Multilocus Sequence Typing Of Lactobacillus Helveticus From Naturally Fermented Dairy Products In Different Regions

The system genetics evolutionary relationship of 245 Lactobacillus helveticus isolates collected from Inner Mongolia, Tibet, Qinghai, YunNan, Sichuan, Gansu, Xinjiang and Mongolia was investigated by multilocus sequence typing (MLST) with 11 housekeeping genes(clpX, dnaA, dnaK, groEL, murC, murE,pepN,pepX,pyrG, recA and uvrC). The high quality sequences of these housekeeping genes were acquired by bidirectional sequencing technology, and were trimmed and aligned. Then the allele diagram was gained through the random allele number was given to distinct combination of the eleven alleles, and then each random allele number was assigned as a sequence type (ST). The gene polymorphism and heredity evolution of 245 Lactobacillus helveticus isolates were analyzed based on the STs and composite sequences. The conclusions were showed as follows:The number of alleles per eleven housekeeping gene locus ranged from 5 (clpX) to 18 (uvrC). And the DNA G+C% content of the eleven genes ranged from 35.91%(clpX) to 46.30%(pepN). The number of polymorphic sites ranged from 5(clpX)-S0 (pepX). The nucleotide diversity site (π) ranged from 0.00373 (murE) to 0.06233(pepX). The ds/du ratio ranged from 0.0255 (pepX) to 0.7614 (clpX), and they were less than 1.245 Lactobacillus helveticus isolates were assigned as 108 STs, and ST-1 contains the largest number (21 isolates). IAS of 0.1948 was greater than the IAS value of 0, expecting that linkage disequilibrium was present in the population.The 108 STs were grouped into eight clonal complexes (CC7, CC21, CC25, CC27, CC46, CC63, CC67 and CC104) and twenty-seven singletons by the eBURST. Among eight CCs, CC27 was the major CC, including the most strains, and CC27 was considered as founder clonal complex.The Minimum-spanning Tree realuts analysed the heredity evolution relationship of 245 Lactobacillus helveticus isolates, and showed that the kinship of strains was positive correlation to the distance of regions. The minimum spanning tree of STs of 245 Lactobacillus helveticus isolates based on regions and sources revealed all isolates were divided into four branchs in which contained the same STs.The phylogenetic analysis realuts revealed that 245 Lactobacillus helveticus isolates identified three separate lineages, namely the koumiss, kurut-tarag, and panmictic lineages, respectively. The heredity distance of strains from Xinjiang province was smaller, and the population evolution of these srains was shorter. During the processing and preservation of naturally fermented dairy products, the genes of L. helveticus mutated based on selection pressures. The conservative genes was reserved, composing its own unique feature.This research analyzed the genetic background of Lactobacillus helveticus isolated from different regions, enriched the Lactobacillus helveticus MLST global database, and then deducted the Lactobacillus helveticus evolutionary process. The diversity of Lactobacillus helveticus provided theory basis for industrial food starter culture design.