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bioRxiv · 10.1101/2025.11.24.689971

Comparative transcriptomic profiling of human conjunctival epithelial cells and macrophages in response to Chlamydia trachomatis genovars A and B in early- and mid-infection cycles

Ghasemian, E.·Holland, M. J.
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Abstract

Chlamydia trachomatis (Ct), an obligate intracellular bacterium, is the primary infectious cause of blindness through trachoma. Ct undergoes a unique biphasic developmental cycle between infectious elementary bodies and replicating reticulate bodies, manipulating host cells via secreted effector proteins. Whilst previous studies have characterised host-pathogen interactions including transcriptomes of urogenital Ct genovars E and L2, limited studies exist on ocular Ct genovars. This study examined transcriptomic responses of human conjunctival epithelial (HCjE) cells and PMA-differentiated THP-1 macrophages to infection with ocular Ct strains A/2497 or B/Tunis864 (live or heat-inactivated) at 4 and 24 hours post-infection (hpi). Transcriptomic profiling was performed using Lexogen QuantSeq 3 mRNA-Seq, with differential gene expression analysis conducted using DESeq2. Gene Ontology Biological Process and KEGG pathway enrichment analyses were performed using Gene Set Enrichment Analysis via clusterProfiler to identify strain-specific and cell type-specific transcriptional signatures. HCjE cells exhibited progressive transcriptional activation, with differentially expressed genes (DEGs) increasing from 4 to 24 hpi (144 to 259; P = 0.0458), whilst THP-1 macrophages showed temporal attenuation (391 to 154; P < 0.0001). B/Tunis864 consistently elicited higher responses than A/2497 in HCjE cells at both time points (4 hpi: 152 vs. 54, P = 0.0003; 24 hpi: 259 vs. 83, P < 0.0001). Conversely, THP-1 macrophages showed higher responses to A/2497 than B/Tunis864 at both time points (4 hpi: 599 vs. 376, P < 0.0001; 24 hpi: 166 vs. 114, P = 0.0221). HCjE cells demonstrated markedly higher proportions of strain-specific DEGs and pathways compared to macrophages. B/Tunis864 infection in HCjE cells induced pronounced interferon-stimulated gene signatures, particularly at 24 hpi. This study revealed contrasting temporal patterns: THP-1 macrophages showed peak-then-decline responses, whilst HCjE cells exhibited progressive activation to 24 hpi. HCjE cells demonstrated predominantly strain-specific responses, whereas macrophages deployed strain-invariant programmes. B/Tunis864s enhanced interferon-stimulated gene and inflammatory pathway activation in HCjE cells may suggest molecular hints for genovar B-associated trachoma severity.

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Ghasemian, E., Holland, M. J.. 2025-11-24. Comparative transcriptomic profiling of human conjunctival epithelial cells and macrophages in response to Chlamydia trachomatis genovars A and B in early- and mid-infection cycles. https://doi.org/10.1101/2025.11.24.689971

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