Characterization of heterogeneous vancomycin-intermediate resistance, MIC and accessory gene regulator (agr) dysfunction among clinical bloodstream isolates of staphyloccocus aureus

Abstract

BackgroundThe development of hVISA has been associated with vancomycin clinical failures and is commonly misidentified in clinical microbiology laboratories. Therefore, the objectives of this present study was to improve the reliability of methodologies and criteria for identifying hVISA, evaluate the prevalence of hVISA among clinical bloodstream isolates of S. aureus and determine if there exists a relationship between accessory gene regulator (agr) dysfunction and the hVISA phenotype.MethodsThe presence of hVISA in 220 clinical S. aureus isolates (121 MSSA, 99 MRSA) from bloodstream infections was examined by CLSI broth microdilution, Macro & Standard Etest. Isolates which were classified as hVISA by Macro Etest, were additionally evaluated using a modified PAP-AUC method using a modified starting inoculum of 1010 CFU/mL, and growth on brain heart infusion agar with 4 mg/L vancomycin (BHIV4) at 108 and 1010 CFU/mL, and agr function was assessed by delta-hemolysin production.ResultsBroth microdilution MIC50/90 of S.aureus and hVISA was 1.0/2.0 and 1.5/2.0 mg/L (p= 0.02), respectively. Macro Etest identified 12 (5.5%) hVISA isolates; higher among MRSA (9.1%) versus MSSA (2.5%) (p = 0.03). The mean modified PAP-AUC ratios (> 0.8) of 7 MRSA strains and 3 MSSA strains were significantly different (p = 0.001). 58% of hVISA strains were found to be agr dysfunctional when 21% of MRSA strains were agr dysfunctional. hVISA was detected among S. aureus bloodstream isolates, which were classified as susceptible among clinical microbiology laboratories.ConclusionsEvaluating the correlation between Etest MICs and modified PAP-AUC ratio values will add further improvement of discriminating hVISA, and agr dysfunction may be predictive of strains which display a greater predilection to display the hVISA phenotype.

What role does the quorum-sensing accessory gene regulator system play during Staphylococcus aureus bacteremia?

“…Similar to methicillin resistance, several studies report reduced or absent Agr activity in isolates with decreased sensitivity to vancomycin, particularly those encoding the type II agr variant [30,[75][76][77]80]. However, it is unknown whether the loss of agr activity is simply a consequence of other changes conferring vancomycin resistance or lack of Agr activity directly contributes to susceptibility.…”

“…agr-defective S. aureus is a frequent cause of bacteremia Despite the importance of Agr in many infections, strains lacking detectable Agr activity have been isolated in 3-82% of cases of S. aureus bacteremia [10,[26][27][28][29][30][31][32]. The methods used to determine Agr functionality vary from simple haemolysis assays on solid media to sequencing of the agr operon, which may contribute to the variation in their reported frequency ( Table 1).…”

Treatment options for methicillin-resistant Staphylococcus aureus (MRSA) infection: Where are we now?

“…Mutations leading to resistance including agr, other polymorphisms and the expression of these, are another key area for ongoing research [42][43][44][45]. Until these mechanisms are more fully understood, current practice will consist of somewhat unsatisfactory phenotypic methods of resistance detection before molecular methods of resistance detection or certainty of susceptibility become available [28].…”

A Novel δ-Hemolysis Screening Method for Detecting Heteroresistant Vancomycin-Intermediate Staphylococcus aureus and Vancomycin-Intermediate S. aureus

“…Our aim was to develop a simple and rapid screening method for heteroresistant vancomycin-intermediate Staphylococcus aureus (hVISA) and vancomycin-intermediate Staphylococcus aureus (VISA), based on ␦-hemolysin production, to correctly identify and distinguish hVISA and VISA from vancomycin-susceptible S. aureus (VSSA), based on the observations that hVISA and VISA are characterized by a decreased functionality of the regulatory agr (accessory gene regulator) locus encoding ␦-hemolysin (1,2,7,8,(11)(12)(13)(14)(15).…”