Effects of Co-culture of Lactobacillus casei NA-2 and Lactobacillus rhamnosus LGG on Antibacterial Activity

doi:10.13304/j.nykjdb.2022.0983

Abstract

Abstract:

The application of lactic acid bacteria in food， medicine and other industry depend on their antibacterial activities. To improve antibacterial activity by optimizing the co-cultural conditions of different lactic acid bacteria， the antibacterial activity of Lactobacillus casei NA-2 was determined by double agar diffusion method using Escherichia coli K88 as indicator bacteria， and the culture system was supplemented with Lactobacillus rhamnosus LGG in order to monitor any changes in antibacterial activity. The co-culture conditions of L. casei NA-2 and L. rhamnosus LGG including inoculation ratio， culture temperature， time， aerobic， carbon and nitrogen sources and pH were optimized by single factor test. The total viable cell counts and lactic acid contents of L. casei NA-2 and L. rhamnosus LGG were detected by quantitative real-time PCR method and high performance liquid chromatography， respectively. The results showed that the antibacterial activity was significantly increased after L. rhamnose LGG was added into L. casei NA-2 for co-culture. By comparing L. casei NA-2 pure culture with co-culture without optimization in optimal co-culture conditions， it revealed increases in antibacterial activity of 20.7% and 11.0% and lactic acid production of 15.2% and 7.3%， respectively. The overal viable cell counts decreased from 2.74×10⁸ to 1.57×10⁸ CFU·mL^-1 after co-culture conditions optimized， but the proportion of L. casei NA-2 increased from 7.0% to 65.6%. Above results provided data support for further research and application of lactic acid bacteria co-culture system.

Key words: co-culture, Lactobacillus, antibacterial activity, condition optimization

摘要：

乳酸菌的抑菌活性决定了其在食品、医药等领域的应用效果。为通过不同乳酸菌的共培养提高其抑菌活性，采用双层琼脂扩散法以大肠杆菌K88为指示菌测定干酪乳杆菌NA-2及其添加鼠李糖乳杆菌LGG共培养后的抑菌活性，首先通过单因素试验对2株菌的共培养体系包括接种比例、培养温度、时间、通气方式、碳氮源、pH等条件进行优化，然后应用实时荧光定量PCR和高效液相色谱法分别对共培养条件优化前后的活菌数和乳酸含量进行测定。结果表明，干酪乳杆菌NA-2与鼠李糖乳杆菌LGG共培养后抑菌活性显著增加；与干酪乳杆菌NA-2单菌株培养和优化前共培养体系相比，优化后共培养体系的抑菌活性分别提高20.7%和11.0%，乳酸产量分别提高15.2%和7.3%；干酪乳杆菌NA-2和鼠李糖乳杆菌LGG的总活菌数由优化前共培养体系中的2.74×10⁸ CFU·mL^-1下降到优化后的1.57×10⁸ CFU·mL^-1，但干酪乳杆菌NA-2的比例从7.0%提高到65.6%。以上研究结果为乳酸菌共培养体系的深入研究和应用提供数据支撑。

关键词: 共培养, 乳酸菌, 抑菌, 条件优化

CLC Number:

S816.3

Linbo SHI, Qing PENG, Xiaoqing XU, Yuwei ZHANG, Shuo YANG, Dandan TIAN, Mengxin HE, Bo SHI, Yu QIAO. Effects of Co-culture of Lactobacillus casei NA-2 and Lactobacillus rhamnosus LGG on Antibacterial Activity[J]. Journal of Agricultural Science and Technology, 2024, 26(6): 63-71.

施琳波, 彭晴, 徐小轻, 张宇微, 杨硕, 田丹丹, 何孟欣, 石波, 乔宇. 共培养干酪乳杆菌NA-2和鼠李糖乳杆菌LGG对抑菌活性的影响[J]. 中国农业科技导报, 2024, 26(6): 63-71.

Figures/Tables 6

Fig. 1 Growth of Lactobacillus casei NA-2 and Lactobacillus rhamnosus LGG by pure culture and co-cultureA： Growth curve； B： Colony of forming unit. LGG— Lactobacillus rhamnosus LGG； NA-2— Lactobacillus casei NA-2

Fig. 2 Antibacterial activity of Lactobacillus casei NA-2 and Lactobacillus rhamnosus LGG on K88 cultured alone and co-cultured at different concentrationsA： Undiluted culture supernatant；B： Double dilution of culture supernatant；C： Quadruple dilution of culture supernatant. a— Lactobacillus casei NA-2 pure culture； b—Lactobacillus rhamnosus LGG pure culture； c—Co-culture without optimization； d— Co-culture with optimization

Fig. 3 Antibacterial activity of co-culture under different inoculation ratios of 2 LAB strainsNote： Different lowercase letters indicate significant differences between different treatments at P<0.05 level.

Fig. 4 Antibacterial activity under different culture conditionsNote： Different lowercase letters indicate significant differences between different treatments at P<0.05 level.

Fig. 5 Antibacterial activity under different medium conditionNote： Different lowercase letters indicate significant differences between different treatments at P<0.05 level.

Table 1 Growth and antibacterial activity of two strains of lactic acid bacteria under different culture conditions

指标

Index

NA-2单培养

NA-2 pure culture

LGG单培养

LGG pure culture

共培养-优化前

Co-culture without optimization

共培养-优化后

Co-culture with optimization

活菌数

Colony forming unit/ （10⁸ CFU·mL^-1）

1.20±0.11

2.80±0.03

NA-2 0.19±0.09

LGG 2.55±0.06

合计Total 2.74±0.06

NA-2 1.03±0.11

LGG 0.54±0.01

合计Total 1.57±0.12

抑菌圈直径

Inhibition zone diameter/mm

乳酸

Lactic acid/（g·L^-1）

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