METHODS: A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed using different protocols. In total, 712 sequencing libraries were investigated for viral sequence contamination.
RESULTS: Among sequences showing similarity to viruses, 493 were significantly associated with the use of laboratory components. Each of these viral sequences had sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components.
CONCLUSIONS: We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover, we show that detection can be problematic due to stochastic appearance and limited non-template controls. Although the exact origin of these viral sequences requires further research, our results support laboratory-component-linked viral sequence contamination of both biological and synthetic origin.
METHODS: The qLAMP assay was optimized targeting the HPV-16 E7 gene. The analytical sensitivity and specificity of the assay were determined using HPV-18 (ATCC® 45152D™), HPV-35 (ATCC® 40330™), HPV-43 (ATCC® 40338™) and HPV-56 (ATCC® 40549™) viral strains and oral bacteria. HPV-16 standard curve was constructed for determination of HPV-16 viral load. The diagnostic performance of the assay was evaluated from 63 OSCC patients comprising 63 tissue, 13 saliva and 49 blood samples, in comparison with p16 immunohistochemistry (IHC), in-house PCR and nested PCR assays.
RESULTS: The detection limit of developed LAMP and PCR assays was 4.68 × 101 and 4.68 × 103 copies/μl, respectively. qLAMP assay enabled detection of positive results as early as 23 min at 67 °C. This assay can detect HPV-16 positivity in 23 % (3/13) saliva and 4.8 % (3/63) tissue samples with the viral load ranging from 4.68 × 101 to 4.68 × 104 copies/μl. HPV-16 positivity was not detected in all the blood samples. The sensitivity and specificity of qLAMP were 100 % in comparison with that of p16 IHC and nested PCR.
CONCLUSION: This study reports for the first time on the use of qLAMP assay for detection of HPV-16 in OSCC in both tissue and saliva as the sample matrix which holds promise in improving the diagnostic application owing to its rapidity, simplicity, high sensitivity and specificity.
MATERIALS AND METHODS: Total RNA was extracted from three formalin-fixed paraffin-embedded (FFPE) samples each of normal cervix, HPV-infected low-grade squamous intraepithelial lesion (LSIL), high-grade SIL (HSIL) and squamous cell carcinoma (SCC). Transcriptomic profiling by microarrays was conducted followed by downstream Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses.
RESULTS: We examined the difference in GOs enriched for each transition stage from normal cervix to LSIL, HSIL, and SCC, and found 307 genes to be differentially expressed. In the transition from normal cervix to LSIL, the extracellular matrix (ECM) genes were significantly downregulated. The MHC class II genes were significantly upregulated in the LSIL to HSIL transition. In the final transition from HSIL to SCC, the immunoglobulin heavy locus genes were significantly upregulated and the ECM pathway was implicated.
CONCLUSION: Deregulation of the immune-related genes including MHC II and immunoglobulin heavy chain genes were involved in the transitions from LSIL to HSIL and SCC, suggesting immune escape from host anti-tumour response. The extracellular matrix plays an important role during the early and late stages of cervical carcinogenesis.
MATERIALS AND METHODS: To examine this hypothesis, in the present study, the associations between CYP3A5 variants (rs776746 and rs1419745) and response to carbamazepine and valproic acid monotherapy in Malaysian epileptic patients were evaluated.
RESULTS: A total of 288 Malaysian epileptic patients were recruited and further reviewed, of whom 63 patients were on carbamazepine monotherapy, and 85 patients were on valproic acid monotherapy. There was no patient with drug hypersensitivity syndrome within the population. Subjects were genotyped by using Sequenom MassARRAY platform. This study found a significant association of CYP3A5 rs776746 with the carbamazepine treatment response in total patients (p = 0.026) and Malay ethnic subgroup (p = 0.006). In addition, a marginal significant association of CYP3A5 rs1419745 with carbamazepine treatment response was reported in the Malays. Similarly, CYP3A5 rs776746 was associated with valproic acid response in total patients (p = 0.037) and Malays (marginal p = 0.05).
CONCLUSION: Our findings suggest that CYP3A5 polymorphisms affect carbamazepine and valproic acid response in Malaysian epileptic patients.
METHODS: Haemoglobin variants were identified by HbA1c analysis in 93 of 3522 samples sent to our laboratory in a period of 1 month. Haemoglobin analysis identified HbE trait in 81 of 93 samples. To determine the influence of HbE trait on HbA1c analysis by Variant II Tubo 2.0, boronate affinity high performance liquid chromatography (HPLC) method (Primus PDQ) was used as the comparison method. Two stage linear regression analysis, Bland Altman plot and Deming regression analysis were performed to analyse whether the presence of HbE trait produced a statistically significant difference in the results. The total allowable error for HbA1c by the Royal Australasian College of Pathologists (RCPA) external quality assurance is 5%. Hence clinically significant difference is more than 5% at the medical decision level of 6% and 9%.
RESULTS: Statistically and clinically significant higher results were observed in Variant II Turbo 2.0 due to the presence of HbE trait. A positive bias of ∼10% was observed at the medical decision levels.
CONCLUSION: Laboratories should be cautious when evaluating HbA1c results in the presence of haemoglobin variants.