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Phycology |
Department of Botany, University of British Columbia, Vancouver, British Columbia V6T 1Z4 Canada
Received for publication August 25, 2000. Accepted for publication February 15, 2001.
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Key Words: Acrosiphonia • Chlorochytrium • Chlorophyta • Codiolum • endophyte • life history phases • phenology
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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The endophytic diploid phase of Acrosiphonia was previously identified as Chlorochytrium inclusum Kjellman and Codiolum petrocelidis Kuckuck. These are morphologically distinct unicells, which were initially placed, separate from Acrosiphonia, in the Order Chlorococcales (e.g., Fritsch, 1956
). Extensive culture studies, primarily in the 1960s and 1970s (Hollenberg, 1958
; Fan, 1959
; Jónsson, 1959a, 1962, 1966, 1970
; Chihara, 1969
; Kornmann, 1970a, 1972
; Hudson, 1974
; Miyaji and Kurogi, 1976
; Miyaji, 1984, 1996
), implicated the unicellular endophytes in the life history of Acrosiphonia and also of Spongomorpha (a closely related green algal genus not present in the northeast Pacific). More recently, DNA sequences of the ITS regions of the nuclear ribosomal cistron (Sussmann et al., 1999
) confirmed that the morphologically different endophytes Chlorochytrium inclusum and Codiolum petrocelidis and Acrosiphonia constitute alternating phases in the same life history in southern British Columbia (for this reason, all reference to the Chlorochytrium and Codiolum phases will appear in single quotes).
Description of Acrosiphonia's phases in nature
Acrosiphonia plants are composed of branched filaments that tend to be bound together by hooked branchlets and rhizoids to form large tangled masses. We recognize two species, A. arcta (Dillwyn) Agardh and A. mertensii Ruprecht, in southern British Columbia based on morphological (Hudson, 1974
) and molecular (Sussmann et al., 1999
) data. Acrosiphonia mertensii is present only in the northeast Pacific, whereas A. arcta has been recorded in both the northern and southern hemispheres, including the northwest Atlantic. Filamentous Acrosiphonia plants are found in the low-to-mid-intertidal zone on boulders or epiphytic on other algae. Seasonality and reproductive phenology is unknown for gametophytic Acrosiphonia plants.
Similarly, few studies (none from the northeast Pacific) have described seasonal abundance and reproductive phenology of ‘Chlorochytrium’ and ‘Codiolum.’ ‘Chlorochytrium inclusum’ is a spherical unicell, reported to be 80–100 µm in diameter and endophytic in foliose red algae such as Weeksia, Mazzaella, Schizymenia, Dilsea, and Farlowia. Some of these unicells have been identified as the sporophyte phase of Acrosiphonia or Spongomorpha in culture and molecular studies (Kornmann, 1961, 1964
; Jónsson, 1962, 1966
; Chihara, 1969
; Hudson, 1974
; Miyaji and Kurogi, 1976
; Sussmann et al., 1999
). Two studies on Arctic and European ‘C. inclusum’ have found it to be fertile in winter (Kjellman, 1883
; Kornmann, 1964
).
‘Codiolum petrocelidis,’ unlike ‘Chlorochytrium,’ consists of an ovoid vesicle portion and a colorless stalk. It is found embedded within the filaments of the crustose red algae Haemescharia hennedyi (Harvey) Vinogradova and Yakoleva and Petrocelis (= tetrasporophytic phase of the bladed alga, Mastocarpus, and so further reference to this crust will appear in single quotes). Like ‘Chlorochytrium’ a number of culture and molecular studies suggest that these ‘Codiolum’ cells represent a phase in the life history of Acrosiphonia or Spongomorpha (Hollenberg, 1958
; Fan, 1959
; Jónsson, 1959a, 1962
; Kornmann, 1961, 1964
; Hudson, 1974
; Sussmann et al., 1999
). A study by Kornmann (1961)
suggested the youngest ‘Codiolum’ stages are found within H. hennedyi in Helgoland, Germany, in July, and pass the following winter in a vegetative state, becoming fertile the subsequent winter. In the northeast Pacific, Dethier (1987)
noted dense ‘Codiolum’ colonization of ‘Petrocelis franciscana’ Setchell and Gardner [= tetrasporophytic phase of Mastocarpus papillatus (Agardh) Kützing] in summer in Washington State, USA.
Environmental requirements for gametophyte and sporophyte
Hanic (1965)
established that the macroscopic (1–2 mm in length), free-living ‘Codiolum’ phase of the filamentous green alga Urospora is fertile in the winter, fertility being induced by cold temperatures. He observed that Urospora is present year-round, but dies off substantially in the summer months. It is the ‘Codiolum’ that presumably better survives high summer temperatures and desiccation (L. Hanic, University of British Columbia, personal communication). Environmental requirements for the production and growth of Acrosiphonia's sporophytes, ‘Chlorochytrium’ and ‘Codiolum,’ have not been identified. However, Hudson (1974)
and Miyaji (1996)
have shown in culture that the sporophytes of A. arcta and A. spiralis Sakai, respectively, did not produce zoospores at temperatures 
15°C (but did at 5°C and 10°C). Hudson (1974)
also found that long-day (16 h light: 8 h dark photoregime) conditions were required for the germination of Acrosiphonia filaments, and growth of gametophytic plants in culture was inhibited at 15–20°C. This implies seasonality of Acrosiphonia's gametophyte and is supported by the fact that this phase has never been collected in the winter.
Although the two Acrosiphonia species, A. arcta and A. mertensii, can be distinguished in the field, it is not possible to identify the species in the endophytic diploid phase. The two morphologically distinct endophytes, ‘Chlorochytrium’ and ‘Codiolum,’ are alternate phenotypes of the sporophyte of a single Acrosiphonia species, i.e., morphological variation is solely attributed to the different nature of the two hosts (Sussmann et al., 1999
). The purpose of this study was to elicit the complete life history of the genus Acrosiphonia in southern British Columbia, rather than to detail life history events among species. More specifically, we investigated (1) seasonal abundance of two species of Acrosiphonia and of the sporophytes, ‘Codiolum’ and ‘Chlorochytrium,’ (2) reproductive phenology of both phases, and (3) when the sporophytic endophytes colonize their hosts and how long they remain within their hosts.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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‘Chlorochytrium’
‘Chlorochytrium’ cells were identified in Mazzaella splendens (Setchell and Gardner) Fredericq, the most abundant foliose red alga at Sooke. Seasonal abundance of ‘Chlorochytrium’ was based on density estimates of cells within M. splendens blades. Thirty-two M. splendens blades were collected, alternating between the largest and smallest (>5 cm) blade, from each quadrant of eight random points along a 12-m transect. An area, between 0.2 and 0.8 m above zero tidal level, was sampled each time. Collection occurred approximately monthly, except on sampling dates where tides were not low enough to expose M. splendens.
Mazzaella splendens blades were examined fresh. Drying of blades was not feasible, because shrunken ‘Chlorochytrium’ cells did not adequately hydrate; frozen blades often deteriorated. Although large cells of ‘Chlorochytrium’ (120 µm) could be seen in the field by holding thin, nonepiphytized blades of M. splendens up to the light, magnification was usually required to detect the bright green endophytic cells. For blades with <100 cells (or where single cells were scattered), individual cells could be counted, but densely colonized patches of endophytes needed to be estimated. Estimates were done by counting the number of groups of 
100 cells. Estimates were checked by actually counting five groups of 100 estimated cells from a blade where 1000 cells and 10 000 cells were estimated. This procedure was replicated five times. Absolute numbers of ‘Chlorochytrium’ cells were recorded per blade, as were frequency and range of cell size. To calculate ‘Chlorochytrium’ densities per square centimeter, M. splendens blades were photocopied and the surface area determined as a percentage of the mass of a sheet of paper of known area.
‘Codiolum’
‘Codiolum’ cells were present in ‘Petrocelis franciscana,’ a conspicuous red algal crust at the study site. Seasonal abundance of ‘Codiolum’ was determined by counting numbers of cells present in patches of ‘Petrocelis.’ Monthly, 30 small patches of 5 x 5 mm were randomly scraped by razor blade from ‘Petrocelis' growing on boulders within the low-to-high intertidal zone (from 0.1 to 5.1 m above zero tidal level). ‘Codiolum’ cells were detected by gently squashing the 25-m2 ‘Petrocelis' patch onto a microscope slide. Density, ‘Codiolum’ cell size, polymorphism among cells, and reproductive state were recorded. Due to the extreme polymorphism exhibited by ‘Codiolum’ cells, cell size for each sampling date was based on vesicle size only (excluding the stalk) and recorded as either large (80 µm in length) or small (<80 µm in length). In the summer and fall when densities exceeded 10 000 cells/cm2, it was necessary to estimate numbers present in the field of view under the 10x objective lens. This method of estimation was verified by actually counting the cells visible in a field of view for ten different fields of view for five individual crust patches, i.e., the average cell count per field of view was used for the estimation.
Data analysis
Statistical analyses were performed using SPSS 9.0 for Windows (1999)
. Transformations were performed to reduce the Levene statistic, but did not reduce heterogeneity to nonsignificant levels. Nevertheless, since standard nonparametric tests are inherently less powerful than parametric tests (Zar, 1996
) and should not be used as a simple means to avoid the problem of unequal variances (Day and Quinn, 1989
), ANOVAs (which are generally robust to variance heterogeneity [Zar, 1996
]) were used. One-way ANOVAs and the Games-Howell post hoc test were performed on square-root transformed data to test for significant differences in Acrosiphonia, ‘Chlorochytrium,’ and ‘Codiolum’ abundance over time. The Games-Howell test is more powerful than other post hoc tests for unequal variances (Games, Keselman, and Rogan, 1983
) and is recommended when number of treatments is small and sample size 7 individuals (Day and Quinn, 1989
).
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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for plants in the Puget Sound region, Washington State, USA, and commonly used by other workers (Kjellman, 1893
; Collins, 1909
; Setchell and Gardner, 1920
; Scagel, 1966
). Those criteria we deemed dependable for distinguishing between species are thallus morphology, filament diameter, hooked branchlet type, and number of fertile cells in a branch (Table 1). DNA sequences of the ITS regions of specimens identified as A. arcta and A. mertensii from Sooke showed consistent base pair differences, thus supporting their status as distinct species (Sussmann et al., 1999
).
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Reproductive phenology was also different for Acrosiphonia arcta and A. mertensii. Twenty percent of A. mertensii plants were fertile in early March, with 100% reproductive plants sampled in mid-April (and variable numbers through to July/August). Fertile A. arcta plants (30%), on the other hand, were not found until the end of April (variable percentage reproductive through to July/August). In Acrosiphonia vegetative intercalary cells located either in side branches or in the main filament of the plant differentiate into gametangia. These gametangia change from dark green to brown as they reach maturity (Figs. 5–8), and evidence of gamete release is seen by empty gametangia (Fig. 3). Here we refer to fertile thalli as those where brown gametangia were visible.
‘Chlorochytrium’ seasonal abundance
Seasonal differences of ‘Chlorochytrium’ density (Fig. 10) were statistically significant (Table 2): from September to late March densities were significantly lower (P < 0.05) than densities in April/May 1997 (sampling days 177/204 from start) and June 1998 (sampling day 576) and coincide with Acrosiphonia absence. No other significant differences were detected with the post hoc test performed on square-root transformed data. Peak densities of ‘Chlorochytrium’ (53 cells/cm2 or 5573 cells per blade in 1998) occurred in May (sampling days 177 and 545), 1 mo after establishment of Acrosiphonia.
‘Chlorochytrium’ morphology and location in host
‘Chlorochytrium’ cells were found embedded among the cells of Mazzaella splendens. In transverse section they were consistently located in the cortical layer of the blade (Fig. 17). Cell morphology ranged from spherical (most common) to ovoid to rarely elongated (Figs. 13–15), evidently due to the surrounding host tissue differentially impeding growth. These elongate cells were found in very thin tetrasporophytic blades collected in winter 1998. Cell size ranged from <40 to 300 µm. A net-like chloroplast and numerous pyrenoids were visible in vegetative cells (Figs. 13–14). Based on a small number of observations of blades collected in late February 1998 and of November 1998 blades maintained in a seawater tank for observation, fertile cells tended to (1) form protuberances that extended toward the surface of the host (Fig. 15), (2) darken at the apex, (3) change from bright green to olive colored, and (4) become homogeneous and bumpy with chloroplasts and pyrenoids no longer distinguishable. A single ‘Chlorochytrium’ cell, 160 µm in diameter, from material collected in late February, was observed releasing >800 zoospores (Fig. 16). The zoospores were 5 µm in diameter and possessed red eyespots and four equal length flagella.
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‘Codiolum’ seasonal abundance
The ‘Codiolum’ density curve (Fig. 11) supported by an ANOVA (Table 2) clearly illustrates seasonality. Unlike ‘Chlorochytrium’ abundance, ‘Codiolum’ densities peaked in late June in 1997 (sampling day 234 from start) and in mid-July in 1998 (day 619). This is 2 mo later than ‘Chlorochytrium’ peaked and 2–3 mo after Acrosiphonia establishment. The highest peak ‘Codiolum’ density of 22 400 cells/cm2 of ‘Petrocelis' occurred in 1998.
‘Codiolum’ morphology and location in host
‘Codiolum’ cells were found attached to and completely embedded in the filamentous system of ‘Petrocelis franciscana’ (Figs. 17–19). They are generally distinguished from ‘Chlorochytrium’ by their differentiation into an ovoid vesicle and colorless stalk. However, extreme polymorphism exhibited by individual cells included ‘Codiolum’ cells where the stalk was lacking. In general, the ovoid vesicle was slender (120–160 x 30 µm, 50–80 x 15 µm) or fat (100 x 60 µm, 80 x 40 µm, 50 x 30 µm); the stalk relatively long (70–100 µm), short (20–50 µm), or nonexistent (Figs. 18–23). Vegetative cells varied from yellow-green to bright green. The presence of oil droplets obscured any internal structures, e.g., chloroplast and pyrenoids. Although no fertile cells were observed discharging zoospores, mature ‘Codiolum’ were identified by darkening of the cell and division of the entire contents of the cell into spores 5 µm in length. Fertile cell vesicles were generally 80 µm in length and 40 µm in width, and reproductive phenology of ‘Codiolum’ coincided with that of ‘Petrocelis,’ from late fall to spring (Fig. 18).
‘Codiolum’ size
Large ‘Codiolum’ cells (many of them fertile) were consistently most abundant during fall and winter, whereas only small cells were found in ‘Petrocelis' patches in the spring. Summer sampling revealed variable numbers of large and small cells, but no fertile cells.
A simplified sporophyte timeline, illustrated in Fig. 24, integrates the dynamics of ‘Chlorochytrium’ and ‘Codiolum.’ Large, fertile cells are present in fall and winter; zoospores are released in winter/early spring; (Acrosiphonia germinates and becomes fertile); endophyte colonization and growth take place throughout the spring and summer.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Acrosiphonia identification in the field
Distinguishing among Acrosiphonia species in the field is problematic, primarily due to the difficulty in identifying juveniles. The branching pattern of A. arcta was generally not evident until plants were >2 cm in height, and compound hooks (a hooked branchlet upon another hooked branchlet, Fig. 4) tended not to be developed on very young A. mertensii. Furthermore, some mature A. arcta plants were found without any hooked branchlets. Others were found with short branchlets beginning to form on simple hooked branchlets and so resembled compound hooks, a diagnostic character of A. mertensii. Hudson (1974)
concluded that the presence or absence of hooked branchlets is a character that should not be given much weight in distinguishing among plants with otherwise similar appearance. Furthermore, light intensity and day length were found to affect such characters as cell diameter, length to width ratios, and amount of branching (Kornmann, 1965
; Hudson, 1974
). Consequently we gave weight to habitat and overall morphology in distinguishing species in the field.
Factors affecting Acrosiphonia seasonal abundance and distribution
Seasonal abundance of Acrosiphonia documented in this study (March–August) is consistent with the fact that only spring and summer collections of plants are reported in the literature (Fan, 1959
; Jónsson, 1959a, 1964, 1986
; Kornmann, 1970a, b
; Hudson, 1974
). It is not unusual that A. mertensii plants were found in October in California (Fan, 1959
), because specimens were also found in early October 1997 from Seppings Island (Barkley Sound), British Columbia (Sussmann, 2000
).
Temperature and photoperiod are important factors in the germination and growth of Acrosiphonia. Hudson (1974)
found that in culture upright filaments of Acrosiphonia sprouted from rhizoid pieces under long-day (16:8 photoregime) conditions, but not under short-day (8:16) conditions at the same temperatures. Also, at 15°C, A. arcta plants grew not as well as at 5°C and 10°C under the same photoregime, and sporophytes (‘Chlorochytrium’ or ‘Codiolum’) died before division into zoospores could occur. Miyaji (1996)
found this to also be true for A. spiralis plants collected in Japan. Vegetative growth of A. mertensii, on the other hand, was only inhibited at 20°C. Furthermore, when both A. arcta and A. mertensii plants were grown under low-light intensity (below 9.2 µmol · m–2 · s–1), motile gametes were not produced. These studies suggest longer days in the spring trigger germination of Acrosiphonia, increased light intensity plays a role in the production of viable gametes, and the effects of high summer temperatures contribute to the mortality of Acrosiphonia.
Variable seasonal abundance and distribution of Acrosiphonia were observed during the study period at different sites in southwestern British Columbia (Sussmann, 2000
). Any number of microhabitat factors such as salinity, nutrient levels, herbivore species, and abundance may result in this difference. In addition, variable endophyte survivorship, variable dispersal of endophytes, drift of reproductive Acrosiphonia thalli, and variable survivorship of overwintering rhizoids of Acrosiphonia may play a role. It is still unproven whether Acrosiphonia gametophytes can overwinter by means of rhizoids (as suggested by Scagel, 1966
), and, if they do, how important this is to the maintenance of Acrosiphonia's population. Purely vegetative propagation by rhizoids was shown under short-day conditions in culture (Hudson, 1974
). Yet the inability to detect rhizoids after disappearance of the upright plant and establishment of Acrosiphonia in new areas from year to year lead us to speculate that overwintering of the filamentous gametophyte via rhizoids may not be very important in Acrosiphonia's life history at our study site.
Dispersal patterns of gametes, spores, or propagules of marine algae have been little studied (Anderson and North, 1966
; Amsler and Searles, 1980
; Kendrick and Walker, 1991, 1995
). The appearance of Acrosiphonia arcta plants at Kitsilano Beach, 49°34' N, 123°10' W, Vancouver, British Columbia, despite there being no hosts present for their sporophyte (Sussmann, 2000
), suggests that dispersal of sporophytes, zoospores, or reproductive Acrosiphonia thalli may be long range, e.g., from an established site populated by Acrosiphonia and its endophytic sporophyte 6 km away. This also implies that free-living sporophytes may survive and overwinter, perhaps in rock crevices. A number of researchers (Jónsson, 1959a, b, 1962, 1966
; Kornmann, 1961, 1964
; Chihara, 1969
; Hudson, 1974
; Miyaji, 1996
) have demonstrated the ability of ‘Chlorochytrium’ and ‘Codiolum’ to survive free-living in culture. On the other hand, vegetative propagation of Acrosiphonia by overwintering rhizoids of the filamentous gametophyte may be important in maintenance of populations in habitats devoid of hosts for ‘Chlorochytrium’ and ‘Codiolum’ (such as at Kitsilano Beach, Vancouver).
Acrosiphonia arcta and A. mertensii differences in the field
The two Acrosiphonia species, A. arcta and A. mertensii, can occupy the same habitat, but show variable patterns of abundance and reproductive phenology. Hudson's culture work (1974)
showed different effects of environmental factors such as temperature on the two species: A. mertensii tolerates higher temperatures than A. arcta. Hudson (1974)
noted that A. mertensii persisted longer than A. arcta in the intertidal zone of the Puget Sound region, Washington State, and that A. arcta had not been reported south of Washington State. Although A. arcta did not consistently disappear from the intertidal zone earlier than A. mertensii at our site, A. arcta's abundance decreased in the high intertidal zone and increased in the low intertidal zone as the summer progressed. However, A. arcta was always the dominant species higher in the intertidal zone, suggesting that in southwestern British Columbia, temperature tolerance alone does not determine species distribution and abundance. Contrary to observations in the field, in culture A. arcta became fertile before A. mertensii (Hudson, 1974
). Fertile A. mertensii plants were found at Sooke, British Columbia, almost immediately after establishment of plants, whereas fertile A. arcta were not detected until the following sampling date.
Endophyte colonization
Colonization of ‘Codiolum’ and ‘Chlorochytrium’ was deduced by examining both cell density and cell size over time. An important assumption is that the smallest cells represent the youngest cells. Based on cell density increase (and the monthly sampling regime), the onset of ‘Chlorochytrium’ colonization occurred in late April 1 mo after Acrosiphonia establishment and 1 mo earlier than for ‘Codiolum’ (Figs. 9–11). This is somewhat counterintuitive, since A. mertensii became fertile before A. arcta, and its sporophyte appears to colonize only ‘Petrocelis' in British Columbia (Sussmann et al., 1999
). The greatest abundance of smallest cells of ‘Chlorochytrium’ and ‘Codiolum’ in spring also implies onset of colonization. Kornmann (1964)
also found the greatest abundance of small ‘Codiolum’ cells (within Haemescharia hennedyi from Germany) in late spring, 4–6 wk after reproductive Spongomorpha lanosa (Roth) Kützing plants were detected. The presence of the smallest ‘Chlorochytrium’ and ‘Codiolum’ cells from spring to early fall in the present study implies colonization is continuous and synchronized with Acrosiphonia gamete release. High densities of ‘Codiolum’ within ‘Petrocelis' from Washington State in summer (Dethier, 1987
) supports spring/summer colonization. The time required and the mechanism for zygote/endophyte penetration of Mazzaella splendens and ‘Petrocelis' remain poorly understood. More detailed culture and microscopy studies are needed to shed light on these events and to better understand the time-lag between Acrosiphonia gamete release and endophyte establishment.
Endophyte growth, fertility, and duration in host
Growth of ‘Chlorochytrium’ and ‘Codiolum’ is rapid, judging by the range of cell sizes observed from one sampling date to the next. Most ‘Chlorochytrium’ cells first observed in early spring were 40 µm in diameter, but cells up to 120 µm were also already present. Less than 2 mo later cells of 200–240 µm were commonly found. Setchell and Gardner (1920)
, Scagel (1966)
, Chihara (1969)
, and Hudson (1974)
described ‘Chlorochytrium inclusum’ cells from the northeast Pacific with a size range between 75 and 100 µm. Many of the collections were, however, made early in the spring. ‘Chlorochytrium inclusum,’ endophytic in Farlowia (Rhodophyta) from Japan, and associated with three Acrosiphonia species unknown to southwestern British Columbia, was also reported to be 
100 µm in diameter (Miyaji and Kurogi, 1976
). Kjellman's (1883)
description of ‘C. inclusum’ within the arctic alga Sarcophyllis arctica Kjellman (Rhodophyta) is the only case where ‘Chlorochytrium’ cells are noted to reach 275 µm, in agreement with the size attained by ‘Chlorochytrium’ cells in Mazzaella splendens in this study.
‘Chlorochytrium’ fertility and zoospore release occur in winter/early spring (Fig. 24) coinciding with Mazzaella splendens reproductive phenology (90–100% of of M. splendens blades collected from October to February were reproductive). Kjellman (1883)
found ‘Chlorochytrium’ fertile within Sarcophyllis arctica in winter, but did not observe zoospore release (unlike this study, however, he noted that ‘Chlorochytrium’ was most abundant in winter). Kornmann (1964)
also collected fertile ‘Chlorochytrium’ (from Polyides, a foliose red seaweed, in Helgoland, Germany) in the winter, just months before Spongomorpha lanosa establishment. ‘Chlorochytrium’ of 80–100 µm in diameter, obtained from Schizymenia (a bladed red seaweed) in Washington State, became fertile in culture and gave rise to 32, 64, or more zoospores 10 µm long (Chihara, 1969
). Unlike Chihara's fertile ‘Chlorochytrium,’ the few fertile cells detected in material collected from Sooke were much larger, 160–200 µm, and >800 zoospores, 5 µm long, were released from one mature sporophyte.
Juvenile cells of ‘Codiolum,’ first detected in ‘Petrocelis' in spring, had a vesicle size of 40–50 µm x 10–20 µm. Due to polymorphism among cells, it is difficult to comment on growth. Fertile ‘Codiolum,’ generally 80 µm in vesicle length, with the width at least one-half the length, were detected in ‘Petrocelis' samples collected from fall through early spring. ‘Codiolum’ fertility coincides with the reproductive phenology of its host (from October/November to February/March). Hanic (1965)
found fertile ‘C. petrocelidis' in reproductive ‘Petrocelis franciscana’ collected from Sooke in December 1963, and Kornmann (1961, 1964)
reported fertile ‘Codiolum’ associated with Spongomorpha lanosa in reproductive Haemescharia hennedyi from Helgoland in December 1963 and February 1964.
The absence of large endophyte cells in spring, after zoospore release, leads us to believe ‘Chlorochytrium’ and ‘Codiolum’ cells spend <1 yr in their host. This is in contrast with Kornmann's (1961, 1964)
finding for ‘Codiolum’ in Helgoland, Germany: a mixture of ‘Codiolum’ cell sizes were found together in Haemescharia hennedyi at the time of ‘Codiolum’ colonization, indicating some cells may spend 18 mo in their host, not releasing zoospores until the second winter. It seems unlikely that the very small cells observed in our study in greatest abundance in the spring and summer were comprised of both newly colonized cells and cells that had colonized ‘Petrocelis' the previous spring or summer. Furthermore, seasonal fluctuations in abundance of both ‘Petrocelis' and Mazzaella splendens (Sussmann, 2000
) potentially affect endophyte survival. A decrease in Mazzaella density was noted in the winter, primarily due to winter storms dislodging blades; loss of tissue of ‘Petrocelis' crusts also occurred in (fall) winter and is attributed to a combination of herbivory, adverse environmental conditions such as rainfall, water temperature, and light levels and tissue decay from senescence. Release of endophyte zoospores appears to coincide with the decrease in host abundance, in which case the endophytes spend 1 yr in their host. In the case of ‘Chlorochytrium,’ M. splendens blade dislodgement prior to endophyte maturity may be compensated by the survival of ‘Chlorochytrium’ in drift blades (Sussmann, 2000
) and endophytes persisting in perennial basal crusts of Mazzaella not dislodged. ‘Codiolum’ survivorship may be affected by the ability of cells present in ‘Petrocelis' tissue consumed by limpets to survive partial digestion. Defecation of still viable seaweeds is known from sea urchins (Santelices, Correa, and Avila, 1983
) and molluscs (Jernakoff, 1985
). Also, an epilithic or planktonic existence for the endophytes would contribute to the survival of ‘Chlorochytrium’ and ‘Codiolum.’
The role the endophytic sporophytes play in Acrosiphonia's complex life history is still poorly understood, i.e., it is unknown whether the sporophyte phase is actually required for the maintenance of Acrosiphonia populations in southwestern British Columbia. Although evidence is lacking in the field, Acrosiphonia may successfully recycle through rhizoid overwintering of gametophytic plants or by gametophytic plants giving rise to haploid zoospores (shown in culture [Kornmann, 1962
]) that survive the winter. The relative contribution of sexual reproduction (and the role of sporophytes) may be less significant compared to asexual reproduction and/or vegetative propagation in Acrosiphonia's life history. In some algae, e.g., the brown alga Analipus japonicus (Harvey) Wynne, and the coralline alga, Lithothrix aspergillum Gray, loss of sexual reproduction may actually occur at the northern range of the species (Nelson, 1980
; DeWreede and Vandermeulen, 1988
).
Despite the above unanswered questions, it is evident that the evolution of Acrosiphonia's alternation of heteromorphic generations has enabled this alga to persist in an environment where it normally could not. Acrosiphonia is clearly successful in southwestern British Columbia, as indicated by its abundance and the appearance of filamentous plants in new areas from year to year. The morphologically different gametophyte and sporophyte phases show strong seasonal differences, i.e., variable tolerance to environmental extremes, and occupy different habitats, i.e., free-living vs. endophytic condition. One can only speculate on the adaptive significance of the sporophyte evolving an endophytic condition: protection from herbivory, less light and nutrient limitation, decreased competition from other algal species, and prevention of overgrowth. We propose that Acrosiphonia's complex alternation of heteromorphic generations is a bet-hedging strategy. Not only have two morphologically different phases adapted to a seasonally variable evnvironment, but the endophytic sporophyte successfully colonizes both foliose and crustose red algae, and ‘Chlorochytrium’ and ‘Codiolum’ appear to have evolved to synchronize their duration as endophytes with host availability.
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FOOTNOTES |
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2 Author for reprint requests (phone: 604-822-6785, fax: 604-822-6089, dewreede{at}interchange.ubc.ca
).
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LITERATURE CITED |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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