Characterization of the Vaginal DNA Virome in Health and Dysbiosis

Rasmus Riemer Jakobsen, Thor Haahr, Peter Humaidan, Jørgen Skov Jensen, Witold Piotr Kot, Josue Leonardo Castro-Mejia, Ling Deng, Thomas Dyrmann Leser, Dennis Sandris Nielsen, Rasmus Riemer Jakobsen, Thor Haahr, Peter Humaidan, Jørgen Skov Jensen, Witold Piotr Kot, Josue Leonardo Castro-Mejia, Ling Deng, Thomas Dyrmann Leser, Dennis Sandris Nielsen

Abstract

Bacterial vaginosis (BV) is characterized by a reduction in Lactobacillus (L.) spp. abundance and increased abundance of facultative anaerobes, such as Gardnerella spp. BV aetiology is not fully understood; however, bacteriophages could play a pivotal role in the perturbation of the vaginal bacterial community. We investigated the vaginal viral community, including bacteriophages and the association to the bacterial community and BV-status. Vaginal samples from 48 patients undergoing IVF treatment for non-female factor infertility were subjected to metagenomic sequencing of purified virus-like particles. The vaginal viral community was characterized and correlated with the BV-status by Nugent score, bacterial community, structure, and the presence of key vaginal bacterial species. The majority of identified vaginal viruses belonged to the class of double-stranded DNA bacteriophages, with eukaryotic viruses constituting 4% of the total reads. Clear links between the viral community composition and BV (q = 0.006, R = 0.26) as well as the presence of L. crispatus (q = 0.001, R = 0.43), L. iners, Gardnerella spp., and Atopobium vaginae were found (q < 0.002, R > 0.15). The eukaryotic viral community also correlated with BV-status (q = 0.018, R = 0.20). In conclusion, the vaginal virome was clearly linked with bacterial community structure and BV-status.

Trial registration: ClinicalTrials.gov NCT02042352.

Keywords: bacterial vaginosis; bacteriophages; dysbiosis; vaginal microbiome; vaginal virome.

Conflict of interest statement

T.H. has received honoraria for lectures from Ferring, IBSA, Besins, and Merck. P.H. received unrestricted research grants from MSD, Merck, and Ferring as well as honoraria for lectures from MSD, Merck, Gedeon-Richter, Theramex, and IBSA. J.S.J. has received speaker’s fee from Hologic, B.D. and Cepheid and serves on scientific advisory boards of Roche Molecular Systems, Abbott Molecular, and Cepheid. P.H., T.H. and J.S.J. received a research grant from Osel inc. which produces LACTIN-V, a live biotherapeutic product with Lb. crispatus. P.H. and T.H. are listed as inventors in an international patent application (PCT/US2018/040882) involving “Use of vaginal lactobacilli for improving the success rate of in vitro fertilization”.

Figures

Figure 1
Figure 1
Composition of the vaginal virome by bacterial community status. Viral community composition by relative abundance, grouped by bacterial vaginosis (BV), abnormal vaginal microbiota (AVM), and the community state type (CST) of the sample bacterial community. Taxonomy based on viral database sequence match. Taxonomic entities are ordered top to bottom from most to least abundant.
Figure 2
Figure 2
The vaginal viral community was significantly correlated with the BV-status. (A) Vaginal virome composition (Bray Curtis dissimilarity metric) by BV-status and (B) Lactobacillus crispatus presence/absence (determined by qPCR). (C) The relative abundance of WISH host genus predictions of vOTUs by BV status. The significance was calculated using the Kruskall–Wallis Test (* p < 0.05, ** p < 10−3). (D) A scatter plot showing the Shannon diversity against the percentage of raw reads mapping to integrase genes by sample. Significance levels were calculated using the Pearson correlation.
Figure 3
Figure 3
Abundance of phages with beneficial hosts correlated negatively with an abundance of pathogenic bacteria and vice versa. The clustered image map (CIM) of regularized canonical correlation analysis (rCCA) between the relative abundances of bacterial and viral operational taxonomic units (vOTUs). The colour grade shows the strength of correlation between individual bacterial and viral OTUs. Viral clusters were summarized based on their bacterial host genus as predicted by WISH as only a minority had matches in the viral databases. Bacterial OTUs with several entries had distinct 16S sequences and were possibly different strains. Correlations above 0.7 are shown. CV-score = 0.82.

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