汉赛巴尔通体感染SD大鼠实验研究

An experiment of Bartonella henselae infection in Sprague-Dawley rats

目的用汉赛巴尔通体菌株感染SD大鼠,检测大鼠体内菌血症及抗体的动态变化。方法用不同浓度的汉赛巴尔通体菌液经皮下多点注射感染SD大鼠,采集其全血稀释培养,心、肝、脾和肾脏组织匀浆后分离培养,提取疑似汉赛巴尔通体菌落核酸检测,用PCR扩增枸橼酸合酶基因(gltA),进行测序分析,间接免疫荧光抗体测定法测定抗体变化情况。应用SPSS 19.0软件进行统计分析,采用χ^(2)检验和配伍组方差分析,分析不同感染浓度、不同组织间阳性率和不同感染天数的阳性率差异。结果共取样品全血90份、组织82份、血清21份,培养均为阴性。核酸检测全血阳性率为30.56%(22/72),其中第1、3、5天均有阳性样品,最高阳性率为100%(第3天的低剂量组和高剂量组),组织阳性率为22.86%(16/70),其中第3、5天均有阳性,阳性率最高的是肾脏,检测的16份样品中7份为阳性,其次是肝脏,检测的18份样品中5份为阳性,肾脏阳性率持续时间最长为5 d。检测血清18份,抗体阳性6份,最早出现在感染的第7天,持续到第14天实验结束。经配伍组方差分析,不同剂量组间阳性率差异无统计学意义(F=3.243,P=0.082);经Fisher’s确切概率法检验,不同组织样品阳性率差异无统计学意义(χ^(2)=7.655,P=0.057)。经配伍组方差分析,不同感染天数阳性率差异有统计学意义(F=11.770,P=0.001),其中第3天与第1、5、7、10和14天的阳性率差异有统计学意义(均P<0.05)。结论 SD大鼠可短期感染汉赛巴尔通体,感染的高峰值出现在感染后的第3天,抗体至少可持续到感染后第2周。

Objective To infect Sprague-Dawley(SD) rats with Bartonella henselae strains and measure the dynamic changes of bacteremia and antibody in rats. Methods SD rats were infected by subcutaneous multi-point injection of different concentrations of B. henselae suspension. Whole blood was collected and diluted for culture. The heart, liver,spleen, and kidney were homogenized and isolated for culture. The probable B. henselae colony nucleic acids were extracted, and the citrate synthase gene(gltA) was amplified by PCR for sequence analysis. The antibody changes were measured by the indirect immunofluorescence antibody assay. SPSS 19.0 software was used to analyze the differences in the positive rate between different tissues, different suspension concentrations, and different days of infection using the Chi-square test and two-way analysis of variance(ANOVA). Results A total of 90 whole blood samples, 82 tissue samples, and 21 serum samples were collected, all of which were negative for culture. For whole blood samples, the positive rate of the nucleic acid was 30.56%(22/72), and positive samples were found on days 1, 3, and 5, with the highest positive rate of 100% occurring in the low-dose group and high-dose group on day 3. For tissue samples, the positive rate of the nucleic acid was 22.86%(16/70), and positive samples were found on days 3 and 5;the renal tissues showed the highest positive rate(7/16) and the most persistent positive status(lasting 5 days), and the liver tissues had the second highest positive rate(5/18). The antibody was present in 6 of 18 serum samples detected, beginning on the 7 th day since infection till the 14 th day when the experiment ended. No significant difference was observed in the positive rate between different dose groups(F=3.243, P=0.082, by two-way ANOVA) or between different types of samples(χ^(2)=7.655, P=0.057, by the Fisher’s exact test). The positive rate showed a significant difference between different times of infection(F=11.770, P=0.001, by two-way ANOVA), and specifically between the 3 rd day and the 1 st, 5 th, 7 th, 10 th, or14 th day(all P<0.05). Conclusion B. henselae can infect SD rats for a short time, with nucleic acid positive rate peaking on the 3 rd day of infection. The antibody can last at least two weeks since infection.