Hiroko Miyoshi

Seroepidemiological study of human herpesvirus-6 and -7 in children of different ages and detection of these two viruses in throat swabs by polymerase chain reaction

The presence of human herpesvirus 6 (HHV‐6) and human herpesvirus 7 (HHV‐7) in throat swabs of 62 children of different age groups (group I, ages 0–5 month; group II, ages 6–11 months, group III, ages 12–23 months, group IV, age 2–8 years) and 28 adults was detected by polymerase chain reaction. The detection rate of HHV‐6 DNA was the highest (87%) in children aged 1‐year‐old and decreased with age, whereas the detection rate of HHV‐7 increased with age and reached a maximum in adults. HHV‐6B was detected in almost all samples except for two children who secreted only HHV‐6A. When the antibody prevalence was determined in the four groups of children, HHV‐6 antibody was detected in 8/12 (66.7%), 10/12 (83.3%), 15/16 (93.8%), and 13/14 (92.9%), respectively. Antibody to HHV‐7 in these groups was detected in 6/12 (50.0%), 4/12 (33.3%), 12/16 (75.0%), and 13/14 (92.9%), respectively. Detection of HHV‐6 DNA in throat swabs of triplets who had the sequential onset of exanthem subitum was attempted by using samples sequentially collected from these children after the onset of the disease in the first patient. HHV‐6 DNA with high copy numbers was detectable during the acute and convalescent phases of the disease in all patients, but no DNA was detected in samples collected before the onset of disease.

Thrombotic microangiopathy associated with reactivation of human herpesvirus-6 following high-dose chemotherapy with autologous bone marrow transplantation in young children

Thrombotic microangiopathy (TMA) is a serious complication of BMT. Several factors are important in the etiology of TMA, such as cyclosporin A, GVHD, irradiation, intensive conditioning chemotherapy and infection, which cause damage to vascular endothelial cells leading to activation of these cells. We describe two young children with TMA following high-dose chemotherapy with autologous BMT. Development of TMA was accompanied by reactivation of HHV-6, which was identified by both an increase in the copy number of HHV-6 DNA in the peripheral blood and a significant increase in antibody titers to HHV-6. Thus, it was suggested that reactivation of HHV-6 together with high-dose chemotherapy played an important role in the pathogenesis of TMA in these patients. Since HHV-6 is known to infect vascular endothelial cells, and CMV which is virologically closely related to HHV-6, has been reported to be a pathogen that causes TMA, infection with HHV-6 of vascular endothelial cells may induce TMA via damage and activation of these cells.

Reactivation of human herpesvirus 6 by infection of human herpesvirus 7.

We have attempted to reactivate human herpesvirus 6 (HHV‐6) by infection with HHV‐7 using childhood exanthem subitum patients in vitro. Peripheral blood mononuclear cells (PBMCs) were collected from children who had a history of exanthem subitum(ES) by HHV‐6 and were infected by human herpesvirus 7 (HHV‐7) in vitro. The antigen positive rate to HHV‐6 started to increase 7 days after the infection and reached a maximum by Day 15 using an immunofluorescence antibody test. The copy number of HHV‐6 DNA also increased in the samples in 10 days after infection in vitro. No antigen or increase in DNA was detected in PBMCs, that were mock‐infected or infected with supernatant of stock virus after ultracentrifugation, suggesting that an infection by HHV‐7 is necessary to reactivate HHV‐6. In the paired sera samples during the acute and the convalescent phases of ES, seven to ten bands, that were specific for HHV‐6, were recognized in samples from the acute phase, and at least 5 dominant polypeptides were found more intensively after HHV‐7 infection. J. Med. Virol. 60:284–289, 2000.

Detection of human herpesvirus 7 (HHV‐7) DNA in breast milk by polymerase chain reaction and prevalence of HHV‐7 antibody in breast‐fed and bottle‐fed children

Twenty‐nine breast milk mononuclear cell samples were analyzed for human herpesvirus 7 (HHV‐7) DNA, human herpesvirus 6 (HHV‐6) DNA, and human cytomegalovirus (HCMV) DNA by polymerase chain reaction (PCR). In addition, peripheral blood mononuclear cell samples from 13 puerperants were analyzed for HHV‐7 DNA by PCR, and seropositivity of HHV‐7 was also analyzed in breast‐fed and bottle‐fed children. HHV‐7 DNA was detected in 3 of 29 breast milk samples. HCMV DNA was also detected in 3 of 29 breast milk samples, but HHV‐6 DNA was not detected. HHV‐7 DNA was detected in 11 of 13 samples of peripheral blood mononuclear cells. Though the seropositivity rate for HHV‐7 in breast‐fed children was slightly higher than that in bottle‐fed children at 18 and 24 months old, the difference was not statistically significant. From these results, we speculate that breast‐feeding may be one of the transmission routes of HHV‐7, although this is not the main route. J. Med. Virol. 56:275–279, 1998.

Human Herpesvirus 6B Infection of the Large Intestine of Patients with Diarrhea

Four patients had severe diarrhea after undergoing stem cell transplantation. Human herpesvirus 6B (HHV-6B) DNA was detected in large intestine tissue specimens and in peripheral blood mononuclear cells. In situ hybridization was positive for HHV-6B DNA in the nuclei of goblet cells and, sometimes, in the histiocytes in the submucous region of the large intestine, which suggests that HHV-6B may infect and reactivate in these cells.

Monitoring of Human Cytomegalovirus Infections in Pediatric Bone Marrow Transplant Recipients by Nucleic Acid Sequence-Based Amplification

In the diagnosis of human cytomegalovirus (HCMV) infection, it is very important to distinguish symptomatic from asymptomatic infection. The nucleic acid sequence-based amplification (NASBA) technique was compared with single and nested polymerase chainreaction (PCR) methods. For NASBA detection, the beta2.7 transcript was chosen as a target because of its abundant active HCMV-specific expression. Of 20 pediatric bone marrow transplant (BMT) recipients, 8 developed HCMV-related clinical symptoms. The clinical sensitivities and specificities were 50% and 100% for single PCR, 100% and 67% for nested PCR, and 100% and 83% for NASBA, respectively. Follow-up of HCMV infections in pediatric BMT recipients showed that NASBA could both detect viral transcript prior to the onset of clinical symptoms and reflect clinical improvement due to antiviral therapy. These data suggest that NASBA should be useful for both predicting HCMV disease development and monitoring the effect of antiviral therapy.

Allogeneic peripheral stem cell transplantation using positively selected CD34 + cells from HLA-mismatched donors

We examined five children who underwent allogeneic peripheral stem cell transplantation (PSCT) using positively selected CD34+ cells from three or two loci-mismatched donors. CD34+ cells mobilized from peripheral blood were separated by immunomagnetic beads. CD34+ cells at 2.2–6.2 × 106/kg were transplanted into three patients with refractory leukemia, a patient with relapsed medulloblastoma and a patient with Fanconi’s anemia following a conditioning regimen which included irradiation, alkylating agents and antithymocyte globulin treatment. The number of infused CD3+ cells included in grafts was 2.3–22.7 × 104/kg. Four patients achieved engraftment and hematopoietic reconstitution (>5 × 108/l of neutrophils on day 10 or 11). Graft rejection was observed in the patient with Fanconi’s anemia, but a rapid engraftment was obtained after second PSCT. Although no prophylactic agents other than ATG (included in the conditioning regimen) were used, greater than grade I acute GVHD was not observed, but limited chronic GVHD was observed in two patients. The two patients with leukemia relapsed on days 103 and 210, respectively, and the patient with medulloblastoma died of disease on day 159. The patient with Fanconi’s anemia died of fungal infection. CMV and HHV-6 diseases developed in four and two patients, respectively. Thus, although SCT using positively selected peripheral CD34+ cells may be an alternative approach for overcoming graft rejection and GVHD from HLA- mismatched donors, persistent immune deficiency attributing to extremely low numbers of T cells in grafts can potentially lead to reactivation of herpes viruses.

Remission of progressive multifocal leukoencephalopathy following highly active antiretroviral therapy in a patient with HIV infection.

Progressive multifocal leukoencephalopathy (PML) is a demyelinating disease resulting from lytic infection of oligodendrocytes by the papovavirus JC (JCV). PML has also been recognized as an AIDS-defining illness. The incidence of PML has increased since 1987 and it occurs in up to 4% of patients with AIDS. To date, there is no treatment available for PML and it usually results in death within 3-6 months of diagnosis. However, there are some reports of remission of PML after antiretroviral therapy. We report a 12-year-old child with hemophilia B and developing AIDS with the onset of PML. With highly active antiretroviral therapy, PML subsided with an increase of CD4 count from 10 to 300/microl in spite of about 1.0 X 10(4) human immunodeficiency virus (HIV)-1-RNA copies. He has survived more than 1 year without specific therapy against JCV. Highly active antiretroviral therapy appears to have improved his prognosis in HIV-associated PML.

Inverse relationship between human herpesvirus-6 and -7 detection after allogeneic and autologous stem cell transplantation

Human herpesvirus-6 (HHV-6) and -7 were analyzed in 25 and 18 patients with allogeneic (allo) and autologous (auto) stem cell transplantation (SCT), respectively, by weekly examination of viral DNA in peripheral mononuclear cells using semiquantitative PCR and serologic tests up to 12 weeks after SCT. HHV-6 DNA was detected in 29.6% and 27.9% of samples after allo- and auto-SCT, respectively. The proportions of HHV-6-DNA-positive samples increased in week 3 and 4 after allo-SCT, and in week 1 to 3 after auto-SCT. The frequency of HHV-7 DNA detection, however, was higher after auto-SCT (24.7%) than allo-SCT (12.8%) (P < 0.01). antibody titer elevation was accompanied by hhv-6 infection in eight of 10 patients. that for hhv-7 was also frequently observed, but not associated with infection in the majority of patients. five of six patients with >102 copies of HHV-6 DNA (/105 cells) on two consecutive occasions were allo-SCT recipients and three showed clinical episodes. Conversely, three of five patients with continuous reactivation of HHV-7 were auto-SCT recipients. Thus, the frequencies of HHV-6 and -7 DNA detection showed an inverse relationship comparing allo- and auto-SCT, suggesting a different mechanism may regulate HHV-6 and -7 reactivation. Bone Marrow Transplantation (2001) 27, 1065–1070.

Human herpesvirus‐6 infection in neonates: Not protected by only humoral immunity

Background : Infants are usually protected from various viral infections, including human herpesvirus‐6 (HHV‐6) and human herpesvirus‐7 (HHV‐7) infections, during the early infantile period by antibodies transferred from their mothers. However, rare cases of exanthem subitum (ES) in neonates have been described in published reports.