β-lactams and colistin resistance in Enterobacterales is a global public health issue. In this study we aimed to investigate the occurrence and genetic determinants of Extended-Spectrum β-lactamases, carbapenemases and mcr-encoding-genes in Enterobacterales isolates recovered from the migratory bird species Ciconia ciconia in an Algerian city. A total of 62 faecal samples from white storks were collected. Samples were then subjected to selective isolation of β-lactams and colistin-resistant-Enterobacterales. The representative colonies were identified using Matrix-Assisted Laser Desorption-Ionisation Time-of-Flight Mass Spectrometry. Susceptibility testing was performed using the disk-diffusion method. ESBL, carbapenemases, and colistin resistance determinants were searched for by PCR and sequencing. The clonality relationships of the obtained isolates were investigated by multilocus sequence typing assays. Mating experiments were carried out to evaluate the transferability of the carbapenemase and mcr-genes. Forty-two isolates were identified as follows: Escherichia coli (n = 33), Klebsiella pneumoniae (n = 4), Proteus mirabilis (n = 4) and Citrobacter freundii (n = 1). Molecular analysis showed that twelve isolates carried the bla_ESBL genes alone, fifteen E. coli isolates were positive for the bla_OXA-48 gene, six isolates were NDM-5-carriers (two P. mirabilis, two K. pneumoniae and two E. coli) and eight E. coli strains were positive for the mcr-1 gene. MLST results showed a high clonal diversity, where NDM-5-producing strains were assigned to two sequence types (ST167 for E. coli and ST198 for K. pneumoniae), whereas the mcr-1 positive E. coli isolates belonged to ST58, ST224, ST453, ST1286, ST2973, ST5542, ST9815 and the international high-risk resistant lineage ST101. To the best of our knowledge, this is the first report of bla_NDM-5 gene in white storks and also the first describing the mcr-1 gene in white storks in Algeria. This study underlines the important role of migratory white storks as carriers of high-level drug-resistant bacteria, allowing their possible implication as indicators and sentinels for antimicrobial resistance surveillance.

Antibiotic-resistant bacteria can leave hospitals and therefore contaminate the environment and, most likely, humans and animals, through different routes, among which wastewater discharge is of great importance. This study aims to assess the possible role of hospital sewage as reservoir and dissemination pathway of carbapenem-resistant Gram-negative bacilli (GNB). Carbapenem-resistant GNB were selectively isolated from wastewater collected from a public hospital in Batna, Algeria. Species identification was carried out using matrix-assisted laser desorption and ionization time-of-flight mass spectrometry, and antibiotic susceptibility was evaluated by the disc diffusion method. β-Lactamase production was investigated phenotypically using the double-disk synergy assay and the modified CarbaNP test, then the molecular mechanisms of β-lactam-resistance were studied by PCR and sequencing. Ten Enterobacteriaceae and 14 glucose-nonfermenting GNB isolates were obtained. All Enterobacteriaceae isolates were positive for OXA-48 and TEM-1D β-lactamases, where seven of them coproduced an extended-spectrum β-lactamase. VIM-2 carbapenemase was detected in six glucose-nonfermenting GNB isolates. However, three Pseudomonas aeruginosa, one Comamonas jiangduensis and one Acinetobacter baumannii isolates were positive for VIM-4 variant. In addition, NDM-1 enzyme was detected in four A. baumannii isolates. Our findings highlight the potential impact of hospital wastewater in the spread of drug resistance mechanisms outside of hospitals.