Waddlia chondrophila Infects and Multiplies in Ovine Trophoblast Cells Stimulating an Inflammatory Immune Response

Abstract

Background Waddlia chondrophila (W. chondrophila) is an emerging abortifacient organism which has been identified in the placentae of humans and cattle. The organism is a member of the order Chlamydiales, and shares many similarities at the genome level and in growth studies with other well-characterised zoonotic chlamydial abortifacients, such as Chlamydia abortus (C. abortus). This study investigates the growth of the organism and its effects upon pro-inflammatory cytokine expression in a ruminant placental cell line which we have previously utilised in a model of C. abortus pathogenicity.Methodology/Principal FindingsUsing qPCR, fluorescent immunocytochemistry and electron microscopy, we characterised the infection and growth of W. chondrophila within the ovine trophoblast AH-1 cell line. Inclusions were visible from 6 h post-infection (p.i.) and exponential growth of the organism could be observed over a 60 h time-course, with significant levels of host cell lysis being observed only after 36 h p.i. Expression of CXCL8, TNF-α, IL-1α and IL-1β were determined 24 h p.i. A statistically significant response in the expression of CXCL8, TNF-α and IL-1β could be observed following active infection with W. chondrophila. However a significant increase in IL-1β expression was also observed following the exposure of cells to UV-killed organisms, indicating the stimulation of multiple innate recognition pathways.Conclusions/Significance W. chondrophila infects and grows in the ruminant trophoblast AH-1 cell line exhibiting a complete chlamydial replicative cycle. Infection of the trophoblasts resulted in the expression of pro-inflammatory cytokines in a dose-dependent manner similar to that observed with C. abortus in previous studies, suggesting similarities in the pathogenesis of infection between the two organisms.

Experimental challenge of pregnant cattle with the putative abortifacient Waddlia chondrophila

“…This is similar to experimental infection of cattle with C. abortus in which an accumulation of yellow-brown exudate between the uterine and chorionic surfaces was observed12, and in vitro evidence demonstrating a similar pro-inflammatory response to infection with either organism in ovine trophoblasts in vitro 1314. However in the current study, the presence of the W. chondrophila and the associated gross lesions were restricted to the surface of the placenta and little pathology or inflammation was observed within the deeper structures of this organ.…”

“…Waddlia chondrophila strain ATCC VR-1470 was grown at 37 °C in HEp2 (ECACC, Salisbury, UK) cells cultured in GIBCO ® GlutaMAX Iscove’s Modified Dulbecco’s Medium (IMDM) (Thermo Fisher Scientific, Renfrew, UK) as previously described14. Media was supplemented with 2% heat inactivated fetal calf serum (PAA Laboratories Ltd, Yeovil, Somerset, UK) and 1 μg/ml cycloheximide (Sigma-Aldrich, Poole, UK).…”

“…Media was supplemented with 2% heat inactivated fetal calf serum (PAA Laboratories Ltd, Yeovil, Somerset, UK) and 1 μg/ml cycloheximide (Sigma-Aldrich, Poole, UK). The inoculum was titrated on 8-well chamber slides (BD Falcon, Becton Dickinson, Bedford, UK) and visualised according to a previously published protocol14 using polyclonal antisera raised against W. chondrophila elementary bodies (a gift from Professor Gilbert Greub, University of Lausanne).…”

In Contrast to Chlamydia trachomatis, Waddlia chondrophila Grows in Human Cells without Inhibiting Apoptosis, Fragmenting the Golgi Apparatus, or Diverting Post-Golgi Sphingomyelin Transport

“…chondrophila can grow in human cells (12)(13)(14), and experimental growth was recently also reported in an ovine trophoblast cell line (15). We here infected HeLa human cervical epithelial cells (the standard host cell line for the study of C. trachomatis) with W. chondrophila and tested for the morphology of the vacuole, association with the endolysosomal pathway, cytoskeletal recruitment, inhibition of apoptosis, cleavage of host cell proteins, the ability to compete with C. trachomatis in coinfection experiments, fragmentation of the Golgi apparatus, and transport of sphingomyelin to the Waddlia inclusion.…”

Waddlia: An emerging pathogen and a model organism to study the biology of chlamydiae

“…A model of infection of ovine trophoblasts showed that W. chondrophila, similar to Chlamydia abortus, was able to induce an inflammatory immune response [33]. However, the pathogenesis of W. chondrophila and other members of the Chlamydiaceae family may substantially differ.…”