Geertrui Rasschaert

The presence of biofilms on the surfaces of drinking water systems in livestock housing can compromise the microbiological quality of drinking water. Understanding the biofilm microbiome and the biofilm-forming capacity of its dominant species is therefore essential, particularly for identifying species that coexist and enhance biofilm formation through interactions. This study investigated 248 bacterial isolates representing the dominant microbiota of biofilm samples collected from drinking water systems in broiler houses and pig nursery units. Biofilm formation was assessed in both single-species cultures and 341 multispecies combinations. Of the tested isolates, 80% formed biofilms in single culture; however, only 25% were classified as strong biofilm formers. In multispecies combinations with up to four species or isolates, antagonistic or competitive interactions were dominant, whereas biofilm mass enhancement was observed in only a limited number of combinations. These included opportunistic pathogens, with Citrobacter freundii and Staphylococcus nepalensis identified as key species driving the interactions. Our research highlights the importance of specific microbial interactions in biofilm development and provides a potentially practice relevant four-species model to guide future research aimed at improving strategies for controlling biofilms in livestock drinking water systems. IMPORTANCE: Ensuring the quality of drinking water on livestock production farms benefits animal production and health. Persistent biofilms on water line surfaces can lead to microbial contamination, resulting in lower-quality drinking water. This study examines bacteria commonly found in biofilms within livestock drinking water systems in pig nursery units and broiler houses. Most bacterial isolates can form biofilms in monoculture. Specific interactions among certain species primarily mediate bacterial interactions that enhance biofilm biomass in multispecies communities. Identifying these key interactions enables the development of a synthetic model to guide research on controlling biofilms in livestock drinking water systems.

Antibiotic resistance in Gram-negative bacteria poses a global health threat, and poultry farming provides an important reservoir for multidrug-resistant pathogens. Our study aimed to characterize the faecal microbiota and acquired resistome of Ross-308 roosters in Hungary. Amplicon and shotgun metagenomics revealed a faecal microbiota dominated by the Firmicutes, Bacteroidota, and Proteobacteria and a diverse faecal resistome, including qnrB and an aadA1-bearing integron. Culture-based screening of an antibiotic-free rooster yielded the MDR Escherichia coli strain K1G, displaying resistance also to third-generation cephalosporins and fluoroquinolones. Whole-genome sequencing classified K1G as a serotype O23:H16-ST453 avian pathogenic E. coli (APEC) strain featuring a set of chromosomal virulence factors (including astA, hlyE, lpfA, and iss) and three plasmids: a phage-like plasmid, a mosaic virulence plasmid (carrying bla_TEM-1b, hlyF, iutA, ompT, iucD, and cvaC), and an IncC type 1 resistance plasmid harbouring bla_CMY-2. The detection of identical or closely related ST453 E. coli strains also in broiler meat in Hungary highlights a potential risk of transmission to humans through the food chain. Moreover, the carriage of multiple acquired antibiotic resistance genes in E. coli K1G indicates that individual chickens can harbour or transmit antibiotic resistance even in the absence of direct antibiotic exposure.

Antimicrobial resistance is a major global health concern, with extended-spectrum β-lactamase (ESBL)-producing bacteria posing a significant threat. Food production animals have been indicated as an important source of ESBL-producing bacteria. This study examined the occurrence of ESBL Escherichia coli in Belgian livestock (cattle, calves, pigs feces, and broiler ceca) and farm environments. Antimicrobial susceptibility of E. coli isolates ( n = 1,219) was screened using microbroth dilution (EUVSEC), and bla _{TEM/SHV/CTX-M} genes were further identified by PCR and sequencing. ESBL-producing E. coli isolates were more common in broiler samples (10% ceca, 2.5% environment) compared to those of pigs (1% feces, 2% environment), cattle (1% feces), and calves (1% feces). Among the 67 ESBL isolates, 63% carried bla _CTX-M genes, and 55% bla _TEM genes. bla _SHV genes (13%) appeared only in isolates from broiler origin (ceca and farm environment). All ESBL-producing E. coli isolates were resistant to between 3 and 13 out of the 19 antimicrobials tested. In addition, a rapid PCR-based screening method in manure DNA extracts was evaluated for its ability to detect bla _{TEM/SHV/CTX-M} genes compared to the conventional culture-based approach. These β-lactamase genes were detected in 98% of the samples ( bla _TEM : 95/101, bla _CTX-M : 6/101, bla _SHV : 0/101). The low prevalence of bla _CTX-M and bla _SHV in manure DNA extracts compared to the corresponding E. coli isolates may be due to PCR sensitivity issues or matrix inhibitory effects. The presence of ESBL-producing E. coli in broiler ceca, livestock feces, and manure raises concerns about the potential transmission of resistance genes in agriculture, highlighting the need for ongoing surveillance and responsible antibiotic use. IMPORTANCE: This research addresses the pressing global health concern of antimicrobial resistance, focusing on extended-spectrum β-lactamase (ESBL)-producing bacteria. The study investigates the occurrence and distribution of ESBL-producing Escherichia coli in Belgian livestock, including cattle, calves, pigs, and broiler chickens, as well as in their farm environments. The findings provide valuable data to inform farmers, veterinarians, and policymakers to raise awareness and support evidence-based antimicrobial stewardship.

Background: Drinking water systems (DWS) are often an overlooked source of microbial contamination of drinking water in broiler and piglet production. Persistent biofilms within water lines can act as reservoirs of contamination, reintroducing microorganisms into the flowing water and potentially compromising animal health. This study investigates the microbial composition of biofilms in the DWS of broiler houses and pig nursery units, their impact on drinking water quality, and the influence of the source water on both water quality and biofilm communities. Results: The bacterial load of DWS biofilm swabs, collected at the end of production cycles before cleaning and disinfection was evaluated, and the dominant bacterial taxa were identified. Furthermore, 16S gene metabarcoding was applied to the biofilm samples. No significant differences in microbial load were observed between the two sectors, with a median total aerobic count of 3.6 log CFU/cm ² . Enterococci, a faecal indicator, were detected in 80% of all samples. Moreover, Escherichia coli was found more frequently in broiler houses (47%) than in pig nursery units (27%). The two dominant identified genera were Staphylococcus and Pseudomonas . The Staphylococcus saprophyticus species was the most frequently identified isolate, accounting for 10.6% of all isolates across both broiler houses and pig nursery units. In broiler houses, the next most frequently identified species were Pseudomonas aeruginosa (5.6%) and Stenotrophomonas maltophilia (5.3%). In contrast, in pig nursery units, Pseudomonas fluorescens (6.3%) and Psychrobacter faecalis/pulmonis (5.5%) were most frequently identified. Research showed that the drinking water microbial community not only depended on the source water but was also influenced by biofilms in DWS, as similar bacterial taxa were found in both the drinking water at the drinking nipples and in biofilms on water-contact surfaces. Conclusions: The presence of faecal indicator bacteria and potential animal pathogens underscores the risks associated with the biofilms. These biofilms can contaminate drinking water to animals, underscoring the need for targeted strategies to monitor and mitigate biofilm formation.

For several years, Infantis was the most common Salmonella serovar circulating in the Belgian broiler sector and persisting on broiler farms. To gain insight into its prevalence and circulation on broiler farms in Belgium, five farms (14 flocks) with a S. Infantis contamination history were monitored during two consecutive production rounds. In total, ten sampling events were conducted using moist sponge sticks after cleaning and disinfection, during the delivery of the one-day-old chicks and during production until slaughter or until positive for S. Infantis. Salmonella presence on samples was determined based on the ISO 6579:2017 standard, and the isolated strains were typed using PFGE. The results showed that current cleaning and disinfection practices were unable to completely remove S. Infantis from the farms. Cleaning equipment (3 out of 9 sample times) and the floor (5 out of 10 sample times) were particularly contaminated. Furthermore, external environmental samples were also frequently contaminated (e.g., mortality containers, concrete driveway). During production, 12 of the 28 sampled flocks were colonized with S. Infantis after one week, indicating that S. Infantis quickly spreads throughout the broiler house, which raises the hypothesis that feeding and/or drinking water systems play a critical role in the circulation of the bacteria. This study gives insights into the circulation and difficulty of controlling S. Infantis in persistently contaminated broiler farms, highlighting the importance of thorough cleaning and disinfection and biosecurity.

1.During transport of broilers from farms to slaughterhouses, the interior of crates may become contaminated with faeces. When these crates are not cleaned and disinfected adequately, they pose a potential risk for the dissemination of pathogens, including Campylobacter spp., between slaughterhouses and farms.2. The aim of the trial was to compare the efficacy of immersion-based crate washing systems with a spray-based washing system in reducing Campylobacter spp. Therefore, the crate washing systems in two slaughterhouses were sampled. In one of these two slaughterhouses, the immersion-based crate washing system was replaced by a new spray-based system which was sampled. Samples were collected from the slaughterhouse equipment, the crate washing system as well as from the crates both before and after cleaning and disinfection (C&D). All samples were screened for the presence of Campylobacter spp.; water and crate samples were used for enumeration. Molecular typing was performed to investigate the source and routes of contamination.3. In all cases, crates were contaminated with Campylobacter spp. prior to washing. However, only the spray-based system achieved a significant reduction after C&D Molecular typing revealed that the same strains were present on the crates after C&D as before cleaning. Additionally, crates could become contaminated by previously cleaned crates via wash water in the immersion-based crate washing system.