Abstract
Colpodella species are free-living close relatives of apicomplexans that were recently reported to cause red blood cell infection in an immunocompromised human host and in a tick-borne human infection resulting in neurological symptoms. Unambiguous identification of the life cycle stages of Colpodella sp. using routine stains for light microscopy will aid rapid diagnosis in infections. Similarly, cells in culture and environmental samples can be rapidly identified by staining. Staining protocols are currently unavailable for cell detection by light microscopy. In this study, we investigated the feasibility of performing routine staining techniques for light microscopy for differentiating Colpodella sp. (ATCC 50594) and Bodo caudatus cysts in Hay medium cultures. We tested different basic and acidic dyes alone and in combination and also utilized a commercial trichrome staining protocol. The nonspecific fluorescent dye Calcofluor white was also evaluated. Staining times, dye concentrations, use of tap or distilled water rinses, use of a mordant and inclusion, or omission of decolorizers after staining were evaluated. We compared the intensity of color, clarity of morphological features, and cytoplasmic structures detected after staining. We report a new trichrome staining technique that allowed clear identification and differentiation of cyst stages of Colpodella sp. and B. caudatus. Immature Colpodella sp. cysts were identified as having an irregular, dual-colored (demilune), dark blue-purple and white appearance. Mature Colpodella sp. cysts stained dark red-blue and were identified in four-way mitotic division, while cysts of B. caudatus in diprotist or monoprotist (ATCC 30905) cultures were detected as spherical and red-pink in appearance.
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Acknowledgments
We gratefully acknowledge Cleveland Clinic Lerner Research Institute, Cleveland Clinic NIH shared instrument grant for Orbitrap Elite LC-MS instrument; Cleveland Clinic NIH grant 1S100D023436-01 for Fusion Lumos instrument; and Cleveland Clinic Lerner Research Institute, Imaging Core. We thank Alberto Williams for assistance in staining and preparation of tables and Cleveland State University BGES stockroom staff Susan Moreno-Molek and Michelle Zinner for stain preparation.
Funding
The study was supported by funds from the Cleveland State University Undergraduate Summer Research Awards 2017 and 2019.
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Sam-Yellowe, T.Y., Addepalli, K., Yadavalli, R. et al. New trichrome stains identify cysts of Colpodella sp. (Apicomplexa) and Bodo caudatus. Int Microbiol 23, 303–311 (2020). https://doi.org/10.1007/s10123-019-00104-1
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DOI: https://doi.org/10.1007/s10123-019-00104-1