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Genomic homogeneity between Mycobacterium avium subsp. avium and Mycobacterium avium subsp. paratuberculosis belies their divergent growth rates.

Bannantine JP, Zhang Q, Li LL, Kapur V - BMC Microbiol. (2003)

Bottom Line: These strategies resulted in the successful amplification and sequencing of a contiguous 11-kb fragment containing the putative Mycobacterium paratuberculosis origin of replication (oriC).Finally, the 16s rRNA gene from these two subspecies is 100% identical.In addition, genomic comparisons between these two mycobacterial subspecies suggest that differences in the oriC region may not be significant enough to account for the diverse bacterial replication rates.

View Article: PubMed Central - HTML - PubMed

Affiliation: National Animal Disease Center, USDA-ARS, 2300 N, Dayton Ave, Ames, IA 50010, USA. jbannant@nadc.ars.usda.gov

ABSTRACT

Background: Mycobacterium avium subspecies avium (M. avium) is frequently encountered in the environment, but also causes infections in animals and immunocompromised patients. In contrast, Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) is a slow-growing organism that is the causative agent of Johne's disease in cattle and chronic granulomatous infections in a variety of other ruminant hosts. Yet we show that despite their divergent phenotypes and the diseases they present, the genomes of M. avium and M. paratuberculosis share greater than 97% nucleotide identity over large (25 kb) genomic regions analyzed in this study.

Results: To characterize genome similarity between these two subspecies as well as attempt to understand their different growth rates, we designed oligonucleotide primers from M. avium sequence to amplify 15 minimally overlapping fragments of M. paratuberculosis genomic DNA encompassing the chromosomal origin of replication. These strategies resulted in the successful amplification and sequencing of a contiguous 11-kb fragment containing the putative Mycobacterium paratuberculosis origin of replication (oriC). This fragment contained 11 predicted open reading frames that showed a conserved gene order in the oriC locus when compared with several other Gram-positive bacteria. In addition, a GC skew analysis identified the origin of chromosomal replication which lies between the genes dnaA and dnaN. The presence of multiple DnaA boxes and the ATP-binding site in dnaA were also found in M. paratuberculosis. The strong nucleotide identity of M. avium and M. paratuberculosis in the region surrounding the origin of chromosomal replication led us to compare other areas of these genomes. A DNA homology matrix of 2 million nucleotides from each genome revealed strong synteny with only a few sequences present in one genome but absent in the other. Finally, the 16s rRNA gene from these two subspecies is 100% identical.

Conclusions: We present for the first time, a description of the oriC region in M. paratuberculosis. In addition, genomic comparisons between these two mycobacterial subspecies suggest that differences in the oriC region may not be significant enough to account for the diverse bacterial replication rates. Finally, the few genetic differences present outside the origin of chromosomal replication in each genome may be responsible for the diverse growth rates or phenotypes observed between the avium and paratuberculosis subspecies.

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DNA matrix analysis of a contiguous 2 million nucleotide section of the M. avium (y-axis) and M. paratuberculosis (x-axis) genomes. Four 500,000 nucleotide matrices are shown with the nucleotide segments indicated above each plot. A long unbroken diagonal line from the upper left corner to the lower right corner indicates that the sequences are collinear. The diagonal line (in blue) that runs from the lower left to the upper right at the 350,000 nucleotide region indicates that one sequence is the reverse complement of the other. The arrows (in red) show sequences present in M. avium but absent in M. paratuberculosis and the arrowhead (in green) shows a sequence represented only in M. paratuberculosis. The initial nucleotide in the dnaA coding sequence was defined as number one in both genomes for this analysis. The parameters for this DNA identity matrix include: a window size of 30, a minimum percent score of 80, and a hash value of 4.
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Figure 4: DNA matrix analysis of a contiguous 2 million nucleotide section of the M. avium (y-axis) and M. paratuberculosis (x-axis) genomes. Four 500,000 nucleotide matrices are shown with the nucleotide segments indicated above each plot. A long unbroken diagonal line from the upper left corner to the lower right corner indicates that the sequences are collinear. The diagonal line (in blue) that runs from the lower left to the upper right at the 350,000 nucleotide region indicates that one sequence is the reverse complement of the other. The arrows (in red) show sequences present in M. avium but absent in M. paratuberculosis and the arrowhead (in green) shows a sequence represented only in M. paratuberculosis. The initial nucleotide in the dnaA coding sequence was defined as number one in both genomes for this analysis. The parameters for this DNA identity matrix include: a window size of 30, a minimum percent score of 80, and a hash value of 4.

Mentions: We next determined if the nucleotide identities would remain consistently high when M. paratuberculosis sequences outside the oriC region were compared with M. avium. Sequencing of the M. paratuberculosis K-10 cattle isolate is nearing completion in our laboratories and TIGR is in the finishing stages of M. avium isolate 104. Beginning with nucleotide number 1 in the dnaA coding region of each genome, a comparison of 2 million bases of M. paratuberculosis with 2 million bases from M. avium by Pustell DNA matrix analysis [25], indicates that genomic similarity continues outside the surrounding oriC region (Fig. 4). When evaluating similarities between two sequences of this size, a matrix comparison is the method of first choice. In addition, the matrix method displays matching regions in the context of the sequence as a whole, making it easy to determine if the regions are repeated or inverted. For example, figure 4 shows a large 56.6 kb genomic inversion of the region surrounding nucleotide 350,000. The DNA identity matrix also identified sequences that were present in one genome, but absent in the other as shown by the broken diagonal lines (Fig. 4). These data show remarkable similarity over large regions in both mycobacterial genomes.


Genomic homogeneity between Mycobacterium avium subsp. avium and Mycobacterium avium subsp. paratuberculosis belies their divergent growth rates.

Bannantine JP, Zhang Q, Li LL, Kapur V - BMC Microbiol. (2003)

DNA matrix analysis of a contiguous 2 million nucleotide section of the M. avium (y-axis) and M. paratuberculosis (x-axis) genomes. Four 500,000 nucleotide matrices are shown with the nucleotide segments indicated above each plot. A long unbroken diagonal line from the upper left corner to the lower right corner indicates that the sequences are collinear. The diagonal line (in blue) that runs from the lower left to the upper right at the 350,000 nucleotide region indicates that one sequence is the reverse complement of the other. The arrows (in red) show sequences present in M. avium but absent in M. paratuberculosis and the arrowhead (in green) shows a sequence represented only in M. paratuberculosis. The initial nucleotide in the dnaA coding sequence was defined as number one in both genomes for this analysis. The parameters for this DNA identity matrix include: a window size of 30, a minimum percent score of 80, and a hash value of 4.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC156888&req=5

Figure 4: DNA matrix analysis of a contiguous 2 million nucleotide section of the M. avium (y-axis) and M. paratuberculosis (x-axis) genomes. Four 500,000 nucleotide matrices are shown with the nucleotide segments indicated above each plot. A long unbroken diagonal line from the upper left corner to the lower right corner indicates that the sequences are collinear. The diagonal line (in blue) that runs from the lower left to the upper right at the 350,000 nucleotide region indicates that one sequence is the reverse complement of the other. The arrows (in red) show sequences present in M. avium but absent in M. paratuberculosis and the arrowhead (in green) shows a sequence represented only in M. paratuberculosis. The initial nucleotide in the dnaA coding sequence was defined as number one in both genomes for this analysis. The parameters for this DNA identity matrix include: a window size of 30, a minimum percent score of 80, and a hash value of 4.
Mentions: We next determined if the nucleotide identities would remain consistently high when M. paratuberculosis sequences outside the oriC region were compared with M. avium. Sequencing of the M. paratuberculosis K-10 cattle isolate is nearing completion in our laboratories and TIGR is in the finishing stages of M. avium isolate 104. Beginning with nucleotide number 1 in the dnaA coding region of each genome, a comparison of 2 million bases of M. paratuberculosis with 2 million bases from M. avium by Pustell DNA matrix analysis [25], indicates that genomic similarity continues outside the surrounding oriC region (Fig. 4). When evaluating similarities between two sequences of this size, a matrix comparison is the method of first choice. In addition, the matrix method displays matching regions in the context of the sequence as a whole, making it easy to determine if the regions are repeated or inverted. For example, figure 4 shows a large 56.6 kb genomic inversion of the region surrounding nucleotide 350,000. The DNA identity matrix also identified sequences that were present in one genome, but absent in the other as shown by the broken diagonal lines (Fig. 4). These data show remarkable similarity over large regions in both mycobacterial genomes.

Bottom Line: These strategies resulted in the successful amplification and sequencing of a contiguous 11-kb fragment containing the putative Mycobacterium paratuberculosis origin of replication (oriC).Finally, the 16s rRNA gene from these two subspecies is 100% identical.In addition, genomic comparisons between these two mycobacterial subspecies suggest that differences in the oriC region may not be significant enough to account for the diverse bacterial replication rates.

View Article: PubMed Central - HTML - PubMed

Affiliation: National Animal Disease Center, USDA-ARS, 2300 N, Dayton Ave, Ames, IA 50010, USA. jbannant@nadc.ars.usda.gov

ABSTRACT

Background: Mycobacterium avium subspecies avium (M. avium) is frequently encountered in the environment, but also causes infections in animals and immunocompromised patients. In contrast, Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) is a slow-growing organism that is the causative agent of Johne's disease in cattle and chronic granulomatous infections in a variety of other ruminant hosts. Yet we show that despite their divergent phenotypes and the diseases they present, the genomes of M. avium and M. paratuberculosis share greater than 97% nucleotide identity over large (25 kb) genomic regions analyzed in this study.

Results: To characterize genome similarity between these two subspecies as well as attempt to understand their different growth rates, we designed oligonucleotide primers from M. avium sequence to amplify 15 minimally overlapping fragments of M. paratuberculosis genomic DNA encompassing the chromosomal origin of replication. These strategies resulted in the successful amplification and sequencing of a contiguous 11-kb fragment containing the putative Mycobacterium paratuberculosis origin of replication (oriC). This fragment contained 11 predicted open reading frames that showed a conserved gene order in the oriC locus when compared with several other Gram-positive bacteria. In addition, a GC skew analysis identified the origin of chromosomal replication which lies between the genes dnaA and dnaN. The presence of multiple DnaA boxes and the ATP-binding site in dnaA were also found in M. paratuberculosis. The strong nucleotide identity of M. avium and M. paratuberculosis in the region surrounding the origin of chromosomal replication led us to compare other areas of these genomes. A DNA homology matrix of 2 million nucleotides from each genome revealed strong synteny with only a few sequences present in one genome but absent in the other. Finally, the 16s rRNA gene from these two subspecies is 100% identical.

Conclusions: We present for the first time, a description of the oriC region in M. paratuberculosis. In addition, genomic comparisons between these two mycobacterial subspecies suggest that differences in the oriC region may not be significant enough to account for the diverse bacterial replication rates. Finally, the few genetic differences present outside the origin of chromosomal replication in each genome may be responsible for the diverse growth rates or phenotypes observed between the avium and paratuberculosis subspecies.

Show MeSH
Related in: MedlinePlus