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September 22, 2019

A comprehensive understanding of the biocontrol potential of Bacillus velezensis LM2303 against Fusarium head blight.

Fusarium head blight (FHB) mainly caused by F. graminearum, always brings serious damage to wheat production worldwide. In this study, we found that strain LM2303 had strong antagonist activity against F. graminearum and significantly reduced disease severity of FHB with the control efficiency of 72.3% under field conditions. To gain a comprehensive understanding of the biocontrol potential of strain LM2303 against FHB, an integrated approach of genome mining and chemical analysis was employed. The whole genome of strain LM2303 was obtained and analyzed, showing the largest number of genes/gene clusters associated with biocontrol functions as compared with the known biocontrol strains (FZB42, M75, CAU B946). And strain LM2303 was accurately determined as a member of the B. velezensis clade using the phylogenomic analysis of single-copy core genes. Through genome mining, 13 biosynthetic gene clusters(BGCs) encoding secondary metabolites with biocontrol functions were identified, which were further confirmed through chemical analyses such as UHPLC-ESI-MS, including three antifungal metabolites (fengycin B, iturin A, and surfactin A), eight antibacterial metabolites (surfactin A, butirosin, plantazolicin and hydrolyzed plantazolicin, kijanimicin, bacilysin, difficidin, bacillaene A and bacillaene B, 7-o-malonyl macrolactin A and 7-o-succinyl macrolactin A), the siderophore bacillibactin, molybdenum cofactor and teichuronic acid. In addition, genes/gene clusters involved in plant colonization, plant growth promotion and induced systemic resistance were also found and analyzed, along with the corresponding metabolites. Finally, four different mechanisms of strain LM2303 involved in the biocontrol of FHB were putatively obtained. This work provides better insights into a mechanistic understanding of strain LM2303 in control of FHB, reinforcing the higher potential of this strain as a powerful biocontrol strain agent (BCA) for FHB control. The results also provide scientific reference and comparison for other biocontrol strains.

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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

Assembly of chromosome-scale contigs by efficiently resolving repetitive sequences with long reads

Due to the large number of repetitive sequences in complex eukaryotic genomes, fragmented and incompletely assembled genomes lose value as reference sequences, often due to short contigs that cannot be anchored or mispositioned onto chromosomes. Here we report a novel method Highly Efficient Repeat Assembly (HERA), which includes a new concept called a connection graph as well as algorithms for constructing the graph. HERA resolves repeats at high efficiency with single-molecule sequencing data, and enables the assembly of chromosome-scale contigs by further integrating genome maps and Hi-C data. We tested HERA with the genomes of rice R498, maize B73, human HX1 and Tartary buckwheat Pinku1. HERA can correctly assemble most of the tandemly repetitive sequences in rice using single-molecule sequencing data only. Using the same maize and human sequencing data published by Jiao et al. (2017) and Shi et al. (2016), respectively, we dramatically improved on the sequence contiguity compared with the published assemblies, increasing the contig N50 from 1.3 Mb to 61.2 Mb in maize B73 assembly and from 8.3 Mb to 54.4 Mb in human HX1 assembly with HERA. We provided a high-quality maize reference genome with 96.9% of the gaps filled (only 76 gaps left) and several incorrectly positioned sequences fixed compared with the B73 RefGen_v4 assembly. Comparisons between the HERA assembly of HX1 and the human GRCh38 reference genome showed that many gaps in GRCh38 could be filled, and that GRCh38 contained some potential errors that could be fixed. We assembled the Pinku1 genome into 12 scaffolds with a contig N50 size of 27.85 Mb. HERA serves as a new genome assembly/phasing method to generate high quality sequences for complex genomes and as a curation tool to improve the contiguity and completeness of existing reference genomes, including the correction of assembly errors in repetitive regions.

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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

Characterization of a novel multidrug resistance plasmid pSGB23 isolated from Salmonella enterica subspecies enterica serovar Saintpaul.

Salmonella enterica subspecies enterica serovar Saintpaul (S. Saintpaul) is an important gut pathogen which causes salmonellosis worldwide. Although intestinal salmonellosis is usually self-limiting, it can be life-threatening in children, the elderlies and immunocompromised patients. Appropriate antibiotic treatment is therefore required for these patients. However, the efficacy of many antibiotics on S. enterica infections has been greatly compromised due to spreading of multidrug resistance (MDR) plasmids, which poses serious threats on public health and needs to be closely monitored. In this study, we sequenced and fully characterized an S. enterica MDR plasmid pSGB23 isolated from chicken.Complete genome sequence analysis revealed that S. Saintpaul strain SGB23 harbored a 254 kb megaplasmid pSGB23, which carries 11 antibiotic resistance genes responsible for resistance to 9 classes of antibiotics and quaternary ammonium compounds that are commonly used to disinfect food processing facilities. Furthermore, we found that pSGB23 carries multiple conjugative systems, which allow it to spread into other Enterobacteriaceae spp. by self-conjugation. It also harbors multiple types of replicons and plasmid maintenance and addictive systems, which explains its broad host range and stable inheritance.We report here a novel MDR plasmid pSGB23 harboured by S. enterica. To our knowledge, it carried the greatest number of antibiotic resistance genes with the broadest range of resistance spectrum among S. enterica MDR plasmids identified so far. The isolation of pSGB23 from food sources is worrisome, while surveillance on its further spreading will be carried out based on the findings reported in this study.

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September 22, 2019

Homogenization of sub-genome secretome gene expression patterns in the allodiploid fungus Verticillium longisporum

Allopolyploidization, genome duplication through interspecific hybridization, is an important evolutionary mechanism that can enable organisms to adapt to environmental changes or stresses. The increased adaptive potential of allopolyploids can be particularly relevant for plant pathogens in their ongoing quest for host immune response evasion. To this end, plant pathogens secrete a plethora of molecules that enable host colonization. Allodiploidization has resulted in the new plant pathogen Verticillium longisporum that infects different hosts than haploid Verticillium species. To reveal the impact of allodiploidization on plant pathogen evolution, we studied the genome and transcriptome dynamics of V. longisporum using next-generation sequencing. V. longisporum genome evolution is characterized by extensive chromosomal rearrangements, between as well as within parental chromosome sets, leading to a mosaic genome structure. In comparison to haploid Verticillium species, V. longisporum genes display stronger signs of positive selection. The expression patterns of the two sub-genomes show remarkable resemblance, suggesting that the parental gene expression patterns homogenized upon hybridization. Moreover, whereas V. longisporum genes encoding secreted proteins frequently display differential expression between the parental sub-genomes in culture medium, expression patterns homogenize upon plant colonization. Collectively, our results illustrate of the adaptive potential of allodiploidy mediated by the interplay of two sub-genomes. Author summary Hybridization followed by whole-genome duplication, so-called allopolyploidization, provides genomic flexibility that is beneficial for survival under stressful conditions or invasiveness into new habitats. Allopolyploidization has mainly been studied in plants, but also occurs in other organisms, including fungi. Verticillium longisporum, an emerging fungal pathogen on brassicaceous plants, arose by allodiploidization between two Verticillium spp. We used comparative genomics to reveal the plastic nature of the V. longisporum genomes, showing that parental chromosome sets recombined extensively, resulting in a mosaic genome pattern. Furthermore, we show that non-synonymous substitutions frequently occurred in V. longisporum. Moreover, we reveal that expression patterns of genes encoding secreted proteins homogenized between the V. longisporum sub-genomes upon plant colonization. In conclusion, our results illustrate the large adaptive potential upon genome hybridization for fungi mediated by genomic plasticity and interaction between sub-genomes.

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September 22, 2019

Analysis of the complete genome sequence of Bacillus atrophaeus GQJK17 reveals its biocontrol characteristics as a plant growth-promoting rhizobacterium

Bacillus atrophaeus GQJK17 was isolated from the rhizosphere of Lycium barbarum L. in China, which was shown to be a plant growth-promoting rhizobacterium as a new biological agent against pathogenic fungi and gram-positive bacteria. We present its biological characteristics and complete genome sequence, which contains a 4,325,818 bp circular chromosome with 4,181 coding DNA sequences and a G+C content of 43.3%. A genome analysis revealed a total of 8 candidate gene clusters for producing antimicrobial secondary metabolites, including surfactin, bacillaene, fengycin, and bacillibactin. Some other antimicrobial and plant growth-promoting genes were also discovered. Our results provide insights into the genetic and biological basis of B. atrophaeus strains as a biocontrol agent for application in agriculture.

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September 22, 2019

Whole-genome comparison of high and low virulent Staphylococcus aureus isolates inducing implant-associated bone infections.

Staphylococcus aureus can cause wide range of infections from simple soft skin infections to severe endocarditis, bacteremia, osteomyelitis and implant associated bone infections (IABI). The focus of the present investigation was to study virulence properties of S. aureus isolates from acute and chronic IABI by means of their in vivo lethality, in vitro osteoblasts invasion, biofilm formation and subsequently whole genome comparison between high and low virulent strains. Application of insect infection model Galleria mellonella revealed high, intermediate and low virulence phenotypes of these clinical isolates, which showed good correlation with osteoblast invasion and biofilm formation assays. Comparative genomics of selected high (EDCC 5458) and low (EDCC 5464) virulent strains enabled the identification of molecular factors responsible for the development of acute and chronic IABI. Accordingly, the low virulent strain EDCC 5464 harbored point mutations resulting in frame shift mutations in agrC (histidine kinase in agr system), graS (histidine kinase in graSR, a two component system) and efeB (peroxidase in efeOBU operon, an iron acquisition system) genes. Additionally, we found a mobile element (present 11 copies in EDCC 5464) inserted at the end of ß-hemolysin (hlb) and sarU genes, which are involved in the pathogenesis and regulation of virulence gene expression in coordination with quorum sensing system. All these results are in good support with the low virulence behavior of EDCC 5464. From the previous literature, it is well known that agr defective S. aureus clinical strains are isolated from the chronic infections. Similarly, low virulent EDCC 5464 was isolated from chronic implant-associated bone infections infection whereas EDCC 5458 was obtained from acute implant-associated bone infections. Laboratory based in vitro and in vivo results and insights from comparative genomic analysis could be correlated with the clinical conclusion of IABIs and allows evidence-based treatment strategies based on the pathogenesis of the strain to cure life devastating implant-associated infections. Copyright © 2018 Elsevier GmbH. All rights reserved.

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September 22, 2019

Complete genome sequencing of exopolysaccharide-producing Lactobacillus plantarum K25 provides genetic evidence for the probiotic functionality and cold endurance capacity of the strain.

Lactobacillus plantarum (L. plantarum) K25 is a probiotic strain isolated from Tibetan kefir. Previous studies showed that this exopolysaccharide (EPS)-producing strain was antimicrobial active and cold tolerant. These functional traits were evidenced by complete genome sequencing of strain K25 with a circular 3,175,846-bp chromosome and six circular plasmids, encoding 3365 CDSs, 16 rRNA genes and 70 tRNA genes. Genomic analysis of L. plantarum K25 illustrates that this strain contains the previous reported mechanisms of probiotic functionality and cold tolerance, involving plantaricins, lysozyme, bile salt hydrolase, chaperone proteins, osmoprotectant, oxidoreductase, EPSs and terpenes. Interestingly, strain K25 harbors more genes that function in defense mechanisms, and lipid transport and metabolism, in comparison with other L. plantarum strains reported. The present study demonstrates the comprehensive analysis of genes related to probiotic functionalities of an EPS-producing L. plantarum strain based on whole genome sequencing.

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September 22, 2019

Biosynthesis of abscisic acid in fungi: identification of a sesquiterpene cyclase as the key enzyme in Botrytis cinerea.

While abscisic acid (ABA) is known as a hormone produced by plants through the carotenoid pathway, a small number of phytopathogenic fungi are also able to produce this sesquiterpene but they use a distinct pathway that starts with the cyclization of farnesyl diphosphate (FPP) into 2Z,4E-a-ionylideneethane which is then subjected to several oxidation steps. To identify the sesquiterpene cyclase (STC) responsible for the biosynthesis of ABA in fungi, we conducted a genomic approach in Botrytis cinerea. The genome of the ABA-overproducing strain ATCC58025 was fully sequenced and five STC-coding genes were identified. Among them, Bcstc5 exhibits an expression profile concomitant with ABA production. Gene inactivation, complementation and chemical analysis demonstrated that BcStc5/BcAba5 is the key enzyme responsible for the key step of ABA biosynthesis in fungi. Unlike what is observed for most of the fungal secondary metabolism genes, the key enzyme-coding gene Bcstc5/Bcaba5 is not clustered with the other biosynthetic genes, i.e., Bcaba1 to Bcaba4 that are responsible for the oxidative transformation of 2Z,4E-a-ionylideneethane. Finally, our study revealed that the presence of the Bcaba genes among Botrytis species is rare and that the majority of them do not possess the ability to produce ABA.© 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

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September 22, 2019

Genome biology of a novel lineage of planctomycetes widespread in anoxic aquatic environments.

Anaerobic strains affiliated with a novel order-level lineage of the Phycisphaerae class were retrieved from the suboxic zone of a hypersaline cyanobacterial mat and anoxic sediments of solar salterns. Genome sequences of five isolates were obtained and compared with metagenome-assembled genomes representing related uncultured bacteria from various anoxic aquatic environments. Gene content surveys suggest a strictly fermentative saccharolytic metabolism for members of this lineage, which could be confirmed by the phenotypic characterization of isolates. Genetic analyses indicate that the retrieved isolates do not have a canonical origin of DNA replication, but initiate chromosome replication at alternative sites possibly leading to an accelerated evolution. Further potential factors driving evolution and speciation within this clade include genome reduction by metabolic specialization and rearrangements of the genome by mobile genetic elements, which have a high prevalence in strains from hypersaline sediments and mats. Based on genetic and phenotypic data a distinct group of strictly anaerobic heterotrophic planctomycetes within the Phycisphaerae class could be assigned to a novel order that is represented by the proposed genus Sedimentisphaera gen. nov. comprising two novel species, S. salicampi gen. nov., sp. nov. and S. cyanobacteriorum gen. nov., sp. nov.© 2018 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

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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.

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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.

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September 22, 2019

Large-scale gene losses underlie the genome evolution of parasitic plant Cuscuta australis.

Dodders (Cuscuta spp., Convolvulaceae) are root- and leafless parasitic plants. The physiology, ecology, and evolution of these obligate parasites are poorly understood. A high-quality reference genome of Cuscuta australis was assembled. Our analyses reveal that Cuscuta experienced accelerated molecular evolution, and Cuscuta and the convolvulaceous morning glory (Ipomoea) shared a common whole-genome triplication event before their divergence. C. australis genome harbors 19,671 protein-coding genes, and importantly, 11.7% of the conserved orthologs in autotrophic plants are lost in C. australis. Many of these gene loss events likely result from its parasitic lifestyle and the massive changes of its body plan. Moreover, comparison of the gene expression patterns in Cuscuta prehaustoria/haustoria and various tissues of closely related autotrophic plants suggests that Cuscuta haustorium formation requires mostly genes normally involved in root development. The C. australis genome provides important resources for studying the evolution of parasitism, regressive evolution, and evo-devo in plant parasites.

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September 22, 2019

A mosaic monoploid reference sequence for the highly complex genome of sugarcane.

Sugarcane (Saccharum spp.) is a major crop for sugar and bioenergy production. Its highly polyploid, aneuploid, heterozygous, and interspecific genome poses major challenges for producing a reference sequence. We exploited colinearity with sorghum to produce a BAC-based monoploid genome sequence of sugarcane. A minimum tiling path of 4660 sugarcane BAC that best covers the gene-rich part of the sorghum genome was selected based on whole-genome profiling, sequenced, and assembled in a 382-Mb single tiling path of a high-quality sequence. A total of 25,316 protein-coding gene models are predicted, 17% of which display no colinearity with their sorghum orthologs. We show that the two species, S. officinarum and S. spontaneum, involved in modern cultivars differ by their transposable elements and by a few large chromosomal rearrangements, explaining their distinct genome size and distinct basic chromosome numbers while also suggesting that polyploidization arose in both lineages after their divergence.

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