Objective A rat model of vaginitis was established to provide a model for developing and evaluating therapeutic drugs for vaginitis. Methods Gardnerella vaginalis, Streptococcus albus and Group B Streptococcus were isolated and purified from clinical vaginal samples. Twenty-six female rats were treated with 20 mg / kg estradiol benzoate and 10 μg / kg streptomycin in the morning, and mixed strains were inoculated into the vagina in the afternoon. Ten female rats serving as the control group were subcutaneously administered 20 mg / kg estradiol benzoate once daily for 3 days. Mixed bacterial infections in the vaginas of the rats were observed by collecting vaginal discharge and lavage fluid, then the rats were grouped according to body weight and strain load. Twenty rats were randomly divided into either the model group (54 mg / day compound metronidazole vaginal suppository once daily for 5 days) or the drug validation group, with 10 animals per group. The control and model groups were injected vaginally with 0. 9% sodium chloride. After the last drug administration, the bacterial strain loads were detected via vaginal lavage, and the vaginal smears, cleanliness and pH were examined from the vaginal discharge. The vaginal tissue was performed to observe the histomorphological changes by HE staining. Results In the model group, the vaginal walls became thickened, the stratified squamous epithelial cells became denatured and necrotic, and the lamina propria became engorged and edematous. Neutrophils were the primary cells, and lymphocyte and monocyte infiltration occurred, with scattered punctiform hemorrhaging, obvious fibroblast proliferation in the mastoid region, and slight reticular fiber proliferation. The mixed bacterial numbers, pH values and cleanliness scores were significantly higher in the model group than in the normal group (P < 0. 01), and the negative conversion rate was significantly reduced ( P < 0. 01). A compound metronidazole vaginal suppository significantly reduced the numbers of mixed bacteria, pH values and cleanliness scores; increased the negative rate for vaginal bacteria and significantly attenuated the degree of the vaginal lesions. Conclusions After pretreatment with estrogen and streptomycin sulfate injections, a rat model of mixed vaginitis was successfully established by inoculating mixed bacteria, thus providing an effective animal model for drug screening and evaluation of female reproductive system.