RNA-seq analysis reveals resistome genes and signalling pathway associated with vancomycin-intermediate Staphylococcus aureus

Context: Vancomycin-intermediate Staphylococcus aureus remains one of the most prevalent multidrug-resistant pathogens causing healthcare infections that are difficult to treat. Aims: This study uses a comprehensive computational analysis to systematically investigate various gene expression profiles of resistant and sensitive S. aureus strains on exposure to antibiotics. Settings and Design: The transcriptional changes leading to the development of multiple antibiotic resistance were examined by an integrative analysis of nine differential expression experiments under selected conditions of vancomycin-intermediate and -sensitive strains for four different antibiotics using publicly available RNA-Seq datasets. Materials and Methods: For each antibiotic, three experimental conditions for expression analysis were selected to identify those genes that are particularly involved in the development of resistance. The results were further scrutinised to generate a resistome that can be analysed for their role in the development or adaptation to antibiotic resistance. Results: The 99 genes in the resistome are then compiled to create a multiple drug resistome of 25 known and novel genes identified to play a part in antibiotic resistance. The inclusion of agr genes and associated virulence factors in the identified resistome supports the role of agr quorum sensing system in multiple drug resistance. In addition, enrichment analysis also identified the kyoto encyclopedia of genes and genomes (KEGG) pathways – quorum sensing and two-component system pathways – in the resistome gene set. Conclusion: Further studies on understanding the role of the identified molecular targets such as SAA6008_00181, SAA6008_01127, agrA, agrC and coa in adapting to the pressure of antibiotics at sub-inhibitory concentrations can help in learning the molecular mechanisms causing resistance to the pathogens as well as finding other potential therapeutics.

Prevalence and Clinical Impact of Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Isolated From Hospitalized Patients

BACKGROUND: We estimated the prevalence and clinical impact of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA). The concordance between macromethod and glycopeptide resistance detection (GRD) E tests was determined. In addition, predictors of clinical outcomes in hospitalized patients with S. aureus bacteremia (SAB) or pneumonia (SAP) were evaluated. METHODS: We obtained 229 consecutive S. aureus isolates from all hospitalized patients at two university hospitals located in Busan and Yangsan, Korea. Standard, macromethod, and GRD E tests were performed. Additionally, we reviewed the medical records of all patients. Among the 229 patients, predictors of clinical outcomes were analyzed for 107 patients with SAB and 39 with SAP. RESULTS: Among the 229 isolates, 34.5% of S. aureus isolates and 50.7% of methicillin-resistant S. aureus isolates exhibited the hVISA phenotype based on the macromethod E test. hVISA was nearly associated with treatment failure in patients with SAB (P=0.054) and was significantly associated with treatment failure in patients with SAP (P=0.014). However, hVISA was not associated with 30-day mortality in patients with SAB or SAP. The concordance between the macromethod and GRD E tests was 84.2%. CONCLUSIONS: hVISA is quite common in the southeastern part of Korea. hVISA is associated with treatment failure in patients with SAP.

Molecular Typing and Resistance Profiles of Vancomycin-Intermediate Staphylococcus aureus in Korea: Results from a National Surveillance Study, 2007-2013 / 대한임상미생물학회지

BACKGROUND: To investigate the national molecular epidemiology and resistance profiles of vancomycin-intermediate Staphylococcus aureus (VISA), we analyzed the characteristics of methicillin-resistant Staphylococcus aureus (MRSA) collected from clinical samples at tertiary or general hospitals participating in a nationwide surveillance program for VISA and vancomycin-resistant Staphylococcus aureus (VRSA) in Korea during an 12-week period in each year from 2007 to 2013. METHODS: VISA was defined by agar dilution, broth dilution and E-test methods with vancomycin minimum inhibitory concentrations of >2 μg/mL. All VISA isolates were characterized by multilocus sequence typing, staphylococcal cassette chromosome mec typing, spa typing, accessory gene regulator typing, Diversilab analysis, and antibiogram analysis. RESULTS: Of 109,345 MRSA isolates, 87,354 were screened and 426 isolates were identified as positive on brain heart infusion agar containing 4 μg/mL vancomycin (BHI-V4). Of 426 isolates, 76 isolates were identified as VISA. No VRSA isolates were detected among the isolates. Overall, a total of 6 genotypes were identified among VISA strains and the predominant clones were ST5-II-t2460, ST72-IV-t324, and ST239-III-t037 (44.7%, 15.8%, and 10.5%, respectively). Of note, ST72-IV-t324 clones are known to be a typical community-associated MRSA. ST239-III-t037 strains were more resistant to trimethoprim-sulfamethoxazole than any other type of strain. ST72-IV-t324 strains were susceptible to all of the antimicrobial agents tested except erythromycin and daptomycin. All of the VISA isolates were susceptible to linezolid and quinupristin-dalfopristin. CONCLUSION: Although VRSA is still rare, continuous monitoring of VRSA occurrence is needed, as well as VISA prevalence, epidemic clonal shift, and antimicrobial resistance.

Epidemiology of heterogeneous vancomycin-intermediate Staphylococcus aureus from sterile body fluid specimens / 国际检验医学杂志

Objective To investigate the prevalence and molecular characteristic of heterogeneous‐ vancomycin intermediate Staphylococcus aureus(hVISA) among methicillin‐resistant Staphylococcus aureus (MRSA)strains isolated from sterile body fluid specimens from 2009 to 2013 in Huangzhou District People′s Hospital in Huanggang City .Methods The minimum inhibitory con‐centrations (MIC) of antibiotics was determined by agar dilution method .The hVISA strains were detected by population analysis profile/area under the curve method (PAP/AUC) .The staphylococcal cassette chromosome mec (SCCmec) ,multilocus‐sequence typing (MLST) ,accessory gene regulator (agr) and Staphylococcus aureus protein A(spa) typing of hVISA strains were detected using PCR method .Results 32 hVISA strains were detected among 285 MRSA strains ,the prevalence rate of hVISA was 11 .2% , and the detection rates of hVISA from 2009 to 2013 were 0 .0% ,6 .4% ,9 .0% ,14 .3% and 18 .8% ,respectively ,showed an increas‐ing trend .The main hVISA epidemic clone was ST239‐SCCmecIII‐ t030‐agrI type(28strains ,accounting for 87 .5% ) .Conclusion The detection rate of hVISA showed an increasing trend in the past 5 years ,should be paid attention to strictly control the utiliza‐tion of glycopeptide drugs .

The incidence and risk factors for heterogeneous vancomycin intermediate Staphylococcus aureus / 中华内科杂志

Objectives To investigate the prevalence of heterogeneous vancomycin intermediate Staphylococcus aureus(hVISA) and the sensitivity of hVISA to novel antibiotics,and to explore the risk factors and infection attributable mortality associated with hVISA infection.Methods A total of 456 methicillin resistant Staphylococcus aureus (MRSA) isolates were isolated in Zhongshan Hospital from January,2008 to November,2010.All MRSA isolates were investigated for hVISA by two agar screening methods BHIA5T (brain-heart infusion containing teicoplanin 5 mg/L)or BHIA6V (brain-heart infusion containing vancomycin 6 mg/L),as well as macroEtest method(MET).Possible hVISA isolates were tested by modified population analysis profile-area under the curve (PAP-AUC).The minimal inhibitory concentrations(MICs) of vancomycin,teicoplanin and linezolid were determined by microbroth dilution as recommended by Clinical Laboratory Standards Institute(CLSI).The contribution difference between hVISA and vancomycin susceptible Staphylococcus aureus (VSSA) in different MIC range was compared.A retrospective case-control study of the patients with hVISA infection or VSSA infection was carried out and statistical analysis was performed using t test,Mann-Whitney test,x2 test and Fisher exact test.Results A total of 105 isolates of hVISA were screened by BHIA5T and BHIA6V (23.0％) with other 23 isolates by MET(5.0％) and 21 by PAP-AUC(4.6％).All isolates were 100％ sensitive to vancomycin,teicoplanin and linezolid.The vancomycin MIC [(1.76 ±-0.16) mg/L] in hVISA group was significantly higher than that in VSSA group[(1.09 ± 0.07)mg/L,P ＜ 0.01],which was a potential risk factor for hVISA infection.The retrospective study showed chronic obstructive pulmonary disease (COPD) was also a risk factor for hVISA infection of the lower respiratory tract.No significant difference in infection attributable mortality was showed between the hVISA group and the VSSA group.Conclusions The overall prevalence of hVISA in Zhongshan Hospital is estimated as 4.6％,while the prevalence of hVISA isolated from blood is as high as 12.5％.All isolates are 100％ sensitive to vancomycin and linezolid.COPD is a risk factor for hVISA infection of the lower respiratory tract.

Be alert to the alterations in the biological characteristics in heterogeneous vancomycin-intermediate Staphylococcus aureus.

The development of reduced vancomycin susceptibility in Staphylococcus aureus in many cases appears to be associated with characteristic changes. These changes may have pitfall of identifying S. aureus by automated testing methods like Vitek 32. In this study, we retested 24 heterogeneous vancomycin-intermediate Staphylococcus haemolyticus (h-VISH) collected in 2008-2010 at the Department of Clinical Microbiology by conventional biochemical tests and polymerase chain reaction (PCR). The heterogeneous vancomycin-intermediate S. aureus (hVISA) reversion test and electron microscopic examination were also used. Six isolates of 24 h-VISH possessed nuc, coa, and 16S rRNA genes, and could be reversed into S. aureus. It suggested that biochemical and morphological changes in hVISA and vancomycin-intermediate S. aureus (VISA) should be considered, and the detection of S. aureus, especially reduced vancomycin susceptibility isolates, requires more attention and different techniques.

Two Cases of Vancomycin-intermediate Staphylococcus aureus Isolated from Joint Tissue or Wound / 대한진단검사의학회지

Since its first isolation in 1997, vancomycin-intermediate Staphylococcus aureus (VISA) has been a clinical concern because it may lead to treatment failure. Up to the present, there were two reports of clinical VISA cases in Korea. We now report two additional cases of VISA with the minimum inhibitory concentration (MIC) of 4 microgram/mL. The first patient was a 59 yr-old man who had undergone total hip replacement arthroplasty in 1999 due to avascular necrosis of femur heads. He had recurrent episodes of infected hip caused by methicillin-resistant Staphylococcus aureus (MRSA) and was treated with vancomycin. He underwent replacement operation of prosthesis. Cultures of joint fluid and joint tissue grew S. aureus. Vancomycin MIC as determined by a broth microdilution method was 4 microgram/mL for the both isolates. The patient was treated with high enough doses of vancomycin to maintain serum trough concentrations at 20-25 microgram/mL for 52 days and was discharged. The second patient was a 57 yr-old man with diabetes. He lost consciousness from drinking. After recovery of consciousness, he was diagnosed with aspiration pneumonia. MRSA and Acinetobacter baumannii were cultured from sputum and the patient was treated with vancomycin and meropenem. During hospitalization, bed sores developed in his ankle and back. A wound culture from the sore grew S. aureus with vancomycin MIC of 4 microgram/mL. Since infection was localized, systemic antibiotics did not seem necessary, and the patient was transferred to another hospital for isolation and management.