Choga, W. T. | bioRxiv Science

bioRxiv2024

OPTIMIZATION OF DRIED BLOOD SPOT ASSAYS FOR HEPATITIS B VIRUS SURFACE ANTIBODY QUANTIFICATION

Dried blood spot (DBS) cards can be used as an alternative sample collection method to plasma, however, there is no optimized elution protocol for DBS cards specifically for hepatitis B surface antibody (anti-HBs) testing. The study aimed to develop a DBS elution protocol for anti-HBs quantification. Our study sought to determine the ideal phosphate-buffered saline (PBS) buffer volume to use by comparing three PBS volumes (300uL, 450uL, and 500uL), and the optimal time to agitate DBS discs on a plate shaker (1hr, 2hrs, 3hrs, and 4hrs) to yield DBS anti-HBs concentrations that are comparable to corresponding plasma anti-HBs concentrations. The optimal DBS storage temperature (25{degrees}C, -20{degrees}C, and -80{degrees}C) was investigated to determine the ideal longterm storage temperature of the cards. Residual samples were used for optimization (2019-2021). A total of 50 DBS-plasma pairs was used throughout the study, with plasma anti-HBs concentrations being used as the golden standard to compare. The analysis of results was carried out by determining the p-values of the Wilcoxon sign rank. A two-way analysis of variance (ANOVA) was also performed to determine the impact of PBS elution volumes, elution time, and storage temperature on the anti-HBs concentration of DBS samples on STATA Version 15.0. No statistically significant difference between the DBS-plasma anti-HBs pairs was observed when using 450 or 500uL of PBS buffer and when samples were agitated for 3 hours (p=0.594, p=0.499 respectively). The optimal storage temperature for DBS cards was 25{degrees}C because the results showed no statistically significant difference between DBS-plasma anti-HBs titers (p=0.594). The two-way ANOVA analysis showed that elution volumes and time had no statistically significant impact on the DBS anti-HBs concentrations, p=0.948 and p=0.381 respectively. Storage temperature had a statistically significant impact on the DBS anti-HBs concentrations, p=0.002. The optimized DBS elution protocol for anti-HBs quantification will help monitor vaccine efficacy in infants due to the low sample volumes required compared to plasma and also can be used for anti-HBs testing in resource-limited areas around the country.

microbiology↗

bioRxiv2020

Whole Genome Sequencing-based Characterization of Human Genome Variation and Mutation Burden in Botswana

The study of human genome variations can contribute towards understanding population diversity and the genetic aetiology of health-related traits. We sought to characterise human genomic variations of Botswana in order to assess diversity and elucidate mutation burden in the population using whole genome sequencing. Whole genome sequences of 390 unrelated individuals from Botswana were available for computational analysis. The sequences were mapped to the human reference genome GRCh38. Population joint variant calling was performed using Genome Analysis Tool Kit (GATK) and BCFTools. Variant characterisation was achieved by annotating the variants with a suite of databases in ANNOVAR and snpEFF. The genomic architecture of Botswana was delineated through principal component analysis, structure analysis and FST. We identified a total of 27.7 million unique variants. Variant prioritisation revealed 24 damaging variants with the most damaging variants being ACTRT2 rs3795263, HOXD12 rs200302685, ABCB5 rs111647033, ATP8B4 rs77004004 and ABCC12 rs113496237. We observed admixture of the Khoe-San, Niger-Congo and European ancestries in the population of Botswana, however population substructure was not observed. This exploration of whole genome sequences presents a comprehensive characterisation of human genomic variations in the population of Botswana and their potential in contributing to a deeper understanding of population diversity and health in Africa and the African diaspora.

genomics↗