Conservation of extrusion as an exit mechanism forChlamydia

Abstract

Chlamydiae exit via membrane-encased extrusion or through lysis of the host cell. Extrusions are novel, pathogen-containing structures that confer infectious advantages to Chlamydia, and are hypothesized to promote cell-to-cell spread, dissemination to distant tissues and facilitate immune evasion. The extrusion phenomenon has been characterized for several Chlamydia trachomatis serovars, but a thorough investigation of extrusion for additional clinically relevant C. trachomatis strains and Chlamydia species has yet to be performed. The key parameters investigated in this study were: (i) the conservation of extrusion across the Chlamydia genus, (ii) the functional requirement for candidate Chlamydia genes in extrusion formation i.e. IncA and CT228 and (iii) extrusion-mediated uptake, and consequent survival of Chlamydia inside macrophages. Inclusion morphology was characterized by live fluorescence microscopy, using an inverted GFP strategy, at early and mid-stages of infection. Enriched extrusions were used to infect bone marrow-derived macrophages, and bacterial viability was measured following macrophage engulfment. Our results demonstrate that extrusion is highly conserved across chlamydiae, including ocular, STD and LGV biovars and divergent Chlamydia species. Consequently, this exit mechanism for Chlamydia may fulfill common advantages important for pathogenesis.

Comparison of Murine Cervicovaginal Infection by Chlamydial Strains: Identification of Extrusions Shed In vivo

“…Consistent with SEM and confocal microscopy data in previous in vitro reports of extrusions (Hybiske and Stephens, 2007 , 2008 ; Lutter et al, 2013 ; Zuck et al, 2016a , b ), the extrusions we collected were filled with Chlamydia , enclosed in the host plasma membrane, and devoid of host nuclei. In vivo host cell exit of all three tested chlamydial strains via the extrusion pathway was hypothesized based on in vitro data demonstrating that all chlamydial species tested to date produce extrusions (Hybiske and Stephens, 2007 , 2008 ; Lutter et al, 2013 ; Zuck et al, 2016a , b ), however documentation was technically challenging in vivo . Extrusions are produced readily and in high numbers by monolayers in cell culture (Lutter et al, 2013 ; Zuck et al, 2016b ); on the other hand, visualization of extrusions produced and shed from the murine cervicovaginal tract was less abundant.…”

“…In vivo host cell exit of all three tested chlamydial strains via the extrusion pathway was hypothesized based on in vitro data demonstrating that all chlamydial species tested to date produce extrusions (Hybiske and Stephens, 2007 , 2008 ; Lutter et al, 2013 ; Zuck et al, 2016a , b ), however documentation was technically challenging in vivo . Extrusions are produced readily and in high numbers by monolayers in cell culture (Lutter et al, 2013 ; Zuck et al, 2016b ); on the other hand, visualization of extrusions produced and shed from the murine cervicovaginal tract was less abundant. The disparity in quantity of extrusions shed in vivo vs. in vitro was expected due to both biological and technical reasons.…”

“…Despite biological and technical challenges of harvesting significant numbers of extrusions in vivo , the fact that they are recovered from the murine vaginal mucosa reveals a previously unappreciated mechanism that may participate in chlamydial pathogenesis in vivo . Recent in vitro studies demonstrate significant advantages for Chlamydia that are packaged into extrusions, including enhanced extracellular survival (Zuck et al, 2016b ) and the ability to avoid the immune system by surviving inside macrophages (Zuck et al, 2016a ). Confirming these advantages in an animal model would greatly advance our understanding of chlamydial strategies for survival.…”

“…Upon the host cell swelling in response to the growing numbers of Chlamydia , the Chlamydia exit the cell by one of two mechanisms: lysis or extrusion (Todd and Caldwell, 1985 ; Scidmore et al, 1996 ; Hybiske and Stephens, 2007 , 2008 ; Lutter et al, 2013 ) for subsequent rounds of infection. Extrusion studies to date have been performed in vitro utilizing many cell types and chlamydial strains (Todd and Caldwell, 1985 ; Hybiske and Stephens, 2007 , 2008 ; Chin et al, 2012 ; Lutter et al, 2013 ; Zuck et al, 2016a , b ); however, the role of extrusions in chlamydial pathogenesis in vivo has not yet been described.…”

Immunobiology of monocytes and macrophages during Chlamydia trachomatis infection

“…SB is supported by grants from the Birthe Meyers Foundation, the Beckett Foundation and, the Hertha Christensen Foundation. [100,101,[110][111][112][102][103][104][105][106][107][108][109]…”