Background: Colistin is a last-resort antibiotic for the treatment of multidrug-resistant Gram-negative bacterial infections. The antibiotic has been used in veterinary medicine, especially in pigs to treat post-weaning diarrhoea caused by Escherichia coli. This study determined the burden of colistin-resistant Enterobacteriaceae (ColR-E) in Belgian broiler and pig farms.
Materials/methods: As part of the i-4-1 Health project, a total of 400 broiler and 400 pig fecal samples were collected from broiler (n=15) and sow farms (n=15) in Belgium between September 2017 and April 2018. A random-stratified design was used to collect twenty to thirty samples per farm (FecalSwab, Copan Italy). The presence of non-intrinsically ColR-E was investigated using selective culturing and colistin MIC testing using broth microdilution (epidemiological cut-off: 2 mg/L). To quantify colistin use, treatment incidence was calculated in accordance with Timmerman et al. (2006). A selection of ColR-E (E. coli n=14, Klebsiella pneumoniae n=15, Klebsiella oxytoca n=2, Enterobacter cloacae n=2) was sequenced (Illumina MiSeq) to determine genotype-phenotype correlations. Core genome multilocus sequence typing (cgMLST) was used to determine clonal relatedness.
Results: The percentage of samples positive for ColR-E was higher in pigs (22%) compared to broilers (0.75%)(Figure 1A). A positive correlation was found between within-farm colistin use and presence of resistance (Kendall’s tau: 0.52, p<0.01). Of all resistant strains, 67% were E. coli exhibiting MIC between 4 and 16 mg/L. Resistance was also detected in K. pneumoniae (28%), K. oxytoca (2%) and E. cloacae complex (3%) exhibiting MIC of 4 to >64 mg/L. Plasmid-mediated resistance genes were detected in E. coli (Figure 1B). Known resistance mechanisms were absent in 9/33 of the ColR-E, suggesting that other mechanisms are involved. Based on PROVEAN scores, we show mutations that possibly influence colistin susceptibility. Disruption of mgrB by stop codon or insertion sequence was predominant in K. pneumoniae (Figure 1C). K. pneumoniae ST101, an emerging clone associated with increased mortality rates in humans, was detected with high-level colistin resistance in pigs.
Conclusions: This study revealed that ColR-E are found in Belgian farms. The presence of high-level colistin-resistant strains, like K. pneumoniae ST101, in pig farms is a potential public health threat.