The entire genome of Lactobacillus casei BL23, a strain with probiotic properties, has been sequenced. The genomes of BL23 and the industrially used probiotic strain Shirota YIT 9029 (Yakult) seem to be very similar.The genome of Lactobacillus casei BL23 was sequenced using a combination of shotgun sequencing (performed by Cogenics, Meylan, France) and 454 pyrosequencing (DNA sequencing was performed by Genoscope, Evry, France). Based on 19,300 reads of the shotgun library, draft assemblies were compiled with Consed, providing 90 contigs and 4.5-fold coverage. The 400,000 pyrosequencing reads were assembled into 132 contigs. The large number of contigs owes to numerous repeated sequences, most of which were already correctly positioned in the shotgun approach. A combined assembly of the two data sets therefore provided only 13 contigs with 34-fold coverage. Twelve gaps were closed by PCR using combinations of 26 primers corresponding to the contig extremities. The last gap was located at the insertion site of a prophage (lcbl23-2), and the pyrosequencing data allowed two different assemblies corresponding to the integrated and circularized phages. For each assembly, about 30 different reads had been obtained. Apparently, only a few cells contain a mobilized prophage, because only a single pyrosequencing read corresponded to the BL23 genome without this phage. Excised lcbl23-2 probably rapidly propagates, explaining why reads corresponding to the circularized prophage were more abundant.The BL23 genome is composed of a single circular chromosome (3,079,196 bp) with an overall GϩC content of 46.34%. Coding sequence (CDS) prediction and annotation were carried out with AGMIAL (2) and provided 3,044 CDSs covering 84% of the genome. BL23 also harbors 5 rRNA operons and 60 tRNAs. The origin of replication was identified based on homology to the L. casei neotype strain ATCC 334 (5, 7). Most genes of BL23 are present on the leading strand. The replication terminus is located almost diametrically opposite to the origin of replication and is accompanied by a sharp transition in the GC skew.Almost all genes present in BL23 and ATCC 334 exhibit nearly (99%) identical sequences, and the synteny is widely conserved in the two chromosomes, with only a few extended homologous regions placed at different locations. Interestingly, a published 15-kb sequence of the probiotic Shirota strain (10) differs from the region covered by the genes LCABL_22200 to LCABL_22350 of BL23 at only two positions. The BL23 CDSs are also nearly (98%) identical to their homologues in Lactobacillus paracasei. The BL23 genome is almost 0.2 Mb bigger than that of ATCC 334 (7), which nevertheless contains numerous DNA regions absent from BL23. The core genome common to both strains is about 2.38 Mb (77% of the BL23 genome). A significant fraction of the accessory genome owes to prophage insertions and insertion elements. Other regions present in only one strain, such as the 25-kb region extending from genes LCABL_28260 to LCABL_28470 in BL23, are often re...
Genome analysis of Lactobacillus casei BL23 revealed that, compared to L. casei ATCC 334, it carries a 12.8-kb DNA insertion containing genes involved in the catabolism of the cyclic polyol myo-inositol (MI). Indeed, L. casei ATCC 334 does not ferment MI, whereas strain BL23 is able to utilize this carbon source. The inserted DNA consists of an iolR gene encoding a DeoR family transcriptional repressor and a divergently transcribed iolTABCDG1G2EJK operon, encoding a complete MI catabolic pathway, in which the iolK gene probably codes for a malonate semialdehyde decarboxylase. The presence of iolK suggests that L. casei has two alternative pathways for the metabolism of malonic semialdehyde: (i) the classical MI catabolic pathway in which IolA (malonate semialdehyde dehydrogenase) catalyzes the formation of acetyl-coenzyme A from malonic semialdehyde and (ii) the conversion of malonic semialdehyde to acetaldehyde catalyzed by the product of iolK. The function of the iol genes was verified by the disruption of iolA, iolT, and iolD, which provided MI-negative strains. By contrast, the disruption of iolK resulted in a strain with no obvious defect in MI utilization. Transcriptional analyses conducted with different mutant strains showed that the iolTABCDG1G2EJK cluster is regulated by substrate-specific induction mediated by the inactivation of the transcriptional repressor IolR and by carbon catabolite repression mediated by the catabolite control protein A (CcpA). This is the first example of an operon for MI utilization in lactic acid bacteria and illustrates the versatility of carbohydrate utilization in L. casei BL23.myo-Inositol (MI) is the most abundant stereoisomer of inositol (1,2,3,4,5,6-cyclohexanehexol). It is common in soils, and it is a constituent of phytic acid (inositol hexaphosphate), which in the form of various salts (phytates) provides a major phosphate storage molecule in plant seeds. Several microorganisms, mostly inhabitants of soil, can utilize MI as a carbon source. For bacteria, the MI catabolic pathway has been elucidated for Enterobacter aerogenes (formerly Aerobacter aerogenes) (3). MI is important for the establishment of symbiosis in legume nodules. The metabolism of MI in Sinorhizobium fredii and Rhizobium leguminosarum is required for efficient nitrogen fixation and nodulation of soybeans (9, 15). MI dehydrogenase genes (e.g., iolG) and inosose dehydratase genes (iolE) have been characterized in S. fredii and Sinorhizobium meliloti (10, 15, 36), and iolA and iolDEB genes have been studied in R. leguminosarum (9). An MI utilization gene cluster has also been described for Clostridium perfringens, which was induced by MI via the IolR regulator (16), and recently, transcriptome analysis permitted the identification of an iol cluster which allowed the rapid growth of Corynebacterium glutamicum on MI (19). However, the first MI catabolic gene cluster was characterized in Bacillus subtilis, and detailed molecular data are available only for this bacterium. The B. subtilis iolRS and iolABC...
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