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July 7, 2019

Hybrid de novo genome assembly of the Chinese herbal fleabane Erigeron breviscapus.

The plants in the Erigeron genus of the Compositae (Asteraceae) family are commonly called fleabanes, possibly due to the belief that certain chemicals in these plants repel fleas. In the traditional Chinese medicine, Erigeron breviscapus , which is native to China, was widely used in the treatment of cerebrovascular disease. A handful of bioactive compounds, including scutellarin, 3,5-dicaffeoylquinic acid, and 3,4-dicaffeoylquinic acid, have been isolated from the plant. With the purpose of finding novel medicinal compounds and understanding their biosynthetic pathways, we propose to sequence the genome of E. breviscapus . We assembled the highly heterozygous E. breviscapus genome using a combination of PacBio single-molecular real-time sequencing and next-generation sequencing methods on the Illumina HiSeq platform. The final draft genome is approximately 1.2 Gb, with contig and scaffold N50 sizes of 18.8 kb and 31.5 kb, respectively. Further analyses predicted 37 504 protein-coding genes in the E. breviscapus genome and 8172 shared gene families among Compositae species. The E. breviscapus genome provides a valuable resource for the investigation of novel bioactive compounds in this Chinese herb.

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July 7, 2019

Biosynthesis of 1a-hydroxycorticosterone in the winter skate Leucoraja ocellata: evidence to suggest a novel steroidogenic route.

The present study explores the ability of intracellular bacteria within the renal-inter-renal tissue of the winter skate Leucoraja ocellata to metabolize steroids and contribute to the synthesis of the novel elasmobranch corticosteroid, 1a-hydroxycorticosterone (1a-OH-B). Despite the rarity of C1 hydroxylation noted in the original identification of 1a-OH-B, literature provides evidence for steroid C1 hydroxylation by micro-organisms. Eight ureolytic bacterial isolates were identified in the renal-inter-renal tissue of L. ocellata, the latter being the site of 1a-OH-B synthesis. From incubations of bacterial isolates with known amounts of potential 1a-OH-B precursors, one isolate UM008 of the genus Rhodococcus was seen to metabolize corticosteroids and produce novel products via HPLC analysis. Cations Zn2+and Fe3+altered metabolism of certain steroid precursors, suggesting inhibition of Rhodococcus steroid catabolism. Genome sequencing of UM008 identified strong sequence and structural homology to that of Rhodococcus erythropolis PR4. A complete enzymatic pathway for steroid-ring oxidation as documented within other Actinobacteria was identified within the UM008 genome. This study highlights the potential role of Rhodococcus bacteria in steroid metabolism and proposes a novel alternative pathway for 1a-OH-B synthesis, suggesting a unique form of mutualism between intracellular bacteria and their elasmobranch host.© 2017 The Fisheries Society of the British Isles.

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July 7, 2019

Genome mining and predictive functional profiling of acidophilic rhizobacterium Pseudomonas fluorescens Pt14.

Pseudomonas fluorescens Pt14 is a non-pathogenic and acidophilic bacterium isolated from acidic soil (pH 4.65). Genome sequencing of strain Pt14 was performed using Single Molecule Real Time (SMRT) sequencing to get insights into unique existence of this strain in acidic environment. Complete genome sequence of this strain revealed a chromosome of 5,841,722 bp having 5354 CDSs and 88 RNAs. Phylogenomic reconstruction based on 16S rRNA gene, Average Nucleotide Identity (ANI) values and marker proteins revealed that strain Pt14 shared a common clade with P. fluorescens strain A506 and strain SS101. ANI value of strain Pt14 in relation to strain A506 was found 99.23% demonstrating a very close sub-species association at genome level. Further, orthology determination among these three phylogenetic neighbors revealed 4726 core proteins. Functional analysis elucidated significantly higher abundance of sulphur metabolism (>1×) which could be one of the reasons for the survival of strain Pt14 under acidic conditions (pH 4.65). Acidophilic bacteria have capability to oxidize sulphur into sulphuric acid which in turn can make the soil acidic and genome-wide analysis of P. fluorescens Pt14 demonstrated that this strain contributes towards making the soil acidic.

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July 7, 2019

Characterization of four endophytic fungi as potential consolidated bioprocessing hosts for conversion of lignocellulose into advanced biofuels.

Recently, several endophytic fungi have been demonstrated to produce volatile organic compounds (VOCs) with properties similar to fossil fuels, called “mycodiesel,” while growing on lignocellulosic plant and agricultural residues. The fact that endophytes are plant symbionts suggests that some may be able to produce lignocellulolytic enzymes, making them capable of both deconstructing lignocellulose and converting it into mycodiesel, two properties that indicate that these strains may be useful consolidated bioprocessing (CBP) hosts for the biofuel production. In this study, four endophytes Hypoxylon sp. CI4A, Hypoxylon sp. EC38, Hypoxylon sp. CO27, and Daldinia eschscholzii EC12 were selected and evaluated for their CBP potential. Analysis of their genomes indicates that these endophytes have a rich reservoir of biomass-deconstructing carbohydrate-active enzymes (CAZys), which includes enzymes active on both polysaccharides and lignin, as well as terpene synthases (TPSs), enzymes that may produce fuel-like molecules, suggesting that they do indeed have CBP potential. GC-MS analyses of their VOCs when grown on four representative lignocellulosic feedstocks revealed that these endophytes produce a wide spectrum of hydrocarbons, the majority of which are monoterpenes and sesquiterpenes, including some known biofuel candidates. Analysis of their cellulase activity when grown under the same conditions revealed that these endophytes actively produce endoglucanases, exoglucanases, and ß-glucosidases. The richness of CAZymes as well as terpene synthases identified in these four endophytic fungi suggests that they are great candidates to pursue for development into platform CBP organisms.

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July 7, 2019

Genetic and functional characterization of blaCTX-M-199, a novel tazobactam and sulbactam resistance-encoding gene located in a conjugative mcr-1-bearing IncI2 plasmid.

The study reported the genetic and functional characterization of a novel CTX-M-199 ß-lactamase, which was encoded by a blaCTX-M-64 variant gene found in a conjugative mcr-1-bearing IncI2 plasmid and exhibited resistance to ß-lactamase inhibitors, tazobactam and sulbactam. Copyright © 2017 American Society for Microbiology.

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July 7, 2019

Divergent and convergent modes of interaction between wheat and Puccinia graminis f. sp. tritici isolates revealed by the comparative gene co-expression network and genome analyses.

Two opposing evolutionary constraints exert pressure on plant pathogens: one to diversify virulence factors in order to evade plant defenses, and the other to retain virulence factors critical for maintaining a compatible interaction with the plant host. To better understand how the diversified arsenals of fungal genes promote interaction with the same compatible wheat line, we performed a comparative genomic analysis of two North American isolates of Puccinia graminis f. sp. tritici (Pgt).The patterns of inter-isolate divergence in the secreted candidate effector genes were compared with the levels of conservation and divergence of plant-pathogen gene co-expression networks (GCN) developed for each isolate. Comprative genomic analyses revealed substantial level of interisolate divergence in effector gene complement and sequence divergence. Gene Ontology (GO) analyses of the conserved and unique parts of the isolate-specific GCNs identified a number of conserved host pathways targeted by both isolates. Interestingly, the degree of inter-isolate sub-network conservation varied widely for the different host pathways and was positively associated with the proportion of conserved effector candidates associated with each sub-network. While different Pgt isolates tended to exploit similar wheat pathways for infection, the mode of plant-pathogen interaction varied for different pathways with some pathways being associated with the conserved set of effectors and others being linked with the diverged or isolate-specific effectors.Our data suggest that at the intra-species level pathogen populations likely maintain divergent sets of effectors capable of targeting the same plant host pathways. This functional redundancy may play an important role in the dynamic of the “arms-race” between host and pathogen serving as the basis for diverse virulence strategies and creating conditions where mutations in certain effector groups will not have a major effect on the pathogen’s ability to infect the host.

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July 7, 2019

Novel plasmid-mediated colistin resistance gene mcr-3 in Escherichia coli.

The mobile colistin resistance gene mcr-1 has attracted global attention, as it heralds the breach of polymyxins, one of the last-resort antibiotics for the treatment of severe clinical infections caused by multidrug-resistant Gram-negative bacteria. To date, six slightly different variants of mcr-1, and a second mobile colistin resistance gene, mcr-2, have been reported or annotated in the GenBank database. Here, we characterized a third mobile colistin resistance gene, mcr-3 The gene coexisted with 18 additional resistance determinants in the 261-kb IncHI2-type plasmid pWJ1 from porcine Escherichia colimcr-3 showed 45.0% and 47.0% nucleotide sequence identity to mcr-1 and mcr-2, respectively, while the deduced amino acid sequence of MCR-3 showed 99.8 to 100% and 75.6 to 94.8% identity to phosphoethanolamine transferases found in other Enterobacteriaceae species and in 10 Aeromonas species, respectively. pWJ1 was mobilized to an E. coli recipient by conjugation and contained a plasmid backbone similar to those of other mcr-1-carrying plasmids, such as pHNSHP45-2 from the original mcr-1-harboring E. coli strain. Moreover, a truncated transposon element, TnAs2, which was characterized only in Aeromonas salmonicida, was located upstream of mcr-3 in pWJ1. This ?TnAs2-mcr-3 element was also identified in a shotgun genome sequence of a porcine E. coli isolate from Malaysia, a human Klebsiella pneumoniae isolate from Thailand, and a human Salmonella enterica serovar Typhimurium isolate from the United States. These results suggest the likelihood of a wide dissemination of the novel mobile colistin resistance gene mcr-3 among Enterobacteriaceae and aeromonads; the latter may act as a potential reservoir for mcr-3IMPORTANCE The emergence of the plasmid-mediated colistin resistance gene mcr-1 has attracted substantial attention worldwide. Here, we examined a colistin-resistant Escherichia coli isolate that was negative for both mcr-1 and mcr-2 and discovered a novel mobile colistin resistance gene, mcr-3 The amino acid sequence of MCR-3 aligned closely with phosphoethanolamine transferases from Enterobacteriaceae and Aeromonas species originating from both clinical infections and environmental samples collected in 12 countries on four continents. Due to the ubiquitous profile of aeromonads in the environment and the potential transfer of mcr-3 between Enterobacteriaceae and Aeromonas species, the wide spread of mcr-3 may be largely underestimated. As colistin has been and still is widely used in veterinary medicine and used at increasing frequencies in human medicine, the continuous monitoring of mobile colistin resistance determinants in colistin-resistant Gram-negative bacteria is imperative for understanding and tackling the dissemination of mcr genes in both the agricultural and health care sectors. Copyright © 2017 Yin et al.

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July 7, 2019

Detection of diazotrophy in the acetylene-fermenting anaerobe, Pelobacter strain SFB93.

Acetylene (C2H2) is a trace constituent of the present Earth’s oxidizing atmosphere, reflecting a mix of terrestrial and marine emissions from anthropogenic, biomass burning, and unidentified biogenic sources. Fermentation of acetylene was serendipitously discovered during C2H2-block assays of N2O reductase, and Pelobacter acetylenicus was shown to grow on C2H2 via acetylene hydratase (AH). AH is a W-containing, catabolic, low redox potential enzyme that unlike nitrogenase (N2ase) is specific for acetylene. Acetylene fermentation is a rare metabolism that is well-characterized only in P. acetylenicus DSM3246 and DSM3247, and Pelobacter sp. strain SFB93. To better understand the genetic controls on AH activity, we sequenced the genomes of the three acetylene-fermenting Pelobacter strains. Genome assembly and annotation produced three novel genomes containing gene sequences for AH, with two copies being present in SFB93. In addition, gene sequences for all five compulsory genes for Mo-Fe nitrogenase were also present in the three genomes, indicating the co-occurrence of 2 acetylene-transformation pathways. Nitrogen fixation growth assays showed that DSM3426 could ferment acetylene in the absence of ammonium, but no ethylene was produced. However, SFB93 degraded acetylene, and in the absence of ammonium, produced ethylene indicating an active N2ase. Diazotrophic growth was observed under N2 but not in experimental controls incubated under Ar. SFB93 exhibits acetylene fermentation and nitrogen fixation, the only known biochemical mechanisms for acetylene transformation. Our results indicate complex interactions between N2ase and AH and suggest novel evolutionary pathways of these relic enzymes from early Earth to modern day.Importance Here we show that a single Pelobacter strain can grow via acetylene fermentation and carry out nitrogen fixation, using the only 2 enzymes known to transform acetylene. These findings provide new insights into acetylene transformations and adaptations for nutrient (C, N) and energy acquisition by microorganisms. Enhanced understanding of acetylene transformations in modern environments (i.e., extent, occurrence, rates, etc.) is important for using acetylene as a potential biomarker for extraterrestrial life and degradation of anthropogenic contaminants. Copyright © 2017 American Society for Microbiology.

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July 7, 2019

Coping with living in the soil: the genome of the parthenogenetic springtail Folsomia candida.

Folsomia candida is a model in soil biology, belonging to the family of Isotomidae, subclass Collembola. It reproduces parthenogenetically in the presence of Wolbachia, and exhibits remarkable physiological adaptations to stress. To better understand these features and adaptations to life in the soil, we studied its genome in the context of its parthenogenetic lifestyle.We applied Pacific Bioscience sequencing and assembly to generate a reference genome for F. candida of 221.7 Mbp, comprising only 162 scaffolds. The complete genome of its endosymbiont Wolbachia, was also assembled and turned out to be the largest strain identified so far. Substantial gene family expansions and lineage-specific gene clusters were linked to stress response. A large number of genes (809) were acquired by horizontal gene transfer. A substantial fraction of these genes are involved in lignocellulose degradation. Also, the presence of genes involved in antibiotic biosynthesis was confirmed. Intra-genomic rearrangements of collinear gene clusters were observed, of which 11 were organized as palindromes. The Hox gene cluster of F. candida showed major rearrangements compared to arthropod consensus cluster, resulting in a disorganized cluster.The expansion of stress response gene families suggests that stress defense was important to facilitate colonization of soils. The large number of HGT genes related to lignocellulose degradation could be beneficial to unlock carbohydrate sources in soil, especially those contained in decaying plant and fungal organic matter. Intra- as well as inter-scaffold duplications of gene clusters may be a consequence of its parthenogenetic lifestyle. This high quality genome will be instrumental for evolutionary biologists investigating deep phylogenetic lineages among arthropods and will provide the basis for a more mechanistic understanding in soil ecology and ecotoxicology.

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July 7, 2019

Genomic characterization of a large plasmid containing a bla NDM-1 gene carried on Salmonella enterica serovar Indiana C629 isolate from China.

The bla NDM-1 gene in Salmonella species is mostly reported in clinical cases, but is rarely isolated from red and white meat in China.A Salmonella Indiana (S. Indiana) isolate was cultured from a chicken carcass procured from a slaughterhouse in China. Antimicrobial susceptibility was tested against a panel of agents. Whole-genome sequencing of the isolate was carried out and data was analyzed.A large plasmid, denoted as plasmid pC629 (210,106 bp), containing a composite cassette, consisting of IS26-bla NDM-1-ble MBL -?trpF-tat-cutA-ISCR1-sul1-qacE?1-aadA2-dfrA12-intI1-IS26 was identified. The latter locus was physically linked with bla OXA-1, bla CTX-M-65, bla TEM-1-encoding genes. A mercury resistance operon merACDEPTR was also identified; it was flanked on the proximal side, among IS26 element and the distally located on the bla NDM-1 gene. Plasmid pC629 also contained 21 other antimicrobial resistance-encoding genes, such as aac(6′)-Ib-cr, aac(3)-VI, aadA5, aph(4)-Ia, arr-3, blmS, brp, catB3, dfrA17, floR, fosA, mph(A), mphR, mrx, nimC/nimA, oqxA, oqxB, oqxR, rmtB, sul1, sul2. Two virulence genes were also identified on plasmid pC629.To the best of our knowledge, this is the first report of bla NDM-1 gene being identified from a plasmid in a S. Indiana isolate cultured from chicken carcass in China.

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July 7, 2019

Maize defective kernel mutant generated by insertion of a Ds element in a gene encoding a highly conserved TTI2 cochaperone.

We have used the newly engineered transposable element Dsg to tag a gene that gives rise to a defective kernel (dek) phenotype. Dsg requires the autonomous element Ac for transposition. Upon excision, it leaves a short DNA footprint that can create in-frame and frameshift insertions in coding sequences. Therefore, we could create alleles of the tagged gene that confirmed causation of the dek phenotype by the Dsg insertion. The mutation, designated dek38-Dsg, is embryonic lethal, has a defective basal endosperm transfer (BETL) layer, and results in a smaller seed with highly underdeveloped endosperm. The maize dek38 gene encodes a TTI2 (Tel2-interacting protein 2) molecular cochaperone. In yeast and mammals, TTI2 associates with two other cochaperones, TEL2 (Telomere maintenance 2) and TTI1 (Tel2-interacting protein 1), to form the triple T complex that regulates DNA damage response. Therefore, we cloned the maize Tel2 and Tti1 homologs and showed that TEL2 can interact with both TTI1 and TTI2 in yeast two-hybrid assays. The three proteins regulate the cellular levels of phosphatidylinositol 3-kinase-related kinases (PIKKs) and localize to the cytoplasm and the nucleus, consistent with known subcellular locations of PIKKs. dek38-Dsg displays reduced pollen transmission, indicating TTI2’s importance in male reproductive cell development.

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July 7, 2019

Expanding landscapes of the diversified mcr-1-bearing plasmid reservoirs.

Polymyxin is a cationic polypeptide antibiotic that can disrupt bacterial cell membrane by interacting with its lipopolysaccharide molecules and is used as a last resort drug against lethal infections by the carbapenem-resistant superbugs (like NDM-1). However, global discovery of the MCR-1 colistin resistance dramatically challenges the newly renewed interest in colistin for clinical use.The mcr-1-harboring plasmids were acquired from swine and human Escherichia coli isolated in China, from 2015 to 2016, and subjected to Illumina PacBio RSII and Hi-Seq2000 for full genome sequencing. PCR was applied to close the gap of the assembled contigs. Ori-Finder was employed to predict the replication origin (oriC) in plasmids. The phenotype of MCR-1-producing isolates was evaluated on the LBA plates with various level of colistin. Genetic deletion was used to test the requirement of the initial “ATG” codon for the MCR-1 function.Here, we report full genomes of over 10 mcr-1-harboring plasmids with diversified replication incompatibilities. A novel hybrid IncI2/IncFIB plasmid pGD17-2 was discovered and characterized from a swine isolate with colistin resistance. Intriguingly, co-occurrence of two unique mcr-1-bearing plasmids (pGD65-3, IncI2, and pGD65-5, IncX4) was detected in a single isolate GD65, which might accelerate dissemination of the mcr-1 under environmental selection pressure. Genetic analyses of these plasmids mapped mobile elements in the context of antibiotic resistance and determined two insertion sequences (ISEcp1 and ISApl1) that are responsible for the mobilization of mcr-1. Gene deletion also proved that the first ATG codon is redundant in the mcr-1 gene.Collectively, our results extend landscapes of the diversified mcr-1-bearing plasmid reservoirs.

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July 7, 2019

Genome sequencing reveals the origin of the allotetraploid Arabidopsis suecica.

Polyploidy is an example of instantaneous speciation when it involves the formation of a new cytotype that is incompatible with the parental species. Because new polyploid individuals are likely to be rare, establishment of a new species is unlikely unless polyploids are able to reproduce through self-fertilization (selfing), or asexually. Conversely, selfing (or asexuality) makes it possible for polyploid species to originate from a single individual-a bona fide speciation event. The extent to which this happens is not known. Here, we consider the origin of Arabidopsis suecica, a selfing allopolyploid between Arabidopsis thaliana and Arabidopsis arenosa, which has hitherto been considered to be an example of a unique origin. Based on whole-genome re-sequencing of 15 natural A. suecica accessions, we identify ubiquitous shared polymorphism with the parental species, and hence conclusively reject a unique origin in favor of multiple founding individuals. We further estimate that the species originated after the last glacial maximum in Eastern Europe or central Eurasia (rather than Sweden, as the name might suggest). Finally, annotation of the self-incompatibility loci in A. suecica revealed that both loci carry non-functional alleles. The locus inherited from the selfing A. thaliana is fixed for an ancestral non-functional allele, whereas the locus inherited from the outcrossing A. arenosa is fixed for a novel loss-of-function allele. Furthermore, the allele inherited from A. thaliana is predicted to transcriptionally silence the allele inherited from A. arenosa, suggesting that loss of self-incompatibility may have been instantaneous.© The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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