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September 22, 2019  |  

A molecular window into the biology and epidemiology of Pneumocystis spp.

Pneumocystis, a unique atypical fungus with an elusive lifestyle, has had an important medical history. It came to prominence as an opportunistic pathogen that not only can cause life-threatening pneumonia in patients with HIV infection and other immunodeficiencies but also can colonize the lungs of healthy individuals from a very early age. The genus Pneumocystis includes a group of closely related but heterogeneous organisms that have a worldwide distribution, have been detected in multiple mammalian species, are highly host species specific, inhabit the lungs almost exclusively, and have never convincingly been cultured in vitro, making Pneumocystis a fascinating but difficult-to-study organism. Improved molecular biologic methodologies have opened a new window into the biology and epidemiology of Pneumocystis. Advances include an improved taxonomic classification, identification of an extremely reduced genome and concomitant inability to metabolize and grow independent of the host lungs, insights into its transmission mode, recognition of its widespread colonization in both immunocompetent and immunodeficient hosts, and utilization of strain variation to study drug resistance, epidemiology, and outbreaks of infection among transplant patients. This review summarizes these advances and also identifies some major questions and challenges that need to be addressed to better understand Pneumocystis biology and its relevance to clinical care. Copyright © 2018 American Society for Microbiology.


September 22, 2019  |  

Comparative analysis reveals unexpected genome features of newly isolated Thraustochytrids strains: on ecological function and PUFAs biosynthesis.

Thraustochytrids are unicellular fungal-like marine protists with ubiquitous existence in marine environments. They are well-known for their ability to produce high-valued omega-3 polyunsaturated fatty acids (?-3-PUFAs) (e.g., docosahexaenoic acid (DHA)) and hydrolytic enzymes. Thraustochytrid biomass has been estimated to surpass that of bacterioplankton in both coastal and oceanic waters indicating they have an important role in microbial food-web. Nevertheless, the molecular pathway and regulatory network for PUFAs production and the molecular mechanisms underlying ecological functions of thraustochytrids remain largely unknown.The genomes of two thraustochytrids strains (Mn4 and SW8) with ability to produce DHA were sequenced and assembled with a hybrid sequencing approach utilizing Illumina short paired-end reads and Pacific Biosciences long reads to generate a highly accurate genome assembly. Phylogenomic and comparative genomic analyses found that DHA-producing thraustochytrid strains were highly similar and possessed similar gene content. Analysis of the conventional fatty acid synthesis (FAS) and the polyketide synthase (PKS) systems for PUFAs production only detected incomplete and fragmentary pathways in the genome of these two strains. Surprisingly, secreted carbohydrate active enzymes (CAZymes) were found to be significantly depleted in the genomes of these 2 strains as compared to other sequenced relatives. Furthermore, these two strains possess an expanded gene repertoire for signal transduction and self-propelled movement, which could be important for their adaptations to dynamic marine environments.Our results demonstrate the possibility of a third PUFAs synthesis pathway besides previously described FAS and PKS pathways encoded in the genome of these two thraustochytrid strains. Moreover, lack of a complete set of hydrolytic enzymatic machinery for degrading plant-derived organic materials suggests that these two DHA-producing strains play an important role as a nutritional source rather than a nutrient-producer in marine microbial-food web. Results of this study suggest the existence of two types of saprobic thraustochytrids in the world’s ocean. The first group, which does not produce cellulosic enzymes and live as ‘left-over’ scavenger of bacterioplankton, serves as a dietary source for the plankton of higher trophic levels and the other possesses capacity to live on detrital organic matters in the marine ecosystems.


September 22, 2019  |  

The Chara genome: Secondary complexity and implications for plant terrestrialization.

Land plants evolved from charophytic algae, among which Charophyceae possess the most complex body plans. We present the genome of Chara braunii; comparison of the genome to those of land plants identified evolutionary novelties for plant terrestrialization and land plant heritage genes. C. braunii employs unique xylan synthases for cell wall biosynthesis, a phragmoplast (cell separation) mechanism similar to that of land plants, and many phytohormones. C. braunii plastids are controlled via land-plant-like retrograde signaling, and transcriptional regulation is more elaborate than in other algae. The morphological complexity of this organism may result from expanded gene families, with three cases of particular note: genes effecting tolerance to reactive oxygen species (ROS), LysM receptor-like kinases, and transcription factors (TFs). Transcriptomic analysis of sexual reproductive structures reveals intricate control by TFs, activity of the ROS gene network, and the ancestral use of plant-like storage and stress protection proteins in the zygote. Copyright © 2018 Elsevier Inc. All rights reserved.


September 22, 2019  |  

Genome survey of the freshwater mussel Venustaconcha ellipsiformis (Bivalvia: Unionida) using a hybrid de novo assembly approach.

Freshwater mussels (Bivalvia: Unionida) serve an important role as aquatic ecosystem engineers but are one of the most critically imperilled groups of animals. Here, we used a combination of sequencing strategies to assemble and annotate a draft genome of Venustaconcha ellipsiformis, which will serve as a valuable genomic resource given the ecological value and unique “doubly uniparental inheritance” mode of mitochondrial DNA transmission of freshwater mussels. The genome described here was obtained by combining high-coverage short reads (65× genome coverage of Illumina paired-end and 11× genome coverage of mate-pairs sequences) with low-coverage Pacific Biosciences long reads (0.3× genome coverage). Briefly, the final scaffold assembly accounted for a total size of 1.54?Gb (366,926 scaffolds, N50?=?6.5 kb, with 2.3% of “N” nucleotides), representing 86% of the predicted genome size of 1.80?Gb, while over one third of the genome (37.5%) consisted of repeated elements and >85% of the core eukaryotic genes were recovered. Given the repeated genetic bottlenecks of V. ellipsiformis populations as a result of glaciations events, heterozygosity was also found to be remarkably low (0.6%), in contrast to most other sequenced bivalve species. Finally, we reassembled the full mitochondrial genome and found six polymorphic sites with respect to the previously published reference. This resource opens the way to comparative genomics studies to identify genes related to the unique adaptations of freshwater mussels and their distinctive mitochondrial inheritance mechanism.


September 22, 2019  |  

The complete mitochondrial genome of the Basidiomycete edible fungus Hypsizygus marmoreus

The complete mitochondrial genome of the edible fungus Hypsizygus marmoreus was published in this paper. It was determined using Pacbio and Illumina sequencing. The complete mitochondrial DNA (mtDNA) is 106,417?bp in length with a GC content of 31.74%, which was the fourth large mitogenome in Agaricales. The circular mitogenome encoded 67 protein-coding genes and one ribosomal RNAs (rns). Among these genes, 13 conserved protein-coding genes were determined in the genome, including 6 subunits of NAD dehydrogenase (nad1-4, 4L and 6), three cytochrome oxidases (cox1-3), one apocytochrome b (cob) and three ATP synthases (atp6, apt 8 and apt 9). The phylogenic analysis confirmed that H. marmoreus (Lyophyllaceae) clustered together with Tricholoma matsutake (Tricholomataceae).


September 22, 2019  |  

Genomic signatures of mitonuclear coevolution across populations of Tigriopus californicus.

The copepod Tigriopus californicus shows extensive population divergence and is becoming a model for understanding allopatric differentiation and the early stages of speciation. Here, we report a high-quality reference genome for one population (~190?megabases across 12 scaffolds, and ~15,500 protein-coding genes). Comparison with other arthropods reveals 2,526 genes presumed to be specific to T. californicus, with an apparent proliferation of genes involved in ion transport and receptor activity. Beyond the reference population, we report re-sequenced genomes of seven additional populations, spanning the continuum of reproductive isolation. Populations show extreme mitochondrial DNA divergence, with higher levels of amino acid differentiation than observed in other taxa. Across the nuclear genome, we find elevated protein evolutionary rates and positive selection in genes predicted to interact with mitochondrial DNA and the proteins and RNA it encodes in multiple pathways. Together, these results support the hypothesis that rapid mitochondrial evolution drives compensatory nuclear evolution within isolated populations, thereby providing a potentially important mechanism for causing intrinsic reproductive isolation.


September 22, 2019  |  

Creating a functional single-chromosome yeast.

Eukaryotic genomes are generally organized in multiple chromosomes. Here we have created a functional single-chromosome yeast from a Saccharomyces cerevisiae haploid cell containing sixteen linear chromosomes, by successive end-to-end chromosome fusions and centromere deletions. The fusion of sixteen native linear chromosomes into a single chromosome results in marked changes to the global three-dimensional structure of the chromosome due to the loss of all centromere-associated inter-chromosomal interactions, most telomere-associated inter-chromosomal interactions and 67.4% of intra-chromosomal interactions. However, the single-chromosome and wild-type yeast cells have nearly identical transcriptome and similar phenome profiles. The giant single chromosome can support cell life, although this strain shows reduced growth across environments, competitiveness, gamete production and viability. This synthetic biology study demonstrates an approach to exploration of eukaryote evolution with respect to chromosome structure and function.


September 22, 2019  |  

Biology and genome of a newly discovered sibling species of Caenorhabditis elegans.

A ‘sibling’ species of the model organism Caenorhabditis elegans has long been sought for use in comparative analyses that would enable deep evolutionary interpretations of biological phenomena. Here, we describe the first sibling species of C. elegans, C. inopinata n. sp., isolated from fig syconia in Okinawa, Japan. We investigate the morphology, developmental processes and behaviour of C. inopinata, which differ significantly from those of C. elegans. The 123-Mb C. inopinata genome was sequenced and assembled into six nuclear chromosomes, allowing delineation of Caenorhabditis genome evolution and revealing unique characteristics, such as highly expanded transposable elements that might have contributed to the genome evolution of C. inopinata. In addition, C. inopinata exhibits massive gene losses in chemoreceptor gene families, which could be correlated with its limited habitat area. We have developed genetic and molecular techniques for C. inopinata; thus C. inopinata provides an exciting new platform for comparative evolutionary studies.


September 22, 2019  |  

Complete sequence of kenaf (Hibiscus cannabinus) mitochondrial genome and comparative analysis with the mitochondrial genomes of other plants.

Plant mitochondrial (mt) genomes are species specific due to the vast of foreign DNA migration and frequent recombination of repeated sequences. Sequencing of the mt genome of kenaf (Hibiscus cannabinus) is essential for elucidating its evolutionary characteristics. In the present study, single-molecule real-time sequencing technology (SMRT) was used to sequence the complete mt genome of kenaf. Results showed that the complete kenaf mt genome was 569,915?bp long and consisted of 62 genes, including 36 protein-coding, 3 rRNA and 23 tRNA genes. Twenty-five introns were found among nine of the 36 protein-coding genes, and five introns were trans-spliced. A comparative analysis with other plant mt genomes showed that four syntenic gene clusters were conserved in all plant mtDNAs. Fifteen chloroplast-derived fragments were strongly associated with mt genes, including the intact sequences of the chloroplast genes psaA, ndhB and rps7. According to the plant mt genome evolution analysis, some ribosomal protein genes and succinate dehydrogenase genes were frequently lost during the evolution of angiosperms. Our data suggest that the kenaf mt genome retained evolutionarily conserved characteristics. Overall, the complete sequencing of the kenaf mt genome provides additional information and enhances our better understanding of mt genomic evolution across angiosperms.


September 22, 2019  |  

Genomic analysis of Sparus aurata reveals the evolutionary dynamics of sex-biased genes in a sequential hermaphrodite fish

Sexual dimorphism is a fascinating subject in evolutionary biology and mostly results from sex-biased expression of genes, which have been shown to evolve faster in gonochoristic species. We report here genome and sex-specific transcriptome sequencing of Sparus aurata, a sequential hermaphrodite fish. Evolutionary comparative analysis reveals that sex-biased genes in S. aurata are similar in number and function, but evolved following strikingly divergent patterns compared with gonochoristic species, showing overall slower rates because of stronger functional constraints. Fast evolution is observed only for highly ovary-biased genes due to female-specific patterns of selection that are related to the peculiar reproduction mode of S. aurata, first maturing as male, then as female. To our knowledge, these findings represent the first genome-wide analysis on sex-biased loci in a hermaphrodite vertebrate species, demonstrating how having two sexes in the same individual profoundly affects the fate of a large set of evolutionarily relevant genes.


September 22, 2019  |  

Whole-genome resequencing and pan-transcriptome reconstruction highlight the impact of genomic structural Variation on secondary metabolite gene clusters in the grapevine Esca pathogen Phaeoacremonium minimum.

The Ascomycete fungus Phaeoacremonium minimum is one of the primary causal agents of Esca, a widespread and damaging grapevine trunk disease. Variation in virulence among Pm. minimum isolates has been reported, but the underlying genetic basis of the phenotypic variability remains unknown. The goal of this study was to characterize intraspecific genetic diversity and explore its potential impact on virulence functions associated with secondary metabolism, cellular transport, and cell wall decomposition. We generated a chromosome-scale genome assembly, using single molecule real-time sequencing, and resequenced the genomes and transcriptomes of multiple isolates to identify sequence and structural polymorphisms. Numerous insertion and deletion events were found for a total of about 1 Mbp in each isolate. Structural variation in this extremely gene dense genome frequently caused presence/absence polymorphisms of multiple adjacent genes, mostly belonging to biosynthetic clusters associated with secondary metabolism. Because of the observed intraspecific diversity in gene content due to structural variation we concluded that a transcriptome reference developed from a single isolate is insufficient to represent the virulence factor repertoire of the species. We therefore compiled a pan-transcriptome reference of Pm. minimum comprising a non-redundant set of 15,245 protein-coding sequences. Using naturally infected field samples expressing Esca symptoms, we demonstrated that mapping of meta-transcriptomics data on a multi-species reference that included the Pm. minimum pan-transcriptome allows the profiling of an expanded set of virulence factors, including variable genes associated with secondary metabolism and cellular transport.


September 22, 2019  |  

The complete mitochondrial genome of the early flowering plant Nymphaea colorata is highly repetitive with low recombination.

Mitochondrial genomes of flowering plants (angiosperms) are highly dynamic in genome structure. The mitogenome of the earliest angiosperm Amborella is remarkable in carrying rampant foreign DNAs, in contrast to Liriodendron, the other only known early angiosperm mitogenome that is described as ‘fossilized’. The distinctive features observed in the two early flowering plant mitogenomes add to the current confusions of what early flowering plants look like. Expanded sampling would provide more details in understanding the mitogenomic evolution of early angiosperms. Here we report the complete mitochondrial genome of water lily Nymphaea colorata from Nymphaeales, one of the three orders of the earliest angiosperms.Assembly of data from Pac-Bio long-read sequencing yielded a circular mitochondria chromosome of 617,195 bp with an average depth of 601×. The genome encoded 41 protein coding genes, 20 tRNA and three rRNA genes with 25 group II introns disrupting 10 protein coding genes. Nearly half of the genome is composed of repeated sequences, which contributed substantially to the intron size expansion, making the gross intron length of the Nymphaea mitochondrial genome one of the longest among angiosperms, including an 11.4-Kb intron in cox2, which is the longest organellar intron reported to date in plants. Nevertheless, repeat mediated homologous recombination is unexpectedly low in Nymphaea evidenced by 74 recombined reads detected from ten recombinationally active repeat pairs among 886,982 repeat pairs examined. Extensive gene order changes were detected in the three early angiosperm mitogenomes, i.e. 38 or 44 events of inversions and translocations are needed to reconcile the mitogenome of Nymphaea with Amborella or Liriodendron, respectively. In contrast to Amborella with six genome equivalents of foreign mitochondrial DNA, not a single horizontal gene transfer event was observed in the Nymphaea mitogenome.The Nymphaea mitogenome resembles the other available early angiosperm mitogenomes by a similarly rich 64-coding gene set, and many conserved gene clusters, whereas stands out by its highly repetitive nature and resultant remarkable intron expansions. The low recombination level in Nymphaea provides evidence for the predominant master conformation in vivo with a highly substoichiometric set of rearranged molecules.


September 22, 2019  |  

Assembly and comparative analysis of the complete mitochondrial genome sequence of Sophora japonica ‘JinhuaiJ2’.

Sophora japonica L. (Faboideae, Leguminosae) is an important traditional Chinese herb with a long history of cultivation. Its flower buds and fruits contain abundant flavonoids, and therefore, the plants are cultivated for the industrial extraction of rutin. Here, we determined the complete nucleotide sequence of the mitochondrial genome of S. japonica ‘JinhuaiJ2’, the most widely planted variety in Guangxi region of China. The total length of the mtDNA sequence is 484,916 bp, with a GC content of 45.4%. Sophora japonica mtDNA harbors 32 known protein-coding genes, 17 tRNA genes, and three rRNA genes with 17 cis-spliced and five trans-spliced introns disrupting eight protein-coding genes. The gene coding and intron regions, and intergenic spacers account for 7.5%, 5.8% and 86.7% of the genome, respectively. The gene profile of S. japonica mitogenome differs from that of the other Faboideae species by only one or two gene gains or losses. Four of the 17 cis-spliced introns showed distinct length variations in the Faboideae, which could be attributed to the homologous recombination of the short repeats measuring a few bases located precisely at the edges of the putative deletions. This reflects the importance of small repeats in the sequence evolution in Faboideae mitogenomes. Repeated sequences of S. japonica mitogenome are mainly composed of small repeats, with only 20 medium-sized repeats, and one large repeat, adding up to 4% of its mitogenome length. Among the 25 pseudogene fragments detected in the intergenic spacer regions, the two largest ones and their corresponding functional gene copies located in two different sets of medium-sized repeats, point to their origins from homologous recombinations. As we further observed the recombined reads associated with the longest repeats of 2,160 bp with the PacBio long read data set of just 15 × in depth, repeat mediated homologous recombinations may play important role in the mitogenomic evolution of S. japonica. Our study provides insightful knowledge to the genetic background of this important herb species and the mitogenomic evolution in the Faboideae species.


September 22, 2019  |  

PacBio-based mitochondrial genome assembly of Leucaena trichandra (Leguminosae) and an intrageneric assessment of mitochondrial RNA editing.

Reconstructions of vascular plant mitochondrial genomes (mt-genomes) are notoriously complicated by rampant recombination that has resulted in comparatively few plant mt-genomes being available. The dearth of plant mitochondrial resources has limited our understanding of mt-genome structural diversity, complex patterns of RNA editing, and the origins of novel mt-genome elements. Here, we use an efficient long read (PacBio) iterative assembly pipeline to generate mt-genome assemblies for Leucaena trichandra (Leguminosae: Caesalpinioideae: mimosoid clade), providing the first assessment of non-papilionoid legume mt-genome content and structure to date. The efficiency of the assembly approach facilitated the exploration of alternative structures that are common place among plant mitochondrial genomes. A compact version (729 kbp) of the recovered assemblies was used to investigate sources of mt-genome size variation among legumes and mt-genome sequence similarity to the legume associated root holoparasite Lophophytum. The genome and an associated suite of transcriptome data from select species of Leucaena permitted an in-depth exploration of RNA editing in a diverse clade of closely related species that includes hybrid lineages. RNA editing in the allotetraploid, Leucaena leucocephala, is consistent with co-option of nearly equal maternal and paternal C-to-U edit components, generating novel combinations of RNA edited sites. A preliminary investigation of L. leucocephala C-to-U edit frequencies identified the potential for a hybrid to generate unique pools of alleles from parental variation through edit frequencies shared with one parental lineage, those intermediate between parents, and transgressive patterns.


September 22, 2019  |  

The genome of Naegleria lovaniensis, the basis for a comparative approach to unravel pathogenicity factors of the human pathogenic amoeba N. fowleri.

Members of the genus Naegleria are free-living eukaryotes with the capability to transform from the amoeboid form into resting cysts or moving flagellates in response to environmental conditions. More than 40 species have been characterized, but only Naegleria fowleri (N. fowleri) is known as a human pathogen causing primary amoebic meningoencephalitis (PAM), a fast progressing and mostly fatal disease of the central nervous system. Several studies report an involvement of phospholipases and other molecular factors, but the mechanisms involved in pathogenesis are still poorly understood. To gain a better understanding of the relationships within the genus of Naegleria and to investigate pathogenicity factors of N. fowleri, we characterized the genome of its closest non-pathogenic relative N. lovaniensis.To gain insights into the taxonomy of Naegleria, we sequenced the genome of N. lovaniensis using long read sequencing technology. The assembly of the data resulted in a 30 Mb genome including the circular mitochondrial sequence. Unravelling the phylogenetic relationship using OrthoMCL protein clustering and maximum likelihood methods confirms the close relationship of N. lovaniensis and N. fowleri. To achieve an overview of the diversity of Naegleria proteins and to assess characteristics of the human pathogen N. fowleri, OrthoMCL protein clustering including data of N. fowleri, N. lovaniensis and N. gruberi was performed. GO enrichment analysis shows an association of N. fowleri specific proteins to the GO terms “Membrane” and “Protein Secretion.”In this study, we characterize the hitherto unknown genome of N. lovaniensis. With the description of the 30 Mb genome, a further piece is added to reveal the complex taxonomic relationship of Naegleria. Further, the whole genome sequencing data confirms the hypothesis of the close relationship between N. fowleri and N. lovaniensis. Therefore, the genome of N. lovaniensis provides the basis for further comparative approaches on the molecular and genomic level to unravel pathogenicity factors of its closest human pathogenic relative N. fowleri and possible treatment options for the rare but mostly fatal primary meningoencephalitis.


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