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Dengue and dengue hemorrhagic fever are increasing public health problems with an estimated 50–100 million new infections each year.Aedes aegyptiis the major vector of dengue viruses in its range and control of this mosquito would reduce significantly human morbidity and mortality. Present mosquito control methods are not sufficiently effective and new approaches are needed urgently. A “sterile-male-release” strategy based on the release of mosquitoes carrying a conditional dominant lethal gene is an attractive new control methodology. Transgenic strains ofAedes aegyptiwere engineered to have a repressible female-specific flightless phenotype using either two separate transgenes or a single transgene, based on the use of a female-specific indirect flight muscle promoter from theAedes aegypti Actin-4gene. These strains eliminate the need for sterilization by irradiation, permit male-only release (“genetic sexing”), and enable the release of eggs instead of adults. Furthermore, these strains are expected to facilitate area-wide control or elimination of dengue if adopted as part of an integrated pest management strategy.

Cold-adapted species in the Northern Hemisphere frequently show arctic-alpine discontinuous ranges at high latitudes and on mountains farther south, but area connectivity through current and historical gene flow remains unclear. We used the coalescent-based program IMa (Isolation with Migration-analytic) to test for migration among disjunct European areas of arctic-alpine wolf spiders of the Pardosa saltuaria group. Mitochondrial (ND1) and nuclear (ITS1, 5.8S rDNA, ITS2) markers were analyzed simultaneously. Gene flow was unidirectional from Scandinavia to the Alps and the Carpathians, complex with respect to intermediate relict areas in central Europe, and very limited in outlying areas in the Balkans and Pyrenees. Population connectivity may have been greater during glacial events that might alternatively account for the inferred arctic-alpine links. A simulation study under various demographic histories (using a new module in the Mesquite package, which models episodic migration) showed that the empirical results are equally consistent with moderate levels of ongoing (continuous) migration or, alternatively, with strong migration bursts at the last glacial maximum but not at earlier times. Habitat connectivity was probably maximal during glacial events, illustrating the potential influence of ecology and life history on organismal responses to past climatic change.

In Europe, 10 among the 15 known species of the Paramecium aurelia complex have been recorded including P. primaurelia, P. biaurelia, P. triaurelia, P. tetraurelia, P. pentaurelia, P. sexaurelia, P. septaurelia, P. novaurelia, P. dodecaurelia, and P. tredecaurelia. The occurrence of species of this complex in Europe has been estimated on the basis of the number of habitats and the ratio value (r.v.), i.e. the number of habitats for a defined species to the total number of habitats with species of the P. aurelia complex. The r.v. presents the relative frequency of the particular species to others of the complex. In total, 483 habitats were considered. The dominant species is P. novaurelia (r.v. 0.38, found in 186 habitats), followed by P. biaurelia (r.v. 0.26 in 125 habitats) and P. primaurelia (0.23 in 113 habitats). Other species are less frequent, such as P. tetraurelia (r.v. 0.06 found in 29 habitats), P. triaurelia (r.v. 0.04 in 22 habitats), or rare such as P. septaurelia (r.v. 0.025 found in 12 habitats), as well as P. pentaurelia and P. sexaurelia (r.v. 0.02 for both, found in 11 and 10 habitats, respectively). P. dodecaurelia was found in three localities in Europe (r.v. 0.006) and P. tredecaurelia is known from a single habitat (r.v. 0.002). This paper presents new populations, found at present, of species of the P. aurelia complex in Europe (Germany, Italy, Spain, Iceland, Poland, Great Britain) and one new from the USA.

We investigated mitochondrial Cytochrome b sequences from all 15 members of the enigmatic Paramecium aurelia species complex (Ciliophora). The analysis revealed high genetic distances between the different P. aurelia species (6.1-19.8%) and a largely unresolved, star-like phylogenetic tree. This result strongly supports a rapid radiation in the evolutionary history of this species complex and it correlates well with the hypothesis that the extant species diversity may have originated from the neutral consequences of a whole genome duplication in the common ancestor of P. aurelia.

To date the awareness of the temporal population structure in eukaryotic microbes is very limited. This is exemplified in the scarce knowledge about the intraspecific genetic variation in ciliates. To elucidate the genetic variation of Coleps (Ciliophora: Prostomatida), we employed the analysis of the mitochondrial apocytochrome b gene of the Coleps community in a young lake in Germany. The analysis of 111 isolates, sampled from April 2005 to September 2006, revealed a high genetic variation for the two dominant Coleps species (11 mitochondrial haplotypes in Coleps spetai, nine in Coleps hirtus hirtus). The study represents one of the largest datasets of intraspecific diversity in a microbial eukaryote and demonstrates for the first time the suitability of a mitochondrial gene for the detection of genetic variation within populations of eukaryotic microbes. However, the results of our study warrant caution in the application of such an approach, as we amplified some non-orthologous cob-like sequences, whose uncritical acceptance would have led to the erroneous discovery of cryptic species.

For the first time the complete mitochondrial genome was sequenced for a member of Lacertidae. Lacerta viridis viridis was sequenced in order to compare the phylogenetic relationships of this family to other reptilian lineages. Using the long-polymerase chain reaction (long PCR) we characterized a mitochondrial genome, 17,156 bp long showing a typical vertebrate pattern with 13 protein coding genes, 22 transfer RNAs (tRNA), two ribosomal RNAs (rRNA) and one major noncoding region. The noncoding region of L. v. viridis was characterized by a conspicuous 35 bp tandem repeat at its 5' terminus. A phylogenetic study including all currently available squamate mitochondrial sequences demonstrates the position of Lacertidae within a monophyletic squamate group. We obtained a narrow relationship of Lacertidae to Scincidae, Iguanidae, Varanidae, Anguidae, and Cordylidae. Although, the internal relationships within this group yielded only a weak resolution and low bootstrap support, the revealed relationships were more congruent with morphological studies than with recent molecular analyses.

Studies of intraspecific genetic diversity of ciliates, such as population genetics and biogeography, are particularly hampered by the lack of suitable DNA markers. For example, sequences of the non-coding ribosomal internal transcribed spacer (ITS) regions are often too conserved for intraspecific analyses. We have therefore identified primers for the mitochondrial cytochrome c oxidase I (COI) gene and applied them for intraspecific investigations in Paramecium caudatum and Paramecium multimicronucleatum. Furthermore, we obtained sequences of the ITS regions from the same strains and carried out comparative sequence analyses of both data sets. The mitochondrial sequences revealed substantially higher variation in both Paramecium species, with intraspecific divergences up to 7% in P. caudatum and 9.5% in P. multimicronucleatum. Moreover, an initial survey of the population structure discovered different mitochondrial haplotypes of P. caudatum in one pond, thereby demonstrating the potential of this genetic marker for population genetic analyses. Our primers successfully amplified the COI gene of other Paramecium. This is the first report of intraspecific variation in free-living protozoans based on mitochondrial sequence data. Our results show that the high variation in mitochondrial DNA makes it a suitable marker for intraspecific and population genetic studies.

Aquatic invertebrates experience strong trade-offs between habitats due to the selective effects of different predators. Diel vertical migration and small body size are thought to be effective strategies against fish predation in lakes. In the absence of fish in small ponds, migration is ineffective against invertebrate predators and large body size is an advantage. Although widely discussed, this phenomenon has never been tested in a phylogenetic context. We reconstructed a mitochondrial DNA (mtDNA) tree to investigate the phylogenetic distribution of pond and lake lifestyles among 10 species of northern temperate Chaoborus midge larvae. The mtDNA tree is similar to previous morphological trees for Chaoborus, the only difference being the disruption of the subgenus Chaoborus sensu stricto. At least three shifts have occurred between pond and lake lifestyles, each time associated with evolution of diel vertical migration in the lake taxon. The trend in larval body size with habitat type is sensitive to tree and character reconstruction methods, only weakly consistent with the effects of fish predation. Despite long time periods over which adaptation to each habitat type could have occurred, there remains significant phylogenetic heritability in larval body size. The tree provides a framework for comparative studies of the metapopulation genetic consequences of pond and lake lifestyles.

Background Ectothermic organisms are thought to be severely affected by global warming since their physiological performance is directly dependent on temperature. Latitudinal and temporal variations in mean temperatures force ectotherms to adapt to these complex environmental conditions. Studies investigating current patterns of thermal adaptation among populations of different latitudes allow a prediction of the potential impact of prospective increases in environmental temperatures on their fitness. Methodology/Principal Findings In this study, temperature reaction norms were ascertained among 18 genetically defined, natural clones of the microbial eukaryote Paramecium caudatum. These different clones have been isolated from 12 freshwater habitats along a latitudinal transect in Europe and from 3 tropical habitats (Indonesia). The sensitivity to increasing temperatures was estimated through the analysis of clone specific thermal tolerances and by relating those to current and predicted temperature data of their natural habitats. All investigated European clones seem to be thermal generalists with a broad thermal tolerance and similar optimum temperatures. The weak or missing co-variation of thermal tolerance with latitude does not imply local adaptation to thermal gradients; it rather suggests adaptive phenotypic plasticity among the whole European subpopulation. The tested Indonesian clones appear to be locally adapted to the less variable, tropical temperature regime and show higher tolerance limits, but lower tolerance breadths. Conclusions/Significance Due to the lack of local temperature adaptation within the European subpopulation, P. caudatum genotypes at the most southern edge of their geographic range seem to suffer from the predicted increase in magnitude and frequency of summer heat waves caused by climate change.

To determine the gene flow of Chaoborus crystallinus, populations throughout Europe were sampled. To test if the gene flow is higher within regions than between regions and to investigate if regional populations may act as metapopulations, four regions with several populations each were examined. For a detailed analysis of the regional gene flow, subregions within one region were analysed. Allozymes and mitochondrial restriction fragment length polymorphism (RFLP) were used to estimate the relative amounts of gene flow. On the European scale gene flow between populations within regions is higher than between regions. On the regional scale, gene flow between subregions is higher than between populations within subregions. Generally, the genetic differentiation between populations within regions is higher for the mitochondrial RFLP data than for the allozyme data. These results suggest that most of the dispersal is female dominated and mostly takes place between populations within regions. Due to this extensive regional gene flow, local adaptation may be influenced by regional processes. This might have interesting implications for the coevolution of the predator C. crystallinus and its plankton prey.

We investigated the morphology, phylogeny of the 18S rDNA, and pH response of Oxytricha acidotolerans sp. nov. and Urosomoida sp. (Ciliophora, Hypotricha) isolated from two chemically similar acid mining lakes (pH similar to 2.6) located at Langau, Austria, and in Lusatia, Germany. Oxytricha acidotolerans sp. nov. from Langau has 18 frontal-ventral-transverse cirri but a very indistinct kinety 3 fragmentation so that the assignment to Oxytricha is uncertain. The somewhat smaller species from Lusatia has a highly variable cirral pattern and the dorsal kineties arranged in the Urosomoida pattern and is, therefore, preliminary designated as Urosomoida sp. The pH response was measured as ciliate growth rates in laboratory experiments at pH ranging from 2.5 to 7.0. Our hypothesis was that the shape of the pH reaction norm would not differ between these closely related (3% difference in their SSU rDNA) species. Results revealed a broad pH niche for O. acidotolerans, with growth rates peaking at moderately acidic conditions (pH 5.2). Cyst formation was positively and linearly related to pH. Urosomoida sp. was more sensitive to pH and did not survive at circumneutral pH. Accordingly, we reject our hypothesis that similar habitats would harbour ciliate species with virtually identical pH reaction norm.

Water temperature, ice cover, and lake stratification are important physical properties of lakes and reservoirs that control mixing as well as bio-geo-chemical processes and thus influence the water quality. We used an ensemble of vertical one-dimensional hydrodynamic lake models driven with regional climate projections to calculate water temperature, stratification, and ice cover under the A1B emission scenario for the German drinking water reservoir Lichtenberg. We used an analysis of variance method to estimate the contributions of the considered sources of uncertainty on the ensemble output. For all simulated variables, epistemic uncertainty, which is related to the model structure, is the dominant source throughout the simulation period. Nonetheless, the calculated trends are coherent among the five models and in line with historical observations. The ensemble predicts an increase in surface water temperature of 0.34 K per decade, a lengthening of the summer stratification of 3.2 days per decade, as well as decreased probabilities of the occurrence of ice cover and winter inverse stratification by 2100. These expected changes are likely to influence the water quality of the reservoir. Similar trends are to be expected in other reservoirs and lakes in comparable regions. Projekt DEAL; European Social Fund (ESF); Saxonian state parliament; National Science Foundation ABI development Grant [DBI 1759865]; European Union's Horizon 2020 Research and Innovation Programme under the Marie Sklodowska-Curie Grant [722518]; WATExR project, ERA4CS, an ERA-NET by JPI Climate; MINECO (ES); FORMAS (SE); BMBF (DE); EPA (IE); RCN (NO); IFD (DK); European Union [690462]; NSF [DEB-1926050, DBI-1933016]; BMBF [FKZ 01LR 2005A] Published version Open Access funding enabled and organized by Projekt DEAL. Johannes Feldbauer is holder of a scholarship from the European Social Fund (ESF) and his work is co-financed by public tax funds as decided by the Saxonian state parliament. Robert Ladwig was funded through a National Science Foundation ABI development Grant (#DBI 1759865). Jorrit P. Mesman was funded by the European Union's Horizon 2020 Research and Innovation Programme under the Marie Sklodowska-Curie Grant Agreement No. 722518 (MANTEL ITN). Tadhg N. Moore was funded by: the WATExR project which is part of ERA4CS, an ERA-NET initiated by JPI Climate, and funded by MINECO (ES), FORMAS (SE), BMBF (DE), EPA (IE), RCN (NO), and IFD (DK), with co-funding by the European Union (Grant Number: 690462) and also by NSF Grants DEB-1926050 and DBI-1933016. Thomas U Berendonk, Thomas Petzoldt, and Johannes Feldbauer also received funding from the BMBF project FKZ 01LR 2005A-Forderma ss nahme "Regionale Informationen zum Klimahandeln" (RegIKlim).

Two sister species of Chaoborus exist in Europe. They are found in two distinct habitats that differ strongly in habitat persistence time. Chaoborus flavicans is found primarily in lakes inhabited by fish, whereas Chaoborus crystallinus is found exclusively in small ponds without fish. Chaoborus flavicans has evolved an adaptive response to predatory fish. In the presence of fish chemicals it can migrate vertically in the water column and can thus avoid predation. Chaoborus flavicans can therefore coexist with fish. Chaoborus crystallinus avoids oviposition into water bodies that are inhabited by fish and hence lives only in small fish-free ponds. These differences lead to several testable hypotheses: (1) The extinction risk of local populations of C. crystallinus is higher than that for C. flavicans. (2) C. crystallinus has a higher dispersal probability than C. flavicans. (3) The spatial population structure of each species differ. The results from a 3-yr population survey verified the first hypothesis that the extinction risk for C. crystallinus is higher than for C. flavicans. Field and laboratory experiments confirmed the second hypothesis. The results showed that, although C. flavicans disperses faster, the probability that C. crystallinus individuals disperse farther than 1 km is almost an order of magnitude higher compared to C. flavicans. The comparison of the spatial population structure of the two species shows that Chaoborus live in two different population structures that are subject to different metapopulation processes. Chaoborus crystallinus exists in a Levins-metapopulation, while C. flavicans exhibits a mainland-island population structure; these findings confirmed the third hypothesis. These results show that two species of the same genus live in two very different metapopulation structures and therefore present an ideal system to test comparative hypotheses within metapopulation ecology.

Treated wastewater (TWW) irrigation is a useful counter-measure against the depletion of freshwater (FW) resources. However, TWW contains several contaminants of emerging concern, such as antibiotic resistant bacteria (ARB) and antibiotic resistant genes (ARGs). Thus, TWW irrigation might promote the spread of antimicrobial resistance in soil environments. In the present work, we hypothesized that the ARG load and irrigation intensity define the effect of TWW irrigation on ARG spread dynamics in soil. This hypothesis was tested using a multiphase approach: a) comparing soil from a full-scale, commercially operated, TWW irrigated field with non-irrigated soil, b) long-term sampling of the TWW irrigated field over one year with different irrigation intensities and intercepted by irrigation breaks and c) laboratory-scale soil microcosms irrigated with TWW compared to FW. Six ARGs, the integrase gene intI1 and the 16S rRNA were quantified using qPCR. In addition, effects of TWW irrigation on bacterial community composition of microcosm-samples were analysed with 16S rRNA amplicon sequencing. The genes sul1, qnrS, blaOXA-58, tet(M) and intI1 were significantly more abundant in the TWW irrigated field soil, whereas blaCTX--M-32 and blaTEM, the least abundant genes in the TWW irrigation, showed higher abundance in the non-irrigated soil. The relative abundance of sul1, qnrS, blaOXA-58, tet(M) and intI1 correlated with TWW irrigation intensity and decreased during irrigation breaks. Despite the decrease, the levels of these genes remained consistently higher than the non-irrigated soil indicating persistence upon their introduction into the soil. Microcosm experiments verified observations from the field study: TWW irrigation promoted the spread of ARGs and intI1 into soil at far elevated levels compared to FW irrigation. However, the impact of TWW irrigation on 16S rRNA absolute abundance and the soil microbial community composition was negligible. In conclusion, the impact of TWW irrigation depends mainly on the introduced ARG load and the irrigation intensity.