2. Universidad Cardenal Herrera-CEU

Antimicrobial resistance poses a major threat to global health and food security and is primarily driven by antimicrobial use in human and veterinary medicine. Understanding its epidemiology at farm level is crucial for effective control measures. Despite the significant reduction in antibiotic use in conventional livestock production, the swine sector traditionally has a higher level of antibiotic use in veterinary medicine. Consequently, multidrug resistance (MDR) among microbial isolates of swine origin has been relatively frequent. The aim of this study was to assess the presence of multidrug-resistant (MDR) bacteria, enteric pathogens and resistance genes to the main antibiotics used in clinical practice, both within the environment and in animals across pig farms characterized by varying degrees of sanitary status. A total of 274 samples were collected. Of these, 34 samples were collected from the environment (wall swabs, slat swabs and slurry pit), and 240 samples were collected from animals (sows’ and piglets’ rectal faeces). All samples were analysed for MDR bacteria and enteric pathogens. The study revealed a high frequency of extended-spectrum beta-lactamases (ESBL)-producing Enterobacterales and Campylobacter spp., with ESBL-producing Enterobacterales predominating in high health status farms (environment and animals) and Campylobacter spp. in both high health status and low health status environments. Additionally, a high percentage of methicillin-resistant Staphylococcus aureus (MRSA) was found, mainly in environmental samples from high health status farms, and Clostridioides difficile was distributed ubiquitously among farms and samples. Furthermore, though less frequently, vancomycin-resistant Enterococcus faecium (VRE) was isolated only in high health status farms, and Gram-negative bacilli resistant to carbapenems were isolated only in environmental samples of high health status and low health status farms. This study underscores the importance of surveillance for MDR bacteria in farm animals and their environment, including their waste. Such ecosystems serve as crucial reservoirs of bacteria, requiring national-level surveillance to promote responsible antibiotic use and pandemic control.

Control strategies to minimize pathogenic bacteria in food animal production are one of the key components in ensuring safer food for consumers. The most significant challenges confronting the food industry, particularly in the major poultry and swine sectors, are antibiotic resistance and resistance to cleaning and disinfection in zoonotic bacteria. In this context, bacteriophages have emerged as a promising tool for zoonotic bacteria control in the food industry, from animals and farm facilities to the final product. Phages are viruses that infect bacteria, with several advantages as a biocontrol agent such as high specificity, self-replication, self-limitation, continuous adaptation, low inherent toxicity and easy isolation. Their development as a biocontrol agent is of particular interest, as it would allow the application of a promising and even necessary “green” technology to combat pathogenic bacteria in the environment. However, bacteriophage applications have limitations, including selecting appropriate phages, legal restrictions, purification, dosage determination and bacterial resistance. Overcoming these limitations is crucial to enhance phage therapy’s effectiveness against zoonotic bacteria in poultry. Thus, this review aims to provide a comprehensive view of the phage-biosanitation strategies for minimizing persistent Salmonella and Campylobacter bacteria in poultry.

Failure in antibiotic therapies due to the increase in antimicrobial-resistant (AMR) bacteria is one of the main threats to public and animal health. In recent decades, the perception of companion animals has changed, from being considered as a work tool to a household member, creating a family bond and sharing spaces in their daily routine. Hence, the aim of this study is to assess the current epidemiological situation regarding the presence of AMR and multidrug resistance (MDR) in companion animals in the Valencia Region, using the indicator bacteria Escherichia coli as a sentinel. For this purpose, 244 samples of dogs and cats were collected from veterinary centres to assess antimicrobial susceptibility against a panel of 22 antibiotics with public health relevance. A total of 197 E. coli strains were isolated from asymptomatic dogs and cats. The results showed AMR against all the 22 antibiotics studied, including those critically important to human medicine. Moreover, almost 50% of the strains presented MDR. The present study revealed the importance of monitoring AMR and MDR trends in companion animals, as they could pose a risk due to the spread of AMR and its resistance genes to humans, other animals and the environment they cohabit.

Mycobacteriosis is an important disease that affects captive and wild aquatic fish. Syngnathids are susceptible to infection by non-tuberculous mycobacteria. The aim of this study was to describe clinical signs, and macroscopic and histological lesions in 25 syngnathids and the molecular characterization of the causative mycobacteria. Clinical presentation ranged from sudden death to non-specific signs, including anorexia, poor body condition, weight loss and marked dyspnea with increased respiratory effort and rate. Gross lesions were mostly ulcers on the tail and small white nodules in the liver, coelomic cavity and inside the eye. The most affected organs were gills, liver, intestine and coelomic mesentery. Microscopic lesions consisted of areas of multifocal to diffuse granulomatous inflammation and bacterial emboli with numerous intralesional acid-fast bacilli. Epithelioid cells, multinucleated giant cells, lymphocytes and fibrous connective tissue, which are commonly observed in granulomatous inflammation, were not observed here. In the real-time PCR, M. fortuitum, M. chelonae and M. marinum common primers, Mycobacterium spp. were detected in 4, 7 and 14 individuals, respectively. In addition, this is the first description of mycobacteriosis found in Syngnathus acus.

Antimicrobial resistance (AMR) and the increase in multi-resistant bacteria are among the most important threats to public health worldwide, according to the World Health Organisation (WHO). Moreover, this issue is underpinned by the One Health perspective, due to the ability of AMR to be transmitted between animals and humans living in the same environment. Therefore, since 2014 different surveillance and control programmes have been established to control AMR in commensal and zoonotic bacteria in production animals. However, public health authorities’ reports on AMR leave out companion animals, due to the lack of national programmes and data collection by countries. This missing information constitutes a serious public health concern due to the close contact between companion animals, humans and their surrounding environment. This absence of control and harmonisation between programmes in European countries leads to the ineffectiveness of antibiotics against common diseases. Thus, there is a pressing need to establish adequate surveillance and monitoring programmes for AMR in companion animals and further develop alternatives to antibiotic use in this sector, considering the impact this could have on the gut microbiota. In this context, the aim of this review is to evaluate the current control and epidemiological situations of AMR in companion animals in the European Union (EU), as well as the proposed alternatives to antibiotics.

Salmonella is mostly noted as a food-borne pathogen, but contact with chelonians has also been reported as a source of infection. Moreover, high levels of antimicrobial resistance (AMR) have been reported in Salmonella isolated from wild and captive reptiles. The aim of this study was to assess the occurrence of Salmonella AMR carriage by chelonians admitted to two zoological institutions in Spain, characterizing the isolates to assess the Salmonella AMR epidemiology in wildlife. To this end, 152 chelonians from nine species were sampled upon their arrival at the zoological nuclei. Salmonella identification was based on ISO 6579-1:2017 (Annex D), isolates were serotyped and their AMR analysed according to the EU Decision 2013/652. Moreover, the genetic relationship of the isolates was assessed by pulsed-field gel electrophoresis (PFGE). Results showed 19% (29/152) of the chelonians positive to Salmonella, all of them tortoises. For all isolates, 69% (20/29) were resistant and 34% (10/29) multidrug-resistant (MDR) strains. PFGE clustered isolates according to the serovar, confirming a low genetic diversity. In conclusion, this study shows a high presence of MDR Salmonella strains in tortoises at their entry into zoological nuclei. This condition highlights the need to establish Salmonella detection protocols for the entry of animals into these centres.

Pork is considered a major source of Salmonella Typhimurium infection in humans in the EU, including monophasic strains. Widespread distribution of virulent serotypes such as monophasic variants of S. Typhimurium have emerged as a public health threat. Despite the current situation, within the EU there is no mandatory programme for the control of Salmonella at pork production level. In this context, the aims of this study were: to examine the presence of Salmonella in the swine production system from arrival at the slaughterhouse until the end of processing, and investigate the genetic relationship among the Salmonella serovars isolated. During the study, a total of 21 pig herds were intensively sampled during processing at the slaughterhouse. ERIC-PCR was performed among isolates recovered at the different steps in the slaughterhouse to assess the genetic relationship. Then, PFGE was done to study the pulsotypes among the different Salmonella serovars isolated. The results showed a high level of Salmonella pork batch contamination upon arrival at the slaughterhouse (71.4%) and at the end of the slaughtering process (66.7%), with mST the main serovar isolated from both origins (53.1% and 38.2%, respectively). The slaughter environment poses a potential risk for carcass contamination and it is considered an important source of Salmonella spp. Similarly, this study shows that 14.3% of the strains isolated from carcasses have the same Xbal-PFGE profile as those previously recovered in the slaughterhouse environment, but not in the live animals from that same batch. In conclusion, there is a high level of Salmonella swine batch contamination upon arrival at the slaughterhouse and at the end of the slaughtering process, mST being the most frequently isolated serovar. Moreover, a strong genetic relationship has been observed between Salmonella strains isolated from the batch on arrival at the slaughterhouse, the processing environment and pork carcass contamination. In this sense, it would be necessary to implement a control programme to reduce the bacterium from pork farms and raise the awareness of biosecurity measures.

Background: Salmonella enterica serovar Infantis is a zoonotic pathogen isolated in broilers causing great economic losses in the European poultry sector. It is demonstrated that an investment in management measures at farm level could directly affect the control of food chain microorganisms. The aim of this study was to investigate the development of S. Infantis antimicrobial resistance (AMR) patterns during the growing period, according to flock density and ventilationmanagement, without antibiotic administration. Methods: The experiment was performed in two identical poultry houses, evaluating commercial and optimal farm conditions. At 24 h of rearing, 20% of the animals were orally infected with a S. Infantis strain susceptible to all the antibiotics tested. To study Salmonella shedding, faeces samples from each experimental groupwere takenweekly and analysed as per ISO/TS 6579- 2:2017. Antibiotic susceptibilitywas assessed according toDecision 2013/653. Results: Salmonella shedding showed that the lowest counts were observed in the first week post-infection and highest at slaughter day for both groups. Moreover, 100% of the isolates were multi-resistant. Conclusion: The acquisition of AMR by S. Infantis starts at the onset of the production cycle and is maintained until the end, demonstrating the importance of transmission of AMR in zoonotic bacteria at farmlevel.

Poultry is one of the main agricultural sub-sectors worldwide. However, public concern regarding animal welfare and antimicrobial resistance has risen in recent years. Due to the influence of management practices on microbiota, it might be considered to evaluate poultry welfare and health. Therefore, the objective of this research was to analyse the influence on microbiota balance of broilers under commercial and optimal farm conditions, using 16S rRNA sequencing analysis. The research was performed in two identical poultry houses (commercial vs. optimal). Results showed a higher level of microbiota complexity in the group reared under optimal farm conditions at the end of rearing. Regarding microbiota composition, Firmicutes was the dominant phylum during the entire growing period. However, the second most prevalent phylum was Proteobacteria at the arrival day, and Bacteroidetes from the mid-period onward in both groups. Moreover, the most predominant genera identified were Oscillospira, Ruminococcus, Bacteroides, and Coprococcus. In conclusion, it is necessary to optimize farm management as much as possible. Using gut microbiota diversity and composition as biomarkers of animal health could be an important tool for infectious disease control, with the aim of reducing the administration of antibiotics at field level.

New measures applied to reduce antimicrobial resistances (AMR) at field level in broiler production are focused on improving animals’ welfare and resilience. However, it is necessary to have better knowledge of AMR epidemiology. Thus, the aim of this study was to evaluate AMR and multidrug resistance (MDR) dynamics during the rearing of broilers under commercial (33 kg/m2 density and max. 20 ppm ammonia) and improved (17 kg/m2 density and max. 10 ppm ammonia) farm conditions. Day-old chicks were housed in two poultry houses (commercial vs. improved), and no antimicrobial agents were administered at any point. Animals were sampled at arrival day, mid-period and at slaughter day. High AMR rates were observed throughout rearing. No statistical differences were observed between groups. Moreover, both groups presented high MDR at slaughter day. These results could be explained by vertical or horizontal resistance acquisition. In conclusion, AMR and MDR are present throughout rearing. Moreover, although a lower level of MDR was observed at mid-period in animals reared under less intensive conditions, no differences were found at the end. In order to reduce the presence of AMR bacteria in poultry, further studies are needed to better understand AMR acquisition and prevalence in differing broiler growing conditions.