Occurrences of similar viral diversity in campus wastewater and reclaimed water of a university dormitory.

Water reuse from wastewater sources still remain some critical safety concerns associated with treacherous contaminants like pathogenic viruses. In this study, viral diversities in campus wastewater (CWW) and its reclaimed water (RCW) recycled for toilet flushing and garden irrigation of a university dormitory were assessed using metagenomic sequencing for acquisition of more background data. Results suggested majority (>80%) of gene sequences within assembled contigs predicted by open reading frame (ORF) finder were no-hit yet believed to be novel/unrevealed viral genomic information whereas hits matched bacteriophages (i.e., mainly Myoviridae, Podoviridae, and Siphoviridae families) were predominant in both CWW and RCW samples. Moreover, few pathogenic viruses (<1%) related to infections of human skin (e.g., Molluscum contagiosum virus, MCV), digestion system (e.g., hepatitis C virus, HCV), and gastrointestinal tract (e.g., human norovirus, HuNoV) were also noticed raising safety concerns about application of reclaimed waters. Low-affinity interactions of particular viral exterior proteins (e.g., envelope glycoproteins or spike proteins) for disinfectant ligand (e.g., chlorite) elucidated treatment limitations of current sewage processing systems even with membrane bioreactor and disinfectant contactor. Revolutionary disinfection approaches together with routine monitoring and new regulations are prerequisite to secure pathogen-correlated water quality for safer reuse of reclaimed waters.

Variations of SARS-CoV-2 in the Iranian population and candidate putative drug-like compounds to inhibit the mutated proteins.

The first cases of the novel coronavirus, SARS-CoV-2, were detected in December 2019 in Wuhan, China. Nucleotide substitutions and mutations in the SARS-CoV-2 sequence can result in the evolution of the virus and its rapid spread across the world. Therefore, understanding genetic variants of SARS-CoV-2 and targeting the conserved elements responsible for viral replication have great benefits for detecting its infection sources and diagnosing and treating COVID-19. In this study, we used the SARS-CoV-2 sequence isolated from a 59-year-old man in Ardabil, Iran, in April 2020 and sequenced using Oxford Nanopore technology. A meta-analysis comparing the sequence under study with other sequences from Iran indicated long nucleotide insertions/deletions (indels) that code for NSP15, the NSP14-NSP10 complex, open reading frame ORF9b, and ORF1ab polyproteins. In addition, replicating the NSP8 protein in the study sequence is another topic that can affect viral replication. Then using the DNA structure of NSP8, NSP15, NSP14-NSP10 complex, and ORF1ab as a genetic target can help find drug-like compounds for COVID-19. Potential drug-like compounds reported in this study for their mechanism of action and interactions with SARS-CoV-2 genes using drug repurposing are resveratrol, erythromycin, chloramphenicol, indomethacin, ciclesonide, and PDE4 inhibitor. Ciclesonide appears to show the best results when docked with chosen viral proteins. Therefore, different proteins isolated from nucleotide mutations in the virus sequence can indicate distinct inducers for antibodies and are important in vaccine design.

Discovering SARS-CoV-2 genes and mutations adapted for humans in 2594 genomes

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a positive-sense single-stranded virus approximately 30 kb in length, is the cause of the ongoing global life-threatening novel coronavirus disease-2019 (COVID-19) outbreak. Studies confirmed significant genome differences between SARS-CoV-2 and SARS-CoV, suggesting that the distinctions in pathogenicity and virulence might be related to genomic diversity. However, the relationship between genomic differences and SARS-CoV-2 fitness has not been fully explained, especially for open reading frame (ORF)-encoded accessory proteins. RNA viruses have a high mutation rate, but how SARS-CoV-2 mutations accelerate host adaptation is not clear. This study shows that the host-genome similarity (HGS) of SARS-CoV-2 is significantly higher than that of SARS-CoV, especially in the ORF6 and ORFS genes that encode proteins antagonizing innate immunity in vivo. A power law relationship was discovered between the HGS of ORF3b, ORF6, and N and the expression of interferon (IFN)-sensitive response element (ISRE)-containing promoters. This finding implies that the increase in HGS in the SARS-CoV-2 genome may further inhibit FN I synthesis and cause delayed host innate immunity. An ORF1ab mutation, 1081SG>T, which occurred in virus populations with high HGS but rarely in low-HGS populations, was identified in 2594 genomes with geolocations of China, the USA and Europe. The genomic mutation caused the amino acid mutation M37F in the transmembrane protein nsp6. The results suggest that the ORF6 and ORFS genes and the residue mutation M37F may play important roles in SARS-CoV-2 adaptation to humans. However, the underlying basis by which the mutations mediate adaptation to humans is still unknown. The findings demonstrate that HGS analysis is a reliable way to identify important genes and mutations in adaptive strains, which may help in the search for potential targets for pharmaceutical agents. © 2021 IEEE.

Variable levels of spike and ORF1ab RNA in post-mortem lung samples of SARS-CoV-2-positive subjects: comparison between ISH and RT-PCR.

Post-mortem examination plays a pivotal role in understanding the pathobiology of the SARS-CoV-2; thus, the optimization of virus detection on the post-mortem formalin-fixed paraffin-embedded (FFPE) tissue is needed. Different techniques are available for the identification of the SARS-CoV-2, including reverse transcription polymerase chain reaction (RT-PCR), immunohistochemistry (IHC), in situ hybridization (ISH), and electron microscopy. The main goal of this study is to compare ISH versus RT-PCR to detect SARS-CoV-2 on post-mortem lung samples of positive deceased subjects. A total of 27 samples were analyzed by RT-PCR targeting different viral RNA sequences of SARS-CoV-2, including envelope (E), nucleocapsid (N), spike (S), and open reading frame (ORF1ab) genes and ISH targeting S and Orf1ab. All 27 cases showed the N gene amplification, 22 out of 27 the E gene amplification, 26 out of 27 the S gene amplification, and only 6 the ORF1ab gene amplification. The S ISH was positive only in 12 out of 26 cases positive by RT-PCR. The S ISH positive cases with strong and diffuse staining showed a correlation with low values of the number of the amplification cycles by S RT-PCR suggesting that ISH is a sensitive assay mainly in cases carrying high levels of S RNA. In conclusion, our findings demonstrated that ISH assay has lower sensitivity to detect SARS-CoV-2 in FFPE compared to RT-PCR; however, it is able to localize the virus in the cellular context since it preserves the morphology.

Genetic characterization of structural and open reading Fram-8 proteins of SARS-CoV-2 isolates from different countries.

Since December 2019, a severe pandemic of pneumonia, COVID-19 associated with a novel coronavirus (SARS-CoV-2), have emerged in Wuhan, China and spreading throughout the world. As RNA viruses have a high mutation rate therefore we wanted to identify whether this virus is also prone to mutations. For this reason we selected four major structural (Spike protein (S), Envelope protein (E), Membrane glycoprotein (M), Nucleocapsid phosphoprotein (N)) and ORF8 protein of 100 different SARS-CoV-2 isolates of fifteen countries from NCBI database and compared these to the reference sequence, Wuhan NC_045512.2, which was the first isolate of SARS-CoV-2 that was sequenced. By multiple sequence alignment of amino acids, we observed substitutions and deletion in S protein at 13 different sites in the isolates of five countries (China, USA, Finland, India and Australia) as compared to the reference sequence. Similarly, alignment of N protein revealed substitutions at three different sites in isolates of China, Spain and Japan. M protein exhibits substitution only in one isolates from USA, however, no mutation was observed in E protein of any isolate. Interestingly, in ORF8 substitution of Leucine, a nonpolar to Serine a polar amino acid at same position (aa84 L to S) in 23 isolates of five countries i.e. China, USA, Spain, Taiwan and India were observed, which may affect the conformation of peptides. Thus, we observed several mutations in the isolates thereafter the first sequencing of SARS-CoV-2 isolate, NC_045512.2, which suggested that this virus might be a threat to the whole world and therefore further studies are needed to characterize how these mutations in different proteins affect the functionality and pathogenesis of SARS-CoV-2.