Publications

ABSTRACT Antibiotic therapy is commonly used in various medical scenarios, and some antibiotics will stay in the blood serum for days. Previous studies have demonstrated that the fluoroquinolone antibiotic norfloxacin exerts mutagenic effects on the whole genomes of target bacteria. However, whether and to what degree these effects compromise non-target eukaryotic genomes remains unclear. Here, we explored this using mutation accumulation experiments with the fission yeast Schizosaccharomyces pombe ATCC-16979, treated with or without norfloxacin at a dose comparable to the therapeutic concentration found in patient blood. Based on a de novo assembly and the mutation accumulation lines without treatment, ATCC-16979 demonstrates a strong mutation bias in the A/T direction (4.33). Furthermore, norfloxacin treatment did not significantly elevate the genomic mutation rate of the fission yeast, based on the analysis of 94 mutation accumulation lines run for ~169,000 cell divisions in total. Nucleotide excision repair was induced by the norfloxacin treatment and might help to counteract the possible mutagenic effects of norfloxacin, as revealed by RNAseq-based differential gene-expression analyses. Norfloxacin thus poses a negligible genotoxic threat to eukaryotic and potentially human genomes, bolstering the safe clinical use of this important antibiotic. IMPORTANCE This study addresses concerns about the potential mutagenic side effects of antibiotics, specifically norfloxacin, which is widely used in clinical settings. While previous research has shown that norfloxacin can cause mutations in bacteria, it was unclear whether it could also harm human or other eukaryotic genomes. By using the fission yeast Schizosaccharomyces pombe as a model, we found that norfloxacin treatment did not significantly increase the mutation rate in eukaryotic cells, possibly resulting from a cellular repair mechanism counteracting potential DNA damage. These findings provide reassurance that, at therapeutic levels, norfloxacin does not pose a significant genetic risk to eukaryotic organisms, supporting its continued safe use in medical treatments.

Abstract Bacteria and eukaryotes are involved in many types of interaction in nature, with important ecological consequences. However, the diversity, occurrence, and mechanisms of these interactions often are not fully known. The obligate bacterial endosymbionts of Paramecium provide their hosts with the ability to kill sensitive Paramecium strains through the production of R-bodies, highly insoluble coiled protein ribbons. R-bodies have been observed in a number of free-living bacteria, where their function is unknown. We have performed an exhaustive survey of genes coding for homologs of Reb proteins (R-body components) in complete bacterial genomes. We found that reb genes are much more widespread than previously thought, being present in representatives of major Proteobacterial subdivisions, including many free-living taxa, as well as taxa known to be involved in various kinds of interactions with eukaryotes, from mutualistic associations to pathogenicity. Reb proteins display very good conservation at the sequence level, suggesting that they may produce functional R-bodies. Phylogenomic analysis indicates that reb genes underwent a complex evolutionary history and allowed the identification of candidates potentially involved in R-body assembly, functioning, regulation, or toxicity. Our results strongly suggest that the ability to produce R-bodies is likely widespread in Proteobacteria. The potential involvement of R-bodies in as yet unexplored interactions with eukaryotes and the consequent ecological implications are discussed.

Abstract Mutations are the ultimate source of variation used for evolutionary adaptation, while also being predominantly deleterious and a source of genetic disorders. Understanding the rate of insertion-deletion mutations (indels) is essential to understanding evolutionary processes, especially in coding regions, where such mutations can disrupt production of essential proteins. Using direct estimates of indel rates from 14 phylogenetically diverse eukaryotic and bacterial species, along with measures of standing variation in such species, we obtain results that imply an inverse relationship of mutation rate and effective population size. These results, which corroborate earlier observations on the base-substitution mutation rate, appear most compatible with the hypothesis that natural selection reduces mutation rates per effective genome to the point at which the power of random genetic drift (approximated by the inverse of effective population size) becomes overwhelming. Given the substantial differences in DNA metabolism pathways that give rise to these two types of mutations, this consistency of results raises the possibility that refinement of other molecular and cellular traits may be inversely related to species-specific levels of random genetic drift.

ABSTRACT Population structure can be described by genotypic correlation coefficients between groups of individuals, the most basic of which are the pair-wise relatedness coefficients between any two individuals. There are nine pair-wise relatedness coefficients in the most general model, and we show that these can be reduced to seven coefficients for biallelic loci. Although all nine coefficients can be estimated from pedigrees, six coefficients have been beyond empirical reach. We provide a numerical optimization procedure that estimates them from population-genomic data. Simulations show that the procedure is nearly unbiased, even at 3× coverage, and errors in five of the seven coefficients are statistically uncorrelated. The remaining two coefficients have a negative correlation of errors, but their sum provides an unbiased assessment of the overall correlation of heterozygosity between two individuals. Application of these new methods to four populations of the freshwater crustacean Daphnia pulex reveal the occurrence of half-siblings in our samples, as well as a number of identical individuals that are likely obligately asexual clone-mates. Statistically significant negative estimates of these pair-wise relatedness coefficients, including inbreeding coefficents that were typically negative, underscore the difficulties that arise when interpreting genotypic correlations as estimations of the probability that alleles are identical by descent.

Abstract Whole-genome duplications (WGDs) have shaped the gene repertoire of many eukaryotic lineages. The redundancy created by WGDs typically results in a phase of massive gene loss. However, some WGD–derived paralogs are maintained over long evolutionary periods, and the relative contributions of different selective pressures to their maintenance are still debated. Previous studies have revealed a history of three successive WGDs in the lineage of the ciliate Paramecium tetraurelia and two of its sister species from the Paramecium aurelia complex. Here, we report the genome sequence and analysis of 10 additional P. aurelia species and 1 additional out group, revealing aspects of post-WGD evolution in 13 species sharing a common ancestral WGD. Contrary to the morphological radiation of vertebrates that putatively followed two WGD events, members of the cryptic P. aurelia complex have remained morphologically indistinguishable after hundreds of millions of years. Biases in gene retention compatible with dosage constraints appear to play a major role opposing post-WGD gene loss across all 13 species. In addition, post-WGD gene loss has been slower in Paramecium than in other species having experienced genome duplication, suggesting that the selective pressures against post-WGD gene loss are especially strong in Paramecium. A near complete lack of recent single-gene duplications in Paramecium provides additional evidence for strong selective pressures against gene dosage changes. This exceptional data set of 13 species sharing an ancestral WGD and 2 closely related out group species will be a useful resource for future studies on Paramecium as a major model organism in the evolutionary cell biology.

Mitochondrial DNA (mtDNA) variants are widely used in evolutionary genetics as markers for population history and to estimate divergence times among taxa. Inferences of species history are generally based on phylogenetic comparisons, which assume that molecular evolution is clock-like. Between-species comparisons have also been used to estimate the mutation rate, using sites that are thought to evolve neutrally. We directly estimated the mtDNA mutation rate by scanning the mitochondrial genome of Drosophila melanogaster lines that had undergone approximately 200 generations of spontaneous mutation accumulation (MA). We detected a total of 28 point mutations and eight insertion-deletion (indel) mutations, yielding an estimate for the single-nucleotide mutation rate of 6.2 x 10(-8) per site per fly generation. Most mutations were heteroplasmic within a line, and their frequency distribution suggests that the effective number of mitochondrial genomes transmitted per female per generation is about 30. We observed repeated occurrences of some indel mutations, suggesting that indel mutational hotspots are common. Among the point mutations, there is a large excess of G-->A mutations on the major strand (the sense strand for the majority of mitochondrial genes). These mutations tend to occur at nonsynonymous sites of protein-coding genes, and they are expected to be deleterious, so do not become fixed between species. The overall mtDNA mutation rate per base pair per fly generation in Drosophila is estimated to be about 10x higher than the nuclear mutation rate, but the mitochondrial major strand G-->A mutation rate is about 70x higher than the nuclear rate. Silent sites are substantially more strongly biased towards A and T than nonsynonymous sites, consistent with the extreme mutation bias towards A+T. Strand-asymmetric mutation bias, coupled with selection to maintain specific nonsynonymous bases, therefore provides an explanation for the extreme base composition of the mitochondrial genome of Drosophila.