Guinea pig genital tract lipidome reveals in vivo and in vitro regulation of phosphatidylcholine 16:0/18:1 and contribution to Chlamydia trachomatis serovar D infectivity

Abstract

Introduction: Chlamydia trachomatis (Ct), is the leading cause of sexually transmitted infections worldwide. Host transcriptomic- or proteomic profiling studies have identified key molecules involved in establishment of Ct infection or the generation of anti Ct-immunity. However, the contribution of the host metabolome is not known.

Objectives: The objective of this study was to determine the contribution of host metabolites in genital Ct infection.

Methods: We used high-performance liquid chromatography-mass spectrometry, and mapped lipid profiles in genital swabs obtained from female guinea pigs at days 3, 9, 15, 30 and 65 post Ct serovar D intravaginal infection.

Results: Across all time points assessed, 13 distinct lipid species including choline, ethanolamine and glycerol were detected. Amongst these metabolites, phosphatidylcholine (PC) was the predominant phospholipid detected from animals actively shedding bacteria i.e., at 3, 9, and 15 days post infection. However, at days 30 and 65 when the animals had cleared the infection, PC was observed to be decreased compared to previous time points. Mass spectrometry analyses of PC produced in guinea pigs (in vivo) and 104C1 guinea pig cell line (in vitro) revealed distinct PC species following Ct D infection. Amongst these, PC 16:0/18:1 was significantly upregulated following Ct D infection (p < 0.05, >twofold change) in vivo and in vitro infection models investigated in this report. Exogenous addition of PC 16:0/18:1 resulted in significant increase in Ct D in Hela 229 cells.

Conclusion: This study demonstrates a role for host metabolite, PC 16:0/18:1 in regulating genital Ct infection in vivo and in vitro.

Keywords: Chlamydia trachomatis serovar D; Guinea pig; Intravaginal infection; Lipids; Phosphatidylcholine.

Fig. 1

Establishment of Chlamydia trachomatis serovar D infection in guinea pigs. Guinea pigs (n = 5) were challenged i.vag. with 10⁵ IFU Ct D. a Chlamydial shedding was monitored every fourth day post infection until day 40 and presented as mean ± SD for each group at each time point. Number of guinea pigs shedding Ct D after genital challenge is indicated in parenthesis. The stages of infection are denoted as early, intermediate and late. b Histopathological lesions in genital tracts at day 65 post chlamydial infection. The genital tract of each guinea pig was removed sectioned, H&E stained, and analyzed microscopically. Histopathological injury in the oviducts was scored by two distinct morphological parameters (inflammation cell infiltration and congestion). Significant increase between Ct D infected and mock infected guinea pigs are indicated as calculated using student’s t test in data of two replicate studies

Fig. 2

Distribution of lipids in Ct D infected guinea pigs. Lipidome profiles of genital swabs collected at early (day 3), intermediate (days 9 and 15) and late (days 30 and 65) time points post Ct D infection were determined. Using HPLC–MS/MS, the most abundantly detected lipids were annotated according to the PRISM Data Analysis system. Pie chart representation of Ct D infected lipids to have >twofold abundance in Ct D infected compared to mock infected guinea pigs. The blow-out section (black, section 7) represents the phosphatidyl choline which were observed to be most abundant during infection. Significant increase between Ct D infected and mock infected guinea pigs are indicated as calculated using student’s t test in data from 1 of 2 replicates as mean ± SD

Fig. 3

Role of phosphatidyl choline 16:0/18:1 in Ct D infection. a Venn diagram representation of PC species with >twofold increase in Ct D infection compared to mock infection. Numbers in each circle denote total number of PC in vivo (guinea pig swabs) and in vitro (104C1 guinea pig cell line). Phosphatidyl choline 16:0/18:1 was observed to be statistically significant and >twofold in both groups and indicated in the overlapping area. b Relative fold change of PC 16:0/18:1 in guinea pigs (in vivo) and 104C1 (in vitro) and total abundance. c Increase in Ct D IFUs in Hela (human cell line) and 104C1 cells following exogenous addition of PC 16:0/18:1 encapsulated liposomes. Significant increase between Ct D infected and mock infected cells are indicated as calculated using student’s t test in data from 1 of 3 replicates as mean ± SD