Application of a vital staining method to measure feeding rates of field ciliate assemblages on a harmful alga

Abstract

The fluorescently labeled algae (FLA) technique using a vital green fluorescent stain CMFDA (5-chloromethylfluorescein diacetate) was applied to investigate the feeding activities of a field ciliate assemblage on a harmful alga, Heterocapsa circularisquama. Using epifluorescence under interference blue-light excitation improved the limitations noted in the original protocol (Li et al. 1996; Aquat Microb Ecol 10: 139-147), which are due to the occurrence of green auto-fluorescence similar to that of CMFDA-labeled prey in natural assemblages and in food vacuoles of field protists. CMFDA-labeled H. circulanrisquama were ingested by the tintinnid ciliate Favella taraikaensis at the same rate as non-labeled alga, indicating that the staining with CMFDA did not influence the feeding activity of the ciliates. In an in situ feeding experiment, more than 6 species of heterotrophic and mixotrophic ciliates were shown to be able to feed on H. circularisquama, and the ingestion rates of 2 species were estimated (1.08 and 1.83 cells ind.-1 h-1). The FLA method with CMFDA is effective in measuring the feeding activities of ciliates and is applicable to the estimation of the grazing impact of ciliate assemblages on H. circularisquama populations in natural assemblages.