Macromolecular crowding links ribosomal protein gene dosage to growth rate in Vibrio cholerae.

BACKGROUND: In fast-growing bacteria, the genomic location of ribosomal protein (RP) genes is biased towards the replication origin (oriC). This trait allows optimizing their expression during exponential phase since oriC neighboring regions are in higher dose due to multifork replication. Relocation of s10-spc-α locus (S10), which codes for most of the RP, to ectopic genomic positions shows that its relative distance to the oriC correlates to a reduction on its dosage, its expression, and bacterial growth rate. However, a mechanism linking S10 dosage to cell physiology has still not been determined. RESULTS: We hypothesized that S10 dosage perturbations impact protein synthesis capacity. Strikingly, we observed that in Vibrio cholerae, protein production capacity was independent of S10 position. Deep sequencing revealed that S10 relocation altered chromosomal replication dynamics and genome-wide transcription. Such changes increased as a function of oriC-S10 distance. Since RP constitutes a large proportion of cell mass, lower S10 dosage could lead to changes in macromolecular crowding, impacting cell physiology. Accordingly, cytoplasm fluidity was higher in mutants where S10 is most distant from oriC. In hyperosmotic conditions, when crowding differences are minimized, the growth rate and replication dynamics were highly alleviated in these strains. CONCLUSIONS: The genomic location of RP genes ensures its optimal dosage. However, besides of its essential function in translation, their genomic position sustains an optimal macromolecular crowding essential for maximizing growth. Hence, this could be another mechanism coordinating DNA replication to bacterial growth.

Scavengers as Prospective Sentinels of Viral Diversity: the Snowy Sheathbill Virome as a Potential Tool for Monitoring Virus Circulation, Lessons from Two Antarctic Expeditions.

Antarctica is a unique environment due to its extreme meteorological and geological conditions. In addition to this, its relative isolation from human influences has kept it undisturbed. This renders our limited understanding of its fauna and its associated microbial and viral communities a relevant knowledge gap to fill. This includes members of the order Charadriiformes such as snowy sheathbills. They are opportunistic predator/scavenger birds distributed on Antarctic and sub-Antarctic islands that are in frequent contact with other bird and mammal species. This makes them an interesting species for surveillance studies due to their high potential for the acquisition and transport of viruses. In this study, we performed whole-virome and targeted viral surveillance for coronaviruses, paramyxoviruses, and influenza viruses in snowy sheathbills from two locations, the Antarctic Peninsula and South Shetland. Our results suggest the potential role of this species as a sentinel for this region. We highlight the discovery of two human viruses, a member of the genus Sapovirus GII and a gammaherpesvirus, and a virus previously described in marine mammals. Here, we provide insight into a complex ecological picture. These data highlight the surveillance opportunities provided by Antarctic scavenger birds. IMPORTANCE This article describes whole-virome and targeted viral surveillance for coronaviruses, paramyxoviruses, and influenza viruses in snowy sheathbills from the Antarctic Peninsula and South Shetland. Our results suggest an important role of this species as a sentinel for this region. This species' RNA virome showcased a diversity of viruses likely tied to its interactions with assorted Antarctic fauna. We highlight the discovery of two viruses of likely human origin, one with an intestinal impact and another with oncogenic potential. Analysis of this data set detected a variety of viruses tied to various sources (from crustaceans to nonhuman mammals), depicting a complex viral landscape for this scavenger species.

The Myotis chiloensis Guano Virome: Viral Nucleic Acid Enrichments for High-Resolution Virome Elucidation and Full Alphacoronavirus Genome Assembly.

Bats are widespread mammals of the order Chiroptera. They are key for ecosystem functioning, participating in crucial processes. Their unique ability amongst mammals to fly long distances, their frequently large population sizes, and their longevity favor infectious agent persistence and spread. This includes a large variety of viruses, encompassing many important zoonotic ones that cause severe diseases in humans and domestic animals. Despite this, the understanding of the viral ecological diversity residing in bat populations remains unclear, which complicates the determination of the origins of zoonotic viruses. To gain knowledge on the viral community of a widely distributed insectivorous bat species, we characterized the guano virome of a native Chilean bat species (Myotis chiloensis (Waterhouse, 1840)). By applying a novel enrichment strategy, we were able to secure a consequent percentage of viral reads, providing unprecedented resolution for a bat virome. This in turn enabled us to identify and assemble a new bat alphacoronavirus from Chilean bats closely related to PEDV, an important viral pathogen with high mortality rates in suckling piglets. This study highlights the importance of applying and improving high-resolution virome studies in this vital order to ultimately enhance epidemiological surveillance for potentially zoonotic pathogens.

RadD Contributes to R-Loop Avoidance in Sub-MIC Tobramycin.

We have previously identified Vibrio cholerae mutants in which the stress response to subinhibitory concentrations of aminoglycoside is altered. One gene identified, VC1636, encodes a putative DNA/RNA helicase, recently named RadD in Escherichia coli Here we combined extensive genetic characterization and high-throughput approaches in order to identify partners and molecular mechanisms involving RadD. We show that double-strand DNA breaks (DSBs) are formed upon subinhibitory tobramycin treatment in the absence of radD and recBCD and that formation of these DSBs can be overcome by RNase H1 overexpression. Loss of RNase H1, or of the transcription-translation coupling factor EF-P, is lethal in the radD deletion mutant. We propose that R-loops are formed upon sublethal aminoglycoside treatment, leading to the formation of DSBs that can be repaired by the RecBCD homologous recombination pathway, and that RadD counteracts such R-loop accumulation. We discuss how R-loops that can occur upon translation-transcription uncoupling could be the link between tobramycin treatment and DNA break formation.IMPORTANCE Bacteria frequently encounter low concentrations of antibiotics. Active antibiotics are commonly detected in soil and water at concentrations much below lethal concentration. Although sub-MICs of antibiotics do not kill bacteria, they can have a major impact on bacterial populations by contributing to the development of antibiotic resistance through mutations in originally sensitive bacteria or acquisition of DNA from resistant bacteria. It was shown that concentrations as low as 100-fold below the MIC can actually lead to the selection of antibiotic-resistant cells. We seek to understand how bacterial cells react to such antibiotic concentrations using E. coli, the Gram-negative bacterial paradigm, and V. cholerae, the causative agent of cholera. Our findings shed light on the processes triggered at the DNA level by antibiotics targeting translation, how damage occurs, and what the bacterial strategies are to respond to such DNA damage.