| Sauerkraut is a traditional food which commonly fermented by lactic acid bacteria(LAB),and has unique flavor and a long history.Its manufacturing process could date back to more than 2,000 years ago."The Book of Songs · Xiaoya·Shennan Mountain"had recorded that "there is a residential house in the middle of the field,which melons and vegetables were scattered and grown at the edge of the field.Peel and cut into pieces to make‘ZU',and dedicated to great ancestors",and the vocabulary of 'ZU' is the old name for sauerkraut.Sauerkraut is very popular in the world due to its characteristics of sour and delicious,crisp and tender,refreshing,appetizing,and adjusting the microbiota balance of the gastrointestine.As a typical representative of traditional fermented food in China,researched on sauerkraut still focused on the screening and separation of fermentation strains,and fermentation process.However,reports about the systematic analysis of the genome of fermentation strains is still few.Lactobacillus plantarum NCU116 was a typical dominant strain isolated from traditional Chinese sauerkraut(TCS)brine.In the past decade,our team has done lots of researches to analyze its characteristics.Which has showed that NCU116 exhibits excellent fermentation performance and has great prebiotic function(including constipation alleviation effect,modulation effect on immune system and intestinal microbiota,etc.)In this study,we first illustrated the complete genome of the TCS derived fermentation stain NCU116,and comparative genomics were applied to in-depth information mining,including gene composition,metabolic characteristics,fermentation characteristics and probiotic functions associated genes.These studies were of great exemplary significance to improve the theoretical research of fermentation and probiotic strains with independent intellectual property rights,as well as the promotion and protection of Chinese traditional fermented foods and culture.The main findings in this thesis were as follows:1.The complete genome of NCU116 is comprised of one circular chromosome(3,245,809 bp),one small(48,567 bp)and two large plasmids(48,567 bp and 48,695 bp,respectively).3,252 coding sequences(CDSs)were identified in the NCU116 genome,of which 67 were tRNA,16 were rRNA and 17 were potential sRNAs,respectivly.2.Various database including NR,SwissProt,COG,GO,and KEGG were used to annotate the functional genes of NCU116.Results showed that relative abundance of genes associated with membrane transport,carbohydrate metabolism,amino acid metabolism is very high,which indicated that NCU116 is a strain with strong metabolic potential.According to the results of annotation by PHI-base and VFDB database,no pathogenic or virulence genes was retrieved in NCU116,suggesting that NCU116 is a safe and reliable strain at the genetic level.3.The key enzymes related to the carbohydrate metabolic pathways(including glycosis,pentose phosphate pathway and pyruvate metabolism,etc.)were found in the NCU116.In addition,high abundance of genes related to pyruvate metabolic pathway and 5 lactate dehydrogenases ldh were found,suggesting that NCU116 is a strain which endows an efficient lactic acid synthesis ability,and this could be a main reason of good fermentation performance and strong acid production capacity in sauerkraut fermentation process.4.Multiple glycoside hydrolases related to oligo-/poly-saccharide hydrolyases were enriched in NCU116,which indicated that the NCU116 has the ability to degradate sugar-chain linked by ?/?-glycosidic bonds.Of which 12 genes were belong to GH13 family amylase,which is the basis that the NCU116 uptakes and utilizes the starch and polysaccharides of plant tissue,and it's also the important basis that NCU116 adapts to plant-based fermentation.5.Relatively complete amino acid and protein metabolic pathway related genes were found in NCU116,suggesting that NCU116 has the potential to ferment in protein-rich substrates.Besides,the lack of asdA decarboxylase related genes lead that the metabolic pathways of alanine and arginine are incomplete,suggesting that alanine could be an essential amino acid for NCU116.6.A glutamic acid decarboxylase gadB and two bifunctional glutathione(GSH)synthase gshF were identified in NCU116,which indicated that it has the potential to synthesize ?-aminobutyric acid(GABA)and GSH.Besides,5 gor reductases were retrieved in NCU116,and the gor reductases could transfer the oxidized GSSG to reduced GSH for coping with oxidative stress,which was consistent with that these genes played an important for the antioxidant capacity and alleviated organ oxidative stress injury in vivo.In addition,4 genes related to bile salt hydrolase BSH and cholesterol lowering in NCU116 were also found.7.Re-annotation of functional genes in NCU116 has been completed.Which was performed to obtain more accurate and feature-rich information for NCU116.The evolution status of NCU116 were identified through the analysis of whole-genome-based phylogenetic and synteny.After that,we compared the whole genome of NCU116 and Lactobacillus from different sources,and found that 171 unique genes in NCU116.Subsequently,the evolutionary mechanism of adaption to pickle fermentation environment was analyzed according the evolutionary level,and these evolutionary mechanisms may include obtaining foreign functional genes by horizontal gene transfer and immuning to the invasion of foreign mobile elements by pre phage elements and restriction/modification system.8.Based on the functional genes from different Lactobacillus,key metabolic pathways were reconstructed,and results showed that the most abundant genes related to carbohydrate transport and metabolism was found in NCU116,which indicated that NCU116 is more suitable for the fermentation of plant substrate.Besides,a novel bacteriocin synthetic cluster responsable for ?-b plantaricin was also identified in NCU116,which is the basis that It plays a group reaction in the complex pickle system.9.Four extracellular protein secretion pathways and 345 potential extracellular protein coding genes were found in NUC116.Subsequently,and these genes were classified based on function and anchor type,and then we constructed its extracellular protein encoding genes library.45 of these extracellular proteins were identified to be membrane-anchored,and of which 11 were GH13 family amylases,which could play critical roles in utilization of plant-based macromolecular sugar,especially the oligosaccharides in plant tissues,and were also one of the reasons that the NCU116 could adapt to the fermentation of plant substrate.In addition,17 potential adhesin in these extracellular proteins could provide a theoretical basis for its colonization and probiotic effects in gastrointestinal.Furthermore,EPS116c1-4 clusters responsible for EPS116 synthesis and specific eps116c1 gene cluster were found,which is also the theoretical basis that the exopolysaccharide in NCU116 differs from that in other L.plantarum strains. |
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