Tim J. Harrison

The complete nucleotide sequence of the genome of a hepatitis B virus isolated from a naturally infected chimpanzee

The complete nucleotide sequence of a strain of hepatitis B virus, originally isolated from a naturally infected chimpanzee, has been determined. Interesting features of the sequence include the presence of an in-phase stop codon in the pre-core region of the core antigen open reading frame. The sequence shows approximately 10% nucleotide divergence from all of the other hepatitis B virus sequences previously published and the possibility that this divergence is the result of passage through chimpanzees is discussed.

Fermentation of cereals for reduction of bacterial contamination of weaning foods in Ghana.

Unfermented and fermented maize dough weaning foods prepared by mothers in a Ghanaian village were examined for gram-negative bacilli (GNB) immediately after preparation and during storage to assess the antimicrobial effect of fermentation. GNB were cultured from all samples of unfermented dough (51) and from 16 of 51 samples of fermented dough. The extent of contamination was significantly higher in the unfermented dough than in fermented dough (5.9 [SEM 0.1] vs 4.0 [0.4] log10 colony forming units/g). After 6 h and 12 h storage, a higher proportion of samples of porridge made from unfermented dough contained GNB than did those made with fermented dough (45/51 vs 22/55; 49/51 vs 20/51, respectively) and levels of GNB were significantly higher in the porridge made from unfermented dough after 6 h (4.2 [0.2] vs 3.8 [0.2]). Fermentation of maize dough is an effective method to reduce contamination of maize dough weaning foods with GNB.

Diversity of genotypes of hepatitis C virus in southern India.

A second generation assay for antibody to hepatitis C virus (anti-HCV) was used to screen 78 southern Indian individuals with a high risk of infection. RT-PCR targeted at the 5 end untranslated region (5UTR) of the HCV genome was used to evaluate evidence of viraemia in 32 anti-HCV positive sera. The PCR products amplified from the 5UTR of the HCV genome from 24 patients were sequenced, revealing the existence of two distinct groups of sequences: 21 corresponded to HCV type 1 while the other three sequences had 95% to 99% identity to HCV type 3. Two of these three isolates had more than 90% nucleotide identity in the NS5 region to established 3b sequences whereas the other had less than 74% nucleotide identity to any of the published genotype 3 (3a, 3b, 3c, 3d, 3e and 3f) sequences. However, a search of the EMBL nucleotide database revealed 91% identity to the unpublished sequence of an isolate of HCV from Indonesia. We provide evidence that these two isolates may represent a novel subtype within genotype 3. Our data also suggest that HCV genotype 1 predominates over HCV genotype 3 in southern India.

Hepatitis B virus DNA in the hepatocyte: a series of 160 biopsies

DNA-DNA hybridisation was used to examine 160 liver biopsies for the presence of the hepatitis B virus genome. HBeAg-positive HBsAg carriers were found to have replicating viral DNA in the hepatocytes and, very occasionally, HBV DNA was also integrated into the chromosomes. A high proportion of the anti-HBe-positive HBsAg carriers also have replicating HBV DNA and in the remainder integrated sequences are often, but not always, seen. No evidence was found, however, to implicate HBV in HBsAg-negative patients with alcoholic liver disease, nor in patients with a variety of other liver diseases including non-A, non-B hepatitis.

Effect of fermentation of Ghanaian maize dough on the survival and proliferation of 4 strains of Shigella flexneri

Fermented and non-fermented Ghanaian maize dough was seeded with approximately 10(7) colony forming units of 4 strains of Shigella flexneri which had been isolated from patients with dysentery. In the non-fermented maize dough (pH 6.2) the shigellae were detectable in large numbers for up to 24 h after exposure. In the maize dough that had been fermented for 3 d (pH 3.2) 3 strains were detectable in small numbers for up to 6 h after inoculation. Thereafter none was isolated. The fourth strain, though detectable for up to 24 h after inoculation, had its numbers reduced considerably. This suggests that traditional methods of food preparation using fermentation have important anti-diarrhoeal functions and the current decline in popularity of such food technologies in certain developing countries may increase the risk of childhood diarrhoea.

Exclusion in liver by polymerase chain reaction of hepatitis B and C viruses in acute liver failure attributed to sporadic non-A, non-B hepatitis

Hepatitis B and C viruses have been implicated in a few cases of acute liver failure attributed to sporadic (community acquired) non-A, non-B hepatitis, but reports are conflicting. We determined whether hepatitis B virus and hepatitis C virus were detectable in prospectively stored hepatectomies from seven British patients grafted for acute liver failure attributed to sporadic non-A, non-B hepatitis. For hepatitis B virus, we used nested polymerase chain reaction with primers to the core and surface regions. For hepatitis C virus, we used one round of reverse transcription polymerase chain reaction with primers to the 5 untranslated region and Southern hybridization using an internal oligonucleotide probe as well as nested PCR for the E1 region. Positive controls were native livers from two patients with unequivocal fulminant hepatitis B and from four patients with cirrhosis attributed to hepatitis C virus. Our negative findings suggest that, in the UK, acute liver failure attributed to sporadic non-A, non-B hepatitis most likely is caused by agent/s other than hepatitis B and C viruses.

Detection of hepatitis B virus DNA in hepatocellular carcinoma: methylation of integrated viral DNA.

In order to determine whether integrated hepatitis B virus DNA sequences in primary liver tumours are methylated we have analysed tumour DNA digested with either MspI or HpaII restriction endonuclease by Southern hybridization. Our results demonstrate methylation in 11 of 17 tumour DNA samples. Where possible, we have examined the tumour tissues for expression of HBsAg and HBcAg using the indirect immunoperoxidase technique. One tumour was positive for both HBsAg and HBcAg and a second was positive for HBsAg alone. Both of these tumours were in the group in which methylation of integrated HBV DNA sequences could not be detected. We postulate that methylation of integrated HBV DNA sequences may influence HBV gene expression in hepatocellular carcinoma.

Assay of HBV DNA in the plasma of HBV-carrier chimpanzees superinfected with non-A, non-B hepatitis

A dot hybridisation technique was used to monitor the levels of hepatitis B virus (HBV) DNA in the plasma of two HBV-carrier chimpanzees which had been inocula ed with documented infectious non-A, non-B hepatitis agents. A marked decrease in the quantity of HBV DNA in the plasma during the acute phase of the non-A, non-B hepatitis was observed in both carriers. The possible role of interferon or a similar antiviral agent in modulation of the HBV-carrier state is discussed. Hybridisation may become, in due course, the method of choice for examining blood samples for infectious hepatitis B virus.

Randomised controlled trial of lymphoblastoid interferon for chronic active hepatitis B

Thirty male patients (27 homosexual) with biopsy proven chronic active hepatitis B were randomised to receive lymphoblastoid interferon (Wellferon) or no treatment. All patients were HBeAg positive and had continuing viral replication. Patients receiving treatment were given a single daily intramuscular injection of interferon for 28 days at a starting dose of 2.5 MU/m2 increasing to a maximum of 7.5 MU/m2/day. Transient side effects of malaise and influenza like symptoms occurred in all patients and resolved rapidly after treatment. Hepatitis B viral replication was suppressed during interferon treatment in all patients but the effect was limited to the period of therapy. After one year there was no appreciable difference in viral markers between the two groups of patients and this treatment schedule appears less effective than the thrice weekly, three month regimes recently reported from other centres.

Candidate Hepatitis F Virus in Sporadic Non-A, Non-B Acute Liver Failure: Exclusion in Liver of Hepatitis Viruses A, E, C and B by Polymerase Chain Reaction

Sporadic non-A, non-B hepatitis is the most common, presumed viral, cause of acute liver failure in the UK and USA. Hepatitis E, C, or B viruses have been implicated in a few cases but reports are conflicting. We determined whether HAV, HEV, HCV, or HBV were detectable in prospectively stored hepatectomies from seven British patients grafted for acute liver failure attributed to sporadic non-A, non-B hepatitis. We used first- and second-round primers in a polymerase chain reaction (PCR) to amplify conserved regions of HAV, HEV, and HCV, the El/S (gp35) region of HCV, and surface and core regions of HBV. False-negative results were minimized by using other fulminant livers of comparable quality as positive controls and reconstruction experiments for sensitivity (nested PCR). Our negative findings suggest that acute liver failure attributed to sporadic non- A, non-B hepatitis in the UK is most likely caused by agent/s other than HAV, HEV, HCV, or HBV.