title: Amoebal endosymbiont Parachlamydia acanthamoebae Bn9 can grow in immortal human epithelial HEp-2 cells at low temperature; an in vitro model system to study chlamydial evolution.
abstract: Ancient chlamydiae diverged into pathogenic and environmental chlamydiae 0.7-1.4 billion years ago. However, how pathogenic chlamydiae adapted to mammalian cells that provide a stable niche at approximately 37 °C, remains unknown, although environmental chlamydiae have evolved as endosymbionts of lower eukaryotes in harsh niches of relatively low temperatures. Hence, we assessed whether an environmental chlamydia, Parachlamydia Bn9, could grow in human HEp-2 cells at a low culture temperature of 30 °C. The assessment of inclusion formation by quantitative RT-PCR revealed that the numbers of bacterial inclusion bodies and the transcription level of 16SrRNA significantly increased after culture at 30 °C compared to at 37 °C. Confocal microscopy showed that the bacteria were located close to HEp-2 nuclei and were actively replicative. Transmission electron microscopy also revealed replicating bacteria consisting of reticular bodies, but with a few elementary bodies. Cytochalasin D and rifampicin inhibited inclusion formation. Lactacystin slightly inhibited bacterial inclusion formation. KEGG analysis using a draft genome sequence of the bacteria revealed that it possesses metabolic pathways almost identical to those of pathogenic chlamydia. Interestingly, comparative genomic analysis with pathogenic chlamydia revealed that the Parachlamydia similarly possess the genes encoding Type III secretion system, but lacking genes encoding inclusion membrane proteins (IncA to G) required for inclusion maturation. Taken together, we conclude that ancient chlamydiae had the potential to grow in human cells, but overcoming the thermal gap was a critical event for chlamydial adaptation to human cells.
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan.
authors: Chikayo
authors: C
authors: Yamane
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan; Research Fellow of Japan Society for the Promotion of Science, Kojimachi Business Center Building, 5-3-1 Kojimachi, Chiyoda-ku, Tokyo, Japan.
authors: Tomohiro
authors: T
authors: Yamazaki
authors: Division of Biomedical Imaging Research, Juntendo University Graduate School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, Japan.
authors: Shinji
authors: S
authors: Nakamura
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan.
authors: Junji
authors: J
authors: Matsuo
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan.
authors: Kasumi
authors: K
authors: Ishida
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan.
authors: Sumire
authors: S
authors: Yamazaki
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan; Hokkaido University Hospital, North-14, West-5, Kita-ku, Sapporo, Japan.
authors: Satoshi
authors: S
authors: Oguri
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan.
authors: Natsumi
authors: N
authors: Shouji
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan; Hokkaido University Hospital, North-14, West-5, Kita-ku, Sapporo, Japan.
authors: Yasuhiro
authors: Y
authors: Hayashi
authors: Division of Ultrastructural Research, Juntendo University Graduate School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, Japan.
authors: Mitsutaka
authors: M
authors: Yoshida
authors: Department of Advanced Medicine, Graduate School of Medicine, Hokkaido University, Nishi-7, Kita-15, Kita-ku, Sapporo, Japan.
authors: Yimin
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, Japan.
authors: Hiroyuki
authors: H
authors: Yamaguchi
mesh_terms: D000222:Adaptation, Physiological; D000656:Amoeba / Q000382:microbiology*; D002695:Chlamydiales / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D004847:Epithelial Cells / Q000166:cytology / Q000382:microbiology*; D019143:Evolution, Molecular; D023281:Genomics; D006801:Humans; D042541:Intracellular Space / Q000382:microbiology; D013559:Symbiosis; D013696:Temperature
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