The co-occurrence of tet(X4) and tmexCD2-toprJ2 mediated tigecycline resistance in Raoultella ornithinolytica

Abstract

Objectives

This study aimed to characterize the co-occurrence of the tigecycline resistance determinants tet(X4) and tmexCD2-toprJ2 in a Raoultella ornithinolytica isolate collected from a pig rectal swab at the slaughterhouse.

Methods

The R. ornithinolytica isolate WS60 was subjected to antimicrobial susceptibility testing. Whole-genome sequencing (WGS) was performed to analyze the genetic features of the plasmids carrying tet(X4) and tmexCD2-toprJ2. Additionally, a conjugation assay was conducted to evaluate the transferability of these plasmids, followed by a 15-day stability test to assess the persistence of the two resistance determinants.

Results

R. ornithinolytica WS60 exhibited high-level tigecycline resistance, with a minimum inhibitory concentration (MIC) of 32 μg/mL, and was also resistant to ampicillin, ampicillin-sulbactam, chloramphenicol, tetracycline, sulfamethoxazole-trimethoprim, florfenicol, and streptomycin. WGS analysis revealed that WS60 harbored three plasmids, including a 384,249-bp tmexCD2-toprJ2-carrying IncQ plasmid (pWS60–1) and a 78,159-bp tet(X4)-carrying IncFII plasmid (pWS60–2). Interestingly, pWS60–2 was identical to several plasmids found in Klebsiella spp. isolated from animals, animal-derived food, and humans. Moreover, pWS60–2 was successfully transferred to Klebsiella spp. via conjugation, whereas pWS60–1 failed to transfer. Notably, no significant fitness cost was observed in the transconjugants carrying pWS60–2. Additionally, a 15-day stability assay demonstrated that both resistance determinants were stably maintained in the bacterial population without significant loss, underscoring their persistence over time.

Conclusions

This is the first report of the co-occurrence of tet(X4) and tmexCD2-toprJ2 in R. ornithinolytica. Enhanced surveillance in slaughterhouses, along with targeted interventions, should be implemented to mitigate the potential spread of mobile tigecycline resistance throughout the food production chain.