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研究不同免疫状态下耐甲氧西林金黄色葡萄球菌(methicillin resistant Staphylococcus aureus,MRSA)毒力因子的表达情况,为结合宿主免疫状态对金黄色葡萄球菌毒力因子基因表达的影响提供新的思路和依据。将SPF清洁级Wistar大鼠随机分成对照组和模型组,每组12只,雌雄各半。以40 mg/kg的环磷酰胺连续肌肉注射3 d,造成大鼠免疫抑制,在此基础上通过滴鼻方式滴入小剂量高浓度的新鲜MRSA菌悬液,连续滴注3 d,造成免疫抑制MRSA定植模型。对照组将正常大鼠直接鼻腔滴入小剂量高浓度的新鲜MRSA菌悬液。细菌定植一定时间后取大鼠鼻粘膜组织,运用实时荧光定量PCR技术分析不同免疫状态下大鼠鼻粘膜组织中分离的MRSA毒力因子在基因水平上的表达差异。这表明与肌注环磷酰胺前比较,肌注环磷酰胺后3 d、7 d时,大鼠血液学检查可见外周血WBC、PLT数量迅速下降,同时RBC、HGB亦明显降低(p<0.01);外周血中包括CD4~+T淋巴细胞和CD8~+T淋巴细胞比例以及B淋巴细胞、NK细胞的比例均明显降低(p<0.01);外周血血清中免疫球蛋白Ig G、Ig M、Ig A的含量明显下降(p<0.01);不同免疫状态下细菌毒力因子基因相对表达含量表明,定植在模型组大鼠鼻粘膜组织中的MRSA毒力因子hlα和spa的表达水平要高于对照组,p<0.05,其中目的基因spa的表达水平显著升高(p<0.01)。说明当宿主处于免疫抑制状态时,可能通过促进金黄色葡萄球菌毒力因子的表达增强金黄色葡萄球菌的致病力。
To investigate the expression of virulence factors of methicillin-resistant Staphylococcus aureus (MRSA) under different immunocompetent conditions, and to provide new ideas for the effect of host immune status on the expression of virulence factor genes of Staphylococcus aureus. in accordance with. The SPF clean Wistar rats were randomly divided into control group and model group, with 12 mice in each group. Continuous intramuscular injection of cyclophosphamide at 40 mg / kg for 3 days resulted in immunosuppression in rats. On the basis of this, intranasal instillation of low dose and high concentration of fresh MRSA suspension was carried out for 3 days, resulting in immunization Inhibit MRSA colonization model. In the control group, the nasal cavity of normal rats was instilled with a suspension of fresh MRSA bacteria at a low dose and high concentration. Bacterial colonization after a certain period of time to take nasal mucosa of rats, the use of real-time fluorescence quantitative PCR analysis of different immune status of nasal mucosa tissue isolated MRSA virulence factor at the gene level differences. This indicates that compared with the intramuscular injection of cyclophosphamide, the number of PLT and WBC in peripheral blood decreased rapidly and the RBC and HGB were significantly decreased (P <0.01) at the 3rd and 7th day after intramuscular injection of cyclophosphamide ). The proportion of CD4 ~ + T lymphocytes and CD8 ~ + T lymphocytes and the ratio of B lymphocytes and NK cells in peripheral blood were significantly decreased (p <0.01). The contents of immunoglobulin Ig G, Ig M in peripheral blood , Ig A decreased significantly (p <0.01). The relative expression of bacterial virulence factors in different immune states showed that the MRSA virulence factor hlα and spa expression in nasal mucosa of rats in model group were higher In the control group, p <0.05, in which the expression level of the target gene spa was significantly increased (p <0.01). It indicates that the pathogenicity of Staphylococcus aureus may be enhanced by promoting the expression of Staphylococcus aureus virulence factors when the host is immunosuppressed.