Biomedical subjects
S Mishiro
Publications and source records attributed to S Mishiro.
Complete circular DNA genome of a TT virus variant (isolate name SANBAN) and 44 partial ORF2 sequences implicating a great degree of diversity beyond genotypes.
Information on the entire genome of TT virus (TTV) has been scarce. The circular ssDNA genome of a variant (isolate name SANBAN) that we sequenced was only 56.7% homologous to the prototype isolate (TA278), with even lower homology at the amino acid level: 34.2% for ORF1 and 39.7% for ORF2. Regarding the ORF1, SANBAN was only very distantly related to the six major TTV genotypes reported to date. In partial ORF2 sequences determined on 44 isolates taken together, TTV has a broad range of genetic diversity and the SANBAN isolate may represent a new TTV-like viral species or genus and not merely a genotype of TTV.
Clinical implications of mutations C-to-T1653 and T-to-C/A/G1753 of hepatitis B virus genotype C genome in chronic liver disease.
Among many mutational "hot spots" on hepatitis B virus (HBV) genome, A-to-T1762 and G-to-A1764 within the core promoter have been underscored in view of disease association as well as viral expression/replication. Although to a lesser extent, C-to-T1653 and T-to-V(C/A/G)1753 were also noteworthy in our previous study. To assess the clinical significance of these mutations, we determined the nucleotide sequence of an HBV DNA fragment covering these sites in HBsAg-positive blood donors (n = 160) and patients with chronic hepatitis (n = 66), liver cirrhosis (n = 45), and hepatocellular carcinoma (n = 58), most of whom were infected with genotype C HBV (subtype adr). In cases where HBe antigen was positive, the frequency of T1653 and/or V1753 showed a striking increment from chronic hepatitis patients (18%) to liver cirrhosis and/or hepatoma patients (82%), whereas that of T1762/A1764 was already high in chronic hepatitis patients (76%). In HBe antigen-negative cases, by contrast, significant difference in the frequency of T1653/V1753 mutants was found between blood donors (22%) and chronic hepatitis patients (67%). Our results suggest that T1653/V(particularly C)1753 mutants are more closely associated than T1762/A1764 with the progression of liver disease from chronic hepatitis to cirrhosis in HBe antigen-positive patients. A system of site-directed mutagenesis PCR RFLP was constructed to diagnose T1653 and C/A1753 more conveniently. Detecting T1653 and C/A1753 by this method would contribute to the differential diagnosis of HBV-associated liver disease.
Genotypes of TT virus (TTV) compared between liver disease patients and healthy individuals using a new PCR system capable of differentiating 1a and 1b types from others*.
TT virus (TTV) lacks obvious pathogenicity; almost all of the infected hosts are symptom-free. A possibility remains, however, that certain strains can cause liver disease while most others are non-pathogenic. Genotypes 1 a and 1 b have been proposed to contain such pathogenic strains. To test this possibility, we constructed a PCR system capable of detecting TTV of the 1 a and 1 b genotypes differentially from the other genotypes, and compared the frequencies of these genotypes between patients with liver disease of unknown etiology (n=42) and healthy individuals (n=50). The assay comprised 3 steps: i) PCR to amplify a 3.2-kb fragment using universal primers; ii) 2nd-round PCR, starting from the 3.2-kb amplicon, for a approximately 280-nt fragment by use of genotype 1-specific primers; and iii) digestion of the approximately 280-nt amplicon with MboI and BanI to discriminate between 1 a and 1 b. Eventually, 40 (95%) of the patients and 47 (94%) of the controls were positive for the 3.2-kb amplicon, and the 1 a, 1 b, 1 a+1 b, and non-1 genotypes of TTV were found in 2 (5%) vs 4 (9 percent), 5 (13%) vs 4 (9%), 1 (3%) vs 1 (2%) and 32 (80%) vs 38 (81%) of the 40 patients and 47 controls, respectively: the distribution was almost identical between the two groups. The hypothesis that the genotype 1 of TTV may be more closely associated with liver disease than other genotypes was not supported by this study.
Ethnic difference in the prevalence of monoclonal B-cell proliferation in patients affected by hepatitis C virus chronic liver disease.
BACKGROUND/AIM: In previous studies we demonstrated that all patients affected by HCV-positive type II mixed cryoglobulinaemia have a monoclonal B-cell population in peripheral blood mononuclear cells, and that a large fraction of HCV-infected patients develop a monoclonal B-cell expansion, even in the absence of dosable serum cryoglobulins. However, the prevalence of Type II mixed cryoglobulinaemia in HCV-infected individuals seems to be high in Italy, whereas it is very low in Japan. This study was performed to investigate whether there are ethnic differences in the prevalence of asymptomatic HCV-associated monoclonal B-cell expansions. METHODS: Forty-four Japanese patients affected by HCV-positive chronic liver disease (two healthy carriers, 31 chronic hepatitis and 11 cirrhosis) were compared with a group of 60 Italian patients (one healthy carrier, 49 chronic hepatitis, and 10 cirrhosis) without dosable levels of cryoglobulins. The monoclonality of peripheral blood mononuclear cells was investigated by RT/PCR analysis of Immunoglobulin gene rearrangements. Liver function tests, rheumatoid factor, cryocrit level, anti-HCV antibodies, HCV-RNA, and HCV genotype were performed according to standard methodology. RESULTS: A B-cell monoclonal population was found in 26% of Italian patients, whereas all Japanese patients were negative. No correlation was found between B-cell monoclonality and severity of liver disease, length or source of the infection, HCV genotype, sex, clinical and biochemical parameters. CONCLUSIONS: This study indicates that a monoclonal B-cell proliferation in peripheral blood mononuclear cells is common in HCV infection, but only in Italy, whereas it is absent in Japan. This explains the very low prevalence of Type II mixed cryoglobulinaemia in HCV-positive Japanese subjects, and suggests that HCV is able to determine a B-cell expansion only in the presence of, presently undetermined, host factors.
GBV-C/HGV and HCV infection in mixed cryoglobulinaemia.
Recently, a new, suspected hepatotropic virus has been identified. Named GBV-C/HGV, this virus shares with the hepatitis C virus (HCV) routes of transmission and molecular organization. Indeed, a proportion of HCV-infected patients (10-25%) are also carriers of GBV-C/HGV. Since mixed cryoglobulinaemia (MC) is closely associated with HCV infection, the aim of this study was to determine the prevalence of GBV-C/HGV infection in MC patients, and to investigate whether the double infection influenced the clinical and/or laboratory aspects of the disease. 52 patients affected by MC were studied. 100 patients affected by HCV-positive chronic liver disease (CLD) without MC were used as control group. To determine the prevalence of GBV-C/HGV infection in general population, 150 blood donors were studied, as well as 80 patients affected by non-A-E CLD. Among the MC patients, only five (9.6%) were positive for both HCV and GBV-C/HGV infection. No difference was found between patients with and without double infection as regards main clinical and laboratory aspects. Among HCV-positive CLD cases, 27 were positive for double infection. Among blood donors, the prevalence of GBV-C/HGV infection was 8.0%, whereas in cases with cryptogenetic CLD the prevalence was 5.0%. In conclusion, these data show that GBV-C/HGV infection does not play any role in the pathogenesis of MC.
Epidemiology of hepatitis C virus in japan.
Blood screening for hepatitis C virus (HCV) antibodies at Japanese Red Cross blood centers has indicated an age-dependent prevalence of HCV infection in the general population of Japan: the older the age, the higher the prevalence. The same pattern was corroborated by the sentinel study conducted in an HCV-endemic area. The high prevalence of HCV in the elderly is most likely due to a spread of HCV infection during the turmoil period just after World War II: in particular by illicit intravenous amphetamine abuse. Fortunately, however, since the rate of newly acquired infection is currently too low to influence HCV prevalence in each age group, the total number of HCV carriers (estimated to be 691,852 for the age group 20-64 years) would decrease spontaneously with generation takeover to approximately 50% of the current number during the decade to come.
The Nineteenth United States-Japan Joint Hepatitis Panel Meeting.
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Mother-to-infant transmission occurs more frequently with GB virus C than hepatitis C virus.
A total of 107 hepatitis C virus (HCV)-infected pregnant women were screened for GB virus C (GBV-C) RNA in their sera, and 11 (10.3%) were positive. Among 11 infants born to these HCV/GBV-C co-infected mothers, GBV-C RNA was detected in 7 (63.6%) while HCV RNA was found in 1 (9.1%) within 1 year after birth: this difference was statistically significant (p = 0.023). The mothers of infected infants had significantly higher serum titers of GBV-C RNA than those of uninfected infants: 10(6.7 +/- 0.5) vs 10(4.0 +/- 1.0) copies/ml in average (p = 0.001). The baby in whom HCV RNA was found was also positive for GBV-C RNA, and had an elevation in serum transaminase levels, whereas all the other GBV-C infected infants showed no evidence for hepatitis. A family study, performed on 2 of the 7 infected cases, revealed that all the elder siblings of the index infants were also GBV-C RNA-positive. Nucleotide sequence of GBV-C RNA, amplified by PCR from an NS3 region, was completely identical between the mother and the infant within each family, but varied significantly across different families. These results suggest that GBV-C is more easily transmitted from mother to infant than HCV, although hepatitis is not caused thereby.
Hepatitis C virus infection and mutations of mannose-binding lectin gene MBL.
We assessed the genetic polymorphism of mannose-binding lectin (MBL) in 93 patients with chronic hepatitis C (45 responders and 48 nonresponders to interferon) and 218 healthy controls. Mutant allele was identified only at codon 54 (Gly-->Asp), leading to three genotypes (54 m/m, 54 W/m, and 54 W/W). Frequency of 54 m/m was significantly lower in interferon-responders (2.2%), compared to those in nonresponders (14.6%) and controls (10.6%): p < 0.05. Our results suggest that homozygous carriage of the variant allele of codon 54 of MBL may predict poor response to interferon in chronic hepatitis C patients.
High-dose (9 MU) long-term (60 weeks) alfa-interferon therapy for chronic hepatitis patients infected with HCV genotype 1b.
Efficacy of standard regimens (e.g., 3-6 MU for 24 weeks) of alfa-IFN therapy for chronic hepatitis C has been limited, particularly in patients with HCV/1b. To see if higher-dose longer term treatment is more effective, we tried a 9 MU 60-week regimen. HCV/1b-infected chronic hepatitis patients received 9 MU IFN alpha 2a everyday but Sunday for 2 weeks and thrice a week for next 10 weeks, and 76 patients became HCV RNA-negative while 81 remained positive. The RNA-negative patients were then randomized to receive 3 MU (group I, n = 37) or 9 MU (group II, n = 39) for 48 weeks. Of the RNA-positive patients, only those with normal ALT received another 9 MU 48-week treatment (group III, n = 45). Sustained responders (SR) were defined as those with negative RNA and normal ALT 6 months after the therapy. SR rates based on intent-to-treat principle did not differ significantly between groups I and II (30% vs 41%), but those based on the protocol-compatible cases showed a significantly lower than those in group II. Adverse effects of IFN, developed more frequently in groups II and III than in group I, were mostly reversible. In conclusion, our results encourage 9 MU 60-week IFN alpha treatment in HCV/1b-infected patients with careful attention to adverse effects, and suggest that the treatment should be discontinued if HCV RNA does not disappear within 12 weeks.
Hepatitis B virus genomic sequence in the circulation of hepatocellular carcinoma patients: comparative analysis of 40 full-length isolates.
We determined full-length nucleotide sequence of hepatitis B virus (HBV) genome in sera from 40 Japanese patients with HBsAg-positive hepatocellular carcinoma (HCC), in order to obtain information on HCC-specific characteristics, if any, of the HBV genome. Direct sequencing of the long distance PCR products starting from 50 microliters of serum samples revealed that 95% of our isolates were of genotype C, and that mutations and deletions/insertions were very common. With respect to envelope protein genes, deletions and missense mutations were frequent in preS2, and the determinant a domain of HBsAg was rich in "antibody-escape" mutations. Within the precore/core region, the most remarkable mutation was the replacement of proline of wild type by other amino acids at codon 130 of the core gene, which was found in 58% of our isolates, while precore-stop mutation was found in 45%. Most interestingly, however, about 90% of our isolates had mutations at nt positions 1762 (A-to-T) and 1764 (G-to-A) within the core promoter, which had been implicated in "e-suppressive" phenotype of HBV genome. G-to-A at nt 1613 and C-to-T at nt 1653 within enhancer II and T-to-C/A at nt 1753 within core promoter were also evident: 38%, 53%, and 40%, respectively. It was interesting that some of the characteristics observed in our isolates form HCC patients had been previously implicated in fulminant hepatitis and/or acute exacerbation of chronic hepatitis.
Association of mannose-binding lectin gene haplotype LXPA and LYPB with interferon-resistant hepatitis C virus infection in Japanese patients.
BACKGROUND/AIMS: Mannose-binding lectin, a key factor of the innate immune system, has genetic polymorphism, and individuals who carry certain genotypes of mannose-binding lectin are known to be more prone to severe or prolonged infectious diseases. We aimed to find any relevance of mannose-binding lectin polymorphism to hepatitis C virus infection. METHODS: We determined the mannose-binding lectin genotypes by sequence specific priming-polymerase chain reaction in 159 hepatitis C virus-infected chronic hepatitis patients and 218 healthy controls in Japan by looking at 4 polymorphic loci: 2 (H/L and X/Y) within the promoter region and 2 (P/Q and A/B) within exon-1 of the mannose-binding lectin gene. RESULTS: A group of mannose-binding lectin genotypes designated "XB-type" (containing LXPA or LYPB haplotype at least heterozygously) was less frequently found in interferon-responsive patients (38.5%) than in interferon-resistant patients (60.7%, p=0.008) and controls (57.3%, p=0.014). Individuals with the "XB-type" had lower serum concentrations of mannose-binding lectin, compared to those with "YA-type", which is defined by homozygous carriage of both Y and A alleles: 0.63+/-0.61 vs 2.06+/-1.17 mg/l, p<0.001. CONCLUSIONS: Our results suggest that the mannose-binding lectin-related innate immune system plays an important role in elimination of hepatitis C virus during interferon therapy. Determining mannose-binding lectin genotypes may help in selecting the hepatitis C virus-infected patients to be treated with interferon.
[Clinical significance of hepatitis G virus (GBV-C/HGV)].
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The 18th United States-Japan Joint Hepatitis Panel Meeting.
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Distribution of the hepatitis B surface antigen subtypes in Indonesia: implications for ethnic heterogeneity and infection control measures.
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High prevalence of hepatitis C virus infection in Japanese patients with porphyria cutanea tarda.
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Prevalence of hepatitis C virus and distribution of its genotypes in Northern Eurasia.
We tested hepatitis C virus (HCV) antibody in 4216 sera collected from healthy people living in European part of Russia (including Northern, North-Western, Central, Central-Blacksoil, Volga-Vyatka, Volga, and North-Caucasian regions), non-European part of Russia (the Urals, East-Siberia, and the Far-East regions) and Mongolia. Prevalence of HCV antibody varied significantly by regions, ranging from 0.7% in Central region of European part of Russia to 10.7% in Mongolia. Genotyping of HCV (into 1a, 1b, 2a, 2b, and 3a) was performed on 469 sera from blood donors and patients (in Russia, Moldova, Turkmenistan, and Mongolia) who were positive for both HCV antibody and RNA. Genotype 1b was the most dominant genotype irrespective of regions (68.9%), with the highest rate in Moldova (96%). HCV unclassifiable into genotypes 1a-to-3a was found in 28 (6.0%) samples: particularly 4 of 10 samples from Lipetzk were untypable. Overall, HCV genotypes in European part of Russia were more similar to those in European countries, while those in Eastern part of Russia more similar to China or Japan. Genotype distribution was not associated with the clinical expression of HCV disease: acute hepatitis, chronic hepatitis or liver cirrhosis.