Diverse phenotypic and genetic responses to short‐term selection in evolving Escherichia coli populations

Beneficial mutations fuel adaptation by altering phenotypes that enhance the fit of organisms to their environment. However, the phenotypic effects of mutations often depend on ecological context, making the distribution of effects across multiple environments essential to understanding the true nature of beneficial mutations. Studies that address both the genetic basis and ecological consequences of adaptive mutations remain rare. Here, we characterize the direct and pleiotropic fitness effects of a collection of 21 first‐step beneficial mutants derived from naïve and adapted genotypes used in a long‐term experimental evolution of Escherichia coli. Whole‐genome sequencing was able to identify the majority of beneficial mutations. In contrast to previous studies, we find diverse fitness effects of mutations selected in a simple environment and few cases of genetic parallelism. The pleiotropic effects of these mutations were predominantly positive but some mutants were highly antagonistic in alternative environments. Further, the fitness effects of mutations derived from the adapted genotypes were dramatically reduced in nearly all environments. These findings suggest that many beneficial variants are accessible from a single point on the fitness landscape, and the fixation of alternative beneficial mutations may have dramatic consequences for niche breadth reduction via metabolic erosion.     

Adaptation,Clonal Interference,and Frequency-Dependent Interactions in a Long-Term Evolution Experiment with Escherichia coli

Twelve replicate populations of Escherichia coli have been evolving in the laboratory for >25 years and 60,000 generations. We analyzed bacteria from whole-population samples frozen every 500 generations through 20,000 generations for one well-studied population, called Ara−1. By tracking 42 known mutations in these samples, we reconstructed the history of this population’s genotypic evolution over this period. The evolutionary dynamics of Ara−1 show strong evidence of selective sweeps as well as clonal interference between competing lineages bearing different beneficial mutations. In some cases, sets of several mutations approached fixation simultaneously, often conveying no information about their order of origination; we present several possible explanations for the existence of these mutational cohorts. Against a backdrop of rapid selective sweeps both earlier and later, two genetically diverged clades coexisted for >6000 generations before one went extinct. In that time, many additional mutations arose in the clade that eventually prevailed. We show that the clades evolved a frequency-dependent interaction, which prevented the immediate competitive exclusion of either clade, but which collapsed as beneficial mutations accumulated in the clade that prevailed. Clonal interference and frequency dependence can occur even in the simplest microbial populations. Furthermore, frequency dependence may generate dynamics that extend the period of coexistence that would otherwise be sustained by clonal interference alone.     

Results of the seventh joint pesticide testing programme carried out by the IOBC/WPRS-Working Group ‘Pesticides and Beneficial Organisms’

The side effect of 10 insecticides, 5 fungicides and 5 herbicides on 24 different species of beneficial organisms was tested by members of the Working Group Pesticides and Beneficial Organisms of the International Organization for Biological Control (IOBC), West Palaearctic Regional Section (WPRS). The tests were conducted by 32 members in 12 countries according to internationally approved guidelines.The microbial insecticides Bacillus thuringiensis var. kurstaki (Delfin), B. thuringiensis var. tenebrionis (Novodor) and Verticillium lecanii (Micro Germin), the fungicides cyproconazol (Alto), difenoconazol (Score), lecithin (Bioblatt Mehltau) and penconazol (Omnex), and the herbicides ethofumesat (Tramat), fluroxypyr (Starane), haloxyfop (Gallant), isoproturon (Arelon) and metamitron (Goltix) were harmless to nearly all the beneficial arthropods. The benzoylurea's teflubenzuron (Nomolt) and flufenoxuron (Cascade) affected predators such as anthocorids, earwigs, coccinellids and lacewings. The remaining preparations were more toxic and should therefore be further tested in semi-field and field experiments on relevant organisms. Most tested fungicides were toxic for the entomopathogenic fungi.     

Effects of 45 insecticides,acaricides and molluscicides on the rove beetleAleochara bilineata (Col.: Staphylinidae) in the laboratory

Effects of 45 insecticides, acaricides and molluscicides on adult females of the rove beetleAleochara bilineata were investigated in the laboratory. The pesticides were tested in concentrations equivalent to the highest recommended dosages for practical use. Mortality, egg production and hatch of the eggs were measured. The reproducibility of the test and the suitability of the strain ofA. bilineata used are discussed. The toxicity of the cholinesterase inhibitors tested was varying, but in general high. Only pirimicarb and acephate showed low toxicity. The pyrethroids tested were highly toxic, even though fluvalinate allowed a few eggs to be laid. The test did not give conclusive results for insect growth regulators. The ecological relevance of laboratory results is discussed, and it is concluded that only for pesticides with no or low toxicity can the results be used for prediction of effects in the field.     

Oryzacystatin I expressed in transgenic potato induces digestive compensation in an insect natural predator via its herbivorous prey feeding on the plant

We observed recently that the rice cysteine proteinase inhibitor, oryzacystatin I (OCI) expressed in transgenic potato does not affect growth and development of the two-spotted stinkbug predator (Perillus bioculatus) via its herbivorous prey feeding on the plant. Here we monitored the inhibitory activity of recombinant OCI along this potato --> herbivore --> predator continuum, to determine if the absence of effect was associated with a digestive compensatory response of the predator following inhibition of its proteinases by the recombinant cystatin. After confirming that OCI is present in the plant, and ingested in an active form by potato beetle larvae, quantitative and electrophoretic assays allowed us to determine that the recombinant cystatin (representing about 0.8% of total soluble proteins in leaves) was entirely bound to a approximately 30-kDa target proteinase in the prey's midgut, forming a sodium dodecyl sulphate (SDS)-stable complex detected on immunoblots with an anti-OCI polyclonal antibody. Despite the apparent absence of free, residual OCI in the beetle's midgut, digestive protease activity in the predator, known to include OCI-sensitive activity, was altered negatively when the prey was fed the modified plant. This inhibitory process at the third trophic level was accompanied by a compensatory response in the predator, by which serine-type proteinases were synthesized de novo. Overall, our data suggest that the affinity between OCI and the predator's OCI-sensitive proteinases is: (i) as strong as (or stronger than) the affinity between OCI and the potato beetle 30-kDa-sensitive proteinase; and (ii) stronger than the affinity between these enzymes and the plant endogenous homologue of OCI, potato multicystatin, induced in the plant by potato beetle feeding. Our results also show that predatory organisms can adapt their digestive metabolism to the presence of plant antidigestive proteins ingested by their herbivorous preys. In a broader context, this study stresses the need to monitor the inhibitory effects of PI-expressing plants not only on the herbivorous insects targeted, but also on the organisms likely to consume these pests in the environment.     

Parallel genetic changes and nonparallel gene-environment interactions characterize the evolution of drug resistance in yeast

Beneficial mutations are required for adaptation to novel environments, yet the range of mutational pathways that are available to a population has been poorly characterized, particularly in eukaryotes. We assessed the genetic changes of the first mutations acquired during adaptation to a novel environment (exposure to the fungicide, nystatin) in 35 haploid lines of Saccharomyces cerevisiae. Through whole-genome resequencing we found that the genomic scope for adaptation was narrow; all adapted lines acquired a mutation in one of four late-acting genes in the ergosterol biosynthesis pathway, with very few other mutations found. Lines that acquired different ergosterol mutations in the same gene exhibited very similar tolerance to nystatin. All lines were found to have a cost relative to wild type in an unstressful environment; the level of this cost was also strongly correlated with the ergosterol gene bearing the mutation. Interestingly, we uncovered both positive and negative effects on tolerance to other harsh environments for mutations in the different ergosterol genes, indicating that these beneficial mutations have effects that differ in sign among environmental challenges. These results demonstrate that although the genomic target was narrow, different adaptive mutations can lead populations down different evolutionary pathways, with respect to their ability to tolerate (or succumb to) other environmental challenges.