Dataset Information

Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Uses the VraSR Regulatory System to Modulate Autophagy for Increased Intracellular Survival in Macrophage-Like Cell Line RAW264.7.

ABSTRACT: The VraSR two-component system is a vancomycin resistance-associated sensor/regulator that is upregulated in vancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) strains. VISA/hVISA show reduced susceptibility to vancomycin and an increased ability to evade host immune responses, resulting in enhanced clinical persistence. However, the underlying mechanism remains unclear. Recent studies have reported that S. aureus strains have developed some strategies to survive within the host cell by using autophagy processes. In this study, we confirmed that clinical isolates with high vraR expression showed increased survival in murine macrophage-like RAW264.7 cells. We constructed isogenic vraSR deletion strain Mu3?vraSR and vraSR-complemented strain Mu3?vraSR-C to ascertain whether S. aureus uses the VraSR system to modulate autophagy for increasing intracellular survival in RAW264.7. Overall, the survival of Mu3?vraSR in RAW264.7 cells was reduced at all infection time points compared with that of the Mu3 wild-type strain. Mu3?vraSR-infected RAW264.7 cells also showed decreased transcription of autophagy-related genes Becn1 and Atg5, decreased LC3-II turnover and increased p62 degradation, and fewer visible punctate LC3 structures. In addition, we found that inhibition of autophagic flux significantly increased the survival of Mu3?vraSR in RAW264.7 cells. Together, these results demonstrate that S. aureus uses the VraSR system to modulate host-cell autophagy processes for increasing its own survival within macrophages. Our study provides novel insights into the impact of VraSR on bacterial infection and will help to further elucidate the relationship between bacteria and the host immune response. Moreover, understanding the autophagic pathway in vraSR associated immunity has potentially important implications for preventing or treating VISA/hVISA infection.

SUBMITTER: Dai Y

PROVIDER: S-EPMC6554704 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Uses the VraSR Regulatory System to Modulate Autophagy for Increased Intracellular Survival in Macrophage-Like Cell Line RAW264.7.

Dai Yuanyuan Y Gao Caihong C Chen Li L Chang Wenjiao W Yu Wenwei W Ma Xiaoling X Li Jiabin J

Frontiers in microbiology 20190531

The VraSR two-component system is a vancomycin resistance-associated sensor/regulator that is upregulated in vancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) strains. VISA/hVISA show reduced susceptibility to vancomycin and an increased ability to evade host immune responses, resulting in enhanced clinical persistence. However, the underlying mechanism remains unclear. Recent studies have reported that S. aureus strains have developed some strateg ...[more]

PMID: 31214151

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Project description:Vancomycin (VAN)-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) isolates are considered to have emerged from VAN-susceptible S. aureus (VSSA) by spontaneous mutation during VAN exposure. We previously reported that laboratory mutant H14, obtained from VSSA strain Delta IP by exposure to imipenem (IPM), showed overexpression of the vraSR two-component system and a typical hVISA phenotype. In the present study, to elucidate the mechanism of VSSA conversion to hVISA, we further characterized strain H14 by determining its whole-genome sequence, morphology, cell wall synthetic activity, and gene expression. Genome sequencing revealed that H14 harbored a mutated vraS (designated vraS(H14)) that caused an amino acid substitution (S(329)-->L). This mutation is different from the VraS mutation (N(5)-->I) identified in representative clinical hVISA strain Mu3. However, H14 exhibited a phenotype similar to that of Mu3, including heterogeneous resistance to VAN, enhanced cell wall synthetic activity, and vraSR overexpression. Replacement of the vraS gene of DeltaIP with the mutated vraS(H14) gene confirmed that the S(329)-->L substitution was responsible for both the upregulation of vraSR and conversion to the hVISA phenotype. This conversion was also achieved by using the vraS gene of Mu3, which carries a mutation (N(5)-->I), but not with the native vraS gene of strain N315. Finally, we carried out a study to analyze the appearance of hVISA from VSSA by exposure of Delta IP to selective concentrations of VAN and beta-lactam antibiotics. A total of 8 and 5 hVISA isolates were detected among 50 isolates selected with VAN and IPM, respectively. Among the 13 hVISA mutants, mutation in vraSR was detected only in mutant strain H14, suggesting that additional mutational mechanisms can be responsible for evolution to the hVISA phenotype. We conclude that exposure not only to VAN but also to beta-lactams may select for reduced glycopeptide susceptibility in S. aureus.

Project description:BackgroundVancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) are associated with vancomycin treatment failure, and are becoming an increasing public health problem. Therefore, we undertook this study of 91 published studies and made subgroup comparisons of hVISA/VISA incidence in different study years, locations, and types of clinical samples. We also analyzed the genetic backgrounds of these strains.MethodsA systematic literature review of relevant articles published in PubMed and EMBASE from January 1997 to August 2014 was conducted. We selected and assessed journal articles reporting the prevalence rates of hVISA/VISA.ResultsThe pooled prevalence of hVISA was 6.05% in 99,042 methicillin-resistant S. aureus (MRSA) strains and that of VISA was 3.01% in 68,792 MRSA strains. The prevalence of hVISA was 4.68% before 2006, 5.38% in 2006-2009, and 7.01% in 2010-2014. VISA prevalence was 2.05%, 2.63%, and 7.93%, respectively. In a subgroup analysis of different isolation locations, the prevalence of hVISA strains was 6.81% in Asia and 5.60% in Europe/America, and that of VISA was 3.42% and 2.75%, respectively. The frequencies of hVISA isolated from blood culture samples and from all clinical samples were 9.81% and 4.68%, respectively, and those of VISA were 2.00% and 3.07%, respectively. The most prevalent genotype was staphylococcal cassette chromosome mec (SCCmec) II, which accounted for 48.16% and 37.74% of hVISA and VISA, respectively. Sequence Type (ST) 239 was most prevalent.ConclusionThe prevalence of hVISA/VISA has been increasing in recent years, but has been grossly underestimated. Its incidence is higher in Asia than in Europe/America. hVISA is isolated from blood culture samples more often than from other samples. These strains are highly prevalent in epidemic MRSA strains. This study clarifies the epidemiology of hVISA/VISA and indicates that the detection of these strains and the control of nosocomial infections must be strengthened.

Project description:Although methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) strains with reduced susceptibility to vancomycin (RVS-MRSA; including vancomycin-intermediate S. aureus [VISA] and heterogeneous VISA [hVISA]) have been linked with vancomycin treatment failure, it is unclear whether they are more pathogenic than vancomycin-susceptible MRSA (VS-MRSA). We prospectively assessed patients with clinical MRSA isolates during a 10-month period to determine clinical status (infection versus colonization) and therapeutic outcome before correlating these findings with the results of detailed in vitro assessment of vancomycin susceptibility, including population analysis profile (PAP) testing. hVISA and VISA were defined by standard PAP criteria (area-under-the-curve ratio compared to that of the reference hVISA strain Mu3 [>or=0.9]) and routine CLSI criteria (vancomycin MIC, 4 to 8 microg/ml), respectively. Among the 117 patients assessed, 58 had RVS-MRSA isolates (56 hVISA and 2 VISA) and 59 had VS-MRSA isolates; the patient demographics and comorbidities were similar. RVS-MRSA was associated with a lower rate of infection than VS-MRSA (29/58 versus 46/59; P = 0.003), including a lower rate of bacteremia (3/58 versus 20/59, respectively; P < 0.001). The cure rates in RVS-MRSA and VS-MRSA groups were not statistically different (16/26 versus 31/42; P = 0.43), but the post hoc assessment of treatment regimes and study size made detailed conclusions difficult. The results of the macro method Etest correlated well with the PAP results (sensitivity, 98.3%, and specificity, 91.5%), but broth microdilution and our preliminary RVS-MRSA detection method correlated poorly. All isolates were susceptible to linezolid and daptomycin. These data suggest that detailed prospective laboratory identification of RVS-MRSA isolates may be of limited value and that, instead, such in vitro investigation should be reserved for isolates from patients who are failing appropriate anti-MRSA therapy.

Dataset Information

Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Uses the VraSR Regulatory System to Modulate Autophagy for Increased Intracellular Survival in Macrophage-Like Cell Line RAW264.7.

Publications

Heterogeneous Vancomycin-Intermediate <i>Staphylococcus aureus</i> Uses the VraSR Regulatory System to Modulate Autophagy for Increased Intracellular Survival in Macrophage-Like Cell Line RAW264.7.

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