Human herpesvirus 6‐A, 6‐B,and 7 in vitreous fluid samples

Human herpesvirus 6 and 7 (HHV‐6, HHV‐7) have been associated with several neurologic syndromes and have been detected in nervous tissue from healthy persons; however, only two cases of HHV‐6A have been reported to be associated with intraocular inflammatory disease. Vitreous fluid was tested from 101 patients, including 69 samples from patients with ocular inflammation including CMV retinitis, idiopathic retinitis, iritis, and vitritis, for HHV‐6A, HHV‐6B, and HHV‐7 DNA by PCR. HHV‐6A DNA (4,950 copies per ml) was detected in vitreous fluid from one patient with CMV retinitis, HHV‐6B DNA (10,140 copies per ml) was detected in vitreous fluid from one patient with idiopathic ocular inflammation in the absence of CMV DNA, and HHV‐7 was not detected in any of the vitreous samples. HHV‐6A, HHV‐6B, and HHV‐7 DNA are detectable in less than 2% of vitreous samples in patients with ocular inflammation. J. Med. Virol. 82:996–999, 2010. © 2010 Wiley‐Liss, Inc.     

Case of fatal encephalitis by HHV-6 variant A

A fatal case is reported of encephalitis in an 85-year-old man caused by the human herpesvirus 6 variant A. The virological diagnosis was based on the findings of the virus variant genomic sequences both in the cerebrospinal fluid and serum of the patient. Moreover, virus replication in nervous tissue was suggested by a viral load higher in the cerebrospinal fluid than in the peripheral blood. The association of a central nervous system infection with the A variant of human herpesvirus 6 is interesting because of the difficulty in establishing a pathological role for this virus strain. Epstein-Barr virus DNA was detected in the patient's cerebrospinal fluid in association with human herpesvirus 6 DNA. The presence of the Epstein-Barr virus genomic sequences in the cerebro-spinal fluid was considered to be unimportant clinically.     

Chromosomally integrated human herpesvirus 6 in the Japanese population

The objectives of the work are to elucidate the incidence and virological findings of chromosomally integrated human herpesvirus 6 (ciHHV‐6) in Japanese population and to analyze an association between ciHHV‐6 and the clinical manifestation of exanthema subitum (ES). Real‐time polymerase chain reaction was performed to determine HHV‐6 DNA loads in 2347 cord blood samples from healthy neonates (cohort A), febrile children less than 5 years old (cohort B), and hematopoietic cell transplant recipients (cohort C). CiHHV‐6 was confirmed by detection of high copy numbers of viral DNA in somatic cells. The integration site was determined by fluorescent in situ hybridization analysis. In the ciHHV‐6 subjects of cohorts A and B, HHV‐6 antibody titers were measured, the history of ES was obtained, and the incidence of ES was compared with non–ciHHV‐6 children without primary HHV‐6B infection in the cohort B. CiHHV‐6 was detected in 14 (0.60%) of the 2347 samples: A (6/1006, 0.60%), B (6/790, 0.76%), and C (2/551, 0.36%). The integration sites were on chromosome 22q in seven cases, Yp in two cases, and 17q and Xp in one case. No past history of ES was observed in 11 of the 12 subjects. Nine children with ciHHV‐6 underwent serological analysis and were found to be positive for HHV‐6 IgG antibodies. Incidence of ES was statistically higher in the control subjects than the ciHHV‐6 subjects (P = 0.0039). In Japan, the frequency of ciHHV‐6 was 0.60%. A high incidence of ciHHV‐6A, specifically in chromosome 22, is a characteristic finding among the Japanese. CiHHV‐6 may interfere with the clinical symptoms of primary HHV‐6B infection.     

Human herpesvirus 6 downregulates major histocompatibility complex class I in dendritic cells

The expression of major histocompatibility complex (MHC) class I, class II, CD1a, and CD 83 in dendritic cells (DCs) after infection with human herpesvirus 6 (HHV-6) was examined. Whereas there was no significant change in the expression of CD1a, CD83, and MHC class II in infected DCs, MHC class I expression was downregulated after infection with HHV-6 variant A but not HHV-6B. The expression of HHV-6 immediate-early or early genes was required for the downregulation of MHC class I. The de novo synthesis of MHC class I was greatly suppressed by infection with HHV-6A in DCs, while its rate of degradation was only slightly elevated. These results suggest that HHV-6A may escape from the host immune system in DCs by causing the downregulation of MHC class I synthesis.     

Human herpesvirus 6 variant a infects human term syncytiotrophoblasts in vitro and induces replication of human immunodeficiency virus type 1 in dually infected cells

Human herpesvirus 6 (HHV-6) and human immunodeficiency virus type 1 (HIV-1) may interact during transplacental transmission of HIV-1. The placental syncytiotrophoblast layer serves as the first line of defense of the fetus against viruses. Patterns of replication of HHV-6 variant A (HHV-6A) and HIV-1 were analyzed in singly and dually infected human term syncytiotrophoblast cells cultured in vitro. For this purpose, the GS strain of HHV-6A and the Ba-L and IIIB strains of HIV-1 were used. HHV-6A replication was restricted at the level of early gene products in singly infected syncytiotrophoblasts, whereas no viral protein expression was found in cells infected with HIV-1 alone. Coinfection of syncytiotrophoblast cells with HHV-6A and HIV-1 resulted in production of infectious HIV-1. In contrast, no enhancement of HHV-6A expression was observed in cell cultures infected with both viruses. Uninfected syncytiotrophoblast cells were found to express CXCR4 and CCR3 but not CD4 or CCR5 receptors. Infection of syncytiotrophoblasts with HHV-6A did not induce CD4 expression and had no influence on chemokine receptor expression. Activation of HIV-1 from latency in coinfected cells was mediated by the immediate-early (IE)-A and IE-B gene products of HHV-6A. Open reading frames U86 and U89 of the IE-A region were able to activate HIV-1 replication in a synergistic manner. The data suggest that in vivo double infection of syncytiotrophoblast cells with HHV-6A and HIV-1 could contribute to the transplacental transmission of HIV-1 but not HHV-6A.     

Clinical applications of two-color telomeric fluorescence in situ hybridization for prenatal diagnosis: Identification of chromosomal translocation in five families with recurrent miscarriages or a child with multiple congenital anomalies

Two-color fluorescence in situ hybridization (FISH) analysis using human chromosome arm-specific telomeric probes (telomeric probes) was used successfully to detect each derivative chromosome of a translocation carrier in five couples who requested a prenatal diagnosis in future pregnancies. Most of the human chromosome terminal bands are G-band-negative, and even FISH analysis using whole-chromosome painting (wcp) probes are often of insufficient complexity to detect subtle chromosomal changes. A complete set of human telomeric probes was developed to improve the sensitivity of diagnosis of microdeletions or other cryptic rearrangements in telomeric regions. Two-color telomeric FISH was the only possible method for precise prenatal diagnosis of one of the couples, because the carrier's chromosomal aberration was too subtle to be detected by wcp FISH or conventional methods. We have demonstrated that two-color telomeric FISH has the potential to be a powerful new tool in the detection of cryptic chromosomal rearrangements involving telomeric regions in prenatal diagnosis precisely and in time. Received: September 9, 1998 / Accepted: October 23, 1998     

Human herpesvirus 6 infection of human epidermal cell line: pathogenesis of skin manifestations

In order to elucidate the pathogenesis of variant B human herpesvirus 6 (HHV-6) infection in skin tissues, an A431 cell line was inoculated with variant B HHV-6. HHV-6 causes abortive infection in the A431 cells, because neither late antigen (OHV-3 antigen) nor progeny virus is produced. Maximum levels of HHV-6 antigen (IEA/ex3 antigen)-positive cells (36.4%) were observed 48 hr after viral infection. Cocultivation of HHV-6-infected cord blood mononuclear cells with A431 cells was necessary for the establishment of a sufficient level of viral infection. Cell-to-cell contact between the infected cord blood mononuclear cells and A431 cells was crucial for increasing infection efficiency. To determine the biological effect of HHV-6 infection, flow cytometric analysis was carried out in HHV-6- and mock-infected A431 cells. Although no alteration was observed in VCAM-1 and ELAM-1 expression, that of HLA-ABC, HLA-DR, and ICAM-1 was upregulated after infection with HHV-6.     

Comparative genomic hybridization as a tool in tumour cytogenetics

The quality of cytogenetic analysis of solid tumours has greatly improved in the past decade, but a number of technical difficulties remain which limit the characterization of solid tumour chromosomes by conventional cytogenetics alone. The identification of regions of chromosomal abnormality has been aided by the introduction of molecular cytogenetic techniques such as fluorescence in situ hybridization (FISH). Of these, a recently developed approach, comparative genomic hybridization (CGH), has had a particular impact on the cytogenetic analysis of solid tumours. It incorporates the sensitivity of in situ techniques and overcomes many of the drawbacks of conventional cytogenetic analysis. This review first outlines the CGH method, giving details for the preparation of DNA probes and target human metaphase chromosomes together with information on the in situ technique and data handling criteria used in our laboratory. It then presents an overview of some of the current applications of CGH, together with a discussion of future directions in the field. Copyright © 1999 John Wiley & Sons, Ltd.     

Detection of a human chromosomal translocation t(8;9) in a baby with multiple malformations using two-color fluorescence in situ hybridization

A human chromosomal translocation t(8;9) was detected using two-color fluorescence in situ hybridization with probes capable of staining the entire lengths of each of these chromosomes. The chromosome 8 probe was labeled with biotin and detected with Texas red, while the chromosome 9 probe was labeled with AAF and detected with FITC . In normal metaphase spreads, two metaphases from the proband, two red, one green and one part red and part green derivative chromosome were seen. The bicolor chromosome corresponded to translocation of a chromosome 8 segment to the distal part of the q region of one chromosome 9, as originally indicated by banding analysis. In interphase nuclei of the proband, four domains with bright fluorescence were recognized in many nuclei. Two were red, one was green, and the fourth had portions of both colors, indicating the presence of the translocation.     

Fine‐needle aspiration biopsy of prostate synovial sarcoma: A case report and review of the literature

We describe a case of synovial sarcoma originating from prostate gland. The diagnosis was confirmed by fluorescent in situ hybridization analysis (FISH) for SYT rearrangement on the cell block. Synovial sarcoma is a high grade soft tissue malignancy with exceedingly rare involvement of genitourinary tract. However this entity should be considered in the differential diagnosis when dealing with aspiration biopsies of particularly deep seated lesions with spindle cell or small round blue cell cytomorphology. Diagn. Cytopathol. 2017;45:168–172. © 2016 Wiley Periodicals, Inc.     

Predominant human herpesvirus 6 variant A infant infections in an HIV‐1 endemic region of Sub‐Saharan Africa

Human herpesvirus 6, HHV‐6, commonly infects children, causing febrile illness and can cause more severe pathology, especially in an immune compromised setting. There are virulence distinctions between variants HHV‐6A and B, with evidence for increased severity and neurotropism for HHV‐6A. While HHV‐6B is the predominant infant infection in USA, Europe and Japan, HHV‐6A appears rare. Here HHV‐6 prevalence, loads and variant genotypes, in asymptomatic compared to symptomatic infants were investigated from an African region with endemic HIV‐1/AIDS. DNA was extracted from blood or sera from asymptomatic infants at 6 and 18 months age in a population‐based micronutrient study, and from symptomatic infants hospitalised for febrile disease. DNA was screened by qualitative and quantitative real‐time PCR, then genotyped by sequencing at variable loci, U46 (gN) and U47 (gO). HIV‐1 serostatus of infants and mothers were also determined. HHV‐6 DNA prevalence rose from 15% to 22% (80/371) by 18 months. At 6 months, infants born to HIV‐1 positive mothers had lower HHV‐6 prevalence (11%, 6/53), but higher HCMV prevalence (25%, 17/67). HHV‐6 positive febrile hospitalized infants had higher HIV‐1, 57% (4/7), compared to asymptomatic infants, 3% (2/74). HHV‐6A was detected exclusively in 86% (48/56) of asymptomatic HHV‐6 positive samples genotyped. Co‐infections with both strain variants were linked with higher viral loads and found in 13% (7/56) asymptomatic infants and 43% (3/7) HIV‐1 positive febrile infants. Overall, the results show HHV‐6A as the predominant variant significantly associated with viremic infant‐infections in this African population, distinct from other global cohorts, suggesting emergent infections elsewhere. J. Med. Virol. 81:779–789, 2009. © 2009 Wiley‐Liss, Inc.     

hTERC基因与高危型人乳头瘤病毒分型的相关性研究

目的：探讨宫颈上皮内瘤变石蜡标本中人染色体端粒酶基因( human telomerase gene, hTERC)表达情况及其与高危型人乳头瘤病毒测定的临床意义。方法收集2010年10月至2013年10月上海第八人民医院148例宫颈石蜡组织标本,其中对照组20例(正常宫颈),研究组128例( CINⅠ级30例, CINⅡ级53例, CINⅢ级45例)。应用荧光原位杂交( FISH)技术检测其hTERC基因的表达并采用凯普HPV-DNA分型测定上述受试者高危型人乳头瘤病毒感染情况。结果①随着宫颈病变级别增高, hTERC基因阳性表达率增加。不同级别宫颈上皮内瘤变hTERC基因扩增率与正常对照组比较,差异有显著统计学意义(χ2=63．707, P<0．05); CINⅡ级及以上病变的hTERC基因扩增率明显高于CINⅠ级,差异有显著统计学意义(χ2=36．973, P<0．05)。②受试者中高危型HPV感染阳性率为74．3%(110/148),高危型HPV感染阳性组hTERC基因扩增率和阴性组相比较,差异具有统计学意义(χ2=26．9, P<0．05)。结论宫颈上皮内瘤变组织中hTERC基因扩增与HPV感染具有一定相关性。     

Reassessment of a chromosome 12q + marker by fluorescent in situ hybridization (FISH)

We present a case previously described by Jenkins et al. (1983) as atypical Down syndrome (DS). The initial diagnosis was first made on the basis of phenotypic and cytogenetic data. This analysis was supported by studies of superoxide dismutase (SOD1) activity that maps to band 21q22.1. Results from phenotypic, chromosome banding and SODI studies suggested a karyotype of 46,XX,—12, + t(12pter to 12qter::21q21 to 21q22.?2). Using fluorescent in situ hybridization (FISH) for chromosome painting with DNA libraries derived from sorted human chromosomes to stain selectively the chromosomes No. 21 and No. 12, we demonstrate that the marker chromosome 12q+ has no chromosome 21 content but it is derived from chromosome 12.     

Detection of numerical alterations of chromosome 1 in cytopathological specimens of breast tumors by chromogen in situ hybridization

To investigate the effectiveness of chromogen in situ hybridization (CISH) in the diagnosis of breast tumors, numerical alterations of chromosome 1 were examined by CISH and fluorescence in situ hybridization (FISH) methods, and the presence of der(16)t(1;16) was also examined by FISH in imprinted cytology specimens from resected tissues of 14 carcinomas and five non-malignant lesions. The modal signal counts of chromosome 1 were compared between the specimens processed by CISH and FISH for each case. Aneusomies of the long arm of chromosome 1 were detected in 10 (71%) carcinomas as the major clones by both methods. In addition, one atypical papilloma demonstrated tetrasomy of 1q12 as a major clone by CISH, but such a clone was at first overlooked by FISH. Four other benign lesions showed disomic 1q12 signals as a major clone by both CISH and FISH. As additional information from FISH, eight cancers showed structural or numerical alterations of chromosome 16, and four showed der(16)t(1;16). In total, 10 carcinomas showed chromosome 16 alterations, and all of these overlapped with the carcinomas with 1q12 aneusomies. The CISH method provided almost the same results as the FISH method, and both methods were considered applicable in supportive diagnosis of cytological specimens of breast tumors. In addition, the CISH method was superior in the detection of numerical alterations in carcinoma cells by referring to the morphology of cells and in the detection of significant clones which might be missed under dark-field microscopy.     

Demonstration of the Philadelphia translocation by fluorescence in situ hybridization (FISH) in paraffin sections and identification of aberrant cells by a combined FISH/immunophenotyping approach

The Philadelphia translocation was demonstrated by two-colour fluorescence in situ hybridization (FISH) in decalcified paraffin sections of bone marrow from patients with chronic myelogenous leukaemia. FISH was combined with immunocytochemical detection of different membrane-bound or cytoplasmic antigens. With this new technique, the cells bearing the 9;22 translocation can be identified morphologically, as well as immunocyto-chemically, in tissue sections.     

Correlation between the detection of viral DNA by the polymerase chain reaction in peripheral blood leukocytes and serological responses to human herpesvirus 6, human herpesvirus 7, and cytomegalovirus in renal allograft recipients

Diagnosis of significant infections by human herpesvirus 6 (HHV6) and 7 (HHV7) in transplant patients has proved difficult because both viruses are ubiquitous and can cause persistent infections in their hosts. The significance of viral DNA detected in peripheral blood leukocytes (PBLs; DNAemia) by PCR is therefore unclear. The interpretation of serological results is complicated by the fact that both primary and secondary infections with other herpesviruses may be associated with a concurrent antibody response to HHV6. Fifty-four renal allograft recipients were studied prospectively and their serological response to HHV6, HHV7 and CMV were compared with the detection of viral DNAemia from the homologous and heterologous viruses. Serum and heparinised blood samples were collected prospectively from 54 renal allograft recipients. DNA was extracted from PBLs and tested for the presence of HHV6, HHV7 and CMV DNA by PCR. Antibodies to HHV6 and HHV7 were measured by an indirect immunofluorescence test and to CMV by an anticomplement immunofluorescence (ACIF) test. CMV IgM antibodies were detected by a commercial enzyme immunoassay. CMV and HHV7 DNAemia were each significantly associated with serological responses to the homologous virus but no such association was found for HHV6 DNAemia. However, patients with consecutively positive DNAemia to any of the viruses (including HHV6) were more likely to have a homologous serological response. Patients who had detectable CMV IgM without a concurrent rise in CMV antibodies were significantly less likely to have CMV DNAemia (odds ratio = 0.16; 95% CI 0.02–0.9). CMV IgM antibodies may be associated with HHV6 or HHV7 DNAemia (odds ratio 2.3; 95% CI 0.5–15). This serological profile may reflect a cross-reactive response to HHV6, HHV7 or other herpesviruses. CMV IgM should not be used in isolation for the diagnosis of CMV infection or disease in this group of patients. J. Med. Virol. 53:288–294, 1997. © 1997 Wiley-Liss, Inc.     

天然菌斑生物被膜中变形链球菌、远缘链球菌和血链球菌的荧光原位检测

目的 检测菌斑生物被膜初期形成过程中的变形链球菌(Streptpcoccus mutans)、远缘链球菌(Streptpcoccus sobrinus)和血链球菌(Streptpcoccus sanguis).方法 从植入釉质磨片表面获得完整的菌斑生物被膜标本,应用激光共聚焦扫描显微镜和荧光原位杂交技术相结合的方法,对天然菌斑生物被膜形成初期的变形链球菌、远缘链球菌和血链球菌进行原位的、实时的动态观察,通过连续断层扫描及三维重建,观察这3种菌在天然菌斑生物被膜中的空间分布,测量3种细菌的扫描厚度.实验中的扫描厚度结果采用方差分析方法进行统计处理,SPSS11.5统计软件辅助完成,α设在0.05.结果 在激光共聚焦扫描显微镜下观察生物被膜呈三维立体结构,形态多样,生物被膜内层和外层细菌较稀疏,中间层较密集,细菌之间可见许多黑色空隙,贯穿整个生物被膜.菌斑生物被膜变形链球菌、远缘链球菌和血链球菌在菌斑生物被膜形成初期的平均扫描厚度随时间延长而增加,1 h时平均扫描厚度分别为20.43、11.50、14.76 μm,到24 h时平均厚度最大,分别为70.25、75.40、79.98 μm.结论 应用荧光原位杂交技术结合激光共聚焦扫描显微镜,可以快速、灵敏地检测出菌斑生物被膜变形链球菌、远缘链球菌和血链球菌.     

Trisomy rescue by postzygotic unbalanced (X;14) translocation in a girl with dysmorphic features

In this report we present the clinical features and molecular and cytogenetic findings in a female with partial trisomy 14q. Molecular and cytogenetic studies allowed us to determine that the extra 14q material (of paternal origin) was translocated postzygotically onto the maternal X chromosome. Consequently, only the derivative X chromosome was inactivated, although inactivation apparently did not spread over the entire chromosome 14q. This partial inactivation makes the present case unusual, giving rise to phenotypic features absent in other patients with partial trisomy 14q, typically restricted to the distal part of the chromosome.