Studies On Integrative Taxonomy And Molecular Phylogeny Of Some Species Of Myxobolus

Myxosporea is a cosmopolitan group of metazoan species that primarily infects and harms fish.The genus Myxobolus is the largest group of known species among Myxosporea.Based on the traditional morphological classification,Myxobolus species often cannot be accurately defined,and their phylogenetic relationships and evolutionary history are still unclear.This study integrated the morphological,histological,molecular biology,and phylogenetic approaches to identify the Myxobolus species collected from some regions in China,demonstrate the phylogenetic position of these species and elucidate the phylogenetic relationships and evolutionary history of Myxobolidae species.The main research results and conclusions are as follows:A total of 32 Myxobolus species were studied in the present work.Among these,three new species were identified,namely Myxobolus shuifuensis n.sp.,Myxobolus pseudowulii n.sp.,and Myxobolus distalisensis n.sp.We supplemented the description of 25 known species,and the validity of eight species was confirmed.The classification data of other species were further improved,and four species were regarded as synonymous with other species.The nomination issues of three species were corrected.Furthermore,four Myxobolus species were temporarily not assigned due to lack of definitive classification evidence,and only taxonomic information was provided.The hosts and infection sites of each species were determined.Among them,nine species infecting multiple hosts were supported by the results of this study.Four species were found infecting multiple organs of their hosts,and the tissue tropism of 18 species was elucidated in the present study.A total of 31 SSU rDNA molecular markers for 26 Myxobolus species were provided for correct identification of the concerned species.The conspecific nature was identified based on over 99% sequence identity.The molecular sequences of other species were compared with those in the Genbank database,and the identity was the highest at 97.24%but did not reach the threshold for characterizing the intraspecific variation.Moreover,the LSU r DNA sequence of 16 isolates was supplemented in this study to implement the phylogenetic analysis.Phylogenetic analysis of Myxobolidae species demonstrated that:(1)Compared to single gene analysis based on the SSU r DNA sequences,the analysis of LSU r DNA sequences and SSU-LSU r DNA concatenated sequences lessly improved bootstrap value of phylogenetic relationships,and the SSU r DNA sequence was still appliable for use in the phylogenetic analysis of Myxobolidae species;(2)In this study,the present species were placed in all major branches of the phylogenetic tree,enriching the branches of the phylogenetic tree.Among them,the clustering relationship of those Myxobolidae species infecting either Tachysurus or Hypophthalmichthys fish was apparent;(3)The traditional taxa of Myxobolidae failed to form a monophyletic clade,and the Myxobolus,Henneguya,Thelohanellus,and Unicauda genera were intermixed in the present analysis;(4)The host’s taxonomic affiliation at the order level and its geographical distribution to some extent affected the clustering relationship of Myxobolidae species.Cypriniform fish significantly affected the clustering of the concerned species,and species within this clade seemed to undergo rapid population expansion;(5)The phylogenetic relationship of Myxobolidae species infecting cypriniform fish showed that the host’s family is significantly correlated with the clustering relationship.Species with similar morphology and infecting the same organs in the host were clustered in individual branches,which may be related to their adaptability to either host or organ during evolution.In summary,this work was based on an integrative taxonomic approach to identify the species of Myxobolus,supplementing the taxonomic information on some species and enhancing our understanding of Myxosporea diversity in China.The construction of a phylogenetic tree based on the SSU r DNA sequences confirmed the phylogenetic positions of the studied species,enriched the branches of the phylogenetic tree,and confirmed the non-monophyletic origins of various groups within the family Myxobolidae.It provided preliminary insights into the complex evolutionary processes of Myxobolidae species.