S. Afr. J . Bot., 1987, 53(5): 405-407 405

A choanoflagellate (craspedomonad) epiphytic on Microcystis aeruginosa

W.E. Scott*, R.D. Robarts and L.M. Sephton National Institute for Water Research, C.S.I.R., P.O. Box 395, Pretoria, 0001 Republic of South Africa

Accepted 17 June 1987

A large population of the choanoflagellate, Salpingaeca sp., developed on Microcystis aeruginasa KOtz. emend. Elenkin which had been collected from a hypertrophic lake and maintained for several days in the laboratory. The choanoflagellate appears to be related to, but not identical to, S. ampul/aides Bicudo and Bicudo which occurs in Brazil on Sa/vinia. Staining of the flagellates with fluorescent dye demonstrated the ingestion of bacteria.

'n Groot bevolking van die koanoflagellaat het op kolonies van Microcystis ae'ruginasa KOtz. emend. Elenkin ontwikkel nadat die sianobakterium in 'n hipertrofe dam versamel is en 'n paar dae in die laboratorium gehou is. Die koanoflagellaat is verwant aan, maar is nie identies met S. ampul/aides Bicudo en Bicudo wat in Brasilie op Sa/vinia voorkom nie. Kleuring van die flagellate met 'n fluoressensiekleurstof het die inname van bakteriee aangetoon .

Keywords: Cyanobacteria, Hartbeespoort Dam, hypertrophic, Salpingaeca

*To whom correspondence should be addressed

Introduction The choanoflagellates comprise a group of uniflagellated organisms of uncertain taxonomic position characterized by the presence of a collar at the flagellar base. In the zoological classification system the organisms are placed in the zooflagel­late class Zoomastigophorea (Norris 1982; Leadbeater 1985) while some phycologists have placed them in the order Protomonadales (Fott 1971) or in the order Craspedomona­dales of the Chrysophyceae (Bourrelly 1968). However, it is now generally accepted that the organisms should not be regarded as algae but as protozoans and thus be excluded from the plant kingdom (Lewin & Gibbs 1982).

Choanoflagellates are often recorded as epiphytes on planktonic and benthic algae, on macrophytes, detritus and on aquatic animals (Lemmermann 1910). Free-living choano­flagellates are often neustonic, attaching themselves upside down (the flagellum pointing downwards) to surface films (Norris 1965). They collect food by generating water move­ments with their flagellum, carrying particulate matter into the extensible collar for ingestion and digestion (Ellis 1929). Ultrastructurally the collar consists of anastomosing radiating microvillae (Laval 1971).

During the course of the National Institute for Water Research's studies on Hartbeespoort Dam we noticed the development of a choanoflagellate population epiphytic on colonies of the cyanobacterium Micracystis aeruginasa. We believe this to be the first record of the choanoflagellate Salpingaeca sp. in South Africa and since there are only limited references and no illustrations of Salpingoeca epiphytic on Micracystis we supply photographic illustrations of the association between the two organisms.

Methods Water samples, with and without preservative [a mixture of water, ethyl alcohol and formalin in a ratio of 6:3:1 (Standard Methods 1985)], were examined by phase contrast microscopy with a Zeiss Research Microscope and photographed with an Olympus automatic photomicrograph camera model PM-10AD. Samples were also stained with the fluorochrome 4',6-diarnidino-2-phenylindole (DAPI), which is DNA-specific, and examined by epifluorescence microscopy as described by Robarts & Sephton (1981).

Results In July 1986 a 5-l sui-face water sample from Hartbeespoort Dam was kept in the laboratory with gentle stirring over the weekend. On Monday numerous specimens of the flagellates were observed on the Micracystis colonies (Figures 1 - 3). The flagellates were not noticeable in the sample at the time of collection.

We compared our species with illustrations and/or descrip­tions in the literature of more than 50 other species of Sal­pingaeca but could not identify our specimens as belonging to species already described. It resembles S. qmpullaides Bicudo & Bicudo (1983; 1984a, b) but the neck of the vase­like lorica (or theca) in our specimens are shorter. The neck is approximately half the total length of the lorica in our specimens compared with the two-thirds length for S. ampul­/aides by Bicudo & Bicudo (1983). LikeS. ampul/aides, our specimens possess a round base with no pedicel with the cell completely filling the lorica up to the neck (Figure 3). The flagella in our specimens also appear shorter than those illustrated for S. ampul/aides (Bicudo & Bicudo 1983). The diameter of the lorica is 6 to 6,5 J.lm at the base and it is 10 J.lm long. The length of the cell including the collar is 16 J.lm excluding the flagellum.

The collar and flagellum disappeared after fixation with either the alcohol:formalin fixative used for preservation or the formalin-only fixative used to preserve samples for DAPI staining. DAPI staining clearly demonstrated the nucleus of the organism as well as the DNA of ingested bacteria (Figure 4).

Discussion Although several authors reported the presence of various Salpingaeca species on phytoplankton and water plants in general, we could find only two authors specifically men­tioning the presence of Salpingaeca on Micracystis colonies (Skuja 1956; Tauscher 1980). The report from Sweden by Skuja (1956) mentioned S. vaginicala Stein, S. gracilis Clark fa. and S. umula Skuja on various species of Microcystis while Tauscher (1980) foundS. frequentissima (Zach.) Lemm. on the sheaths of M. aeruginasa and M. viridis (A. Br.) Lemm. in East Germany. Three of these four species are stalked and only S. vaginica/a is sessile like the Salpingaeca sp. from

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Figure 1 A single Sa/pingoeca sp. epiphytic on a small Microcystis aeruginosa colony. Several bacteria (dark spots) can be noticed in the Microcystis mucilage. Phase contrast; scale bar = 15 ~m.

Figure 2 A group of Sa/pingoeca sp. cells partly immersed in a Microcystis colony. a . Extensible collar (part of the cell), b. neck of the lorica. Phase contrast; scale bar = 10 ~m.

Figure 3 Sa/pingoeca sp. showing that the cell completely fills the lorica and that a pedicel is absent. a. Extensible collar (part of the cell), b. neck of the lorica. Gas vacuoles in Microcystis cells (arrow c) have been collapsed by centrifugation. Phase contrast; scale bar = 5 ~m.

Hartbeespoort Dam. Salpingoeca marssonii Lemm. (not stalked), S. frequentissima and S. elegans (Bachmann) Lemm. have been observed on the related cyanobacterial genus Coelosphaerium by earlier authors (see Huber-Pestalozzi 1941). All of the above species are quite different from the Salpingoeca found in Hartbeespoort Dam. A similar species of choanoflagellate to that found in Hartbeespoort Dam is S. ampulloides from Brazil which was found on the leaves of Salvinia herzogii De la Sota (Bicudo & Bicudo 1983).

S.-Afr. Tydskr. Plantk ., 1987, 53(5)

Figure 4 Cells of Sa/pingoeca sp. stained with DAPI showing the choanoflagellate nucleus (arrow) and fluorescent bacteria present both in the protoplasm and as background. Note the disappearance of collar and flagellum as a result of fixing the sample with formalin. Epi­fluorescence; scale bar = 3 ~m.

Microcystis samples collected from Hartbeespoort Dam at approximately two-weekly intervals for the remainder of 1986 (August to December) were routinely examined for the pre­sence of further Salpingoeca sp. None were observed. We assume the epiphytic relationship to be opportunistic rather than facultative. It therefore appears from our observations, and the relative paucity of published reports of Salpingoeca on Microcystis, that the epiphytic attachment of this genus on Microcystis is not as common as one would expect it to be considering the widespread and common nature of Micro­cystis globally and of Salpingoeca at least in Europe and in America.

Acknowledgements This is contribution No. 52 to the Hartbeespoort Dam Eco­system Programme. We thank Prof. R.E. Norris for initially identifying the choanoflagellate as a species of Salpingoeca.

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