The emergence of drug resistant pathogens is a serious public health problem. It is a long-standing goal to predict rates of resistance evolution and design optimal treatment strategies accordingly. To this end, it is crucial to reveal the underlying causes of drug-specific differences in the evolutionary dynamics leading to resistance. However, it remains largely unknown why the rates of resistance evolution via spontaneous mutations and the diversity of mutational paths vary substantially between drugs. Here we comprehensively quantify the distribution of fitness effects (DFE) of mutations, a key determinant of evolutionary dynamics, in the presence of eight antibiotics representing the main modes of action. Using precise high-throughput fitness measurements for genome-wide Escherichia coli gene deletion strains, we find that the width of the DFE varies dramatically between antibiotics and, contrary to conventional wisdom, for some drugs the DFE width is lower than in the absence of stress. We show that this previously underappreciated divergence in DFE width among antibiotics is largely caused by their distinct drug-specific dose-response characteristics. Unlike the DFE, the magnitude of the changes in tolerated drug concentration resulting from genome-wide mutations is similar for most drugs but exceptionally small for the antibiotic nitrofurantoin, i.e., mutations generally have considerably smaller resistance effects for nitrofurantoin than for other drugs. A population genetics model predicts that resistance evolution for drugs with this property is severely limited and confined to reproducible mutational paths. We tested this prediction in laboratory evolution experiments using the “morbidostat”, a device for evolving bacteria in well-controlled drug environments. Nitrofurantoin resistance indeed evolved extremely slowly via reproducible mutations—an almost paradoxical behavior since this drug causes DNA damage and increases the mutation rate. Overall, we identified novel quantitative characteristics of the evolutionary landscape that provide the conceptual foundation for predicting the dynamics of drug resistance evolution. When bacteria acquire drug resistance through mutation of their genomes, it renders once-powerful treatments useless. For some antibiotics, bacteria develop resistance rapidly, while for other antibiotics this process can be very slow. The causes of such differences in evolutionary dynamics between drugs remain poorly understood. Determining how these mutations that confer drug resistance affect bacterial fitness is a crucial requirement for developing predictions about evolution. In this study, we systematically measured the fitness effects of thousands of mutations that mimic all viable spontaneous gene disruptions in the bacterium Escherichia coli, when exposed to eight different antibiotics that represent the main modes of drug action. We find that the overall magnitude of these fitness effects varies enormously among antibiotics and show that this phenomenon is explained by the drug-specific response of bacteria to small changes in drug dose. We further show that, unlike the fitness effects—which vary greatly—the changes in tolerated drug concentration generated by resistance-conferring mutations are similar for most drugs, with the notable exception of an outlier drug called nitrofurantoin (which has an extremely small tolerated drug concentration range). Using a combination of mathematical modeling and evolution experiments, we propose new quantitative measures that enable partial predictions about how quickly resistance to a given drug will evolve and whether it will reproducibly occur by exploiting the same mutations.

Fusarium oxysporum is a cross-kingdom fungal pathogen that infects plants and humans. Horizontally transferred lineage-specific (LS) chromosomes were reported to determine host-specific pathogenicity among phytopathogenic F. oxysporum. However, the existence and functional importance of LS chromosomes among human pathogenic isolates are unknown. Here we report four unique LS chromosomes in a human pathogenic strain NRRL 32931, isolated from a leukemia patient. These LS chromosomes were devoid of housekeeping genes, but were significantly enriched in genes encoding metal ion transporters and cation transporters. Homologs of NRRL 32931 LS genes, including a homolog of ceruloplasmin and the genes that contribute to the expansion of the alkaline pH-responsive transcription factor PacC/Rim1p, were also present in the genome of NRRL 47514, a strain associated with Fusarium keratitis outbreak. This study provides the first evidence, to our knowledge, for genomic compartmentalization in two human pathogenic fungal genomes and suggests an important role of LS chromosomes in niche adaptation.

Chili pepper (Capsicum) is known for its unique fruit pungency due to the presence of capsaicinoids. The evolutionary history of capsaicinoid biosynthesis and the mechanism of their tissue specificity remain obscure due to the lack of high-quality Capsicum genomes. Here, we report two telomere-to-telomere (T2T) gap-free genomes of C. annuum and its wild nonpungent relative C. rhomboideum to investigate the evolution of fruit pungency in chili peppers. We precisely delineate Capsicum centromeres, which lack high-copy tandem repeats but are extensively invaded by CRM retrotransposons. Through phylogenomic analyses, we estimate the evolutionary timing of capsaicinoid biosynthesis. We reveal disrupted coding and regulatory regions of key biosynthesis genes in nonpungent species. We also find conserved placenta-specific accessible chromatin regions, which likely allow for tissue-specific biosynthetic gene coregulation and capsaicinoid accumulation. These T2T genomic resources will accelerate chili pepper genetic improvement and help to understand Capsicum genome evolution.

The lack of effective and well-tolerated therapies against antibiotic-resistant bacteria is a global public health problem leading to prolonged treatment and increased mortality. To improve the efficacy of existing antibiotic compounds, we introduce a new method for strategically inducing antibiotic hypersensitivity in pathogenic bacteria. Following the systematic verification that the AcrAB-TolC efflux system is one of the major determinants of the intrinsic antibiotic resistance levels in Escherichia coli, we have developed a short antisense oligomer designed to inhibit the expression of acrA and increase antibiotic susceptibility in E. coli. By employing this strategy, we can inhibit E. coli growth using 2- to 40-fold lower antibiotic doses, depending on the antibiotic compound utilized. The sensitizing effect of the antisense oligomer is highly specific to the targeted gene’s sequence, which is conserved in several bacterial genera, and the oligomer does not have any detectable toxicity against human cells. Finally, we demonstrate that antisense oligomers improve the efficacy of antibiotic combinations, allowing the combined use of even antagonistic antibiotic pairs that are typically not favored due to their reduced activities. Antibiotic resistance is a global health threat. While genome sequencing and genetic manipulation tools have elucidated many resistance mechanisms, these tools have not yet been developed into successful therapeutics. One tool with such potential are peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs), which are synthetic DNA/RNA mimics that function as antisense mRNA translation inhibitors. In this paper we use PPMOs to increase antibiotic susceptibility of bacteria. First, we identify the AcrAB-TolC efflux system as a major intrinsic resistance mechanism in E. coli. Then by targeting the mRNA of each component of this efflux system with PPMOs, we identify an acrA-PPMO as the most effective antisense molecule. Treatment of bacteria with acrA-PPMO resulted in a 2- to 40-fold increase in antibiotic efficacy, prevented translation of the AcrA protein, and inhibited efflux of antibiotic molecules without being cytotoxic to human cells. Finally, we demonstrate that acrA-PPMO is efficacious in several pathogenic bacterial genera and enhances activity of both synergistic and antagonistic antibiotic pairs when used together. This work establishes that PPMOs can potentially be used to treat infections caused by antibiotic resistant bacteria.

Fusarium oxysporum is a pathogenic fungus that infects hundreds of plant species. This paper reports the improved genome assembly of a reference strain, F. oxysporum f.