TY - JOUR
T1 - Disruption of Mycobacterium avium subsp. paratuberculosis-specific genes impairs in vivo fitness
AU - Wang, Joyce
AU - Pritchard, Justin R.
AU - Kreitmann, Louis
AU - Montpetit, Alexandre
AU - Behr, Marcel A.
N1 - Funding Information:
We thank Dr. Eric Rubin (Harvard University, Cambridge, MA) for MycomarT7 phagemid, Génome Québec for optimizing Illumina sequencing conditions and Dr. James Wynne (CSIRO Australian Animal Health Laboratory, Geelong, Australia) for providing resequenced MAP K-10 files to facilitate analysis. We also thank members of the Behr, Reed and Schurr labs for input and suggestions. This work was supported by an operating grant MOP-97813 from the Canadian Institutes of Health Research (CIHR) to MB. MB is Chercheur National of the FRSQ and a William Dawson Scholar of McGill University.
PY - 2014/5/31
Y1 - 2014/5/31
N2 - Background: Mycobacterium avium subsp. paratuberculosis (MAP) is an obligate intracellular pathogen that infects many ruminant species. The acquisition of foreign genes via horizontal gene transfer has been postulated to contribute to its pathogenesis, as these genetic elements are absent from its putative ancestor, M. avium subsp. hominissuis (MAH), an environmental organism with lesser pathogenicity. In this study, high-throughput sequencing of MAP transposon libraries were analyzed to qualitatively and quantitatively determine the contribution of individual genes to bacterial survival during infection. Results: Out of 52384 TA dinucleotides present in the MAP K-10 genome, 12607 had a MycoMarT7 transposon in the input pool, interrupting 2443 of the 4350 genes in the MAP genome (56%). Of 96 genes situated in MAP-specific genomic islands, 82 were disrupted in the input pool, indicating that MAP-specific genomic regions are dispensable for in vitro growth (odds ratio = 0.21). Following 5 independent in vivo infections with this pool of mutants, the correlation between output pools was high for 4 of 5 (R = 0.49 to 0.61) enabling us to define genes whose disruption reproducibly reduced bacterial fitness in vivo. At three different thresholds for reduced fitness in vivo, MAP-specific genes were over-represented in the list of predicted essential genes. We also identified additional genes that were severely depleted after infection, and several of them have orthologues that are essential genes in M. tuberculosis. Conclusions: This work indicates that the genetic elements required for the in vivo survival of MAP represent a combination of conserved mycobacterial virulence genes and MAP-specific genes acquired via horizontal gene transfer. In addition, the in vitro and in vivo essential genes identified in this study may be further characterized to offer a better understanding of MAP pathogenesis, and potentially contribute to the discovery of novel therapeutic and vaccine targets.
AB - Background: Mycobacterium avium subsp. paratuberculosis (MAP) is an obligate intracellular pathogen that infects many ruminant species. The acquisition of foreign genes via horizontal gene transfer has been postulated to contribute to its pathogenesis, as these genetic elements are absent from its putative ancestor, M. avium subsp. hominissuis (MAH), an environmental organism with lesser pathogenicity. In this study, high-throughput sequencing of MAP transposon libraries were analyzed to qualitatively and quantitatively determine the contribution of individual genes to bacterial survival during infection. Results: Out of 52384 TA dinucleotides present in the MAP K-10 genome, 12607 had a MycoMarT7 transposon in the input pool, interrupting 2443 of the 4350 genes in the MAP genome (56%). Of 96 genes situated in MAP-specific genomic islands, 82 were disrupted in the input pool, indicating that MAP-specific genomic regions are dispensable for in vitro growth (odds ratio = 0.21). Following 5 independent in vivo infections with this pool of mutants, the correlation between output pools was high for 4 of 5 (R = 0.49 to 0.61) enabling us to define genes whose disruption reproducibly reduced bacterial fitness in vivo. At three different thresholds for reduced fitness in vivo, MAP-specific genes were over-represented in the list of predicted essential genes. We also identified additional genes that were severely depleted after infection, and several of them have orthologues that are essential genes in M. tuberculosis. Conclusions: This work indicates that the genetic elements required for the in vivo survival of MAP represent a combination of conserved mycobacterial virulence genes and MAP-specific genes acquired via horizontal gene transfer. In addition, the in vitro and in vivo essential genes identified in this study may be further characterized to offer a better understanding of MAP pathogenesis, and potentially contribute to the discovery of novel therapeutic and vaccine targets.
UR - http://www.scopus.com/inward/record.url?scp=84902251612&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84902251612&partnerID=8YFLogxK
U2 - 10.1186/1471-2164-15-415
DO - 10.1186/1471-2164-15-415
M3 - Article
C2 - 24885784
AN - SCOPUS:84902251612
SN - 1471-2164
VL - 15
JO - BMC genomics
JF - BMC genomics
IS - 1
M1 - 415
ER -