Heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) and vancomycin-intermediate S. aureus (VISA) have been associated with treatment failure, and hVISA cannot be detected by routine disc diffusion test. Currently, there is no specific genetic marker for detection of these strains. The aim of the current study was to investigate the differential expression of six genes (viz., isaA, atl, rplN, pbp4, walK, walR) in VISA compared to those of hVISA and vancomycin-susceptible S. aureus (VSSA) isolates. The expression of six genes of 69 MRSA isolates (26 VSSAs, 28 hVISAs, and15 VISAs) was studied by using real-time PCR. The mean respective relative expression (i.e., the relative quantification (RQ) value) for five (isaA, atl, pbp4, walK, walR) vs. four (isaA, atl, walK, walR) genes in the VISA group was significantly higher than those in the VSSA and hVISA groups. In contrast, the respective mean RQ value for all genes in the hVISA and VSSA groups showed no significant difference. The mean RQ value for isaA in the VISA group showed the highest increased level compared with those in the hVISA (5.3-fold) and VSSA (9.1-fold) groups. The expression of six genes in VISA and hVISA compared to VSSA strains were analysed and found that isaA was up-regulated in VISA compared to hVISA and VSSA. The variation in gene expression levels reflects the diversity in genetic alterations of hVISA and VISA, involving the development of reduced vancomycin-susceptibility among hVISA and VISA.