminants are the most important reservoir, STEC human infections have also been attributed to pigs. We examined for the presence of STEC in 234 samples of swine caecal content collected during the year 2015 at Italian abattoirs in the framework of the harmonized monitoring of antimicrobial resistance (Decision 2013/652/EU). The presence of stx genes was detected in 122 (52.1%) samples, which were subsequently subjected to STEC isolation and characterization. The analysis of the 66 isolated STEC strains showed that the majority of the isolates (74.2%) possessed the stx2a gene subtype, in a few cases (16.7%) in combination with stx2b or stx2c. Only 25.8% of isolates possessed the stx2e subtype, typical of swine-adapted STEC. None of the isolates possessed the intimin-coding eae gene and the majority of them did not belong to serogroups commonly associated with human infections. The results of this study suggest that pigs can be considered as potential reservoir of certain STEC types.

Death of captive-bred vultures caused by flunixin poisoning in Italy.

Among non steroidal anti-inflammatory drugs (NSAIDs) diclofenac is considered the main cause for the decline of vulture populations in the Indian subcontinent since the '90 s. Chemical analysis showed high levels of flunixin (31,350 µg/kg) in beef which three captive Gyps vultures fed on, later dying with severe visceral gout. Levels in dead vultures' organs and tissues ranged from 4 to 38.5 µg/kg. The typical lesions and the concentrations found in beef indicate flunixin as the cause of death. This is the first observational study which correlates the concentration of flunixin in the meat ingested with that found in tissues of vultures.