Amplification of GB virus-C/hepatitis G virus RNA with primers from different regions of the viral genome

GB virus-C/hepatitis G virus (GBV-C/HGV) is a newly identified RNA virus. The aim of the study was to compare three primer pairs from the 5′ untranslated region (5′UTR), envelope region 2 (E 2) and nonstructural region 3 (NS 3) of GBV-C/HGV genome for their ability to detect GBV-C/HGV RNA by polymerase chain reaction (PCR) assays. By using PCR with primers from different regions of the viral genome, serum GBV-C/HGV RNA was assayed in 200 at-risk individuals. The sensitivity of this assay was assessed by a titration experiment, and nucleotide sequences of the amplified products were determined directly. Of 200 serum samples, 43 (21.5%) were positive for GBV-C/HGV RNA with at least one of the primer pairs. The positive rates by 5′UTR, NS 3, and E 2 primers were 100%, 98%, and 84%, respectively, and the sensitivity of PCR assays using 5′UTR primers was 10 to 100 times more likely to detect GBV-C/HGV RNA than that of NS 3 and E 2 primers. The average homology of amplified targets to the prototype HGV genome was 89%, 80%, and 85% and the similarity between each amplified target was up to 100%, 90%, and 92% in the 5′UTR, E 2, and NS 3 regions, respectively. Therefore, the 5′UTR of GBV-C/HGV genome is highly conserved and primers deduced from this region can provide a sensitive and specific PCR assay for GBV-C/HGV RNA. J. Med. Virol. 51:284–289, 1997. © 1997 Wiley-Liss, Inc.     

Retrospective analysis of non-A-E hepatitis: possible role of hepatitis B and C virus infection

In an effort to determine the cause of non-A-E hepatitis, a retrospective study was undertaken on a group of patients with hepatitis but without serological infection markers of hepatitis viruses A-E. A total of 60 patients admitted to Beijing Ditan Hospital during the period of September 1997 and September 1999 were chosen for this study. These patients were diagnosed as either acute or chronic hepatitis, but no serological markers of hepatitis viruses A-E were detected. Since TT virus (TTV), human parvovirus B19 (B19), SEN virus (SENV), and GB virus C/HGV were reported to be associated with hepatitis, attempts were made to detect the presence of these viruses in the sera of patients with non-A-E hepatitis by a nested polymerase chain reaction (nPCR) method. Also, more sensitive nPCR and RT-nPCR methods were used to determine HBV DNA and HCV RNA in these patients. Results derived from these analyses demonstrate that HBV DNA was detected in most of these patients (47/60, 78.3%), suggesting that HBV infection played a major role in occult non-A-E hepatitis and detection of HBV DNA by more sensitive PCR methods such as nPCR should be considered for diagnosis of HBV infection. In addition, HCV RNA was detected in three (5%) of these patients. However, GBV-C (HGV) RNA was not detected, and TTV, B19, and SENV appear not to be associated with non-A-E hepatitis, as the prevalence rates of these viruses in patients with non-A-E hepatitis were similar to those in patients with viral hepatitis A-E. The results from this study indicate that co-infection of TTV or B19 with HBV did not increase the severity of the disease.     

Etiology of sporadic acute viral hepatitis in Taiwan: the role of hepatitis C virus, hepatitis E virus and GB virus-C/hepatitis G virus in an endemic area of hepatitis A and B.

The etiology of sporadic acute hepatitis was studied in 334 consecutive patients from Taiwan (237 men and 97 women, aged 16-81 years), with emphasis on the role of hepatitis C virus (HCV), hepatitis E virus (HEV), and GB virus-C/hepatitis G virus (GBV-C/HGV) in acute non-A, non-B (NANB) hepatitis and in HBsAg carriers with superimposed acute hepatitis. According to the conventional diagnostic criteria, there were 12 cases (3.6%) of acute hepatitis A, 17 cases (5.1%) of acute hepatitis B, 128 cases (38.3%) of acute NANB hepatitis, and 177 cases (53.0%) of acute hepatitis in HBsAg carriers (those who were HBsAg positive but IgM anti-HBc negative). Among 128 cases of acute NANB hepatitis, 70 (54.7%) had acute hepatitis C (HCV RNA positive), 5 (3.9%) had acute hepatitis E (IgM anti-HEV positive), and the other 53 (41.4%) were presumably acute hepatitis non-A-E. The prevalence of acute hepatitis A, B, E, and non-A-E showed no significant sex difference, whereas acute hepatitis C was significantly more prevalent in females. The prevalence of acute hepatitis A and B decreased and that of acute hepatitis C increased significantly with increasing age. In contrast, acute hepatitis E and non-A-E showed no significant age predominance. Of 177 HBsAg carriers with acute hepatitis, 64 (36.1%) demonstrated non-B hepatotropic virus superinfection, with HCV being the most common (60.9%), followed by hepatitis D, E, and A viruses, and the other 55 (31.1%) and 58 (32.8%) were presumed to have acute exacerbation of chronic hepatitis B or superimposed acute hepatitis non-A-E, respectively. Serum GBV-C/HGV RNA was detected in 3-4% of acute hepatitis non-A-E cases, suggesting its limited role in these cases.     

庚型肝炎病毒IgM抗体检测方法的建立及其临床意义

目的建立一种早期、快速诊断庚型肝炎的血清学检测方法。方法以辣根过氧化物酶标记庚型肝炎病毒（ＨＧＶ）多肽ＮＳ３,ＮＳ５区段抗原,建立了捕获酶联免疫吸附试验（ＥＬＩＳＡ）,用于检测血清中ＨＧＶＩｇＭ抗体。结果本法不受特异性ＩｇＧ的竞争和类风湿因子的干扰;与其它致肝炎的病毒（ＨＡＶ、ＨＢＶ、ＨＣＶ、ＨＥＶ、ＣＭＶ、ＥＢＶ）无交叉反应。检测４６例非甲、乙、丙、戊型肝炎患者血清,抗－ＨＧＶＩｇＭ阳性１４例,阳性率３０．４３％,其中,６例同时为ＨＧＶＲＮＡ阳性,阳性符合率为４２．８６％（６／１４）,检测１２例庚型肝炎病人双份血清,其中,急性期血ＨＧＶＩｇＭ抗体均为阳性。结论该法检测ＨＧＶＩｇＭ抗体特异性强,敏感性高,且简便快速,适用于临床对庚型肝炎新近感染的早期诊断,有推广应用价值。     

Follow-up of four HIV-infected individuals after administration of hepatitis C virus and GBV-C/hepatitis G virus contaminated intravenous immunoglobulin: Evidence for HCV but not for GBV-C/HGV transmission

In 1994, hepatitis C virus (HCV) infection was transmitted to four HIV seropositive patients attending the Department of Angiology, University Clinics, Frankfurt am Main, by the administration of Gammagard®. The patients were suffering from thrombocytopenia and received betweeen 20 and 30 g of the contaminated lot 93F21AB11. GBV-C/HGV RNA could be amplified from the Gammagard® lot 93F21AB11 using 5′NCR and NS5 primer pairs. All the four patients were negative in the GBV-C/HGV RT-PCR prior to therapy and until the end of the follow-up period. GBV-C/HGV IgG antibodies to the putative envelope (E2) were detected using the E2 HGV-env kit (Boehringer-Mannheim, Germany) in Gammagard® lot 93F21AB11 and in one patient before donation of immunoglobulin. Anti-E2 seroconversion was observed in one recipient, the other two patients remained anti-E2 seronegative until the end of the observation period. It is concluded that there is no direct evidence for transmission of GBV-C/HGV by contaminated intravenous immunoglobulin since GBV-C/HGV RNA was not detected in the recipients up to 1 year after administration. J. Med. Virol. 53:25–30, 1997. © 1997 Wiley-Liss, Inc.     

Identification of GBV-C hepatitis G RNA in chronic hepatitis C patients

Sera from patients with chronic hepatitis C were examined for the presence of GBV-C/HGV RNA by RT-PCR. The amplified products, derived from the 5′ non-coding, NS3, and NS5a regions, were detected in 19 (19%) of the 100 HCV RNA-positive samples. Analysis of GBV-C/HGV prevalence rates revealed that dual infections are related to shared parenteral risk factors. Intravenous drug abuse and multiple transfusions were the factors clearly associated with a simultaneous HCV and GBV-C/HGV infection. Apart from this, patients with dual infections had a statistically significant lower mean age compared to those patients infected solely by HCV. Determination of HCV genotypes involved in GBV-C/HGV coinfection by RFLP analysis showed no correlation between the presence of GBV-C/HGV and a distinct HCV genotype. The study demonstrates that, based on the assessment of risk criteria, GBV-C/HGV is transmitted efficiently parenterally and is frequently linked to hepatitis C coinfection, regardless of HCV genotype. © 1996 Wiley-Liss, Inc.     

用逆转录—套式聚合酶链反应检测我国不同临床型肝？…

目的 为了研究庚型肝炎病毒（ＨＧＶ）在我国的感染状况。方法 概括已发表的ＨＧＶ的５‘端非编码区（５’－ＵＴＲ区）及螺旋酶区（ＮＳ３区）两段高度保守的基因序列分别设计两套引物，用逆转录－套式聚合酶链式反应检测ＨＧＶＲＮＡ。结果 从北京、秦皇岛、河南等地采集各种肝病患者及职业献血员血清３５４份，ＨＧＶＲＮＡ阳性９７份，阳性率为２２．３％。其中已确定的临床型肝炎／肝病患者２５４例，ＨＧＶＲＮＡ阳性者为５     

Detection of GB Virus-C/Hepatitis G Virus RNA in Serum by Reverse Transcription Polymerase Chain Reaction

Three PCR methods based on the GB virus-C/hepatitis G virus (GBV-C/HGV) 5′UTR and NS3 genomic region were used for the detection of GBV-C/HBV RNA in serum of 62 patients with chronic hepatitis C virus (HCV) infection. Ten of 62 (16%) patients were found to have GBV-C/HGV RNA, which was confirmed by sequence analysis of the 5′UTR PCR amplicon. All methods appear to be specific, but methods based on the 5′UTR appear to be more sensitive. J. Med. Virol. 52:92–96, 1997. © 1997 Wiley-Liss, Inc.     

Genetic heterogeneity and molecular epidemiology of GB virus C/hepatitis G virus in China

OBJECTIVE: The inter- and intrapatient genetic variation of GB virus C (GBV-C)/hepatitis G virus (HGV) was investigated to characterize the molecular epidemiologic profile of GBV-C/ HGV infection in China, an area endemic for viral hepatitis. The intrapatient variation of hepatitis C virus (HCV) from the same patients was compared to that of GBV-C/HGV. STUDY DESIGN/METHODS: GB virus C/HGV RNA was amplified by polymerase chain reaction in 88 patients with hepatitis C, hepatitis B or presumed non-A-E hepatitis from three cities in China. Five clones of the GBV-C/HGV NS3 region were sequenced from each GBV-C/HGV RNA-positive patient. The corresponding region of HCV was also sequenced from patients co-infected with HCV. Representative sequences of the GBV-C/HGV NS3 region from each patient and those of isolates from other continents were subjected to phylogenetic analyses. RESULTS: GB virus C/HGV was detected in 22 (25.25%) of 88 patients: 9 (21.4%) of 42 patients with presumed non-A-E hepatitis, 10 (27.7%) of 36 patients with hepatitis C, 3 (30.0%) in 10 patients with hepatitis B and C, and in none of 60 volunteer blood donors. The extent of nucleotide variation was less between Chinese isolates (2.4-17%; median, 10.4%) than between Chinese isolates and seven isolates from outside China (10.5-19.5%; median, 15.3%). Intrapatient sequence variation ranged from 0 to 1.75%, with a mean of 0.57 +/- 0.51%. Phylogenetic analysis grouped most Chinese isolates into four geographically specific clusters with a divergence of 10% to 16% from each other. The ratio of nonsynonymous to synonymous substitutions of GBV-C/HGV (Ka/Ks 0.019) was much lower than for HCV (0.071) in the same patients. CONCLUSION: Chinese isolates of GBV-C/HGV are genetically distinct. There are local strains as well as shared strains between different locales. The extent of amino acid sequence conservation suggests strong selection against nonsynonymous substitutions in the GBV-C/HGV genome.     

庚型肝炎病毒致病性的初步研究

目的 探讨庚型肝炎病毒（ＨＧＶ）的致病性。方法 应用ＲＴ－ｎＰＣＲ检测３６８例肝炎患者血清ＨＧＶ ＲＮＡ和血清酶的变化,并对其中１例单独庚型肝炎肝硬化病例进行肝脏活组织是检测。结果 在７１例急性黄疸型肝炎中检出单纯性ＨＧＶ ＲＮＡ阳性７例,１５５例慢性肝炎中检出单纯性ＨＧＶ ＲＮＡ阳性２２例,５１例肝硬化中检出单纯ＨＧＶ ＲＮＡ阳性３例。其中１例肝穿组织免疫组化证实为ＨＧＶ ＮＳ ５Ａｇ阳性。结论 争性黄疸型肝炎及乙、丙型肝炎病毒携带者,其慢性肝炎、肝硬化和肝癌中均可检出ＨＧＶ ＲＮＡ,ＨＧＶ感染可为单独感中与乙／丙型肝炎病毒混合或重叠感染,肝脏病理和免疫组化检查证实庚型肝炎病毒是一种嗜肝病毒,其定位主要存在于细胞浆内,可引起慢性病毒性肝炎甚至肝硬化。庚型肝炎病毒很可能具有致病性。     

利多组套式引物的测庚型肝炎病毒RNA

利用庚型肝炎病毒ＮＳ３－５区序列合成了四组套式引物，建立了灵敏，特异的 型肝炎病毒ＲＮＡ双扩增聚合酶链反应检测方法，用此方法检测了１０份庚型肝炎病毒抗体阳性患者血清及１０份阴性的健康人血清。前者不同组引物的检出率为ＮＳ３（１）引物９份了是性，ＮＳ３（２）引物８份阳性，ＮＳ４引物４份阳性，ＮＳ５（１）引物５份阳性，ＮＳ５（２）引物９份阳性；后者各组引物匀为阴性。结果表明，庚型肝炎病毒不同区域引物用于     

利用多组套式引物检测庚型肝炎病毒RNA

利用庚型肝炎病毒（ＧＢＶ－Ｃ／ＨＧＶ）ＮＳ３～５区序列合成了四组套式引物，建立了灵敏、特异的庚型肝炎病毒ＲＮＡ双扩增聚合酶链反应（ＰＣＲ）检测方法。用此方法检测了１０份庚型肝炎病毒抗体阳性患者血清及１０份阴性的健康人血清。前者不同组引物的检出率为ＮＳ３（１）引物９份阳性，ＮＳ３（２）引物８份阳性，ＮＳ４引物４份阳性，ＮＳ５（１）引物５份阳性，ＮＳ５（２）引物９份阳性；后者各组引物检测均为阴性。结果表明，庚型肝炎病毒不同区域引物用于ＲＴ－ＰＣＲ检出率差异较大。ＮＳ３（１）及ＮＳ５（２）这两组引物检出率最高，更适合用于ＲＴ－ＰＣＲ检测庚型肝炎病毒ＲＮＡ。     

High prevalence of GBV-C hepatitis G virus infection in a rural South African population

A novel virus, GBV-C/hepatitis G virus (GBV-C/HGV), has been cloned and characterised recently. GBV-C/HGV global epidemiology and risk factors for acquisition are currently unclear. We aimed to establish the determinants of this infection in a rural South African (SA) population. The study population included two samples, namely a community-based sample, and consenting persons from a nonspecialist outpatient department in the same district. A questionnaire regarding demographic details and putative risk factors was administered; blood samples were taken on which a polymerase chain reaction (PCR) was performed for both 5′NCR and NS5a regions of GBV-C/HGV using commercially available primers and probes. Two hundred and forty-nine people were studied with a mean GBV-C/HGV prevalence of 10.4%. Outpatient department and community prevalences differed significantly (18.0% and 6.3%, respectively, P = 0.004). GBV-C/HGV infection was associated with excessive alcohol consumption (P = 0.02; OR, 4.18) and a lack of waterborne sewerage (P = 0.04). PCR amplification of the NS5a region of all but two South African GBV-C/HGV positive samples showed poor reactivity. The prevalence of GBV-C/HGV in rural SA appears to be higher than that reported from Europe and North America. Infection appeared to be associated with excess alcohol intake and a history of previous blood transfusions. The discrepant NS5a and 5′NCR PCR sensitivity in this study raises the possibility of genetic differences in southern African GBV-C/HGV. J. Med. Virol. 53:225–228, 1997. © 1997 Wiley-Liss, Inc.