Identification and Genetic Characterization of MERS-Related Coronavirus Isolated from Nathusius' Pipistrelle (Pipistrellus nathusii) near Zvenigorod (Moscow Region, Russia).

Being diverse and widely distributed globally, bats are a known reservoir of a series of emerging zoonotic viruses. We studied fecal viromes of twenty-six bats captured in 2015 in the Moscow Region and found 13 of 26 (50%) samples to be coronavirus positive. Of P. nathusii (the Nathusius' pipistrelle), 3 of 6 samples were carriers of a novel MERS-related betacoronavirus. We sequenced and assembled the complete genome of this betacoronavirus and named it MOW-BatCoV strain 15-22. Whole genome phylogenetic analysis suggests that MOW-BatCoV/15-22 falls into a distinct subclade closely related to human and camel MERS-CoV. Unexpectedly, the phylogenetic analysis of the novel MOW-BatCoV/15-22 spike gene showed the closest similarity to CoVs from Erinaceus europaeus (European hedgehog). We suppose MOW-BatCoV could have arisen as a result of recombination between ancestral viruses of bats and hedgehogs. Molecular docking analysis of MOW-BatCoV/15-22 spike glycoprotein binding to DPP4 receptors of different mammals predicted the highest binding ability with DPP4 of the Myotis brandtii bat (docking score -320.15) and the E. europaeus (docking score -294.51). Hedgehogs are widely kept as pets and are commonly found in areas of human habitation. As this novel bat-CoV is likely capable of infecting hedgehogs, we suggest hedgehogs can act as intermediate hosts between bats and humans for other bat-CoVs.

Integrated Jingmenvirus Polymerase Gene in Ixodes ricinus Genome.

Members of the jingmenviruses group have been found in arthropods and mammals on all continents except Australia and Antarctica. Two viruses of this group were isolated from patients with fever after a tick bite. Using a nested RT-PCR assay targeting a jingmenvirus polymerase gene fragment, we screened ticks collected in seven regions of Russia and found that the abundant jingmenvirus-positive were of Ixodes ricinus species, with the prevalence ranging from 19.8% to 34.3%. In all cases, DNase/RNase treatment suggested that the detected molecule was DNA and subsequent next generation sequencing (NGS) proved that the viral polymerase gene was integrated in the I. ricinus genome. The copy number of the integrated polymerase gene was quantified by qPCR relative to the ITS2 gene and estimated as 1.32 copies per cell. At least three different genetic variants of the integrated polymerase gene were found in the territory of Russia. Phylogenetic analysis of the integrated jingmenvirus polymerase gene showed the highest similarity with the sequence of the correspondent gene obtained in Serbia from I. ricinus.

Mobile Colistin Resistance Genetic Determinants of Non-Typhoid Salmonella enterica Isolates from Russia.

Polymyxin resistance, determined by mcr genes located on plasmid DNA, currently poses a high epidemiological threat. Non-typhoid Salmonella (NTS) are one of the key pathogens causing diarrheal diseases. Here, we report the isolation and whole genome sequencing of multidrug colistin-resistant/susceptible isolates of non-typhoid Salmonella enterica serovars carrying mcr genes. Non-typhoid strains of Salmonella enterica subsp. enterica were isolated during microbiological monitoring of the environment, food, and diarrheal disease patients between 2018 and 2020 in Russia (n = 586). mcr-1 genes were detected using a previously developed qPCR assay, and whole genome sequencing of mcr positive isolates was performed by both short-read (Illumina) and long-read (Oxford Nanopore) approaches. Three colistin-resistant isolates, including two isolates of S. Enteritidis and one isolate of S. Bovismorbificans, carried the mcr-1.1 gene located on IncX4 and IncI2 conjugative plasmids, respectively. The phenotypically colistin-susceptible isolate of S. Typhimurium carried a mcr-9 gene on plasmid IncHI2. In conclusion, we present the first three cases of mcr gene-carrying NTS isolates detected in Russia with both outbreak and sporadic epidemiological backgrounds.

Utilizing the VirIdAl Pipeline to Search for Viruses in the Metagenomic Data of Bat Samples.

According to various estimates, only a small percentage of existing viruses have been discovered, naturally much less being represented in the genomic databases. High-throughput sequencing technologies develop rapidly, empowering large-scale screening of various biological samples for the presence of pathogen-associated nucleotide sequences, but many organisms are yet to be attributed specific loci for identification. This problem particularly impedes viral screening, due to vast heterogeneity in viral genomes. In this paper, we present a new bioinformatic pipeline, VirIdAl, for detecting and identifying viral pathogens in sequencing data. We also demonstrate the utility of the new software by applying it to viral screening of the feces of bats collected in the Moscow region, which revealed a significant variety of viruses associated with bats, insects, plants, and protozoa. The presence of alpha and beta coronavirus reads, including the MERS-like bat virus, deserves a special mention, as it once again indicates that bats are indeed reservoirs for many viral pathogens. In addition, it was shown that alignment-based methods were unable to identify the taxon for a large proportion of reads, and we additionally applied other approaches, showing that they can further reveal the presence of viral agents in sequencing data. However, the incompleteness of viral databases remains a significant problem in the studies of viral diversity, and therefore necessitates the use of combined approaches, including those based on machine learning methods.

Case report: change of dominant strain during dual SARS-CoV-2 infection.

BACKGROUND: The dual infection with SARS-CoV-2 is poorly described and is currently under discussion. We present a study of two strains of SARS-CoV-2 detected in the same patient during the same disease presentation. CASE PRESENTATION: A patient in their 90 s was hospitalised with fever. Oropharyngeal swab obtained on the next day (sample 1) tested positive for SARS-CoV-2. Five days later, the patient was transferred to the ICU (intensive care unit) of the hospital specialising in the treatment of COVID-19 patients, where the patient's condition progressively worsened and continuous oxygen insufflation was required. Repeated oropharyngeal swab (sample 2), which was taken eight days after the first one, also tested positive for SARS-CoV-2. After 5 days of ICU treatment, the patient died. The cause of death was a coronavirus infection, which progressed unfavourably due to premorbid status. We have performed sequencing of full SARS-CoV-2 genomes from oropharyngeal swabs obtained eight days apart. Genomic analysis revealed the presence of two genetically distant SARS-CoV-2 strains in both swabs. Detected strains belong to different phylogenetic clades (GH and GR) and differ in seven nucleotide positions. The relative abundance of strains was 70% (GH) and 30% (GR) in the first swab, and 3% (GH) and 97% (GR) in the second swab. CONCLUSIONS: Our findings suggest that the patient was infected by two genetically distinct SARS-CoV-2 strains at the same time. One of the possible explanations is that the second infection was hospital-acquired. Change of the dominant strain ratio during disease manifestation could be explained by the advantage or higher virulence of the GR clade strain.