Even distribution of BK polyomavirus subtypes and subgroups in the Japanese Archipelago.

BK polyomavirus (BKV) is ubiquitous among humans, infecting children asymptomatically and then persisting in renal tissue. BKV has four subtypes (I-IV) that can be identified by serological and genotyping methods. Subtypes I and IV are most prevalent in all countries examined to date. Based on nucleotide sequence variation, subtype I is further classified into four subgroups (Ia, Ib-1, Ib-2 and Ic), each of which have a close relationship to a particular human population. To clarify the relationships between BKV and human populations, we investigated the distribution patterns of BKV subtypes and subgroups in the modern Japanese population, which was formed from two distinct ethnic groups. Urine samples were collected from immunocompetent elderly patients in six regions along the Japanese Archipelago. The 287-bp VP1 region of the viral genome from these samples was amplified using the polymerase chain reaction. The amplified VP1 regions were sequenced and a neighbor-joining phylogenetic tree was reconstructed to classify the BKV isolates. We observed a similar pattern of subtype distribution throughout the Japanese Archipelago, with subtype I always detected at high rates (67-75%), followed by subtype IV (19-31%), with rare or no detection of subtypes II and III. Based on phylogenetic and single nucleotide polymorphism analyses, the subtype I isolates were divided into subgroups; the percentage of the Ic subgroup was high in all geographic regions (88-100%). These results suggest that BKV subtypes and subgroups are evenly distributed in the Japanese Archipelago. We discuss the implications of these findings for the relationships between BKV and human populations.

Subtype IV of the BK polyomavirus is prevalent in East Asia.

BK polyomavirus (BKV) is ubiquitous in human populations, infecting children asymptomatically and then persisting in the kidney. Using either serological or genotyping methods, BKV isolates have been classified into four subtypes (I-IV), with subtype I mainly detected in all countries studied so far. To elucidate the subtype of BKV prevalent in East Asia, we examined BKV-positive urine samples collected from immunocompetent elderly patients in Mongolia, Northeast China, Northwest China, Southeast China, Southwest China, Vietnam and Japan. The 287-bp typing region of the viral genome in each of these samples was PCR-amplified and sequenced, and a phylogenetic tree was constructed. According to the tree, BKV isolates in East Asia were unambiguously classified into subtype I or IV (subtypes II and III were not detected). In Japan, subtype I was mainly detected and subtype IV was rare, whereas in the other regions subtype IV was detected frequently, at rates ranging from 24 to 100%. Thus, East Asia (excluding Japan) is a region in which subtype-IV BKV is prevalent, a finding that requires the view of the geographic distribution of BKV subtypes to be revised. Furthermore, we present evidence that the immunological states of urine donors do not affect the pattern of BKV subtypes.

Flow cytometric detection of cell-associated interleukin-8 in alveolar macrophages in vivo from patients with hypersensitivity pneumonitis and sarcoidosis.

In comparison with neutrophil-mediated lung diseases, such as acute respiratory distress syndrome, the involvement of IL-8 in lymphocyte-mediated lung diseases has not been fully investigated. Several reports have shown a slight increase in bronchoalveolar lavage fluid (BALF) IL-8 in patients with hypersensitivity pneumonitis (HP) and sarcoidosis (SAR), but the source of the IL-8 has not been clarified. In the present study, the in vivo production of IL-8 by alveolar macrophages (AMs) is examined in these patients by analyzing the cell-associated IL-8, using the flow cytometric method adopted previously. The IL-8 levels in the epithelial lining fluid (ELF) were also assessed. Initially, slight, but significant, increased levels of ELF IL-8 in HP and SAR were confirmed. Using flow cytometric analysis, a significant increase was found in the cell-associated IL-8 of the freshly isolated AMs in HP, but not in SAR, indicating in vivo production of IL-8 by AMs in HP. The cell-associated IL-8 of the AMs cultured with or without lipopolysaccharide was also analyzed. However, in contrast to previous findings in patients with idiopathic pulmonary fibrosis, no differences were found between SAR and HP patients and control subjects. Based on these findings, it is speculated that ELF IL-8 levels are slightly increased in HP and SAR, and they may contribute to the accumulation of neutrophils and possibly lymphocytes. However, the source of IL-8 may be different and AMs are the candidate source of IL-8 in HP, but not in SAR. The flow cytometric method may be useful in assessing cytokines production by AMs.

Detection of identical JC virus DNA sequences in both human kidneys.

We studied JC virus (JCV) DNA sequence diversity among kidneys derived from cadavers with various causes of death. The 610-bp JCV DNA sequences we evaluated were identical not only among specimens derived from the same kidney but also among those derived from both kidneys of the same cadaver. Because the left and right kidneys are anatomically independent, our findings suggest that the viremia that has been proposed to occur after primary infection distributes the same JCV strain to both kidneys.

Unambiguous identification of JC polyomavirus strains transmitted from parents to children.

JC polyomavirus (JCV), the etiological agent of progressive multifocal leukoencephalopathy, is ubiquitous in humans, infecting children asymptomatically, then persisting in renal tissue. It has been proposed that JCV is transmitted mainly from parents to children through long-term cohabitation. The objective of this study was to further elucidate the mode of JCV transmission. In 5 families, we selected parent/child pairs between whom JCV was probably transmitted (judged on the basis of the identity of a 610-bp JCV DNA sequence between the parent and child). We established 5 to 9 complete JCV DNA clones from the urine of each parent or child. The complete sequences of these clones were determined and compared in each family. Nucleotide substitutions were detected in 4 parents and 1 child, and sequence rearrangements (deletions or duplications) were found in 2 parents and 2 children. Phylogenetic comparison of the detected sequences indicated that the diversity of JCV DNA sequences was generated in each family (i.e. not caused by multiple infection). We found that in 4 of the 5 families, a sequence detected in the parent was completely identical to one in the child. These findings provided further support for the proposed mode of JCV transmission, i.e. parent-to-child transmission during cohabitation.

Comparison of PCR-amplified JC virus control region sequences from multiple brain regions in PML.

The authors report a 14-year-old boy with Wiskott-Aldrich syndrome complicated by progressive multifocal leukoencephalopathy after allogeneic bone marrow transplantation. Several therapeutic approaches were attempted, but there was no response. The patient died 2 months after the onset of neurologic symptoms. We detected three distinct, rearranged regions of JC virus in the cerebellum, occipital lobe, and brainstem. These findings suggest that the brain lesions had three independent origins.

JC virus genotyping offers a new means of tracing the origins of unidentified cadavers.

There has been no reliable means of tracing the origins of unidentified cadavers but the recent finding that JC virus (JCV) can serve as a means of elucidating human migrations suggested that this virus may also be useful to trace the origins of unidentified cadavers. DNA samples extracted from renal tissue and urine were used as the template for PCR amplification of a 610 bp region (IG region) of the viral genome. We detected JCV DNA in 45% of the renal samples and in 33% of the urine samples and was detectable even 10 days after death. The sequences of the amplified IG regions could be used to determine the genotypes. We conclude that the JC virus genotype is a new marker useful for tracing the origins of unidentified cadavers.

Asian domains of four major genotypes of JC virus, Af2, B1-b, CY and SC.

JC virus (JCV) strains worldwide can be classified into various genotypes based on DNA sequence variations. To define the domains of the four major JCV genotypes in Asia, we collected urine samples at six unstudied sites: three in southeastern Asia, two in the central highlands and one in central Asia. DNA was extracted from urine samples, and used to amplify a 610-bp region of the viral genome. For each geographical site, we determined 16 to 31 sequences, from which a phylogenetic tree was constructed to unambiguously classify detected JCV isolates into distinct genotypes. From JCV genotype profiles at the sites studied here and elsewhere, the following conclusions were drawn. Although Af2 is the major genotype in Africa, this genotype also occurs in western and central Asia. B1-b mainly occurs in western and central Asia, including the central highlands. CY occurs in northeastern Asia with the southern boundary between China and southeast Asian countries. Although SC predominates in southeastern Asia, it also occurs in northern and central Asia at lower frequencies. In addition, a few minor JCV genotypes (B1-a, B2 and B3) occur at many sites. We discuss here the anthropological and medical significance of the present findings.

Phylogenetic analysis of dengue-3 viruses prevalent in Guatemala during 1996-1998.

Dengue is an acute viral disease transmitted by the Aedes aegypti mosquito which is present in most tropical urban areas of the world. There are four antigenically distinct serotypes, designated dengue-1 (DEN-1), dengue-2 (DEN-2), dengue-3 (DEN-3) and dengue-4 virus (DEN-4). In this study, we determined the serotypes of dengue viruses isolated in Guatemala in 1995-1998, and found that DEN-3 viruses appeared in 1995 and became predominant in the following three years. We then sequenced cDNAs from fifteen DEN-3 isolates recovered during 1996-1998. From the nucleic acid sequences and previously determined DEN-3 sequences, a phylogenetic tree was constructed using the neighbor joining method. The tree indicated that all fifteen isolates and other DEN-3 viruses isolated in Sri Lanka, India, Samoa and Mozambique formed subtype III. More than two decades ago, DEN-3 virus was prevalent in the Caribbean, but the isolates obtained at that time belonged to subtype IV. Therefore, we concluded that the 1996-1998 dengue epidemic in Guatemala was caused by DEN-3 strains, imported from a tropical area of Asia or Africa or from a Pacific island.

JC virus regulatory region rearrangements in the brain of a long surviving patient with progressive multifocal leukoencephalopathy.

JC virus (JCV) infection of oligodendrocytes causes demyelination in brains of patients with with progressive multifocal leukoencephalopathy (PML). Expansion of demyelination throughout the brain is not fully understood. The opportunity was taken to investigate the postmortem brain of a long surviving patient with PML for whom diagnosis was made 4 years before death based on pathological and virological findings of a brain biopsy. Four distinct regulatory sequences in the JCV genome were detected (designated as JW-1 to 4) from various regions of the necropsied brain. All regulatory sequences were rearranged forms that could be produced from the archetype by deletions and duplications. JW-1 and 2 shared some structural features not present in JW-3 and 4 and vice versa. JW-1 was distributed throughout the brain, whereas JW-2, 3, and 4 were restricted to only part of the brain. JW-1 and 2 had been detected in the initial brain biopsy 4 years earlier. These findings suggested that brain lesions in advanced stages were generated not only by expansion of the original variant (JW-1) of JCV but also by delayed growth of two other variants (JW-3 and 4).

[Chronological changes of lacunar infarctions on fluid-attenuated inversion recovery magnetic resonance images].

In 26 patients with lacunar syndromes, emergence of new lacunar infarctions were identified within 13 days from onset by diffusion-weighted magnetic resonance images. The identified lacunar infarctions were repeatedly imaged using fluid-attenuated inversion recovery (FLAIR) sequence up to 600 days from onset. On FLAIR images taken by 23 days from onset, lacunar infarctions showed homogeneous hyperintensity. On the later FLAIR images beyond 25 days from onset they were observed as heterogeneously hyperintense lesions in half of the patients. In the other patients, lacunar infarctions were observed as hypointense areas with a hyperintense rim beyond 41 days from onset, which indicates cystic transformation with surrounding gliosis. These FLAIR images of lacunar infarction differ from those of dilated perivascular space which is observed as an area of simple hypointensity.

Lack of disease-specific amino acid changes in the viral proteins of JC virus isolates from the brain with progressive multifocal leukoencephalopathy.

Progressive multifocal leukoencephalopathy (PML) is a fatal demyelinating disease of the central nervous system caused by a ubiquitous human polyomavirus designated as JC virus (JCV). JCVs in the brain of PML patients (PML-type JCVs) contain various regulatory regions generated from the archetypal regulatory region during persistence in the patients. We determined the complete DNA sequences of 2 PML-type isolates and 5 archetype isolates. Amino acid sequences of individual viral proteins were deduced from complete DNA sequences, and were compared among 16 isolates (6 PML types and 10 archetypes). From the data obtained, we concluded that PML-associated amino acid changes did not occur in the viral proteins of PML-type JCVs.

[Lung cancer obscured by aspergillus hyphae].

A 66-year-old man was referred to our hospital because of cough. Chest X-ray films showed complete atelectasis of the left lung. Serum CYFRA was elevated. Bronchoscopic examination disclosed a white polypoid lesion occluding the left main bronchus. A biopsy specimen from the lesion revealed numerous aspergillus hyphae. Oral itraconazole (100 mg) and weekly endobronchial instillation of fluconazole were administered. Three months later, on the 12th bronchoscopic examination, the tumor occluding the left main bronchus was detected after the removal of aspergillus, and the biopsy findings yielded a diagnosis of squamous cell carcinoma. Aspergillosis complicated by lung cancer without cavity formation is very rare, and compounded the difficulty of diagnosing lung cancer in this case.

Judgments of emotion by nurses and students given double-bind information on a patient's tone of voice and message content.

We examined the communication process in a situation typical of the nursing setting by use of a double-bind communication. Our objective was to examine which of two cues in communications from a patient, tone of voice or verbal content, was more important in judging the speaker's emotional status and personality traits and in arousing the listening nurse's emotions. Subjects were 82 nurses who worked at the university hospital and 100 students who were studying at the Faculty of Nursing of the university. They were assigned into four groups at random, presented professionally tape-recorded scripts representing a patient's verbal report, and then completed a questionnaire concerning the speaker's emotional status as well as the listener's own emotional status. When the listeners judged the speaker's emotional status, they gave more attention to a negative emotional expression, and when the listeners formed an impression of the speaker's personality traits, they were influenced by the speaker's tone of voice rather than by the content of the speech.

[Acquired immunodeficiency syndrome-associated progressive multifocal leukoencephalopathy treated with highly active anti-retroviral therapy].

We reported a patient with acquired immunodeficiency syndrome (AIDS)-associated progressive multifocal leukoencephalopathy (AIDS-PML), whose condition improved after highly active anti-retroviral therapy (HAART). A 70-year-old man was admitted to our hospital because of worsening left hemiplegia and disturbance of consciousness. During the past 30 years, he frequently traveled to the United States and southeast Asia. On neurological examination, he was somnolent and left hemiplegia with severe rigospasticity was present. The deep tendon reflexes showed hyper-reflexes with extensor plantar responses. Laboratory studies showed pancytopenia and positive HIV-1 antibodies. The CD4 cell count was 38/mm3 and his HIV viral RNA load in the blood was 9,500 copies/ml. T2-weighted magnetic resonance imaging (MRI) of the brain revealed asymmetrical high intensity white matter lesions in the right fronto-parietal, and left frontal regions and in the cerebellar hemisphere. The cerebrospinal fluid (CSF) protein elevated to 91 mg/dl with a normal cell count. The diagnosis of PML was confirmed by the detection of JC virus DNA in the CSF using a nested polymerase chain reaction assay. Three weeks after starting HAART with zidovudine, lamivudine, and indinavir, he was able to respond to simple commands. Two months later, the HIV viral RNA load decreased to less than 400 copies/mm3, and no JC virus DNA was detected in the CSF, with an increase of the CD4 cell count to 285/mm3 in the blood. A follow-up MRI of the brain showed a reduction in the cerebellar and cerebral white matter lesions. The recovering immune function by decreasing of the HIV load after HAART might suppress JC virus replication. It was suggested that HAART would become a beneficial treatment for patients with AIDS-PML.

JC virus genotypes in a Taiwan aboriginal tribe (Bunun): implications for its population history.

The origin of Taiwanese aborigines remains obscure; it has been speculated that they may be from either mainland China or southeastern Asia. We used the JCV genotyping method to elucidate the origin of Bunun aborigines who now live in central mountain areas of Taiwan. We found that Bunun aborigines carried two major (B1-a2 and CY) and two minor JCV genotypes (B1-a1 and SC). This was contrasted with the JCV genotype profile in modern Taiwanese: one major (SC) and two minor genotypes (CY and B1-a1). It thus appears that B1-a2 and CY are indigenous to the Bunun tribe. B1-a2 was first identified in this study as a discrete cluster that contained only Bunun and Philippine JCV isolates and that was closely related to B1-a1, one of the three common JCV genotypes in China. CY predominates in North China, while SC predominates in South China and southeastern Asia. The present findings suggest that the Bunun tribe is an admixture of two ethnic groups, one carrying B1-a2 and the other carrying CY. In other words, it is likely that the Bunun tribe was established by two waves of immigrations from mainland Asia, predating those by southern Chinese which began in the 17th century.

Four geographically distinct genotypes of JC virus are prevalent in China and Mongolia: implications for the racial composition of modern China.

JC polyomavirus (JCV) is ubiquitous in humans, persisting in renal tissue and excreting progeny in urine. It has been shown that the genotyping of urinary JCV offers a novel means of tracing human migrations. This approach was used to elucidate the racial composition of modern China. JCV isolates in the Old World were previously classified into nine distinct genotypes. One of them (B1) has a wide domain, encompassing part of Europe and the entirety of Asia. By constructing a neighbour-joining phylogenetic tree, all B1 isolates detected so far were classified into four distinct groups (B1-a to -d), each occupying unique domains in the world. According to this revised classification system of JCV DNAs, four genotypes (CY, SC, B1-a and -b) were found to be prevalent in China and Mongolia (Mongolia was studied instead of Inner Mongolia, which is part of China). There was a remarkable variation in the incidence of genotypes among the sites of sample collection. CY was more frequently detected in Northern China, SC was predominant in Southern China and B1-b was detected only in Mongolia. B1-a was spread throughout China. These data were statistically analysed and the observed regional differences in the incidence of genotypes were found to be significant. It is likely that these differences in JCV distribution in China reflect the intermingling of different population groups that constitute modern China.

Archetype JC virus efficiently replicates in COS-7 cells, simian cells constitutively expressing simian virus 40 T antigen.

JC polyomavirus (JCV), the causative agent of progressive multifocal leukoencephalopathy (PML), is ubiquitous in humans, infecting children asymptomatically and then persisting in the kidney. Renal JCV is not latent but replicates to excrete progeny in the urine. The renal-urinary JCV DNAs carry the archetype regulatory region that generates various rearranged regulatory regions occurring in JCVs derived from the brains of PML patients. Tissue cultures that support the efficient growth of archetype JCV have not been reported. We studied whether archetype JCV could replicate in COS-7 cells, simian cells transformed with an origin-defective mutant of simian virus 40 (SV40). Efficient JCV replication, as detected by a hemagglutination assay, was observed in cultures transfected with five of the six archetype DNAs. The progeny JCVs could be passaged to fresh COS-7 cells. However, when the parental cells of COS-7 not expressing T antigen were transfected with archetype JCV DNAs, no viral replication was detected, indicating that SV40 T antigen is essential for the growth of JCV in COS-7 cells. The archetype regulatory region was conserved during viral growth in COS-7 cells, although a small proportion of JCV DNAs underwent rearrangements outside the regulatory region. We then attempted to recover archetype JCV from urine by viral culture in COS-7 cells. Efficient JCV production was observed in COS-7 cells infected with five of the six JCV-positive urine samples examined. Thus, COS-7 cells should be of use not only for the production of archetype JCV on a large scale but also for the isolation of archetype JCV from urine.