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Abstract Quantifying the features of mitochondrial genome structural variation is crucial for understanding its contribution to complexity. Accurate quantification and interpretation of organizational diversity can help uncover biological evolutionary laws and patterns. The current qMGR approach accumulates the changes in two adjacent genes to calculate the rearrangement frequency RF of each single gene and the rearrangement score RS for specific taxa in the mitogenomes of a given taxonomic group. However, it may introduce bias, as it assigns scores to adjacent genes rather than to rearranged genes. To overcome this limitation, we propose a novel statistical method called qGO to quantify the diversity of gene organization. The qGO method, which is based on the homology of gene order, provides a more accurate representation of genome organizational diversity by partitioning gene strings and individually assigning weights to genes spanning different regions. Additionally, a comprehensive approach is employed for distance computation, generating an extensive matrix of rearrangement distances. Through experiments on more than 5500 vertebrate mitochondrial genomes, we demonstrated that the qGO method outperforms existing methods in terms of accuracy and interpretability. This method improves the comparability of genomes and allows a more accurate comparison of the diversity of mitochondrial genome organization across taxa. These findings have significant implications for unraveling genome evolution, exploring genome function, and investigating the process of molecular evolution.

Quantifying the features of mitochondrial genome structural variation is crucial for understanding its contribution to complexity. Accurate quantification and interpretation of organizational diversity can help uncover biological evolutionary laws and patterns. The current qMGR approach accumulates the changes in two adjacent genes to calculate the rearrangement frequency RF of each single gene and the rearrangement score RS for specific taxa in the mitogenomes of a given taxonomic group. However, it may introduce bias, as it assigns scores to adjacent genes rather than to rearranged genes. To overcome this limitation, we propose a novel statistical method called qGO to quantify the diversity of gene organization. The qGO method, which is based on the homology of gene order, provides a more accurate representation of genome organizational diversity by partitioning gene strings and individually assigning weights to genes spanning different regions. Additionally, a comprehensive approach is employed for distance computation, generating an extensive matrix of rearrangement distances. Through experiments on more than 5500 vertebrate mitochondrial genomes, we demonstrated that the qGO method outperforms existing methods in terms of accuracy and interpretability. This method improves the comparability of genomes and allows a more accurate comparison of the diversity of mitochondrial genome organization across taxa. These findings have significant implications for unraveling genome evolution, exploring genome function, and investigating the process of molecular evolution. [ABSTRACT FROM AUTHOR]

Invertebrate mitogenomes are generally fixed with formal 37 genes: 13 PCGs encoded subunits of OXPHOS, 2 ribosomal RNA (rRNA) functional in the translation of these PCGs and 22 transfer RNA (tRNA) genes. The order of these genes varies greatly among organisms and named rearrangement. Rearrangement patterns of mitochondrial genomes may shed light on mutation processes and evolutionary relationships of organisms. Mitochondrial gene organization is highly variable among Acari, so rearrangement is a very common mitogenomic pattern in this group. In this study, 258 unique Acari (Acariformes + Parasitiformes) mitogenomes were downloaded from NCBI and studied about rearrangement patterns. Sixty-seven mitotypes were determined among Acari and the most rearranged genes were trnL1 and nad2. Following that, trnI, trnS1, trnN, trnE, trnT, and trnP genes are remarkably mobile (RF > 95%). Conversely, atp6, cox3, trnG, and cytb genes also appears to be quite stable (RF < 20%). Within Acari, mean distance calculations are varied from 1.210 in atp8 to 0.155 in rrnS. Contrary to expectations, among Acari mobile tRNA genes appear to be conserved in sequences, whereas PCGs have higher distance values and seem to be mutated. Consistently, tRNA genes seem saturated, but some PCGs (atp6, cox genes, cytb, nad1, and nad6) are not saturated. These values do not correlate with each other (p > 0.005). This discrepancy may indicate that the genes were rearranged after mutation load; consistent with this, DAMBE saturation values are also not correlated with RF values. Parasitiformes mitogenomes are more mobile than Acariformes mitogenomes and may be under the effect of selective sweeping. [ABSTRACT FROM AUTHOR]

The longhand rubble crab Ratha longimana inhabits coral reefs and rocky shores in the subtropical and tropical western Atlantic Ocean. This species was long considered to be the only chlorodielline species present in the area, but has since been transferred to the genus Ratha Lasley, Lai & Thoma, 2013 and is now thought to belong to an undescribed subfamily within Xanthidae MacLeay, 1838. Here we provide a complete mitochondrial assemblage and characterisation of this species. The assembled mitochondrial genome (hereafter mitogenome) of R. longimana is 15,819 bp in length and contains 13 protein coding genes (PCGs), 22 transfer RNA (tRNA) genes, and two ribosomal RNA (rRNA) genes. The 22 tRNA genes range from 62 to 69 bp in length. A mitochondrial gene order (MGO) analysis revealed a transposition of the tRNA genes Val (V) and 12S rRNA (rrnS) when compared to the ancestral brachyuran gene order, which is in line with earlier studies on xanthoid mitogenomes. Mitogenome data for xanthid crabs (as well as the superfamily Xanthoidea MacLeay, 1838) are currently scarce considering the species richness of the family, limiting the options for phylogenomic studies. This is partially remedied here by providing a new genomic resource for R. longimana. [ABSTRACT FROM AUTHOR]

While originally intending to explore the venom gland microbiome of the desert hairy scorpion Hadrurus arizonensis Ewing, 1928, nanopore sequencing serendipitously recovered complete mitochondrial genomes for this iconic arachnid. Phylogenetic analysis of these high-quality genomes places Hadrurus as sister to Uroctonus, in agreement with some phylogenomic hypotheses. Additionally, we reveal significant genetic variation among individuals from the same population, highlighting the potential of mitogenomics for population genetics and phylogeography. This study showcases the effectiveness and affordability of nanopore sequencing for research with non-model organisms, opening new avenues for investigating arachnid biodiversity, evolution, and biogeography. [ABSTRACT FROM AUTHOR]

Abstract Background Eriophyoid mites (Eriophyoidea) are among the largest groups in the Acariformes; they are strictly phytophagous. The higher-level phylogeny of eriophyoid mites, however, remains unresolved due to the limited number of available morphological characters—some of them are homoplastic. Nevertheless, the eriophyoid mites sequenced to date showed highly variable mitochondrial (mt) gene orders, which could potentially be useful for resolving the higher-level phylogenetic relationships. Results Here, we sequenced and compared the complete mt genomes of 153 eriophyoid mite species, which showed 54 patterns of rearranged mt gene orders relative to that of the hypothetical ancestor of arthropods. The shared derived mt gene clusters support the monophyly of eriophyoid mites (Eriophyoidea) as a whole and the monophylies of six clades within Eriophyoidea. These monophyletic groups and their relationships were largely supported in the phylogenetic trees inferred from mt genome sequences as well. Our molecular dating results showed that Eriophyoidea originated in the Triassic and diversified in the Cretaceous, coinciding with the diversification of angiosperms. Conclusions This study reveals multiple molecular synapomorphies (i.e. shared derived mt gene clusters) at different levels (i.e. family, subfamily or tribe level) from the complete mt genomes of 153 eriophyoid mite species. We demonstrated the use of derived mt gene clusters in unveiling the higher-level phylogeny of eriophyoid mites, and underlines the origin of these mites and their co-diversification with angiosperms.

AbstractThe order Hymenoptera is one of the most species-rich insect orders, with more than 150,000 described extant species. Many hymenopteran insects have very different mitochondrial genome (mitogenome) organizations compared to the putative ancestral organization of insects. In this study, we sequenced 18 mitogenomes of representatives in the order Hymenoptera to increase taxonomic sampling. A total of 475 species were used in phylogenetic analyses, including 18 new mitogenomes and 457 existing mitogenomes. Using a site-heterogeneous model, Bayesian’s inference from amino acid data yielded more resolved relationships among Hymenoptera than maximum-likelihood analysis and coalescent-based species analyses. The monophyly of Symphyta was not supported. The Xyeloidea was the earliest branching clade in the Hymenoptera. The Orussoidea was closely related to Apocrita. Within Apocrita, the Parasitoida was non-monophyletic. The monophyly of most Parasitoida superfamilies received strong support. The Proctotrupomorpha clade was supported in Bayesian’s analysis. The Apoidea was monophyletic when excluding Ampulex compressa from consideration. The superfamilies Vespoidea and Chrysidoidea were found to be non-monophyletic. Comparisons of mitochondrial gene order revealed a higher frequency of gene rearrangement among lineages with a parasitoid lifestyle, particularly prominent in Chalcidoidea. The degree of gene rearrangement ranked second in specific taxa of Cynipoidea and Ichneumonoidea. [ABSTRACT FROM AUTHOR]

Background: Eriophyoid mites (Eriophyoidea) are among the largest groups in the Acariformes; they are strictly phytophagous. The higher-level phylogeny of eriophyoid mites, however, remains unresolved due to the limited number of available morphological characters—some of them are homoplastic. Nevertheless, the eriophyoid mites sequenced to date showed highly variable mitochondrial (mt) gene orders, which could potentially be useful for resolving the higher-level phylogenetic relationships. Results: Here, we sequenced and compared the complete mt genomes of 153 eriophyoid mite species, which showed 54 patterns of rearranged mt gene orders relative to that of the hypothetical ancestor of arthropods. The shared derived mt gene clusters support the monophyly of eriophyoid mites (Eriophyoidea) as a whole and the monophylies of six clades within Eriophyoidea. These monophyletic groups and their relationships were largely supported in the phylogenetic trees inferred from mt genome sequences as well. Our molecular dating results showed that Eriophyoidea originated in the Triassic and diversified in the Cretaceous, coinciding with the diversification of angiosperms. Conclusions: This study reveals multiple molecular synapomorphies (i.e. shared derived mt gene clusters) at different levels (i.e. family, subfamily or tribe level) from the complete mt genomes of 153 eriophyoid mite species. We demonstrated the use of derived mt gene clusters in unveiling the higher-level phylogeny of eriophyoid mites, and underlines the origin of these mites and their co-diversification with angiosperms. [ABSTRACT FROM AUTHOR]

Pilargidae is a family of free-living and burrowing marine annelids. A lack of available molecular data for most of these species has precluded a molecular assessment of their phylogenetic relationships and has left uncertain the placement of Antonbruunia, which is hypothesized to be either a member of Pilargidae or its sister clade, the monotypic family Antonbruunidae. In this study, we describe the new species Antonbruunia milenae sp. nov., found at 845 m of depth off the coast of San Diego, California, USA, and we address the phylogeny of these organisms using 15 novel mitogenomes and multiple Sanger-sequenced loci. Our results show that Antonbruunia falls within Pilargidae, making Antonbruunidae a junior synonym of Pilargidae. Glyphohesione was transferred from Pilarginae to Synelminae, the previously unassigned genera Otopsis and Antonbruunia were shown to belong within Synelminae, and Hermundura was assigned to Phyllodocida incertae sedis. Sigambra was found to be non-monophyletic. Four different mitogenome gene orders were found among Pilargidae. Changes between the gene orders and the ancestral state gene order of the family were inferred. Two species have introns within the COI gene. These efforts represent a significant expansion of the available molecular resources for pilargids, as well as the basis for a more stable taxonomy. [ABSTRACT FROM AUTHOR]

ABSTRACT On circular bacterial chromosomes, the majority of genes are coded on the leading strand. This gene strand bias (GSB) ranges from up to 85% in some Bacillota to a little more than 50% in other phyla. The factors determining the extent of the strand bias remain to be found. Here, we report that species in the phylum Gemmatimonadota share a unique chromosome architecture, distinct from neighboring phyla: in a conserved 600-kb region around the terminus of replication, almost all genes were located on the leading strands, while on the remaining part of the chromosome, the strand preference was more balanced. The high strand bias (HSB) region harbors the rRNA clusters, core, and highly expressed genes. Selective pressure for reduction of collisions with DNA replication to minimize detrimental mutations can explain the conservation of essential genes in this region. Repetitive and mobile elements are underrepresented, suggesting reduced recombination frequency by structural isolation from other parts of the chromosome. We propose that the HSB region forms a distinct chromosomal domain. Gemmatimonadota chromosomes evolved mainly by expansion through horizontal gene transfer and duplications outside of the ancient high strand bias region. In support of our hypothesis, we could further identify two Spiroplasma strains on a similar evolutionary path.IMPORTANCEOn bacterial chromosomes, a preferred location of genes on the leading strand has evolved to reduce conflicts between replication and transcription. Despite a vast body of research, the question why bacteria show large differences in their gene strand bias is still not solved. The discovery of “hybrid” chromosomes in different phyla, including Gemmatimonadota, in which a conserved high strand bias is found exclusively in a region at ter, points toward a role of nucleoid structure, additional to replication, in the evolution of strand preferences. A fine-grained structural analysis of the ever-increasing number of available bacterial genomes could help to better understand the forces that shape the sequential and spatial organization of the cell’s information content.

Here, we present a karyotype, a chromosome-scale genome assembly, and a genome annotation from the ctenophore Hormiphora californensis (Ctenophora: Cydippida: Pleurobrachiidae). The assembly spans 110 Mb in 44 scaffolds and 99.47% of the bases are contained in 13 scaffolds. Chromosome micrographs and Hi-C heatmaps support a karyotype of 13 diploid chromosomes. Hi-C data reveal three large heterozygous inversions on chromosome 1, and one heterozygous inversion shares the same gene order found in the genome of the ctenophore Pleurobrachia bachei. We find evidence that H. californensis and P. bachei share thirteen homologous chromosomes, and the same karyotype of 1n = 13. The manually curated PacBio Iso-Seq-based genome annotation reveals complex gene structures, including nested genes and trans-spliced leader sequences. This chromosome-scale assembly is a useful resource for ctenophore biology and will aid future studies of metazoan evolution and phylogenetics.

We report that epigenetic silencing causes the loss of function of multi-transcript unit constructs that are integrated using CRISPR-Cas9. Using a modular two color reporter system flanked by selection markers, we demonstrate that expression heterogeneity does not correlate with sequence alteration but instead correlates with chromosomal accessibility. We partially reverse this epigenetic silencing via small-molecule inhibitors of methylation and histone deacetylation. We then correlate each heterogeneously-expressing phenotype with its expected epigenetic state by employing ATAC-seq. The stability of each expression phenotype is reinforced by selective pressure, which indicates that ongoing epigenetic remodeling can occur for over one month after integration. Collectively, our data suggests that epigenetic silencing limits the utility of multi-transcript unit constructs that are integrated via double-strand repair pathways. Our research implies that mammalian synthetic biologists should consider localized epigenetic outcomes when designing complex genetic circuits.

Nereididae is one of Annelida's most diverse and well-studied families; despite this, only recently have molecular approaches been used to access phylogenetic hypotheses within the family. At least two nereidid groups can be identified based on mitochondrial gene order; however, this type of molecular information is not available for most taxa, and relationships of subfamilies are still poorly understood. In the present study, we used nuclear and mitochondrial markers to evaluate relationships within Nereididae and verified hypotheses on subfamilies delineation and relationship. Our results suggest the paraphyletic status of Gymnonereidinae and Nereidinae, as well as some genera within these subfamilies. Despite this, a revision of these subfamilies delineation and diagnoses was presented to match current phylogenetic understanding of the group. We suggest reverting to the original definition of Gymnonereidinae to include only the genera originally proposed by Banse in 1977. Our results also support monophyly of a large group that can be defined by mitochondrial gene order. This group includes most taxa identified as Nereidinae, and therefore, we suggest that this feature should be used as diagnostic of the subfamily. [ABSTRACT FROM AUTHOR]

The elkhorn coral crab Domecia acanthophora inhabits shallow-water coral reefs in the Western Atlantic. The species has a wide distribution and, although primarily associated with endangered Acropora corals, has been recorded from a myriad of hosts. Here we conducted the first genomic survey and complete mitochondrial assemblage and characterisation of any species of Domeciidae, as well as the first species within Trapezioidea. The estimated size of the nuclear genome ranged from 0.64 Gbp to 1.76 Gbp, revealing a small genome. Repetitive elements of the genome were estimated here at 66.4% and 74%, respectively, with the majority of the repetitive elements consisting of LINE, LTR, and satellite DNA. The assembled A-T rich mitochondrial genome consisted of 15,568 bp in length, with 13 protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes and two ribosomal RNA (rRNA) genes. A 619 bp long non-coding region was identified as the supposed D-loop/control region, containing eight microsatellites. The 22 tRNA genes, ranging from 65 to 71 bp in length, displayed a typical "cloverleaf" secondary structure, with the exception of tRNA-Ser1 which lacked part of the DHU arm and tRNA-Asp displayed a deletion of the TΨC loop but not the arm. Two transposition events of two tRNA genes were also found when comparing the gene order of D. acanthophora to that of the brachyuran basic gene order, which had not been reported before. Despite belonging to a widely distributed, well-known superfamily of coral-associated crabs, the Trapezioidea, very little was known about this species from a genetics perspective, which is remedied here by providing a new genomic resource for D. acanthophora. [ABSTRACT FROM AUTHOR]

Precisely altering gene expression is critical for understanding molecular processes of embryogenesis. Although some tools exist for transgene misexpression in developing chick embryos, we have refined and advanced them by simplifying and optimizing constructs for spatiotemporal control. To maintain expression over the entire course of embryonic development we use an enhanced piggyBac transposon system that efficiently integrates sequences into the host genome. We also incorporate a DNA targeting sequence to direct plasmid translocation into the nucleus and a D4Z4 insulator sequence to prevent epigenetic silencing. We designed these constructs to minimize their size and maximize cellular uptake, and to simplify usage by placing all of the integrating sequences on a single plasmid. Following electroporation of stage HH8.5 embryos, our tetracycline-inducible promoter construct produces robust transgene expression in the presence of doxycycline at any point during embryonic development in ovo or in culture. Moreover, expression levels can be modulated by titrating doxycycline concentrations and spatial control can be achieved using beads or gels. Thus, we have generated a novel, sensitive, tunable, and stable inducible-promoter system for high-resolution gene manipulation in vivo.

Different from the true oyster (family Ostreidae), the molecular diversity of the gryphaeid oyster (family Gryphaeidae) has never been sufficiently investigated. In the present study, the complete mitochondrial (mt) genome of Hyotissa sinensis was sequenced and compared with those of other ostreoids. The total length of H. sinensis mtDNA is 30,385 bp, encoding 12 protein-coding-genes (PCGs), 26 transfer RNA (tRNA) genes and two ribosomal RNA (rRNA) genes. The nucleotide composition and codon usage preference of H. sinensis mtDNA is similar to that of H. hyotis within the same genus. On the other hand, the presence of three trnM and three trnL genes of H. sinensis was not detected neither in H. hyotis nor other ostroid species. Another unique character of H. sinensis mtDNA is that both rrnS and rrnL have a nearly identical duplication. The PCG order of H. sinensis is identical to H. hyotis and the two congener species also share an identical block of 12 tRNA genes. The tRNA rearrangements mostly happen in the region from Cox1 to Nad3, the same area where the duplicated genes are located. The rearrangements within Gryphaeidae could be explained by a "repeat-random loss model". Phylogenetic analyses revealed Gryphaeidae formed by H. sinensis + H. hyotis as sister to Ostreidae, whereas the phylogenetic relationship within the latter group remains unresolved. The present study indicated the mitogenomic diversity within Gryphaeidae and could also provide important data for future better understanding the gene order rearrangements within superfamily Ostreoidea.

The present paper reports the first occurrence of Seira ferrarii Parona, 1888 from Hungary. On this occasion, molecular analyses were performed on both S. ferrarii and another historical species of the genus, S. pallidipes Reuter, 1895, originally described from Hungary. Using low-coverage whole-genome sequencing, the complete mitogenomes were assembled and annotated using MitoZ. To test the phylogenetic placement of both species, we performed maximum likelihood and Bayesian analyses using a matrix containing 14 Seirinae species and two outgroups. Both resultant trees showed that the European populations of the sampled Seira spp. likely derive from ancestral branches of Seirinae, compared to the Asian and American populations. Our results put in question the monophyly of the genus Seira, as already observed in previous studies. [ABSTRACT FROM AUTHOR]

We analysed the mitochondrial genome of Aristaeopsis edwardsiana (Johnson, 1868) to understand its origin and evolution. Our study uncovered new insights into its phylogenetic relationships and evolutionary history. The mitogenome of A. edwardsiana is 15 913 bp, contains 37 genes, including 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (rRNA) and 22 transfer RNA genes (tRNA). The AT content is 64.98%, with negative AT and GC skews. Our analysis showed that the PCGs of A. edwardsiana and related Aristeidae species underwent purifying selection. The tRNA genes exhibited a typical structure, except trnC and trnS1. The gene order was conserved compared to ancestral decapods. Using species of Dendrobranchiata, we constructed a phylogenetic tree, revealing the relationship of A. edwardsiana within the family Aristeidae. These findings improve our understanding of Decapoda classification and Aristeidae's evolutionary position, facilitating further phylogenetic investigations. This study may serve as a valuable reference for future research on species of Aristeidae. [ABSTRACT FROM AUTHOR]

Cnidarians are the most primitive metazoans, but their evolutionary relationships are poorly understood, although recent studies present several phylogenetic hypotheses. Here, we collected 266 complete cnidarian mitochondrial genomes and re‐evaluated the phylogenetic relationships between the major lineages. We described the gene rearrangement patterns of Cnidaria. Anthozoans had significantly greater mitochondrial genome size and lower A + T content than medusozoans. Most of the protein‐coding genes in anthozoans such as COX 13, ATP6, and CYTB displayed a faster rate of evolution based on selection analysis. There were 19 distinct patterns of mitochondrial gene order, including 16 unique gene orders in anthozoans and 3 mtDNA gene orders pattern in medusozoans, were identified among cnidarians. The gene order arrangement suggested that a linearized mtDNA structure may be more conducive to Medusozoan mtDNA stability. Based on phylogenetic analyses, the monophyly of the Anthozoa was strongly supported compared to previous mitochondrial genome‐based analyses rather than octocorals forming a sister group relationship with medusozoans. In addition, Staurozoa were more closely related to Anthozoa than to Medusozoa. In conclusion, these results largely support the traditional phylogenetic view of the relationships of cnidarians and provide new insights into the evolutionary processes for studying the most ancient animal radiations. [ABSTRACT FROM AUTHOR]

Homing based gene drives (HGD) possess the potential to spread linked cargo genes into natural populations and are poised to revolutionize population control of animals. Given that host encoded genes have been identified that are important for pathogen transmission, targeting these genes using guide RNAs as cargo genes linked to drives may provide a robust method to prevent disease transmission. However, effectiveness of the inclusion of additional guide RNAs that target separate genes has not been thoroughly explored. To test this approach, we generated a split-HGD in Drosophila melanogaster that encoded a drive linked effector consisting of a second gRNA engineered to target a separate host-encoded gene, which we term a gRNA-mediated effector (GME). This design enabled us to assess homing and knockout efficiencies of two target genes simultaneously, and also explore the timing and tissue specificity of Cas9 expression on cleavage/homing rates. We demonstrate that inclusion of a GME can result in high efficiency of disruption of both genes during super-Mendelian propagation of split-HGD. Furthermore, both genes were knocked out one generation earlier than expected indicating the robust somatic expression of Cas9 driven by Drosophila germline-limited promoters. We also assess the efficiency of 'shadow drive' generated by maternally deposited Cas9 protein and accumulation of drive-induced resistance alleles along multiple generations, and discuss design principles of HGD that could mitigate the accumulation of resistance alleles while incorporating a GME.

The editorial in Frontiers in Genetics discusses comparative and evolutionary analyses of organelle genomes, focusing on studies of chloroplast and mitochondrial genomes in non-model species. The paper highlights genomic rearrangements and gene evolution in parasitic plants like Love Vines and the impact of a parasitic lifestyle on chloroplast genomes. Additionally, the editorial covers the sequencing of the mitochondrial genome of the Asian freshwater fish Osteochilus salsburyi and the evolution of the CHS gene in plants. Lastly, it explores population genetics in rats, specifically the Brown Rat and Oriental Rat, to understand genetic diversity and migration patterns. The research topic provides insights into the potential of organelle genome analyses for scientific studies and highlights the importance of new sequencing technologies for future research. [Extracted from the article]

Mitochondrial genome (mitogenome) is a frequently-used tool for phylogenetic and evolutionary studies among metazoans, however, it was still poorly represented in some invertebrate groups, including holothurians. Apodida is one of the most ancient orders of Holothuroidea, with unique characteristics in lacking tube feet and the respiratory tree. Here, we sequenced six mitogenomes from the holothurian order Apodida (Synaptidae and Chiridotidae) and compared them with other available holothurian mitogenomes. The apodan mitogenomes present positive GC skews and negative AT skews, a pattern diametrically opposite to that found in the mitogenomes of other holothurians. The amino acid genetic distances of the 13 protein-coding genes (PCGs) were the highest between Apodida and other holothurians. The mitochondrial gene orders within Apodida were extensively rearranged at the species level, and also were unique from the arrangement patterns of other holothurians. Phylogenetic relationships based on two different criteria: (i) nucleotides sequences with all codon positions (PCG123) and (ii) Neutral Transitions Excluded model (NTE) confirmed the monophyly and basal position of the order Apodida with a long branch. The NTE phylogeny generated a low variation of branch length, but it did not ameliorate the long branch length of Apodida. The divergence time estimation suggested that the Apodida originated in the Middle Carboniferous, implying that they survived the extinction event that occurred in the boundary of the Permian and Triassic. The NTE dataset revealed younger age than that generated by PCG123 dataset. Apodida is accompanied by a faster evolutionary rate than other holothurians (0.0069 vs 0.0043 subs/s/my). The absence of tube feet and the respiratory tree may play an important role in the dramatic evolutionary changes of apodan mitogenomes.

Simple Summary: Coccoidea (scale insects) are sap-sucking hemipterous insects with a high diversity of species. They exhibit dramatically variable appearance and sexual dimorphism and are also closely related to human beings as important economic pests and resource insects. However, we know very little about the phylogenetic relationship within Coccoidea. We reconstructed that relationship among five coccoid families (e.g., Aclerdidae, Cerococcidae, Coccidae, Eriococcidae, and Kerriidae) based on mitogenomes. Aclerdidae and Coccidae were recovered as the sister group, successively sister to Cerococcidae, Kerriidae, and Eriococcidae. In addition, there were gene rearrangements occurring in all mitogenomes of coccoid species studied here. The novel gene rearrangement ND6-trnP and trnI-ND2-trnY supported the monophyly of Coccoidea and the sister relationship of Aclerdidae and Coccidae. Coccoidea (scale insects) are important plant parasites with high diversity of species. However, the phylogenetic relationship within Coccoidea has not been fully determined. In this study, we sequenced mitogenomes of six species belonging to five coccoid families. With the addition of three previously published mitogenomes, a total of 12 coccoid species were adopted for the phylogenetic reconstruction based on the maximum likelihood and Bayesian inference. The monophyly of Coccoidea was recovered and Aclerdidae and Coccidae were recovered as the sister group, successively sister to Cerococcidae, Kerriidae, and Eriococcidae. In addition, there were gene rearrangements occurring in all mitogenomes of coccoid species studied here. The novel gene rearrangement ND6-trnP and trnI-ND2-trnY supported the monophyly of Coccoidea and the sister relationship of Aclerdidae and Coccidae. This implies that data from the mitogenome can provide new insight for clarifying the deeper level of phylogenetic relationship within Coccoidea. [ABSTRACT FROM AUTHOR]

Peptide infusions of peptides the corticotropin releasing factor family, including urocortin 2, stresscopin, and urocortin 3 (UCn3), have favorable acute effects in clinical heart failure (HF), but their short half-lives make them unsuitable for chronic therapy. This study asked whether UCn3 gene transfer, which provides sustained elevation of plasma UCn3 levels, increases the function of the failing heart. HF was induced by transmural left ventricular (LV) cryoinjury in mice. LV function was assessed 3 weeks later by echocardiography. Those with ejection fractions (EF)

The complete nucleotide sequence of the mitochondrial (mt) genome of the demersal zebra seabream Diplodus cervinus (Lowe, 1838) was determined for the first time. The double stranded circular molecule is 16,559 base pairs (bp) in length and encodes for the typical 37 metazoan mitochondrial genes, and 2 non-coding regions (D-loop and L-origin). The gene arrangement of the D. cervinus mt genome follows the usual one for fishes. The nucleotide sequences of the mt protein coding and ribosomal genes of D. cervinus mt genome were aligned with orthologous sequences from representatives of the Sparidae family and phylogenetic relationships were inferred. Maximum likelihood analyses placed D. cervinus as a sister species of Diplodus sargus (Linnaeus, 1758).

Piper nigrum, also known as black pepper, is an economically and ecologically important crop of the genus Piper. It has been titled as the king of spices due to its wide consumption throughout the world. In the present investigation, the chloroplast genome of P. nigrum has been assembled from a whole genome sequence by integrating the short and long reads generated through Illumina and PacBio platforms, respectively. The chloroplast genome was observed to be 161,522 bp in size, having a quadripartite structure with a large single copy (LSC) region of 89,153 bp and a small single copy (SSC) region of 18,255 bp separated by a copy of inverted repeats (IRs), each 27,057 bp in length. Taking into consideration all the duplicated genes, a total of 131 genes were observed, which included 81 protein-coding genes, 37 tRNAs, 4 rRNAs, and 1 pseudogene. Individually, the LSC region consisted of 83 genes, the SSC region had 13 genes, and 18 genes were present in each IR region. Additionally, 216 SSRs were detected and 11 of these were validated through amplification in 12 species of Piper. The features of the chloroplast genome have been compared with those of the genus Piper. Our results provide useful insights into evolutionary and molecular studies of black pepper which will contribute to its further genetic improvement and breeding. [ABSTRACT FROM AUTHOR]

Containing the largest number of species, the orchid family provides not only materials for studying plant evolution and environmental adaptation, but economically and culturally important ornamental plants for human society. Previously, we collected genome and transcriptome information of Dendrobium catenatum, Phalaenopsis equestris, and Apostasia shenzhenica which belong to two different subfamilies of Orchidaceae, and developed user-friendly tools to explore the orchid genetic sequences in the OrchidBase 4.0. The OrchidBase 4.0 offers the opportunity for plant science community to compare orchid genomes and transcriptomes and retrieve orchid sequences for further study. In the year 2022, two whole-genome sequences of Orchidoideae species, Platanthera zijinensis and Platanthera guangdongensis, were de novo sequenced, assembled and analyzed. In addition, systemic transcriptomes from these two species were also established. Therefore, we included these datasets to develop the new version of OrchidBase 5.0. In addition, three new functions including synteny, gene order, and miRNA information were also developed for orchid genome comparisons and miRNA characterization. OrchidBase 5.0 extended the genetic information to three orchid subfamilies (including five orchid species) and provided new tools for orchid researchers to analyze orchid genomes and transcriptomes. The online resources can be accessed at https://cosbi.ee.ncku.edu.tw/orchidbase5/ [ABSTRACT FROM AUTHOR]

Background: Caryophylliidae is one of the most diverse scleractinian families, however it was recovered as polyphyletic in multiple molecular studies. Recently, the mitochondrial gene order was proposed as a character for a taxonomic revision of the family. Here we describe the first mitogenome of the caryophylliid genus Crispatotrochus, whose phylogenetic position remains uncertain. Methods and results: The complete mitochondrial genomes of Crispatotrochus rubescens and Crispatotrochus rugosus were sequenced, assembled, and annotated. The two mitogenomes are identical and circular, have a length of 16,536 bp, a GC content of 35.9%, and contain 13 protein-coding genes, 2 ribosomal RNAs and 2 transfer RNAs. Both species have a transposition of a three gene block - cob, nad2, and nad6 - similarly to a group of caryophylliid genera that were recovered as monophyletic, including the type genus (Caryophyllia) of the family. The phylogenetic analyses recovered Crispatotrochus within the clade that presents the gene rearrangement and specifically as sister taxa of the genus Caryophyllia, a result consistent with previous studies and the similar gross morphology of the two genera. Conclusions: We determined the mitochondrial genomes of the genus Crispatotrochus to investigate their relations within Scleractinia. Results from this study provide insights on the phylogenetic position of the genus and corroborate that the mitochondrial gene order could be used as taxonomic character for the family Caryophylliidae. [ABSTRACT FROM AUTHOR]

Both regulatory sequences and genome organization contribute to the production of diverse transcript isoforms, which can influence how genes, or sets of genes, are expressed. An efficient, modular approach is needed to generate the combinatorial complexity required to empirically test many combinations of different regulatory sequences and different gene orders. Golden Gate assembly provides such a tool for seamless one-pot cleavage and ligation, by using type IIS restriction enzymes, which cleave outside of their recognition site. In addition to reducing the number of steps, this one-pot reaction can improve correct assemblies by the continued cleavage of self-ligation products that retain the recognition site. Switching the specific restriction enzyme used between steps allows for modular assembly of several units. A protocol to perform modular assemblies with two type IIS restriction enzymes, namely BsaI-v2-HF and BsmBI-v2, is described here. This protocol includes a description for generating destination vectors that add loxPsym sites between transcriptional units, allowing for diversification of gene order, orientation, and spacing., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Piper nigrum, also known as black pepper, is an economically and ecologically important crop of the genus Piper. It has been titled as the king of spices due to its wide consumption throughout the world. In the present investigation, the chloroplast genome of P. nigrum has been assembled from a whole genome sequence by integrating the short and long reads generated through Illumina and PacBio platforms, respectively. The chloroplast genome was observed to be 161,522 bp in size, having a quadripartite structure with a large single copy (LSC) region of 89,153 bp and a small single copy (SSC) region of 18,255 bp separated by a copy of inverted repeats (IRs), each 27,057 bp in length. Taking into consideration all the duplicated genes, a total of 131 genes were observed, which included 81 protein-coding genes, 37 tRNAs, 4 rRNAs, and 1 pseudogene. Individually, the LSC region consisted of 83 genes, the SSC region had 13 genes, and 18 genes were present in each IR region. Additionally, 216 SSRs were detected and 11 of these were validated through amplification in 12 species of Piper. The features of the chloroplast genome have been compared with those of the genus Piper. Our results provide useful insights into evolutionary and molecular studies of black pepper which will contribute to its further genetic improvement and breeding.

The complete mitochondrial genome (mitogenome) of Miroplana shenzhensis Yu & Wang, is reported in the present study, representing the second mitogenome recorded in the suborder Maricola. The circular mitogenome is 14,344 bp in length, containing 12 protein-coding genes, 2 ribosomal RNAs and 22 transfer RNAs. Comparative analysis on mitochondrial gene order reveals a rearrangement in the suborder Maricola, indicating that mitochondrial gene order is conserved only in Continenticola, and is divergent across Tricladida. Phylogenetic analysis shows M. shenzhensis is clustered with an another marine triclad, forming a well-supported monophyletic group of Maricloan.

Vernal pool clam shrimp (Eulimnadia texana) are a promising model system due to their ease of lab culture, short generation time, modest sized genome, a somewhat rare stable androdioecious sex determination system, and a requirement to reproduce via desiccated diapaused eggs. We generated a highly contiguous genome assembly using 46× of PacBio long read data and 216× of Illumina short reads, and annotated using Illumina RNAseq obtained from adult males or hermaphrodites. Of the 120 Mb genome 85% is contained in the largest eight contigs, the smallest of which is 4.6 Mb. The assembly contains 98% of transcripts predicted via RNAseq. This assembly is qualitatively different from scaffolded Illumina assemblies: It is produced from long reads that contain sequence data along their entire length, and is thus gap free. The contiguity of the assembly allows us to order the HOX genes within the genome, identifying two loci that contain HOX gene orthologs, and which approximately maintain the order observed in other arthropods. We identified a partial duplication of the Antennapedia complex adjacent to the few genes homologous to the Bithorax locus. Because the sex chromosome of an androdioecious species is of special interest, we used existing allozyme and microsatellite markers to identify the E. texana sex chromosome, and find that it comprises nearly half of the genome of this species. Linkage patterns indicate that recombination is extremely rare and perhaps absent in hermaphrodites, and as a result the location of the sex determining locus will be difficult to refine using recombination mapping.

Deciphering neuronal networks governing specific brain functions is a longstanding mission in neuroscience, yet global manipulation of protein functions pharmacologically or genetically lacks sufficient specificity to reveal a neuronal protein's function in a particular neuron or a circuitry. Photostimulation presents a great venue for researchers to control neuronal proteins with high temporal and spatial resolution. Recently, an approach to optically control the function of a neuronal protein directly in neurons has been demonstrated using genetically encoded light-sensitive Unnatural amino acids (Uaas). Here, we describe procedures for genetically incorporating Uaas into target neuronal proteins in neurons in vitro and in embryonic mouse brain. As an example, a photocaged Uaa was incorporated into an inwardly rectifying potassium channel Kir2.1 to render Kir2.1 photo-activatable. This method has the potential to be generally applied to many neuronal proteins to achieve optical regulation of different processes in brains. Uaas with other properties can be similarly incorporated into neuronal proteins in neurons for various applications.

Genetic switches frequently include DNA loops secured by proteins. Recent studies of the lambda bacteriophage repressor (CI), showed that this arrangement in which the protein links two sets of three operators separated by approximately 2.3 kbp, optimizes both the stability and dynamics of DNA loops, compared to an arrangement with just two sets of two operators. Because adjacent dimers interact pairwise, we hypothesized that the odd number of operators in each set of the lambda regulatory system might have evolved to allow for semi-specific, pair-wise interactions that add stability to the loop while maintaining it dynamic. More generally, additional CI dimers may bind non-specifically to flanking DNA sequences making the genetic switch more sensitive to CI concentration. Here, we tested this hypothesis using spectroscopic and imaging approaches to study the binding of the lambda repressor (CI) dimer protein to DNA fragments. For fragments with only one operator and a short flanking sequence, fluorescence correlation spectroscopy measurements clearly indicated the presence of two distinct DNA-CI complexes; one is thought to have a non-specifically bound CI dimer on the flanking sequence. Scanning force micrographs of CI bound to DNA with all six operators revealed wild-type or mutant proteins bound at operator positions. The number of bound, wild-type proteins increased with CI concentration and was larger than expected for strictly specific binding to operators. In contrast, a mutant that fails to oligomerize beyond a dimer, D197G, only bound to operators. These data are evidence that CI cooperativity promotes oligomerization that extends from operator sites to influence the thermodynamics and kinetics of CI-mediated looping.

Generation of hematopoietic stem cells (HSCs) from pluripotent stem cells (PSCs) could potentially provide unlimited HSCs for clinical transplantation, a curative treatment for numerous blood diseases. However, to date, bona fide HSC generation has been largely unsuccessful in vitro. We have previously described proof of concept for in vivo HSC generation from PSCs via teratoma formation. However, our first-generation system was complex and the output low. Here, we further optimize this technology and demonstrate the following: (1) simplified HSC generation using transcription factor overexpression; (2) improved HSC output using c-Kit-deficient host mice, and (3) that teratomas can be transplanted and cryopreserved. We demonstrate that overexpression of Gfi1b, c-Fos, and Gata2, previously reported to transdifferentiate fibroblasts into hematopoietic progenitors in vitro, can induce long-term HSC formation in vivo. Our in vivo system provides a useful platform to investigate new strategies and re-evaluate existing strategies to generate HSCs and study HSC development.

Lowering total tau levels is an attractive therapeutic strategy for Alzheimer's disease and other tauopathies. High-throughput screening in neurons derived from human induced pluripotent stem cells (iPSCs) is a powerful tool to identify tau-targeted therapeutics. However, such screens have been hampered by heterogeneous neuronal production, high cost and low yield, and multi-step differentiation procedures. We engineered an isogenic iPSC line that harbors an inducible neurogenin 2 transgene, a transcription factor that rapidly converts iPSCs to neurons, integrated at the AAVS1 locus. Using a simplified two-step protocol, we differentiated these iPSCs into cortical glutamatergic neurons with minimal well-to-well variability. We developed a robust high-content screening assay to identify tau-lowering compounds in LOPAC and identified adrenergic receptors agonists as a class of compounds that reduce endogenous human tau. These techniques enable the use of human neurons for high-throughput screening of drugs to treat neurodegenerative disease.

The agricultural pest known as the glassy-winged sharpshooter (GWSS) or Homalodisca vitripennis (Hemiptera: Cicadellidae) harbors two bacterial symbionts, "Candidatus Sulcia muelleri" and "Ca Baumannia cicadellinicola," which provide the 10 essential amino acids (EAAs) that are limited in the host plant-sap diet. Although they differ in origin and symbiotic age, both bacteria have experienced extensive genome degradation resulting from their ancient restriction to specialized host organs (bacteriomes) that provide cellular support and ensure vertical transmission. GWSS bacteriomes are of different origins and distinctly colored red and yellow. While Sulcia occupies the yellow bacteriome, Baumannia inhabits both. Aside from genomic predictions, little is currently known about the cellular functions of these bacterial symbionts, particularly whether Baumannia in different bacteriomes perform different roles in the symbiosis. To address these questions, we conducted a replicated, strand-specific RNA-seq experiment to assay global gene expression patterns in Sulcia and Baumannia Despite differences in genomic capabilities, the symbionts exhibit similar profiles of their most highly expressed genes, including those involved in nutrition synthesis and protein stability (chaperonins dnaK and groESL) that likely aid impaired proteins. Baumannia populations in separate bacteriomes differentially express genes enriched in essential nutrient synthesis, including EAAs (histidine and methionine) and B vitamins (biotin and thiamine). Patterns of differential gene expression further reveal complexity in methionine synthesis. Baumannia's capability to differentially express genes is unusual, as ancient symbionts lose the capability to independently regulate transcription. Combined with previous microscopy, our results suggest that the GWSS may rely on distinct Baumannia populations for essential nutrition and vertical transmission.

We recently developed base editing, the programmable conversion of target C:G base pairs to T:A without inducing double-stranded DNA breaks (DSBs) or requiring homology-directed repair using engineered fusions of Cas9 variants and cytidine deaminases. Over the past year, the third-generation base editor (BE3) and related technologies have been successfully used by many researchers in a wide range of organisms. The product distribution of base editing-the frequency with which the target C:G is converted to mixtures of undesired by-products, along with the desired T:A product-varies in a target site-dependent manner. We characterize determinants of base editing outcomes in human cells and establish that the formation of undesired products is dependent on uracil N-glycosylase (UNG) and is more likely to occur at target sites containing only a single C within the base editing activity window. We engineered CDA1-BE3 and AID-BE3, which use cytidine deaminase homologs that increase base editing efficiency for some sequences. On the basis of these observations, we engineered fourth-generation base editors (BE4 and SaBE4) that increase the efficiency of C:G to T:A base editing by approximately 50%, while halving the frequency of undesired by-products compared to BE3. Fusing BE3, BE4, SaBE3, or SaBE4 to Gam, a bacteriophage Mu protein that binds DSBs greatly reduces indel formation during base editing, in most cases to below 1.5%, and further improves product purity. BE4, SaBE4, BE4-Gam, and SaBE4-Gam represent the state of the art in C:G-to-T:A base editing, and we recommend their use in future efforts.

Gastropoda is the most speciose class in Mollusca, the second largest animal phylum. The internal relationships of major gastropod groups remain largely unsettled, partly due to the insufficient data from key deep-water endemic lineages such as the subclass Neomphaliones. Neomphaliones currently includes two orders: Cocculinida, best known from sunken wood habitats, and Neomphalida, best known from hydrothermal vents and often referred to as the “hot vent clade.” Phylogenetic controversy has also been observed in this subclass across different studies, requesting additional investigations. Here, we assembled nine new mitogenomes from two Cocculinida and seven Neomphalida species and analyzed them with published gastropod mitogenomes, with a particular focus on Neomphaliones. The phylogenetic reconstruction of Gastropoda based on 13 mitochondrial protein-coding genes resulted in a topology largely congruent with previous reconstructions based on morphological characters. Furthermore, we recovered characteristic mitochondrial gene order arrangements of Cocculinida and Neomphalida compared to the hypothetical ancestral gastropod gene order, at a level similar to other subclass-level clades. Divergence time estimation showed that Cocculinida and Neomphalida diverged approximately 322.68 million years ago. In addition to characteristic gene order arrangements for the clade, Cocculinida mitogenomes also exhibit some minor rearrangements even among congeners. Within Neomphalida, our tree adds support to monophyletic Peltospiridae and Neomphalidae, with unique gene arrangement recovered for each family. Our results offer new insights into the rearrangement of mitogenomes in Gastropoda, providing another clue to the evolutionary history of gastropods.

The major histocompatibility complex (MHC) is of central importance to the immune system, and an optimal MHC diversity is believed to maximize pathogen elimination. Birds show substantial variation in MHC diversity, ranging from few genes in most bird orders to very many genes in passerines. Our understanding of the evolutionary trajectories of the MHC in passerines is hampered by lack of data on genomic organization. Therefore, we assembled and annotated the MHC genomic region of the great reed warbler (Acrocephalus arundinaceus), using long‐read sequencing and optical mapping. The MHC region is large (>5.5 Mb), characterized by structural changes compared to hitherto investigated bird orders and shows higher repeat content than the genome average. These features were supported by analyses in three additional passerines. MHC genes in passerines are found in two different chromosomal arrangements, either as single copy MHC genes located among non‐MHC genes, or as tandemly duplicated tightly linked MHC genes. Some single copy MHC genes are old and putative orthologues among species. In contrast tandemly duplicated MHC genes are monophyletic within species and have evolved by simultaneous gene duplication of several MHC genes. Structural differences in the MHC genomic region among bird orders seem substantial compared to mammals and have possibly been fuelled by clade‐specific immune system adaptations. Our study provides methodological guidance in characterizing complex genomic regions, constitutes a resource for MHC research in birds, and calls for a revision of the general belief that avian MHC has a conserved gene order and small size compared to mammals. [ABSTRACT FROM AUTHOR]

The Pleosporales is the most predominant order in the Dothideomycetes class, which contains over 4700 species that function in a variety of ways. The material used in this research was previously isolated from the Chinese white wax scale insect, and it was determined to be a Paraconiothyrium genus species that belonged to the Pleosporales order. For further molecular analysis, we assembled the complete mitochondrial genome of Paraconiothyrium sp. based on short reads of BGISEQ sequencing and subreads from Pacbio sequencing. The results showed that it was 42,734 bp in length and contained 8 open reading frames, 12 protein-coding genes and 31 non-coding genes. Phylogenetic analysis showed it was affiliated to the Pleosporales order and formed a sister relationship with Pithomyces chartarum. Compared to the seven other species in the Pleosporales order, Paraconiothyrium sp. has generally conserved gene content and structure, while the homologous blocks and gene order were shown to be significantly rearranged, in accordance with the species diversity in the Pleosporales order. In this study, we presented the first mitochondrial genome of Paraconiothyrium fungi to be reported, and we also showed gene order diversity in the Pleosporales order. These findings will lay the foundation for further species studies regarding molecular diversity and our understanding of species characteristics in the Paraconiothyrium genus. [ABSTRACT FROM AUTHOR]

The complete mitogenome sequence of the Great Frigatebird, Fregata minor was sequenced for the first time in this study. The mitogenome (16,899 bp) comprises of 13 protein-coding genes (PCGs), two ribosomal RNA (rRNA) genes, and 22 transfer RNA (tRNA) genes, and a control region (CR). The mitogenome was AT-rich (55.60%) with 11 overlapping and 18 intergenic spacer regions. Most of the PCGs were started by a typical ATG initiation codon except for cox1 and nad3. A maximum-likelihood phylogeny of concatenated PCGs resulted in a well-resolved phylogeny of all the species of Suliformes and illuminates the sister relationship of F. minor with F. magnificens. The present mitogenome-based phylogeny clearly enlightens the evolutionary position of Suliformes and Pelecaniformes species. Unique tandem repeats were identified in both F. minor and F. magnificens, which can be employed as a species-specific marker. To illuminate the population structure of this migratory seabirds, the present study advocate more sampling and the generation of additional molecular data to clarify their genetic diversity. The present study also rejects an earlier hypothesis on the mitochondrial gene order of Suliformes and corroborated the typical avian gene order in frigatebirds. [ABSTRACT FROM AUTHOR]

To further understand the origin and evolution of Patellogastropoda, we determined the mitochondrial genome sequence of Cellana toreuma, and compared its mitogenome characteristics with the other four limpets of Nacellidae. The ratio of Ka and Ks indicated that these Nacellidae species were suffering a purifying selection, with exception of the atp6 gene. The gene sequence is basically consistent among families, while there are great differences among Lottidae species. According to the mitogenome sequences of selected gastropod species, we reconstructed a new phylogenetic tree with two methods. The data complement the mitogenome database of limpets and is a favorable research tool for the phylogenetic analysis of Gastropoda. It is found that there is a long-branch attraction (LBA) artefact in the family Lottiidae of Patellogastropoda. Therefore, the Patellogastropoda was separated by Heterobranchia, and Lottiidae is located at the root of the whole phylogenetic tree. Furthermore, we constructed the divergence time tree according to the Bayesian method and discussed the internal historical dynamics, and divergence differences among the main lineages of 12 Patellogastropoda under an uncorrelated relaxed molecular clock. In turn, we made a more comprehensive discussion on the divergence time of limpets at the molecular level. [ABSTRACT FROM AUTHOR]

Since the first description of adenoviruses in bats in 2006, a number of micro- and megabat species in Europe, Africa, and Asia have been shown to carry a wide diversity of adenoviruses. Here, we report on the evolutionary, biological, and structural characterization of a novel bat adenovirus (BtAdV) recovered from a Rafinesque's big-eared bat (Corynorhinus rafinesquii) in Kentucky, USA, which is the first adenovirus isolated from North American bats. This virus (BtAdV 250-A) exhibits a close phylogenetic relationship with Canine mastadenovirus A (CAdV A), as previously observed with other BtAdVs. To further investigate the relationships between BtAdVs and CAdVs, we conducted mass spectrometric analysis and single-particle cryo-electron microscopy reconstructions of the BtAdV 250-A capsid and also analyzed the in vitro host ranges of both viruses. Our results demonstrate that BtAdV 250-A represents a new mastadenovirus species that, in contrast to CAdV, has a unique capsid morphology that contains more prominent extensions of protein IX and can replicate efficiently in a phylogenetically diverse range of species. These findings, in addition to the recognition that both the genetic diversity of BtAdVs and the number of different bat species from disparate geographic regions infected with BtAdVs appears to be extensive, tentatively suggest that bats may have served as a potential reservoir for the cross-species transfer of adenoviruses to other hosts, as theorized for CAdV. Although many adenoviruses are host specific and likely codiverged with their hosts over millions of years, other adenoviruses appear to have emerged through successful cross-species transmission events on more recent time scales. The wide geographic distribution and genetic diversity of adenoviruses in bats and their close phylogenetic relationship to Canine mastadenovirus A (CAdV A) has raised important questions about how CAdV A, and possibly other mammalian adenoviruses, may have emerged. Although most adenoviruses tend to cause limited disease in their natural hosts, CAdV A is unusual in that it may cause high morbidity and sometimes fatal infections in immunocompetent hosts and is thus an important pathogen of carnivores. Here, we performed a comparative evolutionary and structural study of representative bat and canine adenoviruses to better understand the relationship between these two viral groups.

The complete mitochondrial genome was sequenced in two individuals of the Kamchatka grayling Thymallus mertensii. The genome sequences are 16 662 bp in size, and the gene arrangement, composition, and size are very similar to the salmonid fish genomes published previously. The low level of sequence divergence (0.92%) detected between the genome of T. mertensii and the GenBank complete mitochondrial genomes of the Arctic grayling T. arcticus (FJ872559) may likely be due to recent divergence of the species and/or historical hybridization and interspecific replacement of mtDNA.