Background: Molecular research revealed that some of the EuropeanCalypogeiaspecies described on the basis of morphological criteria are genetically heterogeneous and, in fact, are species complexes. DNA barcoding is already commonly used for correct identification of difficult to determine species, to disclose cryptic species, or detecting new taxa. Among liverworts, some DNA fragments, recommend as universal plant DNA barcodes, cause problems in amplification. Super-barcoding based on genomic data, makes new opportunities in a species identification.Results: On the basis of 22 individuals, representing 10Calypogeiaspecies, plastid genome was tested as a super-barcode. It is not effective in 100%, nonetheless its success of species discrimination (95.45%) is still conspicuous. It is not excluded that the above outcome may have been upset by cryptic speciation inC. suecica, as our results indicate. Having the sequences of entire plastomes of EuropeanCalypogeiaspecies, we also discovered that thendhBandndhHgenes and thetrnT-trnLspacer identify species in 100%.Conclusions: This study shows that even if a super-barcoding is not effective in 100%, this method does not close the door to a traditional single- or multi-locus barcoding. Moreover, it avoids many complication resulting from the need to amplify selected DNA fragments. It seems that a good solution for species discrimination is a development of so-called “specific barcodes” for a given taxonomic group, based on plastome data.

A recently presented taxonomical arrangement of the moss genusOrthotrichumHedw. s.l. substantially changed the traditional view of the taxon that had been accepted throughout the twentieth century. This paper provides the results of mitogenomic studies that strongly support the new taxonomical concept. Comparative analyses presented in this study confirmed the stable structure of moss mitogenomes. Moreover, 17 complete mitogenome sequences were used to identify the major evolutionary groups, including 11 newly sequenced ones, for this study. The analysis of mitochondrial hotspots revealed intron 4 of thecox1 gene to be the most variable non-coding region. The most variable protein-coding genes in the tribe Orthotricheae wereccmFC andtatC. The intergenic and intronic hotspots ofOrthotrichums.l. identified in the present study do not correspond to those described in vascular plant mitogenomes.

Aneura pinguisis known as a species complex with several morphologically indiscernible species, which are often reproductively isolated from each other and show distinguishable genetic differences. Genetic dissimilarity of cryptic species may be detected by genomes comparison. This study presents the first complete sequences of chloroplast and mitochondrial genomes of six cryptic species ofA. pinguiscomplex:A. pinguisA, B, C, E, F, J. These genomes have been compared to each other in order to reconstruct phylogenetic relationships and to gain better understanding of the evolutionary process of cryptic speciation in this complex. The chloroplast genome with the nucleotide diversity 0.05111 and 1537 indels is by far more variable than mitogenome with π value 0.00233 and number of indels 1526. Tests of selection evidenced that on about 36% of chloroplast genes and on 10% of mitochondrial genes ofA. pinguisacts positive selection. It suggests an advanced speciation of species. The phylogenetic analyses based on genomes show thatA. pinguisis differentiated and forms three distinct clades. Moreover, on the cpDNA trees,Aneura mirabilisis nested among the cryptic species ofA. pinguis. This indicates that theA. pinguiscryptic species do not derive directly from one common ancestor.

Background: The mitogenomes of vascular plants are one of the most structurally diverse molecules. In the present study we characterize mitogenomes of a rare and endangered speciesPulsatilla patens. We investigated the gene content and its RNA editing potential, repeats distribution and plastid derived sequences.Results: The mitogenome structure of early divergent eudicot, endangeredPulsatilla patensdoes not support the master chromosome hypothesis, revealing the presence of three linear chromosomes of total length 986 613 bp. The molecules are shaped by the presence of extremely long, exceeding 87 kbp repeats and multiple chloroplast-derived regions including nearly complete inverted repeat. Since the plastid IR content of Ranunculales is very characteristic, the incorporation into mitogenome could be explained rather by intracellular transfer than mitochondrial HGT. The mitogenome contains almost a complete set of genes known from other vascular plants with exception ofrps10 andsdh3, the latter being present but pseudogenized. Analysis of long ORFs enabled the identification of genes which are rarely present in plant mitogenomes, including RNA and DNA polymerases, albeit their presence even at species level is variable. Mitochondrial transcripts ofP. patenswere edited with a high frequency, which exceeded the level known in other analyzed angiosperms, despite the strict qualification criteria of counting the editing events and taking into analysis generally less frequently edited leaf transcriptome. The total number of edited sites was 902 andnad4 was identified as the most heavily edited gene with 65 C to U changes. Non-canonical, reverse U to C editing was not detected. Comparative analysis of mitochondrial genes of threePulsatillaspecies revealed a level of variation comparable to chloroplast CDS dataset and much higher infrageneric differentiation than in other known angiosperm genera. The variation found in CDS of mitochondrial genes is comparable to values found amongPulsatillaplastomes. Despite the complicated mitogenome structure, 14 single copy regions of 329 kbp, not splitted by repeats or plastid-derived sequences (MTPT), revealed the potential for phylogenetic, phylogeographic and population genetics studies by revealing intra- and interspecific collinearity.Conclusions: This study provides valuable new information about mitochondrial genome of early divergent eudicots,Pulsatilla patens, revealed multi-chromosomal structure and shed new light on mitogenomics of early eudicots.

Comparative analyses of chloroplast and mitochondrial genomes have shown that organelle genomes in bryophytes evolve slowly. However, in contrast to seed plants, the organellar genomes are yet poorly explored in bryophytes, especially among liverworts. Discovering another organellar genomes of liverwort species by sequencing provides new conclusions on evolution of bryophytes. In this work, the organellar genomes of Gymnomitrion concinnatum liverwort were sequenced, assembled and annotated for the first time. The chloroplast genome displays, typical for most plants, quadripartite structure containing large single copy region (81,701 bp), two inverted repeat regions (8704 bp each) and small single copy region (20,179 bp). The gene order and content of chloroplast are very similar to other liverworts with minor differences observed. A total number of 739 and 222 RNA editing sites were predicted in chloroplast and mitochondrial genes of G. concinnatum. The mitochondrial genome gene content is also in accordance with liverworts except few alterations such as: intron loss in cox1 and atp1 genes. Nonetheless the analysis revealed that G. concinnatum mitogenome structure and gene order are rearranged in comparison with other mitogenomes of liverworts. The causes underlying such mitogenomic rearrangement were investigated and the probable model of recombination was proposed. This study provide the overview of mitochondrial and chloroplast genome structure and gene order diversity of Gymnomitrion concinnatum against the background of known organellar genomes of liverworts. The obtained results cast doubt on the idea that mitogenome structure of early land plants is highly conserved as previous studies suggested. In fact is the very first case of recombination within, evolutionary stable, mitogenomes of liverworts.

Oil bodies are the unique feature of most liverworts. Their shape, color and distribution pattern in leaf and underleaf cells are important taxonomic features of the genus Calypogeia. Most species of the genus Calypogeia have pellucid and colorless oil bodies, whereas colored, including gray to pale brown, purple-brown or blue oil bodies, are rare. To date, C. azurea was the only species with blue oil bodies to have been considered as a species of the Holarctic range. This species has been noted in various parts of the northern hemisphere–from North America, through Europe to the Far East. The aim of this study was to determine the genetic diversity of C. azurea from different parts of its distribution range and to ascertain whether blue oil bodies appeared once or several times in the evolution of the genus Calypogeia. The phylogenetic analyses based on four plastid regions (rbcL, trnG, trnL, trnH-psbA) and one nuclear region (ITS2) revealed that C. azurea is presently a paraphyletic taxon, with other Calypogeia species nested among C. azurea accessions that were clustered into four different clades. Based on the level of genetic divergence (1.03–2.17%) and the observed morphological, ecological and geographical differences, the evaluated clades could be regarded as previously unrecognized species. Four species were identified: C. azurea Stotler & Crotz (a European species corresponding to the holotype), two new species from Pacific Asia—C. orientalis Buczkowska & Bakalin and C. sinensis Bakalin & Buczkowska, and a North American species which, due to the lack of identifiable morphological features, must be regarded as the cryptic species of C. azurea with a provisional name of C. azurea species NA.