目的：研究角膜胶原交联方法联合那他霉素应用时的体内外抗真菌效果，为临床真菌性角膜炎的治疗提供治疗方法及实验基础。

方法：采用黄曲霉菌、茄病镰刀菌、白色念珠菌三种常见致病真菌，实验组分为交联联合那他霉素组、那他霉素联合核黄素组、那他霉素联合紫外照射组、交联组，并用那他霉素组作为对照。将药物涂到有1.5麦氏浊度的真菌孢子悬液的沙堡弱琼脂培养皿的中心，10min后使用胶原交联仪照射10min，之后28℃培养36h，测量抑菌圈并进行统计学分析。制备兔眼茄病镰刀菌感染模型，按随机数字表法将模型兔随机分为模型对照组、交联治疗组、那他霉素治疗组、交联联合那他霉素治疗组，每组5只； 另取5只正常兔按照角膜胶原交联疗法进行照射，5只正常对照。通过前节照相、角膜刮片、共焦显微镜等观察各组治疗结果，并在治疗结束后采用电镜观察角膜超微结构改变情况。

结果：体外单独应用角膜胶原交联术，对三种真菌均无效； 当交联与那他霉素联合应用时，抗真菌效果均优于对照组，且差异具有统计学意义(P<0.05)； 那他霉素与核黄素、那他霉素与紫外线分别联合应用时，抗菌效果与对照组比较，差异均无统计学意义(P>0.05)。针对兔真菌感染模型，那他霉素治疗组及交联联合那他霉素治疗组病程约14d，交联治疗组病程约21d； 经治疗后，各治疗组均已愈合，各组角膜上皮均无缺损，新生血管较多，角膜内无菌丝； 前节照相结果显示，交联联合那他霉素治疗组治疗结果优于其它各治疗组，瘢痕组织较少，角膜愈合较好，病程相对短。

结论：角膜胶原交联方法联合那他霉素，可促进角膜愈合，缩短病程，增强抗真菌效果，有望为眼科临床真菌性角膜炎的治疗提供一种新的治疗技术。

METHODS: Three common pathogenic fungi(Aspergillus flavus, Fusarium Solani and Candida albicans)were used. The experimental group was divided into cross-linking combined natamycin group, natamycin combined riboflavin group, natamycin combined UVA irradiation group, cross-linking group and natamycin group as the control group. The drug was added to the center of the Sabouraud dextrose agar(SDA)plate coated with liquid with each fungal spores with the same maid turbidity of 1.5. Ten minutes later, it was irradiated with collagen cross-linking instrument for 10min and cultured at 28℃ for 36h, and then the inhibition zone size was measured and analyzed statistically. The rabbit model of Fusarium Solani corneal infection was prepared. The model rabbits were randomly divided into model control group, cross-linking treatment group, natamycin treatment group, cross-linking combined natamycin group, 5 rabbits in each group. And another 5 normal rabbits were taken as control, and five rabbits were irradiated in accordance with corneal collagen cross-linking therapy. The results were observed by anterior segment photography, corneal scraping and confocal microscopy, and the ultra micro structural changes of the corneas were observed by electron microscope after the treatment.

RESULTS: Corneal collagen cross-linking alone had shown no effect on each fungus in vitro. Corneal collagen cross-linking combined with natamycin produced significant anti-fungal effect(P<0.05). However, the anti-fungal effect of natamycin combined riboflavin group and natamycin combined ultraviolet light group showed no statistical difference(P>0.05)comparing with the control group. For the model of rabbit fungal infection, the course of disease was about 14d in the natamycin group and CXL combined with natamycin group, and it was about 21d in CXL group. After the treatment, all the groups healed. There were no defects in the corneal epithelium, no mycelium in the corneas, except for more corneal neovascularization. The results of the anterior segment photography showed that the treatment effect of the cross-linking combined natamycin group was better than other groups, with fewer scar tissue, better corneal healing and relatively short course of disease.

CONCLUSION: Corneal collagen cross-linking combined with natamycin treatment is able to enhance anti-fungal effect, promote corneal healing, and shorten the course of disease. So it is a promising therapeutic technique for the clinical treatment of fungal keratitis.