Abstract
Members of the Chlamydiales order all share a biphasic lifecycle alternating between small infectious particles, the elementary bodies (EBs) and larger intracellular forms able to replicate, the reticulate bodies. Whereas the classical Chlamydia usually harbours round-shaped EBs, some members of the Chlamydia-related families display crescent and star-shaped morphologies by electron microscopy. To determine the impact of fixative methods on the shape of the bacterial cells, different buffer and fixative combinations were tested on purified EBs of Criblamydia sequanensis, Estrella lausannensis, Parachlamydia acanthamoebae, and Waddlia chondrophila. A linear discriminant analysis was performed on particle metrics extracted from electron microscopy images to recognize crescent, round, star and intermediary forms. Depending on the buffer and fixatives used, a mixture of alternative shapes were observed in varying proportions with stars and crescents being more frequent in C. sequanensis and P. acanthamoebae, respectively. No tested buffer and chemical fixative preserved ideally the round shape of a majority of bacteria and other methods such as deep-freezing and cryofixation should be applied. Although crescent and star shapes could represent a fixation artifact, they certainly point towards a diverse composition and organization of membrane proteins or intracellular structures rather than being a distinct developmental stage.
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Acknowledgments
This work was supported by the Swiss National Science Foundation (project no. PDFMP3-127302). Brigida Rusconi is supported by the Swiss National Science Foundation within the PRODOC program “Infection and Immunity”. Julia Lienard is supported by SUEZ-Environment (CIRSEE, Paris, France). We thank D. Bardy (CHUV) for measurements of osmolarity. We thank the PFMU at the Medical Faculty of Geneva for assisting with electron microscopy.
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The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors declare having no conflict of interest related to the content of this contribution.
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Brigida Rusconi and Julia Lienard have contributed equally to this study.
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Rusconi, B., Lienard, J., Aeby, S. et al. Crescent and star shapes of members of the Chlamydiales order: impact of fixative methods. Antonie van Leeuwenhoek 104, 521–532 (2013). https://doi.org/10.1007/s10482-013-9999-9
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DOI: https://doi.org/10.1007/s10482-013-9999-9