Detection of hepatitis B surface antigen in pregnant women attending a public hospital for delivery: implication for vaccination strategy in Bangladesh

Routine antenatal hepatitis B surface antigen (HBsAg) screening and immunization of risk babies is very effective in preventing perinatal transmission of hepatitis B virus (HBV). We studied 1,800 parturients attending a public hospital to assess the rationale for such vaccination in Bangladesh. In one in every 29 deliveries (63 of 1,800 or 3.5%), the mother was found to be HBsAg positive. All were asymptomatic and many (41 of 63 or 65%) without risk factors would remain undetected if HBsAg screening were performed on selected groups. Most of the HBsAg-positive mothers (54 of 63 or 85.7%) were found to be chronic carriers and 30.2% (19 of 63) were also hepatitis B e antigen (HBeAg) positive, indicating high infectivity. Although 23 cord blood were positive for HBsAg or HBeAg, none were positive for IgM antibody to hepatitis B core antigen (IgM anti-HBc), suggesting transplacental transmission of the antigens rather than intrauterine infection. These findings are discussed in relation to the cost-effectiveness of routine prenatal screening and immunization of risk babies compared with universal infant immunization.     

Hepatitis B virus infection in dental surgical practice

Sixty-one dental surgeons at King's College Hospital were interviewed to establish the incidence of attacks of viral hepatitis and to relate this to environmental risk factors. Six (10%) had a history of hepatitis, in one case due to infection with the hepatitis B virus. Screening blood for HBsAg by radioimmunoassay showed no carriers of the antigen, but transient antigenaemia was observed in one dentist. Antibody to HBsAg, tested by radioimmunoassay, was detected in four dentists (7%), only one of whom had had clinical hepatitis. Dental surgeons may be more at risk from infection with the hepatitis B virus than the general population, although this should be minimised in hospital practice, where the most infected patients will already have been identified and appropriate precautions can be taken. The risk of transmission from an antigen-positive dentist to his patients is probably much smaller, and there is no evidence to restrict his clinical activities.     

Differing characteristics of hepatitis B and C risk factors among elders in a rural area in Taiwan

BACKGROUND: Both hepatitis B and C are major health concerns in Taiwan. The goal of this study was to determine how risk factors for hepatitis B and C differed in this study population. It was also hoped that the data might help determine how age and place of residence affect hepatitis risk factors. METHODS: Complete serum and hepatitis marker analysis (HBsAg and AntiHCV) was done for 282 individuals over 65 years old. Of these, 254 were interviewed for risk factor analysis. RESULTS: Of the 282 subjects, 8.2% were HBsAg+, 27.3% were AntiHCV+, and 3.2% were both HBsAg+ and AntiHCV+. AntiHCV+ subjects were more likely than AntiHCV-subjects to have had frequent medical injections, odds ratio (OR) = 2.94, 95% confidence interval (CI) (1.68, 5.12), and it was the only independent risk factor for determining AntiHCV+, OR = 3.26, 95% CI (1.85, 6.11) (N = 254). The AntiHCV+ group had higher alanine and asparate aminotransferase levels but lower cholesterol and triglyceride levels than AntiHCV- and HBsAg+ groups (p < .0001). Abnormal ALT existed in 40.3% of AntiHCV+ and 10.7% of AntiHCV- cases. ALT was associated with AntiHCV and sex, although abnormal AST was only associated with AntiHCV. CONCLUSIONS: AntiHCV was closely related with frequent medical injections and was the primary risk factor for abnormal ALT and AST levels in this study population. It appears that frequent medical injections are an important risk factor because of the previously common habit of reusing syringes. This is of major concern to elders in Taiwan because of their much greater likelihood of repeated exposure.     

Prevalence and risk factors for hepatitis B virus infections among visitors to an STD clinic

OBJECTIVE: To determine the prevalence and risk factors for hepatitis B virus (HBV) infections among individuals attending an STD clinic in a low endemic region. STUDY DESIGN: A total of 1228 women and 1648 men attending the STD clinic at the University Hospital Rotterdam, Netherlands, were examined for HBV infection by determination of hepatitis B surface antigen (HBsAg) and antibodies to hepatitis B core antigen (anti-HBc). Demographic characteristics, information on sexual behaviour, and intravenous drug use were recorded. RESULTS: The seroprevalence of HBsAg was 1.4% in women and 2.1% in men (0% in homosexual men). The seroprevalence of anti-HBc was 13% in women and 20% in men (36% in homosexual men). Native country, intravenous drug use, a history of STD, and the number of partners in the past half year (inversely) were independent risk factors for HBsAg positivity in women and heterosexual men. For anti-HBc independent associations were observed for native country, age, intravenous drug use, commercial sex, number of lifetime partners, homosexual contacts, orogenital contact (inverse), and a history of STD. CONCLUSION: The HBV prevalence in the STD clinic attendants was high, exceeding the national estimate, and indicates that the STD clinic population may be considered a high risk group. Our data confirmed an increased risk for HBV infections among established risk groups. Therefore, these risk groups should be routinely screened to identify HBV cases for counselling and contact tracing.     

The complex forensic medical study of the bony remains from a burial long ago]

A cross-sectional study was conducted in a randomly selected village of Rangpur district from June '94 to May '95. Serum from 1000 human subjects irrespective of age and sex were screened for hepatitis B surface antigen (HBsAg). Among the respondents 661 were male and 339 female. The overall seroprevalence of HBsAg was 6.4%. It was 6.66% in male and 5.89% in female. Age, sex, religion, income, occupation, education and marital status did not show any relationship with HBsAg status. Among 64 sero-positive cases, 16 (25%) had no history of exposure to known risk factors and 48 (75%) had one or more exposure to known risk factors (p < 0.05). This study did not depict the national scenario. Well designed studies with more sensitive serological methods are recommended to get the epidemiological information of hepatitis B virus (HBV) infection in our community.     

Differences in risk factors for being either a hepatitis B carrier or anti-hepatitis C+ in a hepatoma-hyperendemic area in rural Taiwan

This is a study of the differences in the risk factors for being either hepatitis B surface antigen positive [HBsAg(+)] or antibody to hepatitis C virus positive [Anti-HCV(+)] in A-Lein, a rural area in southern Taiwan, an area which also has a high hepatoma mortality rate. Three hundred eighty-five patients age > or =40 years participated in hepatoma screening at the A-Lein Community Health Center during 1995. Those who were HBsAg(-) and anti-HCV(-) or had coinfection of HBsAg(+) and anti-HCV(+) were excluded, leaving 293 patients: 109 HBsAg(+) and 184 anti-HCV(+). The anti-HCV(+) patients had a lower socioeconomic status (as defined by level of education and type of occupation) and were older than HBsAg(+) patients (P < 0.05). Those with higher alanine aminotransferase levels (ALT) also had a higher anti-HCV(+) to HBsAg(+) odds ratio (OR), and a dose response relationship was found, P < 0.0001. Anti-HCV(+) patients were more likely than HBsAg(+) patients to have a spouse who shared the infection, OR = 5.11; 95% CI, 2.30-11.28. Anti-HCV(+) patients were more likely than HBsAg(+) patients to have had blood transfusions (OR = 2.66; 95% CI, 1.20-5.89), frequent medical injections (OR = 2.64; 95% CI, 1.62-4.31), or injections by non-licensed medical providers (OR = 1.91; 95% CI, 1.18-3.09). Multiple logistic regression analysis showed that the significant factors for anti-HCV(+) patients vs. HBsAg(+) patients are drinking habit (OR = 3.45; 95% CI, 1.02-11.60), age (OR = 6.33; 95% CI, 2.93-13.68), and frequent medical injections (OR = 2.88; 95% CI, 1.65-5.03). The transmission of hepatitis C in A-Lein is closely related to low socioeconomic status, age, alcohol abuse, spouses being anti-HCV(+), and frequent medical injections, especially from non-licensed medical providers, including both pharmacists and those with no medical licensing whatsoever. These nonlicensed medical providers sometimes reuse needles to save money, which is a likely route of infection.     

Factors associated with intrafamilial transmission of hepatitis B virus infection in Korea

Epidemiologic and serologic data on 137 household contacts of 51 chronic carriers of HBsAg and 111 household contacts of 38 controls who were negative for serologic markers of hepatitis B virus (HBV) were obtained from March 1990 to August 1991. Using this data, possible routes of intrafamilial transmission of hepatitis B virus among household contacts of chronic carriers of hepatitis B surface antigen (HBsAg) were evaluated and analyzed. The HBsAg prevalence among the household contacts of carriers was 14. 1% (95% CI 7.8-24.0) compared to 0.0% (95% CI 0.0-7.0) among those of controls (P < 0.01). The offspring of carriers showed significantly higher risk of HBV infection(relative risk; 6.6). Sharing of towels and handkerchieves, and drinking vessels was associated with an increased risk of HBV infection via intrafamilial transmission in Korea (relative risk 11.5 for towel and handkerchief, 12.1 for drinking vessels).     

Prevalence of hepatitis B and C viruses in healthy Indonesian blood donors

Blood samples were collected from 7572 healthy volunteer blood donors from 21 of the 27 Indonesian provinces, and tested for antibodies to hepatitis C virus (anti-HCV) using the new second-generation enzyme immunosorbent assay, and also tested for hepatitis B surface antigen (HBsAg). We detected anti-HCV in 2.1% of the blood donors. No statistically significant difference was found between males and females or between locations, but there was a statistically significant increasing likelihood of anti-HCV prevalence with increasing age. HBsAg was found in 8.8% of the 3839 tested donors. There was no statistically significant difference between sexes or age groups, but there was a statistically significant higher prevalence in the islands of Sulawesi and eastern Indonesia. Only 7 individuals, from 5 locations, were both anti-HCV and HBsAg positive. Based on responses to a questionnaire, a history of surgery, blood transfusion, intravenous medication, and acupuncture were identified as risk factors for the presence of anti-HCV. No such risk factor was identified for HBsAg prevalence. The combined data suggest separate modes of transmission for the 2 viruses, and indicate the need for continued surveillance for these agents in Indonesian blood banks.     

Emergence of an S gene mutant during thymosin alpha1 therapy in a patient with chronic hepatitis B

The presence of a hepatitis B virus S gene mutant was investigated in a patient being treated with thymosin alpha1. He was seropositive for hepatitis B e antigen throughout therapy but was intermittently seronegative for hepatitis B surface antigen (HBsAg) by an RIA. Sequence analysis revealed an S gene mutant in HBsAg-seronegative serum with two consecutive amino acid substitutions: threonine115-to-isoleucine and threonine116-to-asparagine, whereas no amino acid substitution or deletion was found in the pre-S region. A site-directed mutagenesis experiment confirmed that these mutations were responsible for the failure to detect HBsAg. In summary, an S gene mutant was identified in an HBsAg-seronegative patient. The mutations were located outside the putative "a" determinant. The emergence of an S gene mutant during thymosin alpha1 treatment suggests that enhanced host immunity against HBsAg may play a role in its antiviral activity.     

Long-term hepatitis B vaccine in infants born to hepatitis B e antigen positive mothers

Neonates of hepatitis B surface antigen (HBsAg) positive and hepatitis B encoded antigen (HBeAg) positive mothers received 10 micrograms of recombinant hepatitis B vaccine at months 0, 1, 6, or 0, 1, 2, 12, with or without immunoglobulin at birth, and were followed up to the age of 8 years for HBsAg, anti-HBc, and anti-HBs. Some were boosted at month 60. The overall vaccine protection at month 12 was 96.2%. No child became a chronic carrier beyond the age of 3 years, showing that this vaccine provides immediate protection against HBsAg carriage, and long term protection against fetally acquired HBsAg. After month 60 hepatitis B serological markers without disease, indicating re-exposure to HBV, reappeared in comparable numbers among boosted and non-boosted children (5 for a total of 167 children). This vaccine provides long-term protection against hepatitis B chronic carriage and infection in high risk neonates with or without a month 60 booster. A booster at the age of 5-6 years or 11-12 years would reduce HBV infection, viral circulation and transmission, while ensuring long-term antibody persistence.     

Relation between laboratory test results and histological hepatitis activity in individuals positive for hepatitis B surface antigen and antibodies to hepatitis B e antigen

BACKGROUND: Hepatitis B surface antigen (HBsAg) and antibodies to hepatitis B e antigen (anti-HBe) commonly coexist, and laboratory tests are often requested to assess histological hepatitis activity. An optimum panel of tests has not been found and the usefulness of hepatitis B virus (HBV) DNA assays in this context has not been established. We assessed various blood tests to find which best predicted hepatitis activity. METHODS: Routine plasma biochemical liver tests and serum HBV DNA (hybridisation and PCR assays) were assessed prospectively in 123 patients positive for HBsAg and anti-HBe. We scored histological hepatitis activity (hepatitis activity index) and determined whether chronic active hepatitis (chronic hepatitis with portal and periportal lesions) was present. We analysed the relation between laboratory data and the hepatitis activity index or risk of chronic active hepatitis by multiple regression and multiple logistic regression, respectively. FINDINGS: The analyses provided models for predicting either the hepatitis activity index or the risk of chronic active hepatitis. Aspartate aminotransferase was the most important test in the two models. The contribution of HBV DNA and other assays, especially alanine-aminotransferase activity, were of no practical importance. INTERPRETATION: Because screening by aspartate-aminotransferase activity could not be improved by the addition of other assays or HBV DNA, patients positive for HBsAg and anti-HBe could be screened for chronic active hepatitis with a single assay and counselling of patients can be improved if proper reference values are used.     

Hepatitis B virus transmission in an elementary school setting

CONTEXT: The risk of transmission of hepatitis B virus (HBV) in day care centers and schools is low. OBJECTIVE: To investigate the source of HBV transmission for an elementary school teacher with acute hepatitis B. DESIGN: Serologic survey for HBV infection among elementary school students, school staff, and household members of an HBV-infected teacher and student. SETTING: General community and elementary school. PATIENTS: Elementary school students and staff members and household members of an HBV-infected teacher. MAIN OUTCOME MEASURES: Elementary school students, school staff, and household members of an HBV-infected teacher were tested for markers of HBV infection. Samples positive for hepatitis B surface antigen (HBsAg) were tested for HBsAg subtype using monoclonal antibodies and examined for HBV DNA homology by polymerase chain reaction techniques. RESULTS: An HBV-infected student and the teacher were found to have the same HBV subtype (ayw1-2) and to have identical HBV DNA sequences. The teacher reported none of the usual risk factors for acquiring HBV infection, and none of her family members had been infected prior to her illness. The specific means of HBV transmission from student to teacher was not identified. Of 108 total children in the same grade as the HBV-infected student, 102 (94%) were tested for serologic markers of HBV infection, and none was positive. CONCLUSIONS: This investigation documented transmission from an HBV-infected student to a teacher in an elementary school setting without a reported overt percutaneous or permucosal exposure to blood or infectious body fluids. Transmission of HBV to other students or staff members in the school was not observed.     

Risk factors for horizontal transmission of hepatitis B virus in a rural district in Ghana

Most hepatitis B virus (HBV) infections in sub-Saharan African infants and children are acquired through horizontal transmission, but the exact mechanisms of spread have not been documented. The authors conducted a study in rural Ghana which determined seroprevalence in a probability sample of 1,385 individuals of all ages, and evaluated risk factors for horizontal transmission of HBV in a subsample of 547 children aged 1-16 years who were not hepatitis B surface antigen (HBsAg) carriers. Most residents in this district live in compounds which typically contain 2-4 households each. Overall prevalence of HBV seropositives (any HBV marker) was 74.7% (95% confidence interval (CI) 72.5%-76.9%). Prevalence of HBsAg was 20.9% (95% CI 18.8%-23.1%). The data suggest a continuous nonuniform acquisition of HBV infection with advancing age predominantly through horizontal transmission in childhood, with the household, rather than the domestic compound, being the primary place for transmission. The behaviors most strongly associated with prevalence of HBV were sharing of bath towels (OR = 3.1, 95% CI 2.1-4.5), sharing of chewing gum or partially eaten candies (OR = 3.4, 95% CI 2.3-5.0), sharing of dental cleaning materials (OR = 2.5, 95% CI 1.3-4.6), and biting of fingernails in conjunction with scratching the backs of carriers (OR = 2.5, 95% CI 1.6-4.3).     

Enhanced HBsAb production in pathogenesis of fulminant viral hepatitis type B

The possible importance of humoral immunity in the pathogenesis of fulminant hepatitis was investigated by comparing 17 patients with fulminant hepatitis type B with 20 patients with severe but non-fulminant disease. Hepatitis B surface antigen (HBsAg) was cleared from the serum significantly faster (P less than 0-001) in those with fulminant hepatitis, and in 41% anti-HBsAg (HBsAb) was detectable by radioimmunoassay (RIA) at presentation. In all 11 sera from patients with fulminant hepatitis that were examined by electron microscopy aggregates of HBsAg and HBsAb were seen. In contrast, HBsAb was never detected by RIA in those with non-fulminant hepatitis, and in only one serum specimen (5%) were aggregates seen on electron microscopy. A significant sex difference between fulminant and non-fulminant hepatitis was observed, 65% of patients with fulminant hepatitis but only 15% of patients with non-fulminant hepatitis being women (P less than 0-01). An enhanced production of HBsAb in fulminant hepatitis, by leading to free HBsAb in portal blood, may cause an Arthus reaction in the sinusoids of the liver with ensuing ischaemic necrosis of hepatocytes.     

Processing of exogenous hepatitis B surface antigen particles for Ld-restricted epitope presentation depends on exogenous beta2-microglobulin

Processing of exogenous hepatitis B surface antigen (HBsAg) particles in an endolysosomal compartment generates peptides that bind to the major histocompatibility complex (MHC) class I molecule Ld and are presented to CD8+ cytotoxic T lymphocytes. Surface-associated 'empty' MHC class I molecules associated neither with peptide, nor with beta2-microglobulin (beta2m) are involved in this alternative processing pathway of exogenous antigen for MHC class I-restricted peptide presentation. Here, we demonstrate that internalization of exogenous beta2m is required for endolysosomal generation of presentation-competent, trimeric Ld molecules in cells pulsed with exogenous HBsAg. These data point to a role of endocytosed exogenous beta2m in the endolysosomal assembly of MHC class I molecules that present peptides from endosomally processed, exogenous antigen.     

False-positive hepatitis B surface antigen screening test results in patients receiving granulocyte-colony-stimulating factor

BACKGROUND: Granulocyte-colony-stimulating factor (G-CSF) is used for the mobilization of progenitor cells and granulocytes. False-positive hepatitis B surface antigen (HBsAg) enzyme-linked immunosorbent assays (ELISAs) (NML) from one manufacturer in individuals receiving G-CSF have been observed. STUDY DESIGN AND METHODS: Sixty-six autologous peripheral blood progenitor cell donors from 1994 were retrospectively reviewed. Donors typically received 5 to 10 micrograms of G-CSF per kg subcutaneously for 5 days before collection. Additional ELISA dilutional studies (1-in-10, 1-in-100, 1-in-1000) with known HBsAg-negative serum were made with G-CSF. Testing was performed by the University of North Carolina, the American Red Cross in Charlotte, NC, or the National American Red Cross, Washington, DC. RESULTS: Of the 66 patients, none reacted for antibody to hepatitis B core antigen, and 30 (45%) had a positive reaction in the ELISA. Surface antigen positivity was "confirmed" on 6 of the 30 patients by neutralizing ELISA reactivity with an antibody to HBsAg test from the same manufacturer. In all cases, the clinical presentation was not suggestive of hepatitis, and these individuals were not at high risk for hepatitis B. Twenty-seven of the 30 cases were tested with a monoclonal HBsAg ELISA (AUSZYME) from another manufacturer in the peridonation period and did not react. In 1994, 256 autologous whole-blood donors not receiving G-CSF were similarly tested and only 1 (0.4%) had a positive reaction with the second manufacturer's HBsAg ELISA (p < 0.001). Of this group, 41 patients with histories of malignancy were identified, which is comparable to the history of the peripheral blood progenitor cell donors in this study, and none of these blood donors tested positive for HBsAg (p < 0.001). Dilutional studies with G-CSF produced no reactions. CONCLUSION: The NML HBsAg ELISA studied has an unacceptably high false-positive rate in patients or donors receiving G-CSF. The false reactivity of this assay appears to be an indirect consequence of G-CSF administration, which can also lead to spurious confirmation by the HBsAg neutralization assay from the same manufacturer.     

The prevalence of hepatitis C in patients admitted with acute hepatitis to Fairfield Infectious Diseases Hospital, 1971-1975

OBJECTIVE: To identify and determine trends in the prevalence of hepatitis C virus (HCV) antibody in stored sera from 1971 to 1975 and to determine associations with HCV seropositivity, including markers for other hepatitis infections and possible routes of transmission. DESIGN: A retrospective cross-sectional study. PATIENTS AND SETTING: 1511 adults admitted to Fairfield Infectious Diseases Hospital, Victoria, with a clinical and biochemical diagnosis of hepatitis between 1 January 1971 and 31 December 1975. MAIN OUTCOME MEASURES: Prevalence over study period of hepatitis A virus antibody (anti-HAV) IgM, hepatitis B core antibody (anti-HBc), hepatitis B surface antigen (HBsAg) and hepatitis C virus antibody (anti-HCV) in stored sera; sociodemographic data and risk factors for blood-borne viruses documented in original medical records. RESULTS: Anti-HCV was detected in 17% of adults admitted with hepatitis from 1971 through 1975. Prevalence increased significantly over this period. Most cases were in young men who had a history of injecting drug use. HCV seropositivity was also significantly associated with markers for hepatitis B infection. CONCLUSIONS: Given the 20-30-year period between infection with hepatitis and the development of liver disease, our findings predict significant liver-related morbidity in Australia in the next decade. The increase in prevalence over the five years studied suggests rapid spread of HCV through susceptible populations, principally injecting drug users.     

Lymphocytotoxins in leprosy and in asymptomatic hepatitis B virus infection

Serum lymphocytotoxic antibodies (LCAs) were detected in 67% of Papua New Guinean lepromatous leprosy patients who were persistent carriers of hepatitis B surface antigen (HBsAg). Lymphocytotoxins were not associated with asymptomatic HBsAg in either healthy controls or tuberculoid leprosy patients. It was apparent that, although HBsAg itself is a poor indicator of in vitro lymphocytotoxicity, when the antigen occurred in a host with impaired immune response, lymphocytotoxicity, when the antigen occurred in a host with impaired immune response, lymphocytotoxicity was enhanced. In contrast to this finding, lepromatous leprosy patients without HBsAg had significantly depressed LCA production in comparison with tuberculoid patients and controls. The interaction between leprosy and hepatitis B virus was highly significant (P = 0.001) in an analysis of variance of cytotoxicity scores. It is proposed that the previously reported equivocal results regarding autoantibodies in leprosy patients may be explained by this unusual interaction between lepromatous leprosy and hepatitis B virus infection.     

Bloodborne viral infection in Irish injecting drug users

AIMS: In Dublin, harm reduction strategies have greatly expanded since 1990. We sought to determine the prevalence of, and factors associated with, hepatitis C (HCV), hepatitis B (HBV) and HIV infections among injecting drug users (IDUs) against this background. DESIGN: Cross-sectional survey. SETTING: Addiction treatment clinic. PARTICIPANTS: Seven hundred and thirty-five IDU tested for antibody to HCV (anti-HCV) between September 1992 and September 1997. MEASUREMENTS: Socio-demographic and drug use characteristics. Serology tests for anti-HCV, HBV surface antigen (HBsAg) and HIV. FINDINGS: The vast majority (89%) commenced injecting since 1990. Prevalence of anti-HCV was 61.8% (453/733), of HBsAg was 1.0% (7/729) and of HIV was 1.2% (7/600). Logistic regression analyses indicated that longer history of injecting and increased daily drug expenditure were the only independent variables associated with significantly increased risk of HCV. The only characteristic associated with increased prevalence of HBsAg was a history of injecting prior to 1990 (3.8%, 3/80). HIV prevalence was significantly higher when aged over 24 years (3.7%, 6/162), when injecting commenced prior to 1990 (6.3%, 4/64) and when injecting over 5 years (6.5%, 4/62). CONCLUSIONS: HIV prevalence has sustained a low level in this population. Interventions which aim to halt transmission of HCV are necessary and will need to target IDU very early in their injecting careers and also those at risk of commencing to inject.     

Hepatitis B injury, male gender, aflatoxin, and p53 expression each contribute to hepatocarcinogenesis in transgenic mice

The major risk factors for human liver cancer: hepatitis B virus (HBV) related liver injury, male gender, aflatoxin exposure, and p53 expression, are evaluated and compared in experimental transgenic mouse models. Transgenic mice that express hepatitis B surface antigen (HBsAg) in their liver and develop liver tumors at 18 months of age (HBV+ mice) were bred to p53 null mice (p53-/-) to produce mice p53+/-, HBV+ mice. These mice and control littermates ([p53+/+, HBV+], [p53+/-, HBV-], and [p53+/+, HBV-) were divided into groups that did or did not receive an injection of aflatoxin at 1 week of age. At sacrifice at 13 months of age, 100% (7/7) of male mice with each of the three risk factors (p53+/-, HBV+, AFB1+) developed high-grade hepatocellular carcinomas (HCC). If any one of the risk factors was absent, the incidence drops: if both p53 alleles are present, 62% (10/16); if HBsAg is not expressed, 14% (1/7); if AFB1 is not given, 25% (2/8). If only one of the risk factors is present no tumors above grade I are found. Similar results were observed in female mice except that HCC incidence in each group is less than in male mice. Some of the tumors in mice with more than one risk factor are of unusual histological types, such as hepatocholangio-carcinomas, adenocarcinomas and undifferentiated carcinomas that are not usually seen in HBV transgenic C57BL/6 mice. No loss or mutation of the p53 gene is detected in any of the tumors. Possibilities of how the four major risk factors for HCC interact to produce malignant liver tumors in these transgenic mouse models of hepatocarcinogenesis are discussed.