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Insights into the Origin of Clostridium botulinum Strains: Evolution of Distinct Restriction Endonuclease Sites in rrs (16S rRNA gene)

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Abstract

Diversity analysis of Clostridium botulinum strains is complicated by high microheterogeneity caused by the presence of 9–22 copies of rrs (16S rRNA gene). The need is to mine genetic markers to identify very closely related strains. Multiple alignments of the nucleotide sequences of the 212 rrs of 13 C. botulinum strains revealed intra- and inter-genomic heterogeneity. Low intragenomic heterogeneity in rrs was evident in strains 230613, Alaska E43, Okra, Eklund 17B, Langeland, 657, Kyoto, BKT015925, and Loch Maree. The most heterogenous rrs sequences were those of C. botulinum strains ATCC 19397, Hall, H04402065, and ATCC 3502. In silico restriction map** of these rrs sequences was observable with 137 type II Restriction endonucleases (REs). Nucleotide changes (NC) at these RE sites resulted in appearance of distinct and additional sites, and loss in certain others. De novo appearances of RE sites due to NC were recorded at different positions in rrs gene. A nucleotide transition A>G in rrs of C. botulinum Loch Maree and 657 resulted in the generation of 4 and 10 distinct RE sites, respectively. Transitions A>G, G>A, and T>C led to the loss of RE sites. A perusal of the entire NC and in silico RE map** of rrs of C. botulinum strains provided insights into their evolution. Segregation of strains on the basis of RE digestion patterns of rrs was validated by the cladistic analysis involving six house kee** genes: dnaN, gyrB, metG, prfA, pyrG, and Rho.

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Acknowledgments

We are thankful to Director of CSIR-Institute of Genomics and Integrative Biology (IGIB), and CSIR project GENESIS (BSC0121) for providing the necessary funds, facilities and moral support.

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  1. Microbial Biotechnology and Genomics, CSIR-Institute of Genomics and Integrative Biology (IGIB), Delhi University Campus, Mall Road, Delhi, 110007, India

    Ashish Bhushan, Tanmoy Mukherjee, Jayadev Joshi, Pratap Shankar & Vipin Chandra Kalia

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  1. Ashish Bhushan
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  2. Tanmoy Mukherjee
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Correspondence to Ashish Bhushan.

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Bhushan, A., Mukherjee, T., Joshi, J. et al. Insights into the Origin of Clostridium botulinum Strains: Evolution of Distinct Restriction Endonuclease Sites in rrs (16S rRNA gene). Indian J Microbiol 55, 140–150 (2015). https://doi.org/10.1007/s12088-015-0514-z

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