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Journal of Bacteriology, June 2005, p. 3739-3751, Vol. 187, No. 11
0021-9193/05/$08.00+0     doi:10.1128/JB.187.11.3739-3751.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Novel Molecular Features of the Fibrolytic Intestinal Bacterium Fibrobacter intestinalis Not Shared with Fibrobacter succinogenes as Determined by Suppressive Subtractive Hybridization{dagger}

Meng Qi,1 Karen E. Nelson,2 Sean C. Daugherty,2 William C. Nelson,2 Ioana R. Hance,2 Mark Morrison,3 and Cecil W. Forsberg1*

Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada,1 The Institute for Genomic Research, Rockville, Maryland,2 Department of Animal Science, Ohio State University, Columbus, Ohio3

Received 18 February 2005/ Accepted 2 March 2005

Suppressive subtractive hybridization was conducted to identify unique genes coding for plant cell wall hydrolytic enzymes and other properties of the gastrointestinal bacterium Fibrobacter intestinalis DR7 not shared by Fibrobacter succinogenes S85. Subtractive clones from F. intestinalis were sequenced and assembled to form 712 nonredundant contigs with an average length of 525 bp. Of these, 55 sequences were unique to F. intestinalis. The remaining contigs contained 764 genes with BLASTX similarities to other proteins; of these, 80% had the highest similarities to proteins in F. succinogenes, including 30 that coded for carbohydrate active enzymes. The expression of 17 of these genes was verified by Northern dot blot analysis. Of genes not exhibiting BLASTX similarity to F. succinogenes, 30 encoded putative transposases, 6 encoded restriction modification genes, and 45% had highest similarities to proteins in other species of gastrointestinal bacteria, a finding suggestive of either horizontal gene transfer to F. intestinalis or gene loss from F. succinogenes. Analysis of contigs containing segments of two or more adjacent genes revealed that only 35% exhibited BLASTX similarity and were in the same orientation as those of F. succinogenes, indicating extensive chromosomal rearrangement. The expression of eight transposases, and three restriction-modification genes was confirmed by Northern dot blot analysis. These data clearly document the maintenance of carbohydrate active enzymes in F. intestinalis necessitated by the preponderance of polysaccharide substrates available in the ruminal environment. It also documents substantive changes in the genome from that of F. succinogenes, which may be related to the introduction of the array of transposase and restriction-modification genes.

* Corresponding author. Mailing address: Department of Molecular and Cellular Biology, University of Guelph, Guelph, N1G 2W1 Ontario, Canada. Phone: (519) 824-4120, ext. 53433. Fax: (519) 837-1802. E-mail: cforsber{at}uoguelph.ca.

{dagger} Supplemental material for this article may be found at http://jb.asm.org/.

Journal of Bacteriology, June 2005, p. 3739-3751, Vol. 187, No. 11
0021-9193/05/$08.00+0     doi:10.1128/JB.187.11.3739-3751.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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