Tetsushi Yoshikawa

Classification of HHV-6A and HHV-6B as distinct viruses

Shortly after the discovery of human herpesvirus 6 (HHV-6), two distinct variants, HHV-6A and HHV-6B, were identified. In 2012, the International Committee on Taxonomy of Viruses (ICTV) classified HHV-6A and HHV-6B as separate viruses. This review outlines several of the documented epidemiological, biological, and immunological distinctions between HHV-6A and HHV-6B, which support the ICTV classification. The utilization of virus-specific clinical and laboratory assays for distinguishing HHV-6A and HHV-6B is now required for further classification. For clarity in biological and clinical distinctions between HHV-6A and HHV-6B, scientists and physicians are herein urged, where possible, to differentiate carefully between HHV-6A and HHV-6B in all future publications.

Dual roles for the telomeric repeats in chromosomally integrated human herpesvirus-6

Approximately 1 percent of healthy individuals carry human herpesvirus-6 within a host chromosome. This is referred to as chromosomally integrated herpesvirus-6 (CIHHV-6). In this study, we investigated the chromosomal integration site in six individuals harboring CIHHV-6B. Using FISH, we found that HHV-6B signals are consistently located at the telomeric region. The proximal endpoints of the integrated virus were mapped at one of two telomere-repeat-like sequences (TRSs) within the DR-R in all cases. In two cases, we isolated junction fragments between the viral TRS and human telomere repeats. The distal endpoints were mapped at the distal TRS in all cases. The size of the distal TRS was found to be ~5 kb which is sufficient to fulfill cellular telomeric functions. We conclude that the viral TRS in the DR regions fulfill dual functions for CIHHV-6: homology-mediated integration into the telomeric region of the chromosome and neo-telomere formation that is then stably transmitted.

Cytokine and chemokine responses in the blood and cerebrospinal fluid of patients with human herpesvirus 6B-associated acute encephalopathy with biphasic seizures and late reduced diffusion

Acute encephalopathy with biphasic seizures and late reduced diffusion has become increasingly common among various types of human herpesvirus 6B (HHV‐6B) encephalitis at the time of primary viral infection. The aim of the present study is to explore the pathophysiology of HHV‐6B‐associated acute encephalopathy with biphasic seizures and late reduced diffusion. Five cytokines and five chemokines were measured in serum and cerebrospinal fluid (CSF) obtained from 12 HHV‐6B‐associated acute encephalopathy with biphasic seizures and late reduced diffusion patients and 19 control exanthem subitum (without complications) patients. Serum interleukin (IL)‐10 (P = 0.007) and IL‐8 (P = 0.025) were significantly higher in the patients with the disease than controls. Serum IL‐1β (P = 0.034) and monocyte chemoattractant protein (MCP)‐1 (P = 0.002) were significantly higher in the controls than patients with the disease. In patients with the disease, IL‐10 (P = 0.012), regulated on activation normal T cell expressed and secreted (RANTES; P = 0.001), and monokine induced by interferon γ (MIG; P = 0.001) were significantly higher in serum than CSF, meanwhile IL‐6 (P = 0.034), IL‐8 (P = 0.034), and MCP‐1 (P = 0.001) were significantly higher in CSF than serum. Additionally, serum IL‐10 was significantly higher in the disease patients with sequelae than those without sequelae (P = 0.016). Several cytokines and chemokines may be associated with the pathogenesis of acute encephalopathy with biphasic seizures and late reduced diffusion. Moreover, the regulation of cytokine networks appears to be different between peripheral blood (systemic) and central nervous system. J. Med. Virol. 86:512–518, 2014.

Pathogenic Role of Human Herpesvirus 6B Infection in Mesial Temporal Lobe Epilepsy

BACKGROUND Human herpesvirus 6B (HHV-6B) is the causative agent for exanthem subitum. HHV-6B was associated with mesial temporal sclerosis (MTS), leading to mesial temporal lobe epilepsy (MTLE). In this study, we sought to elucidate the pathogenic role of HHV-6B in patients with MTLE. METHODS Seventy-five intractable MTLE patients, including 52 MTS patients and 23 non-MTS patients, were enrolled in this study. Resected hippocampus, amygdala, and mixed samples of amygdala and uncus samples were examined by real-time polymerase chain reaction (PCR) and reverse-transcriptase PCR to detect viral DNA and messenger RNA (mRNA), respectively. Host gene expressions, including neural markers, were measured using the TaqMan Gene Expression Assay. RESULTS Detection of HHV-6 DNA was higher in MTS patients than non-MTS patients (median/interquartile range: 19.1/0-89.2 vs 0.0/0.0-0.0 copies/µg DNA; P = .004). HHV-6B viral DNA was determined in 12/27 HHV-6 DNA-positive samples, and no HHV-6B mRNA were detected in all samples. In MTS patients, expression of monocyte chemotactic protein-1 (P = .029) and glial fibrillary acidic protein (P = .043) were significantly higher in the amygdala samples with HHV-6 DNA than those without viral DNA. CONCLUSIONS This study suggests that HHV-6B may play an important role in the pathogenesis of MTS via modification of host gene expression.

Analysis of the shedding of three β-herpesviruses in urine and saliva of children with renal disease.

Cytomegalovirus (CMV), human herpesvirus 6 (HHV‐6) and 7 (HHV‐7) are important pathogens in immunocompromised patients. To elucidate the kinetics of the three β‐herpesviruses in saliva and urine samples were collected serially from children with renal diseases. Twenty children with renal diseases were enrolled in this study. A total of 240 saliva and urine samples were collected monthly from the patients over a 1‐year period. Viral DNAs loads were measured by real‐time PCR. In 10 CMV seropositive patients CMV DNA was detected rarely in saliva and CMV DNA load was lower than the other two β‐herpesviruses DNA loads. All patients were seropositive for HHV‐6B and the virus was detected frequently in saliva. Two of 20 patients were HHV‐7 seronegative. High copies of viral DNA were detected continuously in saliva of the HHV‐7 seropositive patients. Although neither CMV nor HHV‐6B DNA load was different among the three renal diseases, HHV‐7 DNA load was different among the diseases (P = 0.039). HHV‐6B DNA loads were significantly higher in patients with immunosuppressive treatment compared to those without treatment (P = 0.013). Although CMV DNA was detected in urine samples collected from 5 of 10 CMV seropositive patients, HHV‐6B and HHV‐7 DNA were detected at relatively low frequencies in urine. No remarkable temporal associations between viral DNA excretion and proteinuria or immunosuppressive treatment were demonstrated. The pattern of viral DNA excretion in saliva and urine were different among the three viruses. No temporal correlation was observed between viral infection and renal diseases. J. Med. Virol. 86:505–511, 2014.

Nationwide survey of rotavirus-associated encephalopathy and sudden unexpected death in Japan

BACKGROUND Rotavirus can cause severe complications such as encephalopathy/encephalitis and sudden unexpected death. The incidence of rotavirus-associated encephalopathy/encephalitis or sudden unexpected death remains unknown. To clarify the clinical features of rotavirus-associated encephalitis/encephalopathy and sudden unexpected death, we conducted a nationwide survey in Japan. METHOD A two-part questionnaire was designed to determine the number of the cases and the clinical features of severe cases of rotavirus infection, including encephalitis/encephalopathy and sudden unexpected death, between 2009 and 2011. RESULT Of the 1365 questionnaires sent to hospitals, 963 (70.5%) were returned and eligible for analysis. We determined 58 cases of rotavirus-associated encephalitis/encephalopathy and 7 cases of sudden unexpected death. These patients were diagnosed with rotavirus infection by immunochromatography. Although 36/58 (62.1%) encephalitis/encephalopathy patients had no sequelae, 15/58 (25.9%) patients had neurological sequelae, and 7/58 (12.1%) patients had fatal outcomes. Pleocytosis was observed in 9/40 (22.5%) patients and cerebrospinal fluid protein levels were elevated in only 4/40 (10%) patients. Elevated lactate dehydrogenase (LDH) (>500 IU/L) or acidemia (pH<7.15) were related to a poor prognosis. CONCLUSION We estimate that annual cases of rotavirus-associated encephalitis/encephalopathy and sudden unexpected death were 44.0 and 4.9 cases in Japan, respectively. Elevated LDH (>500 IU/L) or acidemia (pH<7.15) were related to a poor prognosis of the encephalitis/encephalopathy.

Virological analysis of inherited chromosomally integrated human herpesvirus-6 in three hematopoietic stem cell transplant patients.

We analyzed 3 hematopoietic stem cell transplant (HSCT) recipients with inherited chromosomally integrated human herpesvirus‐6 (inherited CIHHV‐6). Cases 1 (inherited CIHHV‐6A) and 2 (inherited CIHHV‐6B) were inherited CIHHV‐6 recipients. Case 3 received bone marrow from a donor with inherited CIHHV‐6B. Following HSCT, HHV‐6B was isolated from Case 1. HHV‐6A and ‐6B messenger RNAs were detected in Cases 1 and 3.

Universal varicella vaccine immunization in Japan.

In 1974, Japanese scientists developed a live attenuated varicella vaccine based on the Oka strain. The efficacy of the vaccine for the prevention of varicella has been primarily demonstrated in studies conducted in the United States following the adoption of universal immunization using the Oka strain varicella vaccine in 1996. Although the vaccine was developed by Japanese scientists, until recently, the vaccine has been administered on a voluntary basis in Japan resulting in a vaccine coverage rate of approximately 40%. Therefore, Japan initiated universal immunization using the Oka strain varicella vaccine in November 2014. Given the transition from voluntary to universal immunization in Japan, it will also be important to monitor the epidemiology of varicella and herpes zoster. The efficacy and safety of co-administration of the varicella vaccine and measles, mumps, and rubella vaccine have been demonstrated in many countries; however, there was no data from Japan. In order to adopt the practice of universal immunization using the Oka strain varicella vaccine in Japan, data demonstrating the efficacy and safety of co-administration of varicella vaccine and measles and rubella (MR) vaccine were required. Additionally, we needed to elucidate the appropriate time interval between the first and second administrations of the vaccine. It is also important to differentiate between wild type and Oka vaccine type strains in herpes zoster patient with past history of varicella vaccine. Thus, there are many factors to consider regarding the adoption of universal immunization in Japan to control varicella zoster virus (VZV) infections.

Direct detection of human herpesvirus 6B by the LAMP method using newly developed dry-reagents☆

The reliability of the HHV-6B LAMP using the dry-reagent method was evaluated using serum samples obtained from febrile children. The sensitivity of the original and dry-reagent methods was 10 copies/reaction and 100 copies/reaction, respectively. The dry-reagent LAMP method was highly sensitive (94.0%) and specific (96.0%) for the detection of HHV-6B.

Clinical utility of loop-mediated isothermal amplification assay for the diagnosis of common alpha herpesvirus skin infections.

Loop‐mediated isothermal amplification (LAMP) is a nucleic acid amplification method with a high specificity, efficiency and speed. No reports exist regarding the usefulness of LAMP for clinically suspected skin infections caused by herpes simplex virus (HSV) or varicella zoster virus (VZV). The aim of this study was to evaluate the clinical usefulness of LAMP in the diagnosis of common cutaneous alpha herpesvirus (HSV type 1 and 2, and VZV) infections. LAMP and real‐time polymerase chain reaction (PCR) were performed using swab samples collected from 106 patients with clinically suspected alpha herpesvirus skin infections. The results of LAMP performed with DNA extraction did not differ from those performed without DNA extraction. The sensitivity of LAMP tested against real‐time PCR was 96% in herpes simplex, 78% in eczema herpeticum, 93% in herpes zoster and 100% in varicella. No viral DNA was detected by LAMP in all negative real‐time PCR samples. Viral DNA load was significantly lower in samples with false‐negative LAMP results than in the LAMP‐positive samples. LAMP enables confirmation of clinically suspected cutaneous HSV and VZV infections. However, the sensitivity of LAMP is lower than real‐time PCR. The accuracy of LAMP may increase if sufficient viral DNA is obtained from lesions. LAMP performed without DNA extraction remains sensitive; thus, LAMP represents a quick and economical method for the diagnosis of common alpha herpesvirus skin infections.