DuPont Nutrition and Biosciences, Shanghai 200335, P. R. China.
DuPont Nutrition and Biosciences, Shanghai 200335, P. R. China.
J Dairy Sci. 2020 Sep;103(9):7898-7907. doi: 10.3168/jds.2020-18285. Epub 2020 Jul 1.
Intracellular pH homeostasis through the extrusion of a proton by FF-ATPase is one of the key mechanisms used by lactic acid bacteria in response to acid stress, and also influences their post-fermentation acidification. In this study, the genotypic and phenotypic stability of a low post-fermentation acidification (LPA) mutant (designated as DGCC12411) of Lactobacillus plantarum DGCC12411 was assessed. Compared with its mother strain, the pH of DGCC12411 in De Man, Rogosa, and Sharpe (MRS) broth after 48-h cultivation was 0.35 pH units higher. Incorporation of DGCC12411 in yogurt stored at ambient temperature (ambient yogurt) showed a reduced post-fermentation acidification during storage at 25°C for 120 d. Whole-genome sequencing analysis showed a SNP mutation (GGT > GAT at positions 505 to 507) in DGCC12411, which resulted in the substitution of a highly conserved glycine residue by aspartic acid at the Walker A motif of the FF-ATPase α-subunit. However, degeneration of the LPA phenotype was observed after 5 passages of DGCC12411 in MRS broth. Analysis of DNA sequencing on both the whole population and the isolates showed that a back mutation occurred at the SNP site (GAT changed back to GGT) over the passaging, and the reversion gradually increased from a ratio of 10.8% at P5 to 60.0% at P10. We also found that the LPA phenotype stability of DGCC12411 was improved by supplementing 0.1 M potassium phosphate buffer to the growth medium as well as by reducing the inoculation rate of DGCC12411 to 2% (vol/vol). Such LPA Lactobacillus strains have potential for use as starter cultures in fermented foods with less change in acidity during shelf-life storage.
通过 FF-ATP 酶将质子排出以维持细胞内 pH 平衡是乳酸菌应对酸胁迫的关键机制之一，也会影响其发酵后的酸化程度。本研究评估了植物乳杆菌 DGCC12411 低发酵后酸化（LPA）突变体（命名为 DGCC12411）的遗传和表型稳定性。与母株相比，DGCC12411 在德氏乳杆菌培养基（MRS）中培养 48 小时后的 pH 值高 0.35 个单位。在环境温度下储存时，将 DGCC12411 掺入酸奶中可在 25°C 下储存 120 天期间减少发酵后酸化。全基因组测序分析表明，DGCC12411 中存在 SNP 突变（位置 505 到 507 处的 GGT > GAT），导致 FF-ATP 酶 α 亚基 Walker A 基序中高度保守的甘氨酸残基被天冬氨酸取代。然而，在 MRS 肉汤中传代 5 次后，观察到 LPA 表型的退化。对全群体和分离株的 DNA 测序分析表明，在传代过程中 SNP 位点发生了回复突变（GAT 变回 GGT），并且这种回复突变逐渐增加，从第 5 代的 10.8%增加到第 10 代的 60.0%。我们还发现，通过向生长培养基中补充 0.1 M 磷酸钾缓冲液以及将 DGCC12411 的接种率降低至 2%（体积/体积），可以提高 DGCC12411 的 LPA 表型稳定性。这种具有低发酵后酸化能力的乳酸菌菌株在发酵食品中具有潜力，可以在货架期储存过程中降低酸度变化。
