Showing papers by "Masashi Mizokami published in 2008"

When Should “I” Consider a New Hepatitis B Virus Genotype?

Abstract: Recently, Huy et al. described a new hepatitis B virus (HBV) strain isolated in Vietnam ([3][1]) and claimed it to be a “new genotype,” “HBV genotype I,” with a complex recombination involving genotypes C, A, and G. We refute both claims. Using complete genome sequence analysis of their

Molecular Epidemiology and Interferon Susceptibility of the Natural Recombinant Hepatitis C Virus Strain RF1_2k/1b

Abstract: BACKGROUND Hepatitis C virus (HCV) genotype is an important determinant of virological response to antiviral therapies. Currently, there are no data available on the molecular epidemiology and interferon susceptibility of the natural intergenotypic recombinant RF1_2k/1b (RF1) strain. METHODS Genotyping and RF1-PCR screening were performed on samples from 604 HCV RNA-positive individuals from 7 countries. uPA/SCID mice carrying human hepatocytes (chimeric mice) were infected with the RF1_2k/1b strain, and the susceptibility of the strain to interferon and ribavirin was compared with the susceptibilities of 2 different strains of genotype B, used as references. RESULTS Six new RF1 cases were identified in this study; 5 (2%) of 281 in Russia and 1 (1%) of 90 in Uzbekistan. Phylogenetic analyses based on Core/E1 and NS5b indicated that all RF1 representatives share a common evolutionary ancestor. Infection with RF1 was established in chimeric mice. Reduction of RF1 viral load was observed in response to 3 injections of 3 microg/kg pegylated-interferon alpha-2a alone or in combination with 50 mg/kg of ribavirin (0.5 or 1.4 log-copies/mL). CONCLUSIONS All identified RF1-type strains appear to be introduced from a single source, suggesting that intergenotypic recombination in HCV is sporadic and not associated with cocirculation of different genotypes in a population. The RF1 strain in this study was responsive to interferon in vivo.

Epidemiological and clinical evaluation of hepatitis B, hepatitis C, and delta hepatitis viruses in Tajikistan.

Abstract: The implication of genotypes is recognized increasingly in the clinical course of hepatitis B virus (HBV) and in response to anti-viral drugs of hepatitis C virus (HCV). Genotypic prevalence of both etiological agents varies geographically and no data are available for Tajikistan. To investigate the epidemiology and clinical significance of HBV and HCV genotypes in chronic hepatitis (group 1) and liver cirrhosis/hepatocellular carcinoma (HCC) (group 2) patients in Tajikistan, 124 patients with chronic liver disease (group 1 = 84 and group 2 = 40) were enrolled. Genotypes of HBV, HCV, and delta hepatitis virus (HDV) were determined by sequencing. The overall prevalence of anti-HCV, HCV core antigen (HCVcAg) and HBsAg was 46% (57/124) and 41.1% (51/124), respectively. Coinfection of HCV/HBV, HBV/HDV, and HCV/HBV/HDV was found in 4.8% (6/124), 11.2% (12/124), and 0.8% (1/124) of cases, respectively. HDV genotype 1 was found in 19.6% (10/51) of HBsAg-positive patients. The HBV/HDV coinfection was relatively high in group 2 compared to group 1 (15% vs. 7.1%). HCV/1b detected in 84.6% (44/52) of HCV RNA-positive patients, followed by 3a (7.6%), 2a (5.7%), and 2c (1.9%). HBV/D was detected in 94.1% (48/51) of HBsAg-positive patients, followed by HBV/A [5.8% (3/51)]. T1762/A1764 double mutation was associated with liver cirrhosis/HCC in HBV-infected patients (P = 0.0004). This is the first study on the molecular epidemiology of hepatitis viruses among chronic liver diseases patients in Tajikistan. Among HBV-infected patients, the T1762/A1764 mutation was associated with liver cirrhosis/HCC.

Virological and clinical implication of core promoter C1752/V1753 and T1764/G1766 mutations in hepatitis B virus genotype D infection in Mongolia.

Abstract: Background and Aim: The aim of the present study was to reveal virological and clinical features of hepatitis B virus (HBV) genotype D infection Methods: One hundred and twenty-two Mongolian chronic liver disease (CLD) patients infected with HBV were subjected for serological HBV-markers screening and HBV- enzyme immunoassay (EIA) genotyping Nucleotide sequences were analyzed for 48 HBV/D strains (23 isolated from hepatocellular carcinoma (HCC) and 25 from CLD patients) Results: Prevalence of hepatitis B e antigen (HBeAg) positivity was low (259%) in young patients (30 years old) indicating early HBeAg seroclearance in HBV/D carriers The T1764/G1766 double mutation was the most common basal core promoter (BCP) mutation (292%) and was frequent in HBeAg-negative patients (393%) Patients harboring T1764/ G1766 mutants exhibited lower HBV-DNA and HBV core antigen (HBcAg) levels than those with wild-type BCP strains (P = 0024, 0049, respectively) C1752 and/or V (not T) 1753 mutation was significantly prevalent in HCC patients (HCC vs CLD; 522% vs 20%, P = 0033) T1762/A1764 mutation was detected in 750% of HCC patients with high viral load (5 log copies/mL) Precore stop codon mutation A1896 was detected in (708%) of HBV/D-infected patients Conclusions: In Mongolians infected with HBV/D, C1752 and/or V1753 mutation was associated with HCC

Genotype, serum level of hepatitis C virus RNA and liver histology as predictors of response to interferon‐α 2α therapy in Japanese patients with chronic hepatitis C

Abstract: To determine whether pretreatment HCV-RNA level, hepatitis C virus genotypes, alanine aminotransferase and histology correlate with subsequent response to interferon-alpha therapy or not, serum HCV-RNA levels and genotype were determined by branched DNA signal amplification assay and genotype-specific polymerase chain reaction in 43 patients with chronic active hepatitis C. Response to recombinant interferon-alpha 2a (504 million units in total) was defined as complete and sustained CR-->SR, n = 12), complete response followed by relapse (CR-->Rel, n = 17), and no response (NR, n = 10), excluding dropouts (n = 4). Patients who showed CR-->SR had a lower HCV-RNA level (0.438 x 10(6) eq/ml) compared to CR-->Rel (2.452 x 10(6) eq/ml, p = 0.008) and NR (4.882 x 10(6) eq/ml, p = 0.009). A higher proportion of patients with CR-->SR had type 2a HCV (67%) compared to the CR-->Rel (28%) and the NR (0%). There was a trend for type 1b hepatitis C virus infection to have higher serum HCV-RNA levels. There was no correlation between pretreatment HCV-RNA level and alanine aminotransferase. However, no relation between pretreatment HCV-RNA level and liver histology was observed; a high proportion of patients with CAH2a showed CR-->SR, compared to those with CAH2b (p = 0.001). Moreover, the patients with CAH2b who had low level hepatitis C virus viremia did not show CR-->SR. These data indicate that pre-treatment serum HCV-RNA levels, genotype and liver histology are good predictors of subsequent response to interferon-alpha therapy in Japanese patients with chronic hepatitis C virus infection.

Hepatitis B Virus Transmission Pattern and Vaccination Efficiency in Uzbekistan

Abstract: A national program of universal vaccination for the prevention of chronic hepatitis B virus (HBV) infection was launched in Uzbekistan since 1998. To evaluate the 6 years' outcome of the program, 567 children were enrolled in the study. Among those enrolled, 333 had immunized with adw2 type based Engerix-B (Glaxo Smith Kline Beechem, Rixensart, Belgium) and 48 with adr type based Hepavax-Gene (Green Cross Vaccine Corporation, Korea). A cohort of 186 children born before the immunization program, was also included in the study. When 45 vaccinated children were compared to age/sex-matched 45 unvaccinated children, the sero-prevalence of HBsAg was 0 versus 11% (P = 0.56), and of anti-HBc was 0% versus 44% (P < 0.0001), respectively. Loss of anti-HBs was observed in 18.4% after 5 years. Among 13 HBsAg carriers found in this study, genotype HBV/D was found in 69%, HBV/A in 23% (all in unvaccinated group) and HBV/C in 8% (in vaccinated group). No significant differences were observed in this study between groups which received different vaccine formulation. Phylogenetic analysis of the HBV isolates obtained from family members of the HBsAg-positive children, revealed evidence suggesting that transmission in the vaccinated group was exclusively perinatal, whereas in the unvaccinated group horizontal transmission pattern predominated. In conclusion, HBV universal vaccination is efficient in Uzbekistan irrespective of the vaccine formulation used, because the horizontal transmission pattern predominates currently in this endemic region.

Two simultaneous hepatitis B virus epidemics among injecting drug users and men who have sex with men in Buenos Aires, Argentina: characterization of the first D/A recombinant from the American continent

Abstract: Previous studies have revealed that hepatitis B virus (HBV)/D and HBV/F predominate among blood donors from Buenos Aires, Argentina. In the present study, blood samples from two high-risk groups were analysed: 160 corresponding to street- and hospital-recruited injecting drug users [81.2% showing the 'anti-hepatitis B core antigen (anti-HBc) only' serological pattern] and 20 to hepatitis B surface antigen (HBsAg)(+)/anti-HBc(+) men who have sex with men. HBV genotypes were assigned by polymerase chain reaction amplification followed by restriction fragment length polymorphism and confirmed by nucleotide sequencing of two different coding regions. HBV DNA was detected in 27 injecting drug users (16.9%, occult infection prevalence: 7.7%), and 14 men who have sex with men (70%). HBV/A prevailed among injecting drug users (81.8%) while HBV/F was predominant among men who have sex with men (57.1%). The high predominance of HBV/A among injecting drug users is in sharp contrast to its low prevalence among blood donors (P = 0.0006) and men who have sex with men (P = 0.0137). Interestingly, all HBV/A S gene sequences obtained from street-recruited injecting drug users encoded the rare serotype ayw1 and failed to cluster within any of the known A subgenotypes. Moreover, one of the HBV strains from a hospital-recruited injecting drug user was fully sequenced and found to be the first completely characterized D/A recombinant genome from the American continent. Data suggest that two simultaneous and independent HBV epidemics took place in Buenos Aires: one spreading among injecting drug users and another one sexually transmitted among the homosexual and heterosexual population.

Genotyping of Hepatitis B Virus – Genotypes A to G by Multiplex Polymerase Chain Reaction

Abstract: Objectives: Eight genotypes (A–H) of hepatitis B virus (HBV) are known with variations in nucleotide sequences greater than 8%. Several recent publications found that the clinical c

Hepatitis B caused by a hepatitis B surface antigen escape mutant

Abstract: Amino acid substitutions within the S gene involving the major antigenic a determinant of the hepatitis B virus (HBV) surface antigen (HBsAg) have been detected in cases of failure of immunization against the virus. Our report showed development of clinical hepatitis in presence of antibody to HBsAg in a healthy individual. A single amino acid substitution (G145R) within the a determinant of the HBsAg was determined by sequencing of the isolated HBV strain. Lamivudine treatment efficiently cleared the peripheral HBV DNA, HBsAg, and hepatitis B e antigen. In conclusion, the immune escape mutant in the S gene can cause hepatitis despite pre-existing naturally acquired immunity.

Clearance of serum hepatitis C virus RNA after interferon therapy in relation to virus genotype

Abstract: The effect of recombinant interferon-alfa on serum HCV RNA levels in Japanese patients with chronic hepatitis C was investigated. At 24 weeks of treatment, 41 (32.5%) of 126 patients lost HCV RNA from serum, and aminotransferases were normalized in 31 (75.6%) of these 41 cases. HCV genotypes were categorized into four types (Type I, II, III, IV); the frequencies among the patients were: Type I: 0%, Type II: 70.6%, Type III: 20.6%, and Type IV: 6.3%. At the end of the 24-week treatment, HCV RNA levels were remarkably decreased in Type III patients and became undetectable in 18 (69.2%) of 26. In contrast, only 18 (20.2%) of 89 patients with Type II and two of eight with Type IV lost HCV RNA from sera. The relation between HCV genotype (Type III) and response to IFN therapy was also confirmed using a logistic regression model. HCV genotype seems to be an important factor in determining the response to IFN in patients with chronic hepatitis C.

長期入院がん患児における Clostridium difficile 消化管保有と院内伝播に関する検討

Abstract: Clostridium difficile is a major causative agent of antimicrobial-associated diarrhea, and the leading cause of nosocomial diarrhea. We clarified intestinal colonization and nosocomial spread of C. difficile in pediatric cancer patients undergoing antineoplastic therapy during long-term hospitalization. Subjects were 10 children with pediatric malignant diseases admitted from November 2005 to December 2006, aged 5 to 15 years, who received antineoplastic agents. Stool specimens were examined at hospitalization, after each course of treatment with antineoplastic chemotherapy, and when symptoms such as diarrhea or fever occurred. While C. difficile was detected from stool specimens of 8 of 10 children during their hospital stay, 6 of these 8 children were negative for C. difficile on the day of their admission. These results demonstrate that the use of antimicrobial agents and antineoplastic agents lead to overgrowth of C. difficile in intestinal tract of pediatric cancer patients. Five of the 8 children carried toxin A-positive, toxin B-positive C. difficle and 2 were diagnosed with C. difficile-associated diarrhea (CDAD). This demonstrates that CDAD is not a rare infection in pediatric cancer patients. Nine C. difficile isolates from 8 children were analyzed by PCR ribotyping. Two isolates from 2 children were typed into the same type;banding patterns of the remaining 7 isolates from 6 children were unique.

Expression of pre‐S1, pre‐S2, S and X peptides in relation to viral replication in livers with chronic hepatitis B

Abstract: The expression of large (pre-S1), middle (pre-S2), major S (S) polypeptides of the envelope (HBs) and X peptides of hepatitis B virus (HBV) was investigated in 37 liver specimens with chronic hepatitis B by indirect immunoperoxidase staining. Primary antisera utilized were polyclonal ones against HBs (poly-HBs), core (HBc) and X and monoclonal ones against pre-S1, pre-S2 and S with (particle-S) or without (peptide-S) conformational structure. The localization of HBs proteins in hepatocytes was classified into three types: diffuse, membranous and inclusion. The peptide-S and pre-S2 were expressed at nearly the same frequency as poly-HBs in all types, whereas particle-S was found less frequently (18/29 cases) in the inclusion type, and pre-S1 was recognized relatively rarely (9/33 cases) in the membranous type. As for staining intensity, peptide-S and pre-S2 were almost identical to poly-HBs which stained the most strongly among all three staining types. Particle-S was similar to poly-HBs in the membranous type, but was weak in the inclusion type in the majority. While pre-S1 was stained in a similar intensity to poly-HBs in the diffuse and inclusion types, it was weak or negative in the membranous type. Thus, envelope particles indicated by particle-S staining appeared to be located most frequently in the membranous type, but their assembly might be suppressed in the inclusion type where pre-S1 was well expressed. The X peptide was more frequently detected in the liver with serum HBe antigen and/or HBV DNA. The X peptide was stained exclusively in the cytoplasm of hepatocytes and was correlated with the cytoplasmic HBc antigen. The X peptide was not observed differently between cases with and those without cirrhosis. This suggests that the expression of X peptide tends to occur with virus replication but not with disease progression.

Pulsed Field Gel Electrophoresis Analysis Conditions for Molecular Epidemiological Group B Streptococcus Identification

Abstract: Introduction:In studying PFGE process conditions, using 7 noninfected Group B Streptococcus(GBS)detected in 3 mother-neonate pairs, some type-III serotype GBS, which produce a large amount of pathogenic neuraminidase, were not detected when we applied PFGE to strains detected in neonates developing meningitis, so we investigated PFGE process conditions under which the GBS type-III serotype could be detected. Methods:Subjects were 26 strains of meningitis causing GBS and 22 noninfected strains. Different lysis processes and restriction enzymes were studied under PFGE process conditions, and the mixture of lysozyme(5mg mL [120-02674 ; Wako])and mutanolysin(100U mL [M4782 ; Sigma-Aldrich])was examined in the lysis process. Lysozyme increased from 50 to 4,000μL, and mutanolysin from 20 to 300μL. We prepared and heated several plugs at 90°C for 10 minutes. After restriction enzyme digestion, we applied electrophoresis to the plugs for 19.8 hours(initial ramp time, 5.3 ; final ramp time, 34.9)with the running buffer temperature maintained at 14°C and at an angle of 120 with a CHEF-DRIII system(Bio-Rad, USA). Results:Strains detected using the PFGE analysis (lysozyme 50μL and mutanolysin 20μL for bacteriolysis, restriction enzyme Apa I)were 17 of 22 noninfected strains―type III, 7 strains ; type NT6, 3 strains ; type Ib, JM 9, 2 strains ; and type Ia, II, V, one strain―and 11 of 26 meningitis-causing strains―type Ib, 4 strains ; type III, 3 strains ; type Ia, 2 strains ; and type NT6, 2 strains (Fig. 1, Lane 1-11)―. Strains not detected―5 non-infected, and 15 infected(Fig. 1, Lane 12-26)― were all type-III serotype. We detected these 20 strains by lyzing ― using 50μL of lysozyme and 20μL of mutanolysin, and heating at 90°C for 10 minutes―and by using restriction enzymes ―Sma I and Sal I―(Fig. 2,3). Discussion:Strains not detected by PFGE analysis―5 noninfected, and 15 infected― were all type-III serotype GBS. We detected these 20 strains by improving detection conditions involving heating temperature and restriction enzymes Sma I and Sal I. Further work is need, however, to determine the relationship between results of PFGE analysis here and pathogenic agents of strains examined. PFGE analysis detected 11 of 26 meningitis-causing strains, 3 of which were type-III serotype and the remaining 15 strains all type-III serotype. 短 報