Blended phenotypes exhibited by a patient may present a challenge to the establishment of diagnosis. In this study, we report a seven-year-old Murut girl with unusual features of Williams-Beuren syndrome (WBS), including recurrent infections and skin abscesses. Considering the possibility of a second genetic disorder, a mutation screening for genes associated with inborn errors of immunity (IEI) was conducted using whole exome sequencing (WES). Analysis of copy number variations (CNVs) from the exome data revealed a 1.53Mb heterozygous deletion on chromosome 7q11.23, corresponding to the known WBS. We also identified a biallelic loss of NCF1, which indicated autosomal recessive chronic granulomatous disease (CGD). Dihydrorhodamine (DHR) flow cytometric assay demonstrated abnormally low neutrophil oxidative burst activity. Coamplification of NCF1 and its pseudogenes identified a GT-deletion (ΔGT) at the start of exon 2 in NCF1 (NM_000265.7: c.75_76delGT: p.Tyr26Hisfs*26). Estimation of NCF1-to-NCF1 pseudogenes ratio using ΔGT and 20-bp gene scans affirmed nil copies of NCF1 in the patient. While the father had a normal ratio of 2:4, the mother had a ratio of 1:5, implicating the carrier of ΔGT-containing NCF1. Discovery of a 7q11.23 deletion involving one NCF1 allele and a ΔGT in the second NCF1 allele explained the coexistence of WBS and CGD in our patient. This study highlights the capability of WES to establish a molecular diagnosis for a case with blended phenotypes, enabling the provision of appropriate prophylactic treatment.
In March 2011, an outbreak of acute respiratory disease was reported at the Kuala Lumpur (Malaysia) Police Training Centre. Approximately 100 trainees were hospitalized and 5 were admitted to the intensive care unit. Three of these 5 trainees died. Human adenovirus type 7 was identified as the etiologic agent.
Familial hypercholesterolemia (FH) is a disease implicated with defects in either, Low density lipoprotein receptor gene (LDLR), Apolipoprotein B-100 gene (APOB), the Proprotein convertase subtilisin/kexin type 9 gene (PCSK9) or other related genes of the lipid metabolism pathway. The general characterization of heterozygous FH is by elevated low-density lipoprotein (LDL) cholesterol and early-onset cardiovascular diseases, while the more severe type, the homozygous FH results in extreme elevated levels of LDL cholesterol and usually death of an affected individual by early twenties. We present here a novel non-synonymous, missense mutation in exon 14 of the LDLR gene in two siblings of the Malay ethnicity discovered during an in-house genetic test. We postulate that their elevated cholesterol is due to this novel mutation and they are positive for homozygous FH. This is the first report of a C711Y mutation in patients with elevated cholesterol in Asia.
Li-Fraumeni syndrome (LFS) is a highly penetrant, autosomal dominant disorder where affected individuals carry a 50% risk of developing cancer before 30 years of age. It is most commonly associated with mutations in the tumour suppressor gene, TP53. Adrenocortical carcinoma (ACC) is a very rare paediatric cancer, and up to 80% of affected children are found to carry germline TP53 mutations. Hence, we propose using childhood ACC incidence as selection criteria for referral for TP53 mutation testing, independent of familial cancer history. Under the auspices of the Malaysian Society of Paediatric Haematology-Oncology, four eligible children diagnosed with ACC over a 30-month study period were referred for mutation testing. Three had a germline TP53 mutation. Subsequent TP53 testing in relatives showed two inherited mutations and one de novo mutation. These findings strongly support paediatric ACC as a useful sentinel cancer for initiating a germline TP53/LFS detection programme, particularly in countries where the lack of structured oncogenetic practice precludes the identification of families with LFS features.
We report for the first time on the prevalence, antibiotic resistance and RAPD types of Campylobacter species in ducks and duck related environmental samples in Malaysia. Samples were examined by enrichment in Bolton Broth followed by plating onto modified Charcoal Cefoperazone Deoxycholate agar (mCCDA) and/or plating directly onto mCCDA. A total of 643 samples were screened, and the prevalence of Campylobacter spp. in samples from different sources ranged from 0% to 85%. The method of isolation had a significant (P<0.05) effect on the isolation rate. One hundred and sixteen Campylobacter isolates, comprising of 94 Campylobacter jejuni, 19 Campylobacter coli and three Campylobacter lari, were examined for their sensitivity to 13 antibiotics. Majority of the C. jejuni isolates were resistant to cephalothin (99%), tetracycline (96%), suphamethoxazole/trimethoprim (96%), and very few were resistant to gentamicin (5%), chloramphenicol (7%) and erythromycin (1%). All C. coli isolates were resistant to cephalothin, nalidixic acid, norfloxacin and tetracycline but susceptible to chloramphenicol, erythromycin and gentamicin. The three C. lari isolates were resistant to all the antibiotics tested except chloramphenicol and gentamicin (1/3 and 2/3 susceptible, respectively). Genetic diversity of Campylobacter isolates were determined using random amplification of polymorphic DNA (RAPD). C. jejuni and C. coli isolates belong to fifty-eight and twelve RAPD types, respectively.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Haemophilia B is caused by coagulation defects in the factor IX gene located in Xq27.1 on the X chromosome. Identification of mutations contributing to defective factor IX may be advantageous for precise carrier and prenatal diagnosis. We studied 16 patients from 11 families, consisting of 8 patients of the Malay ethnic group, of which 6 were siblings. Factor IX mutations have not been previously reported in the Malay ethnic group. The functional region of the factor IX gene was sequenced and mutations were identified in either the exon or intronic regions in 15 of the patients. One novel mutation, 6660_6664delTTCTT was identified in siblings with moderate form of haemophilia B. Mutations identified in our patients when linked with disease severity were similar to findings in other populations. In summary, this preliminary data will be used to build a Malaysian mutation database which would facilitate genetic counseling.
The aim of our study was to see the effects of GSTP1 polymorphism on biomarkers of ageing, including micronuclei (MN), comet tail length, and relative telomere length in automobile repair workers, who are exposed to a broad spectrum of potential mutagens. The analysis was performed on buccal cells collected from occupationally exposed and non-exposed (control) subjects. Samples were analysed using cytogenetic and molecular methods, including restriction fragment length polymorphism (RFLP), MN test, comet assay, and real-time PCR. The results confirmed the DNA damaging effects of substances used in the mechanical workshops, but did not confirm the influence of GSTP1 gene polymorphism on DNA damage. However, further studies on both occupationally exposed and control populations are needed to understand the relationship between GSTP1 polymorphism and genome damage.
To date, no genome of any of the species from the genus Spiroplasma has been completely sequenced. Long repetitive sequences similar to mobile units present a major obstacle for current genome sequencing technologies. Here, we report the assembly of the Spiroplasma melliferum KC3 genome into 4 contigs, followed by proteogenomic annotation and metabolic reconstruction based on the discovery of 521 expressed proteins and comprehensive metabolomic profiling. A systems approach allowed us to elucidate putative pathogenicity mechanisms and to discover major virulence factors, such as Chitinase utilization enzymes and toxins never before reported for insect pathogenic spiroplasmas.
Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) is a common autosomal recessive disorder. Our objective was to identify the 21-hydroxylase active gene, CYP21A2 mutations in Malaysian 21-OHD patients using different techniques.
Inophyllin A (INO-A), a pyranoxanthone isolated from the roots of Calophyllum inophyllum represents a new xanthone with potential chemotherapeutic activity. In this study, the molecular mechanism of INO-A-induced cell death was investigated in Jurkat T lymphoblastic leukemia cells. Assessment of phosphatidylserine exposure confirmed apoptosis as the primary mode of cell death in INO-A-treated Jurkat cells. INO-A treatment for only 30 min resulted in a significant increase of tail moment which suggests that DNA damage is an early apoptotic signal. Further flow cytometric assessment of the superoxide anion level confirmed that INO-A induced DNA damage was mediated with a concomitant generation of reactive oxygen species (ROS). Investigation on the thiols revealed an early decrease of free thiols in 30 min after 50 μM INO-A treatment. Using tetramethylrhodamine ethyl ester, a potentiometric dye, the loss of mitochondrial membrane potential (MPP) was observed in INO-A-treated cells as early as 30 min. The INO-A-induced apoptosis progressed with the simultaneous activation of caspases-2 and -9 which then led to the processing of caspase-3. Taken together, these data demonstrate that INO-A induced early oxidative stress, DNA damage and loss of MMP which subsequently led to the activation of an intrinsic pathway of apoptosis in Jurkat cells.
Zika virus (ZIKV) is a mosquito-borne flavivirus distributed throughout much of Africa and Asia. Infection with the virus may cause acute febrile illness that clinically resembles dengue fever. A recent study indicated the existence of three geographically distinct viral lineages; however this analysis utilized only a single viral gene. Although ZIKV has been known to circulate in both Africa and Asia since at least the 1950s, little is known about the genetic relationships between geographically distinct virus strains. Moreover, the geographic origin of the strains responsible for the epidemic that occurred on Yap Island, Federated States of Micronesia in 2007, and a 2010 pediatric case in Cambodia, has not been determined.
Hereditary pancreatitis (HP) is a very rare form of early-onset chronic pancreatitis, which usually begins in childhood with a variable spectrum of severity of disease. HP is commonly caused by variants/mutations in the PRSS1 gene as reported in many studies. Therefore, in this study, we aimed to investigate the possible association of PRSS1 gene variants/mutations in a Malaysian Chinese family with HP.
To measure plasma pro-apoptotic and pro-necrotic activity in severe acute kidney injury (AKI) patients within a randomized controlled trial of continuous veno-venous hemofiltration with high cut-off filters (CVVH-HCO) versus standard filters (CVVH-Std).
Ferritin is a highly-conserved iron-storage protein that has also been identified as an acute phase protein within the innate immune system. The iron-storage function is mediated through complementary roles played by heavy (H)-chain subunit as well as the light (L) in mammals or middle (M)-chain in teleosts, respectively. In this study, we report the identification of five ferritin subunits (H1, H2, M1, M2, M3) in the Atlantic salmon that were supported by the presence of iron-regulatory regions, gene structure, conserved domains and phylogenetic analysis. Tissue distribution analysis across eight different tissues showed that each of these isoforms is differentially expressed. We also examined the expression of the ferritin isoforms in the liver and kidney of juvenile Atlantic salmon that was challenged with Aeromonas salmonicida as well as in muscle cell culture stimulated with interleukin-1β. We found that each isoform displayed unique expression profiles, and in certain conditions the expressions between the isoforms were completely diametrical to each other. Our study is the first report of multiple ferritin isoforms from both the H- and M-chains in a vertebrate species, as well as ferritin isoforms that showed decreased expression in response to infection. Taken together, the results of our study suggest the possibility of functional differences between the H- and M-chain isoforms in terms of tissue localisation, transcriptional response to bacterial exposure and stimulation by specific immune factors.
A study was carried out to isolate Brucella melitensis using established bacteriological and PCR techniques in Brucella seropositive goats in farms in Selangor, Negeri Sembilan, Melaka and Pulau Pinang. Brucella melitensis was isolated from 7 of 134 reactors with the highest isolation from the vaginal swabs (57.14%) followed by the spleen (28.57%), uterine fluid (14.29%). No Brucella was isolated from the lymph nodes. PCR confirmed all the seven isolates as B. melitensis and isolates were phylogenetically related to other isolates from India, Iran, and Israel but most closely related to isolates from Singapore.
Staphylococcus aureus is a persistent human pathogen responsible for a variety of infections ranging from soft-tissue infections to bacteremia. The objective of this study was to determine genetic relatedness between methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) strains. We isolated 35 MRSA and 21 MSSA strains from sporadic cases at the main tertiary hospital in Terengganu, Malaysia, screening them for the presence of virulence genes. Their genetic relatedness was determined by accessory gene regulator (agr) types, PCR-restriction fragment length polymorphism (RFLP) of the coa gene, pulsed-field gel electrophoresis (PFGE), S. aureus protein A (spa), and multilocus-sequence typing (MLST). We found that 57% of MRSA and 43% of MSSA strains harbored enterotoxin genes. The majority (87.5%) of the strains were agr type I. PCR-RFLP and PFGE genotyping of the coa gene revealed that MRSA strains were genetically related, whereas MSSA strains had higher heterogeneity. The combined genotype, MLST-spa type ST239-t037, was shared among MRSA and MSSA strains, indicating that MRSA strains could have evolved from MSSA strains. Two combined MLST-spa types were present in MRSA strains, whereas 7 different MLST-spa types were detected in MSSA strains, including 2 combined types (ST779-t878 and ST1179-t267) that have not been reported in Malaysia. In conclusion, enterotoxin genes were more prevalent in MRSA than in MSSA strains in the Terengganu hospital. The MSSA strains were genetically more diverse than the MRSA strains.
Rubella has been listed as a mandatory notifiable disease in Taiwan since 1988. Because of high coverage rates with an effective vaccine, rubella cases have decreased dramatically in Taiwan since 1994. However, rubella outbreaks still occur due to imported transmission. Five large clusters were detected in Taiwan from 2007 to 2011. In 2007, one cluster was caused by rubella genotype 1E viruses that were imported from Vietnam, whereas another cluster was caused by genotype 2B viruses and was untraceable. In 2008, two clusters were caused by different lineages of genotype 1E viruses that were imported from Malaysia. In 2009, a cluster that was caused by genotype 2B viruses was associated with imported cases from Vietnam. The rubella viruses from 124 confirmed cases from 2005 to 2011 were characterized, and the data revealed that these viruses were distributed in the following four genotypes: 1E (n = 56), 1h (n = 1), 1j (n = 4), and 2B (n = 63). Of these viruses, 93 (75%) were associated with imported cases, and 43 of 56 genotype 1E viruses were associated with imported cases from China, Vietnam, Malaysia, and Indonesia. One genotype 1h virus was imported from Belarus, and three of four genotype 1j viruses were imported from the Philippines. Of 63 rubella genotype 2B viruses, 46 were imported from Vietnam, Thailand, Malaysia, China, Germany, and South Africa. Molecular surveillance allows for the differentiation of circulating rubella viruses and can be used to investigate transmission pathways, which are important to identify the interruption of endemic virus transmission.
Toxoplasmosis is an infection caused by the parasite Toxoplasma gondii. Chronically-infected individuals with a compromised immune system are at risk for reactivation of the disease. In-vivo induced antigen technology (IVIAT) is a promising method for the identification of antigens expressed in-vivo. The aim of the present study was to apply IVIAT to identify antigens which are expressed in-vivo during T. gondii infection using sera from individuals with chronic toxoplasmosis. Forty serum samples were pooled, pre-adsorped against three different preparations of antigens, from each in-vitro grown T. gondii and Escherichia coli XLBlue MRF', and then used to screen a T. gondii cDNA expression library. Sequencing of DNA inserts from positive clones showed eight open reading frames with high homology to T. gondii genes. Expression analysis using quantitative real-time PCR showed that SAG1-related sequence 3 (SRS3) and two hypothetical genes were up-regulated in-vivo relative to their expression levels in-vitro. These three proteins also showed high sensitivity and specificity when tested with individual serum samples. Five other proteins namely M16 domain peptidase, microneme protein, elongation factor 1-alpha, pre-mRNA-splicing factor and small nuclear ribonucleoprotein F had lower RNA expression in-vivo as compared to in-vitro. SRS3 and the two hypothetical proteins warrant further investigation into their roles in the pathogenesis of toxoplasmosis.
Plasmodium knowlesi is a potentially life-threatening zoonotic malaria parasite due to its relatively short erythrocytic cycle. Microscopic identification of P. knowlesi is difficult, with "compacted parasite cytoplasm" being one of the important identifying keys. This report is about a case of hyperparasitaemic human P. knowlesi infection (27% parasitaemia) with atypical amoeboid morphology. A peninsular Malaysian was admitted to the hospital with malaria. He suffered anaemia and acute kidney function impairment. Microscopic examination, assisted by nested PCR and sequencing confirmed as P. knowlesi infection. With anti-malarial treatment and several medical interventions, patient survived and recovered. One-month medical follow-up was performed after recovery and no recrudescence was noted. This case report highlights the extreme hyperparasitaemic setting, the atypical morphology of P. knowlesi in the patient's erythrocytes, as well as the medical interventions involved in this successfully treated case.
A cryptic plasmid, pMWHK1 recovered from an Antarctic bacterium Pedobacter cryoconitis BG5 was sequenced and characterised. The plasmid is a circular 6206bp molecule with eight putative open reading frames designated as orf1, orf2, orf3, orf4, orf5, orf6, orf7 and orf8. All the putative open reading frames of pMWHK1 are found to be actively transcribed. Proteins encoded by orf2 and orf4 are predicted to be responsible for the mobilization and replication of the plasmid respectively. orf4 shares 55% and 61% identities with the theta-type Rep proteins from two strains of Riemerella anatipestifer. This suggests that pMWHK1 could be a member of the theta-type replicating plasmid. The origin of replication is located within the AT-rich region upstream of orf4. orf5 and orf6 encode bacterial toxin-antitoxin proteins predicted to maintain plasmid stability. orf3 encodes an entry exclusion protein that is hypothetically involved in reducing the frequency of DNA transfer through conjugation. orf1, orf7 and orf8 encode proteins with unknown functions. Plasmid, pMWHK1 is stably maintained in P. cryoconitis BG5 at 20°C.