VirulentCoxiella burnetiipathotypes productively infect primary human alveolar macrophages

Abstract

Summary The intracellular bacterial pathogen Coxiella burnetii is a category B select agent that causes human Q fever. In vivo, C. burnetii targets alveolar macrophages wherein the pathogen replicates in a lysosome-like parasitophorous vacuole (PV). In vitro, C. burnetii infects a variety of cultured cell lines that have collectively been used to model the pathogen’s infectious cycle. However, differences in the cellular response to infection have been observed, and virulent C. burnetii isolate infection of host cells has not been well defined. Because alveolar macrophages are routinely implicated in disease, we established primary human alveolar macrophages (hAMs) as an in vitro model of C. burnetii-host cell interactions. C. burnetii pathotypes, including acute disease and endocarditis isolates, replicated in hAMs, albeit with unique PV properties. Each isolate replicated in large, typical PV and small, non-fused vacuoles, and lipid droplets were present in avirulent C. burnetii PV. Interestingly, a subset of small vacuoles harbored single organisms undergoing degradation. Prototypical PV formation and bacterial growth in hAMs required a functional type IV secretion system, indicating C. burnetii secretes effector proteins that control macrophage functions. Avirulent C. burnetii promoted sustained activation of Akt and Erk1/2 pro-survival kinases and short term phosphorylation of stress-related p38. Avirulent organisms also triggered a robust, early pro-inflammatory response characterized by increased secretion of TNF-α and IL-6, while virulent isolates elicited substantially reduced secretion of these cytokines. A corresponding increase in pro- and mature IL-1β occurred in hAMs infected with avirulent C. burnetii, while little accumulation was observed following infection with virulent isolates. Finally, treatment of hAMs with IFN-γ controlled intracellular replication, supporting a role for this antibacterial insult in the host response to C. burnetii. Collectively, the current results demonstrate the hAM model is a human disease-relevant platform for defining novel innate immune responses to C. burnetii.

Right on Q: Genetics begin to Unravel Coxiella Burnetii host cell Interactions

“…All other strains are considered biosafety level-3 organisms. Importantly, the growth kinetics and intracellular trafficking of NMI and NMII in human macrophages appear indistinguishable, making infection by NMII a relevant model for ex vivo pathogen-host interaction studies [31,32]. NMII also has the advantage of being roughly 500-fold more infectious on a per particle basis for nonprofessional phagocytic cells, such as Vero epithelial cells, aiding cell biology studies [33].…”

“…Ligation of TLR-2 triggers synthesis of the future science group www.futuremedicine.com proinflammatory cytokine tumor necrosis factor, which in turn elicits production of the antibacterial effector nitric oxide that inhibits NMII replication [37][38][39]. Although potentiated innate immune signaling also occurs during NMII interactions ex vivo with primary human macrophages and dendritic cells [32,40], no growth defects relative to NMI are observed. However, the resulting robust immune response elicited by NMII during primary infection of mammals is thought to render the strain avirulent [37,40].…”

“…The primary target of Coxiella during natural infection is alveolar macrophages [32,[41][42][43]. The organism can subsequently disseminate to colonize and replicate in resident macrophages of different tissues and organs [44].…”

“…Thus, the organism is adept at maintaining host cell viability by disrupting both intrinsic and extrinsic apoptotic cell death signaling [63,77]. Apoptosis inhibition involves pathogen activation of the prosurvival kinases Akt, Erk1/2 and PKA [32,78,79]. Host cell maintenance requires active Coxiella protein synthesis and a functional T4BSS [63,77,80].…”

“…However, infection with the Coxiella icaA mutant alone fails to activate the inflammasome, suggesting that, like antiapoptotic effectors, Coxiella deploys functionally redundant effectors that contribute to protection [98]. A potential caveat is related to the observation that human primary alveolar macrophages infected with NMII, but not virulent strains of Coxiella, release mature IL-1β [32]. Thus, inflammasome activation may be Coxiella strain and/or host cell specific.…”