Draft Genome Sequences for Clostridium thermocellum Wild-Type Strain YS and Derived Cellulose Adhesion-Defective Mutant Strain AD2

Abstract

Clostridium thermocellum wild-type strain YS is an anaerobic, thermophilic, cellulolytic bacterium capable of directly converting cellulosic substrates into ethanol. Strain YS and a derived cellulose adhesion-defective mutant strain, AD2, played pivotal roles in describing the original cellulosome concept. We present their draft genome sequences.

GENOME ANNOUNCEMENT

Clostridium thermocellum was characterized over 50 years ago (31), and its ability for efficient degradation and utilization of cellulose for ethanol production was recognized early on (1, 2, 21, 26, 33, 34, 36, 37, 39). It is an obligate anaerobic microorganism that has one of the highest growth rates on cellulose. The bacterium possesses productivity advantages associated with thermophilic growth and is capable of producing its own enzymes for lignocellulosic biomass breakdown (see reviews in references 4, 11, 28, and 29). Recently, important progress has been made in understanding C. thermocellum ethanol tolerance (10), a targeted deletion system has been developed (38), and four C. thermocellum genome sequences have been determined (16, 19) (GenBank accession no. CP000568.1).

C. thermocellum strain YS was purified from samples derived from hot springs at Yellowstone National Park in the United States, and it was characterized as a potent cellulolytic strain. Strain YS and a derived cellulose adhesion-defective mutant (AD2) played pivotal roles in describing the original cellulosome concept that recognized that C. thermocellum cellulases and associated polysaccharide-degrading enzymes are packaged in organized, high-molecular-weight, cellulolytic enzyme complexes (5, 23, 24). Strain YS was used in a number of subsequent studies (e.g., references 6, 8, 13–15, 17, 18, 25, 27, 32, and 35), and the cellulosome concept has served as a model for different clostridia and other related anaerobic bacteria, e.g., Clostridium cellulovorans, Clostridium cellulolyticum, Clostridium josui, Clostridium papyrosolvens, Acetivibrio cellulolyticus, Bacteroides cellulosolvens, and Ruminococcus flavefaciens (3, 7, 12, 22).

Draft genome data for strain YS were generated using a combination of 454 (30) and Illumina (9) HiSeq2000 technologies from 3-kb and 500-bp paired-end libraries, respectively. The 454 data consisted of 650,450 reads and generated 207,578,580 bp. After trimming and filtering of Illumina data (CLC Genomics Workbench version 4.9.1), there were 16,806,784,095 and 15,259,925,308 bp of sequence data for strains YS and AD2 from 174,965,956 and 159,631,515 reads, respectively, with an average length of 96 bp. Trimmed Illumina reads were assembled using the CLC Genomics Workbench. The consensus Illumina sequences for strain YS were processed further by generating 1.5-kb overlapping fake reads using the fb_dice.pl script, which is part of the FragBlast module (http://www.clarkfrancis.com/codes/fb_dice.pl). The Newbler application (version 2.6; 454 Life Sciences) was then used to assemble the YS Illumina consensus sequences and the 454 reads into 100 large (≥500-bp) contiguous DNA elements of approximately 3.46 Mb. The average YS contig size was 34,644 bp, the N50 contig size was 126,840 bp, and the largest contig was 330,620 bp. The genome had an overall estimated G+C content of ∼39%. Strain AD2 had 132 large contigs with an average size of 26,020 bp, the N50 contig size was 76,137 bp, and the largest contig was 269,895 bp. Draft genome sequences were annotated at Oak Ridge National Laboratory using an automated annotation pipeline, based on the Prodigal gene prediction algorithm (20). Sequence data for DNA contigs, coding and translation models, annotations, and metabolic reconstructions are available online (http://genome.ornl.gov/microbial/guest/YSORG_Nov2011_Hybrid and http://genome.ornl.gov/microbial/guest/AD2_NovRerun).

This study reveals the C. thermocellum YS and AD2 genome sequences for the first time. Access to these genome sequences, which are linked to important prior observations, will facilitate further studies with this genus and species.

Nucleotide sequence accession numbers.

The C. thermocellum YS and AD2 nucleotide sequences have been deposited in DDBJ/EMBL/GenBank under accession numbers AJGT00000000 and AJGS00000000, respectively, and the versions described in this paper are the first versions. The entire data set has been deposited in the National Center for Biotechnology Information (NCBI) Sequence-Read Archive (SRA) database under accession number SRA049437.