Physical and genetic map of Streptococcus thermophilus A054

Abstract

The three restriction endonucleases Sfil, BssHl, and SmaI were found to generate fragments with suitable size distributions for mapping the genome of Streptococcus thermophilus A054. A total of 5, 8, and 24 fragments were produced with SfiI, BssHI, and SmaI, respectively. An average genome size of 1,824 kb was determined by summing the total fragment sizes obtained by digestions with these three enzymes. Partial and multiple digestions of genomic DNA in conjunction with Southern hybridization were used to map SfI, BssHII, and SmaI fragments. All restriction fragments were arranged in a unique circular chromosome. Southern hybridization analysis with specific probes allowed 23 genetic markers to be located on the restriction map. Among them, six rrn loci were precisely located. The area of the chromosome containing the ribosomal operons was further detailed by mapping some of the ApaI and SgrAI sites. Comparison of macrorestriction patterns from three clones derived from strain A054 revealed two variable regions in the chromosome. One was associated with the tandem rinD and rnnE loci, and the other was mapped in the region of the lactose operon.Streptococcus thermophilus is an economically important lactic acid bacterium used in the manufacture of yoghurt and some hard cheeses. It is a gram-positive, facultative anaerobic microorganism that is exclusively isolated from milk and fermented dairy products. Despite its importance in the dairy industry, genetic studies of S. thermophilus have been limited until recently. To date, basic methods of gene transfer by transformation, transduction, or conjugative mobilization have been reported (34). Several genes have been cloned and analyzed, including genes involved in glucide metabolism (lacZ, lacS, galE, galM, and ldh [5,25,43,44,51]) and DNA recombination (recA [15]), ribosomal sequences (4, 23, 38), and the aspartate racemase gene (62). A marked restriction polymorphism has been detected between different strains of this species by analysis of hybridization patterns obtained with a species-specific probe (13) and with probes by genes coding for rRNA (38). Despite the availability of gene transfer systems and cloned genes, the structure and organization of the S. thermophilus genome are not well defined.Pulsed-field gel electrophoresis (PFGE) (9, 52) has proved to be an efficient method for genome size estimation and construction of chromosomal maps, as well as being a useful tool for the characterization of bacterial species. Preliminary results from PFGE have indicated a genome size of 1.75 Mb for strain STi of S. thermophilus (31) and allowed restriction polymorphism within S. thermophilus to be observed (48).In this study, we used PFGE to estimate the size of the chromosome of S. thermophilus A054 and to construct a physical map by using the restriction enzymes SMaI, Sfil, BssHII, ApaI, and SgrAI. Hybridization probes were used to locate 23 genetic markers on the chromosomal map. Genetic polymorphism within strain A054 was analyzed by comparing the macrorestric...

Construction of a combined physical and genetic map of the chromosome of Lactobacillus acidophilus ATCC 4356 and characterization of the rRNA operons

“…1). Chromosomal clustering of the rrn loci has not been found in L. plantarum (Kleerebezem et al, 2003), but it has been described for L. gasseri (Abs EL-Osta et al, 2002), L. sakei (Dudez et al, 2002) and Lactococcus lactis (Tulloch et al, 1991;Le Bourgeois et al, 1992) and for prokaryotes other then the lactic acid bacteria, such as E. coli, Bacillus subtilis, Haemophilus influenzae, Pasteurella multocida and Streptococcus thermophilus (Lee et al, 1989;Bentley et al, 1991;Roussel et al, 1994;Itaya, 1997;Hunt et al, 1998). In these bacteria, the rrn loci are often located within one-third to one-half of the chromosome.…”

Multireplicon genome architecture of Lactobacillus salivarius

“…Lactacin F production by L. acidophilus strain 88 was shown by conjugation analysis to be linked to a 110-kb plasmid, pPM68 (40). A plasmid of 150 kb was identified in L. gasseri by PFGE and was suggested to be linear on the basis of electrophoretic behavior (41). However, its size and conformation were not confirmed.…”

Physical map of the genome of Planctomyces limnophilus, a representative of the phylogenetically distinct planctomycete lineage

“…It is interesting that in P. limnophilus, each atpD gene is near an rrs locus, suggesting a possible duplication of the whole chromosomal segment containing these two genes. The large repeated elements of the rrn loci have been suggested as good candidates for intrachromosomal homologous recombination, and many intraspecies chromosomal variations are found between rrn loci (5,19,29). In addition, extra copies of the region of the chromosome close to the origin of replication have been reported (16).…”

“…A relatively large genome, facilitating adaptation to changing conditions, has been suggested to be necessary for bacteria which live in nutrient-poor or fluctuating environments (29). Eutrophic lakes, such as that from which P. limnophilus was isolated (13), could be considered to constitute such a demanding environment.…”