LETTERS to the EDITOR

Human herpesvirus-6 associated with fatal haemophagocytic syndrome

Acute infection with HHV-6 and HHV-7 and its reactivation may cause various diseases that deserve medical attention. Pathologic changes show features of a viral infection without specific cytopathic effects for HHV-6A, HHV-6B, and HHV-7. Intranuclear inclusions (preferentially Cowdry type B) and giant cells are seen rarely in tissues, and in all such cases coincident viral infections must be excluded. Dual infections have been reported for HHV-6, HHV-7, EBV, CMV, HSV-2, HIV, parvovirus (heart), and measles (lung), and suggestively for other viruses such as papillomavirus, coxsackievirus (heart), and JC virus (brain). Most patients were immunodeficient (transplant or AIDS patients, tumor carriers, children with immune deficiency syndromes) or suffered from autoimmune disorders (collagen vascular diseases, drug- associated hypersensitivity syndromes). To unequivocally relate pathologic lesions to a given virus infection, meticulous microscopic, immunohistologic, and molecular diagnostics should be combined with serology and viral culture. Identification of HHV-6 or HHV-7 as the specific pathogen in a given case is usually tentative, but effective treatment offers improvement and recovery.

We reported a girl with HHV-6 infection associated with both acute encephalopathy with biphasic seizures and late reduced diffusion, and hemophagocytic syndrome. She had a prolonged convulsion after a one-day history of febrile illness. Cerebrospinal fluid or brain CT showed no abnormalities on admission and her consciousness was recovered on the next day. However, a prolonged seizure and deterioration of consciousness appeared on the sixth day of illness. Diffusion-weighted images revealed marked reduction of water diffusion in the bilateral frontal areas. HHV-6 infection was virologically proven by polymerase chain reaction. She was treated with γ-globulin, steroid pulse therapy, and brain hypothermia. In addition, decrease in white blood cells and platelet counts, and elevation of liver enzymes and ferritin were noted on the fourth day of illness. Hemophagocytic macrophages were revealed by bone marrow aspiration on the sixth day. Her hematological and blood chemistry abnormalities recovered gradually after steroid pulse therapy. An elevation of interleukin-6, -8, and -10, and tumor necrosis factor in the serum and that of interleukin-4, -6, and-8 in the cerebrospinal fluid were observed at the onset of a late seizure. These facts suggested that hypercytokinemia will be related to the pathogenesis of acute encephalopathy of our patient.

A more rapid and easier method is needed for monitoring human herpesvirus 6 (HHV-6) infections. The loop-mediated isothermal amplification method (LAMP) can detect viral DNA with high specificity, efficiency, and speed under isothermal conditions. LAMP requires only simple equipment that is available in hospital laboratories.

We evaluated LAMP as a means of detecting HHV-6 DNA directly from patients’ sera.

The sensitivity of the HHV-6 LAMP protocol without heat denaturation was 1000 copies/tube; with heat denaturation 10 copies/tube were detected. Three hundred serum samples from children with fever were analyzed. Using HHV-6 isolation as a definition of HHV-6 infection, the sensitivity, specificity, positive predictive value, and negative predictive value of the HHV-6 LAMP method without DNA extraction were 95.5%, 95.2%, 94.0%, and 96.4%, respectively.

Direct detection of HHV-6 DNA in serum with a modified HHV-6 LAMP could be used for rapid diagnosis of exanthem subitum (ES).

Primary cytomegalovirus (CMV) infection in healthy children is associated with hepatomegaly and mild elevations of hepatic transaminase. Human herpesvirus-6 (HHV-6) and CMV are highly related pathogens, and like CMV, HHV-6 is associated with a variety of clinical conditions in immunocompromised patients, such as transplant recipients. One report examining hematopoietic stem cell transplant (HSCT) recipients suggests an association between HHV-6 infection and bloody diarrhea. Moreover, CMV infection causes hepatitis and gastrointestinal ulceration in immunocompromised patients. Some patients experience a mild elevation in hepatic transaminase levels during infection. Approximately 4% (four cases) of 89 infants with primary HHV-6 infection had signs of hepatic injury during viral infection. However, one case report described chronic hepatitis caused by HHV-6 infection in one 20-month old, otherwise healthy, boy. Chronic hepatitis in young children can be caused by a variety of viruses, such as hepatitis B virus, hepatitis C virus, and CMV. In addition to these viruses, HHV-6 appears to be an etiologic agent for chronic hepatitis in young children. Moreover, HHV-6 DNA was detected in the endothelium of the portal vein but not in hepatocytes by in situ hybridization. Two additional cases of neonatal HHV-6 infection-associated fulminant hepatitis were not as clinically severe. HHV-6 may be associated with Gianotii–Crosti syndrome, a popular acrodermatitis of childhood causing mild elevations of hepatic transaminase, and hepatitis B virus plays an important role in the pathogenesis of this disease.

Two variants (A and B) of human herpesvirus 6 (HHV-6) can be isolated from humans, with each variant having unique biological properties. HHV-6 variant typing is mainly done following amplification of viral genomic DNA followed by restriction endonuclease digestion. Our objective was to generate a monoclonal antibody (mAb) that would allow us to discriminate between variants A and B of HHV-6. BALB/c mice were immunized with a recombinant glutathione-S-transferase protein fused to the immediate–early (IE) 2 protein from HHV-6 variant A. Following splenocytes fusion, one IgG1 κ light chain mAb (P6H8) was isolated and found to react specifically with variant A IE2 protein in immunofluorescence and western blot assays. The P6H8 antibody represents a useful tool for both fundamental research and clinical applications allowing for the discrimination of infections caused by HHV-6 variants A or B.

Immediate-early (IE) proteins are the first proteins expressed following viral entry and play a crucial role in the initiation of infection. We report the cloning and characterization of a full-length IE1 transcript and protein (IE1B) from human herpesvirus 6 (HHV-6) variant B. The IE1B transcript consists of five exons (3720 nucleotides), three of which are coding for the IE1 protein. The 1078-amino acid-long IE1B protein is 62% identical and 75% similar to the 941-amino acid IE1 from HHV-6 variant A. IE1B protein can be detected at 4 h post-infection (P.I.), and it is distributed as small intranuclear structures. The maximal number of IE1 bodies (∼10–12/nucleus) is detected at 12 h P.I. after which the IE1 bodies condense into 1–3 larger entities by 24–48 h P.I. During infection the IE1B protein is phosphorylated on serine and threonine residues. IE1B undergoes further post-translational modification with its conjugation to the small ubiquitin-like modifier (SUMO-1) peptide. IE1B colocalizes with SUMO-1 and promyelocytic leukemia nuclear bodies during infection as well as in transfection experiments. Finally, IE1 from variant B is a weaker transactivator than IE1 from variant A, when assayed using heterologous promoters. Overall, the characterization of the HHV-6 IE1B protein presented highlights the similarity and divergence between IE1 from both variants and provides useful information pertaining to the early phase of infection.