Association of mprF mutations with cross-resistance to daptomycin and vancomycin in methicillin-resistant Staphylococcus aureus (MRSA)

Yusuke Sato’O,Yuancheng Zhang,Yusuke Taki,Yoshifumi Aiba,Aa Haeruman Azam,Xin-Ee Tan,Feng-Yu Li,Kanate Thitiananpakorn,Tanit Boonsiri,Longzhu Cui,Kotaro Kiga,Teppei Sasahara,Shinya Watanabe

doi:10.1038/s41598-020-73108-x

Abstract

We first reported a phenomenon of cross-resistance to vancomycin (VCM) and daptomycin (DAP) in methicillin-resistant Staphylococcus aureus (MRSA) in 2006, but mechanisms underlying the cross-resistance remain incompletely understood. Here, we present a follow-up study aimed to investigate genetic determinants associated with the cross-resistance. Using 12 sets of paired DAP susceptible (DAPS) and DAP non-susceptible (DAPR) MRSA isolates from 12 patients who had DAP therapy, we (i) assessed susceptibility to DAP and VCM, (ii) compared whole-genome sequences, (iii) identified mutations associated with cross-resistance to DAP and VCM, and (iv) investigated the impact of altered gene expression and metabolic pathway relevant to the cross-resistance. We found that all 12 DAPR strains exhibiting cross-resistance to DAP and VCM carried mutations in mprF, while one DAPR strain with reduced susceptibility to only DAP carried a lacF mutation. On the other hand, among the 32 vancomycin-intermediate S. aureus (VISA) strains isolated from patients treated with VCM, five out of the 18 strains showing cross-resistance to DAP and VCM carried a mprF mutation, while 14 strains resistant to only VCM had no mprF mutation. Moreover, substitution of mprF in a DAPS strain with mutated mprF resulted in cross-resistance and vice versa. The elevated lysyl-phosphatidylglycerol (L-PG) production, increased positive bacterial surface charges and activated cell wall (CW) synthetic pathways were commonly found in both clinical isolates and laboratory-developed mutants that carry mprF mutations. We conclude that mprF mutation is responsible for the cross-resistance of MRSA to DAP and VCM, and treatment with DAP is more likely to select for mprF-mediated cross-resistance than is with VCM.

Highlights

Methicillin-resistant Staphylococcus aureus (MRSA) infections are serious clinical problems causing high morbidity and mortality worldwide
Our results indicated that reduced susceptibility to both DAP and VCM was regulated by mprF mutation via increased L-PG production, subsequent alteration of bacterial surface charge, and cell wall (CW) biosynthetic pathways
This study began with analysis and validation of VCM and DAP susceptibilities for all paired isolates collected from 12 patients from whom D APR methicillin-resistant Staphylococcus aureus (MRSA) strains were generated during DAP therapy (Supplemental Table 1)

Summary

Introduction

Methicillin-resistant Staphylococcus aureus (MRSA) infections are serious clinical problems causing high morbidity and mortality worldwide. The interaction between DAP and the CM causes potassium leakage and membrane depolarization that contribute to cell d eath[12] This means that DAP and VCM differ in chemical structure. The more positively charged CM surface may serve as a protective barrier against DAP b inding[20,21] This remains controversial since only some DAP non-susceptible ( DAPR) strains displayed enhanced L-PG concentration in outer leaflet most DAPR strains carrying mprF mutation showed increased L-PG production[19,22,23,24,25]. A total of 12 sets of DAP susceptible (DAPS) and DAPR MRSA isolates collected from different hospitals in Japan were whole-genome sequenced, and gene mutations associated with the phenotype of cross-reduced susceptibility were identified and functionally characterized. Our results indicated that reduced susceptibility to both DAP and VCM was regulated by mprF mutation via increased L-PG production, subsequent alteration of bacterial surface charge, and CW biosynthetic pathways

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