CHARACTERISTICS OF SEXUAL AND ASEXUAL RESTING CYST (STATOSPORE) FORMATION IN DINOBRYON CYLINDRICUM IMHOF (CHRYSOPHYTA)1

The reproductive significance of siliceous cysts (statospores) produced by the common vernal chrysophyte Dinobryon cylindricum Imhof has been investigated under defined culture conditions. Three types of statospores have thus far been induced in culture: 1) uninucleate, asexual; 2) binucleate, asexual (potentially autogamic); 3) binucleate, sexual (zygotic). The production of each type of cyst responds differently to an array of nutrient deficiencies (P, N, vitamins, micrometals). An individual clone may be capable of participating in the production of all or only a subset of these types of resting cysts. All D. cylindricum statospores are morphologically identical regardless of their reproductive significance. Sexual reproduction leading to zygotic statospore formation is anisogamous, heterothallic, and involves a gametogenic hormone (erogen) that is apparently continuously released from female clones. Only a single bipolar mating group is documented here and clonal compatibility varies considerably within the mating group. The dynamics of encystment for each type of statospore has been determined relative to the growth of the vegetative cell population. Statospores may be produced either during the exponential phase (intrinsic encystment) or stationary phase (extrinsic encystment) of culture growth depending on the clones involved. The effect of both asexual and sexual resting cyst production on the net growth rate and dynamics of natural chrysophyte populations is discussed. Statospores appear to represent a more flexible reproductive strategy than the resistant zygospores produced by the other common groups of planktonic microalgae.     

ENCYSTMENT OF THE DINOFLAGELLATE GYRODINIUM UNCATENUM: TEMPERATURE AND NUTRIENT EFFECTS1

Sexual reproduction and encystment of the marine dinoflagellate Gyrodinium uncatenum Hulburt were induced in nitrogen and phosphorus-limited batch cultures. Sexuality did not occur under nutrient-replete conditions even when growth rate was reduced by non-optimal temperatures. Growth was optimal over a broader temperature range than encystment and virtually no cysts were produced at some low and high temperatures where growth occurred. Most cells initiated sexuality as intracellular pools of each limiting nutrient reached minimum or subsistence levels as much as four days after extracellular nutrients were exhausted. High nitrogen cell quotas during the phosphorus experiment indicate that sexuality was induced by a shortage of phosphorus and not by an indirect effect on nitrogen uptake. Total cyst yield corresponded to successful encystment of 9–13% of the motile populations, yet 60–85% of the plateau-phase motile cells were planozygotes (swimming zygotes formed from fusing gametes). Batch culture studies monitoring total cyst yield may thus seriously underestimate the extent of sexuality. More importantly, the number of cysts produced in a dinoflagellate population may be significantly reduced by environmental factors acting on the cells after sexual induction and fusion.     

A STUDY OF THE SEXUAL REPRODUCTION AND DETERMINATION OF MATING TYPE OF GYMNODINIUM NOLLERI (DINOPHYCEAE) IN CULTURE1

Sexual reproduction of Gymnodinium nolleri ( Ellegaard & Moestrup 1999 ) was studied by intercrossing experiments in all combinations of six clonal strains and backcrossing of five clonal F1 offspring. The results indicated that the conjugation of G. nolleri responded to the existence of more than two sexual types (complex heterothallism) and that compatibility between progeny of one cyst (inbreeding) was the rule. Sexual fusion, planozygote formation and development, cyst formation, and germination and planomeiocyte division were followed using time‐lapse photography. This study revealed many similarities between the sexual stages and life cycle pattern of G. nolleri and the related G. catenatum and the existence under culture conditions of an alternative cycle between vegetative cells and zygotes without a hypnozygote stage. The fate of zygotes, division or encystment, was influenced by the nutritional status of the external medium. The division of G. nolleri planozygotes was promoted by high levels of external nutrients, whereas the maximum percentage of encystment was recorded when phosphates were reduced in the isolation medium. The division of zygotes might be different from both vegetative and planomeiocyte division because it resulted in two‐cell chains with the cells not oriented in parallel.     

海洋浮游纤毛虫包囊的研究进展*

作为微型浮游动物的重要组成部分,海洋浮游纤毛虫是连接微食物环和经典食物链的重要中介。有些浮游纤毛虫在生活周期中会形成包囊,条件适宜时包囊会萌发,这对纤毛虫种群动态有重要的意义。目前国际上对于浮游纤毛虫生态学的研究主要集中在其营养期细胞,浮游纤毛虫包囊的研究尚少,中国还没有这方面的研究。本文对浮游纤毛虫包囊研究进展进行概述,包括包囊的形态、沉积物中包囊的丰度、包囊形成的环境因素、包囊萌发过程及环境因素对包囊萌发的影响等方面,希望对国内开展浮游纤毛虫包囊的研究有所裨益。     

Synchronization of encystment of <Emphasis Type="BoldItalic">Scrippsiella lachrymosa</Emphasis> (Dinophyta)

The encystment of Scrippsiella lachrymosa cells (strain B-10), which can be induced reliably in encystment medium, was inhibited by stirring the culture. 100 mL cultures in glass beakers were stirred at 1 rotation s⁻¹. Stirring inhibited vegetative cells from congregating (swarming) at the walls of the culture container. When stirring was stopped, a rapid induction of sexual reproduction was seen. As soon as stirring stopped (within 2 min), cells were observed swarming near the edges of the glass beaker. Four days after cessation of stirring, large percentages of the cells were mating and, after 7 days, most were zygotes. Cultures were observed after 31, 38, and, 45 days of stirring. When cultures were stirred for 45 days, cysts developed in the stirred treatments, but these cysts were attached to flocculent material that had also formed in the medium. The use of this laboratory method is advantageous for the study of the mating through cyst stages of the dinoflagellate life history. This method may also demonstrate the need for a ‘surface’ as a place for the dinoflagellate to congregate in order to successfully encyst and may help explain environmental observations of encystment at pycnoclines.     

SEXUAL PROCESSES IN THE LIFE CYCLE OF GYRODINIUM UNCATENUM (DINOPHYCEAE): A MORPHOGENETIC OVERVIEW1

Sexual processes in the life cycle of the dinoflagellate Gyrodinium uncatenum Hulburt were investigated in isolated field populations. Morphological and morphogenetic aspects of gamete production, planozygote formation, encystment, excystment, and planomeiocyte division are described from observations of living specimens, Protargol silver impregnated material and scanning electron microscope preparations. The sexual cycle was initiated by gamete formation which involved two asexual divisions of the vegetative organism. Gametes were fully differentiated following the second division and immediately capable of forming pairs. Either isogamous or anisogamous pairs were formed by the mid-ventral union of gametes. Gametes invariably joined with flagellar bases in close juxtaposition. Complete fusion of gametes required ca. 1 h, involved plasmogamy followed by karyogamy and resulted in a quadriflagellated planozygote. Planozygotes encysted in 24–48 h to yield a hypnozygote capable of overwintering in estuarine sediments. Hypnozygotes collected from sediment in late winter readily excysted upon exposure to temperatures above 15°C. A single quadriflagellated planomeiocyte emerged from the cyst and under culture conditions divided one to two days later. The four flagella were not evenly distributed at the first division and both bi- and tri-flagellated daughter cells were formed.     

Sexual and vegetative phases in the planktonic diatom Pseudo-nitzschia multistriata

The planktonic diatom Pseudo-nitzschia multistriata (Takano) Takano is known to produce the toxin domoic acid and it is recorded during late summer and autumn in the Gulf of Naples (Tyrrhenian Sea, Italy). We describe the sexual cycle of this species and report information on the variability of growth and cell size reduction rates at different experimental conditions. We induced sexual reproduction by crossing monoclonal cultures of opposite mating type. P. multistriata has a heterothallic life cycle that follows the general pattern reported for other congeneric species. Sexual stages were detected in cultures with an average apical length between 55 and 39 μm. The size of the initial cells produced at the end of the sexual phase was comprised between 72 and 82 μm and the lower cell size detected in culture was 26 μm. Sexual reproduction was thus recorded within a size window corresponding to 39–71% of the maximum cell apical length. Both growth performances and cell size reduction rates depend on cell size. The largest cells showed slower growth rates and larger size reduction rates at each division, while the relationship was opposite for cells smaller then 60% of the maximum size.     

Experimental sexual selection in Chlamydomonas

Sexual and asexual lines of the unicellular chlorophyte Chlamydomonas reinhardtii were propagated for about 100 sexual cycles and 1000 vegetative cycles in contrasted environments, liquid and solid growth media, in order to generate divergent natural and sexual selection. Sexual lines were transferred by many zygotes or by a single zygote in each sexual generation. By the end of the experiment zygote production was in the order sexual mass-transfer>sexual single-zygote>asexual>ancestor. The direct response to sexual selection was large, with zygote production increasing by about two orders of magnitude, mainly because mating had become spontaneous instead of being invoked by nitrogen starvation. Asexual lines became sexually sterilized by the fixation of a single mating type. Sexual selection caused a radical shift in the gender system, with homothallism spreading to high frequency in all sexual lines of this normally heterothallic species. This may have been caused by the transposition of a mating-type gene to an autosome. No substantial degree of environment-specific mating evolved, however, and thus no sexual isolation indicative of incipient speciation. It is possible that selection experiments of this kind are unlikely to induce sexual isolation because mating-type genes evolve in a saltatory fashion.     

Entamoeba invadens: the requirement for galactose ligands during encystment

During periods of stress, trophozoites of Entamoeba invadens (strain IP-1) undergo a process of differentiation (encystment) that results in a dormant cyst with a chitin-containing cyst wall. Encystment can be induced by resuspension of trophozoites from growth medium into a diluted glucose-free medium (47% LG) containing 5% adult bovine serum (ABS). ABS is thought to be a source of gal-terminated ligands that are required for high levels of encystment. After resuspension of trophozoites in 47% LG, encystment cultures were examined every 2h for responses to the (i) addition of 10mM free-galactose, (ii) resuspension of cells to serum-free medium, (iii) and dilution of encysting cultures to cell densities below that known to support full encystment (from 5 x 10(5) to 1 x 10(4)cells/ml). The role of serum components (and the gal-terminated ligand asialofetuin; ASF) adsorbed onto the surface upon which encystment proceeds, and their effect on the multi-cellular aggregation patterns formed during encystment, were also investigated. The addition of free-galactose reduced the levels of encystment (compared with the control) even when added at 10h after resuspension of trophozoites in 47% LG. The requirement for the presence of ABS during encystment was lost within 6h, with levels of encystment of cells washed free of serum reaching 80% of the control. The ability of cells to encyst when diluted to a cell density below that normally thought to support encystment reached over 50% by 8h. Efficient encystment could be obtained in 47% LG in the absence of ABS or ASF using pre-treated glass culture tubes. Encystment (47% LG; 5% ABS) using ultra low attachment plates was poor, suggesting attachment of cells to a surface via gal-terminated ligands was important for efficient encystment. The results suggest that ABS is probably not the only source of gal-terminated ligands necessary for high levels of encystment in 47% LG. While serum may provide a source of ligands which enhance the levels of encystment initially, other gal-terminated ligands possibly released by the encysting cells are still required for the completion of the encystment process and the formation of mature cysts. In addition, the gal-terminated ligands necessary for encystment efficiency may be adsorbed onto the glass surface of culture tubes and aid the initial aggregation process, as well as be involved in cell signaling during the encystment process.     

Sexual conflict and ecology: Species composition and male density interact to reduce male mating harassment and increase female survival

Sexual conflict is a pervasive evolutionary force that can reduce female fitness. Experimental evolution studies in the laboratory might overestimate the importance of sexual conflict because the ecological conditions in such settings typically include only a single species. Here, we experimentally manipulated conspecific male density (high or low) and species composition (sympatric or allopatric) to investigate how ecological conditions affect female survival in a sexually dimorphic insect, the banded demoiselle (Calopteryx splendens). Female survival was strongly influenced by an interaction between male density and species composition. Specifically, at low conspecific male density, female survival increased in the presence of heterospecific males (C. virgo). Behavioral mating experiments showed that interspecific interference competition reduced conspecific male mating success with large females. These findings suggest that reproductive interference competition between con‐ and heterospecific males might indirectly facilitate female survival by reducing mating harassment from conspecific males. Hence, interspecific competitors can show contrasting effects on the two sexes thereby influencing sexual conflict dynamics. Our results call for incorporation of more ecological realism in sexual conflict research, particularly how local community context and reproductive interference competition between heterospecific males can affect female fitness.     

Encystment of Peridinium gatunense: occurrence, favourable environmental conditions and its role in the dinoflagellate life cycle in a subtropical lake

1. The abundance of cysts of the bloom‐forming dinoflagellate Peridinium gatunense in the sediments of Lake Kinneret and the effects of environmental conditions on encystment were studied in relation to bloom dynamics. Peak cyst formation coincided with the highest growth rate of the population, prior to bloom peak. 2. Peridinium cysts were counted in water and sediment corer samples from 2000 to 2003 and in archived sediment trap samples collected during 1993–94. The cyst data were examined in relation to ambient temperature and nutrient records, and revealed no direct correlation. 3. In laboratory encystment experiments with Peridinium cells collected from the lake, 0.2–3% of the vegetative cells encysted. Temperature, light and cell density had no significant effect on the percentage of encystment. 4. Cysts were always present in the lake sediments but their abundance in ‘non Peridinium’ years was much lower than after a massive bloom. Vegetative cells were always present in the water column after the collapse of the annual dinoflagellate bloom, potentially serving as the inoculum for the next bloom. We propose that the hardy cysts serve as an emergency ‘gene bank’ to initiate population build up following catastrophic die outs.     

MORPHOLOGICAL VARIABILITY IN POPULATIONS OF CHRYSOPHYCEAN RESTING CYSTS. I. GENETIC (INTERCLONAL) AND ENCYSTMENT TEMPERATURE EFFECTS ON MORPHOLOGY1

Many chrysophycean species produce resting cysts (statospores) with purportedly species-specific morphology. I investigated variation in the cyst morphology of a single species that may result from genetic differences among the vegetative clones involved and from variation in the temperature of the environment during cyst development. Populations of Dinobryon cylindricum Imhof cysts were produced under defined conditions in vitro and then sampled for morphological analysis based on SEM micrographs. Morphological data is presented and then used in a multivariate discriminant analysis to determine the utility of each morphological character in distinguishing the six populations studied. Results suggest that some features of cyst morphology (i.e. cyst diameter) are invariant among the populations, while other features show distinctive variation. The density of spines covering the cyst body as well as the morphology of those spines appear correlated to the specific clones involved, and thus may represent useful phenotypic genetic markers. The length and definition of both the spines and the cyst collar, on the other hand, are markedly influenced by encystment temperature. The implications of these findings for paleoecological studies is discussed.     

Life cycle of the cereal aphid Sitobion avenae F.: polymorphism and comparison of life history traits associated with sexuality

1. Clones of Sitobion avenae from the west of France exhibited four types of responses in growth chambers to short days and low temperatures: in addition to permanently parthenogenetic clones (anholocyclic) and clones producing both males and parthenogenetic females (androcyclic), lineages producing both mating females and males were categorized into holocyclic and intermediate types, the latter investing into both sexual and parthenogenetic reproduction.
2. Holocyclic, intermediate and androcyclic clones were tested outdoors for sexual production in autumn and results were consistent with those obtained in growth chambers. Eggs from holocyclic and intermediate clone selfings were laid and hatched for most crosses.
3. Reciprocal crosses within and between life cycle variants showed no effect of the type of cross on the number of eggs laid per mating female, their hatching success, or the survival and fecundity of the parthenogenetic females born from eggs (fundatrices).
4. Eggs from intermediate × intermediate crosses hatched 20 days earlier than those involving at least one holocyclic parent. It is suggested that this short diapausing time gives a selective advantage to intermediate clones in oceanic climates, which could compensate for their lower sexual production in autumn.
5. Selfed offspring showed inbreeding depression, which seems to be common in non host-alternating aphids.
6. Mating with androcyclic males affects percentage (lower) and time (longer) of egg hatching, but not the other investigated traits.
7. The significance of intragenotypic strategy mixing in androcyclic and intermediate clones is discussed as an overall adaptation to the uncertainty of winter climate in oceanic zones.     

SEXUALITY AND CYST FORMATION IN THE DINOFLAGELLATE GONYAULAX TAMARENSIS: CYST YIELD IN BATCH CULTURES1

Encystment of the toxic dinoflagellate Gonyaulax tamarensis Lebour (var. excavata) was monitored in batch cultures exposed to a variety of nutritional and environmental treatments. Limitation by nitrogen (as ammonium or nitrate) or phosphorus (as phosphate) resulted in cyst formation. When the initial concentration of limiting nutrient was varied, total cyst yield (mL^?1) was directly proportional to the cell yield at all but the highest nutrient concentrations (where encystment was minimal). Encystment efficiency was relatively constant (0.1–0.2 cysts · cell^?1) over a 5-fold range of cell densities, indicating that 20 to 40% of the vegetative populations successfully encysted. Cyst formation was negligible in nutrient-replete medium, even with a significant reduction in growth rate due to non-optimal light, temperature, or to high batch culture cell densities. Low light levels did decrease cyst yield once encystment was initiated by nutrient limitation, but this was probably linked to smaller motile cell yield and not to a specific inhibition of encystment. In contrast, encystment was more sensitive to temperature than was growth rate: optimal cyst production occurred over a relatively narrow temperature range and no cysts were formed at [Page missing]     

First evidences of sexual selection by mate choice in marine zooplankton

Sexual selection is potentially important in marine zooplankton, presumably the most abundant metazoans on earth, but it has never been documented. We examine the conditions for sexual selection through mate choice and describe mating preferences in relation to size in a marine zooplankter, the pelagic copepod Acartia tonsa. Males produce spermatophores at a rate (~1 day⁻¹) much lower than known female encounter rates for most of the year and the decision to mate a particular female thus implies lost future opportunities. Female egg production increases with female size, and males mating larger females therefore sire more offspring per mating event. Similarly, females encounter males more frequently than they need to mate. Large males produce larger spermatophores than small males and the offspring production of female increases with the size of the spermatophore she receives. Additionally, large spermatophores allow females to fertilize eggs for a longer period. Thus, mating with large males reduces the female’s need for frequent matings and she may sire sons that produce more offspring because size is heritable in copepods. Finally, we show that both males and females mate preferentially with large partners. This is the first demonstration of sexual selection by mate choice in a planktonic organism.     

Cyst formation and poly-beta-hydroxybutyric acid accumulation in Azotobacter

Stevenson, L. H. (Louisiana State University, Baton Rouge), and M. D. Socolofsky. Cyst formation and poly-beta-hydroxybutyric acid accumulation in Azotobacter. J. Bacteriol. 91:304-310. 1966.-The relationship between cyst formation and the accumulation of poly-beta-hydroxybutyric acid (PHB) in Azotobacter vinelandii (A. agilis) was investigated. After various periods of growth, the cells were harvested, and the amount of PHB and the extent of encystment were determined. The polymer content of the cells increased sharply and reached a maximum on the 2nd day of growth followed by a gradual decline as the culture aged. At maximal accumulation, the PHB content was 35% of the dry weight, and the PHB-nitrogen ratio was 11:1. Those substrates promoting the highest polymer content (glucose, butanol) also promoted 95 to 100% encystment. Manipulation of the carbon and nitrogen supply in the medium indicated that both the maximal PHB content and the extent of cyst formation could be controlled. A direct correlation was noted between the amount of polymer accumulated and the percentage of cysts formed, indicating a possible role of PHB as a carbon or energy source, or both, for the encystment process.     

The life history of the toxic marine dinoflagellate Protoceratium reticulatum (Gonyaulacales) in culture

Asexual and sexual life cycle events were studied in cultures of the toxic marine dinoflagellate Protoceratium reticulatum. Asexual division by desmoschisis was characterized morphologically and changes in DNA content were analyzed by flow cytometry. The results indicated that haploid cells with a C DNA content occurred only during the light period whereas a shift from a C to a 2C DNA content (indicative of S phase) took place only during darkness. The sexual life cycle was documented by examining the mating type as well as the morphology of the sexual stages and nuclei. Gamete fusion resulted in a planozygote with two longitudinal flagella, but longitudinally biflagellated cells arising from planozygote division were also observed, so one of the daughter cells retained two longitudinal flagella while the other daughter cell lacked them. Presumed planozygotes (identified by their longitudinally biflagellated form) followed two life-cycle routes: division and encystment (resting cyst formation). Both the division of longitudinally biflagellated cells and resting cyst formation are morphologically described herein. Resting cyst formation through sexual reproduction was observed in 6.1% of crosses and followed a complex heterothallic pattern. Clonal strains underwent sexuality (homothallism for planozygote formation and division) but without the production of resting cysts. Ornamental processes of resting cysts formed from the cyst wall under an outer balloon-shaped membrane and were fully developed in <1 h. Obligatory dormancy period was of ∼4 months. Excystment resulted in a large, rounded, pigmented, longitudinally biflagellated but motionless, thecate germling that divided by desmoschisis. Like the planozygote, the first division of the germling yielded one longitudinally biflagellated daughter cell and another without longitudinal flagella. The longitudinal biflagellation state of both sexual stages and of the first division products of these cells is discussed.     

Selection of low investment in sex in a cyclically parthenogenetic rotifer

Cyclical parthenogens, which combine asexual and sexual reproduction, are good models for research into the ecological and population processes affecting the evolutionary maintenance of sex. Sex in cyclically parthenogenetic rotifers is necessary for diapausing egg production, which is essential to survive adverse conditions between planktonic growing seasons. However, within a planktonic season sexual reproduction prevents clonal proliferation. Hence, clones with a low propensity for sex should be selected, becoming dominant in the population as the growing season progresses. In this context, we studied the dynamics of the heritable variation in propensity for sexual reproduction among clones of a Brachionus plicatilis rotifer population in a temporary Mediterranean pond during the period the species occurred in plankton. Clonal isolates displayed high heritable variation in their propensity for sex. Moreover, the frequency of clones with low propensity for sex increased during the growing season, which supports the hypothesized short‐term selection for low investment in sex within a growing season. These results demonstrate (1) the inherent instability of the cyclical parthenogenetic life cycle, (2) the cost of sexual reproduction in cyclical parthenogens where sex produces diapausing eggs and (3) the role of the association between sexual reproduction and diapause in maintaining sex in these cyclical parthenogens.     

Sexual reproduction and diversity: Connection between sexual selection and biological communities via population dynamics

Sexual reproduction is a mysterious phenomenon. Most animals and plants invest in sexual reproduction, even though it is more costly than asexual reproduction. Theoretical studies suggest that occasional or conditional use of sexual reproduction, involving facultative switching between sexual and asexual reproduction, is the optimal reproductive strategy. However, obligate sexual reproduction is common in nature. Recent studies suggest that the evolution of facultative sexual reproduction is prevented by males that coerce females into sexual fertilization; thus, sexual reproduction has the potential to enforce costs on a given species. Here, the effect of sex on biodiversity is explored by evaluating the reproductive costs arising from sex. Sex provides atypical selection pressure that favors traits that increase fertilization success, even at the expense of population growth rates, that is, sexual selection. The strength of sexual selection depends on the density of a given species. Sexual selection often causes strong negative effects on the population growth rates of species that occur at high density. Conversely, a species that reduces its density is released from this negative effect, and so increases its growth rate. Thus, this negative density-dependent effect on population growth that arises from sexual selection could be used to rescue endangered species from extinction, prevent the overgrowth of common species and promote the coexistence of competitive species. Recent publications on sexual reproduction provide several predictions related to the evolution of reproductive strategies, which is an important step toward integrating evolutionary dynamics, demographic dynamics and community dynamics.