Abstract
A comparative genomics approach was utilised to compare the genomes of Mycobacterium avium subspecies paratuberculosis (MAP) isolated from early onset paediatric Crohn's disease (CD) patients as well as Johne's diseased animals. Draft genome sequences were produced for MAP isolates derived from four CD patients, one ulcerative colitis (UC) patient, and two non-inflammatory bowel disease (IBD) control individuals using Illumina sequencing, complemented by comparative genome hybridisation (CGH). MAP isolates derived from two bovine and one ovine host were also subjected to whole genome sequencing and CGH. All seven human derived MAP isolates were highly genetically similar and clustered together with one bovine type isolate following phylogenetic analysis. Three other sequenced isolates (including the reference bovine derived isolate K10) were genetically distinct. The human isolates contained two large tandem duplications, the organisations of which were confirmed by PCR. Designated vGI-17 and vGI-18 these duplications spanned 63 and 109 open reading frames, respectively. PCR screening of over 30 additional MAP isolates (3 human derived, 27 animal derived and one environmental isolate) confirmed that vGI-17 and vGI-18 are common across many isolates. Quantitative real-time PCR of vGI-17 demonstrated that the proportion of cells containing the vGI-17 duplication varied between 0.01 to 15% amongst isolates with human isolates containing a higher proportion of vGI-17 compared to most animal isolates. These findings suggest these duplications are transient genomic rearrangements. We hypothesise that the over-representation of vGI-17 in human derived MAP strains may enhance their ability to infect or persist within a human host by increasing genome redundancy and conferring crude regulation of protein expression across biologically important regions.
Highlights
Mycobacterium avium subspecies paratuberculosis (MAP), a Grampositive acid fast bacillus, is a member of the Mycobacterium avium complex and is the causative agent of Johne’s disease (JD), a chronic granulomatous enteritis affecting ruminants
The aim of this study was to investigate the genetic relationship between multiple human and animal derived MAP strains at a genome-wide level
We hypothesised that genetic differences between strains may reveal phylogenetic relationships that provide a better understanding of the processes involved with MAP zoonotic transmission
Summary
Mycobacterium avium subspecies paratuberculosis (MAP), a Grampositive acid fast bacillus, is a member of the Mycobacterium avium complex and is the causative agent of Johne’s disease (JD), a chronic granulomatous enteritis affecting ruminants. Kirkwood et al [3] more recently demonstrated that MAP could be identified by IS900 PCR significantly more often in mucosal biopsies and or peripheral blood mononuclear cells (PBMCs) from paediatric CD patients (47%) not yet receiving therapy, when compared to non-IBD controls (11%). Viable MAP could only be cultured from mucosal biopsies from four of ten CD patients and none of the non-IBD controls. MAP is an extremely persistent pathogen that can survive within the livestock environment (i.e., water, faeces and soil) for long periods [4,5]. While bacilli from these environmental sources may pose some risk to humans, the main source of transmission from animals to humans is more likely to be via contaminated milk. Similar recovery rates have been found elsewhere [7] which indicate a possible transmission route of live MAP from animals to humans is occurring through contaminated milk and possibly through animal derived foodstuff
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