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September 22, 2019  |  

Genomic changes associated with the evolutionary transitions of Nostoc to a plant symbiont.

Cyanobacteria belonging to the genus Nostoc comprise free-living strains and also facultative plant symbionts. Symbiotic strains can enter into symbiosis with taxonomically diverse range of host plants. Little is known about genomic changes associated with evolutionary transition of Nostoc from free-living to plant symbiont. Here, we compared the genomes derived from 11 symbiotic Nostoc strains isolated from different host plants and infer phylogenetic relationships between strains. Phylogenetic reconstructions of 89 Nostocales showed that symbiotic Nostoc strains with a broad host range, entering epiphytic and intracellular or extracellular endophytic interactions, form a monophyletic clade indicating a common evolutionary history. A polyphyletic origin was found for Nostoc strains which enter only extracellular symbioses, and inference of transfer events implied that this trait was likely acquired several times in the evolution of the Nostocales. Symbiotic Nostoc strains showed enriched functions in transport and metabolism of organic sulfur, chemotaxis and motility, as well as the uptake of phosphate, branched-chain amino acids, and ammonium. The genomes of the intracellular clade differ from that of other Nostoc strains, with a gain/enrichment of genes encoding proteins to generate l-methionine from sulfite and pathways for the degradation of the plant metabolites vanillin and vanillate, and of the macromolecule xylan present in plant cell walls. These compounds could function as C-sources for members of the intracellular clade. Molecular clock analysis indicated that the intracellular clade emerged ca. 600 Ma, suggesting that intracellular Nostoc symbioses predate the origin of land plants and the emergence of their extant hosts.

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September 22, 2019  |  

New genomic data and analyses challenge the traditional vision of animal epithelium evolution.

The emergence of epithelia was the foundation of metazoan expansion. Epithelial tissues are a hallmark of metazoans deeply rooted in the evolution of their complex developmental morphogenesis processes. However, studies on the epithelial features of non-bilaterians are still sparse and it remains unclear whether the last common metazoan ancestor possessed a fully functional epithelial toolkit or if it was acquired later during metazoan evolution.To investigate the early evolution of animal epithelia, we sequenced the genome and transcriptomes of two new sponge species to characterize epithelial markers such as the E-cadherin complex and the polarity complexes for all classes (Calcarea, Demospongiae, Hexactinellida, Homoscleromorpha) of sponges (phylum Porifera) and compare them with their homologues in Placozoa and in Ctenophora. We found that Placozoa and most sponges possess orthologues of all essential genes encoding proteins characteristic of bilaterian epithelial cells, as well as their conserved interaction domains. In stark contrast, we found that ctenophores lack several major polarity complex components such as the Crumbs complex and Scribble. Furthermore, the E-cadherin ctenophore orthologue exhibits a divergent cytoplasmic domain making it unlikely to interact with its canonical cytoplasmic partners.These unexpected findings challenge the current evolutionary paradigm on the emergence of epithelia. Altogether, our results raise doubt on the homology of protein complexes and structures involved in cell polarity and adhesive-type junctions between Ctenophora and Bilateria epithelia.

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September 22, 2019  |  

Isolation, functional characterization and transmissibility of p3PS10, a multidrug resistance plasmid of the fish pathogen Piscirickettsia salmonis.

Antibiotic resistance is a major public health concern due to its association with the loss of efficacy of antimicrobial therapies. Horizontal transfer events may play a significant role in the dissemination of resistant bacterial phenotypes, being mobilizable plasmids a well-known mechanism. In this study, we aimed to gain insights into the genetics underlying the development of antibiotic resistance by Piscirickettsia salmonis isolates, a bacterial fish pathogen and causative agent of salmonid piscirickettsiosis, and the main target of antibiotics used in Chilean salmon farming. We provide experimental evidence that the plasmid p3PS10, which harbors multidrug resistance genes for chloramphenicol (cat2), tetracyclines [tet(31)], aminoglycosides (sat1 and aadA1), and sulfonamides (sul2), is carried by a group of P. salmonis isolates exhibiting a markedly reduced susceptibility to oxytetracycline in vitro (128-256 µg/mL of minimal inhibitory concentration, MIC). Antibiotic susceptibility analysis extended to those antibiotics showed that MIC of chloramphenicol, streptomycin, and sulfamethoxazole/trimethoprim were high, but the MIC of florfenicol remained at the wild-type level. By means of molecular cloning, we demonstrate that those genes encoding putative resistance markers are indeed functional. Interestingly, mating assays clearly show that p3PS10 is able to be transferred into and replicate in different hosts, thereby conferring phenotypes similar to those found in the original host. According to epidemiological data, this strain is distributed across aquaculture settings in southern Chile and is likely to be responsible for oxytetracycline treatment failures. This work demonstrates that P. salmonis is more versatile than it was thought, capable of horizontally transferring DNA, and probably playing a role as a vector of resistance traits among the seawater bacterial population. However, the low transmission frequency of p3PS10 suggests a negligible chance of resistance markers being spread to human pathogens.

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September 22, 2019  |  

Comparative genomics analysis of plasmid pPV989-94 from a clinical isolate of Pantoea vagans PV989.

Pantoea vagans, a gram-negative bacterium from the genus Pantoea and family Enterobacteriaceae, is present in various natural environments and considered to be plant endophytes. We isolated the Pantoea vagans PV989 strain from the clinic and sequenced its whole genome. Besides a chromosome DNA molecule, it also harboured three large plasmids. A comparative genomics analysis was performed for the smallest plasmid, pPV989-94. It can be divided into four regions, including three conservative regions related to replication (R1), transfer conjugation (R2), and transfer leading (R3), and one variable region (R4). Further analysis showed that pPV989-94 is most similar to plasmids LA637P2 and pEA68 of Erwinia amylovora strains isolated from fruit trees. These three plasmids share three conservative regions (R1, R2, and R3). Interestingly, a fragment (R4′) in R4, mediated by phage integrase and phage integrase family site-specific recombinase and encoding 9 genes related to glycometabolism, resistance, and DNA repair, was unique in pPV989-94. Homologues of R4′ were found in other plasmids or chromosomes, suggesting that horizontal gene transfer (HGT) occurred among different bacteria of various species or genera. The acquired functional genes may play important roles in the adaptation of bacteria to different hosts or environmental conditions.

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September 22, 2019  |  

Footprints of parasitism in the genome of the parasitic flowering plant Cuscuta campestris.

A parasitic lifestyle, where plants procure some or all of their nutrients from other living plants, has evolved independently in many dicotyledonous plant families and is a major threat for agriculture globally. Nevertheless, no genome sequence of a parasitic plant has been reported to date. Here we describe the genome sequence of the parasitic field dodder, Cuscuta campestris. The genome contains signatures of a fairly recent whole-genome duplication and lacks genes for pathways superfluous to a parasitic lifestyle. Specifically, genes needed for high photosynthetic activity are lost, explaining the low photosynthesis rates displayed by the parasite. Moreover, several genes involved in nutrient uptake processes from the soil are lost. On the other hand, evidence for horizontal gene transfer by way of genomic DNA integration from the parasite’s hosts is found. We conclude that the parasitic lifestyle has left characteristic footprints in the C. campestris genome.

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September 22, 2019  |  

Sea cucumber genome provides insights into saponin biosynthesis and aestivation regulation.

Echinoderms exhibit several fascinating evolutionary innovations that are rarely seen in the animal kingdom, but how these animals attained such features is not well understood. Here we report the sequencing and analysis of the genome and extensive transcriptomes of the sea cucumber Apostichopus japonicus, a species from a special echinoderm group with extraordinary potential for saponin synthesis, aestivation and organ regeneration. The sea cucumber does not possess a reorganized Hox cluster as previously assumed for all echinoderms, and the spatial expression of Hox7 and Hox11/13b potentially guides the embryo-to-larva axial transformation. Contrary to the typical production of lanosterol in animal cholesterol synthesis, the oxidosqualene cyclase of sea cucumber produces parkeol for saponin synthesis and has “plant-like” motifs suggestive of convergent evolution. The transcriptional factors Klf2 and Egr1 are identified as key regulators of aestivation, probably exerting their effects through a clock gene-controlled process. Intestinal hypometabolism during aestivation is driven by the DNA hypermethylation of various metabolic gene pathways, whereas the transcriptional network of intestine regeneration involves diverse signaling pathways, including Wnt, Hippo and FGF. Decoding the sea cucumber genome provides a new avenue for an in-depth understanding of the extraordinary features of sea cucumbers and other echinoderms.

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September 22, 2019  |  

Genome mining of the marine actinomycete Streptomyces sp. DUT11 and discovery of tunicamycins as anti-complement agents.

Marine actinobacteria are potential producers of various secondary metabolites with diverse bioactivities. Among various bioactive compounds, anti-complement agents have received great interest for drug discovery to treat numerous diseases caused by inappropriate activation of the human complement system. However, marine streptomycetes producing anti-complement agents are still poorly explored. In this study, a marine-derived strain Streptomyces sp. DUT11 showing superior anti-complement activity was focused, and its genome sequence was analyzed. Gene clusters showing high similarities to that of tunicamycin and nonactin were identified, and their corresponding metabolites were also detected. Subsequently, tunicamycin I, V, and VII were isolated from Streptomyces sp. DUT11. Anti-complement assay showed that tunicamycin I, V, VII inhibited complement activation through the classic pathway, whereas no anti-complement activity of nonactin was detected. This is the first time that tunicamycins are reported to have such activity. In addition, genome analysis indicates that Streptomyces sp. DUT11 has the potential to produce novel lassopeptides and lantibiotics. These results suggest that marine Streptomyces are rich sources of anti-complement agents for drug discovery.

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September 22, 2019  |  

The complete genome sequence of Vibrio aestuarianus W-40 reveals virulence factor genes.

Vibrio aestuarianus is an opportunistic environmental pathogen that has been associated with epidemics in cultured shrimp Penaeus vannamei. Hepatopancreas microsporidian (HPM) and monodon slow growth syndrome (MSGS) have been reported in cultured P. vannamei. In this study, we sequenced and assembled the whole genome of V. aestuarianus strain W-40, a strain that was originally isolated from the intestines of an infected P. vannamei. The genome of V. aestuarianus strain W-40 contains two circular chromosomes of 483,7307 bp with a 46.23% GC content. We identified 4,457 open reading frames (ORFs) that occupy 86.35% of the genome. Vibrio aestuarianus strain W-40 consists primarily of the ATP-binding cassette (ABC) transporter system and the phosphotransferase system (PTS). CagA is a metabolism system that includes bacterial extracellular solute-binding protein. Glutathione reductase can purge superoxide radicals (O22-) and hydrogen peroxide (H2 O2 ) damage in V. aestuarianus strain W-40. The presence of two compete type I restriction-modification systems was confirmed. A total of 42 insertion sequences (IS) elements and 16 IS elements were identified. Our results revealed a host of virulence factors that likely contribute to the pathogenicity of V. aestuarianus strain W-40, including the virulence factor genes vacA, clpC, and bvgA, which are important for biofilm dispersion. Several bacitracin and tetracycline antibiotic resistance-encoding genes and type VI secretion systems were also identified in the genome. The complete genome sequence will aid future studies of the pathogenesis of V. aestuarianus strain W-40 and allow for new strategies to control disease to be developed.© 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

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September 22, 2019  |  

Comparative genomics provides insights into the marine adaptation in sponge-derived Kocuriaflava S43.

Sponge-derived actinomycetes represent a significant component of marine actinomycetes. Members of the genus Kocuria are distributed in various habitats such as soil, rhizosphere, clinical specimens, marine sediments, and sponges, however, to date, little is known about the mechanism of their environmental adaptation. Kocuria flava S43 was isolated from a coastal sponge. Phylogenetic analysis revealed that it was closely related to the terrestrial airborne K. flava HO-9041. In this study, to gain insights into the marine adaptation in K. flava S43 we sequenced the draft genome for K. flava S43 by third generation sequencing (TGS) and compared it with those of K. flava HO-9041 and some other Kocuria relatives. Comparative genomics and phylogenetic analyses revealed that K. flava S43 might adapt to the marine environment mainly by increasing the number of the genes linked to potassium homeostasis, resistance to heavy metals and phosphate metabolism, and acquiring the genes associated with electron transport and the genes encoding ATP-binding cassette (ABC) transporter, aquaporin, and thiol/disulfide interchange protein. Notably, gene acquisition was probably a primary mechanism of environmental adaptation in K. flava S43. Furthermore, this study also indicated that the Kocuria isolates from various marine and hyperosmotic environments possessed common genetic basis for environmental adaptation.

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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.

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September 22, 2019  |  

Comparative genomics of Pseudomonas sp. strain SI-3 associated with macroalga Ulva prolifera, the causative species for green tide in the Yellow Sea.

Algae-bacteria associations occurred widely in marine habitats, however, contributions of bacteria to macroalgal blooming were almost unknown. In this study, a potential endophytic strain SI-3 was isolated from Ulva prolifera, the causative species for the world’s largest green tide in the Yellow Sea, following a strict bleaching treatment to eliminate epiphytes. The genomic sequence of SI-3 was determined in size of 4.8 Mb and SI-3 was found to be mostly closed to Pseudomonas stutzeri. To evaluate the characteristics of SI-3 as a potential endophyte, the genomes of SI-3 and other 20 P. stutzeri strains were compared. We found that SI-3 had more strain-specific genes than most of the 20 P. stutzeri strains. Clusters of Orthologous Groups (COGs) analysis revealed that SI-3 had a higher proportion of genes assigned to transcriptional regulation and signal transduction compared with the 20 P. stutzeri strains, including four rhizosphere bacteria, indicating a complicated interaction network between SI-3 and its host. P. stutzeri is renowned for its metabolic versatility in aromatic compounds degradation. However, significant gene loss was observed in several aromatic compounds degradation pathways in SI-3, which may be an evolutional adaptation that developed upon association with its host. KEGG analysis revealed that dissimilatory nitrate reduction to ammonium (DNRA) and denitrification, two competing dissimilatory nitrate reduction pathways, co-occurred in the genome of SI-3, like most of the other 20 P. stutzeri strains. We speculated that DNRA of SI-3 may contribute a competitive advantage in nitrogen acquisition of U. prolifera by conserving nitrogen in NH4+ form, as in the case of microalgae bloom. Collectively, these data suggest that Pseudomonas sp. strain SI-3 was a suitable candidate for investigation of the algae-bacteria interaction with U. prolifera and the ecological impacts on algal blooming.

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September 22, 2019  |  

Quorum-quenching bacteria isolated from Red Sea sediments reduce biofilm formation by Pseudomonas aeruginosa.

Quorum sensing (QS) is the process by which bacteria communicate with each other through small signaling molecules such as N-acylhomoserine lactones (AHLs). Certain bacteria can degrade AHL molecules by a process called quorum quenching (QQ); therefore, QQ can be used to control bacterial infections and biofilm formation. In this study, we aimed to identify new species of bacteria with QQ activity. Red Sea sediments were collected either from the close vicinity of seagrass or from areas with no vegetation. We isolated 72 bacterial strains, which were tested for their ability to degrade/inactivate AHL molecules. Chromobacterium violaceum CV026-based bioassay was used for the initial screening of isolates with QQ activity. QQ activity was further quantified using high-performance liquid chromatography-tandem mass spectrometry. We found that these isolates could degrade AHL molecules of different acyl chain lengths as well as modifications. 16S-rRNA sequencing of positive QQ isolates showed that they belonged to three different genera. Specifically, two isolates belonged to the genus Erythrobacter; four, Labrenzia; and one, Bacterioplanes. The genome of one representative isolate from each genus was sequenced, and potential QQ enzymes, namely, lactonases and acylases, were identified. The ability of these isolates to degrade the 3OXOC12-AHLs produced by Pseudomonas aeruginosa PAO1 and hence inhibit biofilm formation was investigated. Our results showed that the isolate VG12 (genus Labrenzia) is better than other isolates at controlling biofilm formation by PAO1 and degradation of different AHL molecules. Time-course experiments to study AHL degradation showed that VG1 (genus Erythrobacter) could degrade AHLs faster than other isolates. Thus, QQ bacteria or enzymes can be used in combination with an antibacterial to overcome antibiotic resistance.

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September 22, 2019  |  

Analysis of the draft genome of the red seaweed Gracilariopsis chorda provides insights into genome size evolution in Rhodophyta.

Red algae (Rhodophyta) underwent two phases of large-scale genome reduction during their early evolution. The red seaweeds did not attain genome sizes or gene inventories typical of other multicellular eukaryotes. We generated a high-quality 92.1 Mb draft genome assembly from the red seaweed Gracilariopsis chorda, including methylation and small (s)RNA data. We analyzed these and other Archaeplastida genomes to address three questions: 1) What is the role of repeats and transposable elements (TEs) in explaining Rhodophyta genome size variation, 2) what is the history of genome duplication and gene family expansion/reduction in these taxa, and 3) is there evidence for TE suppression in red algae? We find that the number of predicted genes in red algae is relatively small (4,803-13,125 genes), particularly when compared with land plants, with no evidence of polyploidization. Genome size variation is primarily explained by TE expansion with the red seaweeds having the largest genomes. Long terminal repeat elements and DNA repeats are the major contributors to genome size growth. About 8.3% of the G. chorda genome undergoes cytosine methylation among gene bodies, promoters, and TEs, and 71.5% of TEs contain methylated-DNA with 57% of these regions associated with sRNAs. These latter results suggest a role for TE-associated sRNAs in RNA-dependent DNA methylation to facilitate silencing. We postulate that the evolution of genome size in red algae is the result of the combined action of TE spread and the concomitant emergence of its epigenetic suppression, together with other important factors such as changes in population size.

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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.

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September 22, 2019  |  

Extensive genomic diversity among Mycobacterium marinum strains revealed by whole genome sequencing.

Mycobacterium marinum is the causative agent for the tuberculosis-like disease mycobacteriosis in fish and skin lesions in humans. Ubiquitous in its geographical distribution, M. marinum is known to occupy diverse fish as hosts. However, information about its genomic diversity is limited. Here, we provide the genome sequences for 15 M. marinum strains isolated from infected humans and fish. Comparative genomic analysis of these and four available genomes of the M. marinum strains M, E11, MB2 and Europe reveal high genomic diversity among the strains, leading to the conclusion that M. marinum should be divided into two different clusters, the “M”- and the “Aronson”-type. We suggest that these two clusters should be considered to represent two M. marinum subspecies. Our data also show that the M. marinum pan-genome for both groups is open and expanding and we provide data showing high number of mutational hotspots in M. marinum relative to other mycobacteria such as Mycobacterium tuberculosis. This high genomic diversity might be related to the ability of M. marinum to occupy different ecological niches.

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