Nobuyoshi Tachibana

Findings from the Miyazaki cohort study

The purpose of the Miyazaki Cohort Study is to describe and analyze the natural history of human T-cell lymphotropic virus type I (HTLV-I) in a highly endemic population in southwestern Japan. As of August 1995, 1,960 individuals have been enrolled, of whom 27% were HTLV-I antibody positive at baseline. Our achievements over the past decade of following this cohort include the identification of several viral markers that characterize high-risk carriers and the documentation that carriers have subclinical evidence of impaired cellular immunity. We have begun to estimate the impact of the infection on the health of carriers and have found that men are at greater risk of HTLV-I-associated diseases than women. We have been able to identify prospectively risk factors associated with sexual transmission. Most important, by identifying subclinical markers of pathogenesis, we hope to provide the foundation for developing interventions to prevent HTLV-I-associated disease.

Role of HTLV‐1 proviral DNA load and clonality in the development of adult T‐cell leukemia/lymphoma in asymptomatic carriers

Akihiko OKAYAMA*, Sherri STUVER, Masao MATSUOKA, Junzo ISHIZAKI, Gen-ichi TANAKA, Yoko KUBUKI, Nancy MUELLER, Chung-cheng HSIEH, Nobuyoshi TACHIBANA and Hirohito TSUBOUCHI Department of Internal Medicine II, Miyazaki Medical College, Miyazaki, Japan Department of Epidemiology, Boston University School of Public Health, Boston, MA, USA Department of Epidemiology, Harvard School of Public Health, Boston, MA, USA Laboratory of Virus Immunology, Institute for Virus Research, Kyoto University, Kyoto, Japan Division of Biostatistics and Epidemiology, University of Massachusetts Medical School Cancer Center, Boston, MA, USA Department of Nursing Science, Miyazaki Prefectural Nursing College, Miyazaki, Japan

Incident hepatitis C virus infection in a community-based population in Japan.

Hepatitis C virus (HCV) is an important cause of liver disease throughout the world. However, the natural history and pathogenesis of this infection is still not completely understood. The aim of this study was to characterize the evolution of incident, asymptomatic HCV infection in a community‐based population in Japan. The Miyazaki Cohort Study is a prospective study of adult residents in two villages, one of which has a very high prevalence of HCV. Nine hundred and seventy‐three people from this village were enrolled in the cohort between 1984 and 1995, with antibodies to HCV (anti‐HCV) found in 23%. During subsequent visits to annual health screens, new HCV seroconverters were identified among susceptible individuals, and their sequential samples were tested for anti‐HCV, HCV‐RNA, and HCV core antigen. Fourteen participants (six males, eight females) acquired anti‐HCV during the first 11 years of study follow‐up, at an incidence rate of 362 per 100 000 person‐years. Detectable HCV‐RNA and high anti‐HCV titres (> 1:2048) were observed for more than 5 years following seroconversion in 80% (8/10) of seroconverters with sufficient information, indicating the development of persistent infection in these subjects. Three (37.5%) of the eight sero converters with persistent infection had fairly consistent, albeit mild, alanine aminotransferase elevations (30–130 IU/L) during the study. Anti‐HCV seroconversions occurred at a very high rate in this community‐based population in Japan, in which this infection is endemic. Persistence also developed at a high frequency among the cases of newly acquired infection, although the associated liver enzyme abnormalities were mild.

Genetic Evidence of Transmission of Human T Cell Lymphotropic Virus Type 1 between Spouses

Sexual transmission of human T cell lymphotropic virus type 1 (HTLV-1) is considered to be an important route of infection in adults. However, no direct evidence has been reported that supports this observation. To address this issue, sequence variations of the gp46 (envelope)-coding region of HTLV-1 were determined in 13 patients infected with HTLV-1 who experienced seroconversion and in their spouses. Twenty-two nucleotide changes that were different from the reference sequence of lambdaATK-1 were identified. However, the gp46 sequences found were identical within each married couple. HTLV-1 proviral DNA loads measured in 11 of these couples varied from 10 to 3430 copies per 10(5) PBMC, and the proviral DNA loads of spouses often differed. This study provides the first genetic confirmation of the transmission of HTLV-1 from a carrier spouse to his/her partner. The findings also suggest that host-related factors play a more important role than do virus-specific factors in determining HTLV-1 proviral DNA load.

Autopsy Findings in 47 Cases of Adult T-cell Leukemia/Lymphoma in Miyazaki Prefecture, Japan

To identify factors that might improve the prognosis of adult T-cell leukemia/lymphoma (ATL), we reviewed data on 47 autopsied cases of ATL with reference to the complications and cause of death. The primary cause of death was respiratory insufficiency due to pulmonary infection. Respiratory insufficiency was also attributed to the diffuse alveolar damage and pulmonary fibrosis resulting from chemotherapy given and oxygen. About 90% of the cases had infections with one or more pathogens. Cytomegalovirus (CMV) was the most frequent pathogen involved in 35/47 (74.5%) while fungal infections were also commonly seen in 25 of the 47 cases (53.2%). Of these, 17 (70%) had pulmonary aspergillosis. Other neoplasias were present in 10 of the 47 cases, while hypercalcemia was evident in 21 patients. These findings suggest that the prevention and treatment of nosocomial infections and of drug-induced pulmonary toxicity may improve the prognosis and quality of life of ATL patients.

Predictors of level of circulating abnormal lymphocytes among human T-lymphotropic virus type I carriers in Japan

Human T‐lymphotropic virus type I (HTLV‐I) carriers often have abnormal lymphocytes (Ably) that resemble malignant cells of adult T‐cell leukemia (ATL). To identify predictors of the level of Ably in a longitudinal study of asymptomatic HTLV‐I carriers, we analyzed data from 215 subjects (67 men and 148 women) with multiple Ably measurements on blood smears. Ably+ (those having Ably > 0.6% of leukocytes counted on a blood smear at least once) was strongly associated with a high proviral<0B> <0R>load (OR 8.9; 95% CI 4.1, 19.5).<0B> <0R>The association among those defined as Ably++ (Ably > 0.6% at all screens or Ably > 1.6% at least once) was higher (19.7; 6.9, 56.1). Ably++ was also significantly associated with male gender (2.8; 1.0, 7.8). Multivariate analysis of Ably level indicates<0B> <0R>that men with a high proviral load, high anti‐HTLV‐I titer and low anti‐Tax reactivity have the highest Ably level. Int. J. Cancer 77:188–192, 1998.© 1998 Wiley‐Liss, Inc.

Sensitivity of Polymerase Chain Reaction Assay for Rickettsia tsutsugamushi in Patients' Blood Samples

We developed a nested polymerase chain reaction (PCR) method to detect Rickettsia tsutsugamushi (R. tsutsugamushi) DNA and determined its sensitivity. Primers were selected from the DNA sequence of the 58‐kDa group‐specific antigen gene of the Karp strain. The target sequence of rickettsial DNA was detectable as the band corresponding to 88 bp in 1.0 μg of the DNA extracted from BS‐C‐1 cells infected with R. tsutsugamushi. Rickettsia‐specific bands were observed not only for the homologous Karp strain, but also for four heterologous strains: two other reference strains (Gilliam and Kato) and two prototype strains prevalent in Miyazaki district (Irie and Hirano). The minimum copy number detectable by this method was estimated to be five rickettsiae. All of nine peripheral blood mononuclear cell samples from patients with tsutsugamushi disease who were seen 2‐11 days after disease onset tested positive for rickettsial DNA. The PCR assay method presented here could be a specific diagnostic tool for tsutsugamushi disease, especially in its early acute stage.

Increased expression of interleukin-2 receptor α on peripheral blood mononuclear cells in HTLV-I tax/rex mRNA-positive asymptomatic carriers

Using the double-nested reverse transcription-polymerase chain reaction, we assayed human T-cell leukemia virus type I (HTLV-I) tax/rex-encoded mRNA in the peripheral blood mononuclear cells (PBMCs) of asymptomatic carriers as an index of the expression of HTLV-I in vivo in relation to the proviral DNA level. HTLV-I tax/rex mRNA was detected in only 1 (3.3%) of 30 samples with medium or lower proviral DNA levels, but it was detected in 11 (39.3%) of 28 samples with high HTLV-I proviral DNA levels, estimated as equal to or more than the proviral DNA of 10 ng of HUT102 (i.e., HUT102 cells were used as positive controls). The mean number of interleukin-2 receptor alpha (IL-2R alpha)-positive cells as a percentage of the total number of PBMCs was higher (13.2%) in the tax/rex mRNA-positive carriers with high proviral DNA levels than in the carriers who were mRNA negative (8.4%) (p = 0.004, Wilcoxon test). These results suggest that virus activation as indicated by the presence of tax/rex mRNA in asymptomatic carriers with high proviral DNA levels is associated with an elevation of the IL-2R alpha-positive cells in vivo.

Sequential Change of Virus Markers in Seroconverters with Community-Acquired Infection of Human T Lymphotropic Virus Type I

Twenty-three human T lymphotropic virus type I (HTLV-I) seroconverters were identified among 1120 HTLV-I-seronegative adults followed up for 11 years in an area of Japan endemic for HTLV-I. The geometric mean titer of anti-HTLV-I was 1:453 in the first year after seroconversion; the titer of each subject did not change significantly during 2-10 years of follow-up. HTLV-I proviral DNA load was quantified in 15 seroconverters, and a broad range of levels was observed-from <10 to >1000 copies/10(5) peripheral blood mononuclear cells. However, there was no obvious change in HTLV-I proviral DNA load over several years within individual subjects. Therefore, both proviral DNA load and humoral response in adult HTLV-I seroconverters were shown to stabilize within a few years after initial infection. In addition, 1 subject tested positive for HTLV-I proviral DNA before antibody seroconversion, which suggests the existence of a window period in community-acquired infection.

Clinical features of OKT4+/OKT8+ adult T-cell leukemia.

Adult T-cell leukemia (ATL) has a range of clinical characteristics. Phenotypically the leukemic cells usually express the helper/inducer associated antigen OKT4 with lack of OKT8. We have observed three patients with acute ATL cytologically indistinguishable from OKT4+/OKT8- ATL but whose neoplastic cells had the unusual phenotype, OKT3+, OKT4+, OKT6-, OKT8+ OKT9+/-, OKT11+, Tac+/-, TdT-. All patients had abnormal karyotypes and antibodies against anti-ATL associated antigens as well as proviral DNA of human T-cell leukemia virus in the leukemic cells. The clinical course was complicated by skin eruptions, hypercalcemia, pulmonary infection and disseminated intravascular coagulopathy. All died of complications shortly after diagnosis. The clinical features of these patients were similar to those of OKT4+/OKT8- ATL. However, their acute course suggests that co-expression surface antigens OKT4 and OKT8 may be a sign of aggressive nature of the disease with poor prognosis.