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September 22, 2019  |  

Comparative genomics of the Baltic Sea toxic cyanobacteria Nodularia spumigena UHCC 0039 and its response to varying salinity.

Salinity is an important abiotic factor controlling the distribution and abundance of Nodularia spumigena, the dominating diazotrophic and toxic phototroph, in the brackish water cyanobacterial blooms of the Baltic Sea. To expand the available genomic information for brackish water cyanobacteria, we sequenced the isolate Nodularia spumigena UHCC 0039 using an Illumina-SMRT hybrid sequencing approach, revealing a chromosome of 5,294,286 base pairs (bp) and a single plasmid of 92,326 bp. Comparative genomics in Nostocales showed pronounced genetic similarity among Nodularia spumigena strains evidencing their short evolutionary history. The studied Baltic Sea strains share similar sets of CRISPR-Cas cassettes and a higher number of insertion sequence (IS) elements compared to Nodularia spumigena CENA596 isolated from a shrimp production pond in Brazil. Nodularia spumigena UHCC 0039 proliferated similarly at three tested salinities, whereas the lack of salt inhibited its growth and triggered transcriptome remodeling, including the up-regulation of five sigma factors and the down-regulation of two other sigma factors, one of which is specific for strain UHCC 0039. Down-regulated genes additionally included a large genetic region for the synthesis of two yet unidentified natural products. Our results indicate a remarkable plasticity of the Nodularia salinity acclimation, and thus salinity strongly impacts the intensity and distribution of cyanobacterial blooms in the Baltic Sea.

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September 22, 2019  |  

Whole genome sequencing of greater amberjack (Seriola dumerili) for SNP identification on aligned scaffolds and genome structural variation analysis using parallel resequencing

Greater amberjack (Seriola dumerili) is distributed in tropical and temperate waters worldwide and is an important aquaculture fish. We carried out de novo sequencing of the greater amberjack genome to construct a reference genome sequence to identify single nucleotide polymorphisms (SNPs) for breeding amberjack by marker-assisted or gene-assisted selection as well as to identify functional genes for biological traits. We obtained 200 times coverage and constructed a high-quality genome assembly using next generation sequencing technology. The assembled sequences were aligned onto a yellowtail (Seriola quinqueradiata) radiation hybrid (RH) physical map by sequence homology. A total of 215 of the longest amberjack sequences, with a total length of 622.8?Mbp (92% of the total length of the genome scaffolds), were lined up on the yellowtail RH map. We resequenced the whole genomes of 20 greater amberjacks and mapped the resulting sequences onto the reference genome sequence. About 186,000 nonredundant SNPs were successfully ordered on the reference genome. Further, we found differences in the genome structural variations between two greater amberjack populations using BreakDancer. We also analyzed the greater amberjack transcriptome and mapped the annotated sequences onto the reference genome sequence.

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September 22, 2019  |  

Repeat-driven generation of antigenic diversity in a major human pathogen, Trypanosoma cruzi

Trypanosoma cruzi, a zoonotic kinetoplastid protozoan with a complex genome, is the causative agent of American trypanosomiasis (Chagas disease). The parasite uses a highly diverse repertoire of surface molecules, with roles in cell invasion, immune evasion and pathogenesis. Thus far, the genomic regions containing these genes have been impossible to resolve and it has been impossible to study the structure and function of the several thousand repetitive genes encoding the surface molecules of the parasite. We here present an improved genome assembly of a T. cruzi clade I (TcI) strain using high coverage PacBio single molecule sequencing, together with Illumina sequencing of 34 T. cruzi TcI isolates and clones from different geographic locations, sample sources and clinical outcomes. Resolution of the surface molecule gene structure reveals an unusual duality in the organisation of the parasite genome, a core genomic region syntenous with related protozoa flanked by unique and highly plastic subtelomeric regions encoding surface antigens. The presence of abundant interspersed retrotransposons in the subtelomeres suggests that these elements are involved in a recombination mechanism for the generation of antigenic variation and evasion of the host immune response. The comparative genomic analysis of the cohort of TcI strains revealed multiple cases of such recombination events involving surface molecule genes and has provided new insights into T. cruzi population structure.

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September 22, 2019  |  

Stress-adaptive responses associated with high-level carbapenem resistance in KPC-producing Klebsiella pneumoniae.

Carbapenem-resistant Enterobacteriaceae (CRE) organisms have emerged to become a major global public health threat among antimicrobial resistant bacterial human pathogens. Little is known about how CREs emerge. One characteristic phenotype of CREs is heteroresistance, which is clinically associated with treatment failure in patients given a carbapenem. Through in vitro whole-transcriptome analysis we tracked gene expression over time in two different strains (BR7, BR21) of heteroresistant KPC-producing Klebsiella pneumoniae, first exposed to a bactericidal concentration of imipenem followed by growth in drug-free medium. In both strains, the immediate response was dominated by a shift in expression of genes involved in glycolysis toward those involved in catabolic pathways. This response was followed by global dampening of transcriptional changes involving protein translation, folding and transport, and decreased expression of genes encoding critical junctures of lipopolysaccharide biosynthesis. The emerged high-level carbapenem-resistant BR21 subpopulation had a prophage (IS1) disrupting ompK36 associated with irreversible OmpK36 porin loss. On the other hand, OmpK36 loss in BR7 was reversible. The acquisition of high-level carbapenem resistance by the two heteroresistant strains was associated with distinct and shared stepwise transcriptional programs. Carbapenem heteroresistance may emerge from the most adaptive subpopulation among a population of cells undergoing a complex set of stress-adaptive responses.

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September 22, 2019  |  

The global distribution and spread of the mobilized colistin resistance gene mcr-1.

Colistin represents one of the few available drugs for treating infections caused by carbapenem-resistant Enterobacteriaceae. As such, the recent plasmid-mediated spread of the colistin resistance gene mcr-1 poses a significant public health threat, requiring global monitoring and surveillance. Here, we characterize the global distribution of mcr-1 using a data set of 457 mcr-1-positive sequenced isolates. We find mcr-1 in various plasmid types but identify an immediate background common to all mcr-1 sequences. Our analyses establish that all mcr-1 elements in circulation descend from the same initial mobilization of mcr-1 by an ISApl1 transposon in the mid 2000s (2002-2008; 95% highest posterior density), followed by a marked demographic expansion, which led to its current global distribution. Our results provide the first systematic phylogenetic analysis of the origin and spread of mcr-1, and emphasize the importance of understanding the movement of antibiotic resistance genes across multiple levels of genomic organization.

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September 22, 2019  |  

Ploidy variation in Kluyveromyces marxianus separates dairy and non-dairy isolates.

Kluyveromyces marxianus is traditionally associated with fermented dairy products, but can also be isolated from diverse non-dairy environments. Because of thermotolerance, rapid growth and other traits, many different strains are being developed for food and industrial applications but there is, as yet, little understanding of the genetic diversity or population genetics of this species. K. marxianus shows a high level of phenotypic variation but the only phenotype that has been clearly linked to a genetic polymorphism is lactose utilisation, which is controlled by variation in the LAC12 gene. The genomes of several strains have been sequenced in recent years and, in this study, we sequenced a further nine strains from different origins. Analysis of the Single Nucleotide Polymorphisms (SNPs) in 14 strains was carried out to examine genome structure and genetic diversity. SNP diversity in K. marxianus is relatively high, with up to 3% DNA sequence divergence between alleles. It was found that the isolates include haploid, diploid, and triploid strains, as shown by both SNP analysis and flow cytometry. Diploids and triploids contain long genomic tracts showing loss of heterozygosity (LOH). All six isolates from dairy environments were diploid or triploid, whereas 6 out 7 isolates from non-dairy environment were haploid. This also correlated with the presence of functional LAC12 alleles only in dairy haplotypes. The diploids were hybrids between a non-dairy and a dairy haplotype, whereas triploids included three copies of a dairy haplotype.

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September 22, 2019  |  

Targeted sequencing by gene synteny, a new strategy for polyploid species: sequencing and physical structure of a complex sugarcane region.

Sugarcane exhibits a complex genome mainly due to its aneuploid nature and high ploidy level, and sequencing of its genome poses a great challenge. Closely related species with well-assembled and annotated genomes can be used to help assemble complex genomes. Here, a stable quantitative trait locus (QTL) related to sugar accumulation in sorghum was successfully transferred to the sugarcane genome. Gene sequences related to this QTL were identified in silico from sugarcane transcriptome data, and molecular markers based on these sequences were developed to select bacterial artificial chromosome (BAC) clones from the sugarcane variety SP80-3280. Sixty-eight BAC clones containing at least two gene sequences associated with the sorghum QTL were sequenced using Pacific Biosciences (PacBio) technology. Twenty BAC sequences were found to be related to the syntenic region, of which nine were sufficient to represent this region. The strategy we propose is called “targeted sequencing by gene synteny,” which is a simpler approach to understanding the genome structure of complex genomic regions associated with traits of interest.

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September 22, 2019  |  

A large-scale comparative metagenomic study reveals the functional interactions in six bloom-forming Microcystis-epibiont communities.

Cyanobacterial blooms are worldwide issues of societal concern and scientific interest. Lake Taihu and Lake Dianchi, two of the largest lakes in China, have been suffering from annual Microcystis-based blooms over the past two decades. These two eutrophic lakes differ in both nutrient load and environmental parameters, where Microcystis microbiota consisting of different Microcystis morphospecies and associated bacteria (epibionts) have dominated. We conducted a comprehensive metagenomic study that analyzed species diversity, community structure, functional components, metabolic pathways and networks to investigate functional interactions among the members of six Microcystis-epibiont communities in these two lakes. Our integrated metagenomic pipeline consisted of efficient assembly, binning, annotation, and quality assurance methods that ensured high-quality genome reconstruction. This study provides a total of 68 reconstructed genomes including six complete Microcystis genomes and 28 high quality bacterial genomes of epibionts belonging to 14 distinct taxa. This metagenomic dataset constitutes the largest reference genome catalog available for genome-centric studies of the Microcystis microbiome. Epibiont community composition appears to be dynamic rather than fixed, and the functional profiles of communities were related to the environment of origin. This study demonstrates mutualistic interactions between Microcystis and epibionts at genetic and metabolic levels. Metabolic pathway reconstruction provided evidence for functional complementation in nitrogen and sulfur cycles, fatty acid catabolism, vitamin synthesis, and aromatic compound degradation among community members. Thus, bacterial social interactions within Microcystis-epibiont communities not only shape species composition, but also stabilize the communities functional profiles. These interactions appear to play an important role in environmental adaptation of Microcystis colonies.

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September 22, 2019  |  

Comparative genomics of smut pathogens: Insights from orphans and positively selected genes into host specialization.

Host specialization is a key evolutionary process for the diversification and emergence of new pathogens. However, the molecular determinants of host range are poorly understood. Smut fungi are biotrophic pathogens that have distinct and narrow host ranges based on largely unknown genetic determinants. Hence, we aimed to expand comparative genomics analyses of smut fungi by including more species infecting different hosts and to define orphans and positively selected genes to gain further insights into the genetics basis of host specialization. We analyzed nine lineages of smut fungi isolated from eight crop and non-crop hosts: maize, barley, sugarcane, wheat, oats, Zizania latifolia (Manchurian rice), Echinochloa colona (a wild grass), and Persicaria sp. (a wild dicot plant). We assembled two new genomes: Ustilago hordei (strain Uhor01) isolated from oats and U. tritici (strain CBS 119.19) isolated from wheat. The smut genomes were of small sizes, ranging from 18.38 to 24.63 Mb. U. hordei species experienced genome expansions due to the proliferation of transposable elements and the amount of these elements varied among the two strains. Phylogenetic analysis confirmed that Ustilago is not a monophyletic genus and, furthermore, detected misclassification of the U. tritici specimen. The comparison between smut pathogens of crop and non-crop hosts did not reveal distinct signatures, suggesting that host domestication did not play a dominant role in shaping the evolution of smuts. We found that host specialization in smut fungi likely has a complex genetic basis: different functional categories were enriched in orphans and lineage-specific selected genes. The diversification and gain/loss of effector genes are probably the most important determinants of host specificity.

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September 22, 2019  |  

Evolution of sequence type 4821 clonal complex meningococcal strains in China from prequinolone to quinolone era, 1972-2013.

The expansion of hypervirulent sequence type 4821 clonal complex (CC4821) lineage Neisseria meningitidis bacteria has led to a shift in meningococcal disease epidemiology in China, from serogroup A (MenA) to MenC. Knowledge of the evolution and genetic origin of the emergent MenC strains is limited. In this study, we subjected 76 CC4821 isolates collected across China during 1972-1977 and 2005-2013 to phylogenetic analysis, traditional genotyping, or both. We show that successive recombination events within genes encoding surface antigens and acquisition of quinolone resistance mutations possibly played a role in the emergence of CC4821 as an epidemic clone in China. MenC and MenB CC4821 strains have spread across China and have been detected in several countries in different continents. Capsular switches involving serogroups B and C occurred among epidemic strains, raising concerns regarding possible increases in MenB disease, given that vaccines in use in China do not protect against MenB.

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September 22, 2019  |  

The genome sequence of “Candidatus Fokinia solitaria”: Insights on reductive evolution in Rickettsiales.

Candidatus Fokinia solitaria is an obligate intracellular endosymbiont of a unicellular eukaryote, a ciliate of the genus Paramecium. Here, we present the genome sequence of this bacterium and subsequent analysis. Phylogenomic analysis confirmed the previously reported positioning of the symbiont within the “Candidatus Midichloriaceae” family (order Rickettsiales), as well as its high sequence divergence from other members of the family, indicative of fast sequence evolution. Consistently with this high evolutionary rate, a comparative genomic analysis revealed that the genome of this symbiont is the smallest of the Rickettsiales to date. The reduced genome does not present flagellar genes, nor the pathway for the biosynthesis of lipopolysaccharides (present in all the other so far sequenced members of the family “Candidatus Midichloriaceae”) or genes for the Krebs cycle (present, although not always complete, in Rickettsiales). These results indicate an evolutionary trend toward a stronger dependence on the host, in comparison with other members of the family. Two alternative scenarios are compatible with our results; “Candidatus Fokinia solitaria” could be either a recently evolved, vertically transmitted mutualist, or a parasite with a high host-specificity.

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September 22, 2019  |  

Developing collaborative works for faster progress on fungal respiratory infections in cystic fibrosis.

Cystic fibrosis (CF) is the major genetic inherited disease in Caucasian populations. The respiratory tract of CF patients displays a sticky viscous mucus, which allows for the entrapment of airborne bacteria and fungal spores and provides a suitable environment for growth of microorganisms, including numerous yeast and filamentous fungal species. As a consequence, respiratory infections are the major cause of morbidity and mortality in this clinical context. Although bacteria remain the most common agents of these infections, fungal respiratory infections have emerged as an important cause of disease. Therefore, the International Society for Human and Animal Mycology (ISHAM) has launched a working group on Fungal respiratory infections in Cystic Fibrosis (Fri-CF) in October 2006, which was subsequently approved by the European Confederation of Medical Mycology (ECMM). Meetings of this working group, comprising both clinicians and mycologists involved in the follow-up of CF patients, as well as basic scientists interested in the fungal species involved, provided the opportunity to initiate collaborative works aimed to improve our knowledge on these infections to assist clinicians in patient management. The current review highlights the outcomes of some of these collaborative works in clinical surveillance, pathogenesis and treatment, giving special emphasis to standardization of culture procedures, improvement of species identification methods including the development of nonculture-based diagnostic methods, microbiome studies and identification of new biological markers, and the description of genotyping studies aiming to differentiate transient carriage and chronic colonization of the airways. The review also reports on the breakthrough in sequencing the genomes of the main Scedosporium species as basis for a better understanding of the pathogenic mechanisms of these fungi, and discusses treatment options of infections caused by multidrug resistant microorganisms, such as Scedosporium and Lomentospora species and members of the Rasamsonia argillacea species complex.

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September 22, 2019  |  

Genomic analyses of unique carbohydrate and phytohormone metabolism in the macroalga Gracilariopsis lemaneiformis (Rhodophyta).

Red algae are economically valuable for food and in industry. However, their genomic information is limited, and the genomic data of only a few species of red algae have been sequenced and deposited recently. In this study, we annotated a draft genome of the macroalga Gracilariopsis lemaneiformis (Gracilariales, Rhodophyta).The entire 88.98 Mb genome of Gp. lemaneiformis 981 was generated from 13,825 scaffolds (=500 bp) with an N50 length of 30,590 bp, accounting for approximately 91% of this algal genome. A total of 38.73 Mb of scaffold sequences were repetitive, and 9281 protein-coding genes were predicted. A phylogenomic analysis of 20 genomes revealed the relationship among the Chromalveolata, Rhodophyta, Chlorophyta and higher plants. Homology analysis indicated phylogenetic proximity between Gp. lemaneiformis and Chondrus crispus. The number of enzymes related to the metabolism of carbohydrates, including agar, glycoside hydrolases, glycosyltransferases, was abundant. In addition, signaling pathways associated with phytohormones such as auxin, salicylic acid and jasmonates are reported for the first time for this alga.We sequenced and analyzed a draft genome of the red alga Gp. lemaneiformis, and revealed its carbohydrate metabolism and phytohormone signaling characteristics. This work will be helpful in research on the functional and comparative genomics of the order Gracilariales and will enrich the genomic information on marine algae.

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September 22, 2019  |  

A whole genome assembly of the horn fly, Haematobia irritans, and prediction of genes with roles in metabolism and sex determination.

Haematobia irritans, commonly known as the horn fly, is a globally distributed blood-feeding pest of cattle that is responsible for significant economic losses to cattle producers. Chemical insecticides are the primary means for controlling this pest but problems with insecticide resistance have become common in the horn fly. To provide a foundation for identification of genomic loci for insecticide resistance and for discovery of new control technology, we report the sequencing, assembly, and annotation of the horn fly genome. The assembled genome is 1.14 Gb, comprising 76,616 scaffolds with N50 scaffold length of 23 Kb. Using RNA-Seq data, we have predicted 34,413 gene models of which 19,185 have been assigned functional annotations. Comparative genomics analysis with the Dipteran flies Musca domestica L., Drosophila melanogaster, and Lucilia cuprina, show that the horn fly is most closely related to M. domestica, sharing 8,748 orthologous clusters followed by D. melanogaster and L. cuprina, sharing 7,582 and 7,490 orthologous clusters respectively. We also identified a gene locus for the sodium channel protein in which mutations have been previously reported that confers target site resistance to the most common class of pesticides used in fly control. Additionally, we identified 276 genomic loci encoding members of metabolic enzyme gene families such as cytochrome P450s, esterases and glutathione S-transferases, and several genes orthologous to sex determination pathway genes in other Dipteran species. Copyright © 2018 Konganti et al.

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September 22, 2019  |  

Inpactor, integrated and parallel analyzer and classifier of LTR retrotransposons and its application for pineapple LTR retrotransposons diversity and dynamics.

One particular class of Transposable Elements (TEs), called Long Terminal Repeats (LTRs), retrotransposons, comprises the most abundant mobile elements in plant genomes. Their copy number can vary from several hundreds to up to a few million copies per genome, deeply affecting genome organization and function. The detailed classification of LTR retrotransposons is an essential step to precisely understand their effect at the genome level, but remains challenging in large-sized genomes, requiring the use of optimized bioinformatics tools that can take advantage of supercomputers. Here, we propose a new tool: Inpactor, a parallel and scalable pipeline designed to classify LTR retrotransposons, to identify autonomous and non-autonomous elements, to perform RT-based phylogenetic trees and to analyze their insertion times using High Performance Computing (HPC) techniques. Inpactor was tested on the classification and annotation of LTR retrotransposons in pineapple, a recently-sequenced genome. The pineapple genome assembly comprises 44% of transposable elements, of which 23% were classified as LTR retrotransposons. Exceptionally, 16.4% of the pineapple genome assembly corresponded to only one lineage of the Gypsy superfamily: Del, suggesting that this particular lineage has undergone a significant increase in its copy numbers. As demonstrated for the pineapple genome, Inpactor provides comprehensive data of LTR retrotransposons’ classification and dynamics, allowing a fine understanding of their contribution to genome structure and evolution. Inpactor is available at https://github.com/simonorozcoarias/Inpactor.

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