160. Microbial Genome Sequencing and Comparative Analysis 

D.R. Smith, M. Ayers, R. Bashirzadeh, H. Bochner, M. Boivin, G. Breton, S. Bross, A. Caron, A. Caruso, R. Cook, P. Daggett, L. Doucette-Stamm, J. Dubois, J. Egan, D. Ellston, J. Ezedi, T. Ho, K. Holtham, P. Joseph, M. LaPlante, H-M. Lee, R. Gibson, K. Gilbert, J. Guerin, D. Harrison, J. Hitti, P. Keagle, J. Kozlovsky, G. LeBlanc, W. Lumm, P. Mank, A. Majeski, J. Nölling, D. Patwell, J. Phillips, B. Pothier, S. Prabhakar, D. Qiu, J.N. Reeve¹, M. Rossetti, M. Sachdeva, P. Snell, ²P. Soucaille, L. Spitzer, R. Vicaire, K. Wall, Y. Wang, L. Wong, A. Wonsey, K. Weinstock, Q. Xu, and L. Zhang 
¹Dept.of Microbiology, The Ohio State University, Columbus, Ohio and ²INSA, Toulose, France; Genome Therapeutics Corp., Waltham, Massachusetts 
doug.smith@genomecorp.com 

This project is applying automated sequencing technology and bioinformatics tools to the analysis of microbial genomes with potential applications in energy production and bioremediation., Efforts have focused on two genomes in particular, those of Methanobacterium thermoautotrophicum strain H, and Clostridum acetobutylicum strain ATCC 824.  

Methanobacterium thermoautotrophicum strain H is a thermophilic archaeon that grows at temperatures from 40-70° C, and was isolated in 1971 from sewage sludge. The complete 1,751,377 bp sequence of the genome of M. thermoautotrophicum was determined by a whole genome shotgun sequencing approach. The results of extensive comparative and functional analysis work were published last year in the Journal of Bacteriology, Volume 179, 7135-7155.  

C. acetobutylicum strain ATCC 824 has a 4.2 Mb, AT-rich genome, and is one of the best-studied solventogenic clostridia (it has been used commercially to produce acetone, butanol and ethanol). The shotgun sequencing phase has been completed, with 4.9 Mb of multiplex and 21.3 Mb of ABI raw sequence reads (6.3 fold total redundancy) that produced 551 contigs spanning 4,030,725 bases when assembled using PHRAP with quality scores. The genome has been finished to 27 ordered contigs, with quality enhancement, at the time of this writing.  

Physical mapping of the C. acetobutylicum genome by P. Soucaille and coworkers (INSA, Toulose) has shown that this strain harbors a large plasmid, designated pSOL1, of about 210 kb in size. Further studies by the same group revealed that loss of this plasmid coincides with the loss of the capacity to produce acetone and butanol and that the genes involved in solvent formation reside on pSOL1. We now have the complete 203 kb sequence of this plasmid. 

C. acetobutylicum contains a variety of genes involved in the utilization of polysaccharides such as starch, hemicelluose and cellulose. The potential to degrade cellulose, indicated by the presence of an entire set of genes predicted to code for a cellulose-hydolysing mutlienzyme complex termed cellulosome, is surprising as cellulolytic activity is unknown for C. acetobutylicum. In addition, a gene similar to the toxin A encoding gene from the pathogenic clostridium C. difficile is present in the non-pathogenic C. acetobutylicum, coding for a polypeptide of ~ 2800 residues the majority of which is organized in ~125 repeats of 20 amino acids each. The genome of C. acetobutylicum ATCC 824 seems to be nearly void of mobile genetic elements. Only a single copy of a transposase gene, belonging to the Tn3 family and located on plasmid pSOL1, could be identified. Two gene clusters of four genes each show similarity to bacteriophage-like elements. There are 11 ribosomal operons. The data are available in GenBank and on our Web page (http://www.genomecorp.com).