【摘要】 目的探讨评价哺乳期无乳链球菌乳腺炎的潜在生物标志物与乳汁微生物及代谢物之间的关系。方法将BALB/C哺乳小鼠30只,随机数字表法分为正常组、模型组、给药组,每组10只。模型组于第3乳注射50μL的无乳链球菌。正常组注射等量的0.9%氯化钠溶液,给药组注射含有青霉素的同等浓度的菌液。采用16S评价菌群多样性差异,气质测定代谢物表达,应用免疫印迹法测定干扰素(IFN-α)、白细胞介素-4(IL-4)和诱导型一氧化氮合成酶(i NOS)蛋白表达量,应用实时荧光定量聚合酶链反应(RT-PCR)检测TNF-αmRNA表达量。结果与正常组相比模型组乳汁中克隆菌落单位(CFU)活菌数、厚壁菌门的相对丰度显著增加,拟杆菌、放线菌、变形菌门水平相对丰度均显著降低(P <0.05);与模型组相比,给药组以上指标变化趋势均与上述相反,差异有统计学意义(P <0.05)。与无乳链球菌乳腺炎关联最重要的4个代谢途径是半乳糖代谢、淀粉蔗糖代谢、戊糖和葡萄糖醛酸酯相互转化和三羧酸(TCA)循环。与空白对照组(1.00±0.03、1.00±0.02、1.00±0.01)相比,模型组小鼠乳汁中的IFN-α、IL-4和i NOS蛋白表达量(4.46±0.20、4.87±0.57、3.31±0.22)显著升高(P <0.05),给药组小鼠乳汁中的各蛋白表达量(2.94±0.37、2.87±0.29、2.34±0.10)与模型组相比显著降低,且更接近于正常组,差异有统计学意义(P <0.05)。与空白对照组(1.00±0.01)相比,模型组小鼠的TNF-αmRNA表达量(0.13±0.04)显著降低,给药组的TNF-αmRNA表达量(0.66±0.07)较模型组恢复明显,略低于正常对照组。结论哺乳期无乳链球菌乳腺炎乳腺组织中炎症因子的表达与乳汁微生物和代谢物密切相关,从而对奶牛产奶质量造成影响。更多还原

【Abstract】 Objective To evaluate the relationship between potential biomarkers of lactation Streptococcus agalactiae mastitis and milk microorganisms and metabolites.Methods Thirty 30 mature BALB/C mice were divided into 3 groups with 10 mice in each group.The mice are divided into normal group,model group and medication group by random number table.The model group was injected with 50 μL streptococcus agalactiae at the third breast.The normal group was injected with the same amount of saline,while the drug group was injected with the same concentration of penicillin.16 S was used to evaluate the diversity of the flora and the temperament was used to determine the expression of metabolites,Western blotting was used to determine the protein expression of interferon(IFN-α),interleukin-4(IL-4) and inducible nitric oxide synthase(i NOS),and Real-time PCR is used to detect the expression of TNF-α mRNA.Results Compared with the normal group,the number of viable clone colony units(CFU) in the milk of the model group and the relative abundance of Firmicutes increased significantly,and the relative abundances of Bacteroides,Actinomycetes,and Proteobacteria decreased significantly(P < 0.05); Compared with the model group,the change trends of the above indicators in the drug group were all opposite to the above,and the differences were statistically significant(P < 0.05).The four most important metabolic pathways associated with Streptococcus agalactiae mastitis were galactose metabolism,starch sucrose metabolism,pentose and glucuronate mutual conversion,and tricarboxylic acid(TCA) cycle.Compared with the blank control group(1.00 ± 0.03,1.00 ± 0.02,1.00 ± 0.01),the expression of IFN-α,IL-4 and i NOS protein in the milk of the model group(4.46 ± 0.20,4.87 ± 0.57,3.31 ± 0.22)increased significantly.The expression of each protein in the milk of mice in the penicillin administration group(2.94 ± 0.37,2.87 ± 0.29,2.34± 0.10) was significantly lower than that of the model group(P < 0.05),and was closer to the normal group.Compared with the blank control group(1.00 ± 0.01),the TNF-α mRNA expression in the model group(0.13 ± 0.04) was significantly reduced,and the TNF-α mRNA expression in the penicillin-administered group(0.66 ± 0.07) recovered significantly compared with the model group,which was slightly lower than the normal control group.Conclusion The expression of inflammatory factors in breast tissues of Streptococcus agalactiae mastitis during lactation is closely related to milk microorganisms and metabolites,which affects the milk quality of dairy cows.更多还原