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Sequence analysis of Malaysian infectious bursal disease virus isolate and the use of reverse transcriptase nested polymerase chain reaction enzyme-linked immunosorbent assay for the detection of VP2 hypervariable region

Phong SF, Hair-Bejo M, Omar AR, Aini I

Avian Dis, 2003 Jan-Mar;47(1):154-62.
PMID: 12713171

The VP2 hypervariable region of P97/302 local infectious bursal disease virus (IBDV) isolate was amplified by the reverse transcriptase (RT) nested polymerase chain reaction (PCR) and cloned. This region of P97/302 local isolate was sequenced and compared with eight other reported IBDV sequences. The result showed that P97/302 IBDV was most identical to the reported very virulent IBDV strains because it has amino acid substitutions at positions 222, 256, 294, and 299, which encode alanine, isoleucine, isoleucine, and serine, respectively. This region can be digested with restriction enzymes of Taq1, Sty1, Ssp1 but not with Sac1. The P97/302 isolate was then used for the optimization of RT nested PCR enzyme-linked immunosorbent assay (ELISA). The RT nested PCR ELISA was able to detect 10(-4) dilution of the infected bursa homogenates and was 10 times more sensitive when compared with the agarose gel detection method. The RT nested PCR ELISA can detect up to 0.48 ng of the PCR product. The specificity of this nested PCR ELISA was also high (100%).
DNA vaccine encoding avian influenza virus H5 and Esat-6 of Mycobacterium tuberculosis improved antibody responses against AIV in chickens

Oveissi S, Omar AR, Yusoff K, Jahanshiri F, Hassan SS

Comp Immunol Microbiol Infect Dis, 2010 Dec;33(6):491-503.
PMID: 19781778 DOI: 10.1016/j.cimid.2009.08.004

The H5 gene of avian influenza virus (AIV) strain A/chicken/Malaysia/5744/2004(H5N1) was cloned into pcDNA3.1 vector, and Esat-6 gene of Mycobacterium tuberculosis was fused into downstream of the H5 gene as a genetic adjuvant for DNA vaccine candidates. The antibody level against AIV was measured using enzyme-linked immunosorbent assay (ELISA) and haemagglutination inhibition (HI) test. Sera obtained from specific-pathogen-free chickens immunized with pcDNA3.1/H5 and pcDNA3.1/H5/Esat-6 demonstrated antibody responses as early as 2 weeks after the first immunization. Furthermore, the overall HI antibody titer in chickens immunized with pcDNA3.1/H5/Esat-6 was higher compared to the chickens immunized with pcDNA3.1/H5 (p<0.05). The results suggested that Esat-6 gene of M. tuberculosis is a potential genetic adjuvant for the development of effective H5 DNA vaccine in chickens.
Fulltext Bioactive compounds and biological activities of Jatropha curcas L. kernel meal extract

Oskoueian E, Abdullah N, Ahmad S, Saad WZ, Omar AR, Ho YW

Int J Mol Sci, 2011;12(9):5955-70.
PMID: 22016638 DOI: 10.3390/ijms12095955

Defatted Jatropha curcas L. (J. curcas) seed kernels contained a high percentage of crude protein (61.8%) and relatively little acid detergent fiber (4.8%) and neutral detergent fiber (9.7%). Spectrophotometric analysis of the methanolic extract showed the presence of phenolics, flavonoids and saponins with values of 3.9, 0.4 and 19.0 mg/g DM, respectively. High performance liquid chromatography (HPLC) analyses showed the presence of gallic acid and pyrogallol (phenolics), rutin and myricetin (flavonoids) and daidzein (isoflavonoid). The amount of phorbol esters in the methanolic extract estimated by HPLC was 3.0 ± 0.1 mg/g DM. Other metabolites detected by GC-MS include: 2-(hydroxymethyl)-2 nitro-1,3-propanediol, β-sitosterol, 2-furancarboxaldehyde, 5-(hydroxymethy) and acetic acid in the methanolic extract; 2-furancarboxaldehyde, 5-(hydroxymethy), acetic acid and furfural (2-furancarboxaldehyde) in the hot water extract. Methanolic and hot water extracts of kernel meal showed antimicrobial activity against both Gram positive and Gram negative pathogenic bacteria (inhibition range: 0-1.63 cm) at the concentrations of 1 and 1.5 mg/disc. Methanolic extract exhibited antioxidant activities that are higher than hot water extract and comparable to β-carotene. The extracts tended to scavenge the free radicals in the reduction of ferric ion (Fe(3+)) to ferrous ion (Fe(2+)). Cytotoxicity assay results indicated the potential of methanolic extract as a source of anticancer therapeutic agents toward breast cancer cells.
Development of a multiplex real-time PCR assay using SYBR Green 1 chemistry for simultaneous detection and subtyping of H9N2 influenza virus type A

Ong WT, Omar AR, Ideris A, Hassan SS

J Virol Methods, 2007 Sep;144(1-2):57-64.
PMID: 17512062

Avian influenza viruses are pathogens of economical and public health concerns. However, infections caused by low pathogenic avian influenza particularly H9N2 subtype are not associated with clear clinical features. Hence, rapid detection and subtyping of the virus will enable immediate measures to be implemented for preventing widespread transmission. This study highlights the development of a multiplex real-time reverse-transcriptase polymerase chain reaction (RRT-PCR) assay using SYBR Green 1 chemistry for universal detection of avian influenza viruses and specific subtyping of H9N2 isolates based on melting temperatures (T(m)) discriminations. Three melting peaks generated simultaneously at temperatures 85.2+/-1.0, 81.9+/-0.9 and 78.7+/-0.9 degrees C represent NP, H9 and N2 gene products, respectively. The RRT-PCR assay was about 10-100-fold more sensitive when compared to the conventional RT-PCR method using reference H9N2 isolate. In addition, the RRT-PCR assay was 100% sensitive as well as 92% specific according to the standard virus isolation method in detecting experimentally infected specific-pathogen-free (SPF) chickens.
Fulltext Cloning and expression of the HN gene from the velogenic viscerotropic Newcastle disease virus strain AF2240 in Sf9 insect cells

Ong HK, Ali AM, Omar AR, Yusoff K

Cytotechnology, 2000 Mar;32(3):243-51.
PMID: 19002985 DOI: 10.1023/A:1008136326756

The haemagglutinin-neuraminidase (HN) gene ofNewcastle disease virus (NDV) strain AF2240, amplifiedfrom the viral genomic RNA ( approximately 1.8 kb) was directionallycloned and inserted into a baculovirus expressionvector system. The recombinant glycoprotein expressedin Spodoptera frugiperda (Sf9) cellsshowed haemagglutinin (HA), neuraminidase (NA) andhemadsorption activities. HA activity was detected inboth extra- and intra-cellular recombinant HN(recHNAF2240) samples. In addition, both HA andhemadsorption activities were inhibited by polyclonalanti-NDV sera. Furthermore, significant expression ofthe recombinant protein was observed on the surface ofinfected cells. SDS-PAGE analysis revealed thepresence of visually distinguishable bands between the70 and 80 kDa in size that were absent in thewild-type samples. Western blot analysis showed thatthe distinct approximately 63 kDa band and a approximately 75 kDa bandcorresponded to the unglycosylated and glycosylated HNglycoprotein respectively as reported in anotherstudy. These observations indicated that the HNrecombinant protein was not only expressed on thesurface of the infected cells as well as with theviral coat protein, but also appears to be functional.
Fulltext An Influenza A Vaccine Based on the Extracellular Domain of Matrix 2 Protein Protects BALB/C Mice Against H1N1 and H3N2

Ong HK, Yong CY, Tan WS, Yeap SK, Omar AR, Razak MA, et al.

Vaccines (Basel), 2019 08 19;7(3).
PMID: 31430965 DOI: 10.3390/vaccines7030091

Current seasonal influenza A virus (IAV) vaccines are strain-specific and require annual reconstitution to accommodate the viral mutations. Mismatches between the vaccines and circulating strains often lead to high morbidity. Hence, development of a universal influenza A vaccine targeting all IAV strains is urgently needed. In the present study, the protective efficacy and immune responses induced by the extracellular domain of Matrix 2 protein (M2e) displayed on the virus-like particles of Macrobrachium rosenbergii nodavirus (NvC-M2ex3) were investigated in BALB/c mice. NvC-M2ex3 was demonstrated to be highly immunogenic even in the absence of adjuvants. Higher anti-M2e antibody titers corresponded well with increased survival, reduced immunopathology, and morbidity of the infected BALB/c mice. The mice immunized with NvC-M2ex3 exhibited lower H1N1 and H3N2 virus replication in the respiratory tract and the vaccine activated the production of different antiviral cytokines when they were challenged with H1N1 and H3N2. Collectively, these results suggest that NvC-M2ex3 could be a potential universal influenza A vaccine.
Efficacy of VP2 protein expressed in E. coli for protection against highly virulent infectious bursal disease virus

Omar AR, Kim CL, Bejo MH, Ideris A

J Vet Sci, 2006 Sep;7(3):241-7.
PMID: 16871018

The ability of a heat-inactivated whole virus from a highly virulent infectious bursal disease virus (hvIBDV) and VP2 protein from hvIBDV expressed in E. coli provided protection against a hvIBDV challenge in specificpathogen-free (SPF) chickens. Six out of seven chickens that were injected three times with crude VP2 protein developed significant antibody titer against IBDV. However, only four out of the seven chickens survived the hvIBDV challenge. Despite showing low antibody titer profiles, all chickens immunized with the heat-inactivated whole virus also survived the challenged with hvIBDV. However, all of these chickens had bursal atrophy and mild to moderate depletion of lymphocytes. Thus, antibodies raised against IBDV VP2 protein expressed in E. coli and denatured IBDV proteins induced some degree of protection against mortality but not against bursal damage following challenge with hvIBDV.
Fulltext Clinical predictors of a positive troponin T test in patients presenting with probable acute coronary syndromes

Omar AR, Ping C, Tan HC, Lim YT

Med J Malaysia, 2005 Mar;60(1):50-3.
PMID: 16250280

Acute coronary syndrome (ACS) patients with positive troponin T (TnT) test are at higher risk for death and myocardial reinfarction. They would significantly benefit from early aggressive pharmacologic and invasive therapy. However, TnT test is not widely available. This retrospective study of 173 patients with ACS showed: that prolonged or repetitive episodes of angina at rest in the previous 24 hours (p = 0.01) and evidence of myocardial ischaemia on ECG (p < 0.001) were associated with positive TnT tests (> or = 0.1 ng/mL). The two variables in combination showed 100% positive predictive value, facilitating early identification and streamlining of therapy.
Fulltext Acute ophthalmic artery occlusion in decompression illness with underlying anterior cerebral artery A1 segment hypoplasia

Omar AR, Ibrahim M, Hussein A

Diving Hyperb Med, 2018 Jun 30;48(2):112-113.
PMID: 29888385 DOI: 10.28920/dhm48.2.112-113

A diver presented with total loss of vision in the left eye and right hemiparesis following a routine no-stop scuba dive to 20 metres' depth. A diagnosis of decompression illness (DCI) with acute ophthalmic artery air embolism and left carotid artery insult causing acute anterior circulatory ischaemia was made. He underwent seven hyperbaric treatments leading to a full recovery. Magnetic resonance angiography revealed an underlying left anterior cerebral artery A1 segment hypoplasia. Making a prompt diagnosis and early hyperbaric oxygen treatment are crucial to halt further tissue damage from ischaemia in central nervous system DCI. In this case, the finding of a left A1 anterior cerebral artery segment hypoplasia variant may have increased the severity of DCI due to deficient collateral circulation.
Fulltext Evaluation of Cyclooxygenase-2 and p53 Expression in Pterygium Tissue Following Preoperative Intralesional Ranibizumab Injection

Omar AR, Ibrahim M, Jaafar H, Siti-Azrin AH, Zunaina E

Front Med (Lausanne), 2021;8:733523.
PMID: 35004714 DOI: 10.3389/fmed.2021.733523

Introduction: Overexpression of vascular endothelial growth factor (VEGF), cyclooxygenase-2 (COX-2), and p53 are the postulated aetiopathogenesis in pterygium. VEGF is responsible for the induction of COX-2 expression, whereas p53 plays an important role in the regulation of VEGF. This study aimed to evaluate the immunohistochemistry of COX-2 and p53 expressions from excised pterygium tissue from patients who received intralesional ranibizumab (anti-VEGF) injection 2 weeks prior to pterygium surgery. Materials and Methods: An interventional comparative study involving patients presenting with primary pterygium was conducted between September 2015 and November 2017. The patients were randomized into either the intervention or control group. Patients in the intervention group were injected with intralesional ranibizumab (0.5 mg/0.05 ml) 2 weeks prior to surgery. Both groups underwent pterygium excision followed by conjunctival autograft. Immunohistochemistry staining was performed to evaluate COX-2 and p53 expressions in the excised pterygium tissue. Results: A total of 50 patients (25 in both the intervention and control groups) were recruited. There were 34 (68%) patients with grade III pterygium and 16 (32%) patients with grade IV pterygium. There was statistically significant difference in reduction of COX-2 expression in the epithelial layer [84.0% (95% CI: 63.9, 95.5)] (p = 0.007) and stromal layer [84.0% (95% CI: 63.9, 95.5)] (p < 0.001) between intervention and control groups. There was no significant difference in the reduction of p53 expression between the two groups. Conclusion: This study demonstrated the possible use of intralesional anti-VEGF treatment prior to pterygium excision as a potential future modality of adjunctive therapy for pterygium surgery.
Fulltext An overview on the development of newcastle disease virus as an anti-cancer therapy

Omar AR, Ideris A, Ali AM, Othman F, Yusoff K, Abdullah JM, et al.

Malays J Med Sci, 2003 Jan;10(1):4-12.
PMID: 23365495

Newcastle disease virus (NDV) is one of the most economically important avian virus which affects the poultry industry worldwide. Although NDV is being very actively studied in Malaysia, there are still no studies on its potential as an anticancer agent, a new approach to treating cancer known as virotherapy. Currently, a collaborative research is being undertaken between Universiti Putra Malaysia (UPM), Universiti Sains Malaysia (USM) and Majlis Kanser Nasional (MAKNA) in characterising various local NDV isolates as anticancer agent. This paper describes an overview of the research that have been carried out worldwide in the use of NDV for cancer treatment and also some of our findings in characterising local NDVs with oncolytic properties.
Molecular characterization of recent infectious bursal disease virus isolates from Malaysia

Nurulfiza I, Hair-Bejo M, Omar AR, Aini I

Acta Virol., 2006;50(1):45-51.
PMID: 16599185

Three isolates of Infectious bursal disease virus (IBDV), designated UPM04178, UPM04190 and UPM04238, were obtained from severe outbreaks of infectious bursal disease (IBD) in Malaysia in 2004. The hypervariable region (HPVR) of VP2 gene of these isolates was sequenced. The obtained sequences were compared with those of other isolates. The highest similarity (98%) concerning both nucleotide and amino acid sequences was found to very virulent IBDV (vvIBDV) strains. Phylogenetic analysis revealed clustering of the three isolates with vvIBDV strains. Evolutionary relatedness of the three isolates to vvIBDV strains was demonstrated by three phylogenetic methods: bootstrap values of 100%, 95% and 90% for nucleotide sequences and those of 58%, 86% and 96% for amino acid sequences were obtained by the distance, maximum parsimony and maximum likehood methods, respectively. It is concluded that UPM04178, UPM04190 and UPM04238 are vvIBDV isolates of serotype 1, which originate from a common ancestor of IBDV strains present in Malaysia.
Immunochromatographic gold-based test strip for rapid detection of Infectious bursal disease virus antibodies

Nurulfiza I, Hair-Bejo M, Omar AR, Aini I

J Vet Diagn Invest, 2011 Mar;23(2):320-4.
PMID: 21398455

The immunochromatographic assay is an alternative method for simple and rapid detection of Infectious bursal disease virus (IBDV) in chickens using colloidal gold-antibody conjugate. The whole-virus antigen of IBDV (UPM04190 isolate) and the high-affinity polyclonal antibodies directed against IBDV were blotted onto nitrocellulose membranes for test and control lines, respectively. Evaluation of the strip was performed using serum samples from experimentally and naturally infected chickens. The results showed that the test strip was more sensitive than the commercial enzyme-linked immunosorbent assay (ELISA) because it could detect a dilution factor up to 120,000 (250 ELISA units) for positive samples. It was also specific, in that it detected IBDV antibodies and did not cross-react with antibodies to other chicken viruses. The method was rapid (2 min) in both clinical and field environments with samples needing only a minimum amount (50 µl) of blood to produce an acceptable detection signal. The pen-site test strip proved successful in monitoring the immune status of chickens against the IBDV infection.
Isolation and Genetic Characterization of Canine Parvovirus in a Malayan Tiger

Nur-Farahiyah AN, Kumar K, Yasmin AR, Omar AR, Camalxaman SN

Front Vet Sci, 2021;8:660046.
PMID: 34414223 DOI: 10.3389/fvets.2021.660046

Naïve Felidae in the wild may harbor infectious viruses of importance due to cross-species transmission between the domesticated animals or human-wildlife contact. However, limited information is available on virus shedding or viremia in the captive wild felids, especially in Malaysia. Four infectious viruses of cat, feline herpesvirus (FHV), feline calicivirus (FCV), canine distemper virus (CDV), and canine parvovirus (CPV), were screened in leopards, feral cats, and tigers in Malaysia based on virus isolation in Crandell-Rees feline kidney (CRFK) cells, PCR/RT-PCR, and whole-genome sequencing analysis of the positive isolate. From a total of 36 sera collected, 11 samples showed three consecutive cytopathic effects in the cell culture and were subjected to PCR using specific primers for FHV, FCV, CDV, and CPV. Only one sample from a Malayan tiger was detected positive for CPV. The entire viral genome of CPV (UPM-CPV15/P. tigris jacksoni; GenBank Accession number MW380384) was amplified using the Sanger sequencing approach. Genome sequencing of the isolate revealed 99.13, 98.65, and 98.40% close similarity to CPV-31, CPV-d Cornell #320, and CPV-15 strains, respectively, and classified as CPV-2a. Time-scaled Bayesian Maximum Clade Credibility tree for the non-structural (NS) genes of CPV showed a close relationship to the isolates CPV-CN SD6_2014 and KSU7-SD_2004 from China and USA, respectively, while the capsid gene showed the same ancestor as the FPV-BJ04 strain from China. The higher evolution rate of the capsid protein (CP) (VP 1 and VP2) [1.649 × 10-5 (95% HPD: 7.626 × 10-3 to 7.440 × 10-3)] as compared to the NS gene [1.203 × 10-4 (95% HPD: 6.663 × 10-3 to 6.593 × 10-3)] was observed in the CPV from this study, and fairly higher than other parvovirus species from the Protoparvovirus genus. Genome sequencing of the isolated CPV from a Malayan tiger in the present study provides valuable information about the genomic characteristics of captive wild felids, which may add information on the presence of CPV in species other than dogs.
Fulltext Virus-like Particle Vaccines: A Prospective Panacea Against an Avian Influenza Panzootic

Ninyio NN, Ho KL, Omar AR, Tan WS, Iqbal M, Mariatulqabtiah AR

Vaccines (Basel), 2020 Nov 19;8(4).
PMID: 33227887 DOI: 10.3390/vaccines8040694

Epizootics of highly pathogenic avian influenza (HPAI) have resulted in the deaths of millions of birds leading to huge financial losses to the poultry industry worldwide. The roles of migratory wild birds in the harbouring, mutation, and transmission of avian influenza viruses (AIVs), and the lack of broad-spectrum prophylactic vaccines present imminent threats of a global panzootic. To prevent this, control measures that include effective AIV surveillance programmes, treatment regimens, and universal vaccines are being developed and analysed for their effectiveness. We reviewed the epidemiology of AIVs with regards to past avian influenza (AI) outbreaks in birds. The AIV surveillance programmes in wild and domestic birds, as well as their roles in AI control were also evaluated. We discussed the limitations of the currently used AI vaccines, which necessitated the development of a universal vaccine. We evaluated the current development of AI vaccines based upon virus-like particles (VLPs), particularly those displaying the matrix-2 ectodomain (M2e) peptide. Finally, we highlighted the prospects of these VLP vaccines as universal vaccines with the potential of preventing an AI panzootic.
Fulltext In Vitro Evaluation of Curcumin-Encapsulated Chitosan Nanoparticles against Feline Infectious Peritonitis Virus and Pharmacokinetics Study in Cats

Ng SW, Selvarajah GT, Hussein MZ, Yeap SK, Omar AR

Biomed Res Int, 2020;2020:3012198.
PMID: 32596292 DOI: 10.1155/2020/3012198

Feline infectious peritonitis (FIP) is an important feline viral disease, causing an overridden inflammatory response that results in a high mortality rate, primarily in young cats. Curcumin is notable for its biological activities against various viral diseases; however, its poor bioavailability has hindered its potential in therapeutic application. In this study, curcumin was encapsulated in chitosan nanoparticles to improve its bioavailability. Curcumin-encapsulated chitosan (Cur-CS) nanoparticles were synthesised based on the ionic gelation technique and were spherical and cuboidal in shape, with an average particle size of 330 nm and +42 mV in zeta potential. The nanoparticles exerted lower toxicity in Crandell-Rees feline kidney (CrFK) cells and enhanced antiviral activities with a selective index (SI) value three times higher than that of curcumin. Feline-specific bead-based multiplex immunoassay and qPCR were used to examine their modulatory effects on proinflammatory cytokines, including tumour necrosis factor (TNF)α, interleukin- (IL-) 6, and IL-1β. There were significant decrements in IL-1β, IL-6, and TNFα expression in both curcumin and Cur-CS nanoparticles. Based on the multiplex immunoassay, curcumin and the Cur-CS nanoparticles could lower the immune-related proteins in FIP virus (FIPV) infection. The single- and multiple-dose pharmacokinetics profiles of curcumin and the Cur-CS nanoparticles were determined by high-performance liquid chromatography (HPLC). Oral delivery of the Cur-CS nanoparticles to cats showed enhanced bioavailability with a maximum plasma concentration (Cmax) value of 621.5 ng/mL. Incorporating chitosan nanoparticles to deliver curcumin improved the oral bioavailability and antiviral effects of curcumin against FIPV infection. This study provides evidence for the potential of Cur-CS nanoparticles as a supplementary treatment of FIP.
Fulltext Cellular Metabolic Profiling of CrFK Cells Infected with Feline Infectious Peritonitis Virus Using Phenotype Microarrays

Ng SW, Selvarajah GT, Cheah YK, Mustaffa Kamal F, Omar AR

Pathogens, 2020 May 25;9(5).
PMID: 32466289 DOI: 10.3390/pathogens9050412

Feline infectious peritonitis (FIP) is a fatal feline immune-mediated disease caused by feline infectious peritonitis virus (FIPV). Little is known about the biological pathways associated in FIP pathogenesis. This is the first study aiming to determine the phenotypic characteristics on the cellular level in relation to specific metabolic pathways of importance to FIP pathogenesis.
METHODS: The internalization of type II FIPV WSU 79-1146 in Crandell-Rees Feline Kidney (CrFK) cells was visualized using a fluorescence microscope, and optimization prior to phenotype microarray (PM) study was performed. Then, four types of Biolog Phenotype MicroArray™ plates (PM-M1 to PM-M4) precoated with different carbon and nitrogen sources were used to determine the metabolic profiles in FIPV-infected cells.
RESULTS: The utilization of palatinose was significantly low in FIPV-infected cells; however, there were significant increases in utilizing melibionic acid, L-glutamine, L-glutamic acid and alanyl-glutamine (Ala-Gln) compared to non-infected cells.
CONCLUSION: This study has provided the first insights into the metabolic profiling of a feline coronavirus infection in vitro using PMs and deduced that glutamine metabolism is one of the essential metabolic pathways for FIPV infection and replication. Further studies are necessary to develop strategies to target the glutamine metabolic pathway in FIPV infection.
Characterization of Malaysian velogenic NDV strain AF2240-I genomic sequence: a comparative study

Murulitharan K, Yusoff K, Omar AR, Molouki A

Virus Genes, 2013 Jun;46(3):431-40.
PMID: 23306943 DOI: 10.1007/s11262-012-0874-y

Newcastle disease virus (NDV) strain AF2240 is a viscerotropic velogenic strain that is used as a vaccine challenge virus in Malaysia. The identification of the full length genome will be a crucial platform for further studies of this isolate. In this study, we fully sequenced the genome of a derivative of this strain named AF2240-I. The 15,192 nt long genome contains a 55-nt leader sequence at the 3' whereas the trailer region consists of 114 nt at the 5'. The intergenic sequences between the NP-P, P-M, M-F, F-HN, and HN-L genes comprise 1, 1, 1, 31, and 47 nt, respectively. The acknowledged cleavage site of fusion protein showed amino acid sequence of 112-R-R-Q-K-R-F-117, which corresponds to those of virulent NDV strains. Phylogenetic analysis of the whole virus genome shows that the strain AF2240-I belongs to genotype VIII and is more closely related to velogenic strains QH1, QH4, Fontana, Largo, and Italienas compared to other strains of NDV. Differences are noticed in the hemagglutinin-neuraminidase (HN) and matrix (M) gene between AF2240 and its derivative AF2240-I. This is the first report of a complete genome sequence of an NDV strain isolated in Malaysia.
Fulltext Rapid Generation of a Recombinant Genotype VIII Newcastle Disease Virus (NDV) Using Full-Length Synthetic cDNA

Murulitharan K, Yusoff K, Omar AR, Peeters BPH, Molouki A

Curr Microbiol, 2021 Apr;78(4):1458-1465.
PMID: 33660046 DOI: 10.1007/s00284-021-02421-z

Rescue of (-)ssRNA viruses involves the sequential assembly and cloning of the full-length cDNA, which is often a challenging and time-consuming process. The objective of this study was to develop a novel method to rapidly clone the full-length cDNA of a very virulent NDV by only one assembly step. A completely synthetic 15 kb cDNA of a Malaysian genotype VIII NDV known as strain AF2240-I with additional flanking BsmBI sites was synthesised. However, to completely follow the rule-of-six, the additional G residues that are traditionally added after the T7 promoter transcription initiation site were not synthesised. The synthetic fragment was then cloned into low-copy number transcription vector pOLTV5-phiX between the T7 promoter and HDV Rz sequences through digestion with BbsI. The construct was co-transfected with helper plasmids into BSRT7/5 cells. A recombinant NDV called rAF was successfully rescued using transfection supernatant harvested as early as 16 h post-transfection. Virus from each passage showed an intracerebral pathogenicity index (ICPI) and a mean death time (MDT) similar to the parent strain AF2240-I. Moreover, rAF possessed an introduced mutation which was maintained for several passages. The entire rescue using the one-step assembly procedure was completed within a few weeks, which is extremely fast compared to previously used methods.
Lactation failure in crossbred Sahiwal Friesian cattle

Murugaiyah M, Ramakrishnan P, Omar AR, Knight CH, Wilde CJ

J Dairy Res, 2001 May;68(2):165-74.
PMID: 11504381

Milk producers in Malaysia make extensive use of crossbred Sahiwal Friesian dairy cattle. These animals have, however, been found susceptible to lactation failure. A survey of cows in an experimental herd of F1 Sahiwal Friesian animals indicated that, in 30% of animals, milk yield decreased to negligible levels within the first 8 weeks post partum. Lactation failure was associated with a progressive increase in the amount of residual milk left in the udder after normal milking. By week 3 of lactation, residual milk volume was significantly greater than that in animals that, based on previous lactation history were not susceptible to lactation failure, and accounted for up to 30% of milk available at the morning milking. The cellular consequences of residual milk accumulation were evident in the activities of acetyl-CoA carboxylase, fatty acid synthetase and galactosyltransferase, key enzyme markers of cellular differentiation, which decreased in glands undergoing lactation failure and were lower than values measured in tissue of control cows. Mammary cell number, estimated by tissue DNA content, was also reduced in animals undergoing lactation failure. These indices of mammary development indicate that lactation failure is the result of premature involution in susceptible animals. Premature involution is a predictable consequence of progressive milk stasis in failing lactation, and attributable to an increase in autocrine feedback by inhibitory milk constituents. The progressive increase in residual milk is, on the other hand, unlikely to be attributable to impaired mammary development. Measurements of milk storage during milk accumulation showed no differences between control and lactation failure cows in the distribution of milk between alveolar and cisternal storage compartments. We conclude that lactation failure in Sahiwal Friesian cows is due to a failure of milk removal, and probably the result of an impaired milk ejection reflex rather than to the glands' milk storage characteristics.

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