Clonal and horizontal transmission of carbapenem-resistant Enterobacterales strains and genes via flies.

Background: Antimicrobial resistance (AMR) is one of the most pressing global public health challenges; in particular, the rapid dissemination of carbapenem-resistant Enterobacterales (CRE) is emerging as a significant concern worldwide. Flies, serving as carriers of pathogens, pose a potential threat in the transmission of antibiotic-resistant bacteria (ARB) between animals and humans. The aim of this study was to evaluate and reveal the potential risk of AMR spread by flies.

Methods: A total of 450 flies were collected from four farms, four rural areas, and four urban areas in Dengfeng, Henan, China. To select CRE strains on the surface of flies, three flies sampled from the same geographical location were arbitrarily selected and placed into one tube of brain heart infusion broth (BHI), and the supernatant was screened using CHROMagar™ mSuperCARBA culture medium. Different colors and shapes of colonies were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and 16S rRNA sequencing. Antimicrobial susceptibility testing for CRE strains was performed using broth microdilution. All CRE strains were whole-genome sequenced. Short-read sequencing was performed using MGISEQ-2000 and long-read sequencing was conducted using GridION.

Results: Totally, 150 BHI tubes were screened for CRE strains, and 33 strains were identified as CRE positive. In 24 mSuperCARBA plates, only one species of CRE strain was isolated from each plate. In three plates, two different species of CRE strains were identified in each plate. In one plate, three different species of CRE strains were simultaneously isolated. Carbapenem resistance genes were detected in 81.8% of CRE strains, and bla_NDM-1 was predominant (66.7%). No significant correlations between carbapenem-resistant phenotypes and carbapenem resistance genes were observed. The complete genomes of all 33 strains were obtained. Genome analysis revealed that clonal transmission events may have occurred among different farms and rural areas. Phylogenetic analysis revealed that bla_NDM-1 IncFII plasmids could break bacterial species barrier for cross-host transmission in diverse areas.

Conclusions: To understand and control the transmission of AMR from the perspective of One Health, it is imperative to enhance surveillance of ARB, antibiotic resistance genes, and antibiotic-resistant plasmids in flies.