目的 系统评价苯酰甲硝唑（BMZ）对抗刚地弓形虫（简称弓形虫）效果和安全性。方法 非洲绿猴肾细胞（Vero细胞）分别加入不同浓度的BMZ（50、100、200、400、500、600、800、1 000 μg/mL），细胞计数试剂盒-8（CCK-8）法检测BMZ对Vero细胞的毒性。Vero细胞接种弓形虫速殖子后分别加入不同浓度的BMZ（50、100、200、400、500、600、800、1 000 μg/mL），CCK-8法检测BMZ对弓形虫的杀伤作用。Vero细胞接种弓形虫速殖子后加入50 μg/mL BMZ，通过吉姆萨染色和扫描电镜观察弓形虫速殖子数量和形态变化。昆明小鼠随机分为低剂量BMZ组（LD-BMZ组）、中剂量BMZ组（MD-BMZ组）、高剂量BMZ组（HD-BMZ组）、磺胺嘧啶钠组（SD-Na组）和阴性对照组（NC组），经腹腔接种1 × 10⁴ 个/mL弓形虫速殖子，4 h后口服灌胃连续给药5 d：LD-BMZ组、MD-BMZ组和HD-BMZ组分别给终浓度为12.5、25和50 mg/kg的BMZ，SD-Na组给终浓度为50 mg/kg的SD-Na，NC组给等体积玉米油。感染后第7天记录小鼠生存率，血细胞计数板计算小鼠腹水荷虫量。收集MD-BMZ组和NC组小鼠的心、肝、脾、肺、肾和脑组织，提取DNA后qPCR反应检测各组织荷虫量，制成病理组织切片后HE染色观察各组织病理学变化。利用GraphPad Prism 9软件进行数据分析并作图，差异显著性分析采用独立样本t检验或单因素方差分析。结果 细胞毒性实验结果显示，BMZ对Vero细胞的CC₅₀为155.67 μg/mL；BMZ浓度为50 μg/mL时细胞存活率为（108.78 ± 1.00）%，100 μg/mL时降至（81.22 ± 3.34）%，将50 μg/mL作为BMZ最大安全浓度。体外抗虫实验结果显示，BMZ可抑制细胞内弓形虫速殖子的增殖，抑制率随浓度升高呈上升趋势；BMZ浓度为50 μg/mL时对弓形虫的抑制率为14.3%。吉姆萨染色结果显示，BMZ组细胞中弓形虫速殖子数量降低。扫描电镜观察结果显示，正常弓形虫速殖子呈典型的梭形或新月形结构，虫体轮廓完整、形态饱满、表面光滑；经BMZ处理后，弓形虫虫体结构完全丧失，轮廓模糊不清，细胞膜完整性被破坏。体内抗虫实验结果显示，感染弓形虫后第7天，LD-BMZ组、MD-BMZ组、HD-BMZ组、SD-Na组和NC组的小鼠存活率分别为2/10、3/10、2/10、2/10和1/10，差异无统计学意义（χ² = 3.19，P > 0.05）；腹水荷虫量分别为（15.00 ± 5.00）× 10⁴、（6.67 ± 2.89）× 10⁴、（6.67 ± 2.89）× 10⁴、（8.33 ± 2.89）× 10⁴和（1 766.67 ± 2.33）× 10⁴个/mL，差异有统计学意义[F(4,10) = 123 320，P < 0.01]。qPCR结果显示，感染后第7天MD-BMZ组小鼠的心、肝、脾、肺、肾和脑中弓形虫相对荷虫量分别为2.32 ± 0.85、2.32 ± 0.76、1.00 ± 0.45、1.00 ± 0.52、4.46 ± 0.98、1.59 ± 0.63，均低于NC组的7.28 ± 1.25、7.11 ± 1.18、3.46 ± 0.95、7.28 ± 1.22、9.37 ± 1.36、8.67 ± 1.29（t = 6.56、6.83、4.68、9.47、5.86、9.86，P < 0.01）。HE染色显示，BMZ治疗可显著减轻脾、肾、肝脏的炎性细胞浸润与肉芽肿病变，且未引起明显药物相关性损伤。结论 BMZ在体内外均具有良好的抗弓形虫活性，中剂量时疗效优于SD-Na且安全性较高。

Objective To systematically evaluate the anti-Toxoplasma gondii activity and safety of benzoyl metronidazole (BMZ). Methods Vero cells were treated with BMZ at different concentrations (50, 100, 200, 400, 500, 600, 800, 1 000 μg/mL), and the cytotoxicity of BMZ to Vero cells was detected by cell counting kit-8 (CCK-8) assay. After inoculation with Toxoplasma tachyzoites, Vero cells were treated with BMZ at the above concentrations, and the killing effect of BMZ on Toxoplasma was determined by CCK-8 assay. Vero cells inoculated with Toxoplasma tachyzoites were treated with 50 μg/mL BMZ, and the quantitative and morphological changes of Toxoplasma tachyzoites were observed via Giemsa staining and scanning electron microscopy. Kunming mice were randomly divided into low-dose BMZ group (LD-BMZ group), medium-dose BMZ group (MD-BMZ group), high-dose BMZ group (HD-BMZ group), sulfadiazine sodium group (SD-Na group) and negative control group (NC group). All mice were intraperitoneally inoculated with 1 × 10⁴ Toxoplasma tachyzoites per mL. Four hours after infection, mice were intragastrically administered continuously for 5 days: the LD-BMZ, MD-BMZ and HD-BMZ groups received BMZ at final doses of 12.5, 25 and 50 mg/kg, respectively, the SD-Na group received SD-Na at a final dose of 50 mg/kg, and the NC group received an equal volume of corn oil. On day 7 post-infection, the survival rates of mice were recorded, and the parasite load in mice ascites was counted using a hemocytometer. The heart, liver, spleen, lung, kidney and brain tissues of mice in the MD-BMZ group and NC group were collected. After DNA extraction, qPCR was performed to detect the parasite load in each tissue, and the tissues were processed into pathological sections for HE staining to observe the histopathological changes. GraphPad Prism 9 software was used for data analysis and plotting, and independent sample t-test or one-way ANOVA was adopted for significance analysis. Results The cytotoxicity assay showed that the CC₅₀ of BMZ to Vero cells was 155.67 μg/mL; the cell viability was (108.78 ± 1.00)% when BMZ concentration was 50 μg/mL, and decreased to (81.22 ± 3.34)% at 100 μg/mL. Therefore, 50 μg/mL was determined as the maximum safe concentration of BMZ. The in vitro anti-parasitic assay showed that BMZ could inhibit the proliferation of intracellular Toxoplasma tachyzoites, and the inhibition rate increased with the elevation of BMZ concentration; the inhibition rate of BMZ against Toxoplasma reached 14.3% at 50 μg/mL. Giemsa staining results showed that the number of Toxoplasma tachyzoites in cells of the BMZ group was reduced. Scanning electron microscopy observation showed that normal Toxoplasma tachyzoites presented typical spindle or crescent structures with complete contours, plump morphology and smooth surface; while after BMZ treatment, the structural integrity of Toxoplasma was completely lost, with blurred contours and destroyed cell membrane. The in vivo anti-parasitic assay showed that on day 7 post Toxoplasma infection, the survival rates of mice in the LD-BMZ, MD-BMZ, HD-BMZ, SD-Na and NC groups were 2/10, 3/10, 2/10, 2/10 and 1/10, respectively, with no statistically significant difference (χ² = 3.19, P > 0.05); the ascites parasite loads were (15.00 ± 5.00) × 10⁴, (6.67 ± 2.89) × 10⁴, (6.67 ± 2.89) × 10⁴, (8.33 ± 2.89) × 10⁴ and (1 766.67 ± 2.33) × 10⁴ parasites per mL respectively, with statistically significant difference [F(4,10) = 123 320, P < 0.01]. The qPCR results showed that on day 7 post infection, the relative Toxoplasma parasite loads in the heart, liver, spleen, lung, kidney and brain of mice in the MD-BMZ group were 2.32 ± 0.85, 2.32 ± 0.76, 1.00 ± 0.45, 1.00 ± 0.52, 4.46 ± 0.98 and 1.59 ± 0.63 respectively, all significantly lower than 7.28 ± 1.25, 7.11 ± 1.18, 3.46 ± 0.95, 7.28 ± 1.22, 9.37 ± 1.36 and 8.67 ± 1.29 in the NC group (t = 6.56, 6.83, 4.68, 9.47, 5.86, 9.86, all P < 0.01). HE staining showed that BMZ treatment significantly reduced the inflammatory cell infiltration and granulomatous lesions in the spleen, kidney and liver, without causing obvious drug-related damage. Conclusion BMZ exhibited excellent anti-Toxoplasma activity both in vivo and in vitro, with superior efficacy to SD-Na and high safety at the medium dose.