In Belgium, the FASFC monitors the evolution of antimicrobial resistance (AMR) in food and food-producing animals (primary production). Resistance in the zoonotic bacteria Salmonella spp. and Campylobacter spp. and in methicillin-resistant Staphylococcus aureus (MRSA) as well as resistance in indicator bacteria Escherichia coli, Enterococcus faecalis and Enterococcus faecium was monitored in 2024. Moreover, a specific monitoring of presumptive extended spectrum β-lactamases/AmpC/carbapenemase producing E. coli is done on strains isolated from food-producing animals and meat derived thereof. Microbiological resistance was assessed using epidemiological cut-off values (ECOFF) according to EUCAST (European Committee on Antimicrobial Susceptibility Testing).

In 2024, the European Commission Implementing Decision 2020/1729 of the 17th of November 2020 was applicable for the determination of the epidemiological cut-off values as well as for the selection of antimicrobial panels (a.o. EUVSEC3/EUCAMP3).

In Campylobacter jejuni isolated from poultry meat, as reported in previous years, the predominant resistance profiles included both, ciprofloxacin combined with tetracycline. Resistance to tetracycline remains stable as in 2023, accounting for 43.3% of the isolates.  Resistance to ertapenem in C. jejuni decreased in 2024 compared to 2023 reaching values <10%.Notably, the proportion of fully susceptible isolates increased in 2024 reaching 31.6%, the highest level of complete susceptibility observed over the last three years.

In Campylobacter coli isolated from poultry meat, similar high patterns of resistance to ciprofloxacin and tetracycline were observed while resistance to erythromycin and gentamicin remain low to very low. Notably, resistance to ertapenem, decreased from extremely high (74.3%) in 2023 to very high (58.3%) in 2024. The proportion of multidrug resistant isolates decreased to 41.7% in 2024 compared to 48.6% in 2023. This decrease is likely explained by the lower level of resistance observed to ertapenem in 2024.

In primary production, the monitoring of Campylobacter coli in fattening pigs showed overall stable resistance levels in comparison with previous years, with extremely high resistance to tetracycline (74.4%) and high to ciprofloxacin (45.1%). Resistance to erythromycin, ertapenem and gentamicin remained at low level (<10%). Resistance patterns in C. coli isolated from caecal content of veal calves,   included extremely high level of resistance to ciprofloxacin and tetracycline combined with very high levels of erythromycin and high level of ertapenem resistance. Only one isolate was susceptible to all antimicrobials tested. In C. jejuni, from veal calves  patterns in ciprofloxacin resistance showed a progressive decrease since 2021, reaching the lowest level in 2024 (50.7%). Resistance to tetracycline remains extremely high. Combined resistance to critically important antimicrobials including ciprofloxacin and erythromycin accounted for 6%. Notably, only 8.6% of the isolates were fully susceptible to all antimicrobials tested  Overall, as seen in previous reports, antimicrobial resistance levels are lower in C. jejuni than in C. coli.

In 2024, Salmonella spp. recovered from food matrices have been analysed by whole genome sequencing (WGS). The most interesting features are subsequently described. Two out of the 236 isolates had a predicted resistance to 3rd generation cephalosporins, one isolated from broiler meat preparation belonging to the serotype Infantis and a second from imported poultry meat from Brazil belonged to the serovar Minnesota. The Infantis  isolate harboured a bla_CTX-M-3 gene together with the following genes: aadA1; aac(6’)-Iaa, dfrA14, sul1 and tet(A), which confer a multidrug resistant profile including aminoglycosides, trimethoprim, sulfamethoxazole and tetracycline. In addition point mutations in the chromosomal genes gyrA (S83Y) and parC (T57S) were also detected. These mutations are associated with resistance to (fluoro)quinolones. The Minnesota isolate carried the bla_CMY-2 gene which confers an AmpC phenotype. It also carried the following genes: aac(6’)-Iaa, sul2, tet(A) and qnrB19. These genes confer resistance to sulfamethoxazole, tetracycline and ciprofloxacin. This isolate is therefore also considered multi-drug resistant. Moreover, it also harboured a mutation of the gene parC (T57S) which also confers resistance to fluoroquinolones.

In Salmonella spp. isolated from food-producing animals, the most prevalent serotypes found in fresh fecal samples from broilers was Paratyphi B var. Java (50.46%) followed by Infantis (15.14%). Notably, this prevalence represents a shift compared to the distribution observed in 2022. Resistance levels to all antimicrobials were lower in 2024 except for amikacin where resistance was detected for the first time in two isolates, S. Chester and S. Infantis. This decline in resistance is likely linked to a shift in dominant Salmonella serovars; S. Infantis — known for higher resistance in 2022 — to the dominant serovar S. Paratyphi B var. Java in 2024, which generally exhibits lower resistance. Resistance to 3^rd generation cephalosporins is rare, one S. Paratyphi B var. Java displayed an ESBL phenotype. Other uncommon resistances to last resorts antimicrobials such as colistin was detected in one S. Typhimurium and one S. Enteritidis isolates.

In 2024, all Salmonella spp. isolates from fattening pigs and veal calves were analysed by WGS. The most predominant serotype on isolates from fattening pigs sampled at slaughterhouse is Monophasic Typhimurium followed by Derby and Typhimurium. Considering all serotypes, a total of 23 different antimicrobial resistance genes or mutations were identified among the isolates. The most frequently detected gene was aac(6’)-Iaa, present in all of the isolates. Other commonly found genes included bla_TEM-1B (47.5%), sul2 (32.5%), aph(3”)-Ib and aph(6)-Id (30% each) conferring a predictive resistance to ampicillin, sulfamethoxazole and aminoglycosides such as streptomycin and spectinomycin. Tetracycline resistance genes such as tet(B) (20%), tet(A) (10%), and tet(M) (7.5%) were also present. Genes conferring resistance to aminoglycosides aadA1, aadA2, aadA2b, sulfamides (sul1, sul3), phenicols (floR, cmlA1), and fluoroquinolones (gyrA p.D87Y, qnrB19) were detected at lower frequencies (2.5–10%).  The mutation in the gene parC p.T57S which  is associated to resistance to fluoroquinolones was found in 35% of the isolates. None of the isolates were predicted to be resistant to highest priority critically important antimicrobials particularly to 3rd generation cephalosporins or to colistin. One S. Typhimurium was recovered from caecal content from veal calves sampled at slaughterhouse. The following resistance genes, aac(6’)-Iaa, aadA2, aadA1, aph(3’)-Ia, cmlA1, sul3, tet(A), tet(M), dfrA12 were detected. Therefore the expected phenotype should include resistance to aminoglycosides (streptomycin and spectinomycin), chloramphenicol sulfamethoxazole, tetracycline and trimethoprim.

Isolates from the official monitoring of Salmonella on broilers, laying hens and turkeys were analysed by WGS as well.

S. Paratyphi B var. Java was the most predominant serotype in broilers, while Enteritidis was the most predominant in laying hens. In turkeys only a single isolate was recovered which belonged to the monophasic variant of S. Typhimurium.

All  S. Enteritidis from laying hens belonged to the ST11 and did not harbour any resistance gene (except the aac(6’)-Iaa) nor exhibit chromosomal point mutations, however S. Paratyphi B var. Java, isolated from broilers, all belonged to ST28 and carried resistance mechanisms to at least 3 classes of antibiotics, including aminoglycosides and trimethoprim/sulfamethoxazole in all of them.

The specific monitoring of ESBL, AmpC and carbapenemase producing E.coli was performed in broilers, turkeys, fattening pigs and veal calves at slaughterhouse and in meat derived from these 4 categories of food producing animals at retail. In 2024, the highest prevalence of ESBL E.coli was found in both caecal content from veal calves (59.3%) and from broilers (59.3%) followed by fattening pigs (20.2%). A progressive decrease has been observed over time, although the differences are not consistently statistically significant. However, since 2015 (60%)  a strong decrease in E. coli ESBL isolated from fattening pigs has been observed reaching significantly lower levels by 2024 (20%). Similar patterns has been observed in fresh meat derived of the food producing animals. Among the fresh meat categories, a gradual decrease of the prevalence of E.coli ESBL/AmpC has been noticed over the years, mainly in broiler meat, from 80% reported in 2016 to 34.73 % in 2024.

No meropenem-resistant isolates were detected in 2024. However, ESBL E.coli isolated from these matrices showed extremely high levels of multidrug resistance (>80%).

The most frequent determinants encoding for ESBL enzymes found in isolates from broilers at slaughterhouse were bla_SHV-12, (40.9%) and bla_TEM-52 (21.6%), conferring an ESBL phenotype, while only 5 isolates had an AmpC predicted phenotype due to the presence of bla_CMY-2 and bla_DHA-1 or a mutation in the ampC promoter. Combination with other critical important antimicrobials such as fluoroquinolones was found in 56% of the isolates. This was found to be mediated by the presence of plasmidic qnrS genes in 67.6% of the isolates or by point mutation in one or more of the of the gyrA, parC and parE genes (32.4%).

Isolates from fattening pigs carried genes encoding for enzyme production CTX-M-1 group (including bla_CTX-M-1, bla_CTX-M-14 , bla_CTX-M-15, bla_CTX-M-32, bla_CTX-M-55). Resistance to fluoroquinolones was detected in 28.1% of the ESBL isolates conferred either by the qnrS gene or by point mutations in the gyrA, parC and parE genes.

Notably, one isolate harboured in addition to ESBL genes the mcr-1.1 colistin resistance gene.

Regarding the ESBL isolates retrieved form caecal content of veal calves, the most common resistance genes detected belonged to the bla_CTX-M group in 88.3% of isolates either alone, in combination with another bla_CTX-M gene, or with the ESBL genes bla_OXA-17 and bla_OXA-74. Predicted resistance in combination with fluoroquinolones was found in 42.8% of the ESBL isolates. The resistance genes were either alone or combined with another qnrS gene or with point mutations of the gyrA, parC or parE genes.

In indicator E.coli, in comparison with 2023 the resistance to most antibiotics remained stable in 2024 in food-producing animals: in broilers, veal calves of less than one year and fattening pigs sampled at the slaughterhouse as well as in laying hens, breeding hens and meat bovines of less than 7 months old sampled at the farm. The highest prevalence to the critically important antimicrobials 3^rd generation cephalosporins was detected in E. coli from veal calves caecal content sampled at the slaughterhouse and was higher than in previous years but remained below 5%. As expected, resistance to (fluoro)quinolones was the highest (48.2%) in E. coli from broilers caecal content. Low or very low rates of resistance to amikacin were detected in E. coli isolated from pig caecal content every year since its addition in 2021. However, for the first time over the previous years, no resistance to colistin was detected in E. coli isolated from veal calves caecal content. At the level of the farm, the highest resistance to (fluoro)quinolones was found in E. coli isolated from breeding hens while resistance to 3^rd generation cephalosporins was detected and confirmed in 3 E. coli isolates from fecal samples of bovines (beef cattle), and in one isolate from breeding hens. Resistance to amikacin was detected in one isolate from breeding hens and in one isolate from laying hens. In indicator E. coli isolated from meat sampled at border control posts, three of the four isolates from beef meat were fully susceptible. Of the four isolates from broiler meat, one was resistant to 3^rd generation cephalosporins and three were resistant to (fluoro)quinolones. No resistance to meropenem was detected in any of the indicator E. coli isolates.

Monitoring of methicillin-resistant Staphylococcus aureus (MRSA) was carried out in veal calves, dairy cows and bovines for meat production on farms in 2024. The aim of this monitoring is to assess the MRSA prevalence in these animal categories and determine the genotypes (STs and spa-types) of  the collected MRSA isolates together with their AMR and virulence genes. In 2024, the prevalence was high in veal calves (44.1%, Ci95% [36.3-52.3%]), low in dairy cows (9.5%, Ci95% [5.5-16.0%]), and low in bovines for meat production (9.4%, Ci95% [4.8-17.7%]). The prevalence of MRSA on farms has decreased in veal calves and remained stable in dairy cows and bovines for meat production since 2012. Among the 84 MRSA collected, all isolates except one, belonged to the CC398, typical of livestock-associated MRSA (LA-MRSA). One isolate from bovine for meat production belonged to the sequence type CC1/ST9381 and t177 spa-type, typical of community-associated and hospital-associated MRSA (CA/HA-MRSA) lineage. Identification of such MRSA CC1/t177 was already reported among pigs, in other countries, confirming that some MRSA types within CC1 have the potential for establishment and spread in livestock (Silva et al., 2023; Elstrom et al., 2019).

All MRSA (n=84/84) were genetically multi-drug resistant (i.e., carrying genes or mutations conferring resistance to at least 3 different antibiotic classes including beta-lactams). Regarding the resistance genes to last-resort antibiotics observed in MRSA in 2024 in Belgium, one ST398/t011 isolate from bovine for meat production carried the cfrAB variant of the cfr gene. This variant is functionally truncated, but cfrAB isolates were found elsewhere to spontaneously mutate to cfr (Murphy 2022, thesis of the Curtin University).

The aur, hlgB, hlgC and selw virulence genes were detected in all MRSA isolates, and hlgA in all but one isolate from dairy cows. The aur gene encodes the aureolysin, a typical exoenzyme from S. aureus (Dubin, 2022); the selw gene was already reported to be commonly found in CC398 MRSA (Vrieling et al., 2020); and the hlgA, hlgB and hlgC are common in Belgian LA-MRSA. The staphylococcal enterotoxin B, encoded by the seb gene, was found in two t011/ST398 isolates from veal calves. This enterotoxin is frequently detected in different retail foods, especially in dairy-related foods (Zhang et al., 2024) and is known to be the source for multiple pathologies in humans (Verreault et al., 2019). The t011/ST6035 isolate from veal calves, belonging to the LA-MRSA CC398 clonal complex carried the sak and scn genes associated with the human immune evasion cluster and several genes associated with toxins (hlgA, hlgB, hlgC, sep and selw) and exoenzyme (aur). The CC1/ST9381/t177 isolate harboured multiple virulence genes, including sak and scn (immune evasion cluster), toxin-associated genes (hlgA, hlgB, hlgC, lukD, lukE, seh, selw and selx), and exoenzyme genes (aur, splA, and splB). Genes from the human immune evasion cluster were not detected in the other isolates.

The presence of MRSA in food-producing animals, along with their carriage of multiple antimicrobial resistance and virulence genes, constitutes a potential public health risk.

The monitoring of Enterococcus faecalis and Enterococcus faecium, organized in Belgium in food-producing animals between 2011 and 2013, and resumed in 2019, continued. Investigation of the AMR prevalence in these commensal indicator bacteria was assessed in order to complete the picture of the situation of antimicrobial resistance within our farms and slaughterhouses. Enterococci are also considered to be reservoirs of antibiotic resistance genes, present in both humans and animals. In 2024, the prevalence of enterococci species by animal category were similar to those observed in previous years. Enterococcus faecium was more frequently isolated than Enterococcus faecalis within the samples of breeding hens (83.8%), laying hens (79.9%), veal calves (69.2%) and pigs (46.3%). In broilers samples, E. faecalis and  E. faecium were equally isolated (68.9% and 76.4%, respectively).

The antimicrobial susceptibility tests carried out this year showed that, in general, the resistance rates observed in Enterococcus faecalis and Enterococcus faecium within the various animal matrices studied have remained stable since 2019, with a few notable exceptions showing significant variations.

Indeed, significant decreases were observed between 2019 and 2024 in erythromycin resistance rate in E. faecalis isolated from breeders and in E. faecium isolated from broilers. Significant decreases were observed in tetracycline resistance rate in E. faecalis isolated from breeders and in E. faecium isolated from breeders and pigs, whereas an increasing trend was observed in E. faecium isolated from veal calves. In addition, a significant decrease in ciprofloxacin resistance rate was observed in E. faecalis isolated from breeders and in E. faecium isolated from layers. A significant decrease was reported in the ampicillin resistance rate in E. faecium isolated from broilers. However, significant increases were also observed in gentamicin and in linezolid resistance rates in E. faecalis isolated from veal calves. No significant variations were observed for the other antimicrobials between 2019 and 2024. Resistance to tetracycline, erythromycin and to quinupristin/dalfopristin were still the most observed resistances, both in E. faecalis and E. faecium. Resistance to linezolid (n=13), a critical antibiotic for human health, was also observed in 2024, in 8 E. faecalis isolated from veal calves (n=7), pigs (n=1) and in 5 E. faecium isolated from pigs (n=2) and veal calves (n=3). Daptomycin resistance was low and observed only in E. faecalis isolated from breeders (n=1, 2.6%). Multidrug resistance was mainly observed in broilers and veal calves with 52.7% of multi-resistant E. faecium and 52.8 % of multi-resistant E. faecalis, respectively. Very low levels of teicoplanin (n=1, 0.6%) and tigecycline (n=1, 0.6%) resistance were observed in E. faecium isolated from veal calves in 2024. No resistance to vancomycin was observed in 2024.

In 2024, 16 enterococci were sequenced by WGS. This report also presents these WGS results and the detailed characterization of these strains, including the detection of cfr (n=1), optrA (n=13) and poxtA (n=3) genes encoding linezolid resistance. The two virulence factors, agg and cyl, associated with high pathogenicity and described in human strains were found in two E. faecalis characterized by an ST179 and isolated from veal calves. Strain typing revealed the identification of 14 different STs (among 16 sequenced isolates), suggesting the presence of some genetic diversity within enterococci exhibiting resistance to critically important antibiotics (i.e. daptomycin, linezolid, tigecycline or teicoplanin).

Taking together all results from both Gram-negative (E. coli) and Gram-positive indicators (enterococci) isolated from food-producing animals, resistance rates between 2019 and 2024 remained overall stable, with some decreasing trend observed in E. coli across several animal categories (broilers, pigs, and slaughtered cattle). Most of the significant changes were decreases, particularly for amikacin, chloramphenicol, sulfamethoxazole, tetracycline, and trimethoprim. In enterococci (E. faecalis and E. faecium), resistance remained globally stable, with some significant decreases seen for erythromycin, tetracycline, ciprofloxacin, and ampicillin in specific animal matrices. Only a limited number of significant increases were observed : gentamicin and nalidixic acid in E. coli respectively isolated from slaughtered cattle and laying hens, and three antimicrobials in enterococci isolated from veal calves (tetracycline in E. faecium, and gentamicin and linezolid in E. faecalis).