Association of Anti-N and -S Seroprevalence in Asymptom atic, Mildly Symptomatic, and Symptomatic SARS-COV-2 Natural Infection

Backgrounds: SARS-COV-2 infection is not always correlated with protection. Antibody seroprevalence in unvaccinated individuals, which is usually measured by N-specific antibodies, is not necessarily correlated with protection, while antibodies against S protein show a better correlation with protection due to its neutralizing epitopes. In this study, we tried to improve our conception of the hidden perspective of SARS-COV-2 in epidemiological reports and investigate anti-S antibody prevalence among anti-N antibody-positive asymptomatic and mildly symptomatic patients. Material(s) and Method(s): Blood samples were collected from asymptomatic or mildly symptomatic volunteer participants and symptomatic hospitalized patients with negative PCR results from May 30 to June 17, 2020. Detection of SARS-COV-2 antibodies was done using an ELISA kit targeting N or S protein. Finding(s): Totally, 716 samples from volunteer participants and 81 samples from symptomatic hospitalized patients with negative PCR results were evaluated. The test performance-adjusted seroprevalence (%95 CI) of SARS-COV-2 antibody was 17.3% (8.8-25.8%) for anti-N IgG in volunteers and 25.5% (12.8-39.7%) for anti-N and anti-S IgM in hospitalized patients. Among anti-N IgG positive infected individuals, %49.2 (21.4 and 78.8%) were anti-S antibody positive. Conclusion(s): The results showed that SARS-COV-2 infection sometimes occurs in individuals without symptoms or with mild symptoms, but in more than half of them, the produced antibody is not protective. The findings of hospitalized patients showed that the combination of IgM assay with real-time PCR improved the disease diagnosis by more than 25% in cases with negative molecular test results.Copyright © 2022, TMU Press.

False-negative results in taqman one-step rt-pcr test: Evaluation of endogenous internal control function used in sars-cov-2 detection tests

Background: Taqman one-step real-time PCR (RT-PCR) has special importance due to its high sensitivity and specificity in the diagnosis of infectious diseases such as viral infections. In the recent pandemic of SARS-CoV-2, diagnostic kits based on this method are commonly used for molecular detection. One of the main systematic errors that misinterpret the results is using inaccurate internal control in RT-PCR diagnostic kits. Designing primers and probes that span exon-exon junction will avoid genomic DNA amplification and lead to obtaining high specific results. Objectives: This study aimed to evaluate the endogenous internal control of primers and probe for RNase P RNA to reduce false-negative results in respiratory samples. Methods: In this study, 30 samples of patients who were negative for SARS-CoV-2, influenza A, and influenza B were re-evaluated for SARS-CoV-2 using newly designed primers and probes for RNase P RNA (ultra-specific primers and probe). We also performed bioinformatics analysis on CDC-approved primers and probes of RNase P endogenous internal control. Results: In this analysis, we specified the location of these newly designed primers and probe on target mRNA and genomic DNA. Then, the Taqman one-step RT-PCR method was performed using both CDC-approved primers and probes along with our ultra-specific primers and probe for RNase P RNA. Based on bioinformatics analysis, the attachment sites of the CDC-approved primers and probe for endogenous internal control of RNase P are located on the first exon of this gene. In addition to identifying the target gene sequence, these primers and probe also non-specifically detect similar sequences on the genomic DNA. Conclusions: The present study showed that the use of specific primers and probes introduced by CDC for SARS-CoV-2 and influenza virus may cause false results due to non-specific binding to the genomic DNA. Therefore, choosing the right internal control for RNase P RNA can be useful in achieving very accurate results.

Importance of SARA-COV-2 serological tests in diagnosis, vaccine productin and treatment

SARS-COV-2, the latest member of Coronaviridea family as the cause of the recent global epidemic, has inflicted sever damages in different fields on most governments. Appropriate strategies such as identifying infected individuals and isolating them, trying to produce an effective vaccine, finding efficient treatment and making correct decisions in the social, economic and health fields are the current priorities of the world. Simultaneous serological tests with molecular tests have led to a reduction in false-negative cases of these tests, which leads to the timely isolation of more patients and less spread of the disease. Evaluation of humoral immune responses by serological tests contribute to effective vaccine production. Serological tests can also be effective in optimizing plasma therapy in patients with SARS-COV-2. Finaly, performing inexpensive serological tests in high-volume caused more correct epidemiological information that leads to accurate decisions at the community level. Due to the effect of serological tests in various aspects, the use of them is effective in SARS-COV-2 epidemic management.

Evaluation of mutation spread in the SARS-CoV2 genome

The Coronaviridae family includes viruses that are considered the causative agents of respiratory infections, and among human RNA viruses have the largest genome. Coronaviruses undergo elusive genetic changes through mutation during replication. A gene mutation is a permanent alteration in the nucleic acid sequence;if it occurs in large numbers, it causes changes in the biological features of a species. Fundamentally, viruses adapt to the human body during replication. Several studies have shown that most mutations do not have much effect on pathogenicity. Sequence diversity in new coronaviruses is very low. However, antigen drift has been observed among some coronaviruses. Most coronavirus mutations occur intermittently in Iran and other countries and have little effect on the pathogenicity of the virus but have increased its rate of transmission. In mutated viruses, deletion of nucleotide sequences has been observed relatively in some reading frames extensively. Studies have shown that the host protein induced mutagenesis through interaction via viral proteins. The most important mutation in SARS-CoV2 compared to the original Wuhan virus was the spike D614G mutation and the lineage of B.1.1.7, 20I/501Y.V1become the dominant and exhibit greater virusspread but did not associate with higher viral loads and morbidity.However, it may affect the effectiveness of the vaccines and mortalityrate.