Draft genome sequencing of Naganishia species isolated from the polar environments.

The draft genomes of five Naganishia strains were sequenced using MinION and annotated using Funannotate pipeline. Phylogenetic and genomic analyses were performed to provide their genetic relationships, diversity, and potential functional capabilities. This approach will aid in understanding their potential to survive under microgravity and their resilience to extreme environments.

Cave Thiovulum (Candidatus Thiovulum stygium) differs metabolically and genomically from marine species.

Thiovulum spp. (Campylobacterota) are large sulfur bacteria that form veil-like structures in aquatic environments. The sulfidic Movile Cave (Romania), sealed from the atmosphere for ~5 million years, has several aqueous chambers, some with low atmospheric O2 (~7%). The cave's surface-water microbial community is dominated by bacteria we identified as Thiovulum. We show that this strain, and others from subsurface environments, are phylogenetically distinct from marine Thiovulum. We assembled a closed genome of the Movile strain and confirmed its metabolism using RNAseq. We compared the genome of this strain and one we assembled from public data from the sulfidic Frasassi caves to four marine genomes, including Candidatus Thiovulum karukerense and Ca. T. imperiosus, whose genomes we sequenced. Despite great spatial and temporal separation, the genomes of the Movile and Frasassi Thiovulum were highly similar, differing greatly from the very diverse marine strains. We concluded that cave Thiovulum represent a new species, named here Candidatus Thiovulum stygium. Based on their genomes, cave Thiovulum can switch between aerobic and anaerobic sulfide oxidation using O2 and NO3- as electron acceptors, the latter likely via dissimilatory nitrate reduction to ammonia. Thus, Thiovulum is likely important to both S and N cycles in sulfidic caves. Electron microscopy analysis suggests that at least some of the short peritrichous structures typical of Thiovulum are type IV pili, for which genes were found in all strains. These pili may play a role in veil formation, by connecting adjacent cells, and in the motility of these exceptionally fast swimmers.

Complete Genome Sequence of Klebsiella quasipneumoniae subsp. similipneumoniae Strain IF3SW-P1, Isolated from the International Space Station.

The 5.2-Mb circular genome of Klebsiella quasipneumoniae subsp. similipneumoniae strain IF3SW-P1, isolated from the International Space Station, was sequenced using Oxford Nanopore Technologies. The genome lacks a megaplasmid typical of hypervirulent and multidrug-resistant Klebsiella strains but does contain a chromosomally encoded OqxAB efflux pump associated with carbapenem resistance.

Molecular Characterization of Increased Amplicon Lengths in SARS-CoV-2 Reverse Transcription Loop-Mediated Isothermal Amplification Assays.

Loop-mediated isothermal amplification (LAMP) is a power tool for the amplification of specific RNA and DNA targets. Much like PCR, LAMP requires primers that surround a target amplification region and generates exponential product through a unique highly specific daisy-chain single-temperature amplification reaction. However, until recently, attempts to amplify targets of greater than 200 base pairs (bp) have been mostly unsuccessful and limited to short amplicon targets of less than 150 bp. Although short amplicons have the benefit of a rapid detection (<40 min), they do not allow for the prediction of RNA integrity based on RNA length and possible intactness. In this study, 8 primer sets were developed using 2 LAMP primer-specific software packages against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid gene with insert lengths ranging from 262 to 945 bp in order to amplify and infer the integrity of viral RNA. Because these amplification lengths are greater than the current methods that use an insert length of 130 or less, they require a longer incubation, modified primer and temperature strategies, and the addition of specific adjuncts to prevent nonspecific amplification. This proof of concept study resulted in successful reverse transcription LAMP reactions for amplicon targets of 262, 687, 693, and 945 bp using a clinical nasopharyngeal NP sample, purified SARS-CoV-2 RNA, and crude lysate containing inactivated virus.