Recombinant lactobacillin PlnK adjusts the gut microbiome distribution in broilers

1. The heterologous expression and biological function of the Lactobacillus bacteriocin plantaricin K (PlnK) remain largely unknown. 2. In this study, PlnK was efficiently expressed in competent E. coli BL21 (used in transformation and protein expression) after 12 h, at 37°C and in 0.4 mmol/l isopropyl β- d-1-thiogalactopyranoside (IPTG). 3. The inhibitory bacterial spectrum of recombinant PlnK was investigated and indicated that levels of PlnK above 0.10 mg/ml produced an obvious inhibitory effect on gram-positive bacteria and gram-negative bacteria in vitro. 4. The effects of PlnK on intestinal immune function and the gut microbiome distribution in broilers were studied. The results revealed that, after consuming 2.50 × 10˗3 mg/ml of PlnK in water for one week, at the phylum level, the abundance of Firmicutes was increased and the abundance of Bacleroidetes was decreased. At the family level, the abundance of Lachnospiraceae, Ruminococcaceae and Streptococcaceae were significantly improved, but the abundance of Bacteroidaceae was reduced. At the genus level, the abundances of Lachnoclostridium, Streptococcus and Ruminococcaceae-UCG-013, were significantly up-regulated, and the abundance of Bacteroides was down-regulated. 5. After oral liquid intake of PlnK for one week, levels of secretory immunoglobulin A (sIgA) in the duodenal mucus were not significantly increased, but the mRNA levels of TLR3, MDA5, IFN-α, IFN-β, IFITM3 and IFITM10 in the duodenum were significantly reduced. 6. This study demonstrated that the recombinant PlnK could adjust the intestinal microbiome distribution and downregulate the IFN pathway.1. 乳杆菌细菌素植物乳杆菌素K（Lactobacillus bacteriocin plantaricin K，PlnK）的外源表达及其生物学功能，目前仍未得到充分阐明。2. 本研究中，在用于转化与蛋白表达的感受态大肠杆菌BL21（E. coli BL21）中实现了PlnK的高效表达：于37℃、0.4 mmol/L异丙基-β-D-硫代半乳糖苷（isopropyl β-d-1-thiogalactopyranoside，IPTG）诱导条件下培养12 h后，即可获得重组表达产物。3. 本研究对重组PlnK的抑菌谱进行了探究，结果显示：当PlnK浓度高于0.10 mg/mL时，可在体外对革兰氏阳性菌与革兰氏阴性菌产生显著抑制效果。4. 本研究探究了PlnK对肉鸡肠道免疫功能与肠道微生物组分布的影响。结果显示：肉鸡连续一周饮水摄入浓度为2.50×10^-3 mg/mL的PlnK后，在门水平上，厚壁菌门（Firmicutes）相对丰度显著升高，拟杆菌门（Bacteroidetes，原文笔误为Bacleroidetes）相对丰度显著降低；在科水平上，毛螺菌科（Lachnospiraceae）、瘤胃球菌科（Ruminococcaceae）与链球菌科（Streptococcaceae）相对丰度显著提升，拟杆菌科（Bacteroidaceae）相对丰度下降；在属水平上，拉氏梭菌属（Lachnoclostridium）、链球菌属（Streptococcus）及瘤胃球菌科UCG-013属（Ruminococcaceae-UCG-013）相对丰度显著上调，拟杆菌属（Bacteroides）相对丰度显著下调。5. 连续一周经口摄入PlnK后，肉鸡十二指肠黏液中的分泌型免疫球蛋白A（secretory immunoglobulin A，sIgA）水平无显著升高，但十二指肠组织中Toll样受体3（TLR3）、黑色素瘤分化相关基因5（MDA5）、干扰素α（IFN-α）、干扰素β（IFN-β）、干扰素诱导跨膜蛋白3（IFITM3）及干扰素诱导跨膜蛋白10（IFITM10）的mRNA表达水平均显著下调。6. 本研究证实，重组PlnK可调控肠道微生物组分布，并下调干扰素通路活性。