Wardah Yusof

The Global Globin Network (GGN) is a project-wide initiative of the Human Variome/Global Variome Project (HVP) focusing on haemoglobinopathies to build the capacity for genomic diagnosis, clinical services, and research in low- and middle-income countries. At present, there is no framework to evaluate the improvement of care, treatment, and prevention of thalassaemia and other haemoglobinopathies globally, despite thalassaemia being one of the most common monogenic diseases worldwide. Here, we propose a universally applicable system for evaluating and grouping countries based on qualitative indicators according to the quality of care, treatment, and prevention of haemoglobinopathies. We also apply this system to GGN countries as proof of principle. To this end, qualitative indicators were extracted from the IthaMaps database of the ITHANET portal, which allowed four groups of countries (A, B, C, and D) to be defined based on major qualitative indicators, supported by minor qualitati...

Accurate diagnosis to limit the spread of SARS-CoV-2 is crucial for the clinical management of this lethal infection. Recently, many low-cost and easy-to-use rapid test kits (RTK) have been developed in many countries for the massive screening of SARS-CoV-2. Thus, evaluating the accuracy and reliability of an RTK is critical. The current study was conducted on 157 individuals to evaluate the performance accuracy of rapid SARS-CoV-2 antigen detection kits using different clinical samples compared with qRT-PCR results. Nasopharyngeal swabs were collected from patients for qRT-PCR and RTK tests, and then buccal and nasal, and nasal swabs were collected for RTK tests separately. The nasal and buccal swabs showed high sensitivity (98%) and specificity (100%) compared with the qRT-PCR results. Meanwhile, for nasal, the sensitivity was 96% with 98% specificity, and nasopharyngeal swabs showed 98% sensitivity and 94% specificity. Fisher’s exact test revealed statistical significance (p <...

CYP2A6 gene encodes the principal enzyme involved in the metabolism of many drugs including artesunate. We developed a simplified duplex nested PCR method for the detection of the CYP2A61B, CYP2A62, CYP2A64, CYP2A67, CYP2A68 and CYP2A69 variant alleles highly prevalent among Malaysian population. Genomic DNA was isolated from peripheral blood of patients treated with artesunate by using a DNA extraction kit. Fragments from TATA box in 5' flanking region, exons 1 to 4 and exon 8 to 3' UTR of the gene were amplified using a first PCR step (PCR1). Products from PCR1 were then used to run a duplex PCR (PCR2) for detecting the selected variant alleles. All 6 CYP2A6 alleles were successfully amplified and confirmed by sequencing. The frequency of the alleles were 14.6% for CYP2A61B, 16.7% for CYP2A64, 4.2% for CYP2A68 and 10.4% for CYP2A69. High frequencies of the CYP2A64 and CYP2A69 alleles which contribute to the poor metabolism status may impact treatment of patients receiving drugs metabolized by CYP2A6 such as artesunate among Malaysians.

Artesunate (AS) and amodiaquine (AQ) are two prodrugs widely used as antimalarial agents and are metabolized by the CYP P450 2A6 (CYP 2A6) and CYP P450 2C8 (CYP 2C8) enzymes, respectively. In this study, we aim to investigate the association of both genes on AS and AQ's tolerabilities in the hope of identifying a pharmacogenetic approach that could be useful in prediction and prevention of adverse drug reactions (ADRs) among Malaysian population. In this randomized crossover study, loose and AS/AQ formulations were administered to normal healthy volunteers (n = 24) over two study phases. The drugs' tolerabilities (incidence of facial flushing, giddiness, headache, nausea, abdominal discomfort, progression of liver enzymes and neutrophil counts) were compared between the two treatment arms. Volunteers were also genotyped for the CYP2C8 and CYP2A6 variants. The frequency of the CYP2A6*1B, CYP2A6*4, CYP2A6*8 and CYP2A6*9 alleles were 54.2%, 16.7%, 4.2% and 10.4%, respectively. No mutations for CYP2C8 gene were, however, detected. Most (96%) of the subjects were of the Malay ethnicity. Subjects having the CYP2A6*1B variants responsible for ultra rapid metabolism of AS suffered a significantly higher incidence of ADRs. Our study is the first to report that CYP2A6 genotyping influences AS's ADR. Gender also plays a role where females reported more incidences of nausea (p < 0.05). It is concluded that genetic polymorphisms of CYP2A6 as well as gender influence the side effect profiles of subjects receiving AS among this Malaysian population.

Abstract. Cytochrome P450 (CYP) 2C8 plays a significant role in metabolizing many clinically important drugs such as paclitaxel, amiodarone and amodiaquine. A simultaneous detection of CYP2C8*2, CYP2C8*3, CYP2C8*4 and CYP2C8*5 alleles by a PCR method will be useful for detecting single nucleotide polymorphisms (SNPs) on the CYP2C8 gene that may contribute to the inter-individual variability in metabolizing these drugs. This is a blinded randomised cross over study conducted in 24 healthy normal volunteers who received the anti-malarial drug amodiaquine and artesunate both as individual agents and in a fixed combination, over two study phases to determine the safety profile of the two preparations. Genomic DNA was isolated from the volunteers using a DNA extraction kit. A nested PCR method was applied to detect all of the CYP2C8 variants. Regions from exons 3, 5 and 8 of the gene were simultaneously amplified using a first PCR step (PCR1). Products from PCR1 were then used to run the...

Application of genome editing have garnered a lot of attention in the scientific world. Its ability to make permanent and precise modification or ‘edits’ in the genomic DNA sequences have open up the possibility of probing the functionality of genes and correcting abnormal genes to treat genetic diseases. The CRISPR/Cas9 which was adapted from Type II bacterial adaptive immune system is the most applied genome editing system due to the ease and efficiency of the system in customising the nucleases needed in editing the targeted sequences. However, the downside of this system is the high frequency of ‘off-target’ editing caused by the nature of the Cas9 protein that can tolerate mismatches between customised nucleases; sgRNA, and the genomic sequences. Despite the set-back, researchers continue applying CRISPR/Cas9 system to edit the genome by minimising the frequency of ‘off-target’ edits. Application of the improved system had successfully produced numerous results in the initial t...

Gene modifiers XmnI have significant association with disease severity in HbE/β-thalassemia patients; however there is no previous study on its association with health related quality of life (HRQoL). The aim of this study is to determine factors affecting HRQoL among transfusion-dependent β-thalassemia and HbE/β- thalassemia adolescents and its association with Xmn1 gene modifier. A cross-sectional study was conducted among transfusion-dependent β-thalassemia adolescents from government hospitals in East Coast Malaysia. HRQoL scores were measured by using the translated PedsQL 4.0 Generic Core Scale while blood was obtained for genetic modifier analysis. The data were analysed using SPSS version 24.0 and its associated factors were analysed using multivariate regression analysis. A total of 151 patients were recruited. The  lowest and highest QOL mean score was school function at 59.69 (16.23) and social function at 77.33 (18.87), respectively. Age of first blood transfusion and Xm...

HbE/β-thalassemia is the most common form of β-thalassemia in Asia. It has various clinical manifestations ranging from very mild to severe. The most reliable and predictive factor of the disease phenotype is the nature of the β-globin gene mutation. Therefore, this study aimed to identify the spectrum of β-globin gene mutations among HbE/β-thalassemia patients, subsequently correlating the identified mutations with hematological parameters. A total of 204 transfusion-dependent HbE/β-thalassemia patients were randomly selected at Hospital USM and Ministry of Health hospitals in Malaysia. MARMS-PCR and CSGE were performed to screen for β-globin gene mutations. All genotyping results were confirmed and validated by DNA sequencing. The results were analyzed using SPSS version 22 and STATA SE.  A total of 152 subjects were diagnosed as HbE/β-thalassemia. Thirteen compound heterozygous mutations were identified. The most prevalent compound heterozygous mutations were CD 26 (G-A) and IVS ...

The antigen rapid diagnostic test (Ag-RDT) is an immunodiagnostic test that detects the presence of viral proteins (antigens) expressed by the COVID-19 virus in a sample from a patient’s respiratory tract. This study focused on evaluating the performance of self-conduct buccal and nasal swabs RTK-antigen test compared to nasopharyngeal swab RTK-based COVID-19 diagnostic assays, Panbio™ COVID-19 Ag Rapid Test Device (Nasopharyngeal) (Abbott Rapid Diagnostics Jena GmbH, Jena, Germany) used in hospitals for first-line screening. The sensitivity and specificity of the paired RTK-Ag test in detecting the an-tigen were calculated at 96.4% and 100%, respectively. Fisher exact tests showed the association between nasopharyngeal swabs RTK-Ag assay and buccal-nasal swabs RTK-Ag from ProdetectTM is significant (p-values < 0.001). The result showed that a self-conducted buccal and nasal RTK-antigen rapid test by the patients is comparable to the results obtained from a rapid test device cond...

Background and objective: Thalassaemia prevention which includes public education, screening and pre-marital counselling, forms important components of a prevention programme. The EduVariome programme under MyHVP Malaysia is an educational intervention programme developed to increase awareness on thalassemia and the importance of thalassemia screening. This study aimed to assess the effectiveness of this an educational intervention programme in increasing the awareness of secondary school students regarding thalassaemia. Method: This study was conducted among secondary school students aged between 14 to 17 years old. The programme was conducted by giving a lecture in the form of a forum related to thalassaemia. Questionnaires to assess the participants’ sunderstanding of the disease were distributed before and after the lecture. The data was analyzed using SPSS Version 22. Results: A total of 422 secondary school students in Kelantan state took part in this study. About thirty six p...

Antimicrobial resistance in companion animals is a major public health concern worldwide due to the animals’ zoonotic potential and ability to act as a reservoir for resistant genes. We report on the first use of meta-analysis and a systematic review to analyze the prevalence of vancomycin-resistant Enterococcus (VRE) in companion animals. Databases such as MedLib, PubMed, Web of Science, Scopus, and Google Scholar were searched. The information was extracted by two independent reviewers and the results were reviewed by a third. Two reviewers independently assessed the study protocol using the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) checklist and the study quality using the Joanna Briggs Institute (JBI) critical appraisal checklist for prevalence data. OpenMeta analyst and comprehensive meta-analysis (CMA) were used for the meta-analysis. The random effect model was used, and publication bias was assessed using the Eggers test and funnel plot. Bet...

Since its first detection in December 2019, more than 232 million cases of COVID-19, including 4.7 million deaths, have been reported by the WHO. The SARS-CoV-2 viral genomes have evolved rapidly worldwide, causing the emergence of new variants. This systematic review and meta-analysis was conducted to provide a global mutational profile of SARS-CoV-2 from December 2019 to October 2020. The review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-analysis (PRISMA), and a study protocol was lodged with PROSPERO. Data from 62 eligible studies involving 368,316 SARS-CoV-2 genomes were analyzed. The mutational data analyzed showed most studies detected mutations in the Spike protein (n = 50), Nucleocapsid phosphoprotein (n = 34), ORF1ab gene (n = 29), 5′-UTR (n = 28) and ORF3a (n = 25). Under the random-effects model, pooled prevalence of SARS-CoV-2 variants was estimated at 95.1% (95% CI; 93.3–96.4%; I2 = 98.952%; p = 0.000) while subgroup meta-an...

This study highlights the development of a multiplex real-time loop-mediated isothermal amplification assay. The developed assay employed a dual-function oligonucleotide (DFO) which simultaneously monitors the emitted amplification signals and accelerates the amplification process. The DFO was a modification of loop primer (LP); the 5'-end and 3'-end of the LP was tagged with fluorophore and quencher, respectively. The DFO was quenched in its unbound state and fluoresces only when it anneals to the specific target during the amplification process. With the same working mechanism as LP, DFO allowed the detection of target genes in less than 1 h in a real time monitoring system. We demonstrated this detection platform with Burkholderia pseudomallei, the causative agent of melioidosis. An internal amplification control (IAC) was incorporated in the assay to rule out false negative result and to demonstrate that the assay was successfully developed in a multiplex system. The assay was 100% specific when it was evaluated against 96 B. pseudomallei clinical isolates and 48 other bacteria species. The detection limit (sensitivity) of the developed assay was 1 fg/μl of B. pseudomallei genomic DNA and 18.2 CFU/ml at the bacterial cell level. In spiked blood samples, the assay's detection limit was 14 CFU/ml. The assay's diagnostic evaluation showed 100% diagnostic sensitivity, diagnostic specificity, positive predictive value, and negative predictive value. An integrated multiplex LAMP and real-time monitoring system was successfully developed, simplifying the workflow for the rapid and specific nucleic acid diagnostic test.

BACKGROUND Beta-thalassemia is a genetic disorder that is inherited in an autosomal recessive pattern. This genetic disease leads to a defective beta-globin hemoglobin chain causing partial or complete beta-globin chain synthesis loss. Beta-thalassemia major patients need a continuous blood transfusion and iron chelation to maintain the normal homeostasis of red blood cells (RBCs) and other systems in the body. Patients also require treatment procedures that are costly and tedious, resulting in a serious health burden for developing nations such as Nepal. METHODS A total of 61 individuals clinically diagnosed to have thalassemia were genotyped with multiplex amplification refractory mutation system-polymerase chain reaction (ARMS-PCR). Twenty-one major mutations were investigated using allele-specific primers grouped into six different panels. RESULTS The most common mutations found (23%) were IVS 1-5 (G-C) and Cd 26 (G-A) (HbE), followed by 619 deletion, Cd 8/9 (+G), Cd 16 (-C), Cd 41/42 (-TTCT), IVS 1-1 (G-T), Cd 19 (A-G), and Cd 17 (A-T) at 20%, 12%, 8%, 6%, 4%, 3%, and 1%, respectively. CONCLUSION The results of this study revealed that Nepal's mutational profile is comparable to that of its neighboring countries, such as India and Myanmar. This study also showed that thalassemia could be detected across 17 Nepal's ethnic groups, especially those whose ancestors originated from India and Central Asia.

BACKGROUND Hereditary nonpolyposis colorectal cancer, or Lynch syndrome, caused by germline mutations or genetic defects in mismatch repair (MMR) genes (MLH1, MSH2, PMS2, MSH6, and epithelial cellular adhesion molecule), is an autosomal dominant condition accounting for 2–5% of all colorectal carcinomas (CRCs). Reports on MMR loss in many populations are available; however, there are no reports on the frequency of MMR protein expression in Nepalese cohorts. Therefore, this study was aimed to assess the expression profiles of MLH1 and MSH2 protein by immunohistochemistry (IHC) in Nepalese CRC patients. METHODS This retrospective study used archived formalin-fixed paraffin-embedded tissue blocks from 43 Nepalese CRC patients. IHC staining was performed using MLH1 and MSH2 antibodies. IHC scoring analysis was assessed using semiquantitative scoring. RESULTS Of the 43 CRC patients, 8 (18.6%) showed loss of staining for MLH1 antibody, 5 (11.6%) showed loss of staining for MSH2 antibody, ...

CYP2A6 gene encodes the principal enzyme involved in the metabolism of many drugs including artesunate. We developed a simplified duplex nested PCR method for the detection of the CYP2A61B, CYP2A62, CYP2A64, CYP2A67, CYP2A68 and CYP2A69 variant alleles highly prevalent among Malaysian population. Genomic DNA was isolated from peripheral blood of patients treated with artesunate by using a DNA extraction kit. Fragments from TATA box in 5' flanking region, exons 1 to 4 and exon 8 to 3' UTR of the gene were amplified using a first PCR step (PCR1). Products from PCR1 were then used to run a duplex PCR (PCR2) for detecting the selected variant alleles. All 6 CYP2A6 alleles were successfully amplified and confirmed by sequencing. The frequency of the alleles were 14.6% for CYP2A61B, 16.7% for CYP2A64, 4.2% for CYP2A68 and 10.4% for CYP2A69. High frequencies of the CYP2A64 and CYP2A69 alleles which contribute to the poor metabolism status may impact treatment of patients receiving drugs metabolized by CYP2A6 such as artesunate among Malaysians.