Displaying publications 1 - 20 of 306 in total

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  1. Fulltext Detection of Strongyloides stercoralis infection among cancer patients in a major hospital in Kelantan, Malaysia
    Zueter AM, Mohamed Z, Abdullah AD, Mohamad N, Arifin N, Othman N, et al.
    Singapore Med J, 2014 Jul;55(7):367-71.
    PMID: 25091885
    INTRODUCTION: Strongyloidiasis is one of the most commonly neglected but clinically important parasitic infections worldwide, especially among immunocompromised patients. Evidence of infection among immunocompromised patients in Malaysia is, however, lacking. In this study, microscopy, real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assays (ELISAs) were used to detect Strongyloides stercoralis (S. stercoralis) infection among cancer patients in a Malaysian hospital.

    METHODS: A total of 192 stool and serum samples were collected from cancer patients who were receiving chemotherapy with or without steroid treatment at a hospital in northeastern Malaysia. Stool samples were examined for S. stercoralis using parasitological methods and real-time PCR. Serology by ELISA was performed to detect parasite-specific immunoglobulin G (IgG), IgG4 and immunoglobulin E (IgE) antibodies. For comparison, IgG4- and IgG-ELISAs were also performed on the sera of 150 healthy individuals from the same area.

    RESULTS: Of the 192 samples examined, 1 (0.5%) sample was positive for S. stercoralis by microscopy, 3 (1.6%) by real-time PCR, 8 (4.2%) by IgG-ELISA, 6 (3.1%) by IgG4-ELISA, and none was positive by IgE-ELISA. In comparison, healthy blood donors had significantly lower prevalence of parasite-specific IgG (2.67%, p < 0.05) and IgG4 (2.67%, p < 0.05) responses.

    CONCLUSION: This study showed that laboratory testing may be considered as a diagnostic investigation for S. stercoralis among immunocompromised cancer patients.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  2. Administration of Bacillus subtilis strains in the rearing water enhances the water quality, growth performance, immune response, and resistance against Vibrio harveyi infection in juvenile white shrimp, Litopenaeus vannamei
    Zokaeifar H, Babaei N, Saad CR, Kamarudin MS, Sijam K, Balcazar JL
    Fish Shellfish Immunol, 2014 Jan;36(1):68-74.
    PMID: 24161773 DOI: 10.1016/j.fsi.2013.10.007
    In this study, vegetative cell suspensions of two Bacillus subtilis strains, L10 and G1 in equal proportions, was administered at two different doses 10(5) (BM5) and 10(8) (BM8) CFU ml(-1) in the rearing water of shrimp (Litopenaeus vannamei) for eight weeks. Both probiotic groups showed a significant reduction of ammonia, nitrite and nitrate ions under in vitro and in vivo conditions. In comparison to untreated control group, final weight, weight gain, specific growth rate (SGR), food conversion ratio (FCR) and digestive enzymatic activity were significantly greater in the BM5 and BM8 groups. Significant differences for survival were recorded in the BM8 group as compared to the control. Eight weeks after the start of experiment, shrimp were challenged with Vibrio harveyi. Statistical analysis revealed significant differences in shrimp survival between probiotic and control groups. Cumulative mortality of the control group was 80%, whereas cumulative mortality of the shrimp that had been given probiotics was 36.7% with MB8 and 50% with MB5. Subsequently, real-time RT-PCR was employed to determine the mRNA levels of prophenoloxidase (proPO), peroxinectin (PE), lipopolysaccharide- and β-1,3-glucan- binding protein (LGBP) and serine protein (SP). The expression of all immune-related genes studied was only significantly up-regulated in the BM5 group compared to the BM8 and control groups. These results suggest that administration of B. subtilis strains in the rearing water confers beneficial effects for shrimp aquaculture, considering water quality, growth performance, digestive enzymatic activity, immune response and disease resistance.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  3. Fulltext Effects of Bacillus subtilis on the growth performance, digestive enzymes, immune gene expression and disease resistance of white shrimp, Litopenaeus vannamei
    Zokaeifar H, Balcázar JL, Saad CR, Kamarudin MS, Sijam K, Arshad A, et al.
    Fish Shellfish Immunol, 2012 Oct;33(4):683-9.
    PMID: 22659618 DOI: 10.1016/j.fsi.2012.05.027
    We studied the effect of two probiotic Bacillus subtilis strains on the growth performance, digestive enzyme activity, immune gene expression and disease resistance of juvenile white shrimp (Litopenaeus vannamei). A mixture of two probiotic strains, L10 and G1 in equal proportions, was administered at two different doses 10(5) (BM5) and 10(8) (BM8) CFU g(-1) feed to shrimp for eight weeks. In comparison to untreated control group, final weight, weight gain and digestive enzyme activity were significantly greater in shrimp fed BM5 and BM8 diets. Significant differences for specific growth rate (SGR) and survival were recorded in shrimp fed BM8 diet as compared with the control; however, no significant differences were recorded for food conversion ratio (FCR) among all the experimental groups. Eight weeks after the start of the feeding period, shrimp were challenged with Vibrio harveyi. Statistical analysis revealed significant differences in shrimp survival between probiotic and control groups. Cumulative mortality of the control group was 63.3%, whereas cumulative mortality of the shrimp that had been given probiotics was 20.0% with BM8 and 33.3% with BM5. Subsequently, real-time PCR was employed to determine the mRNA levels of prophenoloxidase (proPO), peroxinectin (PE), lipopolysaccharide- and β-1,3-glucan-binding protein (LGBP) and serine protein (SP). The expression of all immune-related genes studied was significantly up-regulated (P 
    Matched MeSH terms: Real-Time Polymerase Chain Reaction/veterinary
  4. Fulltext DACH1, a zona glomerulosa selective gene in the human adrenal, activates transforming growth factor-β signaling and suppresses aldosterone secretion
    Zhou J, Shaikh LH, Neogi SG, McFarlane I, Zhao W, Figg N, et al.
    Hypertension, 2015 May;65(5):1103-10.
    PMID: 25776071 DOI: 10.1161/HYP.0000000000000025
    Common somatic mutations in CACNAID and ATP1A1 may define a subgroup of smaller, zona glomerulosa (ZG)-like aldosterone-producing adenomas. We have therefore sought signature ZG genes, which may provide insight into the frequency and pathogenesis of ZG-like aldosterone-producing adenomas. Twenty-one pairs of zona fasciculata and ZG and 14 paired aldosterone-producing adenomas from 14 patients with Conn's syndrome and 7 patients with pheochromocytoma were assayed by the Affymetrix Human Genome U133 Plus 2.0 Array. Validation by quantitative real-time polymerase chain reaction was performed on genes >10-fold upregulated in ZG (compared with zona fasciculata) and >10-fold upregulated in aldosterone-producing adenomas (compared with ZG). DACH1, a gene associated with tumor progression, was further analyzed. The role of DACH1 on steroidogenesis, transforming growth factor-β, and Wnt signaling activity was assessed in the human adrenocortical cell line, H295R. Immunohistochemistry confirmed selective expression of DACH1 in human ZG. Silencing of DACH1 in H295R cells increased CYP11B2 mRNA levels and aldosterone production, whereas overexpression of DACH1 decreased aldosterone production. Overexpression of DACH1 in H295R cells activated the transforming growth factor-β and canonical Wnt signaling pathways but inhibited the noncanonical Wnt signaling pathway. Stimulation of primary human adrenal cells with angiotensin II decreased DACH1 mRNA expression. Interestingly, there was little overlap between our top ZG genes and those in rodent ZG. In conclusion, (1) the transcriptome profile of human ZG differs from rodent ZG, (2) DACH1 inhibits aldosterone secretion in human adrenals, and (3) transforming growth factor-β signaling pathway is activated in DACH1 overexpressed cells and may mediate inhibition of aldosterone secretion in human adrenals.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  5. Optimization and Implementation of the Virus Extraction Method for Hepatitis E Virus Detection from Raw Pork Liver
    Zhao MY, Li D
    Food Environ Virol, 2021 03;13(1):74-83.
    PMID: 33449335 DOI: 10.1007/s12560-020-09452-y
    Hepatitis E virus (HEV) has been frequently detected from pork liver and liver products, which can usually cause self-limiting diseases in healthy adults, yet may result in fatality in immunosuppressed groups. Nevertheless, there is so far no standardized method for HEV detection available from pork liver and/or liver products. The present study aimed to optimize the virus extraction method of HEV from raw pork liver, which is often consumed in Asia undercooked to avoid a grainy texture. By comparing different sample preparation protocols and by applying the selected protocol to 60 samples collected from Singapore retail markets, we demonstrated that homogenization of 0.25 g raw pork liver with FastPrep™ Lysing Matrix Y containing yttria-stabilized zircondium oxide beads in 2 ml tubes and with harsh mechanical force at 6 ms-1, 40 s/cycle, for 5 cycles with 300 s pause time after each cycle is promising in both releasing the potentially intracellular viruses and resulting in satisfactory virus recovery rates (> 1%). A high prevalence (52%) of HEV genome was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) from the 60 samples collected from Singapore retail markets imported from Indonesia, Australia and Malaysia. However, RNase treatment decreased the HEV prevalence to 33.3%, and all of the 20 positive samples were with high RT-qPCR Ct values above 35, suggesting that the positive RT-qPCR signals maybe largely due to the inactive viruses and/or exposed HEV RNA traces in raw pork liver products. Therefore, conscious care should be taken when interpreting molecular detection results of viruses from food samples to be correlated with public health risks.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  6. Fulltext Flavone enhances dengue virus type-2 (NGC strain) infectivity and replication in vero cells
    Zandi K, Lani R, Wong PF, Teoh BT, Sam SS, Johari J, et al.
    Molecules, 2012;17(3):2437-45.
    PMID: 22374315 DOI: 10.3390/molecules17032437
    This study investigates the effects of 2-phenyl-1-benzopyran-4-one (flavone) on DENV-2 infectivity in Vero cells. Virus adsorption and attachment and intracellular virus replication were investigated using a foci forming unit assay (FFUA) and quantitative RT-PCR, respectively. Addition of flavone (100 μg/mL) significantly increased the number of DENV-2 foci by 35.66% ± 1.52 and 49.66% ± 2.51 when added during and after virus adsorption to the Vero cells, respectively. The average foci size after 4 days of infection increased by 33% ± 2.11 and 89% ± 2.13. The DENV-2 specific RNA copy number in the flavone-treated infected cells increased by 6.41- and 23.1-fold when compared to the mock-treated infected cells. Flavone (100 μg/mL) did not promote or inhibit Vero cell proliferation. The CC₅₀ value of flavone against Vero cells was 446 µg/mL. These results suggest that flavone might enhance dengue virus replication by acting antagonistically towards flavonoids known to inhibit dengue virus replication.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  7. Quantitative analysis of fetal DNA in maternal plasma in gestational diabetes mellitus, iron deficiency anemia and gestational hypertension pregnancies
    Zamanpoor M, Rosli R, Yazid MN, Husain Z, Nordin N, Thilakavathy K
    J Matern Fetal Neonatal Med, 2013 Jul;26(10):960-6.
    PMID: 23339569 DOI: 10.3109/14767058.2013.766710
    OBJECTIVE: To quantify circulating fetal DNA (fDNA) levels in the second and third trimesters of normal healthy pregnant individuals and pregnant women with the following clinical conditions: gestational diabetes mellitus (GDM), iron deficiency anemia and gestational hypertension (GHT).
    METHODS: The SRY gene located on the Y chromosome was used as a unique fetal marker. The fDNA was extracted from maternal plasma and the SRY gene concentrations were measured by quantitative real-time polymerase chain reaction (PCR) amplification using TaqMan dual labeled probe system.
    RESULTS: No significant differences were observed in the mean fDNA concentration between normal and GDM pregnancy samples (p > 0.05) and also between normal and anemic pregnancy samples (p > 0.05) in both trimesters, but significant differences were observed between the third trimester normal and GHT pregnancy samples (p = 0.001). GDM and iron deficiency anemia do not affect the levels of fDNA in maternal plasma while GHT significantly elevates the levels of fDNA in maternal plasma.
    CONCLUSIONS: Increased amount of circulating fDNA in maternal plasma could be used for early identification of adverse pregnancies. GDM and anemia do not affect the levels of fDNA in maternal plasma while GHT significantly elevates the levels of fDNA in maternal plasma. Hence, the elevated fDNA values could be used as a potential screening marker in pregnancies complicated with GHT but not with GDM and iron deficiency anemia.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction*
  8. Fulltext HLA-B*15:02 screening in epileptic patients using a high resolution melting-real time PCR (HRM-QPCR) method
    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 (time-consuming when compared to the 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: Real-Time Polymerase Chain Reaction
  9. Fulltext Nucleic Acid-Based Diagnostic Tests for the Detection SARS-CoV-2: An Update
    Yu CY, Chan KG, Yean CY, Ang GY
    Diagnostics (Basel), 2021 Jan 01;11(1).
    PMID: 33401392 DOI: 10.3390/diagnostics11010053
    The coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) began as a cluster of pneumonia cases in Wuhan, China before spreading to over 200 countries and territories on six continents in less than six months. Despite rigorous global containment and quarantine efforts to limit the transmission of the virus, COVID-19 cases and deaths have continued to increase, leaving devastating impacts on the lives of many with far-reaching effects on the global society, economy and healthcare system. With over 43 million cases and 1.1 million deaths recorded worldwide, accurate and rapid diagnosis continues to be a cornerstone of pandemic control. In this review, we aim to present an objective overview of the latest nucleic acid-based diagnostic tests for the detection of SARS-CoV-2 that have been authorized by the Food and Drug Administration (FDA) under emergency use authorization (EUA) as of 31 October 2020. We systematically summarize and compare the principles, technologies, protocols and performance characteristics of amplification- and sequencing-based tests that have become alternatives to the CDC 2019-nCoV Real-Time RT-PCR Diagnostic Panel. We highlight the notable features of the tests including authorized settings, along with the advantages and disadvantages of the tests. We conclude with a brief discussion on the current challenges and future perspectives of COVID-19 diagnostics.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  10. Fulltext Potentiality of a triple microRNA classifier: miR-193a-3p, miR-23a and miR-338-5p for early detection of colorectal cancer
    Yong FL, Law CW, Wang CW
    BMC Cancer, 2013 Jun 08;13:280.
    PMID: 23758639 DOI: 10.1186/1471-2407-13-280
    BACKGROUND: MicroRNAs (miRNAs) are short, non-coding RNA molecules that act as regulators of gene expression. Circulating blood miRNAs offer great potential as cancer biomarkers. The objective of this study was to correlate the differential expression of miRNAs in tissue and blood in the identification of biomarkers for early detection of colorectal cancer (CRC).

    METHODS: The study was divided into two phases: (I) Marker discovery by miRNA microarray using paired cancer tissues (n = 30) and blood samples (CRC, n = 42; control, n = 18). (II) Marker validation by stem-loop reverse transcription real time PCR using an independent set of paired cancer tissues (n = 30) and blood samples (CRC, n = 70; control, n = 32). Correlation analysis was determined by Pearson's test. Logistic regression and receiver operating characteristics curve analyses were applied to obtain diagnostic utility of the miRNAs.

    RESULTS: Seven miRNAs (miR-150, miR-193a-3p, miR-23a, miR-23b, miR-338-5p, miR-342-3p and miR-483-3p) have been found to be differentially expressed in both tissue and blood samples. Significant positive correlations were observed in the tissue and blood levels of miR-193a-3p, miR-23a and miR-338-5p. Moreover, increased expressions of these miRNAs were detected in the more advanced stages. MiR-193a-3p, miR-23a and miR-338-5p were demonstrated as a classifier for CRC detection, yielding a receiver operating characteristic curve area of 0.887 (80.0% sensitivity, 84.4% specificity and 83.3% accuracy).

    CONCLUSION: Dysregulations in circulating blood miRNAs are reflective of those in colorectal tissues. The triple miRNA classifier of miR-193a-3p, miR-23a and miR-338-5p appears to be a potential blood biomarker for early detection of CRC.

    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  11. Increased Expression of Phosphatidylinositol 3-Kinase p110α and Gene Amplification of PIK3CA in Nasopharyngeal Carcinoma
    Yip WK, He PY, Abdullah MA, Yusoff S, Seow HF
    Pathol Oncol Res, 2016 Apr;22(2):413-9.
    PMID: 26581613 DOI: 10.1007/s12253-015-0007-8
    Molecular alterations in PIK3CA oncogene that encodes the p110α catalytic subunit of phosphatidylinositol 3-kinase (PI3K p110α) are commonly found in human cancers. In this study, we examined the expression of PI3K p110α and PIK3CA gene amplification in 74 nasopharyngeal carcinoma (NPC) cases. Immunohistochemical staining demonstrated overexpression of PI3K p110α protein in 44.6% (33/74) of NPCs and 4.8% (2/42) of the adjacent normal nasopharyngeal mucosa. Copy number of PIK3CA gene was successfully analyzed in 51 of the total NPC cases and 19 non-malignant nasopharynx tissues by quantitative real-time PCR. Using mean + 2(standard deviation) of copy numbers in the non-malignant nasopharynx tissues as a cutoff value, PIK3CA copy number gain was found in 10 of 51 (19.6%) NPC cases. High PI3K p110α expression level was correlated with increased PIK3CA copy number (Spearman's rho =0.324, P = 0.02). PI3K p110α expression and PIK3CA copy number did not associate with Akt phosphorylation, and patient and tumor variables. This study suggests that PI3K p110α overexpression, which is attributed, at least in part, to PIK3CA gene amplification, may contribute to NPC pathogenesis. However, these molecular aberrations may not be responsible for activation of Akt signaling in NPC.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  12. Activation of phosphatidylinositol 3-kinase/Akt signaling by EGF downregulates membranous E-cadherin and β-catenin and enhances invasion in nasopharyngeal carcinoma cells
    Yip WK, Seow HF
    Cancer Lett, 2012 May 28;318(2):162-72.
    PMID: 22182447 DOI: 10.1016/j.canlet.2011.12.018
    Dysregulation of E-cadherin and β-catenin function in cell-cell adhesion is common in nasopharyngeal carcinoma (NPC) and correlates with metastatic disease. In this study, we examined the role of EGF-activated phosphatidylinositol 3-kinase (PI3K)-Akt signaling in E-cadherin and β-catenin regulation. We found that reduced membranous E-cadherin and β-catenin expression was positively correlated with Akt phosphorylation in NPC tissues. EGF treatment disrupted cell-cell adhesion and resulted in mesenchymal morphological features in NPC cell lines (TW01, TW04, and TW06). Western blot analysis showed that the E-cadherin protein level was partially reduced in TW04 cells only and the β-catenin levels were not considerably affected upon EGF treatment. In contrast, quantitative real-time RT-PCR showed that the E-cadherin, but not β-catenin, mRNA levels were markedly reduced by EGF in all cell lines. Immunofluorescent staining revealed that E-cadherin and β-catenin appeared to be markedly reduced on the cell surface and more localized in the cytoplasm. Inhibition of PI3K by LY294002 did not abolish the EGF-induced downregulation of E-cadherin protein or mRNA in TW04 cells but moderately increased the β-catenin protein level in TW01 cells and mRNA level in TW06 cells. However, LY294002 substantially restored or increased cell surface E-cadherin and β-catenin in all EGF-treated cell lines, in concordance with the inhibition of cell morphological changes. Moreover, LY294002 significantly blocked EGF-driven cell invasion, correlating with the elevation of membranous E-cadherin and β-catenin levels. In conclusion, EGF-induced epithelial-to-mesenchymal transition may not be only dependent on downregulation of E-cadherin protein/mRNA but also on mislocalization of E-cadherin and β-catenin. The mechanisms involved may be related, at least in part, to the PI3K-Akt pathway.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  13. Epstein-Barr virus-encoded latent membrane protein-1 upregulates 14-3-3σ and Reprimo to confer G(2)/M phase cell cycle arrest
    Yeo KS, Mohidin TB, Ng CC
    C. R. Biol., 2012 Dec;335(12):713-21.
    PMID: 23312294 DOI: 10.1016/j.crvi.2012.11.002
    Epstein-Barr virus (EBV) is a ubiquitous tumor-causing virus which infects more than 90% of the world population asymptomatically. Recent studies suggest that LMP-1, -2A and -2B cooperate in the tumorigenesis of EBV-associated epithelial cancers such as nasopharygeal carcinoma, oral and gastric cancer. In this study, LMPs were expressed in the HEK293T cell line to reveal their oncogenic mechanism via investigation on their involvement in the regulation of the cell cycle and genes that are involved. LMPs were expressed in HEK293T in single and co-expression manner. The transcription of cell cycle arrest genes were examined via real-time PCR. Cell cycle progression was examined via flow cytometry. 14-3-3σ and Reprimo were upregulated in all LMP-1 expressing cells. Moreover, cell cycle arrest at G(2)/M progression was detected in all LMP-1 expressing cells. Therefore, we conclude that LMP-1 may induce cell cycle arrest at G(2)/M progression via upregulation of 14-3-3σ and Reprimo.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  14. In vitro characterization of chicken bone marrow-derived dendritic cells following infection with very virulent infectious bursal disease virus
    Yasmin AR, Yeap SK, Tan SW, Hair-Bejo M, Fakurazi S, Kaiser P, et al.
    Avian Pathol, 2015;44(6):452-62.
    PMID: 26305169 DOI: 10.1080/03079457.2015.1084997
    Infectious bursal disease is caused by infectious bursal disease virus (IBDV), an immunosuppressive virus that targets immune cells such as B cells and macrophages. However, the involvement of dendritic cells (DCs) during IBDV infection is not well understood. In this study the in vitro effects of live and inactivated very virulent IBDV (vvIBDV) UPM0081 on bone marrow-derived DCs (BM-DC) were characterized and compared with BM-DC treated with lipopolysaccharide (LPS). Morphologically, BM-DC treated with LPS and vvIBDV showed stellate shape when compared to immature BM-DC. In addition, LPS-treated and both live and inactivated vvIBDV-infected BM-DC expressed high levels of double positive CD86 and major histocompatibility complex class II antigens (>20%). vvIBDV-infected BM-DC showed significantly higher numbers of apoptotic cells compared to LPS. Replication of vvIBDV was detected in the infected BM-DC as evidenced by the increased expression of VP3 and VP4 IBDV antigens based on flow cytometry, real-time polymerase chain reaction and immunofluorescence tests. Levels of different immune-related genes such as interleukin-1β (IL-1β), CXCLi2 (IL-8), IL-18, interferon gamma (IFN-γ, IL-12α, CCR7 and Toll-like receptor-3 (TLR3) were measured after LPS and vvIBDV treatments. However, marked differences were noticed in the onset and intensity of the gene expression between these two treatment groups. LPS was far more potent than live and inactivated vvIBDV in inducing the expression of IL-1β, IL-18 and CCR7 while expression of Th1-like cytokines, IFN-γ and IL-12α were significantly increased in the live vvIBDV treatment group. Meanwhile, the expression of TLR3 was increased in live vvIBDV-infected BM-DC as compared to control. Inactivated vvIBDV-treated BM-DC failed to stimulate IFN-γ, IL-12α and TLR3 expressions. This study suggested that BM-DC may serve as another target cells during IBDV infection which require further confirmation via in vivo studies.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  15. Fulltext Effect of Ganoderma lucidum polysaccharides on the growth of Bifidobacterium spp. as assessed using Real-time PCR
    Yamin, S., Shuhaimi, M., Arbakariya, A., Khalilah, A. K., Anas, O., Yazid, A. M., et al.
    International Food Research Journal, 2012;19(3):1199-1205.
    MyJurnal
    The use of component from Ganoderma lucidum as prebiotic source is interesting as the G. lucidum itself was known for more than a decade in the traditional Chinese medicine. In this work, Ganoderma lucidum crude polysaccharides (GLCP) and Polysaccharide-fraction number 2 (PF-2) were used as carbon sources in the fermentation with Bifidobacterium sp. The results showed the potential of prebiotic effect of the G. lucidum extract in batch-culture fermentation based on increment in the growth of bacteria used (0.4 – 1.5 log10 CFU/mL) after 18h fermentation. Fermentation was further done using faecal materials as bacterial inocula and bacterial growth changes were examined using real-time PCR. The results showed the ability of GLCP and PF-2 to support the growth of Bifidobacterium genus with 0.3 and 0.7 log10 cells/ml increased, respectively. Interestingly, Lactobacillus which is known as beneficial bacterial genus also showed growth increment with 0.7 and 1 log10 cells/ml increased. The competition for carbon sources thus inhibits the growth of potentially harmful genus, Salmonella (0.3 and 0.5 log10 cells/ml) in comparison to the control.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  16. Male and female NOD mice differentially express peroxisome proliferator-activated receptors and pathogenic cytokines
    Yaacob NS, Goh KS, Norazmi MN
    Exp. Toxicol. Pathol., 2012 Jan;64(1-2):127-31.
    PMID: 20674317 DOI: 10.1016/j.etp.2010.07.005
    The peroxisome proliferator-activated receptors (PPARs) have been implicated in regulating the immune response. We determined the relative changes in the transcriptional expression of PPAR isoforms (α, γ1 and γ2) and cytokines involved in the pathogenesis of type 1 diabetes (T1D) in the immune cells of 5 weeks, 10 weeks and diabetic male non-obese diabetic (NOD) mice compared to those of female NOD mice from our previous studies, "normalized" against their respective non-obese diabetic resistant (NOR) mice controls. Overall PPARα was significantly more elevated in the macrophages of female NOD mice of all age groups whereas PPARγ, particularly the PPARγ2 isoform was more depressed in the macrophages and CD4(+) lymphocytes of female NOD mice compared to their male counterparts. The pro-inflammatory cytokines, IL-1 and TNFα, as well as the Th1 cytokines, IL-2 and IFNγ were more elevated in female NOD mice whereas the Th2 cytokine, IL-4, was more depressed in these mice compared to their male counterparts. These findings suggest that the preponderance of T1D in female NOD mice may be influenced by the more pronounced changes in the expression of PPAR isoforms and pathogenic cytokines compared to those in male NOD mice.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  17. Fulltext Molecular epidemiology of enterovirus 71 infection in the central region of Taiwan from 2002 to 2012
    Wu WH, Kuo TC, Lin YT, Huang SW, Liu HF, Wang J, et al.
    PLoS One, 2013;8(12):e83711.
    PMID: 24391812 DOI: 10.1371/journal.pone.0083711
    Enterovirus 71 (EV71), a causative agent of hand, foot, and mouth disease can be classified into three genotypes and many subtypes. The objectives of this study were to conduct a molecular epidemiological study of EV71 in the central region of Taiwan from 2002-2012 and to test the hypothesis that whether the alternative appearance of different EV71 subtypes in Taiwan is due to transmission from neighboring countries or from re-emergence of pre-existing local strains. We selected 174 EV71 isolates and used reverse transcription-polymerase chain reaction to amplify their VP1 region for DNA sequencing. Phylogenetic analyses were conducted using Neighbor-Joining, Maximum Likelihood and Bayesian methods. We found that the major subtypes of EV71 in Taiwan were B4 for 2002 epidemic, C4 for 2004-2005 epidemic, B5 for 2008-2009 epidemic, C4 for 2010 epidemic and B5 for 2011-2012 epidemic. Phylogenetic analysis demonstrated that the 2002 and 2008 epidemics were associated with EV71 from Malaysia and Singapore; while both 2010 and 2011-2012 epidemics originated from different regions of mainland China including Shanghai, Henan, Xiamen and Gong-Dong. Furthermore, minor strains have been identified in each epidemic and some of them were correlated with the subsequent outbreaks. Therefore, the EV71 infection in Taiwan may originate from pre-existing minor strains or from other regions in Asia including mainland China. In addition, 101 EV71 isolates were selected for the detection of new recombinant strains using the nucleotide sequences spanning the VP1-2A-2B region. No new recombinant strain was found. Analysis of clinical manifestations showed that patients infected with C4 had significantly higher rates of pharyngeal vesicles or ulcers than patients infected with B5. This is the first study demonstrating that different EV 71 genotypes may have different clinical manifestations and the association of EV71 infections between Taiwan and mainland China.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  18. P16INK4a: a potential diagnostic adjunct for prediction of high-grade cervical lesions in liquid-based cytology: with HPV testing and histological correlation
    Wong YP, Abdul Raub SH, Mohd Dali AZ, Kassim F, Visvalingam V, Zakaria Z, et al.
    Malays J Pathol, 2016 Aug;38(2):93-101.
    PMID: 27568665
    Human papillomavirus (HPV) is a necessary cause of cervical cancer and its precursors. Increased expression of high-risk hrHPV viral oncogenes in abnormal cells might increase the expression of p16INK4a. We aimed to determine the role of p16INK4a in detecting hrHPV-transformed epithelial cells in liquid-based cervical cytology, and compared the results with hrHPV DNA testing by realtime polymerase chain reaction (RT-PCR). Fifty-seven cytological samples were tested for p16INK4a immunomarker and hrHPV DNA. Test performance of both tests was determined by comparing sensitivity, specificity and predictive values using available histological follow-up data as gold standard. Of 57 samples, 36 (63.2%) showed immunoreactivity for p16INK4a and 43 (75.4%) were hrHPV-infected. A fairly low concordance rate (k = 0.504) between p16INK4a immunolabelling and hrHPV DNA status was noted. For prediction of cervical intraepithelial neoplasia (CIN) II and worse lesions, p16INK4a had a sensitivity and specificity of 93.5% and 60%; whereas hrHPV DNA testing had a sensitivity and specificity of 100% and 20%. Dual testing by combining p16INK4a and hrHPV showed sensitivity and specificity of 100% and 33.3%. In conclusion, p16INK4a is useful in predicting severity of the cytological abnormalities. Although p16INK4a is more specific but less sensitive than hrHPV in detecting high-grade cervical lesions, a combination of both tests failed to demonstrate significant improvement in diagnostic sensitivity, specificity and predictive value. Larger-scale prospective studies are required to assess further whether this biomarker should be routinely used as primary screening tool independently or in combination with hrHPV testing to improve diagnostic accuracy in cervical cytology.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  19. Parallel ELISAs using crude soluble antigen and excretory-secretory antigen for improved serodiagnosis of amoebic liver abscess
    Wong WK, Foo PC, Olivos-Garcia A, Noordin R, Mohamed Z, Othman N, et al.
    Acta Trop, 2017 Aug;172:208-212.
    PMID: 28506795 DOI: 10.1016/j.actatropica.2017.05.017
    Crude soluble antigen (CSA) produced from Entamoeba histolytica trophozoite is conventionally used for serodiagnosis of invasive amoebiasis. However, high background seropositivities by CSA-assay in endemic areas complicate the interpretation of positive result in clinical settings. Instead, incorporating a second assay which indicates active or recent infection into the routine amoebic serology could possibly complement the limitations of CSA-assay. Hence, the present study aimed to evaluate the diagnostic efficacies of indirect ELISAs using CSA and excretory-secretory antigen (ESA) for serodiagnosis of amoebic liver abscess (ALA). Reference standard for diagnosis of ALA at Hospital Universiti Sains Malaysia is based on clinical presentation, radiological imaging and positive indirect haemagglutination assay (titer ≥256). Five groups of human serum samples collected from the hospital included Group I - ALA diagnosed by the reference standard and pus aspirate analysis using real-time PCR (n=10), Group II - ALA diagnosed by the reference standard only (n=41), Group III - healthy control (n=45), Group IV - other diseases control (n=51) and Group V - other infectious diseases control (n=31). For serodiagnosis of ALA serum samples (Group I and II), CSA-ELISA showed sensitivities of 100% for both groups, while ESA-ELISA showed sensitivities of 100% and 88%, respectively. For serodiagnosis of non-ALA serum samples (Group III, IV and V), CSA-ELISA showed specificities of 91%, 75% and 100%, respectively; while ESA-ELISA showed specificities of 96%, 98% and 100%, respectively. Indirect ELISAs using CSA and ESA have shown distinct strength for serodiagnosis of ALA, in terms of sensitivity and specificity, respectively. In conclusion, parallel analysis by both assays improved the overall efficacies of amoebic serology as compared to either single assay.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  20. Ectopic expression of a Musa acuminata root hair defective 3 (MaRHD3) in Arabidopsis enhances drought tolerance
    Wong GR, Mazumdar P, Lau SE, Harikrishna JA
    J Plant Physiol, 2018 Dec;231:219-233.
    PMID: 30292098 DOI: 10.1016/j.jplph.2018.09.018
    Genetic improvement is an important approach for crop improvement towards yield stability in stress-prone areas. Functional analysis of candidate stress response genes can provide key information to allow the selection and modification of improved crop varieties. In this study, the constitutive expression of a banana cDNA, MaRHD3 in Arabidopsis improved the ability of transgenic lines to adapt to drought conditions. Transgenic Arabidopsis plants expressing MaRHD3 had roots with enhanced branching and more root hairs when challenged with drought stress. The MaRHD3 plants had higher biomass accumulation, higher relative water content, higher chlorophyll content and an increase in activity of reactive oxygen species (ROS) scavenging enzymes; SOD, CAT, GR, POD and APX with reduced water loss rates compared to control plants. The analysis of oxidative damage indicated lower cell membrane damage in transgenic lines compared to control plants. These findings, together with data from higher expression of ABF-3 and higher ABA content of drought-stressed transgenic MaRHD3 expressing plants, support the involvement of the ABA signal pathway and ROS scavenging enzyme systems in MaRHD3 mediated drought tolerance.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction
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