The role of plants in the formation of species-specific features in grass flies (Diptera, Chloropidae, Meromyza)

In the current manuscript, we present the results of comparative analysis of seven species of Meromyza flies in the "variegata" cluster and of the evolutionary close species M. inornata, based the following criteria: 1) 14 external key features; 2) shape and area of the anterior processes of postgonites; 3) mtDNA CO1 region and 4) host plant diversity data. We could demonstrate the primary role of host plants in species formation inside genus Meromyza and calculated the timing of the divergence of M. inornata and the species of "variegata" cluster. Based on our estimates of evolution rate for mtDNA CO1 gene, we could conclude that that divergence of herbs happened before the speciation of grass flies Meromyza. Meromyza species, close to the ancestral species of the cluster, are adapted to the wide range of host plants. We revealed the most informative variables h1, S and Plant analysing data with the following statistical methods: linear discriminant analysis - LDA, regularised discriminant analysis - RDA, flexible discriminant analysis – FDA and probabilistic neural network - PNN. The highest classification accuracy was achieved using PNN (99%) and the lowest when using LDA (95.8%). When the Plant trait was excluded, the classification accuracy decreased by 14%. We revealed the significant trends in size change of the anterior process of the postgonite amongst studies species. This morphological structure is an element of male reproductive apparatus critical for the restriction of interspecies mating. We determined three branches of speciation in the "variegata" cluster and five trends in the evolution of this cluster, based on the external morphological features. We showed that M. variegata and especially M. mosquensis, the species closest to the ancestral haplotype, have the largest number of features typical of those of M. inornata. Based on the external features and the area of the anterior process of the postgonite, we reconstructed the phylogenetic position of M. elbergi in the cluster. In accordance with the obtained outcomes, we could conclude that the distribution, species diversity and the adaptation of the grass flies to narrow oligophagy were directly connected to host plant diversity. The adaptation to different host plants could be the main factor in divergence of grass flies and their evolution started later than the diversification in the Pooideae subfamily of grasses.