Displaying publications 1 - 20 of 85 in total

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  1. Abdul Rahman NA, Mohd Desa MN, Masri SN, Taib NM, Sulaiman N, Hazman H, et al.
    Pol J Microbiol, 2023 Jun 01;72(2):103-115.
    PMID: 37314355 DOI: 10.33073/pjm-2023-023
    Streptococcus pneumoniae (pneumococcus) belongs to the Gram-positive cocci. This bacterium typically colonizes the nasopharyngeal region of healthy individuals. It has a distinct polysaccharide capsule - a virulence factor allowing the bacteria to elude the immune defense mechanisms. Consequently, it might trigger aggressive conditions like septicemia and meningitis in immunocompromised or older individuals. Moreover, children below five years of age are at risk of morbidity and mortality. Studies have found 101 S. pneumoniae capsular serotypes, of which several correlate with clinical and carriage isolates with distinct disease aggressiveness. Introducing pneumococcal conjugate vaccines (PCV) targets the most common disease-associated serotypes. Nevertheless, vaccine selection pressure leads to replacing the formerly dominant vaccine serotypes (VTs) by non-vaccine types (NVTs). Therefore, serotyping must be conducted for epidemiological surveillance and vaccine assessment. Serotyping can be performed using numerous techniques, either by the conventional antisera-based (Quellung and latex agglutination) or molecular-based approaches (sequetyping, multiplex PCR, real-time PCR, and PCR-RFLP). A cost-effective and practical approach must be used to enhance serotyping accuracy to monitor the prevalence of VTs and NVTs. Therefore, dependable pneumococcal serotyping techniques are essential to precisely monitor virulent lineages, NVT emergence, and genetic associations of isolates. This review discusses the principles, associated benefits, and drawbacks of the respective available conventional and molecular approaches, and potentially the whole genome sequencing (WGS) to be directed for future exploration.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  2. Fu JYL, Chong YM, Sam IC, Chan YF
    J Virol Methods, 2022 Mar;301:114462.
    PMID: 35026305 DOI: 10.1016/j.jviromet.2022.114462
    Emerging SARS-CoV-2 variants of concern (VOC) have been associated with enhanced transmissibility and immune escape. Next-generation sequencing (NGS) of the whole genome is the gold standard for variant identification for surveillance but is time-consuming and costly. Rapid and cost-effective assays that detect SARS-CoV-2 variants are needed. We evaluated Allplex SARS-CoV-2 Master Assay and Variants I Assay to detect HV69/70 deletion, Y144 deletion, E484K, N501Y, and P681H spike mutations in 248 positive samples collected in Kuala Lumpur, Malaysia, between January and May 2021. Spike variants were detected in 78/248 (31.5 %), comprising 60 VOC B.1.351 (beta) and 18 B.1.1.7 (alpha). With NGS as reference for 115 samples, the sensitivity for detecting the spike mutations was 98.7 % with the Master Assay and 100 % with the Variants I Assay. The emergence of beta variants correlated with increasing COVID-19 infections in Malaysia. The prevalence of alpha VOC and lineage B.1.466.2 was low. These assays detect mutations present in alpha, beta and gamma VOCs. Of the VOCs which have subsequently emerged, the assays should detect omicron (B.1.1.529) but not B.1.617.2 (delta). In conclusion, spike variant PCR assays can be used to rapidly monitor selected SARS-CoV-2 VOCs in resource-limited settings, but require updates as new variants emerge.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  3. Zahari Z, Salleh MR, Zahri Johari MK, Musa N, Ismail R
    Malays J Med Sci, 2011 Oct;18(4):44-57.
    PMID: 22589672 MyJurnal
    The dopamine D2 receptor gene (DRD2) plays a role in many diseases such as schizophrenia, Parkinson's disease, and addictive behaviour. Methods currently available for the detection of DRD2 polymorphisms are costly and cannot detect all 8 polymorphisms of our research interest simultaneously (Val96Ala, Leu141Leu, Val154Ile, Pro310Ser, Ser311Cys, TaqI A, A-241G, and -141C Ins/Del). Therefore, we developed a nested multiplex polymerase chain reaction (PCR) for simultaneous detection of these polymorphisms.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  4. Thong KL, Teh CS, Chua KH
    Trop Biomed, 2014 Dec;31(4):689-97.
    PMID: 25776594 MyJurnal
    The present study aims to develop a system which consists of four pairs of primers that specifically detects Salmonella spp., Salmonella serovar Typhi and Salmonella serovar Paratyphi A with an internal amplification control. The system, when applied in Polymerase Chain Reaction (PCR) under specific conditions, reaction mixture and cycling temperatures produced four bands; 784 bp, 496 bp, 332 bp and 187 bp. The DNA band 784 bp is present in all Salmonella spp., while the bands of 496 bp and 332 bp are only present in S. Paratyphi A and S. Typhi, respectively. An internal amplification control as indicated by the 187 bp shows the system is working in optimum condition in all the tests. This multiplex PCR was evaluated on 241 bacterial cultures and 691 naturally contaminated samples. Overall, this multiplex PCR detection system provides a single step for simultaneous detection of DNAs of Salmonella spp., S. Typhi and S. Paratyphi A.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*; Multiplex Polymerase Chain Reaction/standards
  5. Hanapi UK, Desa MN, Ismail A, Mustafa S
    J Food Sci Technol, 2015 Jul;52(7):4166-75.
    PMID: 26139881 DOI: 10.1007/s13197-014-1459-7
    A Common Primer Multiplex PCR (CP-M-PCR) was developed to detect meat origin of four groups of animal (pig, ruminant, avian and rabbit). This method demonstrated higher sensitivity and efficiency than the conventional multiplex PCR. In this approach, a common forward primer was designed in the 5' end of a homologous region of mitochondrial NADH dehyrogenase subunit 4 (Nad 4) gene sequences of all the animal groups. Specific adapter reverse primers were designed by adding an adapter sequence at the 5' end. The same adapter sequence was used as the common adapter reverse primer. The primers generated specific fragments of 267, 370, 504, and 548 bp lengths for pig, ruminant, avian and rabbit meats, respectively. The use of adapter sequence at the 5' end of the common adapter reverse primers increased the efficiency of the amplification and the application of a common forward primer solved the complexity in multiplex PCR system. Bands of specific amplification can be detected in the PCR assays containing as low as 10(-6) μM of adapter reverse primer. This result indicated that the sensitivity was tremendously increased as compared to the conventional multiplex PCR (10(-3) μM). CP-M-PCR detection limit of the DNA samples was 0.1 ng for the four groups of meats. CP-M-PCR has greatly improved the sensitivity and efficiency of the PCR system for a more reliable and accurate outcome than conventional multiplex PCR system.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  6. Zeti Norfidiyati Salmuna, Murnihayati Hassan, Habsah Hasan, Zakuan Zainy Deris
    MyJurnal
    Carpanenamase-producing Enterobacteriaceae (CPE) has emerged as a threat to hospitalized patients. Phenotypic test such as Modified hodge test was less sensitive and specific especially to detect blaNDM-1 which is the most predominant genotype in this region. Nucleic acid amplification technology offers improved specificity and sensitivity. Failed amplification due to the presence of inhibitors is a limitation. In this study, we tried to use previous method described by Villumseen et al with some modification using another DNA extraction kit. Methods: Ten mls of sterile whole blood taken from nearly expired blood bag from blood bank was spiked with 200 μl of 0.5mcFarland bacterial suspension from thirty-six confirmed isolates of blaNDM-1 carbapenamase-producing Klebsiella pneumoniae in an aerobic Bactec Plus and incubated until the growth was detected. The blood specimen was subjected to DNA extraction method using Macherey-Nachel, Nucleospin® Blood QuickPure followed with multiplex PCR. Results: Out of the 36 isolates, 12 isolates revealed blaNDM-1 , 9 isolates revealed blaNDM-1 and blaOXA-48, 7 isolates revealed blaNDM-1, blaVIM and blaKPC genotypes that were amplified at cycle threshold of less than 30. Another 8 isolates could not pick up any genotypes possibly due to pipetting error as all the internal control were amplified. Eight true negative gram negative isolates underwent same procedure and none amplified at a cycle threshold less than 30. Conclusion: This modified method was proved to give a high yield of CPE genotypes with the cycle threshold was set at less than or equal to 30 and able to overcome the presence of PCR inhibitors.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  7. Jeshveen, S.S., Chai, L.C., Pui, C.F., Son, R.
    MyJurnal
    The main source of E. coli 0157:H7 is cattle, but recent studies showed high percentage of outbreaks
    contributed by contaminated water. The occurrence of E. coli O157:H7 in environmental water samples poses a potential threat to human health. The aim of this study was to establish a protocol for the detection of the pathogen E. coli O157:H7 and E. coli virulence genes (eaeA, rfbE, hly, stx1, and stx2) in a multiplex PCR protocol using six specific primer pairs. The target genes produced species-specific amplicons at 625 bp, 397 bp, 296 bp, 166 bp, 210 bp and 484 bp for E. coli O157:H7 (fliCh7 gene) and virulence genes (eaeA, rfbE, hly, stx1, and stx2) respectively. The results obtained show that the established PCR protocol is suitable for a rapid and specific analysis of the pathogenic E. coli O157:H7 in environmental water samples for the assessment of microbiological risks.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  8. Tsai MH, Chan CK, Chang YC, Lin CH, Liou CW, Chang WN, et al.
    Front Neurol, 2018;9:515.
    PMID: 30034362 DOI: 10.3389/fneur.2018.00515
    Objective: Focal epilepsy is the most common subtype of epilepsies in which the influence of underlying genetic factors is emerging but remains largely uncharacterized. The purpose of this study is to determine the contribution of currently known disease-causing genes in a large cohort (n = 593) of common focal non-lesional epilepsy patients. Methods: The customized focal epilepsy gene panel (21 genes) was based on multiplex polymerase chain reaction (PCR) and sequenced by Illumina MiSeq platform. Results: Eleven variants (1.85%) were considered as pathogenic or likely pathogenic, including seven novel mutations. There were three SCN1A (p.Leu890Pro, p.Arg1636Ter, and p.Met1714Val), three PRRT2 (two p.Arg217Profs*8 and p.Leu298Pro), two CHRNA4 (p.Ser284Leu, p.Ile321Asn), one DEPDC5 (p.Val516Ter), one PCDH19 (p.Asp233Asn), and one SLC2A1 (p.Ser414Ter) variants. Additionally, 16 other rare variants were classified as unknown significance due to inconsistent phenotype or lack of segregation data. Conclusion: Currently known focal epilepsy genes only explained a very small subset of focal epilepsy patients. This indicates that the underlying genetic architecture of focal epilepsies is very heterogeneous and more novel genes are likely to be discovered. Our study highlights the usefulness, challenges and limitations of using the multi-gene panel as a diagnostic test in routine clinical practice in patients with focal epilepsy.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  9. Ling, S., Noramirah, R., Abidatul, A.A., Nurfarhanah, N.M.J., Noor-Azira, A.M., Jambari, N.N., et al.
    Food Research, 2018;2(3):240-246.
    MyJurnal
    Foodborne illness is a global burden that impacts a country politically, economically and
    socio-economically. The severity of the burden can be unmeasurable as foodborne illness
    is often an underestimated problem. In order to enlighten the burden, appropriate food
    safety control measures should be taken. This study aimed to optimize a multiplex
    Polymerase Chain Reaction (mPCR) detection method to identify foodborne pathogens
    simultaneously. Six foodborne pathogens namely, Salmonella spp., Escherichia coli O157,
    Vibrio parahaemolyticus, Vibrio cholerae, Listeria monocytogenes and Campylobacter
    spp., were targeted in the mPCR detection method. Each mPCR parameter was tested and
    the outcome was analysed to obtain a successful mPCR protocol to detect the targeted
    foodborne pathogens. The amplified PCR products showed that the optimized mPCR
    protocol will be a potential rapid diagnostic tool in foodborne pathogen detection.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  10. Kim JH, Chong CK, Sinniah M, Sinnadurai J, Song HO, Park H
    J Clin Virol, 2015 Apr;65:11-9.
    PMID: 25766980 DOI: 10.1016/j.jcv.2015.01.018
    BACKGROUND: Dengue is a mosquito-borne disease that causes a public health problem in tropical and subtropical countries. Current immunological diagnostics based on IgM and/or nonstructural protein 1 (NS1) antigen are limited for acute dengue infection due to low sensitivity and accuracy.
    OBJECTIVES: This study aimed to develop a one-step multiplex real-time RT-PCR assay showing higher sensitivity and accuracy than previous approaches.
    STUDY DESIGN: Serotype-specific primers and probes were designed through the multiple alignment of NS1 gene. The linearity and limit of detection (LOD) of the assay were determined. The assay was clinically validated with an evaluation panel that was immunologically tested by WHO and Malaysian specimens.
    RESULTS: The LOD of the assay was 3.0 log10 RNA copies for DENV-1, 2.0 for DENV-3, and 1.0 for DENV-2 and DENV-4. The assay showed 95.2% sensitivity (20/21) in an evaluation panel, whereas NS1 antigen- and anti-dengue IgM-based immunological assays exhibited 0% and 23.8-47.6% sensitivities, respectively. The assay showed 100% sensitivity both in NS1 antigen- and anti-dengue IgM-positive Malaysian specimens (26/26). The assay provided the information of viral loads and serotype with discrimination of heterotypic mixed infection.
    CONCLUSIONS: The assay could be clinically applied to early dengue diagnosis, especially during the first 5 days of illness and approximately 14 days after infection showing an anti-dengue IgM-positive response.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  11. Koh SF, Tay ST, Sermswan R, Wongratanacheewin S, Chua KH, Puthucheary SD
    J Microbiol Methods, 2012 Sep;90(3):305-8.
    PMID: 22705921 DOI: 10.1016/j.mimet.2012.06.002
    We have developed a multiplex PCR assay for rapid identification and differentiation of cultures for Burkholderia pseudomallei, Burkholderia thailandensis, Burkholderia mallei and Burkholderia cepacia complex. The assay is valuable for use in clinical and veterinary laboratories, and in a deployable laboratory during outbreaks.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction*
  12. Strobl C, Churchill Cihlar J, Lagacé R, Wootton S, Roth C, Huber N, et al.
    Forensic Sci Int Genet, 2019 09;42:244-251.
    PMID: 31382159 DOI: 10.1016/j.fsigen.2019.07.013
    The emergence of Massively Parallel Sequencing technologies enabled the analysis of full mitochondrial (mt)DNA sequences from forensically relevant samples that have, so far, only been typed in the control region or its hypervariable segments. In this study, we evaluated the performance of a commercially available multiplex-PCR-based assay, the Precision ID mtDNA Whole Genome Panel (Thermo Fisher Scientific), for the amplification and sequencing of the entire mitochondrial genome (mitogenome) from even degraded forensic specimens. For this purpose, more than 500 samples from 24 different populations were selected to cover the vast majority of established superhaplogroups. These are known to harbor different signature sequence motifs corresponding to their phylogenetic background that could have an effect on primer binding and, thus, could limit a broad application of this molecular genetic tool. The selected samples derived from various forensically relevant tissue sources and were DNA extracted using different methods. We evaluated sequence concordance and heteroplasmy detection and compared the findings to conventional Sanger sequencing as well as an orthogonal MPS platform. We discuss advantages and limitations of this approach with respect to forensic genetic workflow and analytical requirements.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction*
  13. Ahmed SA, Raabe CA, Cheah HL, Hoe CH, Rozhdestvensky TS, Tang TH
    Am J Trop Med Hyg, 2019 06;100(6):1328-1334.
    PMID: 30963989 DOI: 10.4269/ajtmh.18-0525
    The diarrheal disease "cholera" is caused by Vibrio cholerae, and is primarily confined to endemic regions, mostly in Africa and Asia. It is punctuated by outbreaks and creates severe challenges to public health. The disease-causing strains are most-often members of serogroups O1 and O139. PCR-based methods allow rapid diagnosis of these pathogens, including the identification of their biotypes. However, this necessitates the selection of specific target sequences to differentiate even the closely related biotypes of V. cholerae. Oligonucleotides for selective amplification of small RNA (sRNA) genes that are specific to these V. cholerae subtypes were designed. The resulting multiplex PCR assay was validated using V. cholerae cultures (i.e., 19 V. cholerae and 22 non-V. cholerae isolates) and spiked stool samples. The validation using V. cholerae cultures and spiked stool suspensions revealed detection limits of 10-100 pg DNA per reaction and 1.5 cells/mL suspension, respectively. The multiplex PCR assay that targets sRNA genes for amplification enables the sensitive and specific detection, as well as the differentiation of V. cholerae-O1 classical, O1 El Tor, and O139 biotypes. Most importantly, the assay enables fast and cheaper diagnosis compared with classic culture-based methods.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction*
  14. Maran S, Faten SA, Lim SE, Lai KS, Ibrahim WPW, Ankathil R, et al.
    Biomed Res Int, 2020;2020:6945730.
    PMID: 33062692 DOI: 10.1155/2020/6945730
    Background: The 22q11.2 deletion syndrome (22q11.2DS) is the most common form of deletion disorder in humans. Low copy repeats flanking the 22q11.2 region confers a substrate for nonallelic homologous recombination (NAHR) events leading to rearrangements which have been reported to be associated with highly variable and expansive phenotypes. The 22q11.2DS is reported as the most common genetic cause of congenital heart defects (CHDs).

    Methods: A total of 42 patients with congenital heart defects, as confirmed by echocardiography, were recruited. Genetic molecular analysis using a fluorescence in situ hybridization (FISH) technique was conducted as part of routine 22q11.2DS screening, followed by multiplex ligation-dependent probe amplification (MLPA), which serves as a confirmatory test.

    Results: Two of the 42 CHD cases (4.76%) indicated the presence of 22q11.2DS, and interestingly, both cases have conotruncal heart defects. In terms of concordance of techniques used, MLPA is superior since it can detect deletions within the 22q11.2 locus and outside of the typically deleted region (TDR) as well as duplications.

    Conclusion: The incidence of 22q11.2DS among patients with CHD in the east coast of Malaysia is 0.047. MLPA is a scalable and affordable alternative molecular diagnostic method in the screening of 22q11.2DS and can be routinely applied for the diagnosis of deletion syndromes.

    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  15. Shakrin NN, Balasubramaniam SD, Yusof HA, Mastuki MF, Masri SN, Taib NM, et al.
    Trop Biomed, 2013 Jun;30(2):338-44.
    PMID: 23959499 MyJurnal
    Determination of Streptococcus pneumoniae serotypes is essential for epidemiological surveillance. Therefore accurate, reliable and cost effective serotyping method is crucial. In this study, we determined the serotypes of 41 pneumococcal isolates recovered from human anterior nares by multiplex Polymerase Chain Reaction (PCR) utilizing published primers. The data was then compared with conventional serology using latex agglutination (LA) and the Quellung reaction. Based on the PCR-approach, 8 different serogroups/serotypes were detected with one isolate classified as non-typeable (cpsA-negative). In reference to the serology-based data, the results were in agreement except for one isolate. For the latter isolate, the LA and Quellung tests failed to show a reaction but the PCR-approach and sequencing identified the isolate as serogroup 15B/C. Based on this experimental setting, we found that the PCR-approach for pneumococcal serotypes determination is reliable to serve as the alternative for determining the pneumococcal serotyping.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  16. Sultana S, Hossain MAM, Naquiah NNA, Ali ME
    PMID: 30028648 DOI: 10.1080/19440049.2018.1500719
    Gelatin is widely used in pharmaceuticals as a protective coating, such as soft and hard capsule shells. However, the animal source of gelatin is a sensitive issue because certain gelatins such as porcine and bovine gelatins are not welcome in Halal, Kosher and Hindus' consumer goods. Recently, we have documented DNA barcoding and multiplex PCR platforms for discriminating porcine, bovine and fish gelatins in various fish and confectionary products; but those assays were not self-authenticating and also not tested in highly refined pharmaceutical products. To address this knowledge gap, here we report a self-authenticating multiplex PCR-restriction fragment length polymorphism (RFLP) assay to identify animal sources of various gelatin in pharmaceutical capsules. Three different restriction enzymes, BsaAI, Hpy188I and BcoDI were used to yield distinctive RFLP patterns for gelatin-based bovine (26, 94 bp), fish (97, 198 bp) and porcine (17, 70 bp) DNA in control experiments. The specificity was cross-tested against 16 non-target species and the optimised assay was used to screen gelatin sources in 30 halal-branded pharmaceuticals capsule shells. Bovine and porcine DNA was found in 27 and 3 of the 30 different capsules products. The assay was suitable for detecting 0.1 to 0.01 ng total DNA extracted from pure and mixed gelatins. The study might be useful to authenticate and monitor halal, kosher, vegetarian and Hindu compliant pharmaceuticals, foods and cosmetics.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  17. Goodwin W, Alimat S
    Electrophoresis, 2017 04;38(7):1007-1015.
    PMID: 28008628 DOI: 10.1002/elps.201600383
    The SNPforID consortium identified a panel of 52 SNPs for forensic analysis that has been used by several laboratories worldwide. The original analysis of the 52 SNPs was based on a single multiplex reaction followed by two single-base-extension (SBE) reactions each of which was analyzed using capillary electrophoresis. The SBE assays were designed for high throughput genetic analyzers and were difficult to use on the single capillary ABI PRISM 310 Genetic Analyzer and the latest generation 3500 Genetic Analyzer, as sensitivity on the 310 was low and separation of products on the 3500 with POP-7™ was poor. We have modified the original assay and split it into four multiplex reactions, each followed by an SBE assay. These multiplex assays were analyzed using polymer POP-4™ on ABI 310 PRISM® and polymers POP-4™, POP-6™ and POP-7™ on the 3500 Genetic Analyzer. The assays were sensitive and reproducible with input DNA as low as 60 pg using both the ABI 310 and 3500. In addition, we found that POP-6™ was most effective with the 3500, based on the parameters that we assessed, achieving better separation of the small SBE products; this conflicted with the recommended use of POP-7™ by the instrument manufacturer. To support the use of the SNP panel in casework in Malaysia we have created an allele frequency database from 325 individuals, representing the major population groups within Malaysia. Population and forensic parameters were estimated for all populations and its efficacy evaluated using 51 forensic samples from challenging casework.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  18. Hussain H, Chong NF
    Biomed Res Int, 2016;2016:8041532.
    PMID: 27995143
    The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. This combined method allows up to 6 base pairs to be mutated at a time and requires a total of 40-45 PCR cycles. A total of eight mutagenesis experiments were successfully carried out, with each experiment mutating between two to six base pairs. Up to four adjacent codons were changed in a single experiment. This method is especially useful for codon optimization, where doublet or triplet rare codons can be changed using a single mutagenic primer set, in a single experiment.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  19. Hossain MA, Ali ME, Abd Hamid SB, Asing, Mustafa S, Mohd Desa MN, et al.
    J Agric Food Chem, 2016 Aug 17;64(32):6343-54.
    PMID: 27501408 DOI: 10.1021/acs.jafc.6b02224
    Beef, buffalo, and pork adulteration in the food chain is an emerging and sensitive issue. Current molecular techniques to authenticate these species depend on polymerase chain reaction (PCR) assays involving long and single targets which break down under natural decomposition and/or processing treatments. This novel multiplex polymerase chain reaction-restriction fragment length polymorphism assay targeted two different gene sites for each of the bovine, buffalo, and porcine materials. This authentication ensured better security, first through a complementation approach because it is highly unlikely that both sites will be missing under compromised states, and second through molecular fingerprints. Mitochondrial cytochrome b and ND5 genes were targeted, and all targets (73, 90, 106, 120, 138, and 146 bp) were stable under extreme boiling and autoclaving treatments. Target specificity and authenticity were ensured through cross-amplification reaction and restriction digestion of PCR products with AluI, EciI, FatI, and CviKI-1 enzymes. A survey of Malaysian frankfurter products revealed rampant substitution of beef with buffalo but purity in porcine materials.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
  20. Nik Zuraina NMN, Mohamad S, Hasan H, Goni MD, Suraiya S
    Pathog Glob Health, 2023 Feb;117(1):63-75.
    PMID: 35331083 DOI: 10.1080/20477724.2022.2028378
    Respiratory tract infections (RTIs), including pneumonia and pulmonary tuberculosis, are among the leading causes of death worldwide. The use of accurate diagnostic tests is crucial to initiate proper treatment and therapy to reduce the mortality rates for RTIs. A PCR assay for simultaneous detection of six respiratory bacteria: Haemophilus influenzae, Klebsiella pneumoniae, Mycobacterium tuberculosis, Pseudomonas aeruginosa, Staphylococcus aureus and Streptococcus pneumoniae, was developed in our lab. The current study aimed to evaluate the performance of this assay along with the retrospective surveillance of respiratory pathogens at a teaching hospital in Kelantan, Malaysia. Leftover sputa (n = 200) from clinical laboratories were collected and undergone DNA template preparation for PCR analysis. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of the PCR assay were determined in comparison with the gold standard sputum culture. Overall, the accuracy performance of this assay was 94.67% (95% CI: 90.87% to 97.21%) with sensitivity, specificity, PPV and NPV of 100%, 91.67%, 87.1% and 100%, respectively. Based on the organisms detected from sputa, K. pneumoniae ranked as the top isolate (n = 48), followed by P. aeruginosa (n = 13) and H. influenzae (n = 10). Surveillance among the patients showed that the associations of bacterial positive with gender and means of acquisition were found significant (p values = 0.049 and 0.001, respectively). Besides the promising performance of this ready-to-use molecular-based assay for the rapid detection of selected bacteria pathogens, this study also highlighted significant spread of K. pneumoniae RTIs in the community.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction*
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