Construction and application of a genechip method for detection of hepatitis B virus lamivudine-resistant mutants and basal core promotor/Pre-C mutants / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>The objective of this research is to construct a clinic-usable genechip method for detection of hepatitis B virus lamivudine-resistant mutants and basal core promotor/Pre-C mutants, compare this method with DNA sequencing to investigate this genechip's character (sensity, specificity, stability and practicability in clinic) and apply it in clinic.</p><p><b>METHODS</b>This genechip detection method can detect the DNA and 8 mutative site of HBV, include 3 lamivudine-resistant mutation site(No. 180, 204, 207 site in DNA polymerase gene), 5 HBeAg escape-related mutation site (nt 1896, 1899, 1862, 1764,1762 site in BCP/Pre-C region).The results of genechip method was verified by DNA sequencing.</p><p><b>RESULTS</b>In detecting HBV DNA, the results of genechip were agree with 100% of the results of DNA sequencing. In detecting HBV mutants, 251 sites (in 32 samples, 256 sites) showed the same results using both methods, and only 5 sites were not completely match (P > 0.05). In these 5 sites, genechip methods got multi-infection results, but sequencing got single-infection results.</p><p><b>CONCLUSION</b>These results suggest that genechip method has the same positive rate and almost these same specificity with DNA sequencing method, and is better than DNA sequencing method in detecting multi-infected HBV strains. [Key words]</p>

Expressive features of HBsAg and HBcAg in the livers of chronic hepatitis B and its clinical significance / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To investigate the necessity of detecting on the expressive intensity and pattern of HBsAg and HBcAg in the livers of chronic hepatitis B.</p><p><b>METHODS</b>HBsAg and HBcAg were detected in paraffin-embedded liver tissue by EnVision immunohistochemistry. Serum hepatitis B virus DNA (HBV DNA) was tested by real-time quantitative polymerase chain reaction. The degrees of hepatic inflammatory activity (grade) and fibrosis (stage) of liver biopsies were determined according to the standard of the Chinese program of prevention and treatment of viral hepatitis.</p><p><b>RESULTS</b>The expression of HBsAg was not correlated with the grade, the stage and the levels of serum HBV DNA (P > 0.05). Liver HBcAg expressive intensity was not correlated with the grade (r=0.02, P > 0.05), while negatively correlated with the stage (r=0.28, P < 0.01) and positively correlated with the serum HBV DNA levels (r=0.53, P < 0.01). Liver HBcAg expressive pattern was negatively correlated with the grade (r=-0.27, P < 0.01). The grade in cytoplasmic pattern group was higher than in nuclear pattern group and in mixed pattern group (P < 0.01), and that in mixed pattern group was higher in nuclear pattern group (P < 0.01). Liver HBcAg expressive pattern was negatively correlated with the stage (r=-0.23, P < 0.01). The stage in cytoplasmic pattern group was higher than in nuclear pattern group and in mixed pattern group (P < 0.05). Liver HBcAg expressive pattern was positively correlated with the levels of serum HBV DNA (r=0.22, P < 0.01).</p><p><b>CONCLUSION</b>Distinguishing the expressive intensity and pattern of HBsAg and HBcAg in the liver of chronic hepatitis B may not help understand the degree of hepatic lesion. The detection of HBcAg in liver tissue of CHB may be beneficial for the antiviral therapy.</p>

High-flux detection of HBV lamivudine resistant strains by genechip and analysis of mutant hotspots in those strains / 中华肝脏病杂志

<p><b>OBJECTIVE</b>Lamivudine resistant HBV strains in Shenzhen were detected at multiple sites and in large amounts to understand further the distribution of lamivudine resistant mutants.</p><p><b>METHODS</b>552 Hepatitis B patients's sera were examined using genechip method. Among them, 192 samples of lamivudine resistant mutant were further analyzed.</p><p><b>RESULTS</b>In those 192 lamivudine resistant samples, 191 were YMDD mutants, 124 mutants of codon 528 and 9 mutants of codon 555. 88% YMDD mutants were multi-mutants of YVDD and codon 528; single mutants of YIDD; multi-mutants of YIDD and codon 528. 91% codon of YMDD mutants were GTG, ATT; the other 9% were ATA, ATC.</p><p><b>CONCLUSIONS</b>These results suggest that mutants of codon 552 (YMDD) are core mutants. Mutants of codon 528 and 555 are incidental mutants, YVDD mutants always emerge with mutants of codon 528, but YIDD mutants appear differently. 9% YMDD mutants's codons are ATA or ATC. This may be the reason for the low positive rate shown by using the conventional PCR methods.</p>