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April 21, 2020

Complete Genome of Bacillus velezensis CMT-6 and Comparative Genome Analysis Reveals Lipopeptide Diversity.

The complete genome sequence of Bacillus velezensis type strain CMT-6 is presented for the first time. A comparative analysis between the genome sequences of CMT-6 with the genome of Bacillus amyloliquefaciens DSM7T, B. velezensis FZB42, and Bacillus subtilis 168 revealed major differences in the lipopeptide synthesis genes. Of the above, only the CMT-6 strain possessed an integrated synthetase gene for synthesizing surfactin, iturin, and fengycin. However, CMT-6 shared 14, 12, and 10 other lipopeptide-producing genes with FZB42, DSM7T, and 168 respectively. The largest numbers of non-synonymous mutations were detected in 205 gene sequences that produced these three lipopeptides in CMT-6 and 168. Comparing CMT-6 with DSM7T, 58 non-synonymous mutations were detected in gene sequences that contributed to produce lipopeptides. In addition, InDels were identified in yczE and glnR genes. CMT-6 and FZB42 had the lowest number of non-synonymous mutations with 8 lipopeptide-related gene sequences. And InDels were identified in only yczE. The numbers of core genes, InDels, and non-synonymous mutations in genes were the main reasons for the differences in yield and variety of lipopeptides. These results will enrich the genomic resources available for B. velezensis and provide fundamental information to construct strains that can produce specific lipopeptides.

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April 21, 2020

Complete genome sequence of Bacillus velezensis JT3-1, a microbial germicide isolated from yak feces

Bacillus velezensis JT3-1 is a probiotic strain isolated from feces of the domestic yak (Bos grunniens) in the Gansu province of China. It has strong antagonistic activity against Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, Salmonella Typhimurium, Mannheimia haemolytica, Staphylococcus hominis, Clostridium perfringens, and Mycoplasma bovis. These properties have made the JT3-1 strain the focus of commercial interest. In this study, we describe the complete genome sequence of JT3-1, with a genome size of 3,929,799 bp, 3761 encoded genes and an average GC content of 46.50%. Whole genome sequencing of Bacillus velezensis JT3-1 will lay a good foundation for elucidation of the mechanisms of its antimicrobial activity, and for its future application.

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April 21, 2020

Genome mining identifies cepacin as a plant-protective metabolite of the biopesticidal bacterium Burkholderia ambifaria.

Beneficial microorganisms are widely used in agriculture for control of plant pathogens, but a lack of efficacy and safety information has limited the exploitation of multiple promising biopesticides. We applied phylogeny-led genome mining, metabolite analyses and biological control assays to define the efficacy of Burkholderia ambifaria, a naturally beneficial bacterium with proven biocontrol properties but potential pathogenic risk. A panel of 64 B.?ambifaria strains demonstrated significant antimicrobial activity against priority plant pathogens. Genome sequencing, specialized metabolite biosynthetic gene cluster mining and metabolite analysis revealed an armoury of known and unknown pathways within B.?ambifaria. The biosynthetic gene cluster responsible for the production of the metabolite cepacin was identified and directly shown to mediate protection of germinating crops against Pythium damping-off disease. B.?ambifaria maintained biopesticidal protection and overall fitness in the soil after deletion of its third replicon, a non-essential plasmid associated with virulence in Burkholderia?cepacia complex bacteria. Removal of the third replicon reduced B.?ambifaria persistence in a murine respiratory infection model. Here, we show that by using interdisciplinary phylogenomic, metabolomic and functional approaches, the mode of action of natural biological control agents related to pathogens can be systematically established to facilitate their future exploitation.

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April 21, 2020

Complete Genome Sequencing of Bacillus velezensis WRN014, and Comparison with Genome Sequences of other Bacillus velezensis Strains.

Bacillus velezensis strain WRN014 was isolated from banana fields in Hainan, China. Bacillus velezensis is an important member of the plant growth-promoting rhizobacteria (PGPR) which can enhance plant growth and control soil-borne disease. The complete genome of Bacillus velezensis WRN014 was sequenced by combining Illumina Hiseq 2500 system and Pacific Biosciences SMRT high-throughput sequencing technologies. Then, the genome of Bacillus velezensis WRN014, together with 45 other completed genome sequences of the Bacillus velezensis strains, were comparatively studied. The genome of Bacillus velezensis WRN014 was 4,063,541bp in length and contained 4,062 coding sequences, 9 genomic islands and 13 gene clusters. The results of comparative genomic analysis provide evidence that (i) The 46 Bacillus velezensis strains formed 2 obviously closely related clades in phylogenetic trees. (ii) The pangenome in this study is open and is increasing with the addition of new sequenced genomes. (iii) Analysis of single nucleotide polymorphisms (SNPs) revealed local diversification of the 46 Bacillus velezensis genomes. Surprisingly, SNPs were not evenly distributed throughout the whole genome. (iv) Analysis of gene clusters revealed that rich gene clusters spread over Bacillus velezensis strains and some gene clusters are conserved in different strains. This study reveals that the strain WRN014 and other Bacillus velezensis strains have potential to be used as PGPR and biopesticide.

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April 21, 2020

Genomic and Functional Characterization of the Endophytic Bacillus subtilis 7PJ-16 Strain, a Potential Biocontrol Agent of Mulberry Fruit Sclerotiniose.

Bacillus sp. 7PJ-16, an endophytic bacterium isolated from a healthy mulberry stem and previously identified as Bacillus tequilensis 7PJ-16, exhibits strong antifungal activity and has the capacity to promote plant growth. This strain was studied for its effectiveness as a biocontrol agent to reduce mulberry fruit sclerotiniose in the field and as a growth-promoting agent for mulberry in the greenhouse. In field studies, the cell suspension and supernatant of strain 7PJ-16 exhibited biocontrol efficacy and the lowest disease incidence was reduced down to only 0.80%. In greenhouse experiments, the cell suspension (1.0?×?106 and 1.0?×?105 CFU/mL) and the cell-free supernatant (100-fold and 1000-fold dilution) stimulated mulberry seed germination and promoted mulberry seedling growth. In addition, to accurately identify the 7PJ-16 strain and further explore the mechanisms of its antifungal and growth-promoting properties, the complete genome of this strain was sequenced and annotated. The 7PJ-16 genome is comprised of two circular plasmids and a 4,209,045-bp circular chromosome, containing 4492 protein-coding genes and 116 RNA genes. This strain was ultimately designed as Bacillus subtilis based on core genome sequence analyses using a phylogenomic approach. In this genome, we identified a series of gene clusters that function in the synthesis of non-ribosomal peptides (surfactin, fengycin, bacillibactin, and bacilysin) as well as the ribosome-dependent synthesis of tasA and bacteriocins (subtilin, subtilosin A), which are responsible for the biosynthesis of numerous antimicrobial metabolites. Additionally, several genes with function that promote plant growth, such as indole-3-acetic acid biosynthesis, the production of volatile substances, and siderophores synthesis, were also identified. The information described in this study has established a good foundation for understanding the beneficial interactions between endophytes and host plants, and facilitates the further application of B. subtilis 7PJ-16 as an agricultural biofertilizer and biocontrol agent.

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April 21, 2020

The role of long-term mineral and organic fertilisation treatment in changing pathogen and symbiont community composition in soil

Application of organic fertilisers to soil prevents erosion, improves fertility and may suppress certain soil-borne plant pathogens, but it is still unclear how different trophic groups of fungi and oomycetes respond to long-term fertilisation treatment. The objective of the study was to examine the effect of different fertilisation regimes on fungal and oomycete pathogen- and mycorrhizal symbiont diversity and community structure in both soil and roots, using PacBio SMRT sequencing. The field experiment included three fertilisation treatments that have been applied since 1989: nitrogen fertilisation (WOM), nitrogen fertilisation with manure amendment (FYM) and alternative organic fertilisation (AOF), each applied at five different rates. Soil samples were collected three times during the growing season, while root samples were collected during the flowering stage. There was no influence of the studied variables on soil and root pathogen richness. Contrary to our hypothesis, pathogen relative abundance in both soil and roots was significantly higher in plots with the AOF treatment. Furthermore, richness and relative abundance of arbuscular mycorrhizal (AM) fungi decreased significantly in the AOF treatment. Permutational analysis of variance (PERMANOVA) demonstrated the effect of fertilisation treatment on pathogen community composition in both soil and roots. Our findings indicate that organic fertilisers may not always benefit soil microbial community composition. Therefore, further studies are needed to understand how fertilisation affects mycorrhizal mutualists and pathogens.

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April 21, 2020

Antimicrobial, plant growth-promoting and genomic properties of the peanut endophyte Bacillus velezensis LDO2.

Peanut suffer from a number of fungal and bacterial pathogens, while plant endophytes were considered excellent candidates as biocontrol agents. In this study, the peanut endophytic bacterium LDO2 was evaluated for the potential of peanut pathogens inhibition and growth-promotion, and the genetic mechanisms were explored by genome mining. Strain LDO2 significantly inhibited the growth of peanut pathogenic fungi and pathogenic bacteria, and specifically, it showed pronounced inhibition on mycelia growth of Aspergillus flavus mycelia and caused mycelial deformity. Gene clusters responsible for antifungal metabolites (fengycin, surfactin, bacilysin) and antibacterial metabolites (butirosin, bacillaene, difficidin, macrolactin, surfactin, bacilysin) were identified. Strain LDO2 also exhibited several growth-promoting related features including phosphate solubilization, siderophore production and growth promotion of peanut root. Genes associated with plant growth promotion were also identified and analyzed, as well as genes related to secreted proteins. These findings suggested that this peanut endophyte could be a potential biocontrol agent in peanut production and a source of antimicrobial compounds for further exploitation. Copyright © 2018 Elsevier GmbH. All rights reserved.

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April 21, 2020

Genome Sequence of Bacillus Velezensis W1, A Strain with Strong Acaricidal Activity against Two-Spotted Spider Mite (Tetranychus Urticae)

Bacillus velezensis W1, isolated from two-spotted spider mites that had died naturally, is a patented strain with strong capability to cause mortality of the phytophagous mite Tetranychus urticae. The whole genome of W1 was completely sequenced with a combination of an Illumina Miseq platform (400-bp paired-end) with 2 × 250 bases and a Pacific Biosciences (PaBio) RS II Single Molecule Real Time (SMRT) sequencing platform using a 20 kb SMRTbellTM template library. Here, we report the complete genome sequence of B. velezensis W1, including one circular chromosome of 4,237,431 bp encoding 4,352 genes with GC content of 45.84%, providing insights into the genomic basis of its acaricidal activity and facilitating its application in red spider mite biocontrol.

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April 21, 2020

Depiction of secondary metabolites and antifungal activity of Bacillus velezensis DTU001.

For a safe and sustainable environment, effective microbes as biocontrol agents are in high demand. We have isolated a new Bacillus velezensis strain DTU001, investigated its antifungal spectrum, sequenced its genome, and uncovered the production of lipopeptides in HPLC-HRMS analysis. To test the antifungal efficacy, extracts of B. velezensis DTU001 was tested against a range of twenty human or plant pathogenic fungi. We demonstrate that inhibitory potential of B. velezensis DTU001 against selected fungi is superior in comparison to single lipopeptide, either iturin or fengycin. The isolate showed analogous biofilm formation to other closely related Bacilli. To further support the biocontrol properties of the isolate, coculture with Candida albicans demonstrated that B. velezensis DTU001 exhibited excellent antiproliferation effect against C. albicans. In summary, the described isolate is a potential antifungal agent with a broad antifungal spectrum that might assist our aims to avoid hazardous pathogenic fungi and provide alternative to toxicity caused by chemicals.

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April 21, 2020

Genome-wide mutational biases fuel transcriptional diversity in the Mycobacterium tuberculosis complex.

The Mycobacterium tuberculosis complex (MTBC) members display different host-specificities and virulence phenotypes. Here, we have performed a comprehensive RNAseq and methylome analysis of the main clades of the MTBC and discovered unique transcriptional profiles. The majority of genes differentially expressed between the clades encode proteins involved in host interaction and metabolic functions. A significant fraction of changes in gene expression can be explained by positive selection on single mutations that either create or disrupt transcriptional start sites (TSS). Furthermore, we show that clinical strains have different methyltransferases inactivated and thus different methylation patterns. Under the tested conditions, differential methylation has a minor direct role on transcriptomic differences between strains. However, disruption of a methyltransferase in one clinical strain revealed important expression differences suggesting indirect mechanisms of expression regulation. Our study demonstrates that variation in transcriptional profiles are mainly due to TSS mutations and have likely evolved due to differences in host characteristics.

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April 21, 2020

Complete genome sequence of a marine-sediment-derived bacterial strain Bacillus velezensis SH-B74, a cyclic lipopeptides producer and a biopesticide.

A marine-sediment sample-derived strain Bacillus velezensis SH-B74 has the capacity to produce cyclic lipopeptides (CLPs), and these CLPs secreted by the strain show biological activities against various pests under both in vitro and in planta conditions, such evidence has supported that the strain SH-B74 is a biopesticide. To get a better insight into the mechanisms on the control of the pesticides by the strain, a genome sequencing project has been applied to the genomic DNA of the strain SH-B74. The results show that the strain SH-B74 has a chromosome size of 4,042,190 bp, with a GC content of 46.5%, in addition, the strain contains a 61,634 bp plasmid pSH-B74, with a GC content of 40.8%. Data from bioinformatic analysis reveal that the strain SH-B74 has genes with the capacity to increase environmental adaptation, promote the rhizosphere fitnesses and secrete a spectrum of antibiotics, including nonribosomal peptide synthetases (NRPSs)-derived CLPs bacillopeptin, plipastatin, and surfactin. The presence of CLPs in the bacterial cultures of the strain SH-B74 was confirmed further by LC-MS analysis. Thus, genome sequencing and analyses together with chemical analysis reveal the promising perspectives of the strain SH-B74 that are of spectacular importance to its trait as a plant beneficial microbe to be used in agriculture practices.

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