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Antimicrobial resistance (AMR) is a global public health threat that affects human, animal and ecosystem health. Antibiotic resistance hot spots are found not only in medical settings but also in environmental compartments that are subjected to anthropogenic pressure and contribute to the reservoir of ARGs collectively constituting the antibiotic resistome. Basing on a holistic and multidisciplinary "One Health" approach to face the AMR, scientific research highlighted the need to reveal the missed link between the increasing of AMR in livestock and humans and the emergence of AMR in wildlife. This PhD thesis was undertaken with the aim to give a broader consideration to this research topic since the discussion of AMR is often focused on human and livestock health outcomes. The studies reported have been performed to assess the antimicrobial resistance profiles of bacteria from wild species (wild birds, reptiles), as well as in other non-traditional species that in recent years have become popular as pets. The detection of multidrug-resistant strains of clinically relevant pathogens in wild and/or captive species of reptiles, birds and small mammals from different settings suggests that they may act as potential carriers of zoonotic agents and resistance determinants, highlighting the potential risks for public health related to the close contact and the sharing of the same environment.

Antimicrobial resistance (AMR) is a current public health issue globally. To counter this phenomenon and prioritize AMR in the health sector, the World Health Organization (WHO) published a list of bacterial pathogens against which the development of new antimicrobial agents is urgently needed, designating the ESKAPE pathogens (i.e., Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) with a ‘priority status’. Moreover, the One Health High Level Expert Panel (OHHLEP) states that human health is closely linked to animal and environmental health, thus promoting a holistic One Health approach in order to be prepared to address possible emerging health threats from the human–animal–environment interface. Wild birds may host and spread pathogens, integrating the epidemiology of infectious diseases. The aim of this study was to examine the role of wild birds as a source of ESKAPE bacteria and other antibiotic-resistant enterobacterales. A total of fifty strains within the ESKAPE group were detected in 40/163 cloacal samples of examined birds (24.5%). Additionally, different strains of enterobacterales were detected in 88/163 cloacal samples (53.9%). Isolated strains exhibited antimicrobial resistance, including towards critically important antibiotics (e.g., third, fourth, fifth generation cephalosporins, fluoroquinolones) for human medicine. Our results confirm that wild birds are potential reservoirs of several pathogens and antimicrobial-resistant bacteria and that they could be involved in the dissemination of those bacteria across different environments, with resulting public health concerns.

Characterizing the gut microbiota of free-range and alternative poultry production systems provides information, which can be used to improve poultry welfare, performance, and environmental sustainability. Gut microbiota influence not only the health and metabolism of the host but also the presence of zoonotic agents contaminating food of animal origin. In this study, the composition and diversity of the cecal microbiota community of free-range grown chickens were characterized by 16S rDNA high-throughput Illumina sequencing. Significant differences were observed in the composition of chicken cecal microbiota at the time points of 28 days of age (Indoor group) and 56 days of age (Outdoor group), i.e., before and after the outdoor access period of chicken groups. The Outdoor group showed a richer and more complex microbial community, characterized by the onset of new phyla such as Deferribacterota and Synergistota, while the Indoor group showed an increase in Campylobacterota. At the species level, it is noteworthy that the occurrence of Mucispirillum schaedleri in Outdoor group is known to potentially stimulate mucus layer formation in the distal intestinal tract, thus being associated with a healthy gut. We also report a significant decrease in the Outdoor group of Helicobacter pullorum, highlighting that the lower abundance at the age of slaughter reduced the possibility to contaminate chickens' carcasses and, consequently, its zoonotic potential. As revealed by a mutual exclusion study in network analysis, H. pullorum was present only if Bacteroides barnesiae, an uncultured organism of the genus Synergistes, and Bacteroides gallinaceum were absent. Finally, microbiome predictive analysis revealed an increase of vitamins and micronutrient biosyntheses such as queuosine (Q) and its precursor pre Q0, in the Outdoor group, suggesting that the outdoor evolved microbiota of chickens do contribute to the vitamin pool of the gut and the biosynthesis of micronutrients involved in vital cell processes.

Antibiotic misuse is greatly contributing to an increase in antimicrobial resistance (AMR) in humans and animals. Natural and synthetic alternative strategies are being investigated in human and veterinary medicine, but little attention is paid to the antimicrobial effects of edible lipids, such as medium-chain fatty acids (MCFAs) and monoglycerides. Among MCFAs, lauric acid (LA) and its monoglyceride derivative, glycerol monolaurate (GML), exhibit the strongest antimicrobial activity. Coconut and palm kernel oils are considered the main sources of LA. On the other hand, some edible insects (e.g., Hermetia illucens) are gaining interest as novel feed ingredients, due to the high amount of LA they contain as well as their numerous bioactive components, which provide many additional benefits to animal health. Although the beneficial effect of both MCFAs and LA is gradually being recognized, their high content within insects and, consequently, their possible role as antimicrobials, has not been well-reported. This mini review focuses on the anti-infective effects of the insect-derived MCFAs LA and its derivatives. We emphasize the potential of insect lipids, compared to the other vegetable sources, in the current global scenario where a sustainable and circular economy is required. Finally, we critically discuss the use and the benefits of edible insects such as favorable options as feed and food from the perspective of animal and human nutrition.

Wild birds may host and spread pathogens, integrating the epidemiology of infectious diseases. Particularly, Larus spp. have been described as responsible for the spread of many enteric diseases, primarily because of their large populations at landfill sites. The aim of this study was to examine the role of yellow-legged gulls as a source of enteropathogenic bacteria such as Campylobacter spp., Salmonella spp., Shiga toxin-producing Escherichia coli and Yersinia spp., with particular attention to antibiotic-resistant strains. Enteropathogenic bacteria were isolated from 93/225 yellow-legged gulls examined from April to July, during a four-year period (2016–2019). Specifically, Campylobacter spp. was isolated from 60/225 samples (26.7%), and identified as C. coli (36/60) and as C. jejuni (24/60). Salmonella spp. was isolated from 3/225 samples (1.3%), and identified as Salmonella arizonae. Shiga toxin-producing E. coli were isolated from 30/225 samples (13.3%) samples, and serotyped as E. coli O128 (12/30) O26 (9/30), O157 (6/30) and O11 (3/30), Yersinia spp. was never detected. Isolated strains exhibited multidrug resistance, including vitally important antibiotics for human medicine (i.e., fluoroquinolones, tetracyclines). Our study emphasizes the importance of yellow-legged gulls as potential reservoirs of pathogenic and resistant strains and their involvement in the dissemination of these bacteria across different environments, with resulting public health concerns.

Antimicrobial resistance is a public health concern worldwide and it is largely attributed to the horizontal exchange of transferable genetic elements such as plasmids carrying integrons. Several studies have been conducted on livestock showing a correlation between the systemic use of antibiotics and the onset of resistant bacterial strains. In contrast, although companion birds are historically considered as an important reservoir for human health threats, little information on the antimicrobial resistance in these species is available in the literature. Therefore, this study was aimed at evaluating the antimicrobial resistance of Escherichia coli and Pseudomonasaeruginosa isolated from 755 companion birds. Cloacal samples were processed for E. coli and P. aeruginosa isolation and then all isolates were submitted to antimicrobial susceptibility testing. P. aeruginosa was isolated in 59/755 (7.8%) samples, whereas E. coli was isolated in 231/755 (30.7%) samples. Most strains showed multidrug resistance. This study highlights that companion birds may act as substantial reservoirs carrying antimicrobial resistance genes which could transfer directly or indirectly to humans and animals, and from a One Health perspective this risk should not be underestimated.

Buffalo mastitis is an important economic problem in southern Italy, causing qualitative/quantitative alterations in milk and resulting in economic losses due to the sub-clinical course and chronic evolution. We investigated 50 udders of slaughtered buffaloes and subjected them to effectual microbiological screening to evaluate macro and microscopic mammary gland changes, immune-characterize the cell infiltrates, and compare the degree of tissue inflammation with somatic cell counts. Numerous Gram-positive and Gram-negative bacteria were isolated from all samples, majority of which were environmental mastitis pathogens. Histological features referable to chronic mastitis were observed in 92% udders. Lymphocytes, plasma cells and macrophages were found to evolve into aggregates in 48% udders, which often organized to form tertiary lymphoid structures (TLSs). A predominance of interstitial CD8+ over CD4+ lymphocytes and, in TLSs, scattered CD8+ lymphocytes in the mantle cells and CD79+ lymphocytes in germinal centers, were evidenced. Environmental pathogens are known to persist and cause chronic inflammatory changes in buffaloes, where CD8+ lymphocytes play an important role by controlling the local immune response. Moreover, the TLSs evidenced here for the first time in buffalo mastitis, could play a role in maintaining immune responses against persistent antigens, thereby contributing in determining the chronic course of mastitis.

The sugar glider (Petaurus breviceps) is a small, arboreal, nocturnal, gliding mammalian possum belonging to the marsupial infraclass. Exotic marsupials, including sugar gliders, are becoming popular companion pets and, consequently, the risk of potential infections that can be transmitted to humans should be investigated. Data on the role of the sugar glider as a possible carrier of pathogenic and zoonotic bacteria are scarce and fragmentary. Therefore, this study is aimed at evaluating the prevalence of potentially zoonotic bacteria (Salmonella spp., Escherichia coli, Campylobacter spp., Pseudomonas spp., Klebsiella spp., Listeria monocytogenes and Yersinia enterocolitica) in 64 sugar gliders kept as pets in Italy. The highest prevalence of infection pertained to members of the family Enterobacteriaceae, in particular Citrobacter spp. (50%), Enterobacter spp. (28·1%) and Klebsiella pneumoniae (15·6%); Pseudomonas aeruginosa was isolated from 10 out of 64 samples (15·6%). All strains of Klebsiella pneumoniae exhibited some level of resistance to multiple antimicrobials (ampicillin, amoxicillin-clavulanic acid and doxycycline). SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study show that sugar gliders may act as carriers of potentially pathogenic agents for humans and other animal species, therefore caution should be exercised in the handling and contact with these animals.

Background New epidemiological data on bacterial and parasitic infections in 24 Italian wall lizards, namely Podarcis sicula (mainland population) and P. sicula klemmerii (insular population) in southern Italy were provided. To achieve this goal, samples were collected from individuals belonging to the two populations and analysed by microbiological and parasitological methods. Results A wide range of bacteria (e.g. Pantoea spp., Citrobacter spp., Morganella spp., Pseudomonas, Enterobacter spp., Staphylococcus spp. and Escherichia coli) and parasites (e.g. Ophionyssus natricis, coccidia, Dicrocoelidae) were detected in both P. sicula and P. sicula klemmerii individuals. Insular population presented similar bacterial and parasitic diversity to its mainland counterpart. Ampicillin was the antimicrobial with the highest resistance rate. Conclusion This study highlighted various bacteria and parasites, some of them potentially zoonotic. Further studies are needed to better understand the epidemiology and transmission routes of these pathogens along with their impact on the welfare and behaviour of Italian wall lizards.

Avian species are considered as the main reservoir of Campylobacter spp. However, few data are available on the presence of this microorganism in pet birds. This study was therefore performed to determine the prevalence of Campylobacter spp. in pet birds bred in southern Italy. Faecal samples were collected from 88 cages housing different species of pet birds and examined by bacteriological culture and polymerase chain reaction. A total of 13.6% of the cage samples were positive for Campylobacter coli. Other Campylobacter spp. were not found. The study shows that C. coli can be isolated from the cages of apparently healthy pet birds, which should therefore be considered as potential carriers of C. coli and a possible source of infection for humans and companion animals.

The incidence of campylobacteriosis have increased in both developed and developing countries over the last 10 years [1]. Several avian species are considered the main reservoirs of Campylobacter spp. [2,3]. Nevertheless, current scientific knowledge on the presence of Campylobacter spp. in pet birds is scarce. To address this lack of information, the present study was undertaken with the aim to evaluate the presence of these microorganisms in pet birds bred in southern Italy. To achieve this goal, 14 bird farms located in the Campania region (southern Italy) were visited. In each farm, bird farm population ranged between 20 to 100 animals belonging to the families of Estrildidae, Fringillidae and Psittacidae, from which 33, 28, and 27 pooled faecal samples were collected respectively. Specifically, eighty-eight cages housing a total of 225 captive birds were examined. The cage was used as an epidemiological unit, and each cage housed from 1 to 5 birds. Each sample was stored in Amies Charcoal Transport Medium at 4°C, transported to the laboratory, and analyzed within 2 h of collection by cultural and molecular methods. A total of 12/88 (13.6%) cages were positive for Campylobacter spp. which was identified as C. coli. In particular, 7/33 (21.2%) cages came from Estrildidae and 5/27 (18.5%) cages came from Psittacidae family. The twentyeight cages coming from Fringillidae family were consistently negative. our results demonstrate that C. coli may be found in the intestines of apparently healthy pet birds which could be considered as a further potential carrier of C. coli for humans and other companion animals. The adoption of good hygiene practices when handling pet birds should be, therefore, promoted.