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Analysis of genomic differences among Clostridium botulinum type A1 strains.

Fang PK, Raphael BH, Maslanka SE, Cai S, Singh BR - BMC Genomics (2010)

Bottom Line: The purpose of this study was to characterize differences among these genomes and compare these differentiating features with two additional unsequenced strains used in previous studies.Several strategies were deployed in this report.Taken together, C. botulinum type A1 strains including Sanger Institute ATCC 3502, ATCC 3502*, ATCC 19397, Hall, Allergan, and UMASS strains demonstrate differences at the level of the neurotoxin gene sequence, in gene content, and in genome arrangement.

View Article: PubMed Central - HTML - PubMed

Affiliation: Botulinum Research Center and Department of Chemistry & Biochemistry, University of Massachusetts Dartmouth, 285 Old Westport Road, North Dartmouth, Massachusetts 02747, USA.

ABSTRACT

Background: Type A1 Clostridium botulinum strains are a group of Gram-positive, spore-forming anaerobic bacteria that produce a genetically, biochemically, and biophysically indistinguishable 150 kD protein that causes botulism. The genomes of three type A1 C. botulinum strains have been sequenced and show a high degree of synteny. The purpose of this study was to characterize differences among these genomes and compare these differentiating features with two additional unsequenced strains used in previous studies.

Results: Several strategies were deployed in this report. First, University of Massachusetts Dartmouth laboratory Hall strain (UMASS strain) neurotoxin gene was amplified by PCR and sequenced; its sequence was aligned with the published ATCC 3502 Sanger Institute Hall strain and Allergan Hall strain neurotoxin gene regions. Sequence alignment showed that there was a synonymous single nucleotide polymorphism (SNP) in the region encoding the heavy chain between Allergan strain and ATCC 3502 and UMASS strains. Second, comparative genomic hybridization (CGH) demonstrated that the UMASS strain and a strain expected to be derived from ATCC 3502 in the Centers for Disease Control and Prevention (CDC) laboratory (ATCC 3502*) differed in gene content compared to the ATCC 3502 genome sequence published by the Sanger Institute. Third, alignment of the three sequenced C. botulinum type A1 strain genomes revealed the presence of four comparable blocks. Strains ATCC 3502 and ATCC 19397 share the same genome organization, while the organization of the blocks in strain Hall were switched. Lastly, PCR was designed to identify UMASS and ATCC 3502* strain genome organizations. The PCR results indicated that UMASS strain belonged to Hall type and ATCC 3502* strain was identical to ATCC 3502 (Sanger Institute) type.

Conclusions: Taken together, C. botulinum type A1 strains including Sanger Institute ATCC 3502, ATCC 3502*, ATCC 19397, Hall, Allergan, and UMASS strains demonstrate differences at the level of the neurotoxin gene sequence, in gene content, and in genome arrangement.

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Identification of the genome organizational patterns of ATCC 3502* and UMASS strains by PCR. Panel A (lanes 1 through 4), PCR products from ATCC 3502* strain genomic DNA amplifications. Panel B (lanes 5 through 8), PCR products from UMASS strain genomic DNA amplifications.
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Figure 5: Identification of the genome organizational patterns of ATCC 3502* and UMASS strains by PCR. Panel A (lanes 1 through 4), PCR products from ATCC 3502* strain genomic DNA amplifications. Panel B (lanes 5 through 8), PCR products from UMASS strain genomic DNA amplifications.

Mentions: To characterize whether the genome organizational pattern of ATCC 3502* and UMASS Hall strain fits into either of above two patterns, a PCR strategy was utilized. Primers were designed to span the boundary between block 3 and block 4 for the ATCC 3502 and ATCC 19397 pattern or between block 3 and block 2 for the Hall pattern (Figure 4). In one set of PCR reactions, using ATCC 3502* genomic DNA as template, the expected PCR product was generated from every PCR reaction with different combinations of upstream and downstream primers for ATCC 3502 and ATCC 19397 pattern (Figure 5 Panel A, lanes 1 to 4) but not from those with different combination of primers for Hall pattern reactions (data not shown). These results demonstrated that the genomic organization of ATCC 3502* indeed was identical to the ATCC 3502 Sanger Institute and ATCC 19397 strain genome organizations.


Analysis of genomic differences among Clostridium botulinum type A1 strains.

Fang PK, Raphael BH, Maslanka SE, Cai S, Singh BR - BMC Genomics (2010)

Identification of the genome organizational patterns of ATCC 3502* and UMASS strains by PCR. Panel A (lanes 1 through 4), PCR products from ATCC 3502* strain genomic DNA amplifications. Panel B (lanes 5 through 8), PCR products from UMASS strain genomic DNA amplifications.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3038992&req=5

Figure 5: Identification of the genome organizational patterns of ATCC 3502* and UMASS strains by PCR. Panel A (lanes 1 through 4), PCR products from ATCC 3502* strain genomic DNA amplifications. Panel B (lanes 5 through 8), PCR products from UMASS strain genomic DNA amplifications.
Mentions: To characterize whether the genome organizational pattern of ATCC 3502* and UMASS Hall strain fits into either of above two patterns, a PCR strategy was utilized. Primers were designed to span the boundary between block 3 and block 4 for the ATCC 3502 and ATCC 19397 pattern or between block 3 and block 2 for the Hall pattern (Figure 4). In one set of PCR reactions, using ATCC 3502* genomic DNA as template, the expected PCR product was generated from every PCR reaction with different combinations of upstream and downstream primers for ATCC 3502 and ATCC 19397 pattern (Figure 5 Panel A, lanes 1 to 4) but not from those with different combination of primers for Hall pattern reactions (data not shown). These results demonstrated that the genomic organization of ATCC 3502* indeed was identical to the ATCC 3502 Sanger Institute and ATCC 19397 strain genome organizations.

Bottom Line: The purpose of this study was to characterize differences among these genomes and compare these differentiating features with two additional unsequenced strains used in previous studies.Several strategies were deployed in this report.Taken together, C. botulinum type A1 strains including Sanger Institute ATCC 3502, ATCC 3502*, ATCC 19397, Hall, Allergan, and UMASS strains demonstrate differences at the level of the neurotoxin gene sequence, in gene content, and in genome arrangement.

View Article: PubMed Central - HTML - PubMed

Affiliation: Botulinum Research Center and Department of Chemistry & Biochemistry, University of Massachusetts Dartmouth, 285 Old Westport Road, North Dartmouth, Massachusetts 02747, USA.

ABSTRACT

Background: Type A1 Clostridium botulinum strains are a group of Gram-positive, spore-forming anaerobic bacteria that produce a genetically, biochemically, and biophysically indistinguishable 150 kD protein that causes botulism. The genomes of three type A1 C. botulinum strains have been sequenced and show a high degree of synteny. The purpose of this study was to characterize differences among these genomes and compare these differentiating features with two additional unsequenced strains used in previous studies.

Results: Several strategies were deployed in this report. First, University of Massachusetts Dartmouth laboratory Hall strain (UMASS strain) neurotoxin gene was amplified by PCR and sequenced; its sequence was aligned with the published ATCC 3502 Sanger Institute Hall strain and Allergan Hall strain neurotoxin gene regions. Sequence alignment showed that there was a synonymous single nucleotide polymorphism (SNP) in the region encoding the heavy chain between Allergan strain and ATCC 3502 and UMASS strains. Second, comparative genomic hybridization (CGH) demonstrated that the UMASS strain and a strain expected to be derived from ATCC 3502 in the Centers for Disease Control and Prevention (CDC) laboratory (ATCC 3502*) differed in gene content compared to the ATCC 3502 genome sequence published by the Sanger Institute. Third, alignment of the three sequenced C. botulinum type A1 strain genomes revealed the presence of four comparable blocks. Strains ATCC 3502 and ATCC 19397 share the same genome organization, while the organization of the blocks in strain Hall were switched. Lastly, PCR was designed to identify UMASS and ATCC 3502* strain genome organizations. The PCR results indicated that UMASS strain belonged to Hall type and ATCC 3502* strain was identical to ATCC 3502 (Sanger Institute) type.

Conclusions: Taken together, C. botulinum type A1 strains including Sanger Institute ATCC 3502, ATCC 3502*, ATCC 19397, Hall, Allergan, and UMASS strains demonstrate differences at the level of the neurotoxin gene sequence, in gene content, and in genome arrangement.

Show MeSH
Related in: MedlinePlus