The strategies by which freshwater teleosts maintain acid-base homeostasis under alkaline stress are attractive and have been explored for a long time. In this study, a cyprinid fish that tolerates extremely alkaline environments (pH 9.6), Leuciscus waleckii, was used as a model to explore the molecular mechanisms of acid-base regulation. Using a lab-controlled alkaline challenge test and 454 sequencing, the transcriptomes of their gills and kidney were profiled and compared. mRNA profiling produced 1 826 022 reads, generated 30 606 contigs with an average length of 1022 bp, of which 19 196 were annotated successfully. Comparative analysis of the expression profiles between alkaline and freshwater L. waleckii habitats revealed approximately 4647 and 7184 genes that were differentially expressed (p < 0.05) in gills and kidney, respectively, of which 2398 and 5127 had more than twofold changes in expression. Gene ontology analysis and KEGG enrichment analysis were conducted. Comprehensive analysis found that genes involved in ion transportation, ammonia transportation, and arachidonic acid metabolism pathways changed dramatically and played important roles in acid-base homeostasis in fish under alkaline stress. These results support the existing hypotheses about candidate genes involved in acid-base regulation under alkaline stress and prompt several new hypotheses. The large transcriptome dataset collected in this study is a useful resource for the exploration of homeostasis modulation in other fish species.

Demographic events and natural selection both influence animal phenotypic and genetic variation; exploring the effects of demography and selection on population divergence is of great significance in evolutionary biology. To uncover the causes behind the patterns of genetic differentiation and adaptation among six populations of Leuciscus waleckii from Dali Basin (two populations, alkaline vs. freshwater) and Amur Basin (four populations, freshwater rivers vs. alkaline lake), a set of 21 unlinked polymorphic microsatellite markers and two mitochondrial DNA sequences (Cytb and D-loop) were applied to examine whether populations from different environments or habitats have distinct genetic differentiation and whether alkalinity is the major factor that caused population divergence. Bayesian analysis and principal component analysis as well as haplotype network analysis showed that these populations are primarily divided into two groups, which are congruent with geographic separation but not inconsistent with the habitat environment (alkalinity). Using three different approaches, outlier detection indicated that one locus, HLJYL017, may be under directional selection and involved in local adaptation processes. Overall, this study suggested that demographic events and selection of local environmental conditions including of alkalinity are jointly responsible for population divergence. These findings constitute an important step towards the understanding of the genetic basis of differentiation and adaptation, as well as towards the conservation of L. waleckii.