Comparison of the vaginal environment in rhesus and cynomolgus macaques pre- and post-lactobacillus colonization

Abstract

Background: Rhesus and cynomologus macaques are valuable animal models for the study of human immunodeficiency virus (HIV) prevention strategies. However, for such studies focused on the vaginal route of infection, differences in vaginal environment may have deterministic impact on the outcome of such prevention, providing the rationale for this study.

Methods: We tested the vaginal environment of rhesus and cynomolgus macaques longitudinally to characterize the normal microflora based on Nugent scores and pH. This evaluation was extended after colonization of the vaginal space with Lactobacilli in an effort to recreate NHP models representing the healthy human vaginal environment.

Results and conclusion: Nugent scores and pH differed significantly between species, although data from both species were suggestive of stable bacterial vaginosis. Colonization with Lactobacilli was successful in both species leading to lower Nugent score and pH, although rhesus macaques appeared better able to sustain Lactobacillus spp over time.

Keywords: lactobacillus; macaque; microbiota; vagina.

Figure 1

(A) Comparison of Nugent scores between cynomolgus and rhesus macaques. Rhesus macaque Nugent Scores were collected during the reproductive period (Oct.–Dec.). (B) Comparison of vaginal pH between cynomolgus and rhesus macaques. Statistical values are presented as **p=0.0072; ***p<0.0001

Figure 2

Nugent Scores (A) and vaginal pH (B) of cynomolgus macaques before and after weekly inoculation of 1~2×10¹⁰ CFU of L. rhamn without pre-conditioning. Statistical values are presented as *p=0.018.

Figure 3

Nugent Scores (A) and vaginal pH (B) of 5 cynomolgus macaques before and after weekly inoculation of 1~2×10¹⁰ CFU of L. rhamn preceded by a 10ml 1% lactic acid flush immediately before L. rhamn. inoculation once a week. No statistically significant differences were observed.

Figure 4

Nugent Scores (A) and vaginal pH (B) of 5 cynomolgus macaques before and after inoculation of 1~2×10¹⁰ CFU of L. rhamn preceded by the vaginal administration of 2 mls of erythromycin solution (40mg/ml) 24 hours before a 10ml 1% lactic acid flush followed by inoculation of L. rhamn. Each animal received 2 such administrations (on week 0 and 1) and were monitored weekly for up for 12 weeks. Statistical values are presented as * p<0.05; ** p<0.01; ***p<0.001; ****p<0.0001.

Figure 5

Nugent score and vaginal pH of rhesus macaques before and after L rhamn. inoculation following erythromycin administration and 1% lactic acid flushing. (A) Nugent scores; after a single L rhamn inoculation, Nugent scores sharply decreased and remained low for 7 weeks and beyond (up to week 42, data not shown).(B) Vaginal pH after a single L. Rhamn. inoculation. Statistical values are presented as * p<0.05; ** p<0.01; ***p<0.001; ****p<0.0001

Figure 6

Relative abundance of Gardnerella, curved Gram⁻ bacilli and Lactobacilli in the vaginal environment of CMs (A) and RMs (B) before and after colonization with L Rhamn. The data are representative of 5 CMs and 20 RMs. The values shown represent Nugent scoring values for Gardnerella, curved Gram⁻ bacilli and inverse scoring for Lactobacilli (1–4).