Genome sequence of Leuconostoc inhae KCTC 3774, isolated from Kimchi

Leuconostoc inhae strain KCTC 3774 is a Gram-positive, non-spore-forming, heterofermentative, spherical or lenticular lactic acid bacterium. Here we announce the draft genome sequence of Leuconostoc inhae KCTC 3774, isolated from traditional Korean kimchi, and describe major findings from its annotation.     

Genome sequence of Leuconostoc fallax KCTC 3537

Leuconostoc fallax is known to be present during the manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the draft genome sequence of the type strain Leuconostoc fallax KCTC 3537 (1,638,971 bp, with a G+C content of 37.5%), which consists of 30 large contigs (>100 bp in size).     

Genome sequence of Leuconostoc argentinum KCTC 3773

Leuconostoc argentinum is one of the most prevalent lactic acid bacteria present during the manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the draft genome sequence of type strain KCTC 3773 of Leuconostoc argentinum (1,720,683 bp, with a G+C content of 42.9%), which consists of 98 large contigs (>100 bp in size).     

Complete genome sequence of Leuconostoc citreum KM20

Leuconostoc citreum is one of the most prevalent lactic acid bacteria during the manufacturing process of kimchi, the best-known Korean traditional dish. We have determined the complete genome sequence of L. citreum KM20. It consists of a 1.80-Mb chromosome and four circular plasmids and reveals genes likely involved in kimchi fermentation and its probiotic effects.     

Genome sequence of Lactobacillus farciminis KCTC 3681

Lactobacillus farciminis is one of the most prevalent lactic acid bacterial species present during the manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the draft genome sequence of the type strain Lactobacillus farciminis KCTC 3681 (2,498,309 bp, with a G+C content of 36.4%), which consists of 5 scaffolds.     

Genome sequence of Lactobacillus coryniformis subsp. coryniformis KCTC 3167

Lactobacillus coryniformis subsp. coryniformis is known to be present during the manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the draft genome sequence of Lactobacillus coryniformis subsp. coryniformis type strain KCTC 3167 (2,964,752 bp, with a G+C content of 42.8%), which consists of 55 scaffolds.     

Genome sequence of Lactobacillus animalis KCTC 3501

Lactobacillus animalis is one of the most prevalent lactic acid bacteria present during the manufacturing process of kimchi, the best-known traditional Korean dish. Here, we present the draft genome sequence of Lactobacillus animalis type strain KCTC 3501 (1,882,795 bp, with a G+C content of 41.1%), which consists of 7 scaffolds.     

Complete Genome Sequence of Leuconostoc carnosum Strain JB16, Isolated from Kimchi

Leuconostoc carnosum strain JB16 was isolated from kimchi, the traditional Korean fermented food. Here, we report the complete genome sequence of L. carnosum strain JB16, consisting of a 1,645,096-bp circular chromosome with a G+C content of 37.24% and four plasmids.     

Metagenomic analysis of kimchi, a traditional Korean fermented food

Kimchi, a traditional food in the Korean culture, is made from vegetables by fermentation. In this study, metagenomic approaches were used to monitor changes in bacterial populations, metabolic potential, and overall genetic features of the microbial community during the 29-day fermentation process. Metagenomic DNA was extracted from kimchi samples obtained periodically and was sequenced using a 454 GS FLX Titanium system, which yielded a total of 701,556 reads, with an average read length of 438 bp. Phylogenetic analysis based on 16S rRNA genes from the metagenome indicated that the kimchi microbiome was dominated by members of three genera: Leuconostoc, Lactobacillus, and Weissella. Assignment of metagenomic sequences to SEED categories of the Metagenome Rapid Annotation using Subsystem Technology (MG-RAST) server revealed a genetic profile characteristic of heterotrophic lactic acid fermentation of carbohydrates, which was supported by the detection of mannitol, lactate, acetate, and ethanol as fermentation products. When the metagenomic reads were mapped onto the database of completed genomes, the Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 and Lactobacillus sakei subsp. sakei 23K genomes were highly represented. These same two genera were confirmed to be important in kimchi fermentation when the majority of kimchi metagenomic sequences showed very high identity to Leuconostoc mesenteroides and Lactobacillus genes. Besides microbial genome sequences, a surprisingly large number of phage DNA sequences were identified from the cellular fractions, possibly indicating that a high proportion of cells were infected by bacteriophages during fermentation. Overall, these results provide insights into the kimchi microbial community and also shed light on fermentation processes carried out broadly by complex microbial communities.     

Monitoring of <Emphasis Type="Italic">Leuconostoc mesenteroides</Emphasis> DRC starter in fermented vegetable by random integration of chloramphenicol acetyltransferase gene

In 2004, Leuconostoc mesenteroides DRC was first used as a starter culture for achieving higher organoleptic effects in Korean kimchi manufacture. For a better understanding of starter growth in a mixed culture system, and for predicting starter predominance in kimchi, a monitoring system for the starter was established. The chloramphenicol resistance marker gene (cat) was randomly integrated into chromosomal DNA of L. mesenteroides DRC using a viral transposon and transposase. The DRC mutant, tDRC2, had a similar growth pattern to the host strain, with no major alteration in phenotypic characteristics. The mutant strain was inoculated into real kimchi, and monitoring of the starter population was successfully achieved. The overall predominance of Leuconostoc in kimchi inoculated with DRC followed the general growth pattern of this genus during kimchi fermentation. Our results also demonstrate the competitive ability of the DRC starter against Leuconostoc from natural flora, maintaining its predominance above 88% during the whole fermentation period. Based on this experiment, the random gene integration method using a transposon was shown to be of utility in transferring any commercial starter into a selectable and monitorable strain for simulation purposes.     

Selective control of lactobacilli in kimchi with nisin

The use of nisin to control the lactobacilli responsible for the over-ripening of kimchi, traditional Korean fermented vegetables, was studied. Of the 40 strains of lactic acid bacteria studied, most were sensitive to nisin at a concentration of 100 IU ml-1, while two strains appeared to be resistant. In MRS broth containing nisin at concentrations of 100-300 IU ml-1, the growth of sensitive strains of Lactobacillus plantarum was delayed for 2-3 d at 20 degrees C. When nisin was added to kimchi at a concentration of 100 IU ml-1, the growth of Lactobacillus spp. was inhibited more than the growth of Leuconostoc spp. Scanning electron micrograph observations confirmed the results, demonstrating the predominance of cocci in kimchi containing nisin. These results suggest that at recommended levels, nisin can be used to preserve kimchi by inhibiting lactobacilli more effectively than other lactic acid bacteria involved in kimchi fermentation.     

Draft Genome Sequence of Weissella koreensis KCTC 3621T

Weissella koreensis is a Gram-positive, rod-shaped, nonmotile, and facultative anaerobic species belonging to the lactic acid bacteria (LAB). The members of this species have been repeatedly isolated from kimchi (a traditional Korean fermented food) and are known for their beneficial effects on human and animal intestinal microflora through producing various clinically important amino acids such as γ-aminobutyric acid and ornithine. Here we report the genome sequence of the type strain of W. koreensis (KCTC 3621^T) to provide taxonomic and functional insights into the species.     

Novel Leuconostoc citreum starter culture system for the fermentation of kimchi, a fermented cabbage product

To determine the dominant microorganisms involved in kimchi fermentation and to examine their effect on kimchi fermentation, we randomly isolated and characterized 120 lactic acid bacteria from kimchi during a 5-day fermentation at 15°C. Leuconostoc citreum was dominant during the early and mid-phases of kimchi fermentation whereas Lactobacillus sake/Lactobacillus curvatus or Lactobacillus brevis were found during later stages. Eighty-two out of 120 isolates (68%) were identified as Leuconostoc citreum by means of a polyphasic method, including 16S rDNA sequencing and DNA/DNA hybridization. A few Weissella confusa-like strains were also isolated during the mid-phase of the fermentation. Strain IH22, one of the Leuconostoc citreum isolates from kimchi, was used as an additive to evaluate growth and acid production in kimchi fermentation. This strain was consistently over 95% of the population in IH22-treated kimchi over a 5-day fermentation, while heterogeneous lactic acid bacteria were observed in the control kimchi. The pH in IH22-treated kimchi dropped rapidly but was stably maintained for 5 days, compared to its slow and prolonged decrease in the control kimchi. These results indicate that Leuconostoc citreum IH22 dominates over and retards the growth of other lactic acid bacteria in kimchi, suggesting it can be used as a bacterial starter culture to maintain the quality of kimchi for prolonged periods. This revised version was published online in June 2006 with corrections to the Cover Date.     

Characterization of Leuconostoc gasicomitatum sp. nov., associated with spoiled raw tomato-marinated broiler meat strips packaged under modified-atmosphere conditions

Lactic acid bacteria (LAB) associated with gaseous spoilage of modified-atmosphere-packaged, raw, tomato-marinated broiler meat strips were identified on the basis of a restriction fragment length polymorphism (RFLP) (ribotyping) database containing DNAs coding for 16S and 23S rRNAs (rDNAs). A mixed LAB population dominated by a Leuconostoc species resembling Leuconostoc gelidum caused the spoilage of the product. Lactobacillus sakei, Lactobacillus curvatus, and a gram-positive rod phenotypically similar to heterofermentative Lactobacillus species were the other main organisms detected. An increase in pH together with the extreme bulging of packages suggested a rare LAB spoilage type called "protein swell." This spoilage is characterized by excessive production of gas due to amino acid decarboxylation, and the rise in pH is attributed to the subsequent deamination of amino acids. Protein swell has not previously been associated with any kind of meat product. A polyphasic approach, including classical phenotyping, whole-cell protein electrophoresis, 16 and 23S rDNA RFLP, 16S rDNA sequence analysis, and DNA-DNA reassociation analysis, was used for the identification of the dominant Leuconostoc species. In addition to the RFLP analysis, phenotyping, whole-cell protein analysis, and 16S rDNA sequence homology indicated that L. gelidum was most similar to the spoilage-associated species. The two spoilage strains studied possessed 98.8 and 99.0% 16S rDNA sequence homology with the L. gelidum type strain. DNA-DNA reassociation, however, clearly distinguished the two species. The same strains showed only 22 and 34% hybridization with the L. gelidum type strain. These results warrant a separate species status, and we propose the name Leuconostoc gasicomitatum sp. nov. for this spoilage-associated Leuconostoc species.     

Effect of Leuconostoc spp. on the formation of Streptococcus mutans biofilm

Insoluble glucans synthesized by Streptococcus mutans enhance the pathogenicity of oral biofilm by promoting the adherence and accumulation of cariogenic bacteria on the surface of the tooth. The objective of this study was to investigate the effect of Leuconostoc spp. on the in vitro formation of S. mutans biofilm. Three strains, Leuconostoc gelidum ATCC 49366, Leuconostoc mesenteroides ssp. cremoris ATCC 19254 and Leuconostoc mesenteroides ssp. mesenteroides ATCC 8293, were used in this study. They exhibited profound inhibitory effects on the formation of S. mutans biofilm and on the proliferation of S. mutans. The water-soluble polymers produced from sucrose were most strongly produced by L. gelidum, followed by L. mesenteroides ssp. cremoris and L. mesenteroides ssp. mesenteroides. The mean wet weights of the artificial biofilm of S. mutans were also significantly reduced as a result of the addition of the water-soluble polymers obtained from Leuconostoc cultures. According to the results of thin-layer chromatographic analysis, the hydrolysates of the water-soluble polymers produced by Leuconostoc were identical to those of dextran T-2000, forming predominately alpha-(1-6) glucose linkages. These results indicate that dextran-producing Leuconostoc strains are able to inhibit the formation of S. mutans biofilm in vitro.     

A rapid method for identification of typical Leuconostoc species by 16S rDNA PCR-RFLP analysis

A polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) method was developed to detect and identify typical Leuconostoc species. This method utilises a set of specific primers for amplification of the 16S rDNA region of typical Leuconostoc species. All Leuconostoc-type strains, all Leuconostoc isolates from kimchi, Korea's traditional, fermented vegetable product, and strains from closely related genera were examined to verify the identification by this method. The primers resulted in amplification only for nine typical Leuconostoc spp., but not for any other genera tested. The size of the amplified products was 976 bp and the amplicons of the different species could be differentiated from each other with MseI, HaeIII and Tsp509I endonucleases, except for the species Leuconostoc argentinum and Leuconostoc lactis, which were indistinguishable. A PCR-RFLP method for the typical Leuconostoc species was optimized to identify a large number of isolates from fermented vegetable product. This PCR-RFLP method enables the rapid and reliable identification of Leuconostoc species and to distinguish them from the other phylogenetically related lactic acid bacteria in food samples.     

Complete genome sequence of Leuconostoc kimchii strain C2, isolated from Kimchi

Leuconostoc kimchii strain C2 was isolated from fermented kimchi in Korea. Here we announce the complete genome sequence of Leuconostoc kimchii strain C2, consisting of a 1,877,174-bp chromosome with a G+C content of 37.9% and no plasmid and describe major findings from its annotation.     

Specific detection of Leuconostoc mesenteroides subsp. mesenteroides with DNA primers identified by randomly amplified polymorphic DNA analysis

Randomly amplified polymorphic DNA analysis using primer 239 (5' CTGAAGCGGA 3') was performed to characterize Leuconostoc sp. strains. All the strains of Leuconostoc mesenteroides subsp. mesenteroides (with the exception of two strains), two strains formerly identified as L. gelidum, and one strain of Leuconostoc showed a common band at about 1.1 kb. This DNA fragment was cloned and sequenced in order to verify its suitability for identifying L. mesenteroides subsp. mesenteroides strains.     

Complete genome sequence of Leuconostoc mesenteroides subsp. mesenteroides strain J18, isolated from kimchi

Leuconostoc mesenteroides subsp. mesenteroides is one of the most predominant lactic acid bacterial groups during kimchi fermentation. Here, we report the complete genome sequence of L. mesenteroides subsp. mesenteroides J18, which was isolated from kimchi. The genome of the strain consists of a 1,896,561-bp chromosome and five plasmids.