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Abstract: We report the creation of a new low-estrogen murine model of concurrent oral and vaginal C. albicans colonization that resembles human candidal carriage at both mucosal sites. Weekly estrogen administration of 5μg intramuscular and subcutaneously was optimal for enhancement of oral colonization and was essential for vaginal colonization. In BALB/c mice, a number of C. albicans clinical isolates (n =3) colonized both oral and/or vaginal sites, but only strain 529L colonized 100% of mice persistently for over 5weeks. Laboratory strains SC5314 and NCPF 3153 did not colonize the model; however, NCPF 3156 showed vaginal colonization up to week 5. Prior passaging through mice enhanced subsequent colonization of SC5314. Intranasal immunization with a C. albicans virulence antigen (secreted aspartyl proteinase 2) significantly reduced or abolished the fungal burden orally and vaginally by week 2 and 7. Our concurrent model of mucosal colonization reduces the numbers of experimental mice by half, can be used to assess potential vaccine candidates, and permits the detailed analysis of host–fungal interactions during the natural state of Candida colonization. [Copyright &y& Elsevier]