Molecular phylogeny and timing of diversification in Alpine Rhithrogena (Ephemeroptera: Heptageniidae).

BACKGROUND: Larvae of the Holarctic mayfly genus Rhithrogena Eaton, 1881 (Ephemeroptera, Heptageniidae) are a diverse and abundant member of stream and river communities and are routinely used as bio-indicators of water quality. Rhithrogena is well diversified in the European Alps, with a number of locally endemic species, and several cryptic species have been recently detected. While several informal species groups are morphologically well defined, a lack of reliable characters for species identification considerably hampers their study. Their relationships, origin, timing of speciation and mechanisms promoting their diversification in the Alps are unknown. RESULTS: Here we present a species-level phylogeny of Rhithrogena in Europe using two mitochondrial and three nuclear gene regions. To improve sampling in a genus with many cryptic species, individuals were selected for analysis according to a recent DNA-based taxonomy rather than traditional nomenclature. A coalescent-based species tree and a reconstruction based on a supermatrix approach supported five of the species groups as monophyletic. A molecular clock, mapped on the most resolved phylogeny and calibrated using published mitochondrial evolution rates for insects, suggested an origin of Alpine Rhithrogena in the Oligocene/Miocene boundary. A diversification analysis that included simulation of missing species indicated a constant speciation rate over time, rather than any pronounced periods of rapid speciation. Ancestral state reconstructions provided evidence for downstream diversification in at least two species groups. CONCLUSIONS: Our species-level analyses of five gene regions provide clearer definitions of species groups within European Rhithrogena. A constant speciation rate over time suggests that the paleoclimatic fluctuations, including the Pleistocene glaciations, did not significantly influence the tempo of diversification of Alpine species. A downstream diversification trend in the hybrida and alpestris species groups supports a previously proposed headwater origin hypothesis for aquatic insects

DNA barcoding and faunistic criteria for a revised taxonomy of Italian Ephemeroptera

<p>Mayfly (Ephemeroptera) systematics has considerably changed over the years, but many questions have yet to be answered. The synergistic connection between traditional knowledge and new data sources, producing increasingly complex information, has become a compelling issue for modern taxonomy. Molecular tests and the use of reliable reference sequence libraries may constitute effective complements to the traditional method in guiding recognition of species and giving information about taxonomic incongruences which require further examination. In the present study, we sought to verify the current Italian mayfly nomenclatural system through DNA barcoding and relevant points to reliably manage the available amount of morpho-ecological and molecular data are discussed. We investigated COI (Cytochrome oxidase I) sequence variation in 163 individuals of Italian mayflies, 126 of which were previously assigned to 24 morphologically recognised species, and 37 could be attributed only to generic taxonomic entities (“sp.”, “cf.” or “gr.”). DNA barcoding statistical tests for species delimitation hypotheses based on genetic distances and inferred gene trees were integrated with GenBank searches and surveys of the historical literature to better understand the knowledge acquired on the status and diversity of the investigated taxa. Combined criteria to define three categories of reliability were then assessed. Concurrent data allowing unambiguous identification were attained for only eight species. High intraspecific genetic distances (> 3%) and a lack of reliable reference material or convincing taxonomic information evidenced 29 critical states, deserving further investigation. Solid species names, potential cryptic species and entities about which little is known are pointed out for a future upgrade/reorganisation of the taxonomy of Italian Ephemeroptera.</p

Towards resolving the double classification in Erythraeus (Actinotrichida: Erythraeidae): matching larvae with adults using 28S sequence data and experimental rearing

The taxonomy of free-living adults and heteromorphic parasitic larvae of Parasitengona mites has in the past been treated independently resulting in a double classification. Correct linkage of names still remains unknown for many species. A holistic understanding of species is imperative for understanding their role in ecosystems. This is particularly true for groups like parasitengone mites with a radically altered lifestyle during development—parasitic to predatory. Here, we infer linkages of three nominal species of Erythraeus, using matching with 28S DNA sequence data from field-collected specimens and through laboratory rearing. The general mixed Yule coalescent method (GMYC) was used to explicitly test if field-collected specimens representing heteromorphic life instars were conspecific. The field-collected larvae were allocated to adults of Erythraeus cinereus and Erythraeus regalis, respectively. Laboratory rearing of the same two species confirmed the matching done by DNA. Rearing was also successful for Erythraeus phalangoides after eggs were treated to an imitated winter diapause. This integrative taxonomic approach of molecular, morphological, and rearing data resulted in the following synonyms: E. phalangoides (De Geer, 1778) [= Erythraeus adrastus(Southcott, 1961), syn. nov.], E. cinereus (Dugès, 1834) [= Erythraeus jowitae Haitlinger, 1987, syn. nov.], and E. regalis (C.L. Koch, 1837) [= Erythraeus kuyperi (Oudemans, 1910), syn. nov., = Erythraeus gertrudae Haitlinger, 1987, syn. nov.]. The molecular evidence confirmed the separate identity of three further members of the genus. We provide redescriptions of E. phalangoides, E. cinereus, and E. regalis after modern standards, and neotypes are designated