- Results: Here we present the de novo sequencing of three strains of the genus Streptomyces isolated from disease- suppressive soils to produce high-quality complete genomes. - GS93 – 23, Streptomyces sp. - In addition, two of the strains were found to have large, linear plasmids. - Ninety percent of the 2000 major diseases of the 31 principle crops in the US are caused by soil-borne path- ogens [2, 3], and soil microbial communities can have a protective effect [4]. - 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0. - Full list of author information is available at the end of the article. - A better under- standing of the composition and ecology of DSSs will fa- cilitate engineering soil communities for crop protection.. - Isolation and phenotypic characterization of strains Each of the strains sequenced for this study were selected because (i) they were isolated from soils with. - S3–4 and 3211–3 were isolated from pathogen suppressive soils located in the Cedar Creek Ecosystem Science Reserve (CCESR), an NSF long-term ecological research site [24]. - We next sought to verify that the short- read corrected sequences were indeed a better representa- tion of the actual genome sequence, as the two sequencing platforms are known to generate different types of errors.. - To determine which sequence variant was correct for each SNP/indel, translated protein sequences at each of the 295 SNP/indel loci in the S3–4 genome were compared against the NCBI GenBank non-redundant database, with the assumption that a frameshift resulting from an indel will result in a worse top blast hit for a stretch of DNA.. - This analysis is only expected to reveal the correct sequence variant when (i) the indel is present within a coding DNA sequence (CDS), (ii) correct protein sequences for close homologs are present in GenBank, and (iii) the 300 base- pair window that is searched is sufficiently focused such that top BLAST hits align to the translated query in the region of the variant locus (i.e. - at the center of the query, not the edges). - General characteristics of the genome sequences. - Annotation of the genomes with the Prokka software tool [35] identified 7188 CDSs, 7 ribosomal RNA op- erons, and 66 tRNAs for GS93–23. - We conservatively assigned specific molecules to these BGCs only when the annotated gene clusters share 100% of the biosynthetic genes from previously character- ized BGCs by manual comparison (Additional file Informa- tion). - The 26 high-confidence BGCs identified in the GS93–23 genome include known pathways for RiPP cyclothiazomy- cin [40], the dienoyltetramic acid streptolydigin [41], and the lipoglycopeptide mannopeptimycin [42]. - confidence BGCs in the S3–4 genome include known pathways for 2-methylisoborneol, and the aminoglyco- side streptothricin [46]. - S3–4 groups. - GS93–23 clusters with the Streptomyces lydicus type strain NRRL-ISP 5461 [54].. - Insertions or deletion events greater than 100 bp account for only 4.5% of the genome sequence as a whole (Fig. - 3b), with a similar proportion being lost/gained in BGCs as in the rest of the genome (Fig. - Interestingly, the position of SNPs relative to CDSs shows a marked de-enrichment in (i) the approxi- mate position of the Shine-Dalgarno sequence in the 5′- UTR, and (ii) the 5 ′ end of the CDS (Fig. - Most of the ~ 33,000 SNPs in CDSs encode silent mutations.. - Of the missense mutations, the majority are conservative in terms of amino acid chemistry (Fig. - Of the genes unique to ISP-5461, only a single gene was of unknown function. - These changes alone account for 9 and 7% of the total genome content, respectively. - The S3–4 genome lacks a close homolog in the sequence databases. - Despite sharing 96.3% sequence identity of the rpoB gene, 26% of the S3 – 4 genome does not align with the WM6372 sequence.. - Our closest pair, GS93–23 and ISP-5461, share 26/26 of the high-confidence BGCs and 61/64 ‘putative’. - Select regions of the atpD, gyrB, recA, rpoB, and trpB genes were concatenated and used to generate a multi-locus alignment in the MEGA7 software package. - katrae (S3 – 4, blue), S. - lydicus (GS93 – 23, red). - similarity of the rpoB gene, have in common only 31/38 of the high-confidence BGC annotations, which is driven mostly by the presence of two plasmids in 3211–3 miss- ing from B-1447. - Between S3–4 and WM6372 (96.3%. - (c) SNP analysis of strain GS93 – 23 and its closest relative ISP-5461. - In the clockwise direction, the first node corresponds to a codon in GS93 – 23 and the second node to ISP-5461. - Each CDS SNP is represented by an arc connecting a codon in GS93 – 23 to a codon in ISP-5461, with the width of the arc indicating number of instances of that mutation. - Black line and grey boxes show average SNP abundance and 1-, 2- standard deviations as calculated for the last 90% of the CDS. - Among 125 complete Streptomyces genomes with antiSMASH 4.1 (Additional file 1: Table S4), the number of high-confidence BGCs in 3211–3 places it in the top 16% in terms of BGC content.. - The large size, repetitive na- ture, and high G + C content of Streptomyces genomes makes them difficult to fully assemble from short reads, and so roughly 90% of the available genomes are only available in draft status. - Area plots show clusterBLAST scores for 3211 – 3 (blue), GS93 – 23 (green), S3 – 4 (red). - Genetic distances between the three newly sequenced isolates and the genome hits from the signaling potential MultiGeneBLAST analysis genomes were obtained by multilocus alignment of the atpD, gyrB, recA, rpoB, and trpB genes. - lydicus ISP-5461, 26 of the 26 high- confidence BGCs found in GS93–23 were also predicted using the short-read only assembly contigs.. - The genomic comparison of GS93–23 and ISP-5461 suggests that these strains are part of the same clonal com- plex, despite being isolated 850 km apart and several decades removed. - Our analysis of the SNP accumulation in relationship to relative location within genes shows a de- enrichment of sequence variation in regions known to control translation initiation rates. - This makes sense in light of the experimental data and ecological models that suggest DSSs community members are selected for their antagonistic phenotypes [7]. - If this is true, it will suggest that evolution of DSS isolates occurs on the level of the genome/strain, not the individual genes, con- trary to what has been observed in other environments [81].. - Strain recruitment is a proposed mechanism of the establish- ment of disease suppressive soils [82], in which plants sup- port the maintenance of those microbial strains which inhibit phytopathogens. - 16S sequencing and denaturing gel electrophoresis of the rhizosphere microbiome of strawberry plants showed that the Actinobacteria community profile was more similar between species of strawberry plant, re- gardless of site, when compared to oil rape rhizosphere com- munities [83]. - For each of the three genomic DNA samples, sequencing was performed using P4 chemistry on two SMRT cells and using P6 chemistry on an add- itional SMRT cell from November 2014 to January 2015.. - For each CDS, start position, end position, strand information, and a unique identifier was provided in tabular format to ensure that Prokka-generated annota- tions would be used for clusters of orthologous genes (COG) assignment in place of the default Glimmer algo- rithm. - GS93 – 23, Table S2.. - The funding bodies played no role in the design of the study and collection, analysis, and interpretation of data and in writing the manuscript.. - 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