牛源无乳链球菌分离及其诱导的乳房炎小鼠模型构建

Isolation of Streptococcus agalactiae from dairy cattle and construction of its induced mastitis mouse model

研究旨在分离并鉴定致病性无乳链球菌并构建小鼠乳房炎模型。试验从病牛乳汁中成功分离并鉴定1株无乳链球菌(命名为HZ-032)。HZ-032以10^(4)CFU菌量感染哺乳期小鼠,同时设相同剂量无乳链球菌ATCC 13813菌悬液组、阴性对照(等体积无菌PBS)组及空白组作为对照,观察攻毒后小鼠乳腺组织病变并进行组织细菌载量的测定;制作乳腺组织石蜡切片,经HE染色镜下观察组织病理变化;采用荧光定量PCR测定乳腺组织中IL-1α、IL-6、IL-10、TNF-α4种炎性因子相对表达量。结果表明,感染组小鼠乳腺组织可分离出大量HZ-032菌株;组织病理切片显示,感染后小鼠乳腺组织较阴性对照和空白组明显观察到乳腺上皮细胞溶解,腺泡内有大量中性粒细胞浸润;荧光定量PCR结果显示ATCC 13813和HZ-032处理组中IL-1α、IL-10、TNF-α表达量均显著升高(P<0.05),IL-6在HZ-032处理组显著升高(P<0.05)。本研究从患病奶牛乳样中分离1株致病性无乳链球菌HZ-032,并使用该菌成功构建小鼠乳房炎模型,为进一步研究无乳链球菌致乳房炎的相关致病机制及防治措施奠定了基础。

This study was to isolate and identify pathogenic Streptococcus agalactiae and to establish a mouse mastitis model. The milk samples were collected from cows that might have mastitis, and a strain of the pathogenic Streptococcus agalactiae(named HZ-032) was isolated and identified from the samples. 10^(4) CFU of HZ-032 and ATCC 13813 were injected into the fourth pair of mammary glands of lactating mice, and sterile PBS with the same dose was injected into the negative control group, to observe the pathology of breast tissue and bacterial load in mice after infection. The tissue sections were observed microscopically after HE staining. The expression levels of IL-1α, IL-6, IL-10, and TNF-α were determined by quantitative real-time PCR. The results showed that a large number of HZ-032 strains could be isolated from the mammary gland tissue of the infected mice. Pathological sections showed that the mammary epithelial cells of the infected mice were lysis and the breast tissues were infiltrated with a large number of neutrophils, the tissue morphology of the control group was normal. Quantitative real-time PCR showed that the expressions of IL-1α, IL-10, and TNF-α in the ATCC13813 and HZ-032 infected groups were significantly increased(P<0.05), while IL-6 was significantly increased in the HZ-032 infected group(P<0.05). In conclusion, a pathogenic Streptococcus agalactiae HZ-032 was successfully isolated from milk samples of cows suffering from mastitis, and a mouse mastitis model was successfully constructed with the isolates, which laid a foundation for further research on the pathogenic mechanism and prevention measures of mastitis caused by Streptococcus agalactiae.