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The impact of cervical cytobrush sampling on cervico-vaginal immune parameters and microbiota relevant to HIV susceptibility.

ABSTRACT: The immunology and microbiota of the female genital tract (FGT) are key determinants of HIV susceptibility. Cervical cytobrush sampling is a relatively non-invasive method permitting the longitudinal assessment of endocervical immune cells, but effects on FGT immunology are unknown. Blood, cervico-vaginal secretions and cervical cytobrushes were collected from sexually transmitted infection (STI)-free women at baseline and after either 6?hours or 48?hours. Endocervical immune cell subsets were assessed by flow cytometry, and pro-inflammatory cytokines by multiplex ELISA. The density of Lactobacillus species and key bacterial vaginosis-associated bacterial taxa were determined by qPCR. Paired changes were assessed before and after cytobrush sampling. After 6?hours there were significant increases in CD4?+?T cell, antigen presenting cell (APC) and neutrophil numbers; APC elevations persisted at 48?hours, while neutrophil and CD4?+?T cell numbers returned to baseline. In addition, pro-inflammatory cytokine levels were increased at 6?hours and returned to baseline by 48?hours. No significant changes were observed in the absolute abundance of Lactobacillus species or BV-associated bacteria at either time point. Overall, cytobrush sampling altered genital immune parameters at 6?hours, but only APC number increases persisted at 48?hours. This should be considered in longitudinal analyses of FGT immunology.

SUBMITTER: Mohammadi A

PROVIDER: S-EPMC7244754 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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The impact of cervical cytobrush sampling on cervico-vaginal immune parameters and microbiota relevant to HIV susceptibility.

Mohammadi A A Bagherichimeh S S Perry M C MC Fazel A A Tevlin E E Huibner S S Tharao W W Coburn B B Kaul R R

Scientific reports 20200522 1

The immunology and microbiota of the female genital tract (FGT) are key determinants of HIV susceptibility. Cervical cytobrush sampling is a relatively non-invasive method permitting the longitudinal assessment of endocervical immune cells, but effects on FGT immunology are unknown. Blood, cervico-vaginal secretions and cervical cytobrushes were collected from sexually transmitted infection (STI)-free women at baseline and after either 6 hours or 48 hours. Endocervical immune cell subsets were a ...[more]

PMID: 32444843

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Project description:BackgroundMenstrual cups (MCs) are increasingly used to collect cervicovaginal secretions to characterise vaginal mucosal immunology, in conjunction with high vaginal swabs (HVS) for metataxonomics, particularly in HIV transmission studies. We hypothesised that both methods of collecting bacterial biomass are equivalent for 16S rRNA gene sequencing.Material and methodsCervicovaginal fluid (CVF) samples from 16 pregnant women with HIV-1 (PWWH) were included to represent the major vaginal bacterial community state types (CST I-V). Women underwent sampling during the second trimester by liquid amies HVS followed by a MC (Soft disc™) and samples were stored at -80°C. Bacterial cell pellets obtained from swab elution and MC (500 µL, 1 in 10 dilution) were resuspended in 120 µL PBS for DNA extraction. Bacterial 16S rRNA gene sequencing was performed using V1-V2 primers and were analysed using MOTHUR. Paired total DNA, bacterial load, amplicon read counts, diversity matrices and bacterial taxa were compared by sampling method using MicrobiomeAnalyst, SPSS and R.ResultsThe total DNA eluted from one aliquot of diluted CVF from an MC was similar to that of a HVS (993ng and 609ng, p=0.18); the mean bacterial loads were also comparable for both methods (MC: 8.0 log10 16S rRNA gene copies versus HVS: 7.9 log10 16S rRNA gene copies, p=0.27). The mean number of sequence reads generated from MC samples was lower than from HVS (MC: 12730; HVS:14830, p=0.05). The α-diversity metrices were similar for both techniques; MC Species Observed: 41 (range 12-96) versus HVS: 47 (range 16-96), p=0.15; MC Inverse Simpson Index: 1.98 (range 1.0-4.0) versus HVS: 0.48 (range 1.0-4.4), p=0.22). The three most abundant species observed were: Lactobacillus iners, Lactobacillus crispatus and Gardnerella vaginalis. Hierarchical clustering of relative abundance data showed that samples obtained using different techniques in an individual clustered in the same CST group.ConclusionThese data demonstrate that despite sampling slightly different areas of the lower genital tract, there was no difference in bacterial load or composition between methods. Both are suitable for characterisation of vaginal microbiota in PWWH. The MC offers advantages, including a higher volume of sample available for DNA extraction and complimentary assays.

Dataset Information

The impact of cervical cytobrush sampling on cervico-vaginal immune parameters and microbiota relevant to HIV susceptibility.

Publications

The impact of cervical cytobrush sampling on cervico-vaginal immune parameters and microbiota relevant to HIV susceptibility.

Similar Datasets

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