Phylogenetic Implications Research Articles

Pantanal flood pulse reveals constitutive and plastic features of two wild rice species (Poaceae, Oryzoideae): implications for taxonomy, systematics, and phylogenetics

Pantanal flood pulse reveals constitutive and plastic features of two wild rice species (Poaceae, Oryzoideae): implications for taxonomy, systematics, and phylogenetics

Characterization of the complete mitochondrial genome of the fluke of turdus, Plagiorchis elegans, and phylogenetic implications

Plagiorchis elegans (Trematoda: Digenea) is mainly parasitic in the intestines of vertebrate animals, including humans, causing irreversible pathological damage and herd-spherical influences. However, little information is available about its molecular epidemiology, population genetics, and phylogeny. In the present study, we sequenced, assembled, and annotated the complete mitochondrial (mt) genome of P. elegans. Combining with the available mitochondrial data of subclass Digenea, phylogenetic analysis was performed based on Bayesian inference (BI). These results showed that the complete length of P. elegans is 13,862 bp, including 12 protein-coding genes (PCGs), 2 ribosomal RNAs (rRNAs), 22 transfer RNAs (tRNAs), and one non-coding region (NCR). There was an obvious A + T content from 61.0% to 71.3% and the values of the Ka/Ks ratio ranged from 0.119 (cox1) to 1.053 (nad6). In the BI analysis, different from previous studies, phylogenetic analysis showed genus Glypthelmins was paraphyletic rather than monophyletic and had a closer relationship with Plagiorchis and Orientocreadium. Additionally, the BI tree also presented that the genus Echinostoma was monophyletic. Our results provided molecular data in the family Plagiorchiidae proposing new insight within Xiphidiata and Echinostomata.

The earliest beetle †Coleopsis archaica (Insecta: Coleo­ptera) – morphological re-evaluation using Reflectance Transformation Imaging (RTI) and phylogenetic assessment

The earliest known fossil beetle †Coleopsis archaicais re-examined using Reflectance Transformation Imaging (RTI). The morphological observations are evaluated with respect to phylogenetic implications and the early evolution of Coleoptera. †Coleopsis archaicabelongs to an early Permian branch of beetles, outside a monophyletic unit comprising Coleoptera (in the widest sense) excluding †Tshekardocoleidae. This clade is mainly characterized by a complex of apomorphic features: elytra with epipleura and with a close fit with the posterior body, thus forming a tightly sealed subelytral space. In contrast to this, the elytra of †C. archaicaand †Tshekardocoleidae cover the metathorax and abdomen in a loose tent-like manner and posteriorly distinctly surpass the abdominal apex. So far, no synapomorphies of the two taxa from the first half of the Permian have been identified. The very short and transverse pronotum is likely an autapomorphy of †C. archaica. A thorough documentation of the structural features of early beetle fossils should have high priority. RTI is a very promising tool to obtain new and well-founded morphological data, which will allow a thorough phylogenetic evaluation of Permian beetles in future studies. We extended the conventional RTI workflow by focus merging and panoramic stitching, in order to overcome previous limitations. Taxonomic re-arrangements of stem group beetles including †C. archaicawere suggested in recent studies by A.G. Kirejtshuk and co-workers. As they are not based on shared derived features they are irrelevant in a phylogenetic and evolutionary context.

Cranial osteology of a juvenile specimen of Acratocnus ye (Mammalia, Xenarthra, Folivora) and its ontogenetic and phylogenetic implications.

The present study comprises a description of the skull and jaw anatomy of a juvenile specimen of the Antillean sloth Acratocnus ye, from the Holocene of Haiti. Detailed descriptions and illustrations are provided of the skull bones and their sutural connections, which normally fuse in adults. Descriptions are also provided for the mandible and ear ossicles, as well as endocranial surfaces and sinuses exposed by breaks. The anatomy of our juvenile A. ye is compared to that of adult A. ye to assess ontogenetic changes in the skull. Several of these ontogenetic features are significant new observations that impact the relationships within Xenarthra as a whole, or between Xenarthrans and other placental mammals, most notably, the presence of a separate mesethmoid element, the presence of alveoli for a lower deciduous canine and anterior incisor, and the presence of separate rostral and caudal entotympanic elements. A full list of such changes are provided. In addition, the specimen provides information on phylogenetically relevant characters, including features unique to the genus Acratocnus, and features of the clade Choloepodini, including Acratocnus, the smaller extinct Antillean sloth Neocnus, and the extant two-toed sloth Choloepus. Contrary to previous studies, Acratocnus shares as many features with Choloepus as it does with its fellow Antillean form Neocnus in the present study, which is consistent with current morphology-based phylogenetic hypotheses regarding the relationships within Choloepodini. The current study highlights the need for further anatomical and phylogenetic investigations of Antillean sloths (Megalocnidae/Megalonychidae), and juvenile sloths in general.

Mitochondrial genomes of two wild silkmoths, Samia watsoni and Samia wangi (Lepidoptera: Saturniidae), and their phylogenetic implications

The wild silkmoth genus Samia HĂźbner, 1819 (Saturniidae) contains a number of economically important species in industrial silk production. However, the interspecific relationships within the genus remain unclear. We sequence the mitogenomes of Samia watsoni OberthĂźr, 1914 and Samia wangi Naumann & Peigler, 2001. Both mitogenomes are annotated and found to be cyclized, with 37 genes (13 PCGs, 2 rRNA genes and 22 tRNA genes). Using maximum likelihood and Bayesian inference methods, we analyze these mitogenomes together with a further 68 downloaded from GenBank (65 Bombycoidea and 5 Lasiocampidae as the outgroup) to investigate the phylogenetic relationships both within the genus and those among the three families of the 'SBS' group: Bombycidae, Saturniidae and Sphingidae. The results show that within Samia, S. ricini is closely related to S. canningi, and not S. cynthia of which it has previously been considered to be a subspecies. Although arguments have been proposed to treat S. ricini and S. canningi as conspecific, we choose to accept the morphological arguments and continue to treat them as two separate species. Samia watsoni is corroborated as the sister group of all other Samia species, but nevertheless should be included within Samia rather than being placed in its own monobasic genus. Our analysis recovers the following relationship among the three families of the 'SBS' group: (Saturniidae + (Bombycidae + Sphingidae)). This agrees with previous studies based on analysis of mitogenomes but continues to contradict the results derived from phylogenomic analysis of nuclear genomes.

Ventral Morphology of the Non-Trilobite Artiopod Retifacies abnormalis Hou, Chen & Lu, 1989, from the Early Cambrian Chengjiang Biota, China.

Simple SummaryThis study reveals the detailed ventral morphology of the enigmatic Cambrian non-biomineralized euarthropod Retifacies abnormalis from the early Cambrian of South China through the use of micro-computer tomography and three-dimensional modelling. The ventral morphology of R. abnormalis includes five pairs of cephalic appendages in three forms (one uniramous antenna pair, three uniramous appendage pairs, one biramous appendage pair), biramous trunk appendages with multilamellar exites, a variable number of pygidial appendage pairs (five or six) and a multi-articulated tailspine with two pairs of short lobe-like accessories. The new anatomical data inform the ecology and evolution of R. abnormalis in the broader evolutionary context of trilobite-like euarthropods known from sites of exceptional preservation. R. abnormalis possessed a higher degree of appendage differentiation along the body than initially thought, in parallel to similar discoveries of other trilobitomorphs such as Naraoia spinosa, Pygmaclypeatus daziensis, and Sinoburius lunaris. This discovery provides additional support to the hypothesis that early trilobitomorphs were ancestrally characterized by heteronomous ventral appendages with various degrees of functional specialization for feeding and respiration. The artiopodans represent a diverse group of euarthropods with a typically flattened dorsal exoskeleton that covers numerous pairs of biramous ventral appendages, and which are ubiquitous faunal components of the 518-million-year-old Chengjiang Lagerstätte in South China. Despite their abundance, several Chengjiang artiopodans remain poorly known, such as the large euarthropoda Retifacies abnormalis, Hou, Chen & Lu, 1989, which is distinguished by the presence of mesh-like ornamentation on its dorsal exoskeleton. Although only a few ventral details were described in a single study in 25 years, it has been frequently featured in phylogenetic analyses that explore the relationships between Cambrian euarthropods. Here, we employ micro-CT and fluorescent microphotography to investigate the exceptionally preserved ventral morphology of R. abnormalis and explore its phylogenetic implications through maximum parsimony and Bayesian inference. Detailed morphology revealed here better supports R. abnormalis as a sister group to the diminutive artiopod Pygmaclypeatus daziensis, also known from Chengjiang, and strengthens the close relationship of these taxa that have been suggested by previous studies as early-branching representatives of Trilobitomorpha. Cephalic appendages suggest this animal might be a scavenger, possibly feeding on soft-bodied organisms. Different pairs of pygidial appendages suggest an anamorphic post-embryonic ontogeny, which adds to the understanding of the developmental mode of Cambrian artiopods, and further supports the statement that post-hatching segment addition occurred in the ancestor of Euarthropoda.

Characterization of the complete mitochondrial genomes of Diplodiscus japonicus and Diplodiscus mehari (Trematoda: Diplodiscidae): Comparison with the members of the superfamily Paramphistomoidea and phylogenetic implication.

Diplodiscus japonicus and Diplodiscus mehari (Trematoda: Diplodiscidae) are two important parasites in wood frogs, which have large infection rates and essential importance of ecology, economy and society. In this study, the complete mitochondrial (mt) genomes of D. japonicus and D. mehari were sequenced, then compared with other related trematodes in the superfamily Paramphistomoidea. The complete circular mt sequence of D. japonicus and D. mehari were 14,210 bp and 14,179 bp in length, respectively. Both mt genomes comprised 36 functional subunits, consisting of 12 protein-coding genes (PCGs), two ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, and one non-coding region. The mt genes of D. japonicus and D. mehari were transcribed in the same direction, and the gene arrangements were identical to those of Paramphistomoidea trematodes. In the 12 PCGs, GTG was the most common initiation codon, whereas TAG was the most common termination codon. All tRNAs had a typical cloverleaf structure except tRNA Ser1. A comparison with related Paramphistomoidea trematode mt genomes suggested that the cox1 gene of D. mehari was the longest in these trematodes. Phylogenetic analyses revealed that Paramphistomoidea trematodes formed a monophyletic branch, Paramphistomidae and Gastrothylacidae were more closely related than Diplodiscidae. And the further analysis with Pronocephalata branch found that the flukes parasitic in amphibians (frogs) formed one group, and the flukes from ruminants (cattle, sheep, ect) formed another group. Our study demonstrated the importance of sequencing mt genomes of D. japonicus and D. mehari, which will provide significant molecular resources for further studies of Paramphistomoidea taxonomy, population genetics and systematics.

The complete chloroplast genome of Abies ernestii Rehder (Pinaceae) and its phylogenetic implications

Abies ernestii Rehder is endemic to the montane regions of Southwest China. Till now, phylogenetic relationships between A. ernestii and other closely related species remain unclear. In this study, we first characterized the complete chloroplast (cp) genome of A. ernestii. The whole cp genome was 121,841 bp in size, including one hundred and thirteen genes. Results of comparative cp genome revealed that only ycf1 and ycf2 was characterized by a considerable variation. Our phylogenetic analyses supported the monophyly of the genus Abies and revealed a clear separation between A. ernestii and A. chensiensis Tiegh. This study highlights the significance of using cp genomes to examine species boundaries among closely related fir species.

Comparative mitochondrial genome analysis of Sesarmidae and its phylogenetic implications

Comparative mitochondrial genome analysis of Sesarmidae and its phylogenetic implications

Insights into genome evolution, pan-genome, and phylogenetic implication through mitochondrial genome sequence of Naegleria fowleri species

In the current study, we have systematically analysed the mitochondrial DNA (mtDNA) sequence of Naegleria fowleri (N. fowleri) isolate AY27, isolated from Karachi, Pakistan. The N. fowleri isolate AY27 has a circular mtDNA (49,541 bp), which harbours 69 genes (46 protein-coding genes, 21 tRNAs and 2 rRNAs). The pan-genome analysis of N. fowleri species showed a Bpan value of 0.137048, which implies that the pan-genome is open. KEGG classified core, accessory and unique gene clusters for human disease, metabolism, environmental information processing, genetic information processing and organismal system. Similarly, COG characterization of protein showed that core and accessory genes are involved in metabolism, information storages and processing, and cellular processes and signaling. The Naegleria species (n = 6) formed a total of 47 gene clusters; 42 single-copy gene clusters and 5 orthologous gene clusters. It was noted that 100% genes of Naegleria species were present in the orthogroups. We identified 44 single nucleotide polymorphisms (SNP) in the N. fowleri isolate AY27 mtDNA using N. fowleri strain V511 as a reference. Whole mtDNA phylogenetic tree analysis showed that N. fowleri isolates AY27 is closely related to N. fowleri (Accession no. JX174181.1). The ANI (Average Nucleotide Identity) values presented a much clear grouping of the Naegleria species compared to the whole mtDNA based phylogenetic analysis. The current study gives a comprehensive understanding of mtDNA architecture as well as a comparison of Naegleria species (N. fowleri and N. gruberi species) at the mitochondrial genome sequence level.

Centres of endemism of Noctuidae (Lepidoptera) in the Palaearctic arid mountains: biogeographical and phylogenetic implications

The oreal fauna is connected with orographically limited non-arboreal habitats. Its chorological centres can be recognised by the high species-diversity of numerous typical genera, and by the accumulated occurrence of endemic species and/or subspecies of disjunct species. The oreal fauna is partitioned to the alpine type, as the faunal type of humid high-mountains with strong connections to the tundral zonobiome, and the xeromontane type, as the faunal type of arid high-mountains with close connections to the eremic zonobiomes. As the results of revisions of several Noctuinae genera, species groups and/or sister species were recognised and their distributions were mapped. The restricted areas of allopatric sister species, often described by us as new for science, fulfil the criteria of the “areas of endemism”. Core areas of the Palaearctic xeromontane Noctuidae, outlined by the distribution of endemic species, have been proven by the occurrence of allopatric subspecies of polytypic species, and/or by the presence of allopatric sister species. In the revised genera of Noctuidae several types of allopatric speciation have been identified based on the analysis of the areas of endemism and of vicariance patterns. As a result of these analyses, it is proved that allopatric sister species, as elementary monophyletic supraspecific units, are suitable for phylogenetic biogeographical surveys. Although the major part of the xeromontane fauna appears to be range-restricted, a considerable fraction of the species could have expanded into the steppic zonobiome due to adaptive changes of their life cycles. High diversity of cold-adapted species originated from the Sino-Himalayan mountains by passing two main filtercorridors. One track of this bifurcation was directed across the “Rhododendron-corridor” to the Holarctic taiga zone while the other one, across the “Xeromontane filter-corridor” to the mountain systems of Central and Inner Asia. This bifurcation becomes apparent from the taxonomic division of the genera, composing both of these main faunal types. Supposedly, the faunal movements of the xeromontanean species in the West Palaearctic had been shaped by the Messinian salinity crisis and, additionally, significantly influenced by the Mid-Pleistocene climatic transition which deeply transformed the zonality of the vegetation by cooling and aridisation of vast areas.

Complete mitochondrial genomes of two catfishes (Siluriformes, Bagridae) and their phylogenetic implications.

The mitochondrial genome (mitogenome) has been widely used as a molecular marker to investigate phylogenetic analysis and evolutionary history in fish. However, the study of mitogenomes is still scarce in the family Bagridae. In this study, the mitogenomes of Tachysurusbrachyrhabdion and T.gracilis were sequenced, annotated, and analyzed. The mitogenomes were found to be 16,532 bp and 16,533 bp, respectively, and each contained 37 typical mitochondrial genes, which are 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a control region. All PCGs begin with the codon ATG, except for the cytochrome c oxidase subunit 1 (COI) gene, while seven PCGs end with an incomplete termination codon. All tRNA genes can fold into their typical cloverleaf secondary structures, except for tRNASer(AGY), which lacks the dihydrouracil arm. The Ka/Ks ratios for all PCGs are far lower than one. Phylogenetic analyses based on Bayesian inference (BI) and maximum likelihood (ML) showed that the two clades in Bagridae excluded Ritarita. The monophyly of Tachysurus supports previous research and the traditional classification that Leiocassis, Pseudobagrus, Pelteobagrus, and Tachysurus belong to one genus (Tachysurus). These findings provide a phylogenetic basis for future phylogenetic and taxonomic studies of Bagridae.

Centres of endemism of Noctuidae (Lepidoptera) in the Palaearctic arid mountains: biogeographical and phylogenetic implications

The oreal fauna is connected with orographically limited non-arboreal habitats. Its chorological centres can be recognised by the high species-diversity of numerous typical genera, and by the accumulated occurrence of endemic species and/or subspecies of disjunct species. The oreal fauna is partitioned to the alpine type, as the faunal type of humid high-mountains with strong connections to the tundral zonobiome, and the xeromontane type, as the faunal type of arid high-mountains with close connections to the eremic zonobiomes. As the results of revisions of several Noctuinae genera, species groups and/or sister species were recognised and their distributions were mapped. The restricted areas of allopatric sister species, often described by us as new for science, fulfil the criteria of the “areas of endemism”. Core areas of the Palaearctic xeromontane Noctuidae, outlined by the distribution of endemic species, have been proven by the occurrence of allopatric subspecies of polytypic species, and/or by the presence of allopatric sister species. In the revised genera of Noctuidae several types of allopatric speciation have been identified based on the analysis of the areas of endemism and of vicariance patterns. As a result of these analyses, it is proved that allopatric sister species, as elementary monophyletic supraspecific units, are suitable for phylogenetic biogeographical surveys. Although the major part of the xeromontane fauna appears to be range-restricted, a considerable fraction of the species could have expanded into the steppic zonobiome due to adaptive changes of their life cycles. High diversity of cold-adapted species originated from the Sino-Himalayan mountains by passing two main filtercorridors. One track of this bifurcation was directed across the “Rhododendron-corridor” to the Holarctic taiga zone while the other one, across the “Xeromontane filter-corridor” to the mountain systems of Central and Inner Asia. This bifurcation becomes apparent from the taxonomic division of the genera, composing both of these main faunal types. Supposedly, the faunal movements of the xeromontanean species in the West Palaearctic had been shaped by the Messinian salinity crisis and, additionally, significantly influenced by the Mid-Pleistocene climatic transition which deeply transformed the zonality of the vegetation by cooling and aridisation of vast areas.

Morphometric analysis of lungfish endocasts elucidates early dipnoan palaeoneurological evolution.

The lobe-finned fish, lungfish (Dipnoi, Sarcoptergii), have persisted for ~400 million years from the Devonian Period to present day. The evolution of their dermal skull and dentition is relatively well understood, but this is not the case for the central nervous system. While the brain has poor preservation potential and is not currently known in any fossil lungfish, substantial indirect information about it and associated structures (e.g. labyrinths) can be obtained from the cranial endocast. However, before the recent development of X-ray tomography as a palaeontological tool, these endocasts could not be studied non-destructively, and few detailed studies were undertaken. Here, we describe and illustrate the endocasts of six Palaeozoic lungfish from tomographic scans. We combine these with six previously described digital lungfish endocasts (4 fossil and 2 recent taxa) into a 12-taxon dataset for multivariate morphometric analysis using 17 variables. We find that the olfactory region is more highly plastic than the hindbrain, and undergoes significant elongation in several taxa. Further, while the semicircular canals covary as an integrated module, the utriculus and sacculus vary independently of each other. Functional interpretation suggests that olfaction has remained a dominant sense throughout lungfish evolution, and changes in the labyrinth may potentially reflect a change from nektonic to near-shore environmental niches. Phylogenetic implications show that endocranial form fails to support monophyly of the 'chirodipterids'. Those with elongated crania similarly fail to form a distinct clade, suggesting these two paraphyletic groups have converged towards either head elongation or truncation driven by non-phylogenetic constraints.

Plastome evolution of Aeonium and Monanthes (Crassulaceae): insights into the variation of plastomic tRNAs, and the patterns of codon usage and aversion.

This study reported 13 new plastomes from Aeonium and Monanthes, and observed new markers for phylogeny and DNA barcoding, such as novel tRNA structures and codon usage bias and aversion. The Macaronesian clade of Crassulaceae consists of three genera: Aichryson, with about 15 species; Monanthes, with about 10 species; Aeonium, with about 40 species. Within this clade, Aeonium, known as "the botanical equivalent of Darwin's finches", is regarded as an excellent model plant for researching adaptive evolution. Differing from the well-resolved relationships among three genera of the Macaronesian clade, the internal branching patterns within the genus Aeonium are largely unclear. In this study, we first reported 13 new plastomes from genus Aeonium and the closely related genus Monanthes. We further performed comprehensive analyses of the plastomes, with focuses on the secondary structures of pttRNAs and the patterns of codon usage and aversion. With a typical circular and quadripartite structure, the 13 plastomes ranged from 149,900 to 151,030bp in size, and the unique pattern in IR junctions might become a family-specific marker for Crassulaceae species. Surprisingly, the π values of plastomes from Monanthes were almost twice those from Aeonium. Most importantly, we strongly recommend that highly polymorphic regions, novel putative pttRNA structures, patterns of codon usage bias and aversion derived from plastomes might have phylogenetic implications, and could act as new markers for DNA barcoding of plants. The results of phylogenetic analyses strongly supported a clear internal branching pattern in Macaronesian clade (represented by Aeonium and Monanthes), with higher nodal support values. The findings reported here will provide new insights into the variation of pttRNAs, and the patterns of codon usage and aversion of the family Crassulaceae.

The complete chloroplast genome of Gynura japonica and its phylogenetic implications

Gynura japonica (Asteraceae) is a folk herbal medicine with multi-pharmacological functions involving analgesic, hemostatic and antiangiogenic activities. The study was conducted to assemble the complete chloroplast (cp) genome of G. japonica through a genome-skimming approach. The assembled cp genome was 151,023 bp in size, with 62.8% AT content, consisting of a large single copy (LSC) of 83,185 bp, two copies of inverted repeat (IRa and IRb) of 24,847 bp, and a small single copy (SSC) of 18,144 bp. The cp genome of G. japonica contained 133 genes, including eight ribosomal RNA genes (rRNAs), 37 transfer RNA genes (tRNAs), 86 protein-coding genes (PCGs), and two pseudogenes (ψycf1 and ψrps19). Our phylogenomic analysis based on whole plastid genomes strongly supports G. japonica is a sister to the clade including Crassocephalum crepidioides and Jacobaea vulgaris.

Comparative mitogenomics of the genus Motacilla (Aves, Passeriformes) and its phylogenetic implications.

The genus Motacilla belongs to Motacillidae (Passeriformes), where mitochondrial features are poorly understood and phylogeny is controversial. Whole mitochondrial genome (mitogenome) data and large taxon sampling are considered to be ideal strategies to obtain this information. We generated four complete mitogenomes of M.flava, M.cinerea, M.alba and Dendronanthusindicus, and made comparative analyses of Motacilla species combined with mitogenome data from GenBank, and then reconstructed phylogenetic trees based on 37 mitochondrial genes. The mitogenomes of four mitogenome sequences exhibited the same gene order observed in most Passeriformes species. Comparative analyses were performed among all six sampled Motacilla mitogenomes. The complete mitogenomes showed A-skew and C-skew. Most protein-coding genes (PCGs) start with an ATG codon and terminate with a TAA codon. The secondary structures of RNAs were similar among Motacilla and Dendronanthus. All tRNAs except for trnS(agy) could be folded into classic clover-leaf structures. Three domains and several conserved boxes were detected. Phylogenetic analysis of 90 mitogenomes of Passeriformes using maximum likelihood (ML) and Bayesian inference (BI) revealed that Motacilla was a monophyletic group. Among Motacilla species, M.flava and M.tschutschensis showed closer relationships, and M.cinerea was located in a basal position within Motacilla. These data provide important information for better understanding the mitogenomic characteristics and phylogeny of Motacilla.

Complete mitochondrial genome of Cultellus attenuatus and its phylogenetic implications.

The mitochondrial genomes of three species in Solenoidea of Heterodonta have been reported, but the mitochondrial genes and phylogenetic relationships of Cultellus attenuatus, which also belongs to this superfamily and has high economic value, are unknown. The complete mitochondrial genome of C. attenuatus was sequenced and compared with mitogenomes of seven species of Heterodonta bivalve mollusks in GenBank. The mitochondrial genome of C. attenuatus has a length of 16,888bp and contains 36 genes, including 12 protein-coding genes, 2 ribosomal RNAs and 22 transfer RNAs. In comparison with C. attenuates, the mitochondrial genes of Sinonovacula constricta from the same family were not rearranged, but those of six other species from different families were rearranged to different degrees. The location, size, and composition of the largest noncoding regions in eight species suggested a closer relationship between C. attenuatus and S. constricta. The phylogenetic analysis showed that C. attenuatus and S. constricta belonging to Cultellidae cluster into one branch and that two species of Solenidae (Solen grandis and Solen strictus) clustered as their sister taxa. Overall, we used mitochondrial genome data to demonstrate that C. attenuatus and S. constricta exhibit the closest relationship in Heterodonta. These data and analyses provide new insights into the phylogenetic relationships in Heterodonta.

Ovary structure and symbiotic associates of a ground mealybug, Rhizoecus albidus (Hemiptera, Coccomorpha: Rhizoecidae) and their phylogenetic implications.

The ovary structure and the organization of its symbiotic system of the ground mealybug, Rhizoecus albidus (Rhizoecidae), were examined by means of microscopic and molecular methods. Each of the paired elongated ovaries of R. albidus is composed of circa one hundred short telotrophic-meroistic ovarioles, which are radially arranged along the distal part of the lateral oviduct. Analysis of serial sections revealed that each ovariole contains four germ cells: three trophocytes (nurse cells) occupying the tropharium and a single oocyte in the vitellarium. The ovaries are accompanied by giant cells termed bacteriocytes which are tightly packed with large pleomorphic bacteria. Their identity as Brownia rhizoecola (Bacteroidetes) was confirmed by means of amplicon sequencing and fluorescence in situ hybridization techniques. Moreover, to our knowledge, this is the first report on the morphology and ultrastructure of the Brownia rhizoecola bacterium. In the bacteriocyte cytoplasm bacteria Brownia co-reside with sporadic rod-shaped smaller bacteria, namely Wolbachia (Proteobacteria: Alphaproteobacteria). Both symbionts are transmitted to the next generation vertically (maternally), that is, via female germline cells. We documented that, at the time when ovarioles contain oocytes at the vitellogenic stage, these symbionts leave the bacteriocytes and move toward the neck region of ovarioles (i.e. the region between tropharium and vitellarium). Next, the bacteria enter the cytoplasm of follicular cells surrounding the basal part of the tropharium, leave them and enter the space between the follicular epithelium and surface of the nutritive cord connecting the tropharium and vitellarium. Finally, they gather in the deep depression of the oolemma at the anterior pole of the oocyte in the form of a 'symbiont ball'. Our results provide further arguments strongly supporting the validity of the recent changes in the classification of mealybugs, which involved excluding ground mealybugs from the Pseudococcidae family and raising them to the rank of their own family Rhizoecidae.

A Unifying Framework for Estimating Generation Time in Age-Structured Populations: Implications for Phylogenetics and Conservation Biology.

AbstractGeneration time is a measure of the pace of life and is used to describe processes in population dynamics and evolution. We show that three commonly used mathematical definitions of generation time in age-structured populations can produce different estimates of up to several years for the same set of life history data. We present and prove a mathematical theorem that reveals a general order relation among the definitions. Furthermore, the exact population growth rate at the time of sampling influences estimates of generation time, which calls for attention. For phylogenetic estimates of divergence times between species, included demographic data should be collected when the population growth rate for each species is most common and typical. In conservation biology, demographic data should be collected during phases of population decline in declining species, contrary to common recommendations to use predisturbance data. The results can be used to improve the International Union for Conservation of Nature's recommendation in parameterizing models for evaluating threat categories of threatened species and to avoid underestimating extinction risk.