Clearance of HBV DNA in immunized children born to HBsAg-positive mothers, years after being diagnosed with occult HBV infection.

In a previous study, we observed immunoprophylaxis failure due to occult hepatitis B virus (HBV) infection (OBI) despite the presence of adequate levels of anti-HBs in 21 (28%) of 75 children born to HBsAg-positive mothers. The aim of the study was to explore the maintenance of this cryptic condition in this population. Of 21 OBI-positive children, 17 were enrolled. HBV serological profiles were determined by enzyme-linked immunosorbent assay. Highly sensitive real-time and standard PCR followed by direct sequencing were applied in positive cases. The mean age (±SD) of studied patients was 6.57 ± 2.75 years. All children still were negative for HBsAg. All but one (94%) were negative for HBV DNA. Only two children were positive for anti-HBc. The results of the most recent anti-HBs titration showed that 4 (23.5%) and 13 (76.5%) had low (<10 IU/mL) and adequate (>10 IU/mL) levels of anti-HBs, respectively. The only still OBI-positive patient had an HBV DNA level of 50 copy/mL, carried the G145R mutation when tested in 2009 and again in 2013 in the 'a' determinant region of the surface protein. Further follow-up showed that after 18 months, he was negative for HBV DNA. In high-risk children, the initial HBV DNA positivity early in the life (vertical infection) does not necessarily indicate a prolonged persistence of HBV DNA (occult infection). Adequate levels of anti-HBs after vaccine and hepatitis B immune globulin immunoprophylaxis following birth could eventually clear the virus as time goes by. Periodic monitoring of these children at certain time intervals is highly recommended.

Hepatitis B virus surface protein mutations clustered mainly in CTL immune epitopes in chronic carriers: results of an Iranian nationwide study.

Mutations within the coding region of hepatitis B surface antigen (HBsAg) have been found naturally in chronic carriers. To characterize the mutations of HBsAg from Iranian chronic carriers who were vaccine and/or medication naive. The surface genes from 360 patients were amplified and directly sequenced. The distribution of amino acid substitutions was classified according to different immune epitopes of the surface protein. All isolates belonged to genotype D. 222 (61.6%) of 360 patients contained at least one amino acid substitution. 404 (74.5%) of 542 amino acid changes occurred in different immune epitopes of HBsAg, of which 112 (27.7%) in 32 residues of B-cell epitopes (62 in the 'a' determinant); 111 (27.4%) in 32 residues of T helper; and 197 (48.7%) in 32 residues inside cytotoxic T lymphocyte (CTL) epitopes. One Th (186-197) and two CTL (28-51 and 206-215) epitopes were found to be hotspot motifs for the occurrence of 213 (52.7%) substitutions. 20 stop codons were identified in different epitopes. There was a significant association between amino acid substitutions and anti-HBe seropositivity; however, the correlation between such changes with viral load and ALT levels was not significant. In chronic hepatitis B virus(HBV) carriers, positive selection in particular outside the 'a' determinant appeared to exert influence on the surface proteins. These changes could be immune escape mutations naturally occurring due to the host immune surveillance especially at the T-cell level.

Hepatitis B vaccine: prophylactic, therapeutic, and diagnostic dilemma.

Hepatitis B virus (HBV) remains a serious public health problem worldwide, accounting for high morbidity and mortality rates as well as significant personal, societal, and economic costs. Hepatitis B is a preventable disease; a safe and effective vaccine has been available for 30 years. The World Health Organization aims to control HBV worldwide by integrating HB vaccination into infant, possibly adolescent, and at-risk adult routine immunization programs. In recent years, a drastic reduction in the mortality and morbidity of chronic HBV, including hepatocellular carcinoma, has occurred, particularly in hyperendemic areas. In addition, a therapeutic vaccine that enhances patient immune response has been considered as a possible alternative to antiviral agents. However, mutant HBV may infect individuals who are anti-HBs positive after immunization (vaccine-escape) and/or fail for detection of HBsAg (diagnostic-escape), which may lead to transmission through donated blood or organs. This review attempts to summarize the prophylactic, therapeutic, and diagnostic concerns on HBV vaccines.

Correlation of Hepatitis B surface antigen mutations with clinical status of the chronically infected patients from Kermanshah, West of Iran.

AIM: Hepatitis B virus (HBV) genetic and protein variations have been found in chronic HBV infected patients who did not receive any treatment and active or passive immunizations. The aims of this study were to determine the genotypes as well as the patterns of variations distribution in chronically infected patients from the west part of Iran. METHODS: Forty-six people with chronic HBV infection were enrolled in the study. The surface genes were amplified, sequenced and subsequently aligned using international and national Iranian database. RESULTS: All strains belonged to genotype D, subgenotype D1 and subtype ayw2. Of all 116 "mutations" that occurred at 59 nucleotide positions, 49 (42.2 %) were missense (amino acid altering) and 67 (57.7%) were silent (no amino acid changing), respectively. At the amino acid level, 38 (79.1%) out of 48 amino acid mutations occurred in different immune epitopes within the surface proteins, of which 2 (5.2%) occurred in B cell; 12 (31.5%) in T helper and 24 (63.1%) inside CTL epitopes. There were significant associations between amino acid mutations (especially within immune epitopes) and anti-HBe positivity and increased ALT levels (P values: 0.014 and 0.04, respectively). CONCLUSION: The distribution of amino acid mutations as well as the ratio between silent and missense nucleotide mutations showed that a narrowly focused immune pressure had already been on the surface protein T cell epitopes (94.9% of mutations), particularly CTL epitopes which led to the emergence of escape mutants in these patients.

Validation of an in-vitro method for Hepatitis B vaccine potency assay: specification setting.

AIM: Production batches of the Hepatitis B vaccine should be tested by the National Control Laboratory (NCL) before being released to the market, in terms of their potency. This can be done either by means of the mouse immunogenicity (in-vivo) method, which is a time-consuming and labor intensive process, or by an in-vitro method with acceptable analytical performance and with specifications determined based on the results obtained from testing some production batches of the vaccine with proven efficacy. Here we report the feasibility of using and validation of a commercial enzyme-linked immunosorbent assay (ELISA) kit replacing the manufacturer's method and setting of different specification for potency of the particular vaccine. METHODS: For the in-vitro potency assay of the Hepavax-Gene®, produced by Berna Biotech Korea Corp, a commercial ELISA kit for hepatitis B surface antigen (HBsAg) quantitation (Hepanostika® HBsAg Ultra from Biomerieux) was used to determine the relative potency. Validation parameters were evaluated following the International Conference on Harmonization (ICH) guidelines. Specification of the vaccine potency was determined based on the results generated by the commercial ELISA kit. Some batches were tested by in-vivo method as well. RESULTS: It was confirmed that the ELISA kit, when used for vaccine potency testing, meets the criteria for accuracy (80% to 110% recovery), precision (repeatability, with a CV% less than 5%; and intermediate precision, with a CV% less than 10%) and Linearity (r2> 98%), as well as being able to detect HBsAg specifically. Specification of the in-vitro method was also determined as having a relative potency of >50%. CONCLUSION: The Hepanostika® HBsAg Ultra kit from Biomerieux can be used to determine the relative potency of the Hepavax-Gene® Hep B vaccine as an alternative to the manufacturer's method and with different specifications.

Hepatitis B virus: origin and evolution.

The pathogenesis of hepatitis B virus (HBV) is complex and it appears that molecular variants play a role in this process. HBV undergoes numerous rounds of error prone production within an infected host. The resulting quasispecies are heterogeneous and in the absence of archaeological records of past infection, the evolution of HBV can only be inferred indirectly from its epidemiology and by genetic analysis. This review gathered the controversies about the HBV origin and factors influencing its quasispecies. Also, it provided some evidence on how HBV genotypes correlated with human history and patterns of migration. It is our belief that this topic deserves further attention and thus it is likely that more critical research work will be performed to elucidate the unknown mechanisms and processes in this area.

EMLA cream for carpal tunnel syndrome: how it compares with steroid injection.

BACKGROUND: A standard treatment option for carpal tunnel syndrome (CTS) is local injection of anesthetic-corticosteroid. This clinical trial was designed to compare the safety and efficacy of daily application of the EMLA cream (lidocaine 2.5% plus prilocaine 2.5%) with that of a single injection of methyl prednisolone acetate (Depo-Medrol) 40 mg. METHODS: In this randomized, parallel-group, open-label, single-center, case-controlled, prospective study, 65 participants (70 hands) aged 18-75 years with clinical & electrodiagnostic evidences of CTS were randomized to receive either the EMLA cream (n = 30 patients, 35 hands, group 1) or one injection of methylprednisolone acetate 40 mg at wrist (n = 35 patients, group 2). Outcome assessments included the visual analog scale and clinical assessment. RESULTS: After 4 weeks of treatment, patients in both groups reported significant changes (P < 0.001) in pain intensity. Both treatments were well tolerated, with treatment-related adverse events (AEs) reported in 2 patients in group 1 (5.7%) and 10 patients in group 2 (28.5%) No systemic treatment-related AEs were observed with the EMLA cream. CONCLUSION: EMLA cream was effective in reducing pain associated with CTS and well tolerated and it may offer patients with CTS an effective, noninvasive symptomatic treatment.

Study of sympathetic skin response in visceral leishmaniasis.

BACKGROUND: Neurologic changes in visceral leishmaniasis are rarely reported. Some articles have reported symptoms suggestive of peripheral neuropathy and showed some degree of axonal degeneration and demyelination. The main purpose of the present study was to identify and quantitatively evaluate sympathetic dysfunction in VL. METHOD: Twenty patients with visceral leishmaniasis and 20 healthy controls were studied. All the patients and controls were examined at first and skin sympathetic response was measured in all of the patients and control group by standard protocol. RESULTS: The patients had mean age of 24.2 +/- 17.8 months. The SSR to the electrical stimulus was absent in 10 patients with VL. In four patients all responses were present and, in four patients only one response from hand or foot was present and, in two cases responses were present from both hands. For right median nerve, median latency was 2.4 (min: 1.19, max: 6.92) seconds. CONCLUSION: In conclusion impairment of SSR was demonstrated electrophysiologically in the patients with visceral leishmaniasis.

A new core promoter mutation and premature stop codon in the S gene in HBV strains from Iranian patients with cirrhosis.

In order to define hepatitis B virus (HBV) mutational patterns in Iran, nucleotide sequences obtained from 91 patients and encompassing the precore, basal core promoter (BCP) and surface (S) regions, were compared. The patients were grouped as asymptomatic carriers, chronic active hepatitis or cirrhotic patients. Genotypes and mutations were determined by sequencing and phylogenetic analysis. All strains belonged to genotype D, and most of them to subgenotype D1. All but two strains specified ayw2, one ayw3 and one adw2 determinants. Two deletions of 8- or 20-bp were found in the X region in eight strains, six from patients with chronic active hepatitis. Eight of 21 strains from patients with cirrhosis harboured unusual mutations such as a stop codon at position 69 in the S region or a previously not described mutation in the BCP region ((1761)TC/ATTTG(1766)). All patients infected by strains with the stop codon mutation had detectable HBsAg and high viral load. The accumulation of mutations found in the BCP and S regions in HBV strains from patients with chronic active hepatitis and cirrhosis may predict disease progression in Iranian HBsAg carriers.

Autologous blood injection in carpal tunnel syndrome (CTS).

INTRODUCTION: Most medical treatments for carpal tunnel syndrome (CTS) have focused on suppressing the inflammatory process. An injection of autologous blood might provide the necessary cellular and humoral mediators to induce a healing cascade. The purpose of this study was to evaluate prospectively autologous blood injection in CTS. METHOD & MATERIALS: Twenty patients with CTS were recruited using strict inclusion and exclusion criterias. All patients were injected with 1 ml of autologous blood and 0.5 ml lidocaeine 1% under the carpal tunnel. Pain intensity (based on VAS) and electrophysiologic parameters of median nerve (transcarpal SNCV, DML and DSL) were recorded before and 3 weeks after autologous blood injection. RESULTS: The average pain scores before and 3 weeks after autologous blood injection was 8.70 +/- 0.92 and 4.30 +/- 0.76 respectively (P < 0.005). Also transcarpal SNCV of median nerve was 33.7 +/- 6.3 m/s and 24.5 +/- 6.8 m/s (P = 0.001); DML of medin nerve was 5.16 +/- 1.04 ms and 4.70 +/- 0.53 ms (P = 0.001) and DSL of median nerve was 4.84 +/- 0.77 ms and 4.2 +/- 0.6 ms (P = 0.001), respectively. CONCLUSIONS: After autologous blood injection, pain intensity and electerophysiologic parameters were significantly improved. This study offers encouraging results for an alternative minimally invasive treatment for CTS. This study cannot prove conclusively whether the blood itself induced an inflammatory cascade or the injury created by the injection was responsible.

HBV core sequence: definition of genotype-specific variability and correlation with geographical origin.

There are eight genotypes and nine subtypes of HBV. Small differences in geographical origin are associated with sequence changes in the surface gene. Here, we compared core gene sequences from different genotypes and geographical regions. Specific combinations of 24 amino acid substitutions at nine residues allowed allocation of a sequence to a subtype. Six of these nine residues were located in different T cell epitopes depending on HBV geographical area and/or genotype. Thirty-seven nucleotide changes were associated uniquely with specific genotypes and subtypes. Unique amino acid and nucleotide variants were found in a majority of sequences from specific countries as well as within subtype ayw2 and adr. Specific nucleotide motifs were defined for Korean, Indian, Chinese, Italian and Pacific region isolates. Finally, we observed amino acid motifs that were common to either South-east Asian or Western populations, irrespective of subtype. We believe that HBV strains spread within constrained ethnic groups, result in selection pressures that define sequence variability within each subtype. It suggests that particular T cell epitopes are specific for geographical regions, and thus ethnic groups; this may affect the design of immunomodulatory therapies.