Rajib Kishore Hazam

Evidence of extrahepatic replication of hepatitis E virus in human placenta.

The incidence and severity of hepatitis E virus (HEV) infection in pregnant women is high in developing countries. Transplacental transmission of HEV in the third trimester of pregnancy has been found to be associated with high fetal mortality. Based on this evidence and in the absence of reports on HEV replication in extrahepatic sites, this study was carried out to investigate if HEV replication occurs in the placenta of infected mothers. The study included 68 acute viral hepatitis (AVH) and 22 acute liver failure (ALF) pregnant patients. Viral RNA was extracted from blood and placenta. HEV replication in placenta was confirmed by a replicative negative-strand-specific reverse transcriptase PCR. Viral load was estimated by real-time PCR. Immunohistochemical studies were also carried out for in situ detection of HEV in placental tissue sections. Replicative HEV RNA was detectable only in the placenta in ALF and AVH cases and not in blood samples. Positive staining of placental tissue sections with HEV antibody against the viral structural protein ORF3 was observed. HEV replication in placenta also correlated with fetal and maternal mortality in ALF patients. It is demonstrated for the first time that HEV replication occurs in human placenta and that placenta may be a site of extrahepatic replication of HEV in humans.

Does high viral load of hepatitis E virus influence the severity and prognosis of acute liver failure during pregnancy

The incidence and mortality in pregnant women with acute liver failure caused by hepatitis E virus (HEV) is high. Data on the viral load of HEV during pregnancy are limited. The study was designed to determine the viral load of HEV and its association with the disease severity in patients with acute liver failure. A total of HEV related 163 patients with acute liver failure which included 105 pregnant, 46 non‐pregnant women and girls and 12 men and 730 patients with acute viral hepatitis which comprised of 220 pregnant women; 282 non‐pregnant women and girls and 228 men were included. Viral load was measured by real‐time PCR. Comparison was made between the pregnant and non‐pregnant women. HEV RNA was detectable in 265 patients (142 pregnant; 75 non‐pregnant and 48 men) and 104 patients with acute liver failure (64 pregnant, 34 non‐pregnant and 6 men). The viral load of HEV in pregnant women with acute liver failure and acute viral hepatitis was significantly higher 129,984.0 ± 103,104.17 and 768.92 ± 1,105.40 copies/ml, respectively compared to the non‐pregnant women which was 189.2 ± 225 and 12.73 ± 7.8 copies/ml (P < 0.0001). The viral load of HEV was also significantly higher in the pregnant patients with acute liver failure compared to the pregnant women with acute viral hepatitis and also men (P < 0.0001). High viral load of HEV during pregnancy could be one of the factors responsible for the severity of the infection during pregnancy. J. Med. Virol. 85:620–626, 2013.

Role of nitric oxide synthase genes in hepatitis E virus infection.

Hepatitis E virus (HEV) is the most common cause of endemic and epidemic acute hepatitis. A correlation between iNOS, eNOS polymorphisms, levels and severity of disease has been reported, and here, we examined the role of iNOS and eNOS gene polymorphisms and their levels in HEV‐related acute viral hepatitis and acute liver failure. Hepatitis E virus‐related cases of acute hepatitis (294 patients) and liver failure (82 patients) and age‐ and sex‐matched healthy controls (331 subjects) were included in the study. PCR‐RFLP was performed to identify the polymorphisms in the iNOS and eNOS genes. iNOS and eNOS levels were studied using ELISA assays and HEV viral load, genotype and combined effects of iNOS genotype, levels and parameters for disease severity were examined. The frequency of iNOS (CT + TT) and eNOS (GT + TT) genotypes was higher in subjects with liver failure compared with controls. iNOS and eNOS levels in patients with acute liver failure (55.51 ± 6.33 IU/mL, 60.2 ± 3.69) cases were significantly increased as compared to patients with acute viral hepatitis (17.8 ± 6.08 IU/mL, 23.7 ± 6.57) and controls (P < 0.05). A significant positive correlation was observed between the iNOS and eNOS levels in our study population when compared with the severity of disease parameters. Hence, the iNOS C150T polymorphism and the eNOS G894T polymorphism and high levels of iNOS and eNOS are associated with an increased risk of HEV‐related acute hepatitis and liver failure. This study supports the possible role of nitric oxide synthase genes (iNOS and eNOS) in determining the severity of HEV infection.

HBV drug resistance : Its relevance in clinical practice

Drug resistance can be considered as a natural response to the selective pressure of the drug. An increase in HBV DNA can be a good indicator of the presence of a resistant HBV mutant population. The nucleoside analogues Lamivudine, Adefovir, Entecavir etc. are oral drugs used for Hepatitis B viral infection. Resistance to HBV drugs arises due to mutations in the polymerase gene. The HBV polymerase can be divided into 4 domains: 1) the terminal protein, 2) the variable spacer domain, 3) the polymerase/reverse transcriptase and 4) the RNase. Drug resistance to Lamivudine is associated with mutations in the very conserved catalytic polymerase /reverse transcriptase domain, located specifically at a locus of four amino acids consisting of tyrosine-methionine-aspartate-aspartate, termed the YMDD motif at position 204:M204V/I. Adefovir resistance mutations are at amino acid residues 181,236/238:A181T/V and N236T/N238D. The Entecavir resistance mutations are at amino acid residues 184, 202 and 250 of the polymerase: T184X, S202I/G/L and M250L/V. There are several assays available that identify resistance mutation like polymerase chain reaction, real time PCR with specific probes, hybridization methods (line probe assay) and restriction fragment length polymorphism (RFLP).The best approach for patients with Lamivudine resistance is to continue Lamivudine and add Adefovir. Lamivudine is effective in suppressing serum HBV DNA levels in patients with Adefovir resistance. Entecavir resistance mutations are sensitive to Adefovir and Tenofovir. The careful selection of a first-line agent is essential to avoid the occurrence of resistance and the development of cross resistance to other agents. The most effective therapy of antiviral-resistant HBV is prevention through judicious use of nucleos(t)ide analogue therapy.