Biology subjects

Ngetsa, C.

Publications and source records attributed to Ngetsa, C..

3 recordsLinked to original sources

bioRxiv2022

Validation of saline, PBS and a locally produced VTM at varying storage conditions to detect the SARS-CoV-2 virus by qRT-PCR

Coronavirus Disease-2019 tests require a Nasopharyngeal (NP) and/or Oropharyngeal (OP) specimen from the upper airway, from which virus RNA is extracted and detected through quantitative reverse transcription-Polymerase Chain Reaction (qRT-PCR). The viability of the virus is maintained after collection by storing the NP/OP swabs in Viral Transport Media (VTM). We evaluated the performance of four transport media: locally manufactured ("REVITAL") Viral Transport Media (RVTM), Standard Universal Transport Media (SUTM), PBS and 0.9% (w/v) NaCl (normal saline). We used laboratory cultured virus to evaluate: i) viral recovery and maintaining integrity at different time periods and temperatures; ii) stability in yielding detectable RNA consistently for all time points and conditions; and iii) their overall accuracy. Four vials of SARS-CoV-2 cultured virus (2 high and 2 low concentration samples) and 1 negative control sample were prepared for each media type (SUTM, RVTM, PBS and normal saline) and stored at the following temperatures, -80{degrees}C, 4{degrees}C, room temperature (25{degrees}C) and 37{degrees}C for 7 days. Viral Ribonucleic acid (RNA) extractions and qRT-PCR were done on the following days after inoculation with the cultured virus, days 1, 2, 3, 4 and 7 to assess virus stability and viral recovery. CT values fell over time at room temperature, but normal saline, PBS, RVTM and SUTM all showed comparable performance in maintaining virus integrity and stability allowing for the detection of SARS-CoV-2 viral RNA. Overall, this study demonstrated that normal saline, PBS and the locally manufactured VTM can be used for COVID-19 sample collection and testing, thus expanding the range of SARS-CoV-2 viral collection media.

molecular biology↗

bioRxiv2021

Improving statistical power in severe malaria genetic association studies by augmenting phenotypic precision

Severe falciparum malaria has substantially affected human evolution. Genetic association studies of patients with clinically defined severe malaria and matched population controls have helped characterise human genetic susceptibility to severe malaria, but phenotypic imprecision compromises discovered associations. In areas of high malaria transmission the diagnosis of severe malaria in young children and, in particular, the distinction from bacterial sepsis, is imprecise. We developed a probabilistic diagnostic model of severe malaria using platelet and white count data. Under this model we re-analysed clinical and genetic data from 2,220 Kenyan children with clinically defined severe malaria and 3,940 population controls, adjusting for phenotype mis-labelling. Our model, validated by the distribution of sickle trait, estimated that approximately one third of cases did not have severe malaria. We propose a data-tilting approach for case-control studies with phenotype mis-labelling and show that this reduces false discovery rates and improves statistical power in genome-wide association studies.

genetics↗

bioRxiv2018

Performance of an IAVI-African Network of Clinical Research Laboratories in Standardized ELISpot and Peripheral Blood Mononuclear Cell Processing in Support of HIV Vaccine Clinical Trials

Immunological assays performed in different laboratories participating in multi-centre clinical trials must be standardized in order to generate comparable and reliable data. This entails standardized procedures for sample collection, processing, freezing and storage. The International AIDS Vaccine Initiative (IAVI) partnered with local institutions to establish Good Clinical Laboratory Practice (GCLP)-accredited laboratories to support clinical trials in Africa, Europe and Asia. Here we report on the performance of seven laboratories based in Africa and Europe in the interferon-gamma enzyme-linked immunospot (IFN-{gamma} ELISpot) assay and peripheral blood mononuclear cell (PBMC) processing over four years. Characterized frozen PBMC samples from 48 volunteer blood packs processed at a central laboratory were sent to participating laboratories. For each stimulus, there were 1751 assays performed over four years. 98% of these ELISpot data were within acceptable ranges with low responses to mock stimuli. There were no significant differences in ELISpot responses at five laboratories actively conducting immunological analyses in support of IAVI sponsored clinical trials or HIV research. In a separate study, 1,297 PBMC samples isolated from healthy HIV-1 negative participants in clinical trials of two prophylactic HIV vaccine candidates were analysed for PBMC yield from fresh blood and cell recovery and viability following freezing and thawing. 94 % and 96 % of samples had fresh PBMC viabilities and cell yields within the pre-defined acceptance criteria while for frozen PBMC, 99 % and 96 % of samples had acceptable viabilities and cell recoveries respectively, along with acceptable ELISpot responses in 95%. These findings demonstrate the competency of laboratories across different continents to generate comparable and reliable data in support of clinical trials.\n\nImportanceThere is a need for the establishment of an African network of laboratories to support large clinical trials across the continent to support and further the development of vaccine candidates against emerging infectious diseases such as Ebola, Zika and dengue viruses and the continued HIV-1 pandemic. This is particularly true in sub-Saharan Africa where the HIV-1 pandemic is most severe. In this report we have demonstrated by using standardized SOPs, training, equipment and reagents that GCLP-accredited clinical trial laboratories based in Africa and Europe can process clinical trial samples and maintain cell integrity and functionality demonstrated by IFN-{gamma} ELISpot testing, producing comparable and reliable data.

immunology↗