上海肝病在线 Shanghai Liver Diseases' Online

从仅抗HBs AB（乙肝表面抗体）阳性的HBV变异株感染者再论---治疗性复合HBV系列疫苗

这几个月会诊，跑得最多的科室是肾炎科，会诊病人中，其中最多的是各种类型的HBV慢性感染者，如慢乙肝和HBV健康携带者。但亦不少的血清抗HBS抗体高滴度阳性的病人，这些病人中或肝功能正常，或肝功能异常。结合前些时间门诊所见一例病人（一女孩因体育课后血尿而检测血清二对半示HBV M中仅抗HBS抗体阳性而肾穿剌活检可见肾内大量HBS AG和HBC AG沉积）和新近科室谈及将收治的一例病人（一例患者长期肝功异常而抗HBS抗体阳性但肝活检示肝内大量HBS AG和HBC AG沉积并肝内HBV DNA（PCR）阳性），今特作一辑，论及ANTI-HBS阳性的一特定类型的HBV感染（HBS区变异型HBV感染，其特征是HBS AG/表面抗原阴性或/和ANTI-HBS阳性而肝脏和肾脏等实质细胞内和/或血清或/和血细胞内HBV DNA（PCR）阳性的HBV感染）。

因免疫压力。如HBV疫苗使用或乙肝丙球使用或免疫偏倚（因TH2/TH3反应而产生无效的抗HBS抗体，如与IgG3和IgG2A型相比，IgG4型抗HBS抗体没有保护作用，不利于HBS AG的清除而反而阻止HBS AG清除），致HBS区变异而表现为乙肝表面抗原（HBS AG）阴性并甚至乙肝表面抗体（ANTI-HBS 或HBS AB）阳性。这类病人多认为是乙肝已愈而最易于忽视，应加以重视，因为这种HBV变异株亦有传染性，故其流行病学意义更大。而肝癌病人因HBV DNA与肝细胞基因整合而致HBS AG阴性，没有明显传染性，但肝细胞内仍HBV DNA阳性。而慢乙肝特别是重症肝炎因HBV变异株与野生株同时存在而出现HBS AG和ANTI-HBS双阳性，则HBV DNA多不显下降，并表明出现毒力更强的HBV变异株即出现致机体免疫损伤的HBV变异株。

事实上只有IgG3和IgG2A型的抗HBS抗体有良好的抗HBV作用即是真正的保护性抗体，而IgG1型的抗HBS抗体作用可疑，而IgG4型的抗HBS抗体不仅不利于抗HBV而且有阻止HBV病毒清除作用，即IgG4型的抗HBS抗体不是保护性抗体而与抗HBC抗体和抗HBE抗体一样，反而有阻止机体的HBV病毒的清除作用。而重症肝炎因HBV变异株与野生株同时存在而出现HBS AG和ANTI-HBS双阳性并HBV DNA阳性，其ANTI-HBS的Ig类型和IgG亚型研究将更有助于HBV感染中所激活的免疫类型（TH1、TH2、TH3等）与预后关系的研究。可叹，国人对此研究不深，故乙肝治疗效果虽多年攻关而成果累累，但真正应用于临床治疗则疗效多不如人意，可谓“雷声大而雨点小”。

由于此类病人对常规化疗性抗病毒药仍多有效，但不能根治。此类病人体内HBV因其HBS区变异，故免疫治疗中常规乙肝疫苗没有治疗意义，而且理论上说，正是因为目前开始大量应用HBS AG型预防性疫苗于肝炎治疗中（作用治疗性疫苗使用），故将导致更多的乙肝表面抗原（HBS AG）阴性并甚至乙肝表面抗体（ANTI-HBS 或HBS AB）阳性的HBV变异株阳性慢乙肝病人产生 ！！！

由于HBV感染的清除有赖于机体针对HBV的HBV S、前S1、前S2、C/E、X、DNA 多聚酶等区域相关抗原（尤其是针对HBC AG）的特异性CD4淋巴细胞辅助下的B细胞体液免疫和CD8+ CTL等细胞免疫，而治疗性复合HBV系列疫苗因其激发／产生多克隆、多位点（针对HBV S、前S1、前S2、C/E、X、DNA多聚酶等多处、多位点） 、强力和全面（细胞与体液）的免疫功能，故对各种HBV感染（包括各种类型HBV变异株感染）均有治疗效果。

关于国内外乙肝表面抗原（HBS AG）阴性并甚至乙肝表面抗体（ANTI-HBS 或HBS AB）阳性的HBS区变异型HBV感染，有大量报道，兹列一些英文文摘于下以供参阅。

Gastroenterology 2002 Mar;122(3):614-24

Resolution of chronic hepatitis B and anti-HBs seroconversion in humans by adoptive transfer of immunity to hepatitis B core antigen.

Lau GK, Suri D, Liang R, Rigopoulou EI, Thomas MG, Mullerova I, Nanji A, Yuen ST, Williams R, Naoumov NV.

BACKGROUND & AIMS: Impaired T-cell reactivity is believed to be the dominant cause of chronic hepatitis B virus (HBV) infection. We characterized HBV-specific T-cell responses in chronic hepatitis B surface antigen carriers who received bone marrow from HLA-identical donors with natural immunity to HBV and seroconverted to antibody to hepatitis B surface antigen. METHODS: T-cell reactivity to HBV antigens and peptides was assessed in a proliferation assay, the frequency of HBV core- and surface-specific T cells was quantified directly by ELISPOT assays, and T-cell subsets were analyzed by flow cytometry. RESULTS: CD4+ T-cell reactivity to HBV core was common in bone marrow donors and the corresponding recipients after hepatitis B surface antigen clearance, whereas none reacted to surface, pre-S1, or pre-S2 antigens. Furthermore, CD4+ T cells from donor/recipient pairs recognized similar epitopes on hepatitis B core antigen; using polymerase chain reaction for the Y chromosome, the recipients' CD4+ T lymphocytes were confirmed to be of donor origin. The frequency of core-specific CD4+ and CD8+ T cells was several-fold higher than those specific for surface antigen. CONCLUSIONS: This study provides the first evidence in humans that transfer of hepatitis B core antigen-reactive T cells is associated with resolution of chronic HBV infection. Therapeutic immunization with HBV core gene or protein deserves further investigation in patients with chronic hepatitis B.

精采华丽篇：接受天然HBV免疫的供体进行BMT致HBV感染者的HBS AG清除，其HBV清除作用与HBC AG特异性TH/CD4T细胞从天然HBV免疫的供体过继给HBV感染的受体有密切关系，即天然HBV免疫的供体的HBC AG特异性TH/CD4T细胞在受体内长期存在于受者体内并发挥其抗HBV免疫的免疫调控作用

J Virol 1995 Jun;69(6):3358-68

Activation of a heterogeneous hepatitis B (HB) core and e antigen-specific CD4+ T-cell population during seroconversion to anti-HBe and anti-HBs in hepatitis B virus infection.

Jung MC, Diepolder HM, Spengler U, Wierenga EA, Zachoval R, Hoffmann RM, Eichenlaub D, Frosner G, Will H, Pape GR.

Institute for Immunology, University of Munich, Germany.

Overcoming hepatitis B virus infection essentially depends on the appropriate immune response of the infected host. Among the hepatitis B virus antigens, the core (HBcAg) and e (HBeAg) proteins appear highly immunogenic and induce important lymphocyte effector functions. In order to investigate the importance of HBcAg/HBeAg-specific T lymphocytes in patients with acute and chronic hepatitis B and to identify immunodominant epitopes within the HBcAg/HBeAg, CD4+ T-cell responses to hepatitis B virus-encoded HBcAg and HBcAg/HBeAg-derived peptides were studied in 49 patients with acute and 39 patients with chronic hepatitis B. The results show a frequent antigen-specific CD4+ T-cell activation during acute hepatitis B infection, a rare HBcAg/HBeAg-specific CD4+ T-cell response among HBeAg+ chronic carriers, and no response in patients with anti-HBe+ chronic hepatitis. An increasing CD4+ T-cell response to HBcAg/HBeAg coincides with loss of HBeAg and hepatitis B virus surface antigen (HBsAg). Functional analysis of peptide-specific CD4+ T-cell clones revealed a heterogeneous population with respect to lymphokine production. Epitope mapping within the HBcAg/HBeAg peptide defined amino acids (aa) 1 to 25 and aa 61 to 85, irrespective of the HLA haplotype, as the predominant CD4+ T-cell recognition sites. Other important sequences could be identified in the amino-terminal part of the protein, aa 21 to 45, aa 41 to 65, and aa 81 to 105. The immunodominant epitopes are expressed in both proteins, HBcAg and HBeAg. Our findings lead to the conclusion that activation of CD4+ T lymphocytes by HBcAg/HBeAg is a prerequisite for viral elimination, and further studies have to focus on the question of how to enhance or induce this type of T-cell response in chronic carriers. The immunodominant viral sequences identified may have relevance to synthetic vaccine design and to the use of peptide T-cell sites as immunotherapeutic agents in chronic infection.

精采篇：HBV清除作用与HBC AG/HBe Ag特异性TH/CD4T细胞激活有密切关系，指示今后的HBV治疗性疫苗应重点注意激活机体的针对HBV的C/E区抗原的免疫作用

Liver 1988 Oct;8(5):307-16

Latent hepatitis B virus infection with full-length viral genome in a patient serologically immune to hepatitis B virus infection.

Blum HE, Offensperger WB, Walter E, Offensperger S, Gerok W.

Department of Medicine, University of Freiburg, West Germany.

The presence, state, physical structure and cellular localization of hepatitis B virus (HBV) DNA were investigated in a patient with hepatitis B surface antigen (HBsAg)-negative chronic liver disease. HBV serology was positive for antibodies to hepatitis B core antigen (anti-HBc), to hepatitis B e antigen (anti-HBe) and to HBsAg (anti-HBs); no HBV DNA was detectable in serum. Southern blot analyses of DNA extracted from the liver demonstrated free monomeric HBV DNA as two distinct species: a predominant species of fully double-stranded relaxed circular molecules and a minor species of linear molecules of 3.2 kilobase pairs (kbp) length. Restriction enzyme analyses identified the HBV genome as HBsAg subtype adw2. Cell fractionation studies further revealed that the free viral DNA species were localized exclusively in liver cell nuclei. These findings in a patient serologically immune to HBV infection demonstrate that in hepatocytes HBV can establish a latent infection, characterized by the extrachromosomal presence of a full-length viral genome without production of infectious virus or synthesis of viral antigens.

PMID: 3200125

J Med Virol 1990 Dec;32(4):257-60

Serum hepatitis B virus DNA in healthy HBsAg-negative Chinese adults evaluated by polymerase chain reaction.

Shih LN, Sheu JC, Wang JT, Huang GT, Yang PM, Lee HS, Sung JL, Wang TH, Chen DS.

Department of Internal Medicine, College of Medicine, National Taiwan University Hospital, Taipei, R.O.C.

Serum hepatitis B virus (HBV) DNA was assayed using polymerase chain reaction, in 107 HBsAg-negative normal Chinese subjects. The results showed that eight subjects (7.5%) had HBV DNA. In the subgroup with antibody to hepatitis B surface antigen (anti-HBs) and to hepatitis B core antigen (anti-HBc), 7.3% (5/68) were positive for HBV DNA; HBV DNA was not detected in six individuals with anti-HBs only and in nine with anti-HBc only. In four persons with anti-HBc and anti-HBe, one had HBV DNA. In 20 subjects negative for all hepatitis B serological markers, two (10%) were found to have HBV DNA. This study indicates that serological markers are not adequate to rule out HBV infection, and it further implies that present blood donor screening methods may need improving.

PMID: 2081972

Zhonghua Nei Ke Za Zhi 1991 Jan;30(1):21-3

[Identification of HBV infection in HBsAg-negative chronic active hepatitis with polymerase chain reaction] [Article in Chinese]

Luo KX, Zhou R, Liang ZS.

Nanfang Hospital of First Military Medical College.

To explore the etiological agent(s) of HBsAg negative chronic active hepatitis, serum HBV DNA was tested in thirty-six of such cases with polymerase chain reaction. In total, viral DNA was detected in 24 cases (67%), including 11 of the 12 cases with positive anti-HBc alone, four of the nine cases with positive anti-HBs and nine of the 15 cases without any HBV markers. The results suggest that in cases of HBsAg-negative chronic active hepatitis, the etiology may still be hepatitis B virus, especially in cases with positive anti-HBc. Furthermore, the cause of activated histology might be due to viral replication.

PMID: 2032490

Liver 1990 Feb;10(1):6-10

Persistence of hepatitis B virus DNA after serological clearance of hepatitis B virus.

Tanaka Y, Esumi M, Shikata T.

Department of Pathology, Nihon University School of Medicine, Tokyo, Japan.

Using Southern blot technique, the state of hepatitis B virus (HBV) DNA in liver tissue was investigated in 16 patients who were sero-negative for hepatitis B surface antigen (HBsAg) but positive for its antibody (anti-HBs). In only one case, was HBV DNA found in liver tissue in a heterogeneously integrated form. In this case, digestion with Taq I demonstrated integrated HBV DNA as two definite bands at 1.8 and 0.5 kbp. This suggests that HBV DNA in some cases persists even after HBV infection has been cleared serologically. It is possible that this persistence of HBV DNA plays an important role in hepatocarcinogenesis.

PMID: 2308481

AIDS 1988 Dec;2(6):443-8

Hepatitis B virus reactivation or reinfection associated with HIV-1 infection.

Waite J, Gilson RJ, Weller IV, Lacey CJ, Hambling MH, Hawkins A, Briggs M, Tedder RS.

Department of Medical Microbiology, University College and Middlesex School of Medicine, London, UK.

Following acute hepatitis B virus (HBV) infection, most individuals develop antibodies to HBV surface (anti-HBs) and core antigen (anti-HBc). Prevalence studies have shown that 10-18% develop anti-HBc in the absence of detectable anti-HBs. We report four such cases, all with persistence of serum anti-HBc, who had evidence of a second period of active HBV replication as demonstrated by the reappearance of serum hepatitis B surface antigen (HBsAg). In one patient, an HBsAg subtype difference indicated that the second period of HBsAg-positivity was due to a reinfection. In the other cases, reactivation may also explain the findings. All cases were anti-HIV-1 seropositive at the time of reappearance of HBsAg. There is experimental evidence that anti-HBc has a protective effect against HBV infection; however, this may require intact cell-mediated immunity to be effective. HIV-1 infection may render such patients susceptible to reinfection. Alternatively, some patients with anti-HBc, but without detectable anti-HBs may have latent HBV infection. Immunosuppression associated with HIV-1 infection may allow reactivation.

PMID: 3149492

Gastroenterol Jpn 1991 Oct;26(5):661-5

A case of HBs antigen negative fulminant hepatitis with IgM antibody to hepatitis B core antigen persisting more than seven years.

Suga M, Shibata K, Kodama T, Arima K, Yamada S, Yachi A.

Department of Internal Medicine (Section 1), Sapporo Medical College, Japan.

A 33-year old dentist developed fulminant hepatitis. At admission, a test for IgM antibody to hepatitis B core antigen (IgM anti-HBc) was positive, while tests for HBsAg and HBeAg were negative. He was cured of the disease, but in follow-up examinations from 1983 to 1990 IgM anti-HBc was continuously detected with radioimmunoassay while HBsAg and HBV-DNA were absent in the serum. However, HBcAg was found in a biopsied liver specimen and a small quantity of HBV-DNA was detectable by polymerase chain reaction assay. These observation suggest that the continuous detection of IgM anti-HBc without HBsAg in serum is due to persistent HBV infection and HBV replication in the liver.

PMID: 1752398

J Med Virol 1991 Oct;35(2):133-5

Acute hepatitis B viral infection in a patient with anti-HBs.

Wang YJ, Lee SD, Wu JC, Lo KJ.

Department of Medicine, Veterans General Hospital, Taipei, Taiwan, Republic of China.

We report a patient with antibody to hepatitis B surface antigen (anti-HBs) but no antibodies to other hepatitis B virus components, who developed acute symptomatic type B hepatitis. The possible explanations for this unusual serological pattern are 1) the antibody-positive status, which developed against only a subdeterminant of hepatitis B surface antigen (HBsAg), arose naturally or as the result of cross-reaction with a variety of antigens; and 2) seroconversion to anti-HBs occurred in response to surface antigen of a mutant strain of hepatitis B virus (HBV). This anti-HBs positivity, in the absence of antibody to hepatitis B core antigen, does not provide natural immunization against HBV infection, and so is not protective. Individuals who are positive to anti-HBs antibody alone which is not elicited by HBV vaccine, should be vaccinated against possible HBV infection.

PMID: 1765776

Scand J Infect Dis 1992;24(5):657-60

Reactivation of hepatitis B virus infection with an unusual pattern of serological markers.

Kidd Ljunggren K, Hansson BG, Wallmark E, Croxson MC, Kidd AH, Nordenfelt E.

Department of Infectious Diseases, University of Lund, Sweden.

A 73-year-old man presented with acute hepatitis, judged to be a reactivation of hepatitis B virus infection. His serum samples during a follow-up time of 16 months showed an unusual pattern of serological markers. He was consistently HBeAg positive, HBsAg fluctuated just under the cut-off value and he had a low level of circulating anti-HBs. By electron microscopy numerous aggregates of surface antigen particles, but not complete virions were seen. He was HBV DNA positive by hybridization. The complete precore and core genes and a region of the surface gene were amplified from his serum by PCR. These findings emphasize the need for expanded serological testing in some patients with acute clinical hepatitis.

PMID: 1465585

Zhonghua Nei Ke Za Zhi 1992 Oct;31(10):629-32, 658-9

[Hepatitis B virus DNA in the serum of anti-HBs positive persons] [Article in Chinese]

Zhang YX, Du SC, Chen P.

Department of Internal Medicine, General Hospital of Beijing Military District.

In order to investigate the significance of HBV DNA in the serum of anti-HBs positive persons, the serum of 76 anti-HBs positive persons was studied for HBV DNA by means of polymerase chain reaction (PCR). The results showed that 21 (32.2%) out of 65 cases without hepatitis B vaccination were positive for HBV DNA detected with PCR (PCR-HBV DNA), but no one was positive for PCR-HBV DNA in 11 cases inoculated against hepatitis B. It was also found that 6 cases were positive for HBsAg-Ab immunocomplex in those positive for PCR-HBV DNA and the liver tissue in 2 of the 5 cases with liver-biopsy were positive for HBVAg determined with immunohistologic ABC method. We believed that persons, who acquired anti-HBs after HBV infection were different from those who were vaccinated, might carry HBV which come from the HBsAg-Ab immunocomplex and HBVAg positive hepatocytes. In addition, the study also proved that the PCR-HBV DNA positive rate correlated significantly with the anti-HBe and or anti-HBc positive rate and with the abnormal rate of liver function in the anti-HBs positive persons. It was suggested that persistent presence of HBV DNA in the bodies should be responsible for the persistent presence of anti-HBe and anti-HBc in the serum and also for the liver damage.

PMID: 1306457

Chin Med J (Engl) 1993 Jan;106(1):7-12

Hepatitis B virus DNA detected by PCR in sera and liver tissues of Chinese patients with chronic liver diseases.

Zhang YY, Guo LS, Li L, Zhang YD, Hao LJ, Hansson BG, Nordenfelt E.

Clinical Immunology Research Unit, Tongji Hospital, Tongji Medical University, Wuhan. To investigate the HBV infection and its replication in Chinese patients with chronic liver disease, polymerase chain reaction (PCR) was employed to detect hepatitis B virus DNA in sera of 410 patients with chronic liver disease and liver samples from 188 patients. The HBV DNA detectability in all serum samples was 58%. Among them 100% HBeAg-positive and 58% anti-HBe cases were HBV DNA detectable respectively. However, HBV DNA was also found in 23% HBsAg-negative/anti-HBc positive chronic cases. Furthermore, 30% anti-HBs positive chronic cases who had neutralizing antibody against HBV infection, continued to contain HBV DNA. Our findings indicate that HBV infection and its replication are dominant cause of chronic liver disease and some HBV variants may escape from the protective antibody to induce chronic liver disease, even anti-HBs antibody circulated.

PMID: 8504690

J Hepatol 1993 Mar;17(3):288-93

Hepatitis B virus occult infection in subjects with persistent isolated anti-HBc reactivity.

Sanchez-Quijano A, Jauregui JI, Leal M, Pineda JA, Castilla A, Abad MA, Civeira MP, Garcia de Pesquera F, Prieto J, Lissen E.

Viral Hepatitis and AIDS Study Group, Virgen del Rocio University Hospital, Seville, Spain.

The aim of this study was to investigate the presence of hepatitis B virus occult infection in asymptomatic subjects with persistent anti-HBc reactivity but no other hepatitis B virus serological markers, including HBsAg, anti-HBs, IgM anti-HBc and HBV-DNA. For this purpose we used both polymerase chain reaction assays in sera and immunohistochemistry for HBsAg and HBcAg in liver biopsy specimens. Twenty-four cases were studied: 15 were drug abusers or homosexuals (eight with normal alanine aminotransferase levels) and nine were heterosexuals with raised alanine aminotransferase levels (> 45 U/l) but with no history of blood transfusion or ethanol intake (< 80 g daily). In all but five cases, liver biopsy was performed in subjects with persistent elevated alanine aminotransferase levels. In 10 out of 24 cases (41.66%) hepatitis B virus infection was demonstrated by polymerase chain reaction or immunohistochemistry, and when results from both procedures were available (n = 11) hepatitis B virus infection was detected in 63.63% of the subjects. The only clinical feature associated with HBV infection was the presence of persistent elevated alanine aminotransferase levels (p < 0.05). In conclusion, persistent isolated anti-HBc reactivity may be a relatively common serologic pattern for hepatitis B virus occult infection, at least in patients with chronic liver disease.

PMID: 8315257

Hepatology 1993 Apr;17(4):538-44

Hepatitis B virus DNA in serum and liver is commonly found in Chinese patients with chronic liver disease despite the presence of antibodies to HBsAg.

Zhang YY, Hansson BG, Kuo LS, Widell A, Nordenfelt E.

Department of Medical Microbiology, University of Lund, Malmo General Hospital, Sweden.

Sera from 410 patients from the Wuhan area in the central part of China with the diagnosis of chronic liver disease were analyzed for markers of hepatitis B, C and D virus infections. All sera, plus liver biopsy specimens from 188 of the patients, were also tested for hepatitis B virus DNA by polymerase chain reaction. Sixty-eight percent were HBsAg positive in serum, whereas 29% showed markers of past hepatitis B virus infection. Hepatitis B virus DNA was detected in all HBeAg-positive sera but also in 58% of patients with HBe antibody. In the liver specimens of the corresponding patient groups, 97% and 78%, respectively, were hepatitis B virus DNA positive. However, more noteworthy was that of the HBsAg-negative/HBs-antibody positive patients 30% had detectable hepatitis B virus DNA in serum and 32% had hepatitis B virus DNA in liver tissue, whereas in a control group of healthy blood donors, of which 90% had HBs antibody, none was hepatitis B virus DNA positive. Our results demonstrate that among patients with chronic liver disease, infections with hepatitis B virus or hepatitis B virus-related virus(es) may frequently occur without being revealed by conventional serological methods. Hepatitis C and D viruses seem to be of only minor importance in the pathogenesis of chronic liver disease in this part of China.

PMID: 7682978

Schweiz Med Wochenschr 1993 Jun 12;123(23):1193-202

[Hepatitis serological finding "anti-HBc alone", circulating viral DNA and interpretation of findings] [Article in German]

Gross A, Joller-Jemelka HI, Wicki AN, Grob PJ.

Departement fur Innere Medizin, Universitatsspital Zurich, Schweiz.

The immunological finding "anti-HBc alone" (without HBsAg, without anti-HBs) leaves open the question of the state of HBV infection it reflects: a transitory stage of an uncomplicated, eventually prolonged but resolving infection, a chronic or a late state of immunity. A finding of this kind is often observed in immunocompromised individuals (e.g. patients on hemodialysis, drug addicts) but also occurs in up to 1% of the Swiss blood donor population. Of 8800 sera tested for HBV marker in a diagnostic laboratory, 153 individuals showed "anti-HBc alone". They were investigated for circulating hepatitis B desoxyribonucleic acid (HBV-DNA) by polymerase chain reaction (PCR). 60 individuals (39%) showed a positive result. Also taking into consideration anamnestic measurements of conventional HBV markers in 95 individuals and consecutive testing for HBV-DNA in 50 individuals, the following conclusions emerged: 1. A positive finding of HBV-DNS by PCR does not necessarily prove an ongoing HBV infection, hence a negative result does not rule it out. Therefore, the indication to test for this parameter is limited for routine use. 2. The finding of "anti-HBc alone" implies that a HBV-infection is still going on until proven otherwise. This not only might be of consequence for the individual involved, but also raises the question of screening of blood donors and of pregnant women for anti-HBc. Publication Types: Review Review, Tutorial

PMID: 8327866

J Clin Invest 1994 Jan;93(1):230-9

Erratum in: J Clin Invest 1994 Aug;94(2):following 905

Hepatitis B virus persistence after recovery from acute viral hepatitis.

Michalak TI, Pasquinelli C, Guilhot S, Chisari FV.

Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037.

Contrary to current opinion, the disappearance of hepatitis B surface antigen (HBsAg) from the serum, the development of anti-HBs antibodies, and normalization of liver function may not reflect complete virological recovery from acute hepatitis B virus (HBV) infection. By using the polymerase chain reaction (PCR), in the current study we demonstrate long-term persistence of HBV DNA in the serum and peripheral blood mononuclear cells (PBMC) of four patients for up to 70 mo after complete clinical, biochemical, and serological recovery from acute viral hepatitis. Serum HBV DNA reactivity co-sedimented with HBsAg in sucrose gradients, and it displayed the size and density characteristics of naked core particles and intact HBV virions, presumably contained within circulating immune complexes in these anti-HBs antibody-positive sera. HBV DNA was also present in PBMC in late convalescent samples from all four patients, and HBV RNA was detected in late convalescent phase PBMC in two of these patients. These results suggest that HBV DNA, and possibly HBV virions, can be present in the serum, and that the viral genome can persist in a transcriptionally active form in PBMC for > 5 yr after complete clinical and serological recovery from acute viral hepatitis.

PMID: 8282792

J Infect Dis 1994 Jun;169(6):1374-6

Breakthrough infections and identification of a viral variant in Gambian children immunized with hepatitis B vaccine.

Fortuin M, Karthigesu V, Allison L, Howard C, Hoare S, Mendy M, Whittle HC.

International Agency for Research on Cancer, The Gambia Hepatitis Intervention Study, Banjul.

Hepatitis B (HB) breakthrough infections, identified by the presence of HB core (c) antibody, were found in 32 of 358 Gambian children vaccinated with plasma-derived HB vaccine. Over 2 years, 15 of these children lost their HBc antibodies. These children had significantly higher HB surface antibody levels before infection than those who retained HBc antibodies. One child, who responded well to the vaccine, had HB viral DNA detected in the presence of HBs antibodies. The S gene sequence of this DNA showed nucleotide changes that resulted in an amino acid substitution at residue 141 (lysine to glutamic acid) of the surface antigen. This finding suggests the child was infected with a variant virus that was not neutralized by antibodies resulting from HB vaccination.

PMID: 8195620

J Hepatol 1994 Aug;21(2):269-72

Detection of HBs antigen in "anti-HBc alone" positive sera.

Joller-Jemelka HI, Wicki AN, Grob PJ.

Department of Internal Medicine, University Hospital, Zurich, Switzerland.

The immunoserological finding "anti-HBc alone" is often observed in defined groups of individuals, such as patients with inflammatory hepatopathies, patients on hemodialyses or with organ transplants, i.v. drug users and homosexuals, but it also occurs in up to 1% of Swiss blood-donors. In order to gain further information about whether "anti-HBc alone" reflects late immunity or points to an ongoing or a recently passed hepatitis B virus infection, 153 serum samples were tested for immune-complex-dissociated HBs-antigen, using acid treatment for complex dissociation. Of the samples tested 31% contained complexed HBsAg, the highest rates being found in individuals with hepatopathies (up to 80%), in i.v. drug users (up to 63%) and in hemodialysis patients (40%). The 153 sera were also tested for HBV-DNA by nested PCR. Sixty (39%) probes yielded positive results, comprising 29 (48%) of 60 sera with immune-complexed HBsAg but only 18 (19%) of 93 probes without complexed HBsAg. The results point to the possibility that at least some of the individuals with "anti-HBc alone" still have an ongoing HBV-infection.

PMID: 7989721

Leber Magen Darm 1994 Sep;24(5):215-7

[A rare etiology for HBs-Ag negative acute hepatitis B--coinfection by hepatitis B and delta] [Article in German]

Korner T, Jaspersen D, Roth J, Hammar CH, Bassler R. Med. Klinik II, Stadt. Klinikums Fulda.

An unusual case of a 25-year-old male Italian is reported. The patient endured an acute hepatitis without detectable HBs-antigen by coinfection with hepatitis-B and Delta. Coincidently, a cured hepatitis-C was present. Firstly hepatitis-B-virus DNA could be demonstrated in a small quantity by serodiagnosis (6 pg/ml, hybridization technique). Subsequently, the identification of B-virus DNA was only possible in liver tissue (PCR-technique), but no longer by serodiagnosis. The probable enduring inhibition of hepatitis-B-virus replication by Delta virus resulted in a self limitation of the disease within 2 months (HDV-RNA negative, HBs-Ag and HBe-Ag negative; Anti-HBs negative, Anti-HBe and Anti-HBc positive). In spite of negativation of replication markers for hepatitis-B a subsequent reactivation of the infection was possible by viral material which persisted in liver tissue.

PMID: 7968181

Przegl Epidemiol 1995;49(3):317-9

[Concurrence of HBs antigen and anti-HBs antibodies in children infected with HBV] [Article in Polish]

Styczynski J, Halota W, Balcar-Boron A, Dorau M.

Katedra i Klinika Chorob Dzieci Akademii Medycznej w Bydgoszczy.

Two cases of concurrence of HBs antigen and antibodies in boys, who acquired HBV infection within the first two years of life are described. Authors discuss possible explanation of that fact.

PMID: 7491429

Liver 1994 Oct;14(5):241-4

Inapparent "wild-type" and "e-minus variant" HBV infection in patients with HCV-related chronic hepatitis.

Sardo MA, Rodino G, Brancatelli S, Pernice M, Campo S, Squadrito G, Russo F, Raimondo G.

Dipartimento di Medicina Interna, Universita di Messina, Italy.

We analysed DNA extracted from liver biopsy specimens and serum samples from 42 HCV-RNA-positive/HBsAg-negative subjects with chronic hepatitis. Twenty-eight of them were anti-HBs/anti-HBc-positive (group A), while 14 were negative for all HBV markers (group B). HBV sequences were found in hepatic DNA of 12 cases (11 of group A, one of group B), but in the serum of only two cases of group A. Sequencing analysis of pre-core region of HBV-DNA showed the presence of wild-type HBV in three cases, HBeAg-defective HBV in three cases, and the coexistence of both viral populations in six cases. These results indicate that HBV and HCV infection may coexist in HBsAg-negative chronic hepatitis, particularly in anti-HBs/anti-HBc-positive patients. However, HBV replication appears suppressed in these cases, and this state of latency may involve both wild and HBeAg-defective HBV types.

PMID: 7997082

J Med Virol 1995 Dec;47(4):410-5

Emergence of hepatitis B virus S gene mutant in a liver transplant recipient.

Cariani E, Ravaggi A, Tanzi E, Romano L, Fiordalisi G, Bellati G, Caccamo L, Galmarini D, Albertini A, Zanetti A.

III Laboratory of Clinical Chemistry, School of Medicine, University of Brescia, Italy.

Immunological and genomic analysis of the "a" determinant was carried out in seven patients with concurrent HBsAg and anti-HBs, four of whom were immunized against hepatitis B virus at liver transplant, two with histologically characterized chronic hepatitis B virus infection, and one HBsAg healthy carrier. The immune reactivity of the HBsAg "a" determinant was evaluated by binding to specific monoclonal antibodies, and the corresponding genomic sequence was studied by differential hybridization in microtiter plates and nucleotide sequence analysis. A double mutation generating an amino acid change (glycine to lysine) at residue 145, able to impair recognition by monoclonal antibodies, was observed in the post-transplant serum from one patient. No significant alteration of the "a" determinant sequence or reactivity was detected in the other patients. Amino acid residue 145 appears therefore to be critical for the recognition by anti-HBs antibodies. A previously undescribed glycine to lysine substitution at this level interferes with the immune reactivity of the "a" determinant.

PMID: 8636711

J Hepatol 1996 Jan;24(1):8-14

Hepatitis B virus surface mutations associated with infection after liver transplantation.

Hawkins AE, Gilson RJ, Gilbert N, Wreghitt TG, Gray JJ, Ahlers-de Boer I, Tedder RS, Alexander GJ.

Academic Department of Genitourinary Medicine, University College London Medi

BACKGROUND/AIMS: Liver transplantation for chronic liver disease due to hepatitis B virus infection is associated with a high risk of graft infection, graft failure and death. Many centres restrict this procedure to those seronegative for HBV-DNA (by hybridisation assay) and use prophylactic polyclonal human hepatitis B specific immunoglobulin to prevent infection of the graft, despite the very high cost. METHODS: We describe three patients who underwent liver transplantation for chronic HBV-related disease in whom death was due to fibrosing cholestatic hepatitis following graft infection with hepatitis B virus, despite receiving hepatitis B specific immunoglobulin. Variation within the immunodominant a epitope of HBsAg was sought by analysis of hepatitis B virus sequences and the use of a point mutation assay, following amplification from serum by the polymerase chain reaction. RESULTS: Prior to transplantation, Cases 1 and 2 had mutations at nucleotide 1902 (codon 145), resulting in G-C substitutions, which persisted at a low level after transplantation. In Case 2 a second mutant type with a G-A substitution at nucleotide 1902, became the predominant viral type post transplant. Case 3 had exclusively wild type virus before and after transplantation. The emergence of mutant type virus in Case 2 may have occurred because of immune pressure exerted by high titre anti-HBs detectable for more than 7 months. Cases 1 and 3 received only brief courses of anti-HBs therapy. The mutant viral surface antigen was not detected by a monoclonal antibody-based assay, and therefore the choice of HBsAg assay for post-transplant monitoring of patients who receive liver grafts for hepatitis B virus disease is important. CONCLUSIONS: A search for mutations affecting the a determinant prior to liver transplantation for HBV-related liver disease may help to identify those at risk of failure of prophylaxis. Monoclonal antibodies specific to the codon 145-mutant surface antigen might prevent graft infection, but other mutations might then emerge.

PMID: 8834018

Scand J Infect Dis 1996;28(1):9-15

Increasing heterogeneity of the 'a' determinant of HBsAg found in the presumed late phase of chronic hepatitis B virus infection.

Zhang YY, Nordenfelt E, Hansson BG.

Department of Medical Microbiology, Section of Clinical Virology, University of Lund, Malmo General Hospital, Malmo, Sweden.

To investigate if the concomitant presence of hepatitis B virus (HBV) DNA and antibodies to hepatitis B surface antigen (anti-HBs) is associated with mutations in the HBV envelope gene, selected sequences of the HBV genome in 54 patients with chronic liver disease, collected from the Wuhan district of China, were amplified by polymerase chain reaction (PCR) and the DNA sequences of the products were determined. The part of the S gene coding for the 'a' determinant of HBsAg was found to be prone to diversity. A total of 19 aberations occurring at 11 of the 69 nucleotide positions of this part of the genome were found in sera from 15 HBsAg-negative but anti-HBs-positive patients. One of 13 HBsAg/HBeAg-positive and 8 of 17 HBsAg/anti-HBe-positive samples also showed point mutations in this gene sequence. Most prevalent was a point mutation from adenine to guanine at nucleotide 530 resulting in a change from threonine to alanine at amino acid position 126. This study highlights that the long time course of chronic HBV infection could favour a selection of escape mutants. The occurrence of such HBV variants in patients with chronic liver disease are not detected by conventional HBV serology, and the patients can therefore easily by be misdiagnosed. If viral mutants like those described here can be transmitted to other patients, there will be difficulties in identifying these infections, and conventional HBV vaccination will presumably not be protective against them.

PMID: 9122641

Acta Virol 1996 Jun;40(3):133-8

Detection of hepatitis B virus DNA in hepatitis B surface antigen-negative serum by polymerase chain reaction: evaluation of different primer pairs and conditions.

Gomes SA, Yoshida CF, Niel C.

Department of Virology, Oswaldo Cruz Foundation, Rio de Janeiro, Brazil.

The presence of hepatitis B virus (HBV) DNA was investigated by polymerase chain reaction (PCR) in the serum of twenty Brazilian blood donors. All sera were negative for hepatitis B surface antigen (HBsAg), 17 of them presented antibodies to the hepatitis B core antigen (anti-HBc) as the unique serological marker of HBV infection, and 3 were positive for antibodies to HBsAg (anti-HBs) and anti-HBc. PCR assays were carried out using different pairs of oligonucleotides designed from conserved sequences of C, pre-S and S regions of the HBV genome. First, all oligonucleotide pairs were tested in PCR using plasmids carrying HBV genome from ayw or adw strains as templates. One-round PCR assays were able to detect 100-25,000 molecules of plasmid DNA, depending on the oligonucleotide pair, while semi-nested PCR assays detected 10-1000 molecules. The frequency of HBV DNA-positive results with HBsAg-sera varied from 0% to 50% depending upon the PCR assay. The results indicated that a number of both isolated anti-HBc and anti-HBs+, anti-HBc+ samples contained HBV DNA at a very low concentration, neighboring the limit of detection.

PMID: 8891092

Hepatology 1996 Sep;24(3):489-93

Hepatitis B virus envelope variation after transplantation with and without hepatitis B immune globulin prophylaxis.

Carman WF, Trautwein C, van Deursen FJ, Colman K, Dornan E, McIntyre G, Waters J, Kliem V, Muller R, Thomas HC, Manns MP.

Institute of Virology, University of Glasgow, Scotland.

Hepatitis B virus (HBV) replicates via an intermediate RNA step. High frequency of polymerase errors with additional selection pressure leads to mutations in the HBV genome. We investigated the number, type, and antigenic effects of mutations in the coding region of the HBV surface antigen in eight patients who underwent orthotopic liver transplantation (OLT) for HBV-related end-stage liver disease and were experiencing infection of the graft and who received hepatitis B surface antigen antibody (anti-HBs) prophylaxis (hepatitis B immune globulin [HBIG]) after OLT. Controls were chronic HBV patients who underwent kidney transplantation and received the same immunosuppressive regime but no HBIG. The S-gene was amplified from serum before and after transplantation, sequenced, and changes in the genome were analyzed. In the five patients who experienced reinfection while receiving anti-HBs, clear mutations occurred in the S-gene. In the patient who did not receive HBIG and those who experienced reinfection only after termination of HBIG, no mutations were found in the S-gene. In the kidney recipients, mutations in the S-gene occurred in only one of eight patients. Because the a determinant contains neutralizing epitopes, this region was chosen for antibody binding to quantify antigenic effects of the mutations. The two patients who selected mutations in the a determinant and became reinfected while receiving HBIG had reduced antibody binding after OLT. Our results suggest that HBIG after OLT imposes a selection pressure on the S-gene, and that mutations are one mechanism for reinfection while receiving HBIG.

PMID: 8781312

Z Gastroenterol 1997 May;35(5):347-55

[Hepatitis B virus mutants--clinical significance] [Article in German]

Blum HE, Moradpour D, von Weizsacker F, Wieland S, Peters T, Rasenack JW.

Abteilung Innere Medizin II, Medizinische Universitatsklinik Freiburg.

Hepatitis B virus (HBV) mutants have recently been identified in patients with acute or fulminant as well as chronic infections. Naturally occurring mutations have been identified in all viral genes and regulatory elements, most notably in the genes coding for the structural envelope and nucleocapsid proteins. Mutations in the gene coding for the hepatitis B surface antigen (HBsAg) may result in infection or viral persistence despite the presence of antibodies against HBsAg (anti-HBs) ("vaccine escape" or "immune escape"). Mutations in the gene encoding the pre-core/ core protein (pre-core stop codon mutant) result in a loss of hepatitis B e antigen (HBeAg) and seroconversion to antibodies to HBeAg (anti-HBe) with persistence of HBV replication (HBeAg minus mutant). Mutations in the core gene may lead among others to an "immune escape" due to a T cell receptor antagonism. Mutations in the gene coding for the polymerase/reverse transcriptase can be associated with viral persistence or resistance to nucleoside analogues. Thus, HBV mutations may affect the natural course of infection, viral clearance and response to antiviral therapy. Apart from the precore/core stop codon mutations, the exact contribution of specific mutations to diagnosis and therapy of HBV infection as well as patient management in clinical practice remain to be established. Publication Types: Review Review, Tutorial

PMID: 9265394

Vaccine 1997 Jul;15(10):1095-100

Erratum in: Vaccine 1998 Oct;16(16):1593

Selective unresponsiveness to HBsAg vaccine in newborns related with an in utero passage of hepatitis B virus DNA.

Lazizi Y, Badur S, Perk Y, Ilter O, Pillot J.

Unite d'Immunologie Microbienne, Hopital Antoine Beclere, WHO Center of Reference and Research for Viral Hepatitis, Clamart, France.

Thirty four out of 158 (22%) newborns to mothers chronically infected by the hepatitis B virus (HBV) did not produce antibodies (Ab) to HBsAg 1 month after the last injection of the HBV vaccine supplemented with HBV specific immunoglobulins. At birth, HBV genome was detected by polymerase chain reaction (PCR) in the peripheral blood mononuclear cells (PBMC) of a large majority (28 out of 34) of these non-responder newborns but never in the other newborns who responded to the HBsAg vaccine. HBV genome was detected in serum, only in some cases (nine out of 34) and never in the absence of HBV DNA in PBMC. For nine out of 14 followed newborns, the absence of response was transitory since anti-HBs Abs appeared after 15 months, without booster, while the HBV genome had disappeared. Unresponsiveness was specific to the HBV envelope protein since all late responders and 15-months-non-responders to the HBsAg vaccine produced normal levels of Abs to the three poliovirus serotypes, to tetanus toxoid and to the pneumococcus polysaccharides. An in utero induced immune tolerance to low doses of HBsAg appears as the most plausible hypothesis to explain this unresponsiveness to HBV vaccine.

PMID: 9269052

Przegl Epidemiol 1998;52(4):469-81

[Humoral response to HBV antigens in acute and chronic hepatitis B] [Article in Polish]

Swiderska H, Nitkiewicz J, Piwowarska A, Jonczyk M, Cholewinska G, Tomaszewska D, Horban A.

Zaklad Immunopatologii PZH, Warszawa.

Sera of patients with acute and chronic hepatitis B and antigenemia were tested for the presence of anti-HBe and anti-HBs antibodies, free or bound in immune complexes. The possible occurrence of immune complexes of HBcAg in these sera was also investigated. Immune complexes were identified by antigen specific enzyme immunoassay, in which polyclonal antibodies against synthetic fragments of proteins S and C of HBV and mono- and polyclonal anti-HBc antibodies were used as a solid phase. Free and/or antigen bound anti-HBe antibodies were detected in 100% of patients with acute (81% HBeAg positive) and in 37% of patients with chronic hepatitis, all HBeAg positive. Anti-HBs antibodies or their immune complexes were found in 83% and 37% of patients, respectively. In not any patient circulating complexes of HBcAg could be identified. The results obtained support the observations that humoral immune response to HBeAg and HBsAg can be detected earlier than generally accepted; they also suggest that the detection of anti-HBs in a single sample of serum should not be considered as the evidence of elimination of infection.

PMID: 10321091

J Med Virol 1999 Jun;58(2):105-10

A novel deletion mutant of hepatitis B virus surface antigen.

Weinberger KM, Zoulek G, Bauer T, Bohm S, Jilg W.

Institute for Medical Microbiology and Hygiene, University of Regensburg, Germany. klaus-michael.weinberger@klinik.uni-regensburg.de

HBsAg is the most important serological marker for acute or chronic hepatitis B. Nevertheless, there are reports of HBsAg-negative virus carriers, either with anti-HBc as the only marker for hepatitis B virus (HBV) infection or even positive for anti-HBs and anti-HBc. We report isolates from a patient, in which a deletion in the HBs-gene was associated with persisting viremia in the presence of anti-HBs. The 62-year-old female, infected most likely by her husband, had detectable markers of chronic active hepatitis B, such as HBsAg, HBeAg, and anti-HBc-IgM, for 2 years. The patient then seroconverted to anti-HBs, although HBeAg and anti-HBc-IgM remained detectable. At this time, semiquantitative polymerase chain reaction showed about 10(4) viral genomes per milliliter of serum. Direct sequencing of the amplified products revealed a major population of DNA molecules with a deletion of nucleotide 31 of the HBs-gene, which up to now has not been described. This deletion led to a frame-shift and introduced a stop-codon after 21 amino acids of the sHBsAg. We suspect that this deletion, and the resulting HBsAg lacking the major epitopes recognized by specific antibodies, could favor ongoing viral replication, despite the presence of anti-HBs. However, because the reading frame of the polymerase was also severely damaged by this deletion, it is assumed that a minor population of intact genomes was present to help in the formation of virus particles.

PMID: 10335855

Liver 1999 Jun;19(3):177-82

Genetic alterations in the S gene of hepatitis B virus in patients with acute hepatitis B, chronic hepatitis B and hepatitis B liver cirrhosis before and after liver transplantation.

Rodriguez-Frias F, Buti M, Jardi R, Vargas V, Quer J, Cotrina M, Martell M, Esteban R, Guardia J.

Biochemistry Department, Hospital General Universitario Valle Hebron, Barcelona, Spain.

BACKGROUND: Several studies have shown that hepatitis B immunoglobulin (HBIG) imposes a selection pressure on the hepatitis B virus (HBV) S gene, and that the emergence of mutations in this region would make reinfection after orthotopic liver transplantation (OLT) possible. AIMS: This study was undertaken to analyze the presence of HBV S-gene mutations in the different stages of HBV infection and the relationship between HBIG therapy and the emergence of mutations in liver transplant recipients. METHODS: The frequency and location of mutations in the coding region of the HBV S gene were studied by PCR and direct sequencing in 30 patients (7 with acute self-limited hepatitis B, 16 with chronic hepatitis B and 7 recipients of (OLT) for HBV-related end stage liver disease who became reinfected). RESULTS: The average number of amino acid changes was higher in patients with a more advanced stage of disease, 0.57 mutations/100 positions in acute hepatitis B and 1.57 in chronic hepatitis B (1.28 in HBeAg-positive and 1.8 in anti-HBe-positive patients). The average number of substitutions in the transplanted patients was 2.7 before OLT and 3 after OLT. No amino acid substitutions were detected in the "a" determinant of HBsAg in acute hepatitis B, however, 8 substitutions were observed in 6 chronic patients. In 3 OLT patients, 4 substitutions were observed in samples before and after OLT. One of these patients, who had protective levels of anti-HBs, showed 3 additional new amino acid substitutions after OLT, suggesting escape mutant selection by the effect of HBIG therapy. No changes were observed between the consensus sequences obtained several years before and after transplantation, indicating consensus sequence stability. CONCLUSION: These results show that there is an accumulation of HBV S-gene mutations in HBV-related end-stage liver disease. Prophylaxis with HBIG mainly obtained from acute self-limited hepatitis patients who have a highly homogeneous viral population, may be one factor underlying the reinfection after liver transplantation.

PMID: 10395035

J Hepatol 1999 Jun;30(6):965-9

The role of HBV DNA quantitative PCR in monitoring the response to interferon treatment in chronic hepatitis B virus infection.

Nagata I, Colucci G, Gregorio GV, Cheeseman P, Williams R, Mieli-Vergani G, Vergani D.

Institute of Hepatology, University College London Medical School and Hospital, Denmark Hill, UK.

BACKGROUND/AIMS: To investigate whether the measurement of HBV DNA by quantitative polymerase chain reaction (PCR) is helpful in monitoring response to interferon treatment in chronic hepatitis B virus infection, we have determined sequentially serum levels of HBV DNA during and up to 18 months after treatment, in 10 patients with a sustained response (all anti-HBe positive, five also HBsAg negative and anti-HBs positive) and, as controls, in 12 non-responders. METHODS: Serum HBV DNA was measured by standard hybrisation assay (Genostics, Abbott) and by quantitative PCR (Amplicor HBV Monitor test, Roche Diagnostic Systems). RESULTS: A clear difference in HBV viral load between responders and non-responders was observed from the fourth week of treatment and was maintained throughout the study period. At the last follow up 16-26 (median 21) months after starting treatment, all the 10 responders were HBV DNA negative by hybridisation. By PCR, however, five (one anti-HBs and four anti-HBe positive) were still HBV DNA positive. In addition, one anti-HBs positive patient HBV DNA negative by PCR at last follow up, had fluctuating levels of HBV DNA by PCR during the observation period, only intermittently falling below the threshold of the assay. CONCLUSIONS: The measurement of HBV DNA by quantitative PCR provides early prediction of response to interferon, allowing prompt modification of treatment. With this technique, HBV DNA is detected in a high proportion of sustained responders, suggesting that HBV may never be completely eliminated by interferon treatment, even after anti-HBs seroconversion.

PMID: 10406171

J Med Virol 1999 Aug;58(4):346-52

A hepatitis B virus variant found in the sera of immunised children induces a conformational change in the HBsAg "a" determinant.

Karthigesu VD, Allison LM, Ferguson M, Howard CR.

Department of Pathology and Infectious Diseases, Royal Veterinary College, London, UK.

The emergence of variants in the outer envelope proteins of hepatitis B virus (HBV) are found among individuals vaccinated against HBV and asymptomatic carriers of the infection. For example, children in The Gambia vaccinated against hepatitis B may show serological evidence of breakthrough infections, particularly if anti-HBs antibodies induced by the vaccine are low in titre. A single-point mutation at nucleotide 421 of the S gene is associated with such breakthrough infections. In the present study, the antigenicity of variant HBV S protein expressed as HBsAg particles in a vaccinia virus expression system has been characterised using a panel of monoclonal antibodies directed against linear and conformational determinations of the S protein. A cellular ELISA procedure using expressed antigen in Vero cells revealed differences in reactivity using four of the six antibodies that had been raised against the adw subtype of HBV and recognise conformational epitopes in the a determinant. In two instances, an enhanced reactivity for the variant antigen was found, confirming that point mutations in the a determinant of the S protein between residues 139 and 147 may result in significant changes in conformation. These findings also demonstrate that there are distinct antigenic differences between the vaccine strains of HBsAg/ adw subtype and the predominant HBsAg subtype circulating in West Africa. The implications of this work are that serodiagnosis of HBV infections may be unreliable in populations where there is a possibility of variant HBV infections emerging in the face of increasing herd immunity to HBV as a result of vaccination, particularly using monoclonal antibody-based diagnostic tests. Such variants may play a role in the maintenance of HBV infections in endemic regions.

PMID: 10421400

Hepatogastroenterology 1999 May-Jun;46(27):1848-54

Liver graft infection by HBV S-gene mutants in transplant patients receiving long-term HBIg prophylaxis.

Santantonio T, Gunther S, Sterneck M, Rendina M, Messner M, Launois B, Francavilla A, Pastore G, Will H.

Heinrich-Pette-Institut fur Experimentelle Virologie und Immunologie an der Universitat Hamburg, Germany. t.santantonio@clininf.uniba.it

BACKGROUND/AIMS: HBV reinfection of transplant livers occurs frequently even in the presence of high doses of anti-HBs immunoglobulins. We analyzed, retrospectively, whether and which type of S-gene variants were selected by long-term polyclonal anti-HBs (HBIg) treatment leading to reinfection of patients transplanted because of chronic HBs-positive end-stage liver disease. METHODOLOGY: The preS2/S gene of the viral genomes obtained from sera before transplantation and during HBV reinfection was amplified by PCR and directly sequenced. RESULTS: According to transaminase and HBV DNA hybridization analysis, 3/18 (17%) liver transplant patients had HBV and hepatitis recurrence during anti-HBs therapy. A HBV S-gene mutant containing a G to A nucleotide mutation at position 587, converting Glycine to Arginine (G145A), was identified in all three patients as the dominant population at reinfection but not pre-transplantation. Contrary to the S-gene, no consistent nucleotide changes were found in the pre-S2 region of HBV genomes when comparing the reinfection and pre-transplantation samples. CONCLUSIONS: These data demonstrate that long-term polyclonal anti-HBs immunoprophylaxis selected the most commonly described G145R S-gene escape HBV variant which became the dominant virus population and was responsible for graft infection. Therefore, immunoglobulins with high affinity for the G145R HBs variant should be included in HBIg to prevent recurrent HBV infection in transplant patients.

PMID: 10430358

J Hepatol 1999 Aug;31(2):195-201

Antigenic characterization of pre- and post-liver transplant hepatitis B surface antigen sequences from patients treated with hepatitis B immune globulin.

Carman WF, Owsianka A, Wallace LA, Dow BC, Mutimer DJ.

Institute of Virology, University of Glasgow, UK. w.carman@vir.gla.ac.uk

BACKGROUND/AIMS: The success of treatment with hepatitis B hyperimmune globulin in preventing recurrence of hepatitis B virus infection in patients undergoing orthotopic liver transplantation depends on maintaining levels of anti-HBs sufficient to neutralise hepatitis B virus and also on patient compliance. Breakthrough infections may occur, and these have been associated with the emergence of variants in HBsAg. METHODS: Three patients, two who relapsed and one who had no evidence of hepatitis B virus infection post-orthotopic liver transplantation were studied. Polymerase chain reaction and sequencing of pre- and post-orthotopic liver transplantation samples was followed by antigenic analysis of the in vitro expressed cloned sequences. RESULTS: In two patients who were treated with hyperimmune globulin, amino acid variation in the region of the immunodominant B cell epitopes of HBsAg occurred. Sequencing of clones revealed fluctuating variant sequences over time. One had clinical relapse and immune escape was evident on in vitro antigenic analysis. Patient two lost HBsAg reactivity post-orthotopic liver transplantation. There was loss of an antigenically critical cysteine molecule; sequencing of clones revealed that this was the dominant species. The third patient relapsed when protective levels of anti-HBs were not maintained; HBsAg showed no variation compared to a standard subtype sequence. CONCLUSION: These data provide strong experimental evidence of immune escape. It appears that hyperimmune globulin provides the selection pressure. In these patients, HBsAg negativity does not exclude infection of the transplanted liver.

PMID: 10453929

Br J Haematol 1999 Oct;107(1):186-95

Evidence that anti-HBc but not HBV DNA testing may prevent some HBV transmission by transfusion.

Allain JP, Hewitt PE, Tedder RS, Williamson LM.

Department of Haematology, Division of Transfusion Medicine, East Anglia Blood Centre, Cambridge. jpa1000@cam.ac.uk

Blood donor screening for antibody to hepatitis B core antigen (anti-HBc) implemented in some countries as a surrogate marker for non-A, non-B hepatitis has been superseded by anti-HCV screening. To assess the value of anti-HBc screening for the detection of hepatitis B surface antigen-negative blood donations that might contain infectious HBV, HBV genomic detection and recipient testing were used. Blood donations were screened and confirmed by multiple anti-HBc assays. Donations containing isolated anti-HBc and those with anti-hepatitis B surface antigen (anti-HBs) level < 0.1 IU/ml were tested for the presence of HBV DNA. Recipients of previous donations from the corresponding donors during the previous 5 years were traced and tested for markers of HBV infection. Of 103 869 donations screened, 586 (0.56%) were anti-HBc positive, two of which contained HBsAg, and 413 (0.4%) had protective (>/= 0.1 IU/ml) levels of anti-HBs. Anti-HBs < 0.1 IU/ml was found in 102 of these donations (0.1%) and isolated anti-HBc in 69 (0.07%). No donations with isolated anti-HBc were HBV DNA confirmed positive. Of 278 recipients of previous donations from 171 donors at risk of HBV carriage, 12 had markers of HBV infection. Six recipients had other identified risk factors. An association with blood transfusion was considered probable in two and possible in four recipients. None of the six corresponding donors had detectable HBV DNA 6-40 months after the implicated donation. The frequency of HBV transmission by chronic carriers negative for hepatitis B surface antigen was estimated in this study to be 1 in 52,000 donations (CI 0.3-7.8/100,000) from HBsAg-negative donors. Such HBV infectious donations may not be detected by DNA amplification.

PMID: 10520040

Bone Marrow Transplant 1999 Dec;24(11):1243-4

Acute hepatitis B after autologous stem cell transplantation in a man previously infected by hepatitis B virus.

Senecal D, Pichon E, Dubois F, Delain M, Linassier C, Colombat P.

Department of Hematology, CHU Bretonneau, Tours, France.

We report a case of acute hepatitis B after autologous stem cell transplantation (ASCT) in a patient with low-grade non-Hodgkin's lymphoma. At diagnosis of the hematological disease, the patient had the characteristic serology of a previous hepatitis B infection, being Ag HBs negative, hepatitis B virus core antibody positive (anti-HBC) and hepatitis B virus surface antibody weakly positive. He developed fatal hepatitis B after autologous stem cell transplantation, suggesting reactivation consequent to immunosuppression.

PMID: 10642815

Hepatology 2000 Feb;31(2):488-95

Latent hepatitis B virus infection in healthy individuals with antibodies to hepatitis B core antigen.

Marusawa H, Uemoto S, Hijikata M, Ueda Y, Tanaka K, Shimotohno K, Chiba T.

Division of Gastroenterology, Department of Medicine, The Institute for Virus Research, Kyoto University, Kyoto, Japan.

Several recent reports have shown that hepatitis B virus (HBV) could be frequently transmitted to the recipients from donors who have antibodies to hepatitis B core antigen (anti-HBc) through liver transplantation. We provide here the molecular evidence of latent HBV infection accompanied with ongoing viral replication in the liver tissue of anti-HBc-positive healthy individuals. HBV DNA was detectable in 13 of 14 healthy donors who were positive for both anti-HBc and antibodies to hepatitis B surface antigen (anti-HBs), but in none of 3 who were positive for anti-HBs alone. The detected HBV genomes from these subjects included covalently closed circular DNA and pregenomic RNA, the replication intermediate of HBV. Notably, 5 of 7 cases tested were predominantly infected with wild type HBV strains without any mutations in the precore and core promoter regions under the presence of circulating antibody to hepatitis B e antigen. Interestingly, a predominant clone detected in one donor showed a 63-nucleotide deletion in the precore region including an encapsidation signal sequence. Our findings indicate that the majority of healthy individuals positive for anti-HBc, which had been assumed to denote a past history of transient HBV infection, were latently infected with the episomal form of HBV accompanied by ongoing viral replication and few nucleotide mutations in the precore and core regions.

PMID: 10655275

Yonsei Med J 2000 Feb;41(1):40-8

Distribution of anti-HBs levels in Korean adults.

Shin BM, Jeong KW.

Department of Clinical Pathology, Inje University School of Medicine, Seoul Paik Hospital, Korea. bmshin@unitel.co.kr

Exact titration of anti-HBs with mIU/mL unit is necessary in evaluating the success of HBV vaccination or in making a decision to increase the dose of HBV vaccination. Data of distribution of anti-HBs titers can contribute to cutting of public health costs by reducing unnecessary HBV booster doses. Moreover, anti-HBc is also an important marker for differentiation of vaccination-induced anti-HBs from infection-acquired anti-HBs. However, not much study about these subjects has been done in Korea. So we evaluated anti-HBs associated with anti-HBc and vaccination history. HBsAg and anti-HBs tests were done in 1,465 cases. The positive rates of HBsAg and anti-HBs were 4.5% and 74.6%, respectively. Anti-HBs positive rate was higher in the vaccinated group than that in the non-vaccinated group. The rates of anti-HBs positive cases with lower titers (10-< 100 mIU/mL) were 31.9%, while cases with higher titers (> or = 100 mIU/mL) were 68.1%. This suggested about 70% of anti-HBs-positive Korean adults (about 53% of the general adult population) have long-lasting immunity against HBV infection and may not require booster doses of HBV vaccination for a long time. Anti-HBs titers in the vaccine-induced anti-HBs group were higher than those in the infection-acquired anti-HBs group. No statistical differences were noted between male and female or among age groups. 25.7% of the HBsAg (-)/anti-HBs (-) group showed anti-HBc positive and HBV-DNA was detected in 11.1% among HBsAg (-)/anti-HBs (-)/anti-HBcAb (+) cases. Further study about post vaccination anti-HBs titer decay in Korean should be performed to help cut vaccination costs.

PMID: 10731918

Scand J Infect Dis 2000;32(3):249-52

Levels of viraemia in subjects with serological markers of past or chronic hepatitis B virus infection.

Noborg U, Gusdal A, Horal P, Lindh M.

Department of Clinical Virology, Goteborg University, Gothenburg, Sweden.

Subjects with serological markers for a past HBV infection may still have HBV DNA in their serum, but the levels of viraemia in such cases are not known. In the present study, of 63 consecutive HBsAg-negative, anti-HBc-positive serum samples with or without anti-HBs, 20 were HBV DNA-positive as analysed by a highly sensitive quantitative PCR, the Cobas Amplicor HBV Monitor test. However, all of these 20 samples had viraemia levels below 1000 copies/ml, compared with median viraemia levels of 10(8.6) and 10(4.3) copies/ml, respectively, in 98 HBeAg-positive and 124 HBeAg-negative HBsAg carriers. There was no difference in viraemia between subjects with anti-HBc alone compared with both anti-HBs and anti-HBc, nor between those with or without hepatitis C virus antibodies. The findings indicate that HBsAg-negative subjects may retain a low infectivity. Their risk for progressive liver damage is probably low, but this deserves further study.

PMID: 10879593

Transplantation 1998 Sep 15;66(5):616-9

Reverse seroconversion of hepatitis B after allogeneic bone marrow transplantation: a retrospective study of 37 patients with pretransplant anti-HBs and anti-HBc.

Dhedin N, Douvin C, Kuentz M, Saint Marc MF, Reman O, Rieux C, Bernaudin F, Norol F, Cordonnier C, Bobin D, Metreau JM, Vernant JP.

Bone Marrow Transplant Unit, Henri Mondor Hospital, Creteil, France.

BACKGROUND: Reverse seroconversion to hepatitis B virus (HBV), i.e., HBV reactivation in patients with pretransplant antibodies to hepatitis B surface antigen (anti-HBs) and to hepatitis B core antigen (anti-HBc), is rarely re-ported after allogeneic bone marrow transplantation. METHODS: To determine this risk, we studied clinical outcome and serological changes in 37 patients with pretransplant anti-HBs and anti-HBc. RESULTS: In 33 cases, no change in HBV markers was observed in the posttransplant period. In four cases, anti-HBs and anti-HBc were lost, and hepatitis B surface antigen, hepatitis B e antigen, and HBV DNA emerged together with acute hepatitis, after cessation of immunosuppression. The actuarial risk of reactivation in the 37 patients was 20.5% (median follow-up 20 months). No reactivation occurred in patients with anti-HBs-positive donors. CONCLUSION: Although few cases of postallogeneic bone marrow transplantation reverse seroconversion to HBV have been reported, this study demonstrates that the actuarial risk is relatively high and suggests that donor vaccination might be proposed prophylactically or that HBs-specific immunoglobulin infusions might be warranted.

PMID: 9753342

Eur J Haematol 2000 Jul;65(1):86-7

Reactivation of hepatitis B virus infection in an anti-HBc and anti-HBs positive patient after allogeneic bone marrow transplantation.

Nordbo SA, Skaug K, Holter E, Waage A, Brinch L. Publication Types: Letter

PMID: 10914949

Transfusion 2000 Aug;40(8):910-6

Hepatitis B virions isolated with antibodies to the pre-S1 domain reveal occult viremia by PCR in Alaska Native HBV carriers who have seroconverted.

Gandhi MJ, Yang GG, McMahon BJ, Vyas GN.

Department of Laboratory Medicine and the Liver Center, University of California, San Francisco, California 94143, USA.

BACKGROUND: Occult viremia occurring before the appearance of HBsAg or after the disappearance of HBsAg is detectable by gene amplification technologies whose efficiency depends on nucleic acid preparation. STUDY DESIGN AND METHODS: To isolate HBV DNA from viremic plasma, immunoaffinity capture (IAC) of intact HBV with biotinylated pre-S1 antibodies coupled to streptavidin-coated magnetic beads was evaluated. IAC was compared with a silica-gel method (Qiagen [QSG]) and its two modifications wherein the samples were heated with lysis buffer at 60(o)C for 10 minutes (QSG-60) or at 58 degrees C for 60 minutes with proteinase-K (QSG-PK). Each HBV DNA sample was tested by heminested PCR amplification of the HBV gene sequences. A total of 36 coded serum samples were tested, including three HBsAg-positive controls and 33 former chronic HBV carriers who had seroconverted (developed antibody to HBsAg [anti-HBs]). Commercially available seroconversion panels (PHM 907, 911, and 922) were similarly tested for window-period viremia. RESULTS: In the 33 former chronic HBV carriers who had seroconverted, IAC revealed HBV DNA in 17 samples, whereas it was revealed in only 11 samples by QSG-PK (p = 0.031), 10 by QSG-60 (p = 0.016), and 9 by QSG (p = 0.0078). However, HBV DNA was not amplified from the 17 samples at 1-in-10 dilutions; thus, they were considered to have low-level viremia. IAC revealed HBV DNA as early as or earlier than the other methods in PHM 907, 911, and 922 panels. CONCLUSION: IAC is apparently an optimal method of sample preparation for amplification of HBV DNA in patients in the pre-HBsAg window period, and for detecting low-level viremia persistent in several individuals who were former chronic HBV carriers who had seroconverted (developed anti-HBs).

PMID: 10960516

Liver 2000 Oct;20(5):411-4

De novo acute hepatitis B infection in a previously vaccinated liver transplant recipient due to a strain of HBV with a Met 133 Thr mutation in the "a" determinant.

Yoshida EM, Ramji A, Erb SR, Davis JE, Steinbrecher UP, Sherlock CH, Scudamore CH, Chung SW, Williams M, Gutfreund KS.

British Columbia Transplant Society and the Department of Medicine, University of British Columbia, Vancouver, Canada.

De novo HBV infection post-liver transplantation (LT) from an anti-HBc seropositive donor rarely presents as acute failure. We report a 42-year-old Caucasian female, HBsAg and anti-HBc seronegative, with primary biliary cirrhosis who received an allograft from a HBsAg negative, anti-HBc seropositive donor. The patient, previously vaccinated years pre-LT, was re-vaccinated against HBV and 1 year post-LT had an anti-HBs titre of 256 IU/l. Two years post-LT, elevated serum aminotransferases and worsening liver function with an INR of 2.0 developed. The HBsAg became positive, anti-HBs undetectable and serum HBV-DNA >2000 pg/ml by hybridisation assay. Liver biopsy revealed significant ballooning degeneration, piecemeal necrosis and positive immunostaining for HBsAg. Progressive liver failure developed followed by sepsis and terminal multi-organ failure. Subsequent analysis of the predominant HBV strain revealed mutations in the "a" determinant: Met 133 Thr (codon change ATG to ACG) and Asn 131 Thr. CONCLUSION:' Acute de novo HBV infection from an anti-HBc sero-positive donor may occur long after LT despite protective anti-HBs titres post-vaccination secondary to the emergence of "a" determinant mutated strains of HBV.

PMID: 11092260

J Infect 2000 Nov;41(3):260-4

Intra-familial evidence of horizontal transmission of hepatitis B virus surface antigen mutant G145R.

Oon CJ, Chen WN, Goo KS, Goh KT.

Ransome Research Laboratory, Singapore General Hospital, Republic of Singapore.

OBJECTIVES: To provide intra-familial evidence on the horizontal transmission of hepatitis B virus surface antigen (HBsAg) mutant G145R. METHODS: Serum samples from family members of 10 vaccinated infants who carried this G145R mutant were collected. The presence of the mutant was analysed by polymerase chain reaction (PCR) and sequencing. RESULTS: The G145R mutant was identified in family members of three of the 10 infants. In family 1, the mutant found initially in child 1 was identified in another child and the father. In families 2 and 3, the G145R mutant detected previously in child 1 was detected in the father. Additional mutations in HBsAg were identified in at least two members in family 1 and 2, suggesting horizontal transmission of the mutant among them. The G145R mutant was found in samples with high levels of neutralizing antibody against HBV (anti-HBs). In addition, liver damage was seen in one G145R carrier infant. CONCLUSIONS: The G145R mutant could be transmitted horizontally among family members, and this could occur in the presence of high levels of anti-HBs. Improvement of detection system for the G145R and other HBsAg mutant will be needed for their effective control. Copyright 2000 The British Infection Society.

PMID: 11120616

J Hepatol 2000 Dec;33(6):992-7

Occult hepatitis B virus after acute self-limited infection persisting for 30 years without sequence variation.

Blackberg J, Kidd-Ljunggren K.

Department of Infectious Diseases, Lund University, Sweden.

BACKGROUND/AIMS: After acute self-limited hepatitis B virus (HBV) infection, serological loss of viral antigens and appearance of anti-HBs is generally believed to signify viral clearance. Latent and occult HBV infection appearing decades after self-limited hepatitis B has not been reported, nor has the evolutionary rate of HBV DNA over the same observation period. METHODS: DNA from serum and leukocytes from 16 patients with acute self-limited hepatitis B 30 years earlier was tested by polymerase chain reaction and positive samples were sequenced. Liver tissue from four patients was also tested. Additionally, another 10 HBV strains isolated from acute HBV cases in 1969-72 were compared to 11 strains isolated from acute cases in 1998-99 in the same community. RESULTS: HBV DNA was detected in liver from two patients, but not in serum or leukocytes. The HBV strains detected in liver showed complete homology, in the sequences analyzed, to the strains originally infecting these patients. Ten strains from 1998-99 were identical in pre-S and core promoter/precore regions to strains from the same community isolated 30 years earlier. CONCLUSIONS: HBV can persist as an occult infection three decades after acute, apparently self-limited hepatitis B, and both the mutation and evolutionary rates of HBV DNA are low.

PMID: 11131464

Med Clin (Barc) 2001 Feb 3;116(4):125-8

[Risk of hepatitis B virus transmission from hepatitis B core antibody-positive liver donors] [Article in Spanish]

Barcena Marugan R, Garcia Garzon S, Lopez San Roman A, Pena Gonzalez E, Nasha R, Fernandez Munoz R, Mateos M, Garcia Plaza A.

Gastroenterologia, Hospital Ramon y Cajal, Madrid.

BACKGROUND: To study the hepatitis B virus (HBV) transmission from donors HBsAg-/AntiHBc+ to liver transplant recipients. PATIENTS AND METHOD: We studied retrospectively the HBV serological markers in 43 donors from our center and also the serological condition of the 41 recipients. The HBV serological markers were analyzed by ELISA and HBV DNA was detected by hybridation assays. RESULTS: 13 donors samples showed some HBV serological markers: 6 anti-HBc and anti- HBs (13.9%), 4 anti-HBc (9%) and 3 anti- HBs (6.9%). There were no cases of hepatitis B among liver recipients from donors with negative serological markers. Among the 13 recipients with HBV serological markers, 9 were followed during 39 (SD 17) months. The 5 recipients with no HBV markers, who received an anti- HBc+ with or without anti- HBs (100%) developed hepatitis B. The two liver recipients with anti-HBs solely, did not developed infection (0%). Of the 41 recipients, 15 had some HBV markers before transplant and two of them received an anti-HBc+ and did not develop the infection (0%). CONCLUSIONS: In our study, the prevalence of serological HBV infection in donors and recipients was of 30.2 and 31.7%, respectively. Anti-HBc with or without anti-HBs donors transmitted the HBV infection in all the cases (100%) to the susceptible recipients. The presence of anti-HBs in recipients protected these against the infection. Only the anti-HBs positive donors did not trasmit the HBV infection.

PMID: 11222157

Transfusion 2001 Mar;41(3):329-32

Infectivity of blood from PCR-positive, HBsAg-negative, anti-HBs-positive cases of resolved hepatitis B infection.

Prince AM, Lee DH, Brotman B.

Laboratory of Virology, the Lindsley F. Kimball Research Institute of The New York Blood Center, New York, New York 10021, USA. aprince@nybc.org

BACKGROUND: Numerous reports have noted the existence of sera, particularly from resolving cases of HBV infection, that are positive for HBV DNA by PCR, despite being negative for HBsAg and IgM anti-HBc. If such blood is infective and detectable by HBV NAT screening, it seems desirable to introduce such screening for transfused blood. STUDY DESIGN AND METHODS: Three chimpanzees were inoculated with serum and lymphocytes from three patients who were HBV DNA PCR positive, but HBsAg negative. The animals were tested over a period of 15 months for HBsAg, anti-HBs, anti-HBc, and HBV DNA by PCR. RESULTS: All animals remained uninfected. CONCLUSION: Small amounts of plasma and MNCs from HBV DNA-positive HBsAg-negative blood do not appear to be infectious; however, further studies with larger volumes of inoculum should be conducted.

PMID: 11274585

J Biomed Sci 2001 May-Jun;8(3):237-47

Vaccine- and hepatitis B immune globulin-induced escape mutations of hepatitis B virus surface antigen.

Cooreman MP, Leroux-Roels G, Paulij WP.

Department of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands. m.cooreman@amc.uva.nl

Hepatitis B virus surface antigen (HBsAg) vaccination has been shown to be effective in preventing hepatitis B virus (HBV) infection. The protection is based on the induction of anti-HBs antibodies against a major cluster of antigenic epitopes of HBsAg, defined as the 'a' determinant region of small HBsAg. Prophylaxis of recurrent HBV infection in patients who have undergone liver transplantation for hepatitis B-related end-stage liver disease is achieved by the administration of hepatitis B immune globulins (HBIg) derived from HBsAg-vaccinated subjects. The anti-HBs-mediated immune pressure on HBV, however, seems to go along with the emergence and/or selection of immune escape HBV mutants that enable viral persistence in spite of adequate antibody titers. These HBsAg escape mutants harbor single or double point mutations that may significantly alter the immunological characteristics of HBsAg. Most escape mutations that influence HBsAg recognition by anti-HBs antibodies are located in the second 'a' determinant loop. Notably, HBsAg with an arginine replacement for glycine at amino acid 145 is considered the quintessential immune escape mutant because it has been isolated consistently in clinical samples of HBIg-treated individuals and vaccinated infants of chronically infected mothers. Direct binding studies with monoclonal antibodies demonstrated a more dramatic impact of this mutation on anti-HBs antibody recognition, compared with other point mutations in this antigenic domain. The clinical and epidemiological significance of these emerging HBsAg mutants will be a matter of research for years to come, especially as data available so far document that these mutants are viable and infectious strains. Strategies for vaccination programs and posttransplantation prophylaxis of recurrent hepatitis need to be developed that may prevent immune escape mutant HBV from spreading and to prevent these strains from becoming dominant during the next decennia. Copyright 2001 National Science Council, ROC and S. Karger AG, Basel Publication Types: Review Review, Tutorial

PMID: 11385295

J Med Virol 2001 Jul;64(3):312-9

Hepatitis B virus markers in anti-HBc only positive individuals.

Weber B, Melchior W, Gehrke R, Doerr HW, Berger A, Rabenau H.

Laboratoires Reunis Kutter-Lieners-Hastert, Junglinster, Luxembourg. laborklh@pt.lu

Isolated reactivity to hepatitis B virus (HBV) core antigen (anti-HBc) is observed relatively frequently in immunocompromised individuals, intravenous drug abusers (IVDA), and in the presence of HCV infection. The reason for the lack of HBsAg is not clear. The aim of the present study was to investigate which factors (genetic variability of S gene, low-level HBsAg, and immune complexes may be responsible for the failure of HBsAg detection with commercial HBsAg screening assays. Dilution series of two recombinant HBsAg escape mutants and dilutions of serum samples from chronic HBV carriers with multiple insertions in the a determinant and different HBsAg subtypes were tested with a highly sensitive assay that detects wild-type HBsAg (Elecsys HBsAg, Roche Diagnostics, Penzberg, Germany) and two assays that detect HBV wild-type and escape mutants (Murex HBsAg Version 3, Murex and Enzygnost HBsAg 5.0, Dade Behring, Marburg, Germany). Elecsys HBsAg showed in comparison to Murex HBsAg Version 3 and Enzygnost HBsAg 5.0 a reduced sensitivity for escape mutant detection. On the other hand, the best performance for HBsAg subtype detection was obtained with Elecsys HBsAg. In the second part of the study, a selected panel of isolated anti-HBc reactive (n = 104) serum samples (AxSYM Core) was submitted to testing by Elecsys HBsAg, Murex HBsAg Version 3, Enzygnost HBsAg 5.0, and HBsAg detection after immune complex dissociation (ICD) and anti-HBs determination with two different assays (AxSYM Ausab and Elecsys Anti-HBs). To assess the specificity of anti-HBc test results, all the samples were tested by a second anti-HBc assay (Elecsys Anti-HBc). Quantitative HBV DNA detection was undertaken with a commercially available HBV PCR assay (Amplicor HBV Monitor). HCV infection was present in 65.4% of anti-HBc only reactive individuals. Five AxSYM Core positive samples were negative by Elecsys Anti-HBc. Overall, 15 (14.4%) AxSYM Ausab negative samples gave positive results with Elecsys Anti-HBs (median value: 21 IU/ml). No low-level HBsAg carrier was detected among the isolated anti-HBc reactive individuals with Elecsys HBsAg. There was no evidence for the presence of immune complexes. Only one sample was repeatedly reactive by the Murex HBsAg, suggesting that the a mutant form of HBsAg was responsible for the isolated anti-HBc reactivity, however neutralisation assay was not interpretable and HBV DNA PCR was negative. Fifteen (14.4%) anti-HBc only positive individuals were HBV DNA carriers with concentrations ranging from 800 to more than >4,000,000 copies of viral DNA/ml. In conclusion, the most probable explanations for isolated anti-HBc reactivity in our study group are a possible interference of HBsAg synthesis by HCV infection (65.4%) and divergence of results of anti-HBs assays (14.4%). There is no evidence for the presence of low-level HBsAg carriers and immune complexes. HBsAg mutants cannot be excluded definitively by the test strategy used in the present evaluation. Copyright 2001 Wiley-Liss, Inc. Publication Types: Evaluation Studies

PMID: 11424120

J Med Virol 2001 Nov;65(3):493-504

Residual hepatitis B virus particles in liver transplant recipients receiving lamivudine: PCR quantitation of HBV DNA and ELISA of preS1 antigen.

Petit MA, Buffello-Le Guillou D, Roche B, Dussaix E, Duclos-Vallee JC, Feray C, Samuel D.

INSERM E99-41 and UPRES 1596, Centre Hepato-Biliaire, Villejuif, France. petit@lyon151.inserm.fr

Lamivudine, an antiviral agent, has a potential role in the treatment of recurrent or acquired de novo hepatitis B virus (HBV) infection after liver transplantation. During lamivudine therapy, residual HBV particles in serum, PBMC, and liver were quantified in 7 patients in whom hepatitis B occurred de novo (n = 4) or recurred (n = 3). HBV DNA and preS1 antigen were measured using a sensitive PCR technique and an in-house ELISA method, respectively. The genetic and antigenic properties of HBV variants that emerged during lamivudine treatment were also examined. One month after the outset of lamivudine treatment, all 7 patients remained positive for both HBV DNA and preS1 antigen in serum, reflecting residual HBV replication. At the end of therapy, four patients were considered to be lamivudine responders, including one who seroconverted to anti-HBs but remained HBV DNA positive in the liver (> 10(3) copies/microg of DNA). Among the three patients who did not respond to lamivudine, one had pol mutations (L450P and S550C) that had not been described previously, in addition to the common mutations within the YMDD locus and B domain. Defective core and preS viral proteins and atypical sedimentation profiles of HBV DNA-positive particles were observed in all three lamivudine-resistant patients. These findings confirm the persistence of HBV in liver transplant recipients despite strong inhibition of replication by lamivudine, and show abnormal viral transcription and/or morphogenesis in lamivudine-resistant patients. Copyright 2001 Wiley-Liss, Inc.

PMID: 11596084