Identification of SARS CoV‐2 Omicron BA.1 and a novel Delta lineage by rapid methods and partial spike protein sequences in Sulaymaniyah Province, Iraq

Background Five variants of concern (VOCs) of severe acute respiratory syndrome coronavirus 2 (SARS CoV‐2) have been globally recorded including Alpha, Beta, Gamma, Delta, and Omicron. The Omicron variant has outcompeted the other variants including the Delta variant. Molecular screenings of VOCs are important for surveillance, treatment, and vaccination programs. This study aimed to identify VOCs by using rapid inexpensive methods and partial sequencing of the virus's spike gene. Methods Mutation‐specific rRT PCR probes were used for both D614G and K417N mutations to potentially discriminate between Delta and Omicron variants. These were followed by sequencing of a fragment of spike gene (748 nucleotides), which covers the most notable VOC mutations in the receptor binding domain of SARS CoV‐2. Results Rapid methods showed that out of 24 SARS CoV‐2 positive samples, 19 carried the N417 mutation, which is present in the Omicron variant. Furthermore, 3 samples carried K417 wildtype, which is present in the Delta variant. Additionally, 2 samples containing both K417 and N417 suggested mixed infections between the two variants. The D614G mutation was present in all samples. Among the 4 samples sequenced, 3 samples carried 13 mutations, which are present in Omicron BA.1. The fourth sample contained the two common mutations (T478K and L452R) present in Delta, in addition to two more rare mutations (F456L and F490S), which are not commonly seen in Delta. Our data suggested that both Omicron variant BA.1 and a novel Delta variant might have circulated in this region that needs further investigations. Investigationos of SARS CoV‐2 variants of concerns (VOCs) have been ignored in developing countries including Iraq. Spike gene sequencings and rapid methods were used to identify VVOCs. Current study identified the Omicron BA1 sub‐variant and a novel Delta variant in Sulaymaniah province of Iraq.

Identification of SARS CoV-2 Omicron BA.1 and a novel Delta lineage by rapid methods and partial spike protein sequences in Sulaymaniyah Province, Iraq.

BACKGROUND: Five variants of concern (VOCs) of severe acute respiratory syndrome coronavirus 2 (SARS CoV-2) have been globally recorded including Alpha, Beta, Gamma, Delta, and Omicron. The Omicron variant has outcompeted the other variants including the Delta variant. Molecular screenings of VOCs are important for surveillance, treatment, and vaccination programs. This study aimed to identify VOCs by using rapid inexpensive methods and partial sequencing of the virus's spike gene. METHODS: Mutation-specific rRT PCR probes were used for both D614G and K417N mutations to potentially discriminate between Delta and Omicron variants. These were followed by sequencing of a fragment of spike gene (748 nucleotides), which covers the most notable VOC mutations in the receptor binding domain of SARS CoV-2. RESULTS: Rapid methods showed that out of 24 SARS CoV-2 positive samples, 19 carried the N417 mutation, which is present in the Omicron variant. Furthermore, 3 samples carried K417 wildtype, which is present in the Delta variant. Additionally, 2 samples containing both K417 and N417 suggested mixed infections between the two variants. The D614G mutation was present in all samples. Among the 4 samples sequenced, 3 samples carried 13 mutations, which are present in Omicron BA.1. The fourth sample contained the two common mutations (T478K and L452R) present in Delta, in addition to two more rare mutations (F456L and F490S), which are not commonly seen in Delta. Our data suggested that both Omicron variant BA.1 and a novel Delta variant might have circulated in this region that needs further investigations.

Association of Toll-like receptor-4 polymorphism with SARS CoV-2 infection in Kurdish Population

Genetic variations are critical for understanding clinical outcomes of infections including server acute respiratory syndrome coronavirus 2 (SARS CoV-2). The immunological reactions of human immune genes with SARS CoV-2 have been under investigation. Toll-like receptors (TLRs), a group of proteins, are important for microbial detections including bacteria and viruses. TLR4 can sense both bacterial lipopolysaccharides (LPS) and endogenous oxidized phospholipids triggered by Covid-19 infection. Two TLR4 single nucleotide polymorphisms (SNPs), Asp299Gly and Thr399Ile have been linked to infectious diseases. No studies have focused on these SNPs in association with Covid-19. This study aims to reveal the association between Covid-19 infection with these SNPs by comparing a group of patients and a general population. Restriction fragment length polymorphisms (RFLP) were used to identify the TLR4 SNPs in both the general population (n = 114) and Covid-19 patient groups (n = 125). The results found no association between the TLR4 polymorphisms and Covid-19 infections as the data showed no statistically significant difference between the compared groups. This suggested that these TLR4 SNPs may not be associated with Covid-19 infections.

Association of tumor necrosis factor alpha ‐308 single nucleotide polymorphism with SARS CoV‐2 infection in an Iraqi Kurdish population

Uncovering risk factors playing roles in the severity of Coronavirus disease 2019 (Covid‐19) are important for understanding pathoimmunology of the disease caused by severe acute respiratory syndrome Coronavirus 2 (SARS CoV‐2). Genetic variations in innate immune genes have been found to be associated with Covid‐19 infections. A single‐nucleotide polymorphism (SNP) in a promoter region of tumor necrosis factor alpha (TNF‐α) gene, TNF‐α −308G>A, increases expression of TNF‐α protein against infectious diseases leading to immune dysregulations and organ damage. This study aims to discover associations between TNF‐α −308G>A SNP and Covid‐19 infection. Polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) was used for genotyping a general Kurdish population and Covid‐19 patients. The homozygous mutant (AA) genotype was found to be rare in the current studied population. Interestingly, the heterozygous (GA) genotype was significantly (p value = 0.0342) higher in the Covid‐19 patients than the general population. This suggests that TNF‐α −308G>A SNP might be associated with Covid‐19 infections. Further studies with larger sample sizes focusing on different ethnic populations are recommended.

Association of Apolipoprotein e polymorphism with SARS-CoV-2 infection.

Coronavirus 2019 (COVID-19) is a viral disease caused by severe acute respiratory syndrome coronavirus-2 (SARS CoV-2). The disease resulted in global morbidity and mortality that led to considering as pandemic. The human body response to COVID-19 infection was massively different from being asymptomatic to developing severe symptoms. Host genetic factors are thought to be one of the reasons for these disparities in body responses. Few studies have suggested that Apolipoprotein Epsilon (Apo E) is a candidate gene for playing roles in the development of the disease symptoms. This work aims to find an association between different Apo E genotypes and alleles to COVID-19 infection comparing a general population and a group of COVID-19 patients. For the first time, the results found that Apo E4 is associated with COVID-19 disease in a Kurdish population of Iraq. Further study is required to reveal this association in different ethnic backgrounds all over the world.

Rapid, inexpensive methods for exploring SARS CoV-2 D614G mutation.

A common mutation has occurred in the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS CoV-2), known as D614G (A23403G). There are discrepancies in the impact of this mutation on the virus's infectivity. Also, the whole genome sequencings are expensive and time-consuming. This study aims to develop three fast economical assays for prompt identifications of the D614G mutation including Taqman probe-based real-time reverse transcriptase polymerase chain reaction (rRT PCR), an amplification refractory mutation system (ARMS) RT and restriction fragment length polymorphism (RFLP), in nasopharyngeal swab samples. Both rRT and ARMS data showed G614 mutants indicated by the presence of HEX probe and 176 bp, respectively. Additionally, the results of the RFLP data and DNA sequencings confirmed the prevalence of the G614 mutants. These methods will be important, in epidemiological, reinfections and zoonotic aspects, through detecting the G614 mutant in retro-perspective samples to track its origins and future re-emergence of D614 wild type.