WalK gene mutation in Vancomycin Intermediate Staphylococcus aureus (VISA) clinical strains

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WalK gene mutation in Vancomycin Intermediate Staphylococcus aureus (VISA) clinical strains

 

Author: Chow, Miu Yee Mandy
Title: WalK gene mutation in Vancomycin Intermediate Staphylococcus aureus (VISA) clinical strains
Degree: M.Sc.
Year: 2013
Subject: Vancomycin resistance.
Methicillin resistance.
Staphylococcus aureus.
Mutation (Biology)
Hong Kong Polytechnic University -- Dissertations
Department: Dept. of Health Technology and Informatics
Pages: xi, 98 leaves : ill. (some col.) ; 30 cm.
Language: English
InnoPac Record: http://library.polyu.edu.hk/record=b2637224
URI: http://theses.lib.polyu.edu.hk/handle/200/7062
Abstract: Background: Although Clinical and Laboratory Standards Institute defines isolates with minimal inhibitory concentration (MIC) ≤ 2 mg/L as vancomycin susceptible Staphylococcus aureus (S. aureus), treatment failure and poor clinical outcomes has been reported even in apparently susceptible strains. Routine identification of clinical isolates with reduced vancomycin susceptibility remains challenging and no standardizes method or molecular marker exists to aid in diagnosis of vancomycin intermediate S. aureus (VISA) strains. Aims: VISA is thought to be generated from vancomycin-susceptible S. aureus in the presence of vancomycin by multiple spontaneous mutations. A mutation in the two-component system (TCS), vraSR and graRS, has previously been reported to be responsible for the VISA phenotype in studies of Hong Kong and Japanese isolates. Researchers elsewhere have observed genes in another TCS, walk, which has not been investigated in clinical isolates locally. Hence, this study was investigated changes in walK, during development of vancomycin non-susceptibility in a patient with methicillin-resistant S. aureus (MRSA) in Hong Kong. Methods: The vancomycin MIC of clinical MRSA strains was determined using spiral gradient endpoint technique. The deoxyribonucleic acid of the samples were extracted and amplified by polymerase chain reaction using primers for walk. The amplicons were sent out for sequencing and then compared with the MRSA control strain (N315) to look for walK gene mutation in these clinical isolates. Results: Two of the ten clinical MRSA strains were VISA. Their vancomycin MICs were 6.002 mg/L and 4.12 mg/L. These strains were the only ones of the ten examined that exhibited mutations in the walK gene. One of the strains carried five non- synonymous (NS) mutations and the other carried three. Comparison of this study with others investigating changes in TCSs it appeared that changes in walK occurred with higher frequency than those in other TCS. Conclusion: The mechanism of vancomycin resistance is quite complicated as the development of VISA includes a gradual process rather than a critical threshold. But this study supported the hypothesis that mutation in the walK TCS occurred with high frequency in the clinical VISA strains. The major mutation regions found in this study were in the HAMP and PAS areas of the walK gene, again reflecting similar findings to other studies. It is hoped that further knowledge of common mutation may allow for the development of a molecular marker as a diagnostic tool and possibly for new antibiotics for the treatment of reduced susceptibility to vancomycin of S. aureus.

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