7 research outputs found
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A Python script to construct FASTA files containing reconstructed genome sequences of evolved clones after DNA rearrangements
Scanning the Landscape of Genome Architecture of Non-O1 and Non-O139 <i>Vibrio cholerae</i> by Whole Genome Mapping Reveals Extensive Population Genetic Diversity
<div><p>Historically, cholera outbreaks have been linked to <i>V</i>. <i>cholerae</i> O1 serogroup strains or its derivatives of the O37 and O139 serogroups. A genomic study on the 2010 Haiti cholera outbreak strains highlighted the putative role of non O1/non-O139 <i>V</i>. <i>cholerae</i> in causing cholera and the lack of genomic sequences of such strains from around the world. Here we address these gaps by scanning a global collection of <i>V</i>. <i>cholerae</i> strains as a first step towards understanding the population genetic diversity and epidemic potential of non O1/non-O139 strains. Whole Genome Mapping (Optical Mapping) based bar coding produces a high resolution, ordered restriction map, depicting a complete view of the unique chromosomal architecture of an organism. To assess the genomic diversity of non-O1/non-O139 <i>V</i>. <i>cholerae</i>, we applied a Whole Genome Mapping strategy on a well-defined and geographically and temporally diverse strain collection, the Sakazaki serogroup type strains. Whole Genome Map data on 91 of the 206 serogroup type strains support the hypothesis that <i>V</i>. <i>cholerae</i> has an unprecedented genetic and genomic structural diversity. Interestingly, we discovered chromosomal fusions in two unusual strains that possess a single chromosome instead of the two chromosomes usually found in <i>V</i>. <i>cholerae</i>. We also found pervasive chromosomal rearrangements such as duplications and indels in many strains. The majority of <i>Vibrio</i> genome sequences currently in public databases are unfinished draft sequences. The Whole Genome Mapping approach presented here enables rapid screening of large strain collections to capture genomic complexities that would not have been otherwise revealed by unfinished draft genome sequencing and thus aids in assembling and finishing draft sequences of complex genomes. Furthermore, Whole Genome Mapping allows for prediction of novel <i>V</i>. <i>cholerae</i> non-O1/non-O139 strains that may have the potential to cause future cholera outbreaks.</p></div
Pulse field gel electrophoresis of chromosomal DNAs of <i>V</i>. <i>cholerae</i> 1154–74 (O49) and 10432–62 (O27) strains.
<p>PFGE of intact <i>V</i>. <i>cholerae</i> DNA isolated from different <i>V</i>. <i>cholerae</i> strains. Lanes from left to right: 1) Molecular weight marker (Mbases) <i>H</i>. <i>wingeii</i> chromosomes, 2) <i>V</i>. <i>cholerae</i> O1 N16961 (the bands corresponding to Chr I and Chr II are marked by an asterisk), 3) <i>V</i>. <i>cholerae</i> 10432–62 (O27) and 4) <i>V</i>. <i>cholerae</i> 1154–74 (O49). In lanes 3 and 4, the band corresponding to the single chromosome is marked by a triangle.</p
UPGMA method based dendrograms of Sakazaki serogroup strains.
<p>Whole Genome Mapping data based distance matrix with default parameters was used to generate the dendrograms. The three clusters (epidemic <i>V</i>. <i>cholerae</i> Classical and El Tor, one group of environmental isolates from rat, and <i>V</i>. <i>mimicus</i>) are indicated on the branches. The distribution of VPI and CTX on chromosome I in non-O1/non-O139 strains are indicated as well. The different colors of the highlighted strains indicate the following characteristics: No highlight- Clinical; Gray-environmental; Light green (serogroups O2-O4) source unknown; Yellow: epidemic O1 Classical and El Tor; Green: non-O1 epidemic strains; Pink: <i>V</i>. <i>mimicus</i>; Blue: One <i>V</i>. <i>mimicus</i> isolate (serogroup O115) that has the VPI cluster; Orange: Clinical and carry both VPI and CTX clusters; Light red (O77, O49, O80, O53): Clinical and carry VPI only. Scale: 0.2 = 20% dissimilarity.</p
Strain collection features of the Sakazaki serogroup set.
<p>A) Breakdown of strains based on geographical location (country where the strains were isolated). B) Breakdown of strains based on isolation source. The number of strains for which Whole Genome Maps were generated in this study is indicated in parentheses.</p
Whole Genome Mapping using different restriction enzymes supports the Chr I and Chr II fusions in <i>V</i>. <i>cholerae</i> 1154–74 (O49) and 10432–62 (O27) strains.
<p>A) The top four panels of maps are derived from strain 1154–74 (O49) and B) the bottom 3 panels are from strain 10432–61 (O27). In each panel, the top and bottom maps indicate <i>in silico</i> generated Chr II and Chr I restriction maps of M66–2 sequences respectively, compared to the experimentally generated maps (middle) of 1154–74 or 10432–62 using indicated restriction enzymes. The fusion in 1154–74 (O49) is around 1.29 Mb of Chr I (size 2.89 Mb) to 0.32 Mb of Chr II (size 1.05) of M66–2. The fusion in 10432–62 (O27) has occurred around 2.80 Mb of Chr I (2.89 Mb) to around 0.83 Mb of Chr II (1.05 Mb) in M66–2. The blue region indicates a single copy match between the chromosomes compared.</p
Whole Genome Maps using different restriction enzymes show putative tandem duplication of chromosomal regions.
<p>A: Top four panels of maps generated using <i>Bam</i>HI, <i>Nhe</i>I, <i>Kpn</i>I and <i>Afl</i>III respectively show the duplicated region in <i>V</i>. <i>cholerae</i> 1154–74 (O49). B: The bottom three panels of maps are that of 10432–62 (O27) using <i>Bam</i>HI, <i>Nhe</i>I and <i>Afl</i>III. In both cases, the location of duplication was found to be around 1240 kb to 1506 kb on reference M66–2 Chr I. The duplicated genome segments are indicated by the orange box in the <i>in silico</i> map of the reference strain. In some panels both assemblies with the single and two copies (resulting from re-analyses of the single molecule data from this region) are shown. The duplicated copies where they could be resolved are indicated by the red circles. The exact lengths of the duplications in the two strains cannot be unequivocally determined by this WGM data. It is also possible that the maps represent a mixed population of cells containing single and two copies of the duplication. (Scale bar 100 Kb).</p