Tetsu Mukai

Human herpesvirus 7: Another causal agent for roseola (exanthem subitum)

Human herpesvirus 7 (HHV-7) was isolated from peripheral blood mononuclear cells of two infants with typical exanthem subitum. The HindIII-, BamHI-, and EcoRI-digested DNA patterns of the isolated viruses were very similar to that of the prototype HHV-7 (RK strain), but different from that of human herpesvirus 6 (HHV-6). During the convalescent period of the first patient, the titer of antibody to HHV-7 rose from < 1:10 to 1:320 by an immunofluorescence antibody test, whereas the titer of antibody to HHV-6 remained < 1:10. In the second patient, who had two independent episodes of exanthem subitum during 2 months, both HHV-6 and HHV-7 were sequentially isolated; seroconversion to HHV-6 occurred during the first episode and to HHV-7 during the second episode. In addition, sera from another 15 children who had episodes of exanthem subitum were serologically tested for antibodies to HHV-6 and HHV-7 by immunofluorescence antibody test. Five of seven patients had seroconversion to HHV-7 just after having typical signs and symptoms of exanthem subitum. These results suggest that HHV-7 is one of the causative agents of exanthem subitum.

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.

Human herpesvirus 6 and human herpesvirus 7 infections in renal transplant recipients and healthy adults in Turkey

SummaryWe explored the prevalence of human herpesvirus 6 (HHV-6) and human herpesvirus 7 (HHV-7) infections in 16 renal transplant recipients and 16 healthy controls by virus isolation, serology, polymerase chain reaction (PCR) followed by dot blot hybridization. HHV-6 variant A was isolated from one renal transplant recipient. Seven patients (44%) and six controls (38%) had HHV-6 variant B DNA in their peripheral blood mononuclear cells. The prevalence of HHV-7 DNA was found to be the same in patients and controls (19%).

Identification of an Immunomodulating Agent from Mycobacterium leprae

A search for an immunomodulating agent from mycobacteria was carried out using Mycobacterium leprae. The antigenicity of each fraction of the bacterial membrane, which contains the most antigenic components of M. leprae, was assessed by using sera from paucibacillary leprosy. N-terminal sequencing of the serum-reactive protein and functional assessment of the membrane fractions using monocyte-derived dendritic cells (DCs) identified major membrane protein II (MMP-II) as one of the efficient T-cell-activating candidates. Purified MMP-II stimulated DCs from healthy individuals to produce interleukin-12 p70 and up-regulated the surface expression of major histocompatibility complex class I and II, CD86, and CD83 molecules. Also, there was an increase in the percentage of CD83(+) cells in the DC population. Furthermore, MMP-II-pulsed DCs expressed their derivatives on their surfaces. Using Toll-like receptor 2 (TLR-2)-dependent receptor constructs, we found that TLR-2 signaling was involved in DC maturation induced by MMP-II. Taken together, MMP-II can be recognized as an immunomodulating protein in terms of activation of antigen-presenting cells and innate immunity.

Isolation of human herpesvirus 7 from a child with symptoms mimicking chronic Epstein‐Barr virus infection

Summary. Human herpesvirus‐7 (HHV‐7), which is a newly identified human herpesvirus with an unknown pathologic role, was isolated from a 5‐year‐old boy suffering from fever, hepatosplenomegaly and pancytopenia. Although the clinical course was similar to that of chronic active Epstein‐Barr virus infection, no viruses other than HHV‐7 were isolated. This finding raises the possibility that HHV‐7 played a pathogenic role in the present patient.

Outbreak of exanthem subitum in an orphanage.

Human herpesvirus type 6 is now recognized as the causal agent of exanthem subitum (roseola infantum). Roseola does not have the contagious characteristics of measles, rubella, and chickenpox; outbreaks thus occur infrequently. Recently, we observed an outbreak of exanthem subitum in an orphanage. We report a comparison of the DNA profiles of the strains of HHV-6 isolated and discuss the possibility of horizontal infection.

Experimental infection of cynomolgus and African green monkeys with human herpesvirus 6

Cynomolgus and African green monkeys were inoculated with human herpesvirus 6 (HHV-6). An antibody response was first observed 10 days and 5 days after inoculation of cynomolgus monkeys and African green monkeys, respectively, and was detectable for the duration of the experiment (33 days). HHV-6 DNA was first detected by the polymerase chain reaction in mononuclear cells of one cynomolgus monkey and one African green monkey 10 days after virus inoculation, and in a total of three of four cynomolgus monkeys (75%) and four of five African green monkeys (80%) later after inoculation. Furthermore, HHV-6 DNA was detected in the lymph nodes and spleen of monkeys killed 33 days after virus inoculation. A rash was observed on the trunk of one African green monkey 13 days after virus inoculation, otherwise the infection was asymptomatic. When mononuclear cells from both groups of monkeys were cultured in medium containing concanavalin A and interleukin 2, and infected with HHV-6 in vitro, virus replication was observed. The data suggest that HHV-6 infects these species of monkey and that this system could be useful as an animal model of HHV-6 infection.

Identification and Characterization of the Genes Involved in Glycosylation Pathways of Mycobacterial Glycopeptidolipid Biosynthesis

Glycopeptidolipids (GPLs) are major components present on the outer layers of the cell walls of several nontuberculous mycobacteria. GPLs are antigenic molecules and have variant oligosaccharides in mycobacteria such as Mycobacterium avium. In this study, we identified four genes (gtf1, gtf2, gtf3, and gtf4) in the genome of Mycobacterium smegmatis. These genes were independently inactivated by homologous recombination in M. smegmatis, and the structures of GPLs from each gene disruptant were analyzed. Thin-layer chromatography, gas chromatography-mass spectrometry, and matrix-assisted laser desorption ionization-time-of-flight mass spectrometry analyses revealed that the mutants Deltagtf1 and Deltagtf2 accumulated the fatty acyl-tetrapeptide core having O-methyl-rhamnose and 6-deoxy-talose as sugar residues, respectively. The mutant Deltagtf4 possessed the same GPLs as the wild type, whereas the mutant Deltagtf3 lacked two minor GPLs, consisting of 3-O-methyl-rhamnose attached to O-methyl-rhamnose of the fatty acyl-tetrapeptide core. These results indicate that the gtf1 and gtf2 genes are responsible for the early glycosylation steps of GPL biosynthesis and the gtf3 gene is involved in transferring a rhamnose residue not to 6-deoxy-talose but to an O-methyl-rhamnose residue. Moreover, a complementation experiment showed that M. avium gtfA and gtfB, which are deduced glycosyltransferase genes of GPL biosynthesis, restore complete GPL production in the mutants Deltagtf1 and Deltagtf2, respectively. Our findings propose that both M. smegmatis and M. avium have the common glycosylation pathway in the early steps of GPL biosynthesis but differ at the later stages.

Impaired maturation and function of dendritic cells by mycobacteria through IL-1β

Dendritic cells (DC) are pivotal for initiation and regulation of innate and adaptive immune responses evoked by vaccination and natural infection. After infection, mycobacterial pathogens first encounter monocytes, which produce pro‐inflammatory cytokines, including IL‐1β, TNF‐α and IL‐6. The role of these cytokines in DC maturation remains incompletely understood. Here, we show that maturation of DC from monocytes was impaired by pretreatment of monocytes with low doses of IL‐1β. Under these conditions, Mycobacterium leprae‐infected DC failed to stimulate antigen‐specific T cell responses. Expression of CD86 and CD83 and production of IL‐12 in response to lipopolysaccharide and peptidoglycan were diminished. In contrast, these DC functions were not impaired by pretreatment with TNF‐α, IL‐6 or IL‐10. When monocytes were infected with M. bovis Bacillus Calmette‐Guérin, and subsequently differentiated to DC, the activity of these DC was suppressed as well. Thus, IL‐1β acts at early stages of differentiation of DC and impairs biological functions of DC at later stages. Therefore, production of IL‐1β by mycobacteria‐infected antigen‐presenting cells counteracts effective stimulation of innate and adaptive immune responses.

Interferon and natural killer cell activity in patients with exanthem subitum.

Early immune response was studied by assessing interferon (IFN) and natural killer cell activity in 13 patients with exanthem subitum asociated with human herpesvirus 6 infection during the acute and convalescent phases. Only IFN-alpha showed a significant increase in the plasma of patients during the acute febrile phase compared with the convalescent period. The inhibitory effect of IFN-alpha and IFN-beta on human herpesvirus 6 replication was demonstrated in vitro with cord blood mononuclear cells. Natural killer cell activity was also significantly augmented in the acute phase, especially in the exanthem period, rather than in the convalescent phase (P < 0.01). These results suggest that the enhanced IFN-alpha response and natural killer cell activity in the acute early phase of the disease may play pivotal roles in the recovery from exanthem subitum.