|Author:||Un, Cheuk Hang Henry|
|Title:||WalK gene : mutation detection in non-susceptible Staphylococcus aureus|
Hong Kong Polytechnic University -- Dissertations
|Department:||Department of Health Technology and Informatics|
|Pages:||iv, 121 leaves : col. ill. ; 30 cm.|
|Abstract:||Vancomycin is the drug of choice to treat methicillin resistant Staphylococcus aureus (MRSA). However, wide use of vancomycin leads to the formation of heterogeneous vancomycin intermediate Staphylococcus aureus (hVISA) and vancomycin intermediate Staphylococcus aureus (VISA) from MRSA or other S. aureus strains and results in a problem of successful vancomycin therapy hVISA and VISA. The main reason of vancomycin treatment failure is that it is hardly to be penetrated into the S. aureus cytoplasmic content due to the increased thickness of its cell wall. Reports have revealed that gene expression play an important role in cell wall formation and it was also demonstrated that genetic mutation of some genes can alter cell wall formation. Therefore, investigations of these related genes are essential for understanding the relationship between the genotypic and phenotypic effect. This study aimed to investigate mutational detection in walK gene across 29 laboratory induced hVISA and VISA strains. The analyzed data was then compared to a normal MSSA strain N315.|
The 6 Laboratory induced strains in 5 different stages were cultured. The DNA from each strain was then amplified with satisfactory concentration and purity. A polymerase chain reaction across all strains were done and the amplified products were sent to Genome Research Centre, The University of Hong Kong for sequencing. Finally, the sequenced data were analyzed using multiple sequence alignment software Multalin. Results in this study indicated that different strains have different degree of mutations, however, the type of mutations are quite inconsistent. While comparing with the normal strain VSSA N315, amino acid deletion at position _354G is the most common mutation found across the selected strains. In addition, amino acid substitution at positions R222K, W358G and Y367D are the most common mutations observed. Finally, positions at Q219STOP and Q448STOP are the mostly found stop codon mutations in the studies strains. It was already reported previously that mutations in walK play an important role in cell wall biosynthesis and cell autolytic activity (by only amino acid substitution). However, stop codon mutations were first observed in our study. WalK presented in these strains with an incomplete amino acid sequences would predictably have an impact on cell wall formation. Further investigations are necessary to check on how stop codon impacts the development and loss of vancomycin non-susceptibility. It was summarized that stop codon mutation is still an area to be addressed whether terminating of protein synthesis would whether delaying the development of VISA formation or enhancing cell wall biosynthesis and increase vancomycin non-susceptibility. This will be the first goal working on impact on stop codon and it is hopefully that further investigation in this study can focus on tested strains' phenotypic changes (in terms of cell wall thickness).
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