Protocols for Vaginal Inoculation and Sample Collection in the Experimental Mouse Model of <em>Candida</em> vaginitis

Abstract

Vulvovaginal candidiasis (VVC), caused by Candida species, is a fungal infection of the lower female genital tract that affects approximately 75% of otherwise healthy women during their reproductive years 18,[32][33][34] . Predisposing factors include antibiotic usage, uncontrolled diabetes and disturbance in reproductive hormone levels due to pregnancy, oral contraceptives or hormone replacement therapies 33,34 . Recurrent VVC (RVVC), defined as three or more episodes per year, affects a separate 5 to 8% of women with no predisposing factors 33 .An experimental mouse model of VVC has been established and used to study the pathogenesis and mucosal host response to Candida 3,4,11,16,17,19,21,25,37 . This model has also been employed to test potential antifungal therapies in vivo 13,24 . The model requires that the animals be maintained in a state of pseudoestrus for optimal Candida colonization/infection 6,14,23 . Under such conditions, inoculated animals will have detectable vaginal fungal burden for weeks to months. Past studies show an extremely high parallel between the animal model and human infection relative to immunological and physiological properties 3,16,21 . Differences, however, include a lack of Candida as normal vaginal flora and a neutral vaginal pH in the mice.Here, we demonstrate a series of key methods in the mouse vaginitis model that include vaginal inoculation, rapid collection of vaginal specimens, assessment of vaginal fungal burden, and tissue preparations for cellular extraction/isolation. This is followed by representative results for constituents of vaginal lavage fluid, fungal burden, and draining lymph node leukocyte yields. With the use of anesthetics, lavage samples can be collected at multiple time points on the same mice for longitudinal evaluation of infection/colonization. Furthermore, this model requires no immunosuppressive agents to initiate infection, allowing immunological studies under defined host conditions. Finally, the model and each technique introduced here could potentially give rise to use of the methodologies to examine other infectious diseases of the lower female genital tract (bacterial, parasitic, viral) and respective local or systemic host defenses. Video LinkThe video component of this article can be found at

Candida albicans Pathogenesis: Fitting within the Host-Microbe Damage Response Framework

“…Interestingly, adaptive cell-mediated or humoral immunity appears to play no role in protection against infection, although certain epitope-specific antibodies characterized as protective antibodies can be used therapeutically against infection (134,135,(150)(151)(152). Another interesting caveat is that while Th17 responses are critical to neutrophil responses at other anatomical sites (oral, bloodstream), the role for Th17 responses in the vaginal neutrophil response is controversial (139,153). The strongest evidence to date has shown that mice deficient in several cytokines encompassing the Th17 axis showed no change in the C. albicans-in-FIG 6 Rat model of Candida-associated denture stomatitis.…”

“…(Reprinted from reference 166. ) duced neutrophil response, suggesting little to no role for the Th17 response in VVC (139).…”

“…For study of VVC, a well-established estrogen-dependent mouse model is available (139). Unlike humans, laboratory rodents do not naturally harbor C. albicans as a commensal; however, the experimental infection model closely parallels the human infection, and findings from the animal model are translatable to the human host (4).…”

Nanostructured lipid system as a strategy to&nbsp;improve the anti-Candida albicans activity of&nbsp;Astronium sp.

“…On days 2, 6, and 10 after the infection, the vaginal load of C. albicans was evaluated through vaginal lavage with 0.1 mL of PBS, and the fungal count was determined by the CFU assay on sabouraud dextrose agar (SDA). 35 Eight days after the treatment period, the animals were subjected to recurrent control of the infection to further identify and to diagnose the infectious states that may eventually emerge after the previously performed treatment. At day 24, the animals were euthanatized by CO 2 chamber intoxication.…”

Immune Response, Detection of IgE and PGE2 during Vaginal Candidiasis in Mice

“…Currently, rats and mice are considered the most used animal as models to study candidiasis (Carrara et al, 2010;Yano and Fidel, 2011). Fidel et al (1993) established the following steps are necessary to the persistent vaginal infection development, which constitutes fungus penetration, tissue invasion, development and proliferation of this fungus in these animals and the estrogen treatment to induce a pseudo-estrus state.…”