Abstract
The mechanisms of linezolid resistance among 13 E. faecalis and 6 E. faecium isolates, recovered from six Spanish hospitals during 2017–2018, were investigated. The presence of acquired linezolid resistance genes and mutations in 23S rDNA and in genes encoding for ribosomal proteins was analyzed by PCR and amplicon sequencing. Moreover, the susceptibility to 18 antimicrobial agents was investigated, and the respective molecular background was elucidated by PCR-amplicon sequencing and whole genome sequencing. The transferability of the linezolid resistance genes was evaluated by filter-mating experiments. The optrA gene was detected in all 13 E. faecalis isolates; and one optrA-positive isolate also carried the recently described cfr(D) gene. Moreover, one E. faecalis isolate displayed the nucleotide mutation G2576T in the 23S rDNA. This mutation was also present in all six E. faecium isolates. All linezolid-resistant enterococci showed a multiresistance phenotype and harbored several antimicrobial resistance genes, as well as many virulence determinants. The fexA gene was located upstream of the optrA gene in 12 of the E. faecalis isolates. Moreover, an erm(A)-like gene was located downstream of optrA in two isolates recovered from the same hospital. The optrA gene was transferable in all but one E. faecalis isolates, in all cases along with the fexA gene. The cfr(D) gene was not transferable. The presence of optrA and mutations in the 23S rDNA are the main mechanisms of linezolid resistance among E. faecalis and E. faecium, respectively. We report the first description of the cfr(D) gene in E. faecalis. The presence of the optrA and cfr(D) genes in Spanish hospitals is a public health concern.
Highlights
On the one hand, enterococci, especially the species Enterococcus faecalis (Efs) and Enterococcus faecium (Efm) can be harmless colonizers of the human intestinal tract, but on the other hand, they are one of the most important bacterial genera related to hospital-associated infections worldwide
The optrA gene was identified in all Efs isolates (n = 13), and the X528 isolate harbored the cfr(D) gene
The nucleotide point mutation G2576T within domain V of the 23S rDNA gene was detected in one optrA-positive isolate
Summary
Enterococci, especially the species Enterococcus faecalis (Efs) and Enterococcus faecium (Efm) can be harmless colonizers of the human intestinal tract, but on the other hand, they are one of the most important bacterial genera related to hospital-associated infections worldwide. Enterococci possess intrinsic resistance to different antimicrobial agents commonly used to treat infections caused by Gram-positive pathogens, such as cephalosporins and aminoglycosides (low-level resistance). They have a great capacity to acquire mobile genetic elements carrying antimicrobial resistance genes, which limits the therapeutic options [1]. Linezolid was the first oxazolidinone approved for clinical use in human medicine and is considered as a last resort antimicrobial agent It is an important treatment option for serious infections (such as nosocomial and community-acquired pneumonia, and complicated skin and soft tissue infections) caused by multiresistant Gram-positive bacteria, including vancomycin-resistant enterococci [2,3,4]. Different amino acid exchanges, deletions, and insertions in the ribosomal proteins L3 (rplC), L4 (rplD), and L22 (rplV)— of lesser significance—have been associated with decreased susceptibility to linezolid [1,3]
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