Comparison of Automated and Manual Viral Nucleic Acid Extraction Kits for Covid-19 Detection Using qRT-PCR

@#Introduction: The emergence of a novel Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in a pandemic. Rapid and accurate diagnosis method is crucial to reduce the disease burden and to improve early diagnosis approaches to control of the disease. Real time Reverse transcriptase PCR (qRT-PCR) has been identified by the World Health Organization as the most sensitive and specific method of detection. However, the success of this assay relies on the quantity and quality of the extracted viral RNA. Methods: Various methods have been developed for nucleic acid extraction however, the methods have not been assessed. RNA extraction was performed from 24 nasopharyngeal swab samples using a manual extraction kit (GF-1) and an automated extraction kit (Genolution). The concentration and purity of the extracted RNA samples were measured, and its performance were tested using qRT-PCR. Results: The average concentration and purity of the RNA samples extracted using GF-1 kit was higher compared to Genolution. Similarly, the qRT-PCR assay using the RNA samples extracted using manual extraction was better compared to automated kit. Conclusion: Both the manual and automated extraction kits have its advantages and disadvantages in terms of yield and purity. However, with proper optimization, both methods may be used for routine molecular diagnostic of COVID-19 in laboratories.

Detection of Pathogenic Leptospira in Rats and Phylogenetic Analysis Using Outer Membrane Lipoprotein Lipl32 Gene at Two Major Public Markets

@#Introduction: Limited studies have been documented on the presence of pathogenic Leptospira in public markets serving the community in sub-districts of Selangor. The aim of this study was to detect the presence of pathogenic Leptospira in rats using a gene encoding an outer membrane lipoprotein LipL32. Methods: Polymerase chain reaction (PCR) was performed using LipL32 primers on sixty kidney samples of rats trapped at two locations of study; Pasar Borong Selangor in Seri Kembangan and Pasar Basah Bandar Baru Bangi in Bangi. Results: Out of 60 samples analysed, 36.7% were positive for the presence of LipL32. All positive samples highly matched (>94%) nucleotide sequence for LipL32 of pathogenic Leptospira and related to the pathogens through phylogenetic analysis. Conclusion: The detection of LipL32 indicates the potential presence of pathogenic Leptospira species at public markets. Although only 60 rats were successfully trapped, the rats are mobile and might further transmit the pathogenic organisms to other areas.

Feasibility and Acceptability of MyWarung©: A Food Poisoning Prevention Smartphone-Apps During Dining Out

@#Introduction: Eating out has always been associated with increasing cases of food poisoning. These problems can be minimized through mobile applications and technology development. A mobile application called MyWarung© was developed to provide an alternative, improved tool for improving food poisoning knowledge and preventive behaviour. Methods: This cross-sectional study aims to assess the feasibility and acceptance of the MyWarung© application for consumers in Terengganu. The 50 consumers were selected based on the inclusion and exclusion criteria using convenience non-probability sampling. The data were collected through a questionnaire that included three components: socio-demographic, feasible (6 components) and acceptable (7 components). The scoring above 80.0% indicates an acceptable, while lower than 80.0% show unacceptable for both feasibility and acceptability sections. SPSS 22.0 has analyzed the data. Results: The results showed excellent feasibility with a median score of 27.5 (IQR 6.0) out of 30.0, and acceptance with 32.0 (IQR 7.0) out of 35.0. Majority of the respondents agreed that the app is easy to use (94.0%), easy to understand (88.0%), attractive (84.0%), catchy (88.0%), provides more information (96.0%), efficient (96.0%), knowledge improvement (96.0%), beneficial (100.0%), useful application (88.0%), and recommend to the other people (84.0%). The overall result showed that most respondents agreed that MyWarung© application was feasible and acceptable with 90.0% and 86.0% feasibility and acceptability rate. Conclusion: The MyWarung© application among consumers can be highly feasible and acceptable in preventing food poisoning during dining out.

Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples

Objective: To compare the efficiency of routine diagnostic PCR assays in detecting pathogenic Leptospira isolated from water and soils. Methods: Seven routine assays targeting six genes (lipL32, flaB, gyrB, lfb1, secY and ligB) were evaluated and compared on the cultures of two groups of pathogenic Leptospira from different sources. One group included 19 described reference strains recovered from infected human or animals, and another group included 22 environmental isolates from recreational and residential sites in Malaysia. The latter have been confirmed for presence of pathogenic Leptospira DNA. PCR positivity or detection sensitivity of each assay was determined and compared between the two groups. Results: Validation on reference strains showed 100.0% PCR sensitivity for all assays except ligB-PCR (95.0%) that failed to amplify Leptospira interrogans serovar Pomona. In marked contrast, there was a notable decline in sensitivity in the environmental isolates (lipL32-PCR, 95.5%;flaB-PCR, 90.9%; gyrB-PCR, 77.3%; lfb1-PCR, 59.1%; secY-PCRs, 40.9% G1/G2-PCR, 36.4%; ligB-PCR, 13.6%), implying a large genetic distance between the two groups, as well as nucleotide polymorphism among environmental isolates. Conclusions: High proportion of false-negative PCR results suggests a need of prudent selection of primers in detecting environmental pathogenic Leptospira. These findings offer valuable insights on the extensive biodiversity of genus Leptospira and its impact on the efficacy and development of molecular detection tool.