Metabolic Features of Protochlamydia amoebophila Elementary Bodies – A Link between Activity and Infectivity in Chlamydiae

Abstract

The Chlamydiae are a highly successful group of obligate intracellular bacteria, whose members are remarkably diverse, ranging from major pathogens of humans and animals to symbionts of ubiquitous protozoa. While their infective developmental stage, the elementary body (EB), has long been accepted to be completely metabolically inert, it has recently been shown to sustain some activities, including uptake of amino acids and protein biosynthesis. In the current study, we performed an in-depth characterization of the metabolic capabilities of EBs of the amoeba symbiont Protochlamydia amoebophila. A combined metabolomics approach, including fluorescence microscopy-based assays, isotope-ratio mass spectrometry (IRMS), ion cyclotron resonance Fourier transform mass spectrometry (ICR/FT-MS), and ultra-performance liquid chromatography mass spectrometry (UPLC-MS) was conducted, with a particular focus on the central carbon metabolism. In addition, the effect of nutrient deprivation on chlamydial infectivity was analyzed. Our investigations revealed that host-free P. amoebophila EBs maintain respiratory activity and metabolize D-glucose, including substrate uptake as well as host-free synthesis of labeled metabolites and release of labeled CO2 from 13C-labeled D-glucose. The pentose phosphate pathway was identified as major route of D-glucose catabolism and host-independent activity of the tricarboxylic acid (TCA) cycle was observed. Our data strongly suggest anabolic reactions in P. amoebophila EBs and demonstrate that under the applied conditions D-glucose availability is essential to sustain metabolic activity. Replacement of this substrate by L-glucose, a non-metabolizable sugar, led to a rapid decline in the number of infectious particles. Likewise, infectivity of Chlamydia trachomatis, a major human pathogen, also declined more rapidly in the absence of nutrients. Collectively, these findings demonstrate that D-glucose is utilized by P. amoebophila EBs and provide evidence that metabolic activity in the extracellular stage of chlamydiae is of major biological relevance as it is a critical factor affecting maintenance of infectivity.

Integrating metagenomic and amplicon databases to resolve the phylogenetic and ecological diversity of the Chlamydiae

“…The absence of representative isolates for most chlamydial families and the lack of specific information about their actual hosts illustrate the huge gap we are facing in studying and understanding chlamydial biology and evolution. Closing this gap will be a major challenge requiring the application of novel approaches and techniques such as single-cell genomics (Woyke et al, 2009;Bruns et al, 2010;Wang and Bodovitz, 2010;Siegl et al, 2011;Li et al, 2012;Stepanauskas, 2012;Seth-Smith et al, 2013) and host-free cultivation and analysis of Chlamydiae (Haider et al, 2010;Omsland et al, 2013;Sixt et al, 2013).…”

Chlamydial metabolism revisited: interspecies metabolic variability and developmental stage-specific physiologic activities

“…The exchange of D‐glucose with the nonmetabolizable stereoisomer L‐glucose, in case of P. amoebophila , or the exchange of D‐glucose‐6‐phosphate by D‐glucose, in case of C. trachomatis , leads to a more rapid decrease in infectivity during host‐free incubation of chlamydiae (Sixt et al ., ). Additional evidence for a biological role of host‐free EB activity was provided by the experiments with the ionophore CCCP described above.…”

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

“…Using a general metabolomics approach, Muller et al, have reported a molecular cartograph of C. pneumoniae infected HEp2 cells, and supporting the role of previously non-annoted metabolites as biomarkers for infection (Muller et al 2013). Nutrient requirements, maintenance of respiratory activity and metabolism of d -glucose in elementary bodies (EB) of the amoeba symbiont, Protochlamydia amoebophila has been demonstrated using metabolomics approaches (Sixt et al 2013). Using a high-performance liquid chromatography–mass spectrometric method (HPLC–MS/MS), we examined the genital lipidome profile of Ct serovar D infected female Dunkin Hartley guinea pigs.…”