Mutational analysis of reverse transcriptase and surface proteins of patients with partial virological response during mono and combination antiviral therapies in genotype D chronic hepatitis B.

INTRODUCTION: The mutational pattern of chronic Hepatitis B virus (HBV) is unclear in patients who show incomplete response to antiviral therapy. The aims of this study were 1) to determine the benefit of combination therapy with adefovir dipivoxil (ADV) and Lamivudine (LAM) versus ADV or LAM alone in maintaining virological, biochemical and histological responses and 2) to investigate the patterns of mutations in the reverse transcriptase and surface proteins of HBV with LAM and/or ADF-resistant in partially-responded chronic hepatitis B (CHB) patients. METHODS: The study group consisted of 186 chronic HBV carriers who were admitted to the Tehran Hepatitis Network from 2010 to 2013. We retrospectively selected 86 patients who partially responded to different nucleoside analogue regimens. After 48 weeks of therapy, five groups of patients were defined including eight Lamivudine (LAM) Group (I), 30 Adefovir (ADV) Group (II), 16 ADV add on LAM Group (III), 32 ADV+LAM Group (IV), and 100 controls (no therapy). Reverse transcriptase (RT) and surface genes were amplified and sequenced for mutational analysis. RESULTS: All groups showed differences between mean values for age, gender, alanine transaminase (ALT), aspartate transaminase (AST), and HBV DNA levels groups showed significant differences than other groups (p < 0.05). The mutation frequencies for groups were I (1.7%), II (1.39%), III (2.28%), IV (2.0%), and V (0.38%). T54N, L80I/V, I91L/V, L180M, M204I/V, Q215P/S, and F221Y/S showed the highest number of mutations in all groups with different frequencies. Four new, unreported mutations were found. CONCLUSION: Those patients who failed to respond in the first 48 weeks, whether they were receiving mono or combination therapy, should be tested genotypically, for the early modification of treatment.

Detection of Epstein-Barr virus in synovial fluid of rheumatoid arthritis patients.

INTRODUCTION: Rheumatoid arthritis (RA) is one of the most common chronic inflammatory disorders. Genes and environmental factors contribute to RA. Epstein-Barr Virus (EBV) has been considered as one the RA pathogeneses. The aim of this study was to detect of the EBV genome in patients with RA. METHODS: In this cross-sectional study, 50 samples of synovial fluid were obtained from patients with RA from 2010-2012. Using a standard of the EBV genome and EBNA-1-specific primers, the method of PCR was set up. Then, all of the samples of synovial fluids separately were subjected to DNA extraction and Polymerase Chain Reaction (PCR) amplification. Data were analyzed using SPSS version 18.0. The statistical analysis was performed by the t-test. RESULTS: The demographic and laboratory characteristic assay revealed that the mean age of patients was 49, and the patients were 60% males and 40% females. In addition, in all cases, the mean rheumatoid factor (RF) levels of the patients were below the normal level. The results of this study showed that the PCR was able to detect EBV DNA in > 60% of the cases. CONCLUSION: The results of this study indicated that EBV was frequently detected in the synovial fluid of RA patients. Thus, EBV may be a strong candidate that can act at several levels of the pathophysiology of RA. However, these findings also indicated that EBV may play a role in the pathogenesis of RA. However, the possible relationship between RA and EBV must be determined by further research.

Evolution of hepatitis B virus surface gene and protein among Iranian chronic carriers from different provinces.

BACKGROUND AND OBJECTIVES: Iranian chronic HBV carrier's population has shown a unique pattern of genotype D distribution all around the country. The aim of this study was to explore more details of evolutionary history of carriers based on structural surface proteins from different provinces. MATERIALS AND METHODS: Sera obtained from 360 isolates from 12 Different regions of country were used for amplification and sequencing of surface proteins. A detailed mutational analysis was undertaken. RESULTS: The total ratio for Missense/Silent nucleotide substitutions was 0.96. Sistan and Kermanshah showed the lowest rate of evolution between provinces (P = 0.055). On the other hand, Khorasan Razavi and Khoozestan contained the highest ratio (P = 0.055). The rest of regions were laid between these two extremes. Azarbayjan and Guilan showed the highest proportion of immune epitope distribution (91.3% and 96%, respectively). Conversely, Sistan and Tehran harbored the least percentage (66.6% and 68.8%, respectively). Kermanshah province contained only 5.2%, whereas Isfahan had 54.5% of B cell epitope distribution. In terms of T helper epitopes, all provinces showed a somehow homogeneity: 22.58% (Fars) to 46.6% (Khuzestan). On the other hand, distribution of substitutions within the CTL epitopes showed a wide range of variation between 6.6% (Khuzestan) and 63% (Kermanshah). CONCLUSION: Further to low selection pressure found in Iranian population, the variations between different regions designate random genetic drift within the surface proteins. These finding would have some applications in terms of specific antiviral regimen, design of more efficient vaccine and public health issues.

Prevalence and characteristics of Epstein-Barr virus-associated gastric cancer in Iran.

BACKGROUND/AIMS: Gastric cancer (GC) is the second leading cause of cancer-related deaths worldwide and is the most frequent cancer in Iran. Epstein-Barr virus (EBV) has been shown to be associated with gastric cancer. The present study was carried out to investigate the prevalence of Epstein-Barr virus (EBV) associated gastric cancer among Iranian patients. METHODS: Ninety formalin fixed paraffin-embedded cases of gastric cancer were studied. The specimens were investigated for the presence of the EBV genome by quantitative real-time polymerase chain reaction. RESULTS: Of ninety specimens, EBV was detected in six cases (6.66%). The mean age for patients EBV-positive gastric carcinomas was 72.1 years, whereas the mean age for the entire group was 65.7 years. Four out of 64 (6.25%) male patients and 2 out of 26 (7.69%) female cases were positive for EBV. According to anatomic location, EBV was detected in 4 out of 39 (10.25% ) gastric cancer were located in cardia and 2 out of 26 (7.69%) gastric cancer were located in middle/corpus. CONCLUSIONS: The present study shows that the frequency of EBV-associated gastric carcinoma in Iran is  low. Differences of EBV-associated gastric carcinoma incidence in different countries may reflect the epidemiologic factors and dietary habits. Further analysis of clinical pathology features of EBV-associated gastric carcinoma using a larger number of cases would give invaluable insights into its etiology.

Prevalence and molecular analysis of occult hepatitis B virus infection isolated in a sample of cryptogenic cirrhosis patients in iran.

OBJECTIVES: The aims of this study are to investigate the prevalence of occult hepatitis B virus infection among patients with cryptogenic cirrhosis and to analyze the relationship between surface protein variability and occult hepatitis B virus infection, which may be related to the pathogenesis of occult hepatitis B virus infection in cryptogenic cirrhosis. Occult hepatitis B virus infection is a well-recognized clinical entity characterized by the detection of hepatitis B virus DNA in serum and/or liver in the absence of detectable hepatitis B virus surface antigen, with or without any serological markers of a past infection. METHODS: Sera from patients with cryptogenic chronic liver disease were tested for hepatitis B virus DNA using both real-time and nested PCR. In the detected hepatitis B virus DNA samples, the surface gene was analyzed for mutations. RESULTS: Hepatitis B virus DNA was detected in 38% of patients, all of whom had a viral load below 10,000 copies/mL. All hepatitis B virus belonged to genotype D. There were no significant associations between occult hepatitis B virus infection status and age, gender, ALT/AST levels, viral load or serologic markers of previous hepatitis B virus infection. There were 14 mutations found in 5 patients; 6 were in the major hydrophilic region, of which 4 were Y134F assigning for the "a" determinant region. All patients who acquired Y134F contained S207R (within HLA-A2-restricted CTL epitope) as a combination. CONCLUSION: Hepatitis B virus surface antigen variants may arise as a result of natural selection to evade the immune surveillance of the infected host, and subsequently may go undetected by conventional hepatitis B virus surface antigen screening tests. Etiological diagnosis of cryptogenic cirrhosis is significantly underestimated with current serology testing methods alone.

Prospective study of BK virus infection and nephropathy during the first year after kidney transplantation.

INTRODUCTION: The aim of this study was to assess the prevalence and severity of BK virus infection, BK virus nephritis, and related risk factors among kidney transplant recipients. MATERIALS AND METHODS: BK viremia during the first year of kidney transplantation was assessed prospectively in 32 successive recipients. BK virus DNA was extracted and determined in all samples by real-time polymerase reaction assay for 1 year after kidney transplantation. RESULTS: The mean age of the patients was 33.3 ± 15.3 years. Sixteen patients (50%) received antithymocyte globulin for induction therapy. Living donor transplant consisted of 75% of the kidney donations. Maintenance immunosuppressive therapy included cyclosporine A in 27 patients (84.4%), plus tapering prednisolone and mycophenolate mofetil. BK viremia was detected in 8 patients (25%). The highest detected plasma viral load was less than 4000 copies per milliliter. BK virus was respectively positive in 5 (62.5%), 2 (25%), and 1 (12.5%) patients during the first 4, 8, and 12 months after transplantation. Biopsy-proven rejection and antirejection therapy by methylprednisolone pulses were 5 and 2.3 times more common in patients with BK virus infection (P = .01 and P = .01), respectively. CONCLUSIONS: Despite occurrence of BK virus infection in 25% of our patients, BK nephropathy did not develop in any of them. Routine screening of BK virus infection, particularly in centers with low prevalence of BK virus nephritis, may not be cost effective for predicting this disease.

Drug-related mutational patterns in hepatitis B virus (HBV) reverse transcriptase proteins from Iranian treatment-naïve chronic HBV patients.

BACKGROUND: Immunomodulators and Nucleotide analogues have been used globally for the dealing of chronic hepatitis B virus (HBV) infection. However, the development of drug resistance is a major limitation to their long-term effectiveness. OBJECTIVES: The aim of this study was to characterize the hepatitis B virus reverse transcriptase (RT) protein variations among Iranian chronic HBV carriers who did not receive any antiviral treatments. MATERIALS AND METHODS: Hepatitis B virus partial RT genes from 325 chronic in active carrier patients were amplified and directly sequenced. Nucleotide/amino acid substitutions were identified compared to the sequences obtained from the database. RESULTS: All strains belonging to genotype D.365 amino-acid substitutions were found. Mutations related to lamivudine, adefovir, telbivudine, and entecavir occurred in (YMDD) 4% (n = 13), (SVQ) 17.23% (n = 56), (M204I/V + L180M) 2.45% (n = 8) and (M204I) 2.76% (n = 9) of patients, respectively. CONCLUSIONS: RT mutants do occur naturally and could be found in HBV carriers who have never received antiviral therapy. However, mutations related to drug resistance in Iranian treatment-naïve chronic HBV patients were found to be higher than other studies published formerly. Chronic HBV patients should be monitored closely prior the commencement of therapy to achieve the best regimen option.