Cytotaxonomy And Chromosome Evolution Of Panorpidae And Bittacidae (Mecoptera)

Mecoptera may represent one of the basal lineages in the Endopterygota,or specifically a basal taxon of Antliophora,because they possess compound eyes in the larval stages,and their fossil record dated back to the upper Permian.Based on the available characters,the phylogenetic position of the families,especially Bittacidae within Mecoptera is still controversial.The limitation of morphological features causes the difficulty of species delimitation and often leads to synonym problems in Panorpidae,which exhibits intricate intraspecific variation.Thus,it is necessary to add additional characters in Mecoptera.The chromosomes of eukaryotic organisms can provide uniquely important characters for taxonomic analyses and may help reveal the evolutionary relationships of species or higher taxa due to their evolutionary conservation.Chromosome numbers provide substantial information related to phylogeny and karyotype evolution.Differential banding techniques are able to identify the presence of numerous structural rearrangements,which may contribute to speciation.Such studies have been conducted on numerous insect groups,but have been largely neglected in Mecoptera to date.Here we investigated the meiosis and karyotypes of the species in Panorpidae and Bittacidae using C-banding technique and DAPI(4'-6-diamino-2-phenylindole)and Giemsa staining.We also used molecular and cytogenetic approaches to determine the phylogenetic relationships in the evolutionary scenario of chromosomes in Panorpidae.All the studied species of Panorpidae display achiasmate meiosis and the X0(?)/XX(?)mechanism of sex determination.We speculate that the X0(?)/XX(?)mechanism of sex determination may represent the plesiomorphy in Mecoptera,and the achiasmate meiosis is the autopomorphy in Panorpidae.During the pachytene stage,we observed a large number of non-homologous associations,which may relate to the functions of the telomere or attractions among heterochromatin segments.Panorpa Linnaeus,1758 is remarkable for its extensive variation of haploid chromosome number,ranging from n = 18 in Panorpa sp2 to n = 24 in P.kunmingensis Fu & Hua,2009,with 23 being the most frequent number.The variations of chromosome numbers provide additional cytological supports for the paraphyly of the genus Panorpa.Intraspecific variation caused by the B chromosome was observed in P.fulvastra Chou,1981 for the first time.In Neopanorpa Weele,1909 the majority of species show the chromosome number n = 21,but a great reduction of chromosomes occurs in N.lipingensis Cai & Hua,2009 and N.sheni Hua & Chou,1997,which exhibit an extraordinary chromosome number n = 17,the lowest chromosome number ever found in Panorpidae so far.The species of Cerapanorpa Gao,Ma & Hua,2016 exhibit two different chromosome numbers with the central Chinese species having n = 22,but the northeastern Asian species n = 23,indicating that this genus is likely a heterogeneous group.The two species examined in Sinopanorpa Cai & Hua,2008 both show n = 23.Dicerapanorpa magna(Chou,1981)also shows n = 23,but Furcatopanorpa longihypovalva(Hua & Cai,2009)exhibits n = 21.The great variations in the size and number of C-bands suggest that they are reliable traits for species delimitation in Panorpidae.The existence of natural C-banding polymorphism indicates that chromosomal rearrangements likely have contributed to the diversification of chromosomal bands in Panorpidae.All the studied species of Bittacidae had chiasmate meiosis and the X0(?)/XX(?)mechanism of sex determination,which are likely plesiomorphic in Bittacidae.The genus Bittacus Latreille,1805 is reprensentative by relatively low chromosome numbers,ranging from n = 8 in Bittacus sinicus Issiki,1931 to n = 19 in B.ussuriensis Plutenko,1985,and symmetric karyotypes with metacentric and submetacentric chromosomes.However,the species of Terrobittacus Tan & Hua,2009 possesses the highest chromosome number ever observed in Bittacidae and an asymmetric karyotype with mainly subtelocentric and telocentric chromosomes.The differences of karyotypes and meiosis between Bittacus and Terrobittacus may provide cytogenetic supports for the estabnlishment of the genus Terrobittacus.In Bittacus,chiasmata range from 17 to 21 in 50 nuclei counted,with a mean chiasma frequency of 19.5 per nucleus and a mean chiasma frequency of 1.1 per autosomal bivalent,and 77% of nuclei have ring-shaped bivalents in which each chromosomal arm possesses a single chiasma.In Terrobittacus,chiasmata range from 11 to 20 in 50 nuclei counted,with a mean chiasma frequency of 15.3 per nucleus and a mean chiasma frequency of 0.8 per autosomal bivalent.The pronounced variations in cytogenetic traits suggest that they are substantial traits for both species delimitation and systematic analysis in Bittacidae.The molecular phylogenetic analysis exhibits that Panorpidae is a monophyletic group and can be divided into two major clades.Clade I comprises the species of Neopanorpa and Leptopanorpa with haploid chromosome numbers no more than 21 and varied banding types.Clade II consists of the genera Cerapanorpa,Dicerapanorpa,Furcatopanorpa,Panorpa and Sinopanorpa,and is characterized by haploid chromosome numbers no less than 21(except Panorpa sp2)and the uniform pattern of C-banding.Neopanorpa is confirmed paraphyletic with Leptopanorpa for the first time.Cerapanorpa is also paraphyletic and needs further revision.The closely related species of Cerapanorpa are reconfirmed to be evolutionarily independent entities by cytogenetic and molecular data.The special status of Furcatopanorpa is likely attributed to frequent chromosome rearrangements.Our cytogenetic analysis suggests that chromosome fusion and translocation likely play a significant role in chromosome evolution of Panorpidae,and result in reduced chromosome number and complicated banding pattern.Cytogenetic data plotted on the phylogenetic tree show that interspecific ascending and,mainly,descending dysploidy are the major types of chromosome number variations,which contribute significantly to lineage differentiation in Panorpidae.Based on the above investigation,we conclude that chromosome variations are undoubtedly reliable traits for taxonomic and phylogenetic analyses,and contribute significantly to taxon differentiation in Mecoptera.The combined molecular-cytogenetic analyses may provide new insights into ambiguous phylogenetic relationships of organisms that lack sufficient supports.