Accumulating evidence indicates that herpesviruses may be putative pathogens in various types of periodontal diseases. The present study was performed to examine infections with different genotypes of human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) in subgingival samples from a Chinese population and to analyze the correlation with periodontal status. A nested PCR assay was used to identify the presence of HCMV, EBV type 1 (EBV-1), and EBV-2; and the amplicons were further analyzed by restriction fragment length polymorphism analysis. HCMV was detected in 79.0% of 143 chronic periodontitis (CP) patients, 78.5% of 65 gingivitis patients, and 76.3% of 76 periodontally healthy individuals, while EBV was found in 63.6%, 32.3%, and 30.3% of the three groups of subjects, respectively. The HCMV-positive PCR products from all the samples were identified as corresponding to gB genotype I (gB-I) or gB-II. HCMV gB-II (62.9%), EBV-1 (43.4%), and EBV-2 (18.2%) were associated with CP at higher frequencies (P < 0.05), whereas HCMV gB-I was more often observed in gingivitis patients (40.0%) and healthy individuals (40.8%) (P < 0.05). Furthermore, a higher rate of coinfection with HCMV and EBV was shown in CP patients (52.4%), especially dual infections with HCMV gB-II and EBV-1 (30.8%) or HCMV gB-II and EBV-2 (12.6%), compared with the rates of single infections with HCMV or EBV (P < 0.05). Infection with HCMV gB-II, EBV-1, or EBV-2 was correlated with higher rates of bleeding on probing (P < 0.05). In patients infected with HCMV gB-II or both HCMV and EBV, including HCMV gB-II and EBV-1, a deeper probing depth or more serious attachment loss was found (P < 0.05). These findings clearly indicate that HCMV gB-II is the dominant genotype detected in subgingival samples in CP. HCMV gB-II infection and HCMV gB-II coinfection with EBV-1 are closely associated with periodontal tissue inflammation and destruction.