The Life History of Adenocystis utricularis (Bory) H. et H.
[Read before Otago Branch, November 9, 1954; received by Editor, January 11,1955.]
Zoospores from the unilocular sporangia of Adenocystis utricularis develop asexually into heteroblastic, prostrate germlings, from which new plants arise as erect filaments. There is occasional production of plurilocular sporangia by the filamentous germlings. The life history thus appears to consist of direct development of the spores with arrested juvenile phases, capable of both vegetative and accessory reproduction, persisting throughout the winter months.
Adenocystis utricularis (Bory) H. et H. occurs in the littoral zone of southern circumpolar waters. It is recorded from subantarctic South America, Kerguelen's Land, Tasmania, the Auckland and Campbell Islands, the Chatham Islands. Stewart Island, and the South Island of New Zealand (Plates 11A and 11B). Also the south of the North Island.
The plants are present on the shore for only a few months of the year. They reproduce by zoids produced in unilocular sporangia which cover the whole surface, and then die back. In Dunedin the plants are summer annuals, first appearing on the shore in August. By October the first plants are mature, and a succession of plants matures and dies back throughout the summer months until by about April the last plants have disappeared. Probably water temperature affects time of development, since plants within Otago Harbour mature and die back three or four months earlier than those of the open coast where the water is [ unclear: ] colder. This may account for Skottsberg's (1907) description of plants from the subantarctic as shedding their zoids from March to August.
The cultural and cytological methods described below have also been used successfully with a number of other members of the Dietyosiphonales and Chordariales.
Mature plants, collected at low tide, were carried back to the laboratory in glass jars containing no water, washed thoroughly with several changes of boiled sea water and placed in a small beaker of boiled sea water until a suspension of zoids was obtained. It was found best to leave the material in the water for as short a period as possible in order to minimise contamination by diatoms and other epiphytes; in fully mature material five minutes was usually adequate. The suspension of zoids was then poured over sterile slides in a flat container and left for half an hour for the zoids to settle. The slides were then washed with a jet of boiled sea water and placed in culture solution. In this way it was
possible to obtain slides evenly covered with spores and reasonably free from contamination.
For cytological investigation the slides were placed in small pots of culture solution, one slide to each pot of 120 ccs. solution, and the pots placed in a good light and kept cool by circulating water. Erdschreiber solution, as modified by Drew (1952) was used. If the plants were to be grown for long periods, they were established in this manner for a week and then transferred to large, well aerated aquarium tanks, or to running sea water.
Filamentous Phascs. All handling of the filamentous phases was done on the slides on which they had grown. They were usually examined at a week to ten days old, at which stage growth was vigorous and lateral branching and hair formation just beginning. The slides were fixed in 3:1, absolute alcohol: glacial acetic acid for 30 minutes, transferred to 95 per cent. alcohol and left for several hours. After gradual transference to water, they were stained overnight in Heidenhain's iron alum haematoxylin, differentiated with iron alum, very slowly dehydrated and mounted in Sira mountant.
Macroscopic Phases. Small pieces were fixed for 24 hours in 3:1, absolute alcohol: glacial acetic acid and then embedded by the tertiary butyl alcohol method in an artificially hardened wax (SSB4, Waterman, 1939). They were sectioned at thicknesses from 2-4μ and stained with iron alum haematoxylin, using the same schedule as that for the microscopic phases.
Fig. 1.—A. Zoospore of Adenocystis utriculans showing the two flagella (fl), the crescentic eyespot (st), the pyrenoid (py) and the single, saddle-like plastid (pl). B. A group of embryospores (em) 24 hours after settling, showing the random orientation of the germ tubes C. A similar group 40 hours after settling. By this stage most of the eyespots have disappeared.
Fig. 2.—A. Well developed filamentous germling 19 days old from the same culture as those illustrated in Fig. 3 A-J. B. Plurilocular sporangia on 34-day-old filamentous germling in culture. C. One of the plurilocular sporangia of B shown after the escape of the [ unclear: ] , one of which can be seen (z). D. Plurilocular sporangia on filaments scraped from mussel shells on which macroscopic plants were just visible, 17.10.50. All outlines drawn with the and of a camera lucida.
This has already been described in abstract form (Naylor, 1954). The form of the zoospores and early stages in their germination are illustrated in Fig. 1 and in Plates 12 and 13. Correlation is seen between the lack of any phototactic responses of the zoospores and the random orientation of the germ tubes of the embryospores. Although there is no response to the direction of the incident light, a high light intensity is necessary before germmation will commence. Cutting down of light intensity was found to retard, or even prevent, germination.
Stages in the formation of filamentous germlings are illustrated in Plates 13A, 13C and Fig. 2A, and in the formation of discoid germlings in Figure 3 and Plates 12B, 12C and 13B. Usually disc formation results from lateral proliferation
[ unclear: ] 11—A [ unclear: ] growing on [ unclear: ] rocks at Portobello [ unclear: ] Biological [ unclear: ] 221053 showing the typical dusters of adult plants Photograph by Dr. E. J. Bath [ unclear: ] B. [ unclear: ] a belt just above the [ unclear: ] chondrophylla at [ unclear: ] Beach, [ unclear: ] Island 19254 The [ unclear: ] dumps [ unclear: ] again be seen
Plate 12—Photomierographs of living plants in seawater of three different cultures of [ unclear: ] 18 22 and 45 days old respectively Photographed 45 52 A 18-day-old plants from St Clam [ unclear: ] which branching of filaments and [ unclear: ] formation has begun B 22-day-old plants from [ unclear: ] Otago Many of the plants have well formed discs but a number of them have retained then filamentous form Hans can also be seen C 45-day-old plants from St Clan, showing well developed discs D, E—Photomierographs of terminal dividing cells of germlings of [ unclear: ] D 8-day old germling fixed at 01 30 hours and showing several [ unclear: ] and a late prophase [ unclear: ] Scale = × 2,750 E 13-day-old germling fixed at 03 30 hours and showing a prophase nucleus Scale = × 3,650
Plate 13—Photomerographs of fixed and stained plants showing [ unclear: ] A 5-day-old germlings with [ unclear: ] still visible it one end The majority of the cells contain only one of two plastids B 19-day-old discord germling, grown for 12 days in running seawater at Portobello Marine Brological Station C 20-day-old germling which has retained the filamentous form and showing branching and an increased number of [ unclear: ] per cell Two harts can be seen passing out of the [ unclear: ] plane to the left
from the terminal cells of a short filament. The discs increase in size to about 50μ and develop long, colourless hairs.
After about two months erect filaments arose from the prostrate systems both filamentous and discoid. At first these were uniseriate without a terminal hair, but soon became multiseriate with a single terminal hair (Fig. 4), as were the youngest plants from the shore described by Skottsberg (1907). Although these erect structures have not yet been grown to macroscopic dimensions, they are assumed to be the young stages of the macroscopic plants. Each germling gives rise to several erect filaments, which may account for the characteristic clumps in which the mature plants occur (Plate 11A).
After about three weeks, some of the filamentous germlings developed pluri-locular sporangia (Figs. 2B and C). The zoids were liberated in a single apical stream, but their fate was not followed. Plurilocular sporangia have not been seen on the discoid individuals. Filamentous growths scraped from mussel shells on which macroscopic plants were just beginning to reappear, and which showed similar details of cell structure to the germlings grown in culture, also bore plurilocular sporangia (Fig. 2D).
In the macroscopic plant the cells of the surface layer contain a single parietal plastid, but those of the deeper layers may contain several discoid plastids. All the cells of the surface layer are rich in fucosan—as tested for by a freshly made solution of vanillin in 95 per cent. alcohol followed by concentrated hydrochloric acid—and contain several small pyrenoids and an inconspicuous nucleus. The cells of the ectocarpoid hairs which grow out at intervals from depressions in the surface, contain neither plastids nor pyrenoids. Cell details of the microscopic discs and filaments agrees with that of the surface layers of the macroscopic plant. The cells of the very young germlings contain only a single pyrenoid (Plate 13A), but the number per cell increases with age.
Nuclear divisions are found most readily in filamentous germlings about a week to ten days old, and are restricted to the terminal cells, to cells from which lateral branches are developing and to the basal cells of the hairs.
The nucleus first becomes visible as a slightly granular body with a small nucleolus. During early prophase the nucleolus increases considerably in size,
and small, intensely staining granules become differentiated within the nucleus which increases to 3-4μ in diameter. The nucleolus then disappears and the chromosomes become clearly visible as spherical bodies about 0.4μ in diameter (Plate 12, D, E). At this stage it is possible to count the chromosomes and a more detailed account of this aspect is now in preparation. Full metaphase plates have only been found in side view, as is to be expected in horizontally growing filaments, and so far all attempts to turn them over have failed. Typical anaphase and telophase stages follow, the former often showing indications of a spindle.
In the macroscopic plant, stages of division similar to those in the filaments have been found in the basal cells of the ectocarpoid hairs. In the unilocular sporangia uni- and quadri- nucleate stages were found, but no stages to indicate the type of division. During the subsequent subdivision into zoids the nuclei become progressively smaller, and although metaphase plates are frequently seen in the latter stages, no counts have yet been made on account of the extremely small size of the chromosomes.
Heteroblasty—i.e., the production of germlings of two different forms by zoids of similar origin—was first described by Sauvageau (1924a) in Cladosiphon zosterae (J. Ag.) Kylin [Castagnea-zosterae (J. Ag.) Thur.] and later (1924b) by him in two further species of Cladosiphon, C. cylindricus (Sauv.) Kylin (Castagnea cylindrica) Sauv. and C. irregularis (Sauv.) Kylin (Castagnea irregularis Sauv.). It has since been recorded in several members of the Ectocarpales, Dictyosiphonales and Chordariales.
Little is known concerning the conditions determining the form of the germling. Sauvageau (1927, p. 417) believed the form to be entirely independent of the external conditions “car elle se présente en des points quelconques d'une měme goutte d'eau, et aboutit à l'hétéroblastic.” Although in Adenocystis utricularis many slides were obtained showing approximately equal distribution of the two types of germling, there were definite indications that the form of the germling was affected by external conditions. Cultures were established in small pots and, after periods varying from a week to seventeen days, some of the slides were transferred to running seawater at Portobello Marine Biological Station, whilst the remainder were left in the small pots. In every case the plants in the running seawater became predominantly or exclusively discoid, whilst those in the small pots remained filamentous. Only in one series were a few small discs produced in the pots after 29 days. The conditions were not sufficiently controlled, however, to know what was promoting disc formation in the running seawater.
Kylin (1933) suggested that in Scytosiphon lomentaria (Lyngb.) Endl. disc formation did not occur if the culture were overcrowded, but in both A. utricularis and in S. lomentaria I have obtained densely crowded slides in which every germling has become discoid. Hollenberg (1941) believed the heteroblasty in Hapterophycus canaliculatus S. & G. to be cytologically determined, the discoid forms being diploid, the filamentous haploid. There is no evidence to support this idea so far as A. utricularis is concerned.
In A. utricularis the disc is formed from a short filament, but in Cladosiphon zosterae (Sauvageau, 1927, Fig. 6) the embryospore germinates directly into a
disc. Sauvageau (1924a) suggested that the formation of a fertile and ephemeral filament had preceded disc production, and that forms in which the spore germinated directly into a disc were examples of accelerated embryogeny. The discoid form was considered to be an advantage, giving firmer attachment to the plant.
The life history of A. utricularis appears to be of the direct type seen in Asperococcus compressus Griff. ex Hook. (Sauvageau, 1929), Striaria attenuata Grev. (Kylin, 1933), Dictyosiphon chordaria Aresch. (Föyn, 1934) and in Hapterophycus canaliculatus S. & G. (Hollenberg, 1941). The microscopie discs and filaments are probably juvenile phases in which the plants persist during the winter, and which probably undergo accessory reproduction by means of plurilocular sporangia. They also serve as a means of vegetative reproduction since each germling can produce several erect filaments.
This type of life history is usually assumed to be due to the failure of meiosis in the unilocular sporangia, but as yet there is no information concerning the type of nuclear division in the unilocular sporangia of A. utricularis.
Drew, K. M., 1952. Studies in the Bangioideae. 1. Observations on Bangia fusco-puipuiea (Dillw.) Lyngb. in culture. Phytomorph., 2. 38.
Foyn, B. R., 1934. Uber den Lebenscyklus einiger Braunalgen. Vorlaufige Mitteilung. Bergens Museums Arbok, 1: 1.
Hollenberg, G. J, 1941. Culture studies of marine algae. II. Hapterophycus canaliculatus S & G Amer. J. Bot., 28: 676.
Kylin, H., 1933. Über die Entwicklungsgeschichte der Phaeophyceen. Lunds Univ. Arsskr. N.F. Avd. 2, 29: 1.
Naylor, M, 1954. The Life Cycle of Adenocystis utriculans Huitième Congrès International de Botanique, Paris. Rapports et communications a la Section 17: 73.
Sauvageau, C., 1924a. Sur le curieux developpement d'une algue phéasporce [ unclear: ] . Castagnea zosterae Thuret. C. R. Acad. Sci. Paris, 179: 1381.
—— 1924b. Sur quelques exemples d'heteroblastie dans le developpement des algues phéosporées. Ibid., 1576.
—— 1927. Sur le Castagnea zosterae Thuret. Bull. Stat. Biol. Arcachon. 24: 369.
—— 1929. Sur le developpement de quelques phéosporées. Bull. Stat. Biol. Arcachon, 26: 254.
Skottsberg, C., 1907. Zur Kenntnis der subantarktischen und antarktischen Meeresalgen. 1. Phaeophyceen. Wiss. Ergeb. Schwed. Süudpolar Exped. 1901-1903, 4: 1.
Waterman, H. C., 1939. The preparation of hardened embedding paraffins having low melting-points. Stain Technology, 14: 55.
Dr. Margaret Naylor
Department of Botany
Queen Mary College
University of London
Mile End Road
London, E 1.