HbS Gene Research Articles

Determination of birth prevalence of sickle cell disease using point of care test HemotypeSC™ at Rundu Hospital, Namibia

BackgroundSickle cell disease (SCD), a noncommunicable disease, has the greatest burden in sub-Saharan Africa. The majority of children (50–90%) with SCD die before their 5th birthday, with approximately 150,000–300,000 annual SCD child deaths in Africa. In developed countries, newborn screening (NBS) has been shown to improve the survival of children with sickle cell disease, with under5 childhood mortality reduced tenfold due to interventions performed before the development of complications. Point –of-care tests have been developed for resource limited settings to expand NBS. The aim of this study was to determine the birth prevalence of sickle cell disease in Namibia using the HemoTypeSC™ point-of-care test.MethodsA cross-sectional descriptive study was carried out at Rundu Intermediate Hospital in the Kavango East Region. Two hundred and two (202) well newborns within 72 h of birth were recruited for the study from 22 February to the 23th March 2023. Descriptive statistics were used to compute the haemoglobin types of the study participants.ResultsThe majority of the participants (n = 105, 52%) were females, and (n = 97,48%) were males. The median age of the participants was 23 h (Q1, Q3; 11; 33),) with an age range of 2–98 h. Sickle cell trait was present in 9.4% of the screened newborns, no homozygous disease was detected, and 90.6% had Hb AA.ConclusionsThis study is the first to measure HbS gene carriage at birth using HemotypeSC point-of-care testing in Namibia. There was a moderate prevalence of sickle cell traits but no SCD. This baseline study may provide the foundation for larger epidemiological surveys to map HbS gene carriage in Namibia to provide evidence for policy makers to fashion appropriate SCD newborn screening services.

Prevalence of occult hepatitis B virus infection and characterisation of hepatitis B surface antigen mutants among adults in western Croatia

Introduction and ObjectivesOccult hepatitis B virus (HBV) infection (OBI) is characterised by low levels of hepatitis B virus (HBV) DNA in the blood/liver of patients with negative hepatitis B surface antigen (HBsAg). This study aimed to determine the OBI prevalence and virological characteristics (viral genotypes and HBsAg mutants) in patients with an “anti-HBc only” serological profile. Materials and MethodsA total of 24 900 serum samples were routinely screened for hepatitis B markers over a five-year period. All anti-HBc-positive/HBsAg-negative/anti-HBs-negative sera were selected and analysed for the presence of HBV DNA. Mutational analyses of the HBs gene and polymerase gene sequences were performed. Results1749 (7.02%) sera were anti-HBc positive, and 113 (0.45%) sera had an “anti-HBc only” serological profile (HBsAg/anti-HBs negative). HBV DNA was detected in 12/113 (10.61%) “anti-HBc only” positive sera, representing 0.048% of all routinely tested samples. Due to extremely low viremia, HBV genome was successfully sequenced in only two sera where subgenotype D3 was confirmed. Mutational analyses of the S gene revealed multiple missense mutations. In addition to the M133I, Y134F, and G145R mutations, already associated with diagnostic escape, we also found nine novel OBI-related S-gene mutations - S136Y, F158L, K160N, E164G, S167L, A168V, L175S, S210I and F212C. ConclusionsWe detected multiple known and novel S gene mutations in 2/12 (16.6%) OBI cases, nevertheless, further studies are required to determine their role in the pathogenesis of OBI. Understanding the frequencies of clinically relevant HBV mutations may contribute to improvement of diagnostic protocols.

Prevalence of Hepatitis B Virus among Renal Dialysis Patients in Diyala Province, Iraq

Background: One of the main public health challenges in the globe is the prevalence of the hepatitis B virus (HBV). Objectives: The curent study aimed to determine the prevalence of hepatitis B virus in renal dialysis patients in Diyala province, Iraq. Methods: Ninety of renal dialysis patients attended Ibn-Sina Dialysis Center were screened for HBsAg, HBc IgG, HBc IgM and HBe Ab using ELISA technique, and then positive results were retested by conventional PCR for detection of HBs gene and HB core gene. Demographic data of study population were recorded. Simple statistical analysis was done using SPSS Version 25. Results: Among studied samples, the seroprevalence of HBsAg, Anti-HBc Ab, Anti-HBc IgM and Anti-HBe Ab were 4.4%, 17.8%, 2.2% and 5.6% respectively. The results of conventional PCR showed that only 1(1.1%) of HBs gene at 417 bp in renal dialysis patients were positive. Whereas, the HBc gene at 791 bp was not detected in all included study population. Conclusion: The prevalence of HBV was relatively high among renal dialysis in Diyala province/Iraq.

A Novel Insertion in the Hepatitis B Virus Surface Protein Leading to Hyperglycosylation Causes Diagnostic and Immune Escape.

Chronic hepatitis B virus (HBV) infection is a global health threat. Mutations in the surface antigen of HBV (HBsAg) may alter its antigenicity, infectivity, and transmissibility. A patient positive for HBV DNA and detectable but low-level HBsAg in parallel with anti-HBs suggested the presence of immune and/or diagnostic escape variants. To support this hypothesis, serum-derived HBs gene sequences were amplified and cloned for sequencing, which revealed infection with exclusively non-wildtype HBV subgenotype (sgt) D3. Three distinct mutations in the antigenic loop of HBsAg that caused additional N-glycosylation were found in the variant sequences, including a previously undescribed six-nucleotide insertion. Cellular and secreted HBsAg was analyzed for N-glycosylation in Western blot after expression in human hepatoma cells. Secreted HBsAg was also subjected to four widely used, state-of-the-art diagnostic assays, which all failed to detect the hyperglycosylated insertion variant. Additionally, the recognition of mutant HBsAg by vaccine- and natural infection-induced anti-HBs antibodies was severely impaired. Taken together, these data suggest that the novel six-nucleotide insertion as well as two other previously described mutations causing hyperglycosylation in combination with immune escape mutations have a critical impact on in vitro diagnostics and likely increase the risk of breakthrough infection by evasion of vaccine-induced immunity.

Prevalence of Gastrointestinal Parasites among Sickle Cell Patients in Delta State, Nigeria

Sickle cell anemia crisis and death in the under-developed nations constitute one of the most challenging global public health problems. It is estimated that each year out of the three hundred thousand babies born worldwide with severe forms of hemoglobin disorders, sickle cell disease constitute more than half of this figure. Sickle cell disorder is an abnormal hemoglobin condition caused by the inheritance of abnormal hemoglobin (Hbs) genes from both parents. This study was aimed at determining the prevalence of gastrointestinal parasites among sickle cell subjects in Delta State. Stool samples were collected from a total of 320 subjects, 160 each from Delta South (Warri) and Delta North (Agbor). Questionnaires and informed consent forms were given to the subjects that are of age and to the parents of the minors before sample collection. The stool samples were examined both macroscopically and microscopically for gastrointestinal parasites .InWarri, 40% (64/160) of SCA subjects infected with either gastrointestinal heamoparisites were male, while 48.1% were females. In Agbor, 47.5% were males, while 43.8% were females. This was statistically significant (p< 0.05). The greatest frequency of infection was observed in SCA subjects within the age bracket of 1-10. (42.5%) in warri and 56.2%atAgbor. In Warri A.lumbricoides, hookworm, T.trichuria, E coliand malaria parasites were observed, 18.1%, 5.6%, 2.5%, 1.3% and 60.6% respectively among sickle cell subjects infected with either gastrointestinal haemoparasites. No microfilaria was isolated among subjects in warri, but was isolated at Agbor (0.6%).Also at Agbor, A. lumbricoides, hookworm, T.trichuria, E. coli,and malaria parasites had prevalence of16.8%, 2.5%, 1.3%, 1.9% and 68.1%respectively. In Warri, SCA subjects without parasitic infection have a mean PCV of 23.27% as against those with parasitic infection having a mean PCV of 12.88%. At Agbor, those without infection have a mean PCV of 22.84% as against 19.72% among those with parasitic infection. Among SCA subjects with either gastrointestinal or haemoparasites in warri, 24.4%, 50.0% 13.7% resides in rural, urban and riverine Communities respectively. At Agbor 60%, 31.3% resides in rural & urban communities respectively. The prevalence of infection among those who deworm always, occasionally only and those who never deworm were 32.5%, 53.1% and 2.5% respectively in Warri and, 10%, 60% & 21.3% respectively in Agbor. All sampled SCA subjects received blood transfusion always or occasionally as 70.6% and 17.8% respectively in Warri, as 85.0% and 6.3% respectively in Agbor. Prevalence of infection among subjects who use water closet and latrine were 18.7% and 6.9% respectively. Subjects who never wash their hands before and after eating have prevalence of 7.2% while those who wash their hands before and after eating always were 18.4%. The prevalence of malaria parasites 60.6% and 66.8%, Warri and Agbor respectively. Overall, the prevalence of parasitic infections among SCA subjects in Warri was 88.1%. Of this 48.1% were females.SCA subjects between the ages of 1-10 were mostly infected. A. lumbricoides (18.1%) was the gastrointestinal parasite mostly isolated. This study concludes that prevalence of gastrointestinal and haemoparasite among sickle cell subjects is very high in Delta State. Therefore, the need for all sickle cell anemic subjects to strictly use insecticide treated nets is highly advocated.

Lipids and Sickle Cell Disease

Sickle cell disease is a hereditary, autosomal recessive pathology caused by the replacement of adenine by thymin in position 6 of the beta globin gene, producing anomalous hemoglobin (hemoglobin S- HbS). It is characterized by a chronic inflammatory state with hemolytic anemia and vaso-occlusive phenomena and occurs when the HbS gene is in homozygosis (SS) or heterozygosis, in association with other variant hemoglobins (SD, SC, SE) or thalassemia (). Sickle cell disease is recognized by the World Health Organization (WHO) [...]

Relationship between malaria parasitemia and haemoglobin variants in patients attending Rivers State University Teaching Hospital, Port Harcourt, Rivers State, Nigeria

Malaria parasitemia and haemoglobin variants in patients attending Rivers State University Teaching Hospital, Port Harcourt, Rivers State, Nigeria was investigated. Blood samples from 385 patients attending the hospital were collected using sterile syringes into well labelled specimen bottles containing ethylene di-amine tetra acetate (EDTA). The genotype was determined using standard alkaline cellulose acetate electrophoretic technique while standard method was used to determine malaria parasitaemia. Out of the 385 patients examined for haemoglobin variants, the frequency of the genotype was 239(62.1%), 127(33%), 17(4.4%) and 1(0.3%) for HbAA, HbAS, HbSS and HbAC respectively. Malaria parasitemia in HbAA, HbAS, HbSS and HbAC was 170(64.2%), 82(30.9%), 13(4.9%) and 1(0%) respectively. HbAA was significantly (P<0.005) susceptible to malaria than other genotypes. The prevalence of malaria in HbAS 82(30.9%), HbSS 13(4.9%) and HbAC 1(0%) also show significance difference (P<0.05). Patience below the age of 6years with HbS gene had the highest malaria prevalence (21%). Malaria prevalence among males 7(53.8%) and females 6(46.2%) with sickle cell anaemia SCA had no significance difference (p>0.05). Sickle hemoglobin (HbS) gene protection against malaria found in heterozygous hemoglobin (HbAS) was also recorded in patients with homozygous (HbSS) gene. Prevalence of malaria is higher in patients with the normal haemoglobin (HbAA) than in patients with abnormal genes (HbAS and HbSS).

Investigation of the Effect of High-Intensity Interval Training (HIIT) on the Expression of the Genes miR-126, miR-296, HGS and VEGF-A Protein Levels in Tumor Tissue in Female Mice

Background & Objective: The studies of the last two decades have shown that regular training is associated with the reduction of mortality among patients with breast cancer, and has an important role in inhibiting breast cancer progression as well as treatment of the disease. However, the micro-molecular mechanisms involved are not fully understood yet. However, the aim of the present study was to investigate the effect of a high-intensity interval training (HIIT) on the expression of miR-126, miR-296, HGS gene, and VEGF-A protein level in tumor tissue of the mice with breast cancer. Materials & Methods: For this purpose, 12 BALB/c mice (6-8 weeks, weight 19 ± 1.05 g), after induction of cancer (MC4-L2 subcutaneous injection into the right side of the mice), were randomly divided into two groups with 6 in each group: HIIT and control. Each session of HIIT group program includes six intervals of three minutes and 20 seconds with an intensity of 85 to 90 percent of VO2max and 1-minute recovery with 30 to 35 percent VO2max intensity between each interval; and the animals were sacrificed 24 hours after the last training session and the expression of miR-126, miR-296, HBS gene and VEGF protein levels in tumor tissues were examined. Results: The statistical results showed that the implementation of HIIT significantly reduced the expression of miR-296 and subsequently increased the HGS gene expression and led to increased expression levels of miR-126 and decreased levels of VEGF-A protein in the training group compared to the control group (p<0.05). Conclusion: It seems that HIIT, by suppressing tumor angiogenesis signaling pathways, could be an effective intervention to inhibit the growth of breast tumors.

Newborn Screening for Sickle Cell Disease Among Tribal Populations in the States of Gujarat and Madhya Pradesh in India: Evaluation and Outcome Over 6 Years.

Sickle cell disease (SCD) poses considerable public health problems in India. This study was undertaken to understand the clinical course of SCD among children identified during newborn screening programmes in Gujarat and Madhya Pradesh where the frequency of the HbS gene is high. A total of 8,916 newborn babies 8,411 from Gujarat and 505 from Madhya Pradesh were screened over 6 years (2010–2016) using HPLC and the diagnosis was confirmed by molecular analysis in a subset. A total of 128 babies (122 Gujarat, 6 Madhya Pradesh) were identified with sickle cell disease, of whom 87 (69 HbSS, 18 HbS-β thalassemia) from Gujarat were followed for 0.5–6.6 years. Acute painful events, severe anemia and fever with infections were the major complications and 23 babies required hospitalization. Severe to moderate clinical presentation was found in 13.8% babies with SCD whereas, 86.2% babies had a milder presentation. Presence of ameliorating factors (α-thalassemia and Xmn 1 polymorphism) did not have a discernible effect on the clinical severity. Parents of babies with SCD were educated and counseled for home care. Distribution of mobile phones to 44 families having babies with SCD was beneficial as it allowed regular contact with patients and their families. Genetic counseling to the affected families has increased the awareness and acceptance for prenatal diagnosis and 18 couples opted for prenatal diagnosis in subsequent pregnancies. SCD is not always mild among tribal groups in India. Therefore, facilities for early diagnosis and prophylactic treatment in the tertiary care centers should be made available. The difficulties in regular follow up of the babies in remote rural areas have also been highlighted.

Status of erythrocyte genetic disorders in people of desert and humid environments in Rajasthan, India: focus on natural selection in tribals against malaria

In India, Rajasthan is the largest state having two distinct environments separated by presence of Aravali Mountains. One is desert, located in the west region and other is humid in the east region where malaria is endemic. In Rajasthan, various inherited erythrocyte genetic disorders, abnormal haemoglobins (Hbs), thalassaemia (thal) and glucose-6-phosphate dehydrogenase enzyme deficiency (Gd) have been extensively studied in people of scheduled tribe (ST), scheduled caste (SC), general caste (GC) and other backward class (OBC) populations residing in both environments. In these populations, abnormal haemoglobin variants, Hb-S, Hb-D, Hb-E, Hb-J, Hb-C and Hb-H in heterozygous (trait) and/or homozygous state with varying incidence have been detected. Among these Hb variants, sickle cell Hb in trait form (Hb-AS) is common and widely distributed in diverse populations. Genes of β-thalassaemia trait (0.0–9.0%) are more prevalent compared to α-thalassaemic (1.88–3.07%) genes and are also widely distributed. Gd disorder is also common in occurrence and widely distributed with varying incidence (0.64–22.0%) in people of both environments. Relatively a high incidence of Hb-S (31.14%), β-thalassaemia (9.00%) and Gd (22.00%) genes reported in tribals of southern Rajasthan. In present communication, status of various erythrocyte genetic disorders in people of both environments, natural selection in tribals against malaria, mitigation and control of these genetic disorders and effect of fluoride exposure in tribals having sickle cell and β-thalassaemic genes are considered and critically reviewed. These findings may be useful in framing and implementation of health policies for the mitigation and control of erythrocyte genetic disorders in tribal communities.

Detection of Q129H Immune Escape Mutation in Apparently Healthy Hepatitis B Virus Carriers in Southwestern Nigeria.

As the global effort to eradicate hepatitis B continues, immune escape mutations (IEMs) and drug resistance mutations (DRMs) affecting its diagnosis, treatment, and prevention are compromising this goal. However, knowledge about the prevalence and circulation of these mutations in Nigeria is scarce. Serum samples (n = 199) from apparently healthy prospective blood donors, pregnant women, and individuals presenting with fever in southwestern Nigeria were analyzed for the presence of IEMs and DRMs by means of nested PCR in the HBV S (HBs) and HBV polymerase (Pol) genes, followed by phylogenetic and mutational analyses. In total, 25.1% (n = 50/199) of samples were positive for HBV, as measured by PCR. In 41 samples (20.6%), both fragments could be amplified, whereas the HBs gene and the Pol gene fragment alone were detected in 0.5% (n = 1/199) and 4% (n = 8/199) of samples, respectively. Sequences were successfully obtained for all 42 HBs gene fragments but for only 31/49 Pol gene fragments (totaling 73 sequences from 44 individuals). All sequences were identified as HBV genotype E. IEMs were present in 18.2% (n = 8/44) of the sequences of HBV-positive individuals with available sequences. IEM Q129H was detected in eight out of the 44 (18.2%) HBV isolates sequenced in this study; however, no DRMs were observed. This study confirms the circulation of HBV IEMs and reports the presence of Q129H IEM for the first time in Nigeria. Intensified research on the dynamics of IEM is necessary in order to enhance the elimination of HBV.

HbA2 induction: the merit of pancellularity in sickle cell disease.

HbA2 induction: the merit of pancellularity in sickle cell disease.

Surgical plume from tissue infected with human hepatitis B virus can contain viral substances

Introduction Evidence on the biological danger associated with surgical plume is lacking. We examined whether surgical plume, generated by the energy devices ultrasonically activated scalpel (US) or electrocautery (EC) contains virus-related substances. Material and methods Experiment 1, ex-vivo model: Tumor mass of a hepatocellular carcinoma line was prepared in a Nod/SCID mouse. Surgical plume generated on the mass by US or EC was collected and detection of HBs gene fragment and antigens (HBsAg or AFP) was conducted. Experiment 2, clinical specimen: Detection of HBV-DNA and HBsAg was conducted following the collection of surgical plume generated from clinically obtained liver specimens from six HBV-associated hepatocellular carcinoma patients. Results Experiment 1: HBs gene fragment was detected in the solutions regardless of the device used. HBsAg was detected in US and EC solutions and AFP was also detected in a US solution. Experiment 2: HBV-DNA was detected in both devices, in all three cases whose preoperative serum HBV-DNA was positive. In the other serum-negative cases, HBV-DNA was not detected. While serum HBsAg was positive in five of six cases, it was not detected in any solution. Conclusions DNA fragments or antigens of virus can exist in the surgical plume generated by EC or US.

Novel mechanism of hereditary pyropoikilocytosis phenotype due to co‐inheritance of β globin and α spectrin mutations

Novel mechanism of hereditary pyropoikilocytosis phenotype due to co‐inheritance of β globin and α spectrin mutations

Pregnancy-Related RBC Alloimmunization in Sickle Cell Trait Patients; A Single-Center Experience

Abstract Background Sickle Cell Disease (SCD) is an autosomal recessive disorder, which results from a point mutation in the β-globin gene. The production of mutant Hb S (V6E) leads to hemolytic anemia and numerous clinical complications. Patients homozygous for HbS gene (SS) typically rely on life-long transfusion therapy. Alloimmunization to foreign RBC antigens in multiply transfused patients is significantly more frequent in SS patients (observed in 7–47%) compared to patients with HbA (AA), and can pose a significant hurdle in finding compatible RBCs. However, the mechanism for enhanced alloimmunization in SS patients has not yet been elucidated. Patients heterozygous for HbS (SA) have a no distinct clinical phenotype and do not typically require transfusions. We investigated the rate of RBC antigen alloimmunization and specificity in SA patients. Aim The aim of this study was to determine and compare the rate and specificities of RBC antigen pregnancy-related alloimmunization in SA and AA patients. Methods In our retrospective study, we identified females who delivered a newborn between January 2014 to October 2019, for whom prenatal hemoglobin electrophoresis testing, and prenatal and perinatal antibody screening and identification was performed. R (V.3.6.2) statistical computing program was used for analysis. Results A total of 41735 subjects were identified: 40058 (AA) and 1677 (SA). African Americans were more prevalent in the SA compared to the AA group (55.8% vs 29.4%, P<0.01)and were more likely to have received ≥ 1 transfusion during pregnancy (4.5% vs 3.3%, P<0.01). A total number of 267 alloantibodies were detected in 246 patients (0.6% of the studied population) during pregnancy, 228 patients (0.6%) in the AA group and 18 patients (1.1%) in the SA group.229/246 (93%) of patients with antibodies did not receive transfusion during pregnancy, and thus antibodies were considered to represent pregnancy-related alloimmunization. In non-transfused subjects the SA group were more likely to have developed alloantibodies in pregnancy compared to the AA group [OR= 1.94, P=0.015]. The median age of patients with antibodies in the SA was older [33.5 vs 27 (p=0.008)] as compared to those in the AA group [29 vs 27 (p=0.002)]. The most common antibody identified was anti-E (SA 42% vs AA 28%), anti-Lea (SA 21% vs AA 13%) and anti-C (SA 5% vs AA 12%). Subjects with pre-existing alloantibodies had a 5.4x increased risk of forming a new allo-antibody. However only 0.1% of subjects had a history of prior allo-antibodies, with no significant difference identified between AA and SA. Conclusions SA patients were found to have an increased rate of pregnancy-related alloimmunization (OR 1.75, P=0.02) after adjusting for the effects of increasing age, RH type, ethnicity, number of previously existing antibodies and number of transfusions in pregnancy.

Association of HBsAg mutation patterns with hepatitis B infection outcome: Asymptomatic carriers versus HCC/cirrhotic patients

Introduction and ObjectivesThe hepatitis B virus (HBV) surface antigen (HBsAg) variations suggested having some effects on infection outcome. Due to some controversial issues, the aim of this study was to compare the pattern of HBsAg variation between asymptomatic carriers and HCC/cirrhosis patients. Materials and methodsIn this cross-sectional study, 19 HCC/cirrhotic and 26 asymptomatic patients were enrolled. After viral DNA extraction, HBs gene was amplified using an in-house nested-PCR. Then, PCR products were introduced into bi-directional Sanger sequencing. The retrieved sequences were compared with references, to investigate the variation of immunologic sites, major hydrophilic region (MHR) of HBsAg as well as reverse transcriptase (RT), and also to determine genotype/subtype. ResultsThe analysis of MHR and epitopes on HBsAg showed dozens of substitution, which occurred more prevalently in I110, P120, Y134, G159, S193, Y206, S207, I208, L213 and P214 positions. However, Y134N/F/L (P=0.04) and P120T/S (P=0.009) were significantly detected in MHR and B-cell epitope of HCC/Cirrhotic group. A number of truncation-related mutations were higher in HCC/Cirrhotic group (P>0.001), albeit only C69* stop codon was statistically significant (P=0.003). In RT, some potentially resistant substitutions such as Q215S, V191I and V214A, were revealed. Phylogenetic analysis showed that all of isolates belonged to genotype D, and the major serotype was ayw1. ConclusionThe higher frequency of substitutions in MHR and immune epitopes at positions such as Y134 and P120 as well as stop codons such as C69* in HCC/cirrhotic group might candidate them as predictive factors for infection outcome.

Malaria and the Sickle Gene: Polymorphism Balance in favour of eradication

Evolutionally, the single nucleotide mutation responsible for the sickle haemoglobin gene, (HbS gene) developed from the regions of the world where malaria is holoendemic, leading to the explanation that the mutation is in response to the presence of the malaria parasite. Studies eventually showed that individuals that are heterozygous for the HbS gene are protected from the lethal clinical effects of malaria infection. In other words, malaria confers a survival advantage to carriers of the HbS gene, and this is referred to as balanced polymorphism. On the other hand, malaria infection is associated with significant morbidity and mortality, particularly among children. Unfortunately, lack of success in the effort to eradicate the malaria parasite through the elimination of the Anopheles mosquito or efforts to limit its contact with human being has produced little success, hence the resort to roll back malaria, with the aim of reducing the morbidity and mortality associated with it. Therefore, it is attractive to consider what will happen to the sickle gene if malaria were to be eradicated. That is, the possibility is that the sickle gene may also follow suit since its evolution in the first instance was as a response to malaria., This is the hypothesis being propounded by this article. In that case, rather than shy away from the malaria eradication initiative, it should remain the goal of all malaria programs.

Spectrum of Hemoglobinopathies: A New Revelation in a Tertiary Care Hospital of Odisha.

The prevalence of different types of hemoglobinopathies and its spectrum in Odisha state is believed to be high, but its exact prevalence is not known due to lack of large population based study. The present study was undertaken to know the magnitude and spectrum of hemoglobinopathies among the patients attending tertiary care centre for evaluation of anemia. All the patients of various age group without any history of blood transfusion preceding 3months of period attending the Clinical Hematology Department of SCB Medical College, Cuttack for evaluation of anemia were included in this 10year prospective study. Detail history, clinical examination followed by blood sample examination including by HPLC/CzE were done in all cases. Other investigations were done as per need of evaluation of anemia. Out of 21,371 patients with anemia, hemoglobinopathies was detected in 10,745 (50.2%) cases. The profile of hemoglobinopathy was as follows: HbS gene in 52.48% cases, betathalassemia in 54.06% and HbE hemoglobinopathies in 9.19% cases. Hemoglobinopathy was detected in very high percentage (50.2%) of cases in our centre. Various types of β-thalassemia and sickle cell hemoglobinopathies were two major types (54.06% and 52.48% respectively). This needs to be confirmed by large population based study.

Different types of viral‑host junction found in HBV integration breakpoints in HBV‑infected patients.

The present study surveyed the characteristics of hepatitis B virus (HBV) integration in the liver genomes of patients with acute hepatitis B (AHB), carriers of inactive hepatitis B surface antigen (HBsAg), and patients with chronic hepatitis B (CHB) receiving antiviral treatment. ‘Short-read’ whole genome sequencing (WGS) with an average of 4,879× coverage for HBV integration was performed in three patients with AHB, two carriers of inactive HBsAg, and 13 patients with CHB receiving antiviral treatment. Conventional polymerase chain reaction and Sanger sequencing were used to verify integration breakpoints supported by at least two paired-end reads, and viral-host chimeric transcripts were surveyed simultaneously. HBV integration breakpoints were 100% identified with an average of 138.2±379.9 breakpoints per sample. The numbers of HBV integration breakpoints were positively associated with the sequencing depth coverage numbers and levels of intrahepatic covalently closed circular DNA, respectively (P<0.0001 and P<0.0001). Four types of viral-host junction in 14 HBV integration breakpoints were detected (two viral junctions mapped in the HBs gene, one in the Precore gene, and others within the HBx gene): Forward simple junction, reverse simple junction, forward and reverse complicated junction, and microhomology were found in many of the junctions. Expression of viral-human chimeric transcripts was observed in several breakpoints, including the HBs gene. As a result, HBV can integrate into the host gene in the same manner as non-homologous end joining and microhomology-mediated end joining with numerous sites, and a close association may exist between HBV integration and patient prognosis. HBx integration may be indispensable for viral-host chimeric transcription and HBsAg may be produced from integrated DNA.

Pre-S2 Start Codon Mutation of Hepatitis B Virus Subgenotype B3 Effects on NF-κB Expression and Activation in Huh7 Cell Lines.

A cross-sectional study on hepatitis B patients in Indonesia showed association of pre-S2 start codon mutation (M120 V) with cirrhosis and hepatocellular carcinoma (HCC), which was dissimilar from studies from other populations where pre-S2 deletion mutation was more prevalent. Different mutation patterns were attributed to different hepatitis B virus (HBV) subgenotypes in each population study. HBV surface proteins are reported to induce the activation of NF-κB, a transcriptional factor known to play an important role in the development of liver disease. This study aimed to see the effects of HBs variants in HBV subgenotype B3 on the expression and activation of NF-κB as one of the mechanisms in inducing advanced liver disease. HBV subgenotypes B3, each carrying wild-type (wt) HBs, M120 V, and pre-S2 deletion mutation were isolated from three HCC patients. HBs genes were amplified and cloned into pcDNA3.1 and were transfected using Lipofectamine into a Huh7 cell line. NF-κB activation was measured through IκB-α expression, which is regulated by NF-κB. RNA expressions for HBs, IκB-α, and NF-κB subunit (p50) were evaluated using real-time PCR. M120 V mutant had a significantly higher mRNA level compared with wt and pre-S2 deletion mutant; however, there were no significant differences in HBs protein expressions. The transcription level of p50 was higher in M120 V mutation compared with HBs wild-type and pre-S2 deletion mutant. NF-κB activation was higher in HBs wild-type compared with the two mutant variants. Pre-S2 mutations had no effect on the increment of NF-κB activation. However, M120 V mutation may utilize a different pathway in liver disease progression that involves high expression of NF-κB subunit, p50.