Shunji Sakuma

Characterization of glycoprotein C-negative mutants of herpes simplex virus type 1 isolated from a patient with keratitis

SummaryRecently three strains of herpes simplex virus type 1 (HSV-1), which did not react with MicroTrak Herpes (Syva Co.), were isolated by us from a patient with recurrent herpetic keratitis. In this study we characterized these strains of HSV-1 and found them to be HSV-1 gC− mutants which are very rare isolates from humans. The properties of the HSV-1 strains regarding plaque morphology on Vero cells and chick embryo fibroblasts and viral DNA analysis were the same as those of the usual HSV-1 strains. An immunofluorescence study using anti-gC-1 monoclonal antibody and SDS-PAGE analysis of radiolabeled viral glycoproteins showed that these strains are deficient in gC-1. They were virulent for mice and sensitive to acyclovir and bromovinyldeoxyuridine. Furthermore the infectivity of the strains was inactivated by complement though the phenomenon was not observed in the usual HSV-1 strains. This finding suggests that protection from damages by complement is an important function of gC. In keratitis the effects of complement are thought to be minimal because of the scanty blood supply and this may be the reason why these strains were isolated from the cornea.

Herpes simplex virus type 1 infection in mice with severe combined immunodeficiency (SCID)

SummaryHerpes simplex virus type 1 (HSV-1) infection in mutant mice with severe combined immunodeficiency (SCID mice), i.e., mice in which the differentiation of both T and B lymphocytes is severely impaired, was studied.All control (infected and not treated with antibodies or with immune spleen cells) SCID mice were dead by 17 days after intracutaneous injection in the right midflank with 1 × 105 PFU of a virulent HSV-1 strain, Hayashida. Immunization with an avirulent strain of HSV-1 (SKa) did not protect them from death or prolong the survival time. Tissue virus titration of infected mice killed at various times after inoculation detected infectious virus in various organs, dorsal root ganglia, spinal cord, brain, kidney and adrenal gland in addition to the inoculation site of the skin in SCID mice, whereas virus could be detected only in the inoculation site and the nervous tissues in euthymic BALB/c mice, and in the adrenal gland from only one out of 17 nude mice.Human gamma globulin containing neutralizing antibody against HSV-1 prolonged the survival time but did not protect SCID mice from death. Transfer of spleen cells from immunized BALB/c mice protected the infected SCID mice from death. Treatment of spleen cells with anti-Thy 1.2 monoclonal antibody and complement abolished the protection.

An acyclovir-resistant strain of herpes simplex virus type 2 which is highly virulent for mice

SummaryHerpes simplex virus type 2 (HSV-2), strain YS-4 C-1, isolated by plaque cloning from a clinical isolate was found to be resistant to acyclovir (ACV; acycloguanosine) in vitro. It was sensitive to phosphonoacetic acid and 9-β-D-arabinofuranosyladenine. Thymidine kinase (TK) activity of YS-4 C-1 was less than 1% of that of other strains from the same clinical source. However, thymidine plaque autoradiography showed that YS-4 C-1 was not completely deficient in TK activity. YS-4 C-1 showed high virulence for mice like other HSV-2 strains which were sensitive to ACV. YS-4 C-1 was able to establish latent infection in mice. Virus isolated from the brain of a mouse died after being inoculated with YS-4 C-1 was also resistant to ACV. ACV was not effective in mice inoculated with YS-4 C-1. This study shows that not all ACV-resistant strains are avirulent for mice.

Urinary Concentrations of Cytokines in Patients with Pyelonephritis and Cystitis

We measured concentrations of interleukin-1β,-6 and-8, and tumor necrosis factor-α in the urine and blood of patients with urinary tract infections. We also examined the relationships between the severity of pyuria and concentrations of interleukin-6 or interleukin-8 in urine. Concentrations of interleukin-6 and-8 were significantly higher in the urine of patients with urinary tract infections than that of healthy subjects or patients with fever unrelated to the urinary tract infection. A close relationship was observed between the severity of pyuria and the concentration of interleukin-8. Concentrations of interleukin-6 and-8 in the blood were higher in patients with pyelonephritis than in patients with cystitis. These results suggest that the interleukin-6 and-8 were generated in the urinary tract during pyelonephritis and cystitis, and that the interleukin-8 induced leukocyte influx into the urine.

Recurrent Herpetic Keratitis: Failure to Detect Herpes simplex Virus Infection Using the Syva MicroTrakTM HSV1/HSV2 Direct Specimen Identification/Typing Test

A 35-year-old man had developed recurrent herpetic keratitis characterized by dendritic keratitis at intervals of a year. We were able to culture cytopathic agents repeatedly from his lesions by inoculating Vero cells. The cultures yielded definitive evidence of a virus that caused a cytopathic effect within 3 days. However, these virus strains could not be identified as herpes simplex virus (HSV) in immunofluorescence assays using the Syva MicroTrak HSV1/HSV2 direct specimen identification/typing test. Rather they were identified as strains of HSV type 1 (HSV-1) on the basis of plaque morphology, neutralization tests, electron-microscopic examination and DNA restriction endonuclease analysis. Our results allow us to assume the existence of HSV-1 strains isolated clinically that are negative to analysis using the Syva Micro-Trak HSV1/HSV2 direct specimen identification/typing test.

Persistent Infection with Herpes Simplex Virus Type 1 in an Ia Antigen-Positive Murine Macrophage Cell Line

The interaction of herpes simplex virus type 1 (HSV‐1) with murine macrophage cell lines was examined. The cell lines appeared to be moderately permissive for HSV‐1 replication, though the yield of the virus was limited compared with that in Vero cells. Furthermore, the murine macrophage cell line SL‐1, bearing Ia antigen, was persistently infected with HSV‐1 for over one year, and was designated SL‐1/KOS. Persistent infection could not be established in an Ia antigen‐negative macrophage cell line, SL‐4. In the SL‐1/KOS culture, there was a small number of infected cells as revealed by infectious center assay. Treatment with monoclonal antibody against HSV‐1 cured the persistent infection. Therefore maintenance of the persistent infection is considered to be due to a carrier culture consisting of a minority of infected cells and a majority of uninfected cells. In the SL‐1/KOS cultures a low level of interferon (IFN) was found. When a large amount of exogenous recombinant murine IFN‐β (105–106 international units/ml) was added to the culture, virus production diminished to undetectable levels. These results suggest that IFN plays an important role in the maintenance of persistent infection. In long‐term persistently infected cultures, syncytium formation appeared and the virus from such cultures had a different DNA structure from that of the virus originally used for infection as revealed by restriction endonuclease analysis.

Seroepidemiology of Herpes Simplex Virus in Yogyakarta, Indonesia

Sera from 487 individuals in Yogyakarta, Indonesia, were tested for antibodies to herpes simplex virus (HSV) by a passive hemagglutination method. Age‐specific incidence rates for antibodies to HSV were calculated. For sera from persons other than prostitutes, in the age group from 10 to 19, the positive rate was 48% but in the age group higher than 20, it was more than 87%. Fifty of 59 pregnant women (85%) were positive. The positive rate and the distribution of antibody levels in prostitutes were higher than in the general population.