- Volume 2, Issue 10, 2020

High-throughput sequencing has become a standard tool for transcriptome analysis. The depletion of overrepresented RNA species from sequencing libraries plays a key role in establishing potent and cost-efficient RNA-seq routines. Commercially available kits are known to obtain good results for the reduction of ribosomal RNA (rRNA). However, we found that the transfer-messenger RNA (tmRNA) was frequently highly abundant in rRNA-depleted samples of Pseudomonas aeruginosa , consuming up to 25 % of the obtained reads. The tmRNA fraction was particularly high in samples taken from stationary cultures. This suggests that overrepresentation of this RNA species reduces the mRNA fraction when cells are grown under challenging conditions. Here, we present an RNase-H-based depletion protocol that targets the tmRNA in addition to ribosomal RNAs. We were able to increase the mRNA fraction to 93–99% and therefore outperform not only the commercially Ribo-off kit (Vazyme) operating by the same principle but also the formerly widely used Ribo-Zero kit (Illumina). Maximizing the read share of scientifically interesting RNA species enhances the discriminatory potential of next-generation RNA-seq experiments and, therefore, can contribute to a better understanding of the transcriptomic landscape of bacterial pathogens and their used mechanisms in host infection.

Reports on the commensal organism and opportunistic pathogen Staphylococcus schleiferi have largely considered isolates from humans and companion dogs. Two subspecies are recognized: the coagulase-negative S. schleiferi ssp. schleiferi, typically seen in humans, and the coagulase-positive S. schleiferi ssp. coagulans, typically seen in dogs. In this study, we report the isolation, genome sequencing and comparative genomics of three S. schleiferi ssp. coagulans isolates from mouth samples from two species of healthy, free-living Antarctic seals, southern elephant seals (Mirounga leonina) and Antarctic fur seals (Arctocephalus gazella), in the South Orkney Islands, Antarctica, and three isolates from post-mortem samples from grey seals (Halichoerus grypus) in Scotland, UK. This is the first report of S. schleiferi ssp. coagulans isolation from Antarctic fur seal and grey seal. The Antarctic fur seal represents the first isolation of S. schleiferi ssp. coagulans from the family Otariidae, while the grey seal represents the first isolation from a pinniped in the Northern Hemisphere. We compare seal, dog and human isolates from both S. schleiferi subspecies in the first genome-based phylogenetic analysis of the species.

We present an unusual case of a previously healthy 74-year-old man who presented with diffuse weakness, severe myalgias, petechial palmar rash and hypotension, but without fever, altered mental status, nuchal rigidity or headache, who was ultimately found through PCR testing to have meningococcal meningitis.

With increasing resistance to currently used antibiotics, antibiotic combinations are being resorted to. The present study deals with five children with complicated urinary tract infection (UTI) whose urine cultures grew multidrug-resistant (MDR) organisms. In all of these five cases, MDR organisms were the causative agents for UTI and the currently available antibiotics, including colistin, were ineffective, although the organisms were sensitive in vitro. In all of these cases, the isolates reverted to being susceptible to the quinolones and cephalosporins tested, namely ceftriaxone and ceftazidime. All were treated using a combination of fosfomycin with other antibiotics, since it has no interference with other classes of antibiotics. Our observations suggest that the use of a combination of fosfomycin with either a carbapenem or an aminoglycoside in a clinical setting would be a reasonable choice to treat UTIs caused by MDR organisms, especially in complicated cases that require chronic therapy.