Biology subjects

Brochu, H.

Publications and source records attributed to Brochu, H..

4 recordsLinked to original sources

bioRxiv2025

Development of ferret immune repertoire reference resources and single-cell-based high-throughput profiling assays

Domestic ferrets (Mustela putorius furo) are important for modeling human respiratory diseases. However, ferret B and T cell receptors have not been completely identified or annotated, limiting immune repertoire studies. Here we performed long read transcriptome sequencing of ferret splenocyte and lymph node samples to obtain over 120,000 high-quality full-length immunoglobin (Ig) and T cell receptor (TCR) transcripts. We constructed a complete reference set of the constant regions of ferret Ig and TCR isotypes and chain types. We also systematically annotated germline Ig and TCR variable (V), diversity (D), joining (J), and constant (C) genes on a recent ferret reference genome assembly. We designed new ferret-specific immune repertoire profiling assays by targeting positions in constant regions without allelic diversity across 11 ferret genome assemblies, and experimentally validated them using a commercially compatible single-cell-based platform. These improved resources and assays will enable future studies to fully capture ferret immune repertoire diversity.

genomics↗

bioRxiv2024

Cervicovaginal microbial features predicts Chlamydia trachomatis spread to the upper genital tract of infected women

IntroductionChlamydia trachomatis (CT) infection can lead to pelvic inflammatory disease, infertility and other reproductive sequelae when it ascends to the upper genital tract. Factors including chlamydial burden, co-infection with other sexually-transmitted bacterial pathogens and oral contraceptive use influence risk for upper genital tract spread. Cervicovaginal microbiome composition influences CT susceptibility and we investigated if it contributes to spread by analyzing amplicon sequence variants (ASVs) derived from the V4 region of 16S rRNA genes in vaginal samples collected from women at high risk for CT infection and for whom endometrial infection had been determined. ResultsParticipants were classified as CT negative (CT-, n=77), CT positive at the cervix (Endo-, n=77), or CT positive at both cervix and endometrium (Endo+, n=66). Although we were unable to identify many significant differences between CT infected and uninfected women, differences in abundance of ASVs representing Lactobacillus iners and L. crispatus subspecies but not dominant lactobacilli were detected. Twelve informative ASVs predicted endometrial chlamydial infection (AUC=0.74), with CT ASV abundance emerging as a key predictor. We also observed a positive correlation between levels of cervically secreted cytokines previously associated with CT ascension and abundance of the informative ASVs. ConclusionOur findings suggest that vaginal microbial community members may influence chlamydial spread directly by nutrient limitation and/or disrupting endocervical epithelial integrity and indirectly by modulating pro-inflammatory signaling and/or homeostasis of adaptive immunity. Further investigation of these predictive microbial factors may lead to cervicovaginal microbiome biomarkers useful for identifying women at increased risk for disease.

microbiology↗

bioRxiv2021

Single cell based high-throughput Ig and TCR repertoire sequencing analysis in rhesus macaques

Recent advancements in microfluidics and high-throughput sequencing technologies have enabled recovery of paired heavy- and light-chains of immunoglobulins (Ig) and VDJ- and VJ-chains of T cell receptors (TCR) from thousands of single cells simultaneously in humans and mice. Despite rhesus macaques being one of the most well-studied model organisms for the human adaptive immune response, high-throughput single cell immune repertoire sequencing assays are not yet available due to the complexity of these polyclonal receptors. Here we employed custom primers that capture all known rhesus macaque Ig and TCR isotypes and chains that are fully compatible with a commercial solution for single cell immune repertoire profiling. Using these rhesus specific assays, we sequenced Ig and TCR repertoires in over 60,000 cells from cryopreserved rhesus PBMC, splenocytes, and FACS-sorted B and T cells. We were able to recover every Ig isotype and TCR chain, measure clonal expansion in proliferating T cells, and pair Ig and TCR repertoires with gene expression profiles of the same single cells. Our results establish the ability to perform high-throughput immune repertoire analysis in rhesus macaques at the single cell level.

immunology↗

bioRxiv2019

Systematic profiling of full-length immunoglobulin and T-cell receptor repertoire diversity in rhesus macaque through long read transcriptome sequencing

The diversity of immunoglobulin (Ig) and T-cell receptor (TCR) repertoires is a focal point of immunological studies. Rhesus macaques are key for modeling human immune responses, placing critical importance on the accurate annotation and quantification of their Ig and TCR repertoires. However, due to incomplete reference resources, the coverage and accuracy of the traditional targeted amplification strategies for profiling rhesus Ig and TCR repertoires are largely unknown. Here, using long read sequencing, we sequenced four Indian-origin rhesus macaque tissues and obtained high quality, full-length sequences for over 6,000 unique Ig and TCR transcripts, without the need for sequence assembly. We constructed the first complete reference set for the constant regions of all known isotypes and chain types of rhesus Ig and TCR repertoires. We show that sequence diversity exists across the entire variable regions of rhesus Ig and TCR transcripts. Consequently, existing strategies using targeted amplification of rearranged variable regions comprised of V(D)J gene segments miss a significant fraction (27% to 53% and 42% to 49%) of rhesus Ig/TCR diversity. To overcome these limitations, we designed new rhesus-specific assays that remove the need for primers conventionally targeting variable regions and allow single cell-level Ig and TCR repertoire analysis. Our improved approach will enable future studies to fully capture rhesus Ig and TCR repertoire diversity and is applicable for improving annotations in any model organism.

immunology↗