Florfenicol-resistant, Rabbit-derived Providencia Rettgeri Isolate With 4 Copies of floR Genes in a Tandem-repeat Plasmid

Background: This work aimed to investigate the resistance mechanism to antimicrobial agents (especially florfenicol) exhibited by Providencia isolates obtained from various sources.Methods: The polymerase chain reaction (PCR) was performed to detect the distribution of the florfenicol resistance genes. Conjugation, S1-PFGE (S1 nuclease pulsed-field gel electrophoresis), sequencing and other molecular methods were employed to analyze the transferability and structure of the resistance gene-encoding plasmids. Results: It showed that the human pathogens exhibited a wider resistance spectrum, while the animal bacteria had higher MIC values for most antibiotics analyzed. Of the seven florfenicol resistance genes (floR, fexA, fexB, optrA, estDL136, cfr and pexA) screened, only floR was identified in both animal (5/10, 50%) and human bacteria (3/13, 23.1%), and none of them was detected in freshwater fish isolates (0/22). The floR-positive strains exhibited high MIC levels for florfenicol (≥ 64 mg/L). The whole genome sequencing of the multidrug-resistant Providencia rettgeri strain R39 demonstrated that it carried four plasmids, with two larger plasmids encoding 5 copies of the floR genes. The largest plasmid, pR39-33 (32,936 bp) with 4 floR genes, was composed of 4 copies of the second large plasmid pR39-8 which encoded 8 ORFs (open reading frames) including a floR and two mobilization genes (mobAC) in the form of tandem repeat. It indicated that the floR gene represented the primary mechanism underlying the florfenicol resistance of the Providencia isolates. Conclusions: This report is the first to describe an antibiotic-resistant plasmid composed of four copies of another plasmid sequence in the form of a tandem repeat.