Reverse transcription followed by real-time or quantitative polymerase chain reaction (RT-qPCR) is the gold standard for validation of results from transcriptomic profiling studies such as microarray and RNA sequencing. The current need for most studies, especially biomarker studies, is to evaluate the expression levels or fold changes of many transcripts in a large number of samples. With conventional low to medium throughput qPCR platforms, many qPCR plates would have to be run and a significant amount of RNA input per sample will be required to complete the experiments. This is particularly challenging when the size of study material (small biopsy, laser capture microdissected cells, biofluid, etc.), time, and resources are limited. A sensitive and high-throughput qPCR platform is therefore optimal for the evaluation of many transcripts in a large number of samples because the time needed to complete the entire experiment is shortened and the usage of lab consumables as well as RNA input per sample are low. Here, the methods of high-throughput RT-qPCR for the analysis of circulating microRNAs are described. Two distinctive qPCR chemistries (probe-based and intercalating dye-based) can be applied using the methods described here.
MicroRNAs regulate gene expression at the post-transcriptional level. Differential expression of miRNAs can potentially be used as biomarkers for early diagnosis and prediction for outcomes. Failure in validation of miRNA profiles is often caused by variations in experimental parameters. In this study, the performance of five extraction kits and three RT-qPCR systems were evaluated using BioMark high-throughput platform and the effects of different experimental parameters on circulating miRNA levels were determined. Differences in the performance of extraction kits as well as varying accuracy, sensitivity and reproducibility in qPCR systems were observed. Normalisation of RT-qPCR data to spike-in controls can reduce extraction bias. However, the extent of correlation for different qPCR systems varies in different assays. At different time points, there was no significant fold change in eight of the plasma miRNAs that we evaluated. Higher level of miRNAs was detected in plasma as compared to serum of the same cohort. In summary, we demonstrated that high-throughput RT-qPCR with pre-amplification step had increased sensitivity and can be achieved with accuracy and high reproducibility through stringent experimental controls. The information provided here is useful for planning biomarker validation studies involving circulating miRNAs.
This review aimed to evaluate the effectiveness of acupressure in promoting sleep quality among adults. Study findings included in the review showed that acupressure significantly improved sleep quality compared with the control group, but no superior effect of acupressure was found compared with sham acupressure.
Lipid nanoparticles are colloidal carrier systems that have extensively been investigated for controlled drug delivery, cosmetic and pharmaceutical applications. In this work, a cost effective stearic acid-oleic acid nanoparticles (SONs) with high loading of salicylic acid, was prepared by melt emulsification method combined with ultrasonication technique. The physicochemical properties, thermal analysis and encapsulation efficiency of SONs were studied. TEM micrographs revealed that incorporation of oleic acid induces the formation of elongated spherical particles. This observation is in agreement with particle size analysis which also showed that the mean particle size of SONs varied with the amount of OA in the mixture but with no effect on their zeta potential values. Differential scanning calorimetry analysis showed that the SONs prepared in this method have lower crystallinity as compared to pure stearic acid. Different amount of oleic acid incorporated gave different degree of perturbation to the crystalline matrix of SONs and hence resulted in lower degrees of crystallinity, thereby improving their encapsulation efficiencies. The optimized SON was further incorporated in cream and its in vitro release study showed a gradual release for 24 hours, denoting the incorporation of salicylic acid in solid matrix of SON and prolonging the in vitro release.
BACKGROUND: Human leukocyte antigen (HLA) antigens and genes have long been reported associated with systemic lupus erythematosus (SLE) susceptibility in many populations. With the advance in technologies such as genome-wide association studies, many newly discovered SLE-associated single-nucleotide polymorphisms (SNPs) have been reported in recent years. These include HLA-DRB1/HLA-DQA1 rs9271366 and HLA-DQB1/HLA-DQA2 rs9275328. Our aim was to investigate these SNPs in a Malaysian SLE cohort.
MATERIALS AND METHODS: SNPs rs9271366 and rs9275328 were screened across 790 Malaysian citizens from three ethnic groups (360 patients and 430 healthy volunteers) by Taqman SNP genotyping assays. Allele and genotyping frequencies, Hardy-Weinberg equilibrium, Fisher's exact test and odds ratio were calculated for each SNP and ethnic group. Linkage disequilibrium and interaction between the two SNPs were also evaluated.
RESULTS: The minor allele G and its homozygous genotype GG of HLA-DRB1/HLA-DQA1 rs9271366 significantly increased the SLE susceptibility in Malaysian patients, including those of Malay and Chinese ethnicity (odds ratio (OR) > 1, p
Two ticks were collected from a reticulated python (Malayopython reticulatus) caught in Tumpat District, Kelantan, Peninsular Malaysia. The ticks were first identified as Ambylomma sp. through morphological comparison with identification keys. Determination of the tick species was made through PCR and sequencing. However, BLAST analysis revealed 85-88% sequence nucleotide identity with Amblyomma nitidum and Amblyomma geoemydae. Additionally, the morphological features of the ticks collected in this study did not match either A. nitidum or A. geoemydae. Further examination of the ticks confirmed the species as Amblyomma cordiferum. This is the first record of A. cordiferum DNA sequence with morphological support of colour illustrations for adult A. cordiferum. This is also the most recent record of this host association in Peninsular Malaysia. Information from this report can serve as a reference for species identification using the described morphology or molecular sequences.
Amebiasis is one of the major causes of diarrhea in the developing countries and it can present with a wide range of gastrointestinal symptoms depending on the phase of infection. We described a case of 50 year-old male patient who presented with abdominal pain, diarrhea and vomiting. After right hemicolectomy for appendicular abscess with tumour over the ileum, histopathological examinations revealed numerous trophozoites of Entamoeba histolytica in a background of inflammations (Figure 1). Following resection of the ameboma, he received intravenous metronidazole treatment for total of two weeks duration.
Patients with end stage renal disease on hemodialysis are affected by physiological and psychological stressors, which contribute to poor quality of life and negative clinical outcomes. Depression, anxiety, and stress are highly prevalent in this population. Effective interventional strategies are required to manage these psychological symptoms. Acupressure has been believed to be one of the complementary therapies that could promote psychological wellbeing.
Peritonitis is well recognized as the Achilles tendon of peritoneal dialysis (PD). Reoccurrence of peritonitis due to the same organism, defined as either repeat or relapsing peritonitis under the 2005 guidelines by the International Society for Peritoneal Dialysis, often results in PD technique failure. Rothia dentocariosa, a low-virulent human oropharynx commensal, is a rarely reported pathogen in human infection, particularly infective endocarditis. R. dentocariosa PD-related peritonitis is exceedingly uncommon yet potentially results in repeat or relapsing peritonitis which requires catheter removal. We report a case of R. dentocariosa repeat and relapsing peritonitis in a PD patient who was treated successfully with antimicrobial therapy.
Pneumocystis jirovecii (formerly Pneumocystis carinii) pneumonia (PCP) is a rare but serious infection that usually occurs within a year after solid organ transplantation. PCP may occur after 1 year post transplantation, but the rate is reported to be very low. Studies have shown an association between cytomegalovirus (CMV) infection in solid organ transplant patients and an increased risk of opportunistic infection. This increased risk is thought to be a result of the immunomodulatory effects of the CMV infection. We present a case of PCP infection occurring 13 years after a renal transplantation. This occurred following a recurrent CMV infection while the patient was on low-dose immunosuppressants.
Gastrointestinal stromal tumour (GIST) is a rare but most common mesenchymal tumour in the gastrointestinal tract. Although GIST research has been carried out extensively worldwide, it has yet to be studied in Malaysia. To establish the immunohistochemical expression pattern of CD117 (c-KIT), CD34, S-100 and Desmin, the incidence of c-KIT and PDGFRA genes mutation in GISTs, and correlate it with clinicopathological parameters. Eleven clinically diagnosed GISTs were stained for CD117, CD34, Desmin and S-100 protein by immunohistochemical technique, and c-KITand PDGFRA gene mutations were studied by PCR-CSGE-DNA sequencing method. All GISTs (7 cases) stain positive for CD117, and co-expressed CD34. None of these cases express Desmin, and only one expressed S-100 protein focally. Fifty-seven percent (4/7 cases) of GIST harboured mutations at exon 11 of c-KIT gene, and they were all high risk and malignant cases. No mutation was detected at exons 9, 13 and 17 of KIT gene, and exons 12 and 18 of PDGFRA gene. Immunohistochemistry using a panel of antibodies shows consistent pattern of CD117 and CD34 expression in GIST, and mutational study may be a useful prognostic marker for kinase inhibitor treatment of GIST.
BACKGROUND AND AIMS: Colorectal cancers of different subtypes involve different pathogenic pathways like the Wnt and the mutator pathways. In this study, we screened 73 colorectal cancer cases from a multi-racial group for genetic and expression profile defects with the aim of correlating these with patients' clinicopathological characteristics.
METHODS: Mutation screening of the entire coding region of APC and exon 3 of CTNNB1, loss of heterozygosity (LOH) of APC, and microsatellite instability (MSI) status were assessed for 44 patients with available paired frozen normal and tumour tissues. In addition, 29 cases with available paraffin embedded tumour blocks were screened for mutation in exon 3 of CTNNB1, the APC mutation cluster region (codon 1286-1513), and hMLH1, hMSH2, hMSH6 protein expressions by immunohistochemistry method.
RESULTS: In our study, 15/73 cases showed APC mutations (20.5%), 1/73 cases had CTNNB1 mutation (1.4%), 5/32 cases had APC LOH (15.6%), and 16/70 (22.9%) cases revealed at least some form of mismatch repair (MMR) defect. Tumour grade (poor differentiation) was found to correlate significantly with right-sided tumour and mucinous histology (p = 0.01879 and 0.00320, respectively). Patients of younger age (below 45 years) more often had tumours of mucinous histology (p = 0.00014), while patients of older age (above 75 years) more often had tumours on the right side of the colon (p = 0.02448). Tumours of the mucinous histology subtype often had MMR defects (p = 0.02686). There was no difference in the occurrence of APC and CTNNB1 mutations and MMR defects found within our multi-racial colorectal cancer patient cohort.
CONCLUSION: Our findings support the notion that racial factor may not be related to the occurrence of MMR defects and APC and CTNNB1 mutations in our multi-racial patient cohort.
Kelantan is a chiefly agrarian state with abundant small-holder ruminant farms in the East Coast economic Region of Malaysia. Ectoparasitism affects small ruminant production in Malaysia. It often causes reduction in meat quality and milk production which affect the farmers' income. To date, no report for the prevalence of ectoparasitism on small ruminant in Kelantan compared to other state in Malaysia. This study aims to determine the prevalence and associated risk factor of ruminant ectoparasitism in Kelantan. Ectoparasites were collected by manual picking and skin scrapping from 462 sheep and goats in Kelantan between April and September 2017 (during dry season). 60% of the sampled animals were infested with at least one species of the ectoparasites. In this study, lice and ticks were the most prevalent ectoparasites on small ruminant, which were 43.64% and 22.98%, respectively. The high biotic potential of lice population on host might be one of the factors they become the most prevalent species found on the animals. There was no significant relationship between ectoparasitism prevalence and species of small ruminants (χ2 = 1.12, p = 0.293). However, there was significant variations in prevalence between the regions where the animals were sampled from (χ2 = 30.25, p = 0.002) and farm management system for both species. This present study provides baseline epidemiological data on the prevalence of ectoparasitism in small ruminant. This information is useful for the formulation of prevention and control measures in order to enhance ruminant productivity in Kelantan.
Penicilliosis is a rare occurrence among non human immunodeficiency virus (HIV) infected patients. We report here two cases of Penicillium marneffei infection in patients with systemic lupus erythematosus (SLE). Both patients had a recent flare of lupus and were on immunosuppressive drugs when they presented with prolonged fever without an obvious foci of infection, unresponsive to broad-spectrum antibiotics. They were leucopaenic upon admission, with rapid deterioration during the course of the illness. Diagnosis of penicilliosis via fungal isolation from blood culture was delayed resulting in the late initiation of antifungal agents. While both patients ultimately recovered, the delay in diagnosis led to a prolonged hospital stay with increased morbidity. Clinicians should be aware of this uncommon but emerging fungal pathogen in SLE patients and maintain a high index of suspicion in diagnosing this potentially fatal but treatable disease.
Tritrichomonas foetus is known to cause chronic diarrhea in the feline species in many different regions of the world. However, there is a paucity of information on T. foetus among cats in Malaysia. This study was conducted to determine the prevalence of Tritrichomonas foetus in the pet and stray cat population in Klang Valley, Malaysia. A total of 201 pet and stray cats' fecal samples were collected in Klang Valley. 24 samples were cultured in the InPouch® TF Feline to observe for motile trophozoites. A nested PCR protocol was used to screen for T. foetus in the collected samples. The prevalence of T. foetus in the cat population in Klang Valley was 33%. There was no association between Tritrichomonas infection and age, sex, breed or management of the cats. However, statistical analysis revealed that stray cats were more likely to be infected with T. foetus compared to pet cats. This study confirmed for the first time the presence of T. foetus among the cat population in Klang Valley, Malaysia.
Toxoplasma gondii, a zoonotic protozoan that has a worldwide distribution, is known to infect many warm-blooded vertebrates. The feline species including domestic cats are the definitive hosts for Toxoplama gondii and shed the infective oocyst. There is lack of information on the prevalence of Toxoplasma gondii in cats in Malaysia. The objective of this study was to determine both the seroprevalence of T. gondii and the prevalence of T. gondii DNA in cats' feces in Klang Valley, Malaysia. 198 blood and 201 fecal samples were collected from pet and stray cats from the local council, Dewan Bandaraya Kuala Lumpur (DBKL) and University Veterinary Hospital, Universiti Putra Malaysia respectively. The overall seroprevalence of Toxoplasma gondii in cats in the Klang Valley was found to be 5.5%. There was a high prevalence (10.5%) of T. gondii DNA detected in the cat fecal samples in both pet and stray cats suggestive of T. gondii oocyst shedding. Stray cats showed a higher seroprevalence and molecular prevalence of T. gondii than the pet cats. However, comparative analysis using Chi-square test showed no significant difference between both groups (P>0.05). Higher prevalence (10.5%) of cats shedding T. gondii DNA as compared to the seroprevalence (5.5%) was found in the cat population in the Klang Valley. The high prevalence of cats shedding T. gondii DNA is alarming as this may directly reflect the number of oocysts excreted into the environment posing a significant public health hazard.
The Epstein–Barr virus (EBV) can cause a wide variety of cancers upon infection of different cell types and induces a highly variable composition of the tumor microenvironment (TME). This TME consists of both innate and adaptive immune cells and is not merely an aspecific reaction to the tumor cells. In fact, latent EBV-infected tumor cells utilize several specific mechanisms to form and shape the TME to their own benefit. These mechanisms have been studied largely in the context of EBV+ Hodgkin lymphoma, undifferentiated nasopharyngeal carcinoma, and EBV+ gastric cancer. This review describes the composition, immune escape mechanisms, and tumor cell promoting properties of the TME in these three malignancies. Mechanisms of susceptibility which regularly involve genes related to immune system function are also discussed, as only a small proportion of EBV-infected individuals develops an EBV-associated malignancy.
That water may not be an inert medium was indicated by the presence at water's interfaces a negatively charged solute free zone of several hundred microns in thickness called the exclusion zone (EZ). Further evidence was demonstrated by Ovchinnikova's experiments (2009) showing that water can store and release substantial amount of charge. We demonstrate that the charge storage capacity of water arises from highly stable large-scale ionic structures with measurable charge imbalances and discrete levels of charge density. We also show evidence that the charge zones formation requires ionic solutes, and their formation correlate to large change in conductivity, by as much as 250%. Our experiments indicate that large-scale structuring plays a pivotal role in electrolysis and conductivity in ionic solution. We propose that water is an electrochemically active medium and present a new model of electrolysis and conductivity in ionic solution.
This paper investigates the effect of different categories of essential COVID-19 data from 2020 to 2021 towards stock price dynamics and options markets. It applied the hypothetical method in which investors develop depression based on the understanding suggested by various green finance divisions. Furthermore, additional elements like panic, sentiment, and social networking sites may impact the attitude, size, and direction of green finance, subsequently impacting the security prices. We created new emotion proxies based on five groups of information, namely COVID-19, marketplace, lockdown, banking sector, and government relief using Google search data. The results show that (1) if the proportional number of traders' conduct exceeds the stock market, the effect of sentimentality indexes on jump volatility is expected to change; (2) the volatility index component jump radically increases with the COVID-19 index, city and market lockdown index, and banking index; and (3) expanding the COVID-19 index gives rise to the stock market index. Moreover, all indexes decreased in jump volatility but only after 5 days. These findings comply with the hypotheses proposed by our model.