SARS-CoV-2 variants typing using real-time reverse transcription-PCR-based assays in Addis Ababa, Ethiopia
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Date
2023-05
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Addis Ababa University
Abstract
Background: Severe Acute Respiratory Syndrome Coronavirus 2 is a single-stranded positive
RNA that possesses 30,000 base pair in diverse variants. Even though genomic sequencing is a
well-established variant detection method; but due to its cost and longer result turnaround time
attributes less attractive for variant identification in resource-limited countries. These complexities
pose the need for RT-PCR based variant typing.
Objective: This study aimed to determine the type of SARS-CoV-2 variants in the first four
consecutive COVID-19 waves using variant typing PCR in Addis Ababa, Ethiopia.
Methods: A cross-sectional study was conducted from nasopharyngeal samples source from EPHI
COVID-19 biobank repository. Samples were randomly selected from the first four waves based
on their collection dates. A total of 641 NP samples were selected and tested for SARS Cov-2.
RNA was extracted using Bioer NPA-32P instrument (Zhejiang, China) using the extraction kit
from MagaBio Plus RNA Purification Kit II (Hangzhou, China). The SARS-CoV-2 detection
carried out from 10 μl RNA and 20 μl RT-PCR fluorescent mix (Shenzhen, China). The Ct value
less than 38 considered positive per the manufacturer. B.1.617 Lineage and 6 S-gene mutation
(Shenzhen, China) PCR kits used for variant typing.
Result: From the 374 total tested for variant detection, 267 (71.4%) were identified by the variant
typing kits. The remaining, 107(28.6%) were not classified by both variant detection kits. Alpha,
Beta, Delta, and Omicron, were dominantly identified variants from wave one, two, three and four,
respectively. Based on the WHO variant classification, Alpha variant was identified with high
proportion from wave-1 but absent in wave-4. Similarly, Beta variant detected from wave 1 to 3
with varied proportion but dominant in wave-2. Delta variant entirely identified from wave 3 and
4; while dominantly identified in wave 3. Omicron was the dominant variant of wave 4. From the
total identified positive study samples, 243/267 (91%) variants identified from samples had Ct
values less than 30 in diagnostic RT-PCR. Besides to VOC; VOI also identified.
Conclusion: The study data demonstrated that RT-PCR type variants from confirmed SARSCoV-
2-positive sample. RT-PCR based variant typing would provide additional screening opportunity;
where sequencing opportunity is inaccessible. The assays could be implemented in laboratories
that perform SARS-CoV-2 molecular testing.
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Keywords
SARS-CoV-2, whole genome sequence, variants, Ct value