Background/Objectives: Antimicrobial resistance (AMR) in Enterococcus species from poultry production represents a significant public health threat due to the potential transmission of AMR through the food chain. This study aimed to examine the relative prevalence, resistance patterns, and mannitol fermentation capacity of Enterococcus isolates from various poultry production systems in Spain over a seven-year period (2017–2023). Methods: A total of 215 Enterococcus isolates were analyzed. Phenotypic assessments were conducted to determine resistance rates and metabolic capacities, while genotypic characterization focused on detecting vancomycin-resistance genes (vanA, vanB, vanC, and vanD). Results: Enterococcus faecalis (62.3%) and Enterococcus faecium (29.77%) were the predominant species, primarily isolated from broilers (74.88%), with the highest frequency observed in one-week-old chicks (31.16%). High resistance rates to tetracyclines and streptogramins were identified, while resistance to vancomycin (0.47%) and tigecycline (3.03%) was low. Interestingly, a significant reduction in tetracyclines resistance was shown in this period for Enterococcus faecalis (from 100% to 70% (2017–2023) and Enterococcus faecium (from 100% to 40% (2018–2023)). Multidrug resistance (MDR) was detected in 26.98% of isolates. Mannitol fermentation tests revealed high metabolic capacity in Enterococcus faecalis (99.25%) and Enterococcus faecium (96.88%), associated with adaptability and virulence potential. Genotypic analysis showed a very low prevalence of vanB and vanC genes. Conclusions: These findings highlight the critical need for targeted surveillance and intervention strategies in poultry production to mitigate the risks posed by MDR Enterococcus to public health.

Salmonella represents a food safety concern worldwide. Despite the application of National Control Programs (NCP) against Salmonella, regulated by the European Union, every year the European Food Safety Authority reports new cases. On the look for new alternatives to antibiotics, bacteriophages, or phages, rise as a promising alternative to treat multidrug resistance infections. Although they are known to be ubiquitous in the environment, their high specificity to host cells hinders their isolation and usage for phage therapy. The ISO 6579-1:2017 is performed as a reference method in the NCP and uses an unspecific media to enrich the sample the same way most phage isolation protocols do. Later, the protocol uses a more selective media to isolate the Salmonella, Modified Semisolid Rappaport Vassiliadis (MSRV). This paper aims to find out whether, due to the similarity between phage isolation protocols and the ISO 6579-1:2017, this last one could be used as a protocol to also isolate phages against the same bacterium that is being simultaneously isolated. To do so, 2 experiments were performed to assess phage isolation from MSRV media in in-vivo conditions. The results from experiments 1 and 2 proved that the MSRV media was usable for simultaneous phage and pathogen isolation through a single procedure. Additionally, there is a correlation between the antigenic formulae from the bacteria and the phage's host range, seeming to be effective against bacteria with similar antigenic formulae.

Salmonella enterica serovar Infantis (S. Infantis) poses a growing issue in the poultry sector, with phage-based products emerging as a safe and effective control measure. This study investigated the emergence of reduced-phage-susceptibility variants (RPSV) of S. Infantis in PhagoVet-treated broilers, given that RPSV could undermine phage treatment efficacy. The bacteriophages in the PhagoVet product were characterized using transmission electron microscopy (TEM), genome sequencing, and infection profiling. Furthermore, two broiler trials were conducted: a challenge group (T1) and a challenge-and-treated group (T2). The S. Infantis infective dose was set at 10⁴ and 10⁶ colony-forming units (CFUs) per animal, with PhagoVet administration at 10⁶ and 10⁸ plaque-forming units (PFUs) per animal, in Trials 1 and 2, respectively. The results revealed that the four PhagoVet bacteriophages belonged to different genera. PhagoVet evidenced broad-spectrum efficacy against 271 strains representing 18 Salmonella serovars. In Trial 1, PhagoVet reduced bacterial counts in feces to nearly undetectable levels by day 42, with no RPSV detected. However, in Trial 2, three and five RPSVs were detected in feces and ceca, respectively. Consequently, PhagoVet demonstrated efficacy against S. Infantis in broilers, and the potential impact of RPSV is deemed unlikely to compromise its efficacy.

Background/Objectives: As bacteriophage-based strategies to control bacterial pathogens continue to gain momentum, phage therapy is increasingly being explored across various fields. In the poultry industry, efforts to minimize the public health impact of Salmonella have spurred growing interest in phage applications, particularly as prophylactic and disinfecting agents. Although the disinfecting potential of bacteriophages has been recognized, in-depth studies examining their efficacy under varying environmental conditions remain limited. This study focused on evaluating the effectiveness of bacteriophages as disinfecting agents against biofilm-forming Salmonella Infantis under different environments. Methods: A comprehensive screening of biofilm-producing strains was conducted using Congo Red Agar and 96-well plate assays. Two strains with distinct biofilm-forming capacities were selected for further analysis under different environmental conditions: aerobic and microaerobic atmospheres at both 25 °C and 37 °C. The resulting biofilms were then treated with four phage preparations: three individual phages and one phage cocktail. Biofilm reduction was assessed by measuring optical density and CFU/well. Additionally, scanning electron microscopy was used to visualize both untreated and phage-treated biofilms. Results: The results demonstrated that all S. Infantis strains were capable of forming biofilms (21/21). All three phage candidates exhibited biofilm-disrupting activity and were able to lyse biofilm-embedded Salmonella cells. Notably, the lytic efficacy of the phages varied depending on environmental conditions, highlighting the importance of thorough phage characterization prior to application. Conclusions: These findings underscore that the effectiveness of bacteriophages as surface disinfectants can be significantly compromised if inappropriate phages are used, especially in the presence of biofilms.