Displaying publications 21 - 40 of 85 in total

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  1. Al-Herz W, Essa S
    Front Immunol, 2019;10:1231.
    PMID: 31191561 DOI: 10.3389/fimmu.2019.01231
    Objective: To present the frequency and spectrum of viral infections in primary immunodeficient children. Methods: The data was obtained from the Kuwait National Primary Immunodeficiency Disorders (PIDs) Registry during the period of 2004-2018. Results: A total of 274 PID children were registered in KNPIDR during the study period with predominance of immunodeficiencies affecting cellular and humoral immunity, followed by combined immunodeficiencies with associated syndromic features and diseases of immune dysregulation. Overall infectious complications affected 82.4% of the patients, and viral infections affected 31.7% of the registered patients. Forty-five patients (16.4%) developed viral infections caused by at least 2 organisms, among those 20 patients were affected by three or more viral infections. There was a statistically significant association between viral infections and PID category. However, there was no statistically significant association between viral infections and gender or the patients' onset age. There was a total of 170 viral infections during the study period and the causes of these infections were predominated by CMV (22.2%), adenovirus (11.7%), EBV (11.1%), and enteroviruses (7.4%). CMV and parainfluenza infections were more common in the group of immunodeficiencies affecting cellular and humoral immunity while EBV and human papilloma virus (HPV) were more common in the immune dysregulation group and combined immunodeficiencies with associated syndromic features, respectively. The most common presentation was viremia (28.8%) followed by pneumonia (28.2%) and skin infections (17.6%). The most common causes of viremia were CMV followed by adenovirus and EBV, while the most common organisms causing pneumonia were CMV followed by rhinovirus and parainfluenza. There were 80 deaths among the registered patients, 10% were caused by viral infections. Conclusions: Viral infections are common in PIDs and result into a wide-range of clinical manifestations causing significant morbidity and mortality.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  2. Zarina Mohd Zawawi, Tengku Rogayah Tengku Abdul Rashid, Amir Hussien Adiee, Murni Maya Sari, Ravindran Thayan
    MyJurnal
    Introduction: Dengue virus (DENV), Zika virus (ZIKV) and Chikungunya virus (CHIKV) are Arboviruses that are transmitted by the same vector, Aedes aegypti. Dengue has become a global problem since the Second World War and is common in more than 110 countries. In Malaysia, dengue is a major disease burden as total economic costs to the country as a result of dengue is close to RM1.05 billion in 2010 and estimated to rise to 1.3 billion by 2020. Apart from Dengue, Zika and Chikungunya are the other important mosquito borne diseases in Malaysia. The aim of this study was to develop a multiplex real-time assay for simultaneous detection of DENV, ZIKV and CHIKV in clinical specimens. Methods: The published singleplex protocols were used with key modifications to implement a triplex assay. A one-step multiplex real-time RT-PCR assay was developed that can simultaneously detect RNA of DENV, ZIKV and CHIKV with good performance for a routine diagnostic use. The assay was evaluated for inter- and intra-reproducibility by mean CT value. The diagnostic sensitivity was tested with 135 archived samples which had been defined positive or negative by routine singleplex assays. Whole blood, plasma and urines were used in this study. Results: Intra- and inter-reproducibility and sensitivity varied from 0.10% to 4.73% and from 0.45% to 5.98% for each virus respectively. The specificity of detection was 100%. The multiplex real-time RT-PCR assay showed concordance with test results performed by routine singleplex assays. No cross reaction was observed for any of the clinical samples. Conclusion: The development of a rapid, sensitive and specific molecular assay for DENV, ZIKV and CHIKV infections will produce a greater diagnostic capacity in our laboratory. This multiplex approach is cost effective and robust with the concurrent detection of 3 viruses of public health concern.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  3. Hosuru Subramanya S, Bairy I, Nayak N, Padukone S, Sathian B, Gokhale S
    PLoS One, 2019;14(2):e0212042.
    PMID: 30779752 DOI: 10.1371/journal.pone.0212042
    A worldwide increase in the gastrointestinal colonization by extended-spectrum β-lactamase (ESBL)-producing bacteria has been observed. Their prevalence amongst Healthy People Living with HIV (HPLWH) has not been investigated adequately. The aim of this study was to determine and compare the rates of and risk factors for intestinal carriage and acquisition of extended-spectrum β-lactamase producing Enterobacteriaceae (ESBL-E) and carbapenemase-producing Enterobacteriaceae (CPE) among healthy people living with HIV (HPLWH) and healthy HIV negative population in the community. A cross-sectional study was conducted. Rectal swabs from HPLWH (n = 119) and HIV negative individuals (n = 357) from the community were screened for ESBL and CPE. Phenotypically confirmed ESBL-E strains were genotyped by multiplex PCR. The risk factors associated with ESBL-E colonization were analyzed by a multivariable conditional logistic regression analysis. Specimen from 357 healthy volunteers (213 female and 144 male) and 119 HPLWH (82 female and 37 male) with a median age of 30 [IQR 11-50] years were included in the study. ESBL colonization were found in 45 (37.82% [CI 29.09, 47.16]) and 246 (68.91% [CI 63.93, 73.49]), HPLWH and healthy HIV negative participants respectively. HPLWH had lower ESBL carriage rate (odds ratio 0.274 [CI 0.178, 0.423]) compared to healthy HIV negative subject's (p<0.01). In this study, no carbapenemase-producing bacteria were isolated.CTX-M-15 type was the most predominant genotype in both groups. Livestock contact and over-the-counter medications were significantly associated with a higher ESBL-E carriage rate among healthy subjects. This is the first study in Nepal that has demonstrated a high rate of gut colonization by ESBL-E in the community, predominantly of blaCTX-M-15 genotype. This study divulges the low fecal carriage rate of ESBL producing bacteria in HPLWH group compared to healthy individuals in western Nepal. The factors responsible for this inverse relationship of HIV status and gut colonization by ESBL-E are unidentified and require further large-scale study.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  4. Brandon-Mong GJ, Ketzis JK, Choy JS, Boonroumkaew P, Tooba M, Sawangjaroen N, et al.
    Trop Biomed, 2018 Dec 01;35(4):1131-1139.
    PMID: 33601860
    Trichuris trichiura, the whipworm of humans, is one of the most prevalent soiltransmitted helminths (STH) reported worldwide. According to a recent study, out of 289 STH studies in Southeast Asia, only three studies used molecular methods. Hence, the genetic assemblages of Trichuris in Southeast Asia are poorly understood. In this study, we used partial mitochondrial DNA (cytochrome c oxidase subunit 1 or COI) sequences for analysis. Trichuris grouped in a same clade with different hosts indicate the potential of cross infection between hosts. Based on COI, the adult Trichuris isolated from a Malaysian patient was most closely related to Trichuris isolated from Papio anubis (olive baboons) from the USA. The Trichuris isolated from the dog from Malaysia was genetically similar to a Trichuris species isolated from Macaca silenus (lion-tailed macaque) from Czech Republic. Both the human and dog isolated Trichuris grouped in clades with different hosts indicating the potential of cross infection between hosts. Specific PCR primers based on the partial COI of T. trichiura isolated from African green monkey and T. serrata were designed and successfully amplified using multiplex PCR of the pooled DNA samples. Our results suggest a complex parasite-host relationship, and support the theory of cross infection of Trichuris between humans and non-human primates as suggested in previous publications.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  5. 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*
  6. Zarizal S, Yeo CC, Faizal GM, Chew CH, Zakaria ZA, Jamil Al-Obaidi MM, et al.
    Trop Med Int Health, 2018 08;23(8):905-913.
    PMID: 29873865 DOI: 10.1111/tmi.13090
    BACKGROUND: This study aimed to profile the antimicrobial susceptibility and presence of resistance and virulence genes of methicillin-susceptible Staphylococcus aureus (MSSA) and MRSA nasal carriage, by means of genotypic analyses, in students of a tertiary institution in the state of Terengganu, east coast of Malaysia.

    METHODS: A total of 370 agricultural biotechnology students from Universiti Sultan Zainal Abidin in Besut, Terengganu, were enrolled in this study. Antimicrobial susceptibility profiles were evaluated by standard methods. PCR detection of resistance and virulence genes was performed on S. aureus that were methicillin-resistant, macrolide-lincosamide-streptogramin B (MLSB )-positive phenotype and/or positive for the leukocidin (pvl) gene followed by staphylococcal cassette chromosome mec (SCCmec), staphylococcal protein A (spa) and accessory gene regulator (agr) typing.

    RESULTS: One hundred and nineteen of 370 students carried S. aureus (32%); 18 of the isolates were MRSA (15%). Erythromycin resistance was detected in 20% (24/119) of which 15% (18/119) were MRSA and 5% (6/119) MSSA. Among the 24 erythromycin-resistant isolates, D-test was positive in 29% (7/24) displaying inducible MLSB , whereas the remaining 71% (17/24) showed constitutive MLSB phenotypes. Nine (7.6%) of 119 isolates were pvl positive: 44% MRSA (4/9) and 56% MSSA (5/9). Staphylococcal surface protein sasX gene was present in 92% of MRSA and 8% of MSSA isolates. The majority of MRSA isolates were agr type I (15/18; 83%). Five spa types identified with spa t037 were predominant, followed by spa types (t304 and t8696) as newly reported Malaysian MRSA in a community setting.

    CONCLUSION: The presence of MRSA with SCCmec of hospital-associated features and globally recognised spa types in community setting is worrisome. Furthermore, the presence of MLSB strains among multidrug-resistant (MDR) S. aureus with sasX as well as pvl-positive isolates highlights the potential risk of a community setting in facilitating the dissemination of both virulence and resistance determinants.

    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  7. 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
  8. Zam Zureena Mohd Rani, Nor Azian Abdul Murad, Saberi Saimun, Sri Noraima Othman, Rahman Jamal, Sue-Mian Then, et al.
    Neurology Asia, 2018;23(2):137-144.
    MyJurnal
    Background: The HLA-B*15:02 polymorphism in epileptic patients is known to be associated with carbamazepine-induced Stevens-Johnson syndrome (SJS). The prevalence of HLA-B*15:02 polymorphism seemed to be ethnic-specific with a higher frequency of HLA-B*15:02 in Asian compared to the Europeans. This study was performed to determine the frequency of the HLA-B*15:02 polymorphism in epileptic patients at the Chancellor Tuanku Muhriz Hospital-UKM Medical Centre (HCTM-UKMMC) using high resolution melting-real time PCR (HRM-QPCR) method.
    Methods: We performed a fast and effective in-house high resolution melting-real time polymerase chain reaction method and compared it with the conventional multiplex-PCR method. The specificity and sensitivity of each test were also determined using DNA from saliva.
    Results: Using the conventional multiplexPCR approach for screening, 25 out of 64 (39.1%) epileptic patients were positive for HLA-B*15:02. However, using the HRM-QPCR technique, 24/64 (37.5%) of the patients were positive. The one patient who tested positive by the multiplex-PCR but negative using the HRM-QPCR turned out to be negative by DNA sequencing. The HRM-QPCR and DNA sequencing showed 100% sensitivity and specificity. The multiplex-PCR showed 100% sensitivity and 98.4% specificity compared to both HRM-QPCR and DNA sequencing. The HRM-QPCR is also more cost-effective (multiplex-PCR ($25.15 USD/test).
    Conclusion: Our result suggested that multiplex PCR, HRM-QPCR and Sanger sequencing can be used for detection of HLA-B*15:02. However, a qualitative method such as multiplex PCR should be confirmed with other quantitative methods such as HRM-QPCR and Sanger sequencing.
    Keywords: Epilepsy, carbamazepine-induced Steven Johnson syndrome, multiplex-polymerase chain reaction, high resolution melting-real time polymerase chain reaction (HRM-QPCR), DNA sequencing
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  9. Tang, J-Y-H., Farhana Sakinah, M.R., Nakaguchi, Y., Nishibuchi, M., Chai, L-C., New, C.Y., et al.
    Food Research, 2018;2(5):447-452.
    MyJurnal
    This goal of this study was to investigate the presence of Vibrio cholerae in street food,
    namely satar and otak-otak, using Loop-Mediated Isothermal Amplification (LAMP),
    multiplex Polymerase Chain Reaction (mPCR) and conventional plating on Thiosulphate
    Citrate Bile-Salt Sucrose (TCBS) agar methods. A total of 78 satar and 35 otak-otak were
    purchased from different districts of Terengganu (Besut, Setiu, Kuala Terengganu and
    Kemaman). V. cholerae was found in satar with LAMP (10.3%), mPCR (10.3%) and
    plating (0%). No V. cholerae was found in otak-otak using the three methods. This might
    be due to V. cholerae able to survive in satar after grilling due to its thickness which may
    contribute to undercooking. This study concluded that low presence of V. cholerae in satar
    and otak-otak can be detected by molecular methods but not the conventional plating
    method. LAMP assay is a useful tool for rapid detection of pathogens in food due to its
    simplicity, highly sensitive and visual interpretation capability. Though the prevalence of
    V. cholerae was low in the samples, proper handling of this food will help in reducing the
    risk of acquiring infection from V. cholerae in contaminated samples.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  10. 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
  11. Jacinta Santhanam, Mohd Hanif Jainlabdin, Ang LC, Tzar Mohd Nizam
    Sains Malaysiana, 2018;47:489-498.
    Invasive fungal infections (IFIs) have risen dramatically in recent years among high risk immunocompromised patients.
    Rapid detection of fungal pathogens is crucial to timely and accurate antifungal therapy. Two multiplex polymerase
    chain reaction (PCR) assays were developed to detect major fungal species that cause invasive infections and identify
    resistant species. Genus specific primers for Candida, Aspergillus, Fusarium and species specific primers for Candida
    glabrata, Candida krusei and Aspergillus terreus which are known to be clinically resistant species, were designed from
    the internal transcribed spacer (ITS) regions of ribosomal ribonucleic acid (rRNA) gene complex. Both assays were
    performed simultaneously to promote rapid detection of fungal isolates based on distinct amplicon sizes. Inclusion of the
    universal fungal primers ITS 1 and ITS 4 in the genus specific assay produced a second amplicon for each isolate which
    served to confirm the detection of a fungal target. The limit of detection for the genus specific assay was 1 nanogram
    (ng) deoxyribonucleic acid (DNA) for Aspergillus fumigatus and Candida albicans, 0.1 ng DNA for Fusarium solani, while
    the species-specific assay detected 0.1 ng DNA of A. terreus and 10 picogram (pg) DNA of C. krusei and C. glabrata. The
    multiplex PCR assays, apart from universal detection of any fungal target, are able to detect clinically important fungi
    and differentiate resistant species rapidly and accurately, which can contribute to timely implementation of effective
    antifungal regime.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  12. Ahamad MNU, Ali ME, Hossain MAM, Asing A, Sultana S, Jahurul MHA
    PMID: 28748739 DOI: 10.1080/19440049.2017.1359752
    Rabbit meat is receiving increasing attention because it contains a high level of proteins with relatively little fat. On the other hand, squirrel meat is served in upper-class meals in certain countries, so is sold at higher prices. The other side of the coin is rat meat, which has family ties with rabbit and squirrel but poses substantial threats to public health because it is a potential carrier of several zoonotic organisms. Recently, rat meat was mislabelled and sold as lamb after chemical modification. Thus, the chances of rabbit and squirrel meat substitution by rat meat cannot be ruled out. For the first time, a multiplex PCR assay was developed in Malaysia for the discriminatory identification of rat, rabbit and squirrel in the food chain. Rabbit (123 bp), rat (108 bp) and squirrel (243 bp) targets were amplified from ATP6 and cytb genes, along with a eukaryotic internal control (141bp). The products were sequenced and cross-tested against 22 species. A total of 81 reference samples and 72 meatball specimens were screened to validate the assay. Analyte stability was evaluated through boiling, autoclaving and micro-oven cooking. The tested lower limits of detection were 0.01 ng DNA for pure meat and 0.1% for meatballs.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction*
  13. Kanniappan P, Ahmed SA, Rajasekaram G, Marimuthu C, Ch'ng ES, Lee LP, et al.
    J Cell Mol Med, 2017 10;21(10):2276-2283.
    PMID: 28756649 DOI: 10.1111/jcmm.13148
    Technological advances in RNA biology greatly improved transcriptome profiling during the last two decades. Besides the discovery of many small RNAs (sRNA) that are involved in the physiological and pathophysiological regulation of various cellular circuits, it becomes evident that the corresponding RNA genes might also serve as potential biomarkers to monitor the progression of disease and treatment. sRNA gene candidate npcTB_6715 was previously identified via experimental RNomic (unpublished data), and we report its application as potential biomarker for the detection of Mycobacterium tuberculosis (MTB) in patient samples. For proof of principle, we developed a multiplex PCR assay and report its validation with 500 clinical cultures, positive for Mycobacteria. The analysis revealed 98.9% sensitivity, 96.1% specificity, positive and negative predictive values of 98.6% and 96.8%, respectively. These results underscore the diagnostic value of the sRNA gene as diagnostic marker for the specific detection of MTB in clinical samples. Its successful application and the general ease of PCR-based detection compared to standard bacterial culture techniques might be the first step towards 'point-of-care' diagnostics of Mycobacteria. To the best of our knowledge, this is the first time for the design of diagnostic applications based on sRNA genes, in Mycobacteria.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods
  14. Tan JL, Nordin S, Besari AM
    Malays J Med Sci, 2017 Oct;24(5):119-123.
    PMID: 29386980 MyJurnal DOI: 10.21315/mjms2017.24.5.14
    Introduction: Tuberculosis is the second leading cause of death under the category of infectious diseases, after the human immunodeficiency virus (HIV). Tuberculous meningitis (TBM) constitutes about 5% of all extrapulmonary disease worldwide. This report describes a case of Tuberculous meningitis with rare presentation in a 28-year-old woman, who was treated based on a collection of her social background, clinical findings and Multiplex PCR of tuberculosis.

    Case presentation: A 28-year-old Malay woman with no significant medical history presented to HUSM with one month history of on and off fever, two weeks history of generalised limbs weakness and one week history of dysphagia. She was reported to have experienced visual hallucination and significant weight loss. Her laboratory result is significant for leukocytosis, elevated ESR and hypernatremia. Non-enhanced and contrast CT scan of the brain showed severe bilateral frontal cerebral atrophy. Cerebral spinal fluid (CSF) for multiplex PCR for Mycobacterium tuberculosis complex was positive. She was promptly started on anti-TB regime combined with dexamethasone. Subsequent follow-up showed significant improvement.

    Conclusion: This is a rare clinical manifestation of Tuberculous meningitis that demonstrates the importance of recognising and initiating the treatment early to reduce disabilities and improve clinical outcome.

    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  15. Kuan CS, Yew SM, Hooi PS, Lee LM, Ng KP
    Malays J Med Sci, 2017 Oct;24(5):33-43.
    PMID: 29386970 MyJurnal DOI: 10.21315/mjms2017.24.5.4
    Introduction: Acute respiratory tract infections (ARTIs) are a major cause of morbidity and mortality in paediatric patients. Therefore, early detection of the viral aetiologies of ARTIs is essential for patient management and infection control. In this study, we evaluated the performance of a new multiplex polymerase chain reaction (PCR) assay (xTAG Respiratory Viral Panel [RVP] Fast v2) in the detection of respiratory viruses by comparing it with that of viral culture and direct immunofluorescence (IF) staining.

    Methods: Nasopharyngeal swab and aspirate samples were collected prospectively from 199 patients who presented with ARTIs at the University Malaya Medical Centre (UMMC) in Kuala Lumpur, Malaysia during a 10-month period. The PCR assay was conducted in parallel with conventional culture and direct IF staining methods.

    Results: The positive rate of the xTAG RVP Fast v2 assay (78.4%) in detecting respiratory viruses was higher than that of the viral isolation (7.5%) and direct IF (23.1%) methods. Using the xTAG RVP Fast v2 assay, human enterovirus/human rhinovirus (HEV/HRV) was the most frequently detected (46.2%). The xTAG RVP Fast v2 assay revealed mixed infection caused by two or three respiratory viruses in 40 specimens, and these were undetected by the viral isolation and direct IF methods.

    Conclusion: The xTAG RVP Fast v2 assay was superior to conventional methods in the identification of common respiratory viruses, with higher sensitivity and shorter turnaround times for laboratory results.

    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  16. Rahman NA, Sharudin A, Diah S, Muharram SH
    Microb Pathog, 2017 Sep;110:352-358.
    PMID: 28711510 DOI: 10.1016/j.micpath.2017.07.021
    INTRODUCTION: Pneumococcal infections have caused morbidity and mortality globally. Streptococcus pneumoniae (pneumococci) are commensal bacteria that colonize the nasopharynx, asymptomatically. From there, pneumococci can spread in the lungs causing pneumonia and disseminate in the bloodstream causing bacteremia (sepsis) and reach the brain leading to meningitis. Endothelial cells are one of the most important components of the blood-brain barrier that separates the blood from the brain and plays the first protective role against pneumococcal entry. Thus this study aimed to investigate on the ability of non-meningitis pneumococcal clinical strains to adhere and invade a brain endothelium model.

    METHODS: Two pneumococcal Brunei clinical strains were serotyped by multiplex PCR method using oligonucleotide sequences derived from Centers for Disease Control and Prevention. A validated immortalised mouse brain endothelial cell line (bEnd.3) was used as a brain endothelium model for the study of the pneumococcal breach of the blood-brain barrier using an adherence and invasion assay.

    RESULTS: Both of the pneumococcal clinical strains were found to be serotype 19F, a common circulating serotype in Southeast Asia and globally and possess the ability to adhere and invade the brain endothelial cells.

    CONCLUSION: In addition, this is the first report on the serotype identification of pneumococci in Brunei Darussalam and their application on a brain endothelium model. Further studies are required to understand the virulence capabilities of the clinical strains.

    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  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. Khazani NA, Noor NZ, Yean Yean C, Hasan H, Suraiya S, Mohamad S
    J Trop Med, 2017;2017:7210849.
    PMID: 28386286 DOI: 10.1155/2017/7210849
    Klebsiella pneumoniae and Haemophilus influenzae are two common pathogens associated with respiratory tract infections. The identification of these pathogens using conventional molecular diagnostic tests requires trained personnel, cold-chain transportation, and storage-dependance, which does not render them user-friendly. The aim of this study was to develop a thermostabilized, cold-chain-free, one-step multiplex PCR for simultaneous detection of K. pneumoniae and H. influenzae. The multiplex PCR assay was designed to amplify the php gene of K. pneumoniae (202 bp) and p6 gene of H. influenzae (582 bp). In addition, the specific primer to amplify glm gene of Helicobacter pylori (105 bp) was included as an internal amplification control. Subsequently, the designed primers and all PCR reagents were thermostabilized by lyophilization. The stability of the thermostabilized PCR was evaluated using the Q(10) method. The sensitivity and specificity of performances for thermostabilized PCR were evaluated using 127 clinical isolates and were found to be 100% sensitive and specific. The thermostabilized PCR mix was found to be stable for 30 days and the Q10 accelerated stability was found to be 3.02 months. A cold-chain-free, PCR assay for easy, rapid, and simultaneous detection of K. pneumoniae and H. influenzae was successfully developed in this study.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  19. Thung TY, Radu S, Mahyudin NA, Rukayadi Y, Zakaria Z, Mazlan N, et al.
    Front Microbiol, 2017;8:2697.
    PMID: 29379488 DOI: 10.3389/fmicb.2017.02697
    The aim of the present study was to investigate the prevalence of Salmonella spp., Salmonella Enteritidis and Salmonella Typhimurium in retail beef from different retail markets of Selangor area, as well as, to assess their pathogenic potential and antimicrobial resistance. A total of 240 retail beef meat samples (chuck = 60; rib = 60; round = 60; sirloin = 60) were randomly collected. The multiplex polymerase chain reaction (mPCR) in combination with the most probable number (MPN) method was employed to detect Salmonella spp., S. Enteritidis and S. Typhimurium in the meat samples. The prevalence of Salmonella spp., S. Enteritidis and S. Typhimurium in 240 beef meat samples were 7.50, 1.25, and 0.83%, respectively. The microbial loads of total Salmonella was found in the range of <3 to 15 MPN/g. Eight different serovars of Salmonella were identified among the 23 isolates, and S. Agona was the predominant serovar (26.09%). Interestingly, all the Salmonella isolates were resistant to penicillin, erythromycin and vancomycin, but the sensitivity was observed for tetracycline, gentamicin and amoxicillin/clavulanic acid. All 23 isolates were resistant to at least three antibiotics. Two S. Typhimurium isolates (8.70%) exhibited the highest multiple antibiotic resistance (MAR) index value of 0.56 which shown resistance to nine antibiotics. PCR analysis of virulence genes showed that all Salmonella isolates (100%) were positive for the invA gene. Meanwhile, pefA was only identified in S. Enteritidis and S. Typhimurium. The findings in this study indicate that retail beef products tested were widely contaminated with multi-drug resistant (MDR) Salmonella and various virulence genes are present among the isolated Salmonella serovars.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  20. Fadilah N, Hanafiah A, Razlan H, Wong ZQ, Mohamed Rose I, Rahman MM
    Br J Biomed Sci, 2016 Oct;73(4):180-187.
    PMID: 27922429
    BACKGROUND: No gold standard has yet been established for the diagnosis of H. pylori infection. A multiplex polymerase chain reaction (mPCR) was developed in this study for rapid, sensitive and specific detection of H. pylori from gastric biopsies.

    METHODS: H. pylori infections were determined by in-house rapid urease test (iRUT), culture, histology and multiplex PCR.

    RESULTS: A total of 140 (60.9%) from 230 patients were positive for H. pylori infection. H. pylori were detected in 9.6% (22/230), 17% (39/230), 12.6% (29/230) and 60% (138/230) of biopsy specimens by culture, iRUT, histology and mPCR, respectively. mPCR identified H. pylori infection in 100% of biopsies with positive histology and culture. All biopsies with positive iRUT yielded positive PCR except two cases. mPCR also detected H. pylori in additional 116, 101 and 109 biopsies that were negative by culture, iRUT and histology, respectively. Positive samples by mPCR showed lower average in H. pylori density, activity and inflammation scores. The Indians showed the highest prevalence of H. pylori infection compared to the Chinese and the Malays. In addition, Chinese patients with older age were significantly infected compared to other ethnicities.

    CONCLUSION: PCR was able to detect the highest numbers of positive cases although the lowest average scores were recorded in the activity, inflammatory and H. pylori density.

    Matched MeSH terms: Multiplex Polymerase Chain Reaction/methods*
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