Circoviridae Survey in Captive Non-Human Primates, Italy.

Circoviruses (CVs) and cycloviruses (CyVs), members of the family Circoviridae, have been identified only occasionally in non-human primates (NHPs). In this study, we investigated the presence and genetic features of these viruses in 48 NHPs housed in the Bioparco-Rome Zoological Garden (Italy) and in the Anima Natura Wild Sanctuary Semproniano (Grosseto, Italy), testing fecal, saliva, and serum samples with a broadly reactive consensus nested PCR able of amplifying a partial region of the replicase (Rep) gene of members of the family Circoviridae. Viral DNA was detected in a total of 10 samples, including a saliva swab and 9 fecal samples collected, respectively from five Japanese macaques (Macaca fuscata) and four mandrills (Mandrillus sphinx), with an overall prevalence of 18.7% (9/48). On genome sequencing, five strains revealed the highest nucleotide identity (98.3-98.6%) to a CyV strain (RI196/ITA) detected in the intestinal content of a Maltese wall lizard (Podarcis filfolensis) in Italy. Although the origin of the Italian NHP strains, genetically distant from previously detected NHP CyVs, is uncertain, our results also highlight that the virome of captive animals is modulated by the different dietary and environmental sources of exposure.

Feline Morbillivirus: Clinical Relevance of a Widespread Endemic Viral Infection of Cats.

Feline morbillivirus (FeMV) was first isolated in 2012 from stray cats in Hong Kong. It has been found in association with tubulointerstitial nephritis (TIN), the most common cause of feline chronic kidney disease (CKD). However, viral host spectrum and virus tropism go beyond the domestic cat and kidney tissues. The viral genetic diversity of FeMV is extensive, but it is not known if this is clinically relevant. Urine and kidney tissues have been widely tested in attempts to confirm associations between FeMV infection and renal disease, but samples from both healthy and sick cats can test positive and some cross-sectional studies have not found associations between FeMV infection and CKD. There is also evidence for acute kidney injury following infection with FeMV. The results of prevalence studies differ greatly depending on the population tested and methodologies used for detection, but worldwide distribution of FeMV has been shown. Experimental studies have confirmed previous field observations that higher viral loads are present in the urine compared to other tissues, and renal TIN lesions associated with FeMV antigen have been demonstrated, alongside virus lymphotropism and viraemia-associated lymphopenia. Longitudinal field studies have revealed persistent viral shedding in urine, although infection can be cleared spontaneously.

Microbial Community and Abundance of Selected Antimicrobial Resistance Genes in Poultry Litter from Conventional and Antibiotic-Free Farms.

In this study, a culture-independent approach was applied to compare the microbiome composition and the abundance of the antimicrobial resistance genes (ARGs) aadA2 for aminoglycosides, tet(A), tet(B), tet(K), and tet(M) for tetracyclines, and mcr-1 for colistin in broiler litter samples collected from conventional and antibiotic-free flocks located in Central Italy. A total of 13 flocks and 26 litter samples, collected at the beginning and at the end of each rearing cycle, were submitted to 16s rRNA sequence analysis and quantitative PCR for targeted ARGs. Firmicutes resulted in the dominant phylum in both groups of flocks, and within it, the Clostridia and Bacilli classes showed a similar distribution. Conversely, in antibiotic-free flocks, a higher frequency of Actinobacteria class and Clostridiaceae, Lactobacillaceae, Corynebacteriaceae families were reported, while in the conventional group, routinely treated with antibiotics for therapeutic purposes, the Bacteroidia class and the Enterobacteriaceae and Bacillaceae families were predominant. All investigated samples were found to be positive for at least one ARG, with the mean values of aadA2 and tet(A) the highest in conventional flocks by a significant margin. The results suggest that antibiotic use can influence the frequency of resistance determinants and the microbial community in poultry flocks, even though other environmental factors should also be investigated more deeply in order to identify additional drivers of antimicrobial resistance.

Feline Infectious Peritonitis: European Advisory Board on Cat Diseases Guidelines.

Feline coronavirus (FCoV) is a ubiquitous RNA virus of cats, which is transmitted faeco-orally. In these guidelines, the European Advisory Board on Cat Diseases (ABCD) presents a comprehensive review of feline infectious peritonitis (FIP). FCoV is primarily an enteric virus and most infections do not cause clinical signs, or result in only enteritis, but a small proportion of FCoV-infected cats develop FIP. The pathology in FIP comprises a perivascular phlebitis that can affect any organ. Cats under two years old are most frequently affected by FIP. Most cats present with fever, anorexia, and weight loss; many have effusions, and some have ocular and/or neurological signs. Making a diagnosis is complex and ABCD FIP Diagnostic Approach Tools are available to aid veterinarians. Sampling an effusion, when present, for cytology, biochemistry, and FCoV RNA or FCoV antigen detection is very useful diagnostically. In the absence of an effusion, fine-needle aspirates from affected organs for cytology and FCoV RNA or FCoV antigen detection are helpful. Definitive diagnosis usually requires histopathology with FCoV antigen detection. Antiviral treatments now enable recovery in many cases from this previously fatal disease; nucleoside analogues (e.g., oral GS-441524) are very effective, although they are not available in all countries.

Feline Injection-Site Sarcoma and Other Adverse Reactions to Vaccination in Cats.

Vaccine-associated adverse events (VAAEs), including feline injection-site sarcomas (FISSs), occur only rarely but can be severe. Understanding potential VAAEs is an important part of informed owner consent for vaccination. In this review, the European Advisory Board on Cat Diseases (ABCD), a scientifically independent board of feline medicine experts, presents the current knowledge on VAAEs in cats, summarizing the literature and filling the gaps where scientific studies are missing with expert opinion to assist veterinarians in adopting the best vaccination practice. VAAEs are caused by an aberrant innate or adaptive immune reaction, excessive local reactions at the inoculation site, an error in administration, or failure in the manufacturing process. FISS, the most severe VAAE, can develop after vaccinations or injection of other substances. Although the most widely accepted hypothesis is that chronic inflammation triggers malignant transformation, the pathogenesis of FISS is not yet fully understood. No injectable vaccine is risk-free, and therefore, vaccination should be performed as often as necessary, but as infrequently as possible. Vaccines should be brought to room temperature prior to administration and injected at sites in which FISS surgery would likely be curative; the interscapular region should be avoided. Post-vaccinal monitoring is essential.

Antibiotic Resistance in the Apennine Wolf (Canis lupus italicus): Implications for Wildlife and Human Health.

The Apennine wolf (Canis lupus italicus) is a subspecies of gray wolf that is widespread throughout Italy. Due to hunting and habitat loss, their population declined dramatically in the late 19th and early 20th centuries, but conservation efforts improved to restore the species to an estimated population of 3300 individuals. The presence of antibiotic-resistant bacteria in Apennine Wolf may pose a risk to its health and survival, as well as the health of other animals in its environment. In this study, we investigated the antibiotic resistance profiles of bacteria collected from Apennine wolves admitted to the Wildlife Research Center of Maiella National Park (Italy) in 2022. A total of 12 bacteria collected from four wolves were isolated and tested for susceptibility to antibiotics used in veterinary medicine and to critically important antibiotics for human health by means of the Vitek 2 system. All isolates were resistant to at least one antibiotic, and six bacteria were multidrug resistant to critically important antibiotics (third-generation cephalosporins, carbapenems and fluoroquinolones). The results of this pilot study have allowed for the characterization of resistant profiles in Escherichia coli, Enterococcus faecalis and other bacterial species not previously reported in Apennine wolves. Our findings provide important insights into antibiotic resistance in wildlife and its potential implications for the conservation of biodiversity and public health.

Whole Genome Sequencing of Escherichia coli and Enterococcus spp. in wildlife-livestock interface: a pilot study.

OBJECTIVES: This pilot study provides a multidisciplinary investigation to monitor livestock-wildlife interface. Ecological data, microbiological investigations, and whole genome sequencing were used to characterize eight bacterial isolates obtained from sympatric domestic and wild ruminants in Maiella National Park (Italy) in terms of genetic patterns of antimicrobial resistance. METHODS: Using selective culturing of fresh fecal samples of monitored and georeferenced populations of Apennine chamois, goats, red deer, and sheep, Escherichia coli, Enterococcus faecium, and Enterococcus faecalis isolates were isolated and subjected to minimum inhibitory concentration determination and whole genome sequencing. RESULTS: The analyzed isolates showed phenotypic and genotypic resistance to tetracycline and critically important antibiotics such as linezolid and carbapenems. Virulence genes related to biofilm regulation and Shiga toxins were also detected. Furthermore, serotypes related to nosocomial infections, harbouring plasmids recognized as important mobile resistance gene transmitters, were identified. CONCLUSIONS: This multidisciplinary pilot study represents a promising initial step to identify the environmental drivers and the transmission routes of antimicrobial resistance and virulence factors, providing new data on bacteria from rare and endangered species such as Apennine chamois.

Antibiotic-Resistant Bacteria Dissemination in the Wildlife, Livestock, and Water of Maiella National Park, Italy.

Antimicrobial resistance (AMR) is a global health concern that has been linked to humans, animals, and the environment. The One Health approach highlights the connection between humans, animals, and the environment and suggests that a multidisciplinary approached be used in studies investigating AMR. The present study was carried out to identify and characterize the antimicrobial resistance profiles of bacteria isolated from wildlife and livestock feces as well as from surface water samples in Maiella National Park, Italy. Ecological and georeferenced data were used to select two sampling locations, one where wildlife was caught within livestock grazing areas (sympatric group) and one where wildlife was caught outside of livestock grazing areas (non-sympatric group). Ninety-nine bacterial isolates from 132 feces samples and seven isolates from five water samples were collected between October and December 2019. The specimens were examined for species identification, antibiotic susceptibility and molecular detection of antibiotic resistance. Forty isolates were identified as Escherichia coli, forty-eight as Enterococcus spp., eight as Streptococcus spp. and ten as other gram-negative bacteria. Phenotypic antibiotic resistance to at least one antimicrobial agent, including some antibiotics that play a critical role in human medicine, was detected in 36/106 (33.9%, 95% CI: 25-43) isolates and multidrug resistance was detected in 9/106 isolates (8.49%, 95% CI: 3.9-15.5). In addition, genes associated with antibiotic resistance were identified in 61/106 (57.55%, 95% CI: 47.5-67) isolates. The samples from sympatric areas were 2.11 (95% CI: 1.2-3.5) times more likely to contain resistant bacterial isolates than the samples from non-sympatric areas. These data suggest that drug resistant bacteria may be transmitted in areas where wildlife and livestock cohabitate. This emphasizes the need for further investigations focusing on the interactions between humans, wildlife, and the environment, the results of which can aid in the early detection of emerging AMR profiles and possible transmission routes.

Antimicrobial Resistance Profile of Bacterial Isolates from Urinary Tract Infections in Companion Animals in Central Italy.

The available data on antimicrobial resistance in pets are limited compared to those collected for food-producing animals. Bacterial urinary tract infections are some of the most important indications for antimicrobial use in pets, and empiric antimicrobial treatments are often administered in the presence of clinical signs. In this study, the results obtained from the laboratory investigations carried out on dogs and cats with urinary tract infections coming from veterinary clinics and practices in Central Italy were evaluated to provide additional data concerning the bacterial urinary pathogens and their antimicrobial resistance patterns in pets. A total of 635 isolates were collected from urine samples. Escherichia coli was the most common species recovered in dogs and cats, followed by Proteus mirabilis and Enterococcus spp. Furthermore, it was possible to isolate bacteria not usually described in other studies concerning pets such as Pantoea dispersa, Raoultella ornithinolytica, and Pasteurella pneumotropica (also known as Rodentibacter pneumotropicus). Based on the antimicrobial susceptibility results, 472/635 (74.3%) isolates were resistant to at least one antibiotic and 285/635 (44.8%) isolates were classified as multidrug-resistant. Monitoring the antibiotic resistance profiles in pet infections is important not only for the public health implications, but also to collect data useful for the treatment of diseases in pets.

Antibiotic Resistance Genes Occurrence in Conventional and Antibiotic-Free Poultry Farming, Italy.

Antimicrobial resistance is a complex and widespread problem threatening human and animal health. In poultry farms, a wide distribution of resistant bacteria and their relative genes is described worldwide, including in Italy. In this paper, a comparison of resistance gene distribution in litter samples, recovered from four conventional and four antibiotic-free broiler flocks, was performed to highlight any influence of farming systems on the spreading and maintenance of resistance determinants. Conventional PCR tests, targeting the resistance genes related to the most used antibiotics in poultry farming, along with some critically important antibiotics for human medicine, were applied. In conventional farms, n. 10 out of n. 30 investigated genes were present in at least one sample, the most abundant fragments being the tet genes specific for tetracyclines, followed by those for aminoglycosides and chloramphenicol. All conventional samples resulted negative for colistin, carbapenems, and vancomycin resistance genes. A similar trend was observed for antibiotic-free herds, with n. 13 out of n. 30 amplified genes, while a positivity for the mcr-1 gene, specific for colistin, was observed in one antibiotic-free flock. The statistical analysis revealed a significant difference for the tetM gene, which was found more frequently in the antibiotic-free category. The analysis carried out in this study allowed us to obtain new data about the distribution of resistance patterns in the poultry industry in relation to farming types. The PCR test is a quick and non-expensive laboratory tool for the environmental monitoring of resistance determinants identifying potential indicators of AMR dissemination.

Vaccination and Antibody Testing in Cats.

Vaccines protect cats from serious diseases by inducing antibodies and cellular immune responses. Primary vaccinations and boosters are given according to vaccination guidelines provided by industry and veterinary organizations, based on minimal duration of immunity (DOI). For certain diseases, particularly feline panleukopenia, antibody titres correlate with protection. For feline calicivirus and feline herpesvirus, a similar correlation is absent, or less clear. In this review, the European Advisory Board on Cat Diseases (ABCD) presents current knowledge and expert opinion on the use of antibody testing in different situations. Antibody testing can be performed either in diagnostic laboratories, or in veterinary practice using point of care (POC) tests, and can be applied for several purposes, such as to provide evidence that a successful immune response was induced following vaccination. In adult cats, antibody test results can inform the appropriate re-vaccination interval. In shelters, antibody testing can support the control of FPV outbreaks by identifying potentially unprotected cats. Antibody testing has also been proposed to support decisions on optimal vaccination schedules for the individual kitten. However, such testing is still expensive and it is considered impractical to monitor the decline of maternally derived antibodies.

Molecular Detection and Phylogenetic Analysis of Canine Distemper Virus in Marsican Brown Bear (Ursus arctos marsicanus).

In this paper, we report the first molecular detection of the canine distemper virus in the Marsican brown bear (Ursus arctos marsicanus). Three subadults and one adult were live-trapped and checked for the main viral pathogens responsible for infectious diseases in this species. The four bears were found to be negative for all investigated viruses except for one, which resulted in a positive outcome for CDV by means of RT-PCR targeting fragments of viral N and H genes. The sequence analysis revealed the specificity of amplicons for the Europe Wildlife lineage of CDV, the same viral strain recovered from three foxes and two unvaccinated dogs coming from the same territories where the positive bear was captured. These results confirm the receptivity of Marsican brown bear for CDV, apparently without any pathological consequences for the positive animal, and suggest the presence in the studied area of a unique wild host-adapted lineage of the virus, able to spread in domestic animals, too. In this respect, continuous and specifically targeted surveillance systems are necessary in order to highlight any changes in the epidemiology of the infection in the territories where the Marsican brown bear lives, along with a more effective vaccination program for domestic dogs co-existing with this endangered species.

Microbial community and antimicrobial resistance in fecal samples from wild and domestic ruminants in Maiella National Park, Italy.

This study aimed to provide new insights about antimicrobial resistance genes abundance and microbial communities of wild and domestic ruminants in wildlife-livestock interface. In total, 88 fecal samples were recovered from Apennine chamois, red deer, goat, cattle and sheep, and were collected in pools. The populations under study were selected based on ecological data useful to define sympatric and non-sympatric populations. Samples were screened for commonly used in farms under study or critically important antimicrobial resistance genes (aadA2, TetA, TetB, TetK, TetM, mcr-1). The microbial community composition was found to be different based on the species and land use of animals under study. Indeed, it was mostly characterized by phyla Firmicutes in bovine, Bacteroidota in chamois and Proteobacteria in red deer. Additionally, positive correlations between antibiotic resistance genes and microbial taxa (e.g., Tet genes correlated with Firmicutes and Patescibacteria) were described. Of the antimicrobials investigated, the abundance of mcr-1 gene suggests the importance of monitoring the wildlife in order to detect the emerging resistance genes contamination in environment. This study provides new data that highlight the importance of multidisciplinary and uncultured study in order to describe the spreading of antimicrobial resistance and related contamination in the environment.

Vaccination of Immunocompromised Cats.

Immunocompromise is a common condition in cats, especially due to widespread infections with immunosuppressive viruses, such as feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV), but also due to chronic non-infectious diseases, such as tumours, diabetes mellitus, and chronic kidney disease, as well as treatment with immunosuppressive drugs, such as glucocorticoids, cyclosporins, or tumour chemotherapy. In this review, the European Advisory Board on Cat Diseases (ABCD), a scientifically independent board of experts in feline medicine from eleven European countries, discusses the current knowledge and rationale for vaccination of immunocompromised cats. So far, there are few data available on vaccination of immunocompromised cats, and sometimes studies produce controversial results. Thus, this guideline summarizes the available scientific studies and fills in the gaps with expert opinion, where scientific studies are missing. Ultimately, this review aims to help veterinarians with their decision-making in how best to vaccinate immunocompromised cats.

Calicivirus Infection in Cats.

Feline calicivirus (FCV) is a common pathogen in domestic cats that is highly contagious, resistant to many disinfectants and demonstrates a high genetic variability. FCV infection can lead to serious or even fatal diseases. In this review, the European Advisory Board on Cat Diseases (ABCD), a scientifically independent board of experts in feline medicine from 11 European countries, presents the current knowledge of FCV infection and fills gaps with expert opinions. FCV infections are particularly problematic in multicat environments. FCV-infected cats often show painful erosions in the mouth and mild upper respiratory disease and, particularly in kittens, even fatal pneumonia. However, infection can be associated with chronic gingivostomatitis. Rarely, highly virulent FCV variants can induce severe systemic disease with epizootic spread and high mortality. FCV can best be detected by reverse-transcriptase PCR. However, a negative result does not rule out FCV infection and healthy cats can test positive. All cats should be vaccinated against FCV (core vaccine); however, vaccination protects cats from disease but not from infection. Considering the high variability of FCV, changing to different vaccine strain(s) may be of benefit if disease occurs in fully vaccinated cats. Infection-induced immunity is not life-long and does not protect against all strains; therefore, vaccination of cats that have recovered from caliciviral disease is recommended.

Emerging Respiratory Viruses of Cats.

In recent years, advances in diagnostics and deep sequencing technologies have led to the identification and characterization of novel viruses in cats as protoparviruses and chaphamaparvoviruses, unveiling the diversity of the feline virome in the respiratory tract. Observational, epidemiological and experimental data are necessary to demonstrate firmly if some viruses are able to cause disease, as this information may be confounded by virus- or host-related factors. Also, in recent years, researchers were able to monitor multiple examples of transmission to felids of viruses with high pathogenic potential, such as the influenza virus strains H5N1, H1N1, H7N2, H5N6 and H3N2, and in the late 2019, the human hypervirulent coronavirus SARS-CoV-2. These findings suggest that the study of viral infections always requires a multi-disciplinary approach inspired by the One Health vision. By reviewing the literature, we provide herewith an update on the emerging viruses identified in cats and their potential association with respiratory disease.

Evidence of Linezolid Resistance and Virulence Factors in Enterococcus spp. Isolates from Wild and Domestic Ruminants, Italy.

The aim of this study was to evaluate the resistance patterns against selected critically and highly important antibiotics (quinupristin/dalfopristin, vancomycin, and linezolid) in 48 Enterococcus isolates obtained from wild (red deer and Apennine chamois) and domestic (cattle, sheep, and goats) ruminants living with varying degrees of sympatry in the protected area of Maiella National Park (central Italy). According to CLSI breakpoints, 9 out of 48 isolates (18.8%) showed resistance to at least one antibiotic. One Apennine chamois isolate was resistant to all tested antibiotics. The PCR screening of related resistance genes highlighted the occurrence of msrC or cfrD in seven Enterococcus resistant isolates. In addition, msrC and vanC genes were amplified in susceptible isolates. Specific sequences of virulence genes (gelE, ace, efa, asa1, and esp) related to pathogenic enterococci in humans were amplified in 21/48 isolates (43.75%), belonging mostly to wild animals (15/21; 71.42%). This is the first report of linezolid-resistant enterococci harboring virulence genes in Italian wildlife with special regard to the red deer and Apennine chamois species. The results allow us to evaluate the potential role of wild animals as indicators of antibiotic resistance in environments with different levels of anthropic pressure.

Resistance Patterns, mcr-4 and OXA-48 Genes, and Virulence Factors of Escherichia coli from Apennine Chamois Living in Sympatry with Domestic Species, Italy.

The aim of this study was to determine and characterize potential resistance mechanisms against selected Critically Important Antibiotics in Escherichia coli isolates collected from wild and domestic ruminants living in the Maiella National Park, in Central Italy. A total of 38 isolates were obtained from red deer, Apennine chamois, cattle, sheep, and goats grazing in lands with different levels of anthropic pressure. Antimicrobial susceptibility was determined by Minimal Inhibitory Concentration testing, showing phenotypic resistance to colistin, meropenem, or ceftazidime in 9 isolates along with one bacterial strain being resistant to three of the tested antibiotics. In addition, the biomolecular assays allowed the amplification of the genes conferring the colistin (mcr-4), the carbapenems (OXA-48), penicillins and cephalosporins (TEM, SHV, CMY-1, CMY-2) resistance. In order to describe the potential pathogenicity of isolates under study, virulence genes related to Shiga toxin-producing (STEC) and enteropathogenic (EPEC) pathovars were identified. This study is the first report of mcr-4 and OXA-48 genes in resistant E. coli harboring virulence genes in Italian wildlife, with special regard to Apennine chamois and red deer species. The multidisciplinary approach used in this study can improve the early detection of emerging antibiotic resistance determinants in human-animal-environment interfaces by means of wildlife monitoring.

Detection of antibodies against domestic cat hepadnavirus using baculovirus-expressed core protein.

A novel orthohepadnavirus (domestic cat hepadnavirus [DCH]) similar to human hepatitis B virus has been recently detected in serum and liver samples from domestic cats with chronic hepatitis and hepatocellular carcinoma. Molecular investigations by independent research groups around the world have revealed positivity rates ranging from 6.5% to 12.5% in blood samples and up to 14.0% in liver tissue. In this study, we screened an age-stratified collection of feline sera (n = 256) by using an antibody detection enzyme-linked immunosorbent assay based on the recombinant core antigen of DCH (DCHc). Specific antibodies (DCHc Abs) were detected with a prevalence of 25.0%. The DNA of DCH was detected in 35.9% (23/64) of seropositive cats and only in 1.0% (2/192) of seronegative animals. Based on the serological (IgG and IgM anti-DCHc) and virological status, the possible stages of DCH infection were predicted.

Detection and Characterization of Feline Calicivirus Associated with Paw and Mouth Disease.

Feline calicivirus (FCV) infection in cats can led to several diverse clinical presentations, ranging from mild upper respiratory signs to virulent systemic disease. Herein, we report a paw and mouth disease case in a 7-year-old household cat due to an FCV infection. An asymptomatic cat living in the same household was also infected with FCV. Clinical and pathological investigations were combined with the molecular and phenotypical characterization of the FCV strains. The RNA of the FCV was detected using qualitative and quantitative reverse transcription (RT)-PCR assays, and FCV antigen was confirmed by immunohistochemistry. After the whole genome analysis, the strains detected in the two cats appeared to be genetically diverse from FCVs previously detected in association with paw and mouth disease and with virulent systemic disease. Interestingly, the isolates obtained in this study were resistant to low pH conditions and slightly susceptible to bile salts, but they were susceptible to a trypsin treatment, revealing a phenotype pattern that is different from that which has been observed for respiratory FCVs.