Topical microbicides that block the sexual transmission of HIV and herpes simplex virus 2 (HSV-2) are desperately needed to reduce the incidence of HIV infections worldwide. Previously we completed phase 3 testing of the carrageenan-based gel Carraguard. Although the trial did not show that Carraguard is effective in preventing HIV transmission during vaginal sex, it did show that Carraguard is safe when used weekly for up to 2 years. Moreover, Carraguard has in vitro activity against human papillomavirus (HPV) and HSV-2 and favorable physical and rheological properties, which makes it a useful vehicle to deliver antiviral agents such as zinc acetate. To that end, we previously reported that a prototype zinc acetate carrageenan gel protects macaques against vaginal challenge with combined simian-human immunodeficiency virus reverse transcriptase (SHIV-RT). Herein, we report the safety and efficacy of a series of zinc acetate and/or carrageenan gels. The gels protected mice (75 to 85% survival; P < 0.001) against high-dose (10^6-PFU) HSV-2 vaginal or rectal challenge. In contrast, zinc acetate formulated in HEC (hydroxyethylcellulose; or the Universal Placebo) failed to protect mice against the high-dose vaginal HSV-2 challenge (similar to aqueous zinc acetate solution and the placebo controls). The gels were found to be effective spreading gels, exhibited limited toxicity in vitro, caused minimal damage to the architecture of the cervicovaginal and rectal mucosae in vivo, and induced no increased susceptibility to HSV-2 infection in a mouse model. Our results provide a strong rationale to further optimize and evaluate the zinc acetate/carrageenan gels for their ability to block the sexual transmission of HIV and HSV-2.