Restriction endonuclease fragment length polymorphism (RFLP) is useful for the epidemiological study of herpes simplex virus type 1 (HSV-1). We report here the identification of a major BglII RFLP variant of HSV-1, designated BgK L , found in 27.0% of 636 HSV-1 clinical isolates. We have also established its geographic distribution in Japan. BgK L has an unusually large BglII K fragment. SalI cleavage analyses showed that 97% of BgK L variant isolates lack both the SalI C-J and the F-J cleavage sites and have an unusually large SalI D or E fragment, and 91% of the BgK L variants lack both SalI G and H fragments. Furthermore, 96% of BgK L isolates have an unusually small KpnI M fragment. Therefore, BgK L is a marker for these five mutations in most HSV-1 isolates and is a useful HSV-1 RFLP marker. The BgK L variant was found in 59% of HSV-1 isolates from Shikoku Island, 44% of HSV-1 isolates from the Chugoku region of Honshu Island, 31% of HSV-1 isolates from Kyushu Island, 0% of HSV-1 isolates from Okinawa Island, 49% of HSV-1 isolates from Osaka, 27% of HSV-1 isolates from Shiga, 13% of HSV-1 isolates from the Chubu Region, and 9% of HSV-1 isolates from the Tohoku Region of Honshu Island. Differences in the frequency of BgK L between the ShikokuChugoku-Osaka area (49%) and Kyushu, between Kyushu and Okinawa, between the Shikoku-Chugoku-Osaka area and Shiga, and between Shiga and Tohoku are all statistically significant. The BgK L frequency decreases in a geographical gradient suggest that this HSV-1 variant was dispersed from Shikoku to the surrounding regions and then to more distant regions. The BgK L frequency in Tokyo was similar to the nationwide average. These are the first data to suggest a geographic and demographic dispersion pattern of HSV-1. Implications for the epidemiology and diversification of HSV-1 are discussed.DNA restriction endonuclease cleavage patterns and restriction fragment length polymorphism (RFLP) are useful for epidemiological studies of herpes simplex virus type 1 (HSV-1) strains (7,14,15,25,28,29). RFLP profiles have been used to differentiate HSV-1 clinical isolates from epidemiologically unrelated individuals and to determine whether viruses isolated from epidemiologically "connected" individuals are identical (1-3, 5, 9, 12, 13, 16, 18, 19, 23, 26, 28, 36). The data indicate that the genomic DNAs of HSV-1 isolates have accumulated enough base substitutions to allow different isolates to be distinguished. HSV-1 polymorphism also has been observed by differences in the electrophoretic patterns of virion structural proteins of different HSV-1 strains (17,21,28). The lifelong latency and reactivation of HSV-1 infection probably facilitates the accumulation of random mutations and diversification of 37,38). HSV-1 isolates with distinct base substitution mutation(s) at restriction endonuclease cleavage sites are designated RFLP variants in here.It is not known whether an HSV-1 RFLP variant isolated at a certain frequency in a particular population is due to an RFLP variation that aro...

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Attempts were made to improve the rate of isolation of herpes simplex virus (HSV) from clinical specimens by minimizing loss of virus infectivity during transportation and employing the most sensitive cells for isolation. Basical analyses using standard strains of type 1 and type 2 HSV indicated that virus titer decrease was marked even at low temperatures in environments free of proteinous stabilizer such as normal serum or tissue extract, negating the generally held concept that HSV is stable in distilled water. YLE (Earle-lactalbumin hydrolysate-yeast extract) medium containing 20% inactivated calf serum was determined to be a transport medium of choice, because degradation of suspended virus during storage and freeze-thawing was negligible and loss of virus during Millipore filtration was minimal. Special coating of the membrane could also be obviated by the use of this solution. In a cell susceptibility test using clinical specimens, secondary rabbit kidney (SRK) cells were the most sensitive, showing a quick development of cytopathic effect. Vero and RK-13 cells were the second best, whereas monkey kidney, HeLa and L cells were far less sensitive. A total of 136 specimens from suspected cases, sent by dermatologists, were tested using SRK cells, and 99 strains of type 1 and 15 strains of type 2 HSV were isolated. Excluding one case from which vaccinia virus was isolated, the isolation rate of HSV was 84.4%.