二甲酸钾预防沙门菌感染小鼠的效果评价及对肠道菌群的影响

doi:10.11843/j.issn.0366-6964.2023.05.031

畜牧兽医学报 ›› 2023, Vol. 54 ›› Issue (5): 2101-2113.doi: 10.11843/j.issn.0366-6964.2023.05.031

二甲酸钾预防沙门菌感染小鼠的效果评价及对肠道菌群的影响

孙瑜凡¹, 于盼元¹, 陈虹宇¹, 谭怡青¹, 陈夏冰², 张腾飞³, 高婷³, 周锐^1*, 黎璐^1*

1. 华中农业大学动物医学院生猪健康养殖省部共建协同创新中心, 武汉 430070;
2. 武汉市农业科学院畜牧兽医研究所, 武汉 430070;
3. 湖北省农业科学院畜牧兽医研究所, 农业农村部畜禽细菌病防治制剂创制重点实验室, 畜禽病原微生物学湖北省重点实验室, 武汉 430064

收稿日期:2022-07-04 出版日期:2023-05-23 发布日期:2023-05-20
通讯作者: 黎璐,主要从事动物传染病防控技术相关研究,E-mail:lilu@mail.hzau.edu.cn;周锐,主要从事动物传染病防控技术相关研究,E-mail:rzhou@mail.hzau.edu.cn
作者简介:孙瑜凡(1997-),河北邢台人,博士,主要从事动物传染病研究,E-mail:sunyufan@webmail.hzau.edu.cn;于盼元(1998-),男,河北定州人,硕士,主要从事动物传染病研究,E-mail:yupanyuan@webmail.hzau.edu.cn
基金资助:
国家重点研发计划(2021YFD1800404);农业农村部畜禽细菌病防治制剂创制重点实验室/畜禽病原微生物学湖北省重点实验室开放课题(KLPCAAB202201);中央引导地方科技发展专项(2020ZYYD029)

Evaluation of the Efficacy of Potassium Diformate in the Prevention of Salmonella Infection and the Effect on Intestinal Flora

SUN Yufan¹, YU Panyuan¹, CHEN Hongyu¹, TAN Yiqing¹, CHEN Xiabing², ZHANG Tengfei³, GAO Ting³, ZHOU Rui^1*, LI Lu^1*

1. The Cooperative Innovation Center for Sustainable Pig Production Co-sponsored by Province and Ministry, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China;
2. Institute of Animal Husbandry and Veterinary Science, Wuhan Academy of Agricultural Sciences, Wuhan 430070, China;
3. Institute of Animal Husbandry and Veterinary, Hubei Academy of Agricultural Sciences, Key Laboratory of Prevention and Control Agents for Animal Bacteriosis (Ministry of Agriculture and Rural Affairs), Hubei Provincial Key Laboratory of Animal Pathogenic Microbiology, Wuhan 430064, China

Received:2022-07-04 Online:2023-05-23 Published:2023-05-20

摘要/Abstract

摘要： 二甲酸钾(potassium diformate, KDF)为酸化剂的代表之一,在2001年被欧盟批准可作为饲料添加剂使用,在2005年被我国批准作为饲料添加剂。目前已有较多的研究表明KDF可以显著提高动物的生长性能,然而关于KDF对传染病预防作用的直接证据的研究还比较少。本研究旨在通过体外抑菌试验及小鼠感染模型,在体外和体内评价KDF对致病菌的预防效果,并检测KDF对小鼠肠道菌群的影响,以探讨KDF的作用机理。在体外检测KDF对养殖场中常见病原菌的最低抑菌浓度(minimum inhibitory concentration, MIC)后,选取3~4周龄,体重12~15 g的C57/BL6 雌鼠,分组饲喂一周后感染鼠伤寒沙门菌,记录KDF饲喂组、KDF饮水组和对照组小鼠感染后的体重变化(n=6),并在感染后第6天将小鼠安乐死检测其盲肠组织载菌量(n=6)、血清炎性细胞因子水平(n=6)和盲肠病理变化(n=3)。另外选取两组3~4周龄,体重12~15 g的C57/BL6 雌鼠(n=5),饲喂含有KDF的饲料两周后采集其粪便,利用16S rRNA测序技术分析饲喂KDF对小鼠盲肠菌群的影响。结果表明,KDF对沙门菌、大肠杆菌等病原菌的MIC为3.125 mg·mL^-1,对金黄色葡萄球菌和巴氏杆菌等的MIC为6.25 mg·mL^-1。在体内,KDF预处理显著降低了小鼠感染沙门菌后血液和盲肠的载菌量(P<0.05),显著降低了感染小鼠的血液中炎性细胞因子IL-6、IL-12和TNF-α的水平(P<0.05),并缓解了小鼠盲肠的病理变化。同时,KDF显著降低了小鼠盲肠菌群的多样性(P<0.01),表示与对照组相比存在差异物种。此外,KDF组小鼠的盲肠物种丰富度下降,在门水平,KDF组中变形菌门的相对丰度显著升高(P<0.001);在科水平,KDF组中多杆菌科的相对丰度显著下降(P<0.05);在属水平,阿克曼菌属、布劳特氏菌属、乳酸杆菌属、真杆菌属等相对丰度升高。综上表明,KDF在体外对常见病原菌具有显著的抑制效果,在小鼠体内具有预防肠道致病菌沙门菌感染的效果。使用KDF后小鼠的肠道菌群发生了显著变化,阿克曼菌、布劳特菌、乳酸杆菌、真杆菌等有益菌属丰度的升高可能对动物的肠道健康起到积极作用。

关键词: 二甲酸钾, 肠道细菌感染, 预防, 沙门菌, 肠道菌群

Abstract: Potassium diformate (KDF) is one of the representatives of acidifiers, which was approved for use as a feed additive by the European Union in 2001 and by China in 2005. Currently, there are more studies showing that KDF can improve the growth performance of animals. However, there are less researches to provide direct evidence of the prevention effect of KDF to infectious disease. The aim of this study was to evaluate the preventive effect of KDF on pathogenic bacteria in vitro and in vivo through bacterial inhibition assay and mouse infection model, and to detect the effect of KDF on intestinal flora of mice in order to investigate the mechanism of KDF action. First, detection of the minimum inhibitory concentration (MIC) of KDF against common pathogenic bacteria in the farm. Subsequently, C57/BL6 female mice aged 3-4 weeks and weighing 12-15 g were selected and examined for changes in body weight, tissue load, inflammatory cytokine levels and cecum pathology after infection with Salmonella Typhimurium in KDF treatment group (n=6) and control mice (n=6). The effects of feeding KDF on the cecum flora of mice were analyzed using 16S rRNA sequencing technology. The results showed that the MIC of KDF was 3.125 mg·mL^-1 for pathogenic bacteria such as Salmonella and Escherichia coli, and 6.25 mg·mL^-1 for bacteria such as Staphylococcus aureus and Pasteurella. In vivo, KDF pretreatment significantly reduced the bacterial loads in the blood and cecum (P<0.05), significantly reduced the levels of inflammatory cytokines IL-6, IL-12 and TNF-α in the blood (P<0.05), and alleviated the pathological changes in the cecum. Simultaneously, KDF significantly reduced the diversity of the intestinal flora of the cecum (P<0.01), which allowed the differential species presence. In addition, the species richness of the cecum flora of mice in the KDF treatment group decreased. At the phylum level, the relative abundance of the Proteobacteria was significantly higher in the KDF group (P<0.001). At the family level, the relative abundance of the Mogibacteriacceae was markedly lower in the KDF group (P<0.05). At the genus level, the relative abundance of the Akkermansia, Blautia, Lactobacillus and Eubacterium were increased. In summary, KDF showed significant inhibitory effects against common pathogenic bacteria in vitro, and prevents infection with the Salmonella in mice in vivo. The intestinal flora of mice changed significantly after KDF supplementation, and the increase in the abundance of beneficial genera such as Akkermansia, Blautia, Lactobacillus and Eubacterium may have a positive effect on the intestinal health of the animals.

Key words: potassium diformate, intestinal bacterial infection, prevention, Salmonella, intestinal flora

中图分类号:

S816.7

孙瑜凡, 于盼元, 陈虹宇, 谭怡青, 陈夏冰, 张腾飞, 高婷, 周锐, 黎璐. 二甲酸钾预防沙门菌感染小鼠的效果评价及对肠道菌群的影响[J]. 畜牧兽医学报, 2023, 54(5): 2101-2113.

SUN Yufan, YU Panyuan, CHEN Hongyu, TAN Yiqing, CHEN Xiabing, ZHANG Tengfei, GAO Ting, ZHOU Rui, LI Lu. Evaluation of the Efficacy of Potassium Diformate in the Prevention of Salmonella Infection and the Effect on Intestinal Flora[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 2101-2113.

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二甲酸钾预防沙门菌感染小鼠的效果评价及对肠道菌群的影响

Evaluation of the Efficacy of Potassium Diformate in the Prevention of Salmonella Infection and the Effect on Intestinal Flora

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