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Fulltext Determinants of neonatal, infant and under-five mortalities: evidence from a developing country, Bangladesh

Murad MW, Abdullah ABM, Islam MM, Alam MM, Reaiche C, Boyle S

J Public Health Policy, 2023 Jun;44(2):230-241.
PMID: 37117262 DOI: 10.1057/s41271-023-00413-w

We investigated the macroeconomic determinants of neonatal, infant, and under-five mortalities in Bangladesh for the period 1991-2018 and discuss implications of the United Nations' Sustainable Development Goal 3 (SDG 3) and Millennium Development Goal 4 (MDG 4) for developing countries. We used annual time series data and the econometric techniques of Fully Modified Ordinary Least Squares (FMOLS) and Dynamic Ordinary Least Squares (DOLS) regressions for analysis. Determinants most effective in combating neonatal, infant, and under-five mortalities include variables such as 'protecting newborns against tetanus', 'increasing healthcare expenditure', and 'making sure births are attended by skilled healthcare staff'. Employing more healthcare workers and assuring more and improved healthcare provisions can further reduce the neonatal, infant, and under-five mortalities. Developing countries with similar macroeconomic profiles can achieve similar SDG 3 and MDG 4 outcomes by emulating the policies and strategies Bangladesh applied to reducing child mortalities over the last three decades.
Fulltext MicroRNAs and Target Genes As Biomarkers for the Diagnosis of Early Onset of Parkinson Disease

Arshad AR, Sulaiman SA, Saperi AA, Jamal R, Mohamed Ibrahim N, Abdul Murad NA

Front Mol Neurosci, 2017;10:352.
PMID: 29163029 DOI: 10.3389/fnmol.2017.00352

Among the neurodegenerative disorders, Parkinson's disease (PD) ranks as the second most common disorder with a higher prevalence in individuals aged over 60 years old. Younger individuals may also be affected with PD which is known as early onset PD (EOPD). Despite similarities between the characteristics of EOPD and late onset PD (LODP), EOPD patients experience much longer disease manifestations and poorer quality of life. Although some individuals are more prone to have EOPD due to certain genetic alterations, the molecular mechanisms that differentiate between EOPD and LOPD remains unclear. Recent findings in PD patients revealed that there were differences in the genetic profiles of PD patients compared to healthy controls, as well as between EOPD and LOPD patients. There were variants identified that correlated with the decline of cognitive and motor symptoms as well as non-motor symptoms in PD. There were also specific microRNAs that correlated with PD progression, and since microRNAs have been shown to be involved in the maintenance of neuronal development, mitochondrial dysfunction and oxidative stress, there is a strong possibility that these microRNAs can be potentially used to differentiate between subsets of PD patients. PD is mainly diagnosed at the late stage, when almost majority of the dopaminergic neurons are lost. Therefore, identification of molecular biomarkers for early detection of PD is important. Given that miRNAs are crucial in controlling the gene expression, these regulatory microRNAs and their target genes could be used as biomarkers for early diagnosis of PD. In this article, we discussed the genes involved and their regulatory miRNAs, regarding their roles in PD progression, based on the findings of significantly altered microRNAs in EOPD studies. We also discussed the potential of these miRNAs as molecular biomarkers for early diagnosis.
Fulltext Interactions Among Non-Coding RNAs in Diabetic Nephropathy

Loganathan TS, Sulaiman SA, Abdul Murad NA, Shah SA, Abdul Gafor AH, Jamal R, et al.

Front Pharmacol, 2020;11:191.
PMID: 32194418 DOI: 10.3389/fphar.2020.00191

Diabetic Nephropathy (DN) is the most common cause of End-stage renal disease (ESRD). Although various treatments and diagnosis applications are available, DN remains a clinical and economic burden. Recent findings showed that noncoding RNAs (ncRNAs) play an important role in DN progression, potentially can be used as biomarkers and therapeutic targets. NcRNAs refers to the RNA species that do not encode for any protein, and the most known ncRNAs are the microRNAs (miRNAs), long noncoding RNAs (lncRNAs), and circular RNAs (circRNAs). Dysregulation of these ncRNAs was reported before in DN patients and animal models of DN. Importantly, there are some interactions between these ncRNAs to regulate the crucial steps in DN progression. Here, we aimed to discuss the reported ncRNAs in DN and their interactions with critical genes in DN progression. Elucidating these ncRNAs regulatory network will allow for a better understanding of the molecular mechanisms in DN and how they can act as new biomarkers for DN and also as the potential targets for treatment.
Cgl-SLT2 is required for appressorium formation, sporulation and pathogenicity in Colletotrichum gloeosporioides

Yong HY, Bakar FD, Illias RM, Mahadi NM, Murad AM

Braz J Microbiol, 2013 Dec;44(4):1241-50.
PMID: 24688518

The mitogen-activated protein (MAP) kinase pathways has been implicated in the pathogenicity of various pathogenic fungi and plays important roles in regulating pathogenicity-related morphogenesis. This work describes the isolation and characterization of MAP kinase gene, Cgl-SLT2, from Colletotrichum gloeosporioides. A DNA sequence, including 1,633 bp of Cgl-SLT2 open-reading frame and its promoter and terminator regions, was isolated via DNA walking and cloned. To analyze gene function, a gene disruption cassette containing hygromycin-resistant gene was constructed, and Cgl-SLT2 was inactivated via gene deletion. Analysis on Cgl-slt2 mutant revealed a defect in vegetative growth and sporulation as compared to the wild-type strain. When grown under nutrient-limiting conditions, hyperbranched hyphal morphology was observed in the mutant. Conidia induction for germination on rubber wax-coated hard surfaces revealed no differences in the percentage of conidial germination between the wild-type and Cgl-slt2 mutant. However, the percentage of appressorium formation in the mutant was greatly reduced. Bipolar germination in the mutant was higher than in the wild-type at 8-h post-induction. A pathogenicity assay revealed that the mutant was unable to infect either wounded or unwounded mangoes. These results suggest that the Cgl-SLT2 MAP kinase is required for C. gloeosporioides conidiation, polarized growth, appressorium formation and pathogenicity.
Fulltext Conducting Polymers for Optoelectronic Devices and Organic Solar Cells: A Review

R Murad A, Iraqi A, Aziz SB, N Abdullah S, Brza MA

Polymers (Basel), 2020 Nov 09;12(11).
PMID: 33182241 DOI: 10.3390/polym12112627

In this review paper, we present a comprehensive summary of the different organic solar cell (OSC) families. Pure and doped conjugated polymers are described. The band structure, electronic properties, and charge separation process in conjugated polymers are briefly described. Various techniques for the preparation of conjugated polymers are presented in detail. The applications of conductive polymers for organic light emitting diodes (OLEDs), organic field effect transistors (OFETs), and organic photovoltaics (OPVs) are explained thoroughly. The architecture of organic polymer solar cells including single layer, bilayer planar heterojunction, and bulk heterojunction (BHJ) are described. Moreover, designing conjugated polymers for photovoltaic applications and optimizations of highest occupied molecular orbital (HOMO)-lowest unoccupied molecular orbital (LUMO) energy levels are discussed. Principles of bulk heterojunction polymer solar cells are addressed. Finally, strategies for band gap tuning and characteristics of solar cell are presented. In this article, several processing parameters such as the choice of solvent(s) for spin casting film, thermal and solvent annealing, solvent additive, and blend composition that affect the nano-morphology of the photoactive layer are reviewed.
Fulltext Identification of the major allergen of Macrobrachium rosenbergii (giant freshwater prawn)

Yadzir ZH, Misnan R, Abdullah N, Bakhtiar F, Arip M, Murad S

Asian Pac J Trop Biomed, 2012 Jan;2(1):50-4.
PMID: 23569834 DOI: 10.1016/S2221-1691(11)60189-5

OBJECTIVE: To characterize the major allergens of Macrobrachium rosenbergii (giant freshwater prawn).
METHODS: Raw and cooked extracts of the giant freshwater prawn were prepared. The IgE reactivity pattern was identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting technique with the sera of 20 skin prick test (SPT) positive patients. The major allergen identified was then characterized using the proteomics approach involving a combination of two-dimensional (2-DE) electrophoresis, mass spectrometry and bioinformatics tools.
RESULTS: SDS-PAGE of the raw extract showed 23 protein bands (15-250 kDa) but those ranging from 40 to 100 kDa were not found in the cooked extract. From immunoblotting experiments, raw and cooked extracts demonstrated 11 and 5 IgE-binding proteins, respectively, with a molecular mass ranging from 15 to 155 kDa. A heat-resistant 36 kDa protein was identified as the major allergen of both extracts. In addition, a 42 kDa heat-sensitive protein was shown to be a major allergen of the raw extract. The 2-DE gel fractionated the prawn proteins to more than 50 different protein spots. Of these, 10 spots showed specific IgE reactivity with patients' sera. Matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) analysis led to identification of 2 important allergens, tropomyosin and arginine kinase.
CONCLUSIONS: It can be concluded that the availability of such allergens would help in component-based diagnosis and therapy of prawn allergies.
Fulltext First report of Aspergillus niger causing fruit rot of bilimbi in Malaysia

Mohd Zainudin NAI, Abd Murad NB, Aris A, Hussain NH

Plant Dis, 2022 Sep 19.
PMID: 36122197 DOI: 10.1094/PDIS-06-22-1291-PDN

In July 2019 to November 2021, symptoms of fruit rot of Averrhoa bilimbi (commonly known as bilimbi) fruits were observed in Serdang (3°00'05.8"N 101°42'18.4"E) and Tanjong Karang (3°25'55.3"N 101°12'55.7"E), Selangor, Malaysia. External decay showed some yellowish to brownish-red discoloration and inside the fruit there was black powdery sporulation and a brownish decay that measured between 8 - 15 mm wide. Twenty random symptomatic fruits were collected from each location. Small pieces (5 mm) of infected tissues from the fruit rot were surface sterilized for 1 min in 0.5% NaOCl, washed twice with sterile distilled water and cultured onto potato dextrose agar (PDA) and peptone pentachloronitrobenzene agar (PPA). The plates were incubated at 28 ± 1oC under 12 hours light/dark for 7 days. The fungal colonies growing from the plates were purified using hyphal tip technique (Leyronas et al. 2012). A total of 42 fungal isolates were obtained, and the morphology characteristics of six isolates were matched that of Aspergillus niger. The A. niger isolates were further identified based genus and species-specific Internal Transcribed Spacer (ITS) sequencing. Primers ITS1/ITS4, were used to amplify and subsequently sequenced the ITS1-5.8S-ITS2 region (White et al. 1990). Primer ASAP1 (5'-CAGCGAGTACATCACCTTGG-3'), ASAP2 (5'-CCATTGTTGAAAGTTTTAACTGATT-3') was used for Aspergillus species confirmation, primer ASPU (5'-ACTACCGATTGAATGGCTCG-3') / Ni1r (5'-ACGCTTTCAGACAGTGTTCG-3') for A. niger species-specific, ASPU / Af3r (5'-CATACTTTCAGAACAGCGTTCA-3') for A. fumigatus specific-specific and ASPU / Fl2r (5'-TTCACTAGATCAGACAGAGT-3') for A. flavus specific-specific (Sugita et al. 2004). In general, Aspergillus niger isolates grew rapidly on PDA and were visibly white initially then appearing black and powdery on the second day of incubation (Figure 1A). Some isolates grew rapidly (0.71-0.85 cm/day) and have a cottony appearance. The conidia were appeared brown to black, globose and rough with diameter ranging between 4.1-5.2 µm (Figure 1B). The vesicles were hyaline, globose, and brown in color with measurement of 30-75 µm in diameter with uniseriate sterigmata (Figure 1C). The conidial head was brownish black in color and split into several irregular and regular columns of conidial chains (Figure 1D-E). The conidiophores were hyaline, and brown in color. Phylogenetic trees of ITS (Figure 2A) and ASAP sequences (Figure 2B) were constructed using a Neighbor-Joining method showing isolates Aspergillus niger #11, #15, #32, #33, #41 and #42 were grouped into the same clade as A. niger (accession no. MT446087). To examine virulence of A. niger, pathogenicity tests were performed three times by inoculating an asymptomatic fruit with six isolates of A. niger (isolate #11, #15, #32, #33, #41 and #42) and a single isolate for each species of Aspergillus aculeatus, Lasiodiplodia theobromae and Penicillium gerundense. Ten fruits were inoculated by placing a mycelial disc (6 mm) (Kouame et al. 2010) from a 5-day-old culture of each fungal colony while control fruits were non-inoculated with any fungal colony (10 fruits were inoculated with a sterile agar disc and 10 were non-inoculated, respectively). After 3 days, typical symptoms of Aspergillus fruit rot were observed on A. niger inoculated fruits, whereas the control fruits remained asymptomatic (Figure 1F-P). Aspergillus niger was reisolated and reidentified based on morphological and molecular characterization from the inoculated, symptomatic fruits, thus confirming Koch's postulates. A. niger causing widespread diseases in various plant and it is a common contaminant of food. This study shows A. niger to be highly virulent on bilimbi fruits and leads to reduction of fruit quality and its production. To our knowledge, this is the first report of A. niger causing fruit rot on bilimbi and future work on its pathogenesis may provide strategies for disease control against the pathogen.
Mating compatibility and competitiveness of transgenic and wild type Aedes aegypti (L.) under contained semi-field conditions

Lee HL, Vasan S, Ahmad NW, Idris I, Hanum N, Selvi S, et al.

Transgenic Res, 2013 Feb;22(1):47-57.
PMID: 22700207 DOI: 10.1007/s11248-012-9625-z

We conducted the world's first experiments under semi-field conditions (ACL-2 field house) to assess the mating competitiveness of genetically sterile RIDL male mosquitoes (513A strain). The field house is a state-of-the-art, fully-contained trial facility, simulating the living space for a household of 2-4 people in Peninsular Malaysia. Ten genetically sterile RIDL male A. aegypti mosquitoes competed with ten wild type males inside this field house to mate with ten wild type females. Hatched larvae from mated females were screened under a fluorescent microscope for genetic markers to determine if they were fathered by RIDL male or wild type male, and all results were cross-checked by PCR. Two such experiments were conducted, each repeated sufficient number of times. All strains were on a Malaysian lab strain background for the first experiment, while the RIDL males alone were on a recently-colonised Mexican strain background for the second experiment. A total of 52 % of the matings were with RIDL males in the first experiment, while 45 % of the matings were with RIDL (Mexican) males in the second experiment. Statistically, this is not significantly different from 50 % of the matings expected to take place with RIDL males if the latter were as competitive as that of the wild type males. This shows that A. aegypti RIDL-513A has excellent mating competitiveness under semi-field conditions, verifying earlier trends obtained in small lab cages. We also observed high mating compatibility between recently-colonised Mexican RIDL males and lab-reared Malaysian wild type females.
Expression and biochemical characterization of a novel thermostable alkaline β-1,3-1,4-glucanase (lichenase) from an alkaliphilic Bacillus lehensis G1

Mat Yajit NL, Fazlin Hashim NH, Illias RM, Abdul Murad AM

Protein Expr Purif, 2024 Jul;219:106486.
PMID: 38642864 DOI: 10.1016/j.pep.2024.106486

New thermostable β-1,3-1,4-glucanase (lichenase) designated as Blg29 was expressed and purified from a locally isolated alkaliphilic bacteria Bacillus lehensis G1. The genome sequence of B. lehensis predicted an open reading frame of Blg29 with a deduced of 249 amino acids and a molecular weight of 28.99 kDa. The gene encoding for Blg29 was successfully amplified via PCR and subsequently expressed as a recombinant protein using the E. coli expression system. Recombinant Blg29 was produced as a soluble form and further purified via immobilized metal ion affinity chromatography (IMAC). Based on biochemical characterization, recombinant Blg29 showed optimal activity at pH9 and temperature 60 °C respectively. This enzyme was stable for more than 2 h, incubated at 50 °C, and could withstand ∼50 % of its activity at 70 °C for an hour and a half. No significant effect on Blg29 was observed when incubated with metal ions except for a small increase with ion Ca2+. Blg29 showed high substrate activity towards lichenan where Vm, Km, Kcat, and kcat/Km values were 2040.82 μmolmin‾1mg‾1, 4.69 mg/mL, and 986.39 s‾1 and 210.32 mLs‾1mg‾1 respectively. The high thermostability and activity make this enzyme useable for a broad prospect in industry applications.
Fulltext Identification of major and minor allergens of black tiger prawn (Penaeus monodon) and king prawn (Penaeus latisulcatus)

Sahabudin S, Misnan R, Yadzir ZH, Mohamad J, Abdullah N, Bakhtiar F, et al.

Malays J Med Sci, 2011 Jul;18(3):27-32.
PMID: 22135598 MyJurnal

BACKGROUND: Prawns and shrimp are a frequent cause of seafood allergy mediated by IgE antibodies. Penaeus monodon and Penaeus latisulcatus, commonly known as black tiger prawn and king prawn, respectively, are among the most frequently consumed prawns in Malaysia. The aim of this study was to identify the IgE-binding proteins of these 2 prawn species.
METHODS: Raw and boiled prawn extracts were prepared and then resolved by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). IgE-immunoblotting was then performed using sera from patients with positive skin prick tests to the raw prawn extracts.
RESULTS: SDS-PAGE analysis of the raw extracts of both prawn species revealed 23 protein bands; the boiled extracts yielded fewer protein bands. The bands in the range of 40 to 100 kDa were sensitive to heat and therefore were not found in the boiled extracts. Immunoblot of raw extracts of black tiger prawns and king prawns yielded 14 and 11 IgE-binding proteins, respectively, with molecular weights of between 15 and 200 kDa. Proteins at 36, 42, and 49 kDa were detected as the major allergens in both species of prawns. A protein of 75 kDa was also identified as a major allergen in black tiger prawns. Other potential allergens were also observed at various molecular masses.
CONCLUSION: Proteins of 36, 42, and 49 kDa were identified as the major allergens of both species of prawns. The 36 and 42 kDa proteins are hypothesised to be tropomyosin and arginine kinase, respectively. A high molecular weight protein of 75 kDa was found to be an additional major allergen in black tiger prawns.
KEYWORDS: Penaeus; allergens; allergy and clinical immunology; hypersensitivity; immunoblotting; tropomyosin
Cardiovascular complications in a diabetes prediction model using machine learning: a systematic review

Kee OT, Harun H, Mustafa N, Abdul Murad NA, Chin SF, Jaafar R, et al.

Cardiovasc Diabetol, 2023 Jan 19;22(1):13.
PMID: 36658644 DOI: 10.1186/s12933-023-01741-7

Prediction model has been the focus of studies since the last century in the diagnosis and prognosis of various diseases. With the advancement in computational technology, machine learning (ML) has become the widely used tool to develop a prediction model. This review is to investigate the current development of prediction model for the risk of cardiovascular disease (CVD) among type 2 diabetes (T2DM) patients using machine learning. A systematic search on Scopus and Web of Science (WoS) was conducted to look for relevant articles based on the research question. The risk of bias (ROB) for all articles were assessed based on the Prediction model Risk of Bias Assessment Tool (PROBAST) statement. Neural network with 76.6% precision, 88.06% sensitivity, and area under the curve (AUC) of 0.91 was found to be the most reliable algorithm in developing prediction model for cardiovascular disease among type 2 diabetes patients. The overall concern of applicability of all included studies is low. While two out of 10 studies were shown to have high ROB, another studies ROB are unknown due to the lack of information. The adherence to reporting standards was conducted based on the Transparent Reporting of a multivariable prediction model for Individual Prognosis or Diagnosis (TRIPOD) standard where the overall score is 53.75%. It is highly recommended that future model development should adhere to the PROBAST and TRIPOD assessment to reduce the risk of bias and ensure its applicability in clinical settings. Potential lipid peroxidation marker is also recommended in future cardiovascular disease prediction model to improve overall model applicability.
Fulltext Genetic Factors for Coronary Heart Disease and Their Mechanisms: A Meta-Analysis and Comprehensive Review of Common Variants from Genome-Wide Association Studies

Zarkasi KA, Abdullah N, Abdul Murad NA, Ahmad N, Jamal R

Diagnostics (Basel), 2022 Oct 21;12(10).
PMID: 36292250 DOI: 10.3390/diagnostics12102561

Genome-wide association studies (GWAS) have discovered 163 loci related to coronary heart disease (CHD). Most GWAS have emphasized pathways related to single-nucleotide polymorphisms (SNPs) that reached genome-wide significance in their reports, while identification of CHD pathways based on the combination of all published GWAS involving various ethnicities has yet to be performed. We conducted a systematic search for articles with comprehensive GWAS data in the GWAS Catalog and PubMed, followed by a meta-analysis of the top recurring SNPs from ≥2 different articles using random or fixed-effect models according to Cochran Q and I2 statistics, and pathway enrichment analysis. Meta-analyses showed significance for 265 of 309 recurring SNPs. Enrichment analysis returned 107 significant pathways, including lipoprotein and lipid metabolisms (rs7412, rs6511720, rs11591147, rs1412444, rs11172113, rs11057830, rs4299376), atherogenesis (rs7500448, rs6504218, rs3918226, rs7623687), shared cardiovascular pathways (rs72689147, rs1800449, rs7568458), diabetes-related pathways (rs200787930, rs12146487, rs6129767), hepatitis C virus infection/hepatocellular carcinoma (rs73045269/rs8108632, rs56062135, rs188378669, rs4845625, rs11838776), and miR-29b-3p pathways (rs116843064, rs11617955, rs146092501, rs11838776, rs73045269/rs8108632). In this meta-analysis, the identification of various genetic factors and their associated pathways associated with CHD denotes the complexity of the disease. This provides an opportunity for the future development of novel CHD genetic risk scores relevant to personalized and precision medicine.
Fulltext Monitoring of the h275y mutation in pandemic influenza A(H1N1) 2009 strains isolated in Malaysia

Suppiah J, Yusof MA, Othman KA, Saraswathy TS, Thayan R, Kasim FM, et al.

Southeast Asian J Trop Med Public Health, 2011 Jan;42(1):100-4.
PMID: 21323171

The 2009 pandemic influenza A(H1N1) infection in Malaysia was first reported in May 2009 and oseltamivir was advocated for confirmed cases in postexposure prophylaxis. However, there are cases of oseltamivir-resistance reported among H1N1-positive patients in other countries. Resistance is due to substitution of histidine by tyrosine at residue 275 (H275Y) of neuraminidase (NA). In this study, we have employed Sanger sequencing method to investigate the occurrence of mutations in NA segments of 67 pandemic 2009 A(H1N1) viral isolates from Malaysian patients that could lead to probable oseltamivir resistance. The sequencing analysis did not yield mutation at residue 275 for all 67 isolates indicating that our viral isolates belong to the wild type and do not confer resistance to oseltamivir.
Identification of Ige-binding proteins of raw and cooked extracts of Loligo edulis (white squid)

Yadzir ZH, Misnan R, Abdullah N, Bakhtiar F, Arip M, Murad S

Southeast Asian J Trop Med Public Health, 2010 May;41(3):653-9.
PMID: 20578555

Allergy to different classes of mollusks, including squid, which are members of the class Cephalopods has been reported. Tropomyosin, a major muscle protein, is the only well-recognized allergen in squid. The aim of this study was to characterize IgE-binding proteins of local Loligo edulis (white squid) consumed in Malaysia. Protein profiles and IgE-binding proteins were detected by sodium dodecyl sulfate-polyacrylamide gel-electrophoresis (SDS-PAGE) and immunoblotting using sera from 23 patients with positive skin prick test to raw squid extract. SDS-PAGE of the raw extract exhibited 21 protein bands (10-170 kDa) but those ranging from 19 to 29 kDa and 41 to 94 kDa were not found in the cooked extract. Immunoblotting of raw extract demonstrated 16 IgE-binding bands, ranging from 13 to 170 kDa. A heat-resistant 36 kDa protein, corresponding to squid tropomyosin, was identified as the major allergen of both extracts. In addition, a 50 kDa heat-sensitive protein was shown to be a major allergen of the raw extract. Our findings indicate that the allergen extract used for diagnosis of squid allergy should contain both the 36 kDa and 50 kDa proteins.
Fulltext Growth phase-dependent proteomes of the Malaysian isolated Lactococcus lactis dairy strain M4 using label-free qualitative shotgun proteomics analysis

Yap TW, Rabu A, Abu Bakar FD, Rahim RA, Mahadi NM, Illias RM, et al.

ScientificWorldJournal, 2014;2014:642891.
PMID: 24982972 DOI: 10.1155/2014/642891

Lactococcus lactis is the most studied mesophilic fermentative lactic acid bacterium. It is used extensively in the food industry and plays a pivotal role as a cell factory and also as vaccine delivery platforms. The proteome of the Malaysian isolated L. lactis M4 dairy strain, obtained from the milk of locally bred cows, was studied to elucidate the physiological changes occurring between the growth phases of this bacterium. In this study, ultraperformance liquid chromatography nanoflow electrospray ionization tandem mass spectrometry (UPLC- nano-ESI-MS(E)) approach was used for qualitative proteomic analysis. A total of 100 and 121 proteins were identified from the midexponential and early stationary growth phases, respectively, of the L. lactis strain M4. During the exponential phase, the most important reaction was the generation of sufficient energy, whereas, in the early stationary phase, the metabolic energy pathways decreased and the biosynthesis of proteins became more important. Thus, the metabolism of the cells shifted from energy production in the exponential phase to the synthesis of macromolecules in the stationary phase. The resultant proteomes are essential in providing an improved view of the cellular machinery of L. lactis during the transition of growth phases and hence provide insight into various biotechnological applications.
Fulltext Evaluation of reference genes for quantitative real-time PCR in oil palm elite planting materials propagated by tissue culture

Chan PL, Rose RJ, Abdul Murad AM, Zainal Z, Low ET, Ooi LC, et al.

PLoS One, 2014;9(6):e99774.
PMID: 24927412 DOI: 10.1371/journal.pone.0099774

The somatic embryogenesis tissue culture process has been utilized to propagate high yielding oil palm. Due to the low callogenesis and embryogenesis rates, molecular studies were initiated to identify genes regulating the process, and their expression levels are usually quantified using reverse transcription quantitative real-time PCR (RT-qPCR). With the recent release of oil palm genome sequences, it is crucial to establish a proper strategy for gene analysis using RT-qPCR. Selection of the most suitable reference genes should be performed for accurate quantification of gene expression levels.
Silica exposure is associated with an increased risk of developing ACPA-positive rheumatoid arthritis in an Asian population: evidence from the Malaysian MyEIRA case-control study

Yahya A, Bengtsson C, Larsson P, Too CL, Mustafa AN, Abdullah NA, et al.

Mod Rheumatol, 2014 Mar;24(2):271-4.
PMID: 24593203 DOI: 10.3109/14397595.2013.854076

OBJECTIVES: Silica exposure has been associated with an increased risk of developing rheumatoid arthritis (RA), especially among smokers. In this study, we aimed at examining the association between silica exposure (and its interaction with smoking) and the risk of RA in the Malaysian population.
METHODS: In total, 149 cases and 213 matched controls, all men, were included between August 2005 and December 2009. A case was defined as a person with early diagnosed RA using the 1987 American College of Rheumatology criteria for RA. Controls were randomly selected matched on sex, age and residential area. Silica exposure was defined as exposure to stone dust, rock drilling or stone crushing, and smoking status was categorized as ever/never cigarette smoking.
RESULTS: An increased risk of anti-citrullinated protein antibody (ACPA)-positive RA (OR = 2.4, 95 % CI 1.0-5.6) was observed among those exposed to silica. Ever-smokers exposed to silica had a particularly high risk of developing ACPA-positive RA (OR = 7.5, 95 % CI 2.3-24.2), compared with never-smokers not exposed to silica. No association was found regarding ACPA-negative RA.
CONCLUSION: Our data demonstrate that exposure to both silica and cigarette smoke comprise risks for developing RA in the Malaysian genetic context. The findings expand a link between environmental lung exposures and ACPA-positive RA to Asian populations.
Fulltext Development of a pyrG mutant of Aspergillus oryzae strain S1 as a host for the production of heterologous proteins

Ling SO, Storms R, Zheng Y, Rodzi MR, Mahadi NM, Illias RM, et al.

ScientificWorldJournal, 2013;2013:634317.
PMID: 24381522 DOI: 10.1155/2013/634317

The ease with which auxotrophic strains and genes that complement them can be manipulated, as well as the stability of auxotrophic selection systems, are amongst the advantages of using auxotrophic markers to produce heterologous proteins. Most auxotrophic markers in Aspergillus oryzae originate from chemical or physical mutagenesis that may yield undesirable mutations along with the mutation of interest. An auxotrophic A. oryzae strain S1 was generated by deleting the orotidine-5'-monophosphate decarboxylase gene (pyrG) by targeted gene replacement. The uridine requirement of the resulting strain GR6 pyrGΔ0 was complemented by plasmids carrying a pyrG gene from either Aspergillus nidulans or A. oryzae. β -Galactosidase expression by strain GR6 pyrGΔ0 transformed with an A. niger plasmid encoding a heterologous β -galactosidase was at least 150 times more than that obtained with the untransformed strain. Targeted gene replacement is thus an efficient way of developing auxotrophic mutants in A. oryzae and the auxotrophic strain GR6 pyrGΔ0 facilitated the production of a heterologous protein in this fungus.
Sequence and structural investigation of a novel psychrophilic α-amylase from Glaciozyma antarctica PI12 for cold-adaptation analysis

Ramli AN, Azhar MA, Shamsir MS, Rabu A, Murad AM, Mahadi NM, et al.

J Mol Model, 2013 Aug;19(8):3369-83.
PMID: 23686283 DOI: 10.1007/s00894-013-1861-5

A novel α-amylase was isolated successfully from Glaciozyma antarctica PI12 using DNA walking and reverse transcription-polymerase chain reaction (RT-PCR) methods. The structure of this psychrophilic α-amylase (AmyPI12) from G. antarctica PI12 has yet to be studied in detail. A 3D model of AmyPI12 was built using a homology modelling approach to search for a suitable template and to generate an optimum target-template alignment, followed by model building using MODELLER9.9. Analysis of the AmyPI12 model revealed the presence of binding sites for a conserved calcium ion (CaI), non-conserved calcium ions (CaII and CaIII) and a sodium ion (Na). Compared with its template-the thermostable α-amylase from Bacillus stearothermophilus (BSTA)-the binding of CaII, CaIII and Na ions in AmyPI12 was observed to be looser, which suggests that the low stability of AmyPI12 allows the protein to work at different temperature scales. The AmyPI12 amino acid sequence and model were compared with thermophilic α-amylases from Bacillus species that provided the highest structural similarities with AmyPI12. These comparative studies will enable identification of possible determinants of cold adaptation.
Fulltext Optimization of a heterologous signal peptide by site-directed mutagenesis for improved secretion of recombinant proteins in Escherichia coli

Jonet MA, Mahadi NM, Murad AM, Rabu A, Bakar FD, Rahim RA, et al.

J. Mol. Microbiol. Biotechnol., 2012;22(1):48-58.
PMID: 22456489 DOI: 10.1159/000336524

A heterologous signal peptide (SP) from Bacillus sp. G1 was optimized for secretion of recombinant cyclodextrin glucanotransferase (CGTase) to the periplasmic and, eventually, extracellular space of Escherichia coli. Eight mutant SPs were constructed using site-directed mutagenesis to improve the secretion of recombinant CGTase. M5 is a mutated SP in which replacement of an isoleucine residue in the h-region to glycine created a helix-breaking or G-turn motif with decreased hydrophobicity. The mutant SP resulted in 110 and 94% increases in periplasmic and extracellular recombinant CGTase, respectively, compared to the wild-type SP at a similar level of cell lysis. The formation of intracellular inclusion bodies was also reduced, as determined by sodium dodecyl sulfate-polyacrylamyde gel electrophoresis, when this mutated SP was used. The addition of as low as 0.08% glycine at the beginning of cell growth improved cell viability of the E. coli host. Secretory production of other proteins, such as mannosidase, also showed similar improvement, as demonstrated by CGTase production, suggesting that the combination of an optimized SP and a suitable chemical additive leads to significant improvements of extracellular recombinant protein production and cell viability. These findings will be valuable for the extracellular production of recombinant proteins in E. coli.

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