Detection of serum hepatitis B, C, and D viral nucleic acids and its implications in hepatocellular carcinoma patients

The association of viremia, elevated serum alanine aminotransferase (ALT) levels, and hepatocyte inflammatory activity in hepatocellular carcinoma (HCC) patients was studied. Serum samples from 114 HCC patients undergoing surgery were assayed for hepatitis B, C, and D viral nucleic acids by polymerase chain reaction (PCR) prior to surgery. Of these patients, 65 had HBV infection alone, 15 had HCV infection alone, 4 had HDV infection, 20 had HBV and HCV superinfection, 1 had triple viral infection, and 9 were negative for HBV and HCV infections. The prevalence of active viral replication was significantly higher in HCV than in HBV (92% versus 70%; P = 0.006) patients, and significantly higher mean serum ALT levels were also noted in the HCV group than in the HBV group (P = 0.02). The incidence of marked ALT elevation (>200 U/l) was highest in the HCV (27%) and the HDV (25%) groups. Patients in the HCV group were 10 years older than those in the HBV group. Viral superinfection did not accelerate the development of HCC. Viral replication persisted in a significant portion of HCC patients and a higher prevalence of hepatic inflammation was noted in patients with HCV- and, possibly, HDV-related HCC. (Received Sept. 22, 1997; accepted Dec. 19, 1997)     

Heat sensitivity of human parvovirus B19

BACKGROUND AND OBJECTIVES: To date there has been no published report on a systematic evaluation of the heat sensitivity of human parvovirus B19 (B19) and the related safety of the plasma-derived fractionated products. In this study, we examined the heat sensitivity of B19 by using the infectivity assay with cultured cells. MATERIALS AND METHODS: The heat sensitivity of B19 was examined by measuring the reduction in viral infectivity titres after heating liquid containing B19 at 60 degrees C. Viral infectivity was assayed by detection of viral antigens or viral mRNA in infected cells. As a control, canine parvovirus (CPV) was also heat-treated. RESULTS: B19 displayed quite different inactivation kinetics to CPV when both were heated in liquid at 60 degrees C. In sharp contrast to the latter, B19 was rapidly inactivated within 1 h when the virus was suspended in 5% or 25% human serum albumin solution, phosphate-buffered saline, or complete medium. However, B19 appeared to be resistant to heat inactivation in liquid containing 60% sucrose. CONCLUSIONS: The heat sensitivity of B19 in liquid was clearly different from that of CPV. Significantly, the efficiency to inactivate B19 and reduce its infectivity following heating in liquid was mainly affected by the composition of the solutions used for virus suspension.     

The structure of human parvovirus B19

Human parvovirus B19 is the only parvovirus known to be a human pathogen. The structure of recombinant B19-like particles has been determined to approximately 3.5-A resolution by x-ray crystallography and, to our knowledge, represents the first near-atomic structure of an Erythrovirus. The polypeptide fold of the major capsid protein VP2 is a "jelly roll" with a beta-barrel motif similar to that found in many icosahedral viruses. The large loops connecting the strands of the beta-barrel form surface features that differentiate B19 from other parvoviruses. Although B19 VP2 has only 26% sequence identity to VP3 of adeno-associated virus, 72% of the C(alpha) atoms can be aligned structurally with a rms deviation of 1.8 A. Both viruses require an integrin as a coreceptor, and conserved surface features suggest a common receptor-binding region.     

乙型肝炎病毒前S1抗原对判断病毒复制和乙型肝炎预后的价值

目的探讨乙型肝炎病毒前S1抗原与HBVM和HBV DNA的相关性,评价前S1抗原与病毒复制和在判断乙型肝炎预后中的作用。方法采用ELISA法检测前S1抗原;采用时间分辨免疫荧光法检测HBVM;采用酶学速率法检测肝功能;采用荧光定量PCR方法检测HBV DNA。结果在HBeAg阳性的141例患者中,HBV DNA和前S1抗原的检出率分别为97.2%和92.9%,抗-HBe阳性的183例中,分别为39.9%和37.2%;在HB-sAg阳性/抗-HBc阳性的59例中分别为62.7%和54.2%;慢性乙型肝炎轻度159例,中度132例,重度96例,其血清前S1抗原阳性率分别为56.6%、73.5%和61.5%,三组之间无显著性相差。结论检测前S1抗原可以较好地反映HBV的存在和复制,其在乙型肝炎发病中的临床意义还有待探讨。     

Detection of yellow fever viral RNA by nucleic acid hybridization and viral antigen by immunocytochemistry in fixed human liver

Histopathologic examination of liver from patients with yellow fever is often not diagnostic. We therefore compared 2 virus-specific assays applicable to fixed liver, in situ nucleic acid hybridization and an immunocytochemical [alkaline phosphatase-antialkaline phosphatase (APAAP)] technique. Yellow fever structural gene sequences were detected by use of 35S-labeled negative-sense RNA probe (but not by immunocytochemistry) in 11 of 17 livers from children with fatal illness during the 1965 epidemic in Senegal. These fixed liver samples had been stored at ambient temperatures for 23 years. Both techniques were diagnostic on tissues collected 15-37 months before testing. Immunocytochemistry is a practical procedure for rapid specific diagnosis of liver stored for months, whereas RNA-RNA hybridization is a sensitive technique which can be applied to material stored for years.     

节律基因hBmal1在人结肠癌中的表达及临床意义

目的检测hBmal1在人结肠癌中的表达,探讨其临床意义。 方法应用免疫组织化学方法检测湖北省肿瘤医院30例人结肠癌患者中hBmal1的表达,以图像分析软件进行相关测定,并分析其表达与临床及病理常用指标的关系。 结果人结肠癌组织和癌旁正常对照组织中hBmal1表达阳性率为76.67%（23/30） vs 26.67%（8/30）（χ²=15.016,P<0.01）;hBmal1在人结肠癌Dukes分期B、C期中强阳性表达率为46.15%（6/13） vs 88.24%（15/17）（χ²=6.212,P=0.013）。 结论hBmal1在人结肠癌组织中有高表达,可能与癌症的发生发展侵袭转移有关。     

Clinical manifestations of human parvovirus B19 in adults

Human parvovirus B19 has been associated with various clinical effects in a number of uncontrolled reports. To define the usual manifestations of B19 infection in adults and the factors that influence them we present a clinicoepidemiological study of an outbreak of B19 infection centered on a junior school. Four hundred fifty-three of 475 adults in this community were interviewed and blood was obtained for serological diagnosis. Fifty-four cases of recent infection were identified and were HLA typed. Fourteen of the cases were asymptomatic; 32 had an influenzalike illness; 23 a rash; and 26 an acute-onset polyarthropathy that was more common in women and lasted for up to 7 months. HLA-A, -B, and -C antigen frequencies were similar to a local control population and showed no association with symptoms except that HLA-DR1 was absent in those with persistent arthropathy.     

Association of symptomatic acute human parvovirus B19 infection with human leukocyte antigen class I and II alleles

To determine the effect of the major histocompatibility complex on the development of symptoms during acute human parvovirus B19 infection, we compared human leukocyte antigen (HLA) class I and II alleles in 36 patients with symptomatic acute B19 infection with those in >900 control subjects from northwestern England. The frequency of each of HLA-DRB1*01 (P=.016), DRB1*04 (P=.007), and DRB1*07 (P<.0001) alleles was significantly higher in parvovirus B19 patients than in control subjects. In the parvovirus group, 63.9% carried the rheumatoid arthritis-associated shared epitope sequence, compared with 45% of control subjects (odds ratio [OR], 2.2; 95% confidence interval [CI], 0.97-4.8; P=.04), and carriage was associated with fatigue during the acute phase (OR, 4.2; 95% CI, 0.8-23.9; P=.047). All symptomatic parvovirus-associated HLA-DRB1 molecules carry a neutrally charged glutamine at position 10 and a positively charged lysine at position 12 of the first hypervariable region. HLA-B49 was associated with parvovirus infection independently of HLA-DRB1*01, DRB1*04, and DRB1*07.     

B cell memory is directed toward conformational epitopes of parvovirus B19 capsid proteins and the unique region of VP1

BACKGROUND: Loss of antibody reactivity against linear epitopes of parvovirus B19 (B19) capsid proteins VP1 and VP2 occurs after infection; however, it is unclear whether B cell memory is established against linear epitopes. METHODS: B cell enzyme-linked immunospot assay was used to evaluate B19-specific B cell memory in volunteer donors (n=22). RESULTS: B cell memory is maintained against conformational epitopes of VP2 and is absent against linear epitopes of VP2. Individuals seronegative for IgG against the unique region of VP1 have detectable B cell memory, with the potential to mount a humoral response on reexposure to B19. Conversely, in mice immunized with VP2, long-lasting IgG against linear epitopes of VP2 and a strong B cell-memory response are observed. CONCLUSIONS: B cell memory is established and maintained against conformational epitopes of VP2 and against linear epitopes of VP1 but not against linear epitopes of VP2. These findings further our understanding of the immune response to B19 and suggest that analysis of B19-specific B cell memory merits consideration for future B19-vaccine studies.     

Lupus-like presentation of human parvovirus B19 infection.

The diagnosis of systemic lupus erythematosus (SLE) was a leading initial consideration in 2 patients with rash, arthritis and hypocomplementemia. One patient also had leukopenia and thrombocytopenia. Spontaneous regression occurred. In both patients antinuclear antibodies were negative. Serologic studies indicated recent human parvovirus B19 infection. We propose adding human parvovirus B19 infection to the list of conditions that may masquerade as SLE.     

Maternal-fetal transmission of human parvovirus B19 genotype 3

Plasma samples obtained at delivery from 885 pregnant Ghanaian women were tested for human parvovirus B19 DNA and B19-specific antibodies. Maternal-fetal transmission was evaluated by testing paired maternal plasma and umbilical cord blood samples, as well as newborn whole-blood samples when they were available. The B19 DNA seroprevalence rate in women was 1.8% (94% had genotype 3 strains), and the immunoglobulin G (IgG) seroprevalence rate in women was 81%. Two of 3 cases of primary maternal B19 infection resulted in fetal transmission. Coexistence of B19 DNA and B19-specific IgG (persistence) was detected in 13 women (1.5%), but no transmission of the virus was observed. Contrary to the situation in pregnant women with primary B19 infection and high viral loads, pregnant women with low viral loads and B19-specific IgG do not appear to be vertically infectious.     

The presence of parvovirus B19 VP and NS1 genes in the synovium is not correlated with rheumatoid arthritis

OBJECTIVE: To amplify both NS1 and VP genes of Parvovirus B19 DNA in synovial membrane (SM) and serum obtained from patients with rheumatoid arthritis (RA) and to analyze whether the presence of viral DNA is correlated with synovitis. METHODS: DNA obtained from 30 SM and 24 serum samples from RA patients was analyzed using single round-polymerase chain reaction (PCR) and nested PCR for both VP and NS1 genes of parvovirus B19. Twenty-four SM and serum samples from sex and age matched subjects with osteoarthritis (OA) or joint trauma served as controls. RESULTS: The first round PCR was negative for NS1 in RA samples. After nested PCR, NS1 was detected in the SM of 6/30 patients and of 10/24 controls and in the serum of 4/24 patients and controls. Nested PCR for the VP gene detected viral DNA in the SM of 7/30 patients with RA and of 7/24 of the controls and in the serum of 5/24 patients and of 2/24 controls. Altogether parvovirus DNA was found in the SM of 11/30 (36.6%) patients and of 12/24 (50%) controls and in the serum of 8/24 (33.3%) patients with RA and of 5/24 (20.8%) controls. CONCLUSION: Our results suggest that the amplification by nested PCR of both NS1 and VP genes is necessary to define the presence of viral DNA in tissue samples and confirm that the presence of parvovirus B19 DNA is similar in RA and control SM, suggesting that simple detection of viral DNA is not sufficient to confirm a link between the virus and RA.     

The relationship between arthritis and human parvovirus B19 infection

In order to evaluate the role of human parvovirus B19 in the etiopathogenesis of autoimmune diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), synovial fluid and blood specimens were collected at 1-month intervals from 20 patients with early synovitis (ES) and 31 with RA. Blood specimens were also collected from 25 patients with SLE, 25 with osteoarthritis (OA) as the diseased control group, and 50 healthy blood donors (HBD) as the healthy control group. Detection of B19 IgM and B19 IgG were performed by enzyme-linked immunosorbent assay from serum specimens, and B19 DNA was detected by polymerase chain reaction from synovial fluid samples. B19 IgM, B19 IgG, and B19 DNA were found in the three patients of the ES group. Subsequently, two of them were diagnosed with RA and one with SLE. B19 DNA was also detected in the synovial fluid of eight patients in the RA group. Of them, all were positive for B19 IgG and half were positive for B19 IgM. B19 IgM was not detected in either of the control groups. To define the role of B19 in the etiopathogenesis and prognosis of undiagnosed arthritis and other chronic inflammatory diseases such as RA and SLE, we need broader serial and prospective studies based on clinical and laboratory collaboration. In conjunction with case reports, these studies would also serve to detect other possible factors in the etiopathogenesis of chronic inflammatory diseases.