Adaptive Evolution Of Mitochondrial Genome Of Cave Species Of Triplophysa And Oreonectes In Guizhou

The Nemacheilidae of Cypriniformes is widely distributed in the tropics or subtropics of Asia,and inhabits in various water environments,especially in fast-flowing mountain streams or caves.Different living environments may help to explain that there are many cave fishes in Nemacheilidae.At present,more than 20 genera and 200 species of Nemacheilidae have been recorded in China.Triplophysa and Oreonectes are one of the most widely distributed,richest species diversity and most heterogeneous biological groups in the family Loachidae,respectively.In the past,many disputes have been put forward on the classification and origin of these two genera.However,due to the lack of support of molecular data and large-scale sampling,as well as the great similarity in appearance,morphology and habitat,the systematic classification and evolutionary relationship between the two genera in the Nemacheilidae has always been controversial.In this study,there were Triplophysa(7 species from Guizhou)and Oreonectes(7 species from Guizhou and 2 species from Guangxi)collected,and using the mitochondrial genome as a molecular marker,the systematic classification and divergence time of these two genera was basically clarified,and revealed the molecular adaptation mechanism of cave fish.The main research results include the following:1.The basic characteristics of mitochondrial genome of Triplophysa and OreonectesThe mitochondrial genomes of Triplophysa and Oreonectes have a typical closed ring structure,containing 13 encoding proteins,22 trn,2 rRNA and a control region.The light strand only encodes ND6 and 8 trn(trnQ,trn,trnN,trnC,trnY,trnS2,trnE,trnP)genes,the rest are all encoded by the heavy strand.2 genus 9 species full mitochondrial genome analysis,the results show that 9 cave fish mitochondrial genome nucleotide composition content is biased AT,16 S A + T% content is higher than 12 S,in 13 PCGs,except for COI using GTG In addition to the start codon,the rest use the ATG start codon.2.Systematic classification of Triplophysa and OreonectesIn this study,phylogenetic analysis using mitochondrial genome reconstruction based on field sampling and NCBI database data showed that:(1)Vaillantellidae,Botiidae,Ellopostomatidae,Barbuccidae,Balitoridae,SerpentiCytbitidae,Gastromyzontidae,Cytbitidae and Nemacheilidae's monophyly were highly supported(UFB = 100%,BPP = 1.00).The sequence used in this study basically covers all Cytbitoidea taxa and combines with the mitochondrial genome,which basically clarifies the systematic classification status of Cytbitoidea.(2)Based on mitochondrial genome data,this study focuses on the phylogenetic status of Triplophysa and Oreonectes sensu lato in Nemacheilidae.However,there are still some genus species have not been included in this study,this needs to be sampled and analyzed in the next research,such as: Yunnanilus,Paranemachilus and Heminoemacheilus.(3)Based on the mitochondrial genome,this study was collected from Guangxi daxing Troglonectes with Oreonectes platycephalus and did not form a monophyletic group with Troglonectes.On the contrary,this study found that it has a close relationship with Pingtoiling loach but has not gathered into a clade,and has a high support rate(UFB = 100%,BPP = 1.00).Therefore,this study believes that the effectiveness of Troglonectes is valid,but more fork caudal fin and round caudal fin species need to be included for research.(4)The validity of Troglonectes daqikongensis is established,Oreonectes jiarongensis and ParaCytbitis maolanensis synonymy species with Troglonectes longibarbatus.(5)Based on the morphological and molecular data of specimens from the type locality,the Triplophysa lihuensis should belong to Troglonectes,and should be renamed Troglonectes lihuensis.3.The time of divergence between Triplophysa and OreonectesDivergent time analysis based on genomic data calibration shows that the spatial and temporal diversity of the Triplophysa and Oreonectes sensu lato are closely related to past geological changes and climate changes.Triplophysa originated at about 106.54 Ma and began to diversify around 95.28 Ma,which differentiated earlier than most taxa in the Loachidae.The clade(including Oreonectes,Micronemacheilus,Yunnanilus)where the broad loach belongs to originated from 89.52 Ma.In the Triplophysa and Oreonectes flathead species groups,the first diversification event occurred around 65 Ma(~ 50–80 Ma)in the Upper Cretaceous,including the plateau loach clade C1–C2,and the second diversification event of the genus Fortunei of the Loach genus occurred around 35 Ma(~ 30–40 Ma)in the late Eocene,which coincides with the first uplift of the Qinghai-Tibet Plateau caused by the collision between the Indian subcontinent and the Asian plate during this period,as well as the cooling temperature event.Therefore,it can be preliminarily inferred that the formation and diversification of cave-type Triplophysa and Oreonectes species are related to the geological changes caused by the uplift of the Tibetan Plateau,especially the development of karst cave-type landforms.4.Molecular adaptation mechanism of cave fishesThrough the selective pressure analysis of Triplophysa and Oreonectes' cave types,the results showed that some protein coding genes of cave types were positively selected,which indicated that the cave types had accelerated evolution at the level of mitochondrial genome.Two species of Triplophysa(ATP6 ? ND5)and nine species of Oreonectes(ATP6?ATP8?COI?CO2?Cytb?ND3?ND4?ND4L?ND5)were positively selected gene.ND series genes may play an important role in guiding mitochondria to regulate energy storage and storage of lipids in cave fish,while ATP series genes may play an important role in regulating the energy supply of cave fish.