SummaryCommon approaches to antibiotic discovery include small-molecule screens for growth inhibition in target pathogens and screens for inhibitors of purified enzymes. These approaches have a shared intent of seeking to directly target a vital Achilles heel in a pathogen of interest. Here, we report the first screen against a sporulation pathway in a non-pathogenic bacterium as a means of discovering novel antibiotics—this effort has resulted in two important discoveries. First, we show that the sporulation program of Streptomyces venezuelae is exquisitely sensitive to numerous forms of DNA damage. Second, we have identified a DNA gyrase inhibitor. This molecule, EN-7, is active against pathogenic species that are resistant to ciprofloxacin and other clinically important antibiotics. We suggest that this strategy could be applied to other morphogenetic pathways in prokaryotes or eukaryotes as a means of identifying novel chemical matter having scientific and clinical utility.

Small molecule disruption of the bacterial membrane is both a challenge and interest for drug development. While some avoid membrane activity due to toxicity issues, others are interested in leveraging the effects for new treatments. Existing assays are available for measuring disruption of membrane potential or membrane permeability, two key characteristics of the bacterial membrane, however they are limited in their ability to distinguish between these properties. Here, we demonstrate a high throughput assay for detection and characterization of membrane active compounds. The assay distinguishes the effect of small molecules on either the membrane potential or membrane permeability using the fluorescent dyes TO-PRO-3 iodide and DiOC(3) without the need for secondary assays. We then applied this assay to a library of 3520 synthetic molecules previously shown to inhibit growth of in order to determine the frequency of membrane activity within such a biologically active library. From the library, we found 249 compounds that demonstrated significant membrane activity, suggesting that synthetic libraries of this kind do not contain a plurality of membrane active molecules.

Urban stormwater runoff is an important pathway for the introduction of microplastics and other anthropogenic pollutants into aquatic environments. Highly variable concentrations of microplastics have been reported globally in runoff, but knowledge of key factors within urban environments contributing to this variability remains limited. Furthermore, few studies to date have quantitatively assessed the release of microplastics to receiving waters via runoff. The objectives of this study were to assess the influence of different catchment characteristics on the type and amount of microplastics in runoff and to provide an estimate of the quantity of microplastics discharged during rain events. Stormwater samples were collected during both dry periods (baseflow) and rain events from 15 locations throughout the city of Calgary, Canada’s fourth largest city. These catchments ranged in size and contained different types of predominant land use. Microplastics were found in all samples, with total concentrations ranging from 0.7 to 200.4 pcs/L (mean = 31.9 pcs/L). Fibers were the most prevalent morphology identified (47.7 ± 33.0%), and the greatest percentage of microplastics were found in the 125–250 µm size range (26.6 ± 22.9%) followed by the 37–125 µm size range (24.0 ± 22.3%). Particles were predominantly black (33.5 ± 33.8%), transparent (22.6 ± 31.3%), or blue (16.0 ± 21.6%). Total concentrations, dominant morphologies, and size distributions of microplastics differed between rain events and baseflow, with smaller particles and higher concentrations being found during rain events. Concentrations did not differ significantly amongst catchments with different land use types, but concentrations were positively correlated with maximum runoff flow rate, catchment size, and the percentage of impervious surface area within a catchment. Combining microplastic concentrations with hydrograph data collected during rain events, we estimated that individual outfalls discharged between 1.9 million to 9.6 billion microplastics to receiving waters per rain event. These results provide further evidence that urban stormwater runoff is a significant pathway for the introduction of microplastics into aquatic environments and suggests that mitigation strategies for microplastic pollution should focus on larger urbanized catchments.

Bacteria exhibit complex responses to biologically active small molecules. These responses include reductions in transcriptional and translational efficiency, alterations in metabolic flux, and in some cases, dramatic changes in growth and morphology. Here, we describe Min-1, a novel small molecule that inhibits growth of Gram-positive bacteria by targeting the cell envelope. Subinhibitory levels of Min-1 inhibits sporulation in Streptomyces venezuelae and reduces growth rate and cell length in Bacillus subtilis. The effect of Min-1 on B. subtilis cell length is significant at high growth rates sustained by nutrient rich media but drops off when growth rate is reduced during growth on less energy rich carbon sources. In each medium, Min-1 has no impact on the proportion of cells containing FtsZ-rings, suggesting that Min-1 reduces the mass at which FtsZ assembly is initiated. The effect of Min-1 on size is independent of UDP-glucose, which couples cell division to carbon availability, and the alarmone ppGpp, which reduces cell size via its impact on fatty acid synthesis. Min-1 activates the LiaRS stress response, which is sensitive to disruptions in the lipid II cycle and the cell membrane, and also compromises cell membrane integrity. Therefore, this novel synthetic molecule inhibits growth at high concentrations and induces a short-cell phenotype at sub-inhibitory concentrations that is independent of known systems that influence cell length, highlighting the complex interactions between small molecules and cell morphology.