fucus radicans and fucus vesiculosus; fucales ... - Semantic Scholar

J. Phycol. 47, 990–998 (2011) 2011 Phycological Society of America DOI: 10.1111/j.1529-8817.2011.01032.x

FREQUENT CLONALITY IN FUCOIDS (FUCUS RADICANS AND FUCUS VESICULOSUS; FUCALES, PHAEOPHYCEAE) IN THE BALTIC SEA 1 Kerstin Johannesson,2 Daniel Johansson, Karl H. Larsson, Cecilia J. Huenchun~ir Department of Marine Ecology – Tja¨rno¨, University of Gothenburg, SE-452 96 Stro¨mstad, Sweden

Jens Perus Department of Environmental and Marine Biology, A˚bo Akademi University, FI-20520 A˚bo, Finland

Helena Forslund, Lena Kautsky Department of Botany, Stockholm University, SE-106 91 Stockholm, Sweden

and Ricardo T. Pereyra Department of Marine Ecology – Tja¨rno¨, University of Gothenburg, SE-452 96 Stro¨mstad, Sweden

Asexual reproduction by cloning may affect the genetic structure of populations, their potential to evolve, and, among foundation species, contributions to ecosystem functions. Macroalgae of the genus Fucus are known to produce attached plants only by sexual recruitment. Recently, however, clones of attached plants recruited by asexual reproduction were observed in a few populations of Fucus radicans Bergstro¨m et L. Kautsky and F. vesiculosus L. inside the Baltic Sea. Herein we assess the distribution and prevalence of clonality in Baltic fucoids using nine polymorphic microsatellite loci and samples of F. radicans and F. vesiculosus from 13 Baltic sites. Clonality was more common in F. radicans than in F. vesiculosus, and in both species it tended to be most common in northern Baltic sites, although variation among close populations was sometimes extensive. Individual clonal lineages were mostly restricted to single or nearby locations, but one clonal lineage of F. radicans dominated five of 10 populations and was widely distributed over 550 · 100 km of coast. Populations dominated by a few clonal lineages were common in F. radicans, and these were less genetically variable than in other populations. As thalli recruited by cloning produced gametes, a possible explanation for this reduced genetic variation is that dominance of one or a few clonal lineages biases the gamete pool resulting in a decreased effective population size and thereby loss of genetic variation by genetic drift. Baltic fucoids are important habitat-forming species, and genetic structure and presence of clonality have implications for conservation strategies.

Key index words: asexual reproduction; clonal richness; macroalgae; population genetic structure; salinity gradient Abbreviations: MLG, multilocus genotype; PSU, practical salinity units

Asexual reproduction by means of cloning is common in eukaryotic species not least among groups of multi- and unicellular plants and algae (reviewed by Ellstrand and Roose 1987 and Silvertown 2008). Indeed, clonality has evolved repeatedly in various lineages of Tree of Life but mostly fails to persist over long evolutionary timescales (Bengtsson 2009). Different hypotheses have been put forward to explain the evolution of clonality, such as polyploidy, hybridization, physiological stress in marginal environments, and clones being favored by high general fitness and ecological specialization (reviewed in Jokela et al. 2003, Silvertown 2008, Bengtsson 2009). Specifically, asexual reproduction has been suggested as an evolutionary successful strategy in marginal environments of species’ distributions where immigration of individuals with suboptimal phenotypes is relatively more common than elsewhere (Peck et al. 1998). Asexual reproduction has the immediate consequence for species with separate sexes that average reproductive output is substantially increased, with strong implications on the potential of population growth. The observations of short-term success hence contrast with the observed long-term failure of most clonal lineages and explain why these issues are still under debate (see Shcherbakov 2010). The absence of genetic recombination in most asexual lineages (but see Gladyshev et al. 2008 for a notable exception) is suggested as a key factor (Feldman et al. 1997), and recent models have shown that recombination leads

1

Received 25 May 2010. Accepted 14 January 2011. Author for correspondence: e-mail kerstin.johannesson@marecol. gu.se. 2

990

991

CLONALITY IN BALTIC FUCOIDS

to purging of deleterious alleles that otherwise become associated with alleles that contribute to positive fitness, and in this way sexual reproduction will be favored over clonality (Barton and Otto 2005, Keightley and Otto 2006). Interestingly, most species that mainly recruit asexually occasionally switch to sexual recruitment (Silvertown 2008) and maintain genetic variation comparable to fully sexual species (Balloux et al. 2003, Bengtsson 2003). In addition, they retain a capacity for recombination of favorable alleles (Bengtsson 2009). We need to understand the evolution of clonality as well as its genetic and ecological consequences. Under which circumstances, for example, do obligate sexually reproducing evolutionary lineages evolve capacities to reproduce asexually, and how does this affect the population genetic structure, the genetic variation, and the potential for evolutionary change? In addition, genetic changes of key-ecosystem species may have important effects at the ecosystem level (Reusch et al. 2005, Whitham et al. 2006, Reusch and Wood 2007), and detailed analyses of genetic structure of ecosystem key species are therefore warranted. The Baltic Sea is a 377,000 km2 large, semienclosed brackish-water basin with a stable salinity gradient increasing from 2 practical salinity units (PSU) in the north to 20–25 PSU in the entrance to the North Sea. The sea is a postglacial environment, and it has existed in its present form for no more than 8,000 years. Species of both marine and freshwater origin have colonized the sea, but still the Baltic Sea has a noticeably low diversity of most groups of organisms. For example, only 20% of North Atlantic macroalgal species are present inside the Baltic Sea (Wallentinus 1991, Nielsen et al. 1995). Among macrophytes (macroalgae and seagrasses) that have been able to enter the Baltic Sea, there is a trend toward asexual reproduction being more common inside than outside the Baltic Sea (Wallentinus 1991, Gabrielsen et al. 2002, Olsen et al. 2004). A recent discovery found a small proportion of clonal reproduction in populations of bladderwrack (F. vesiculosus) inside the Baltic Sea (Tatarenkov et al. 2005). This is particularly interesting, since this group of alga (fucoids) was earlier known to always produce new attached plants from zygotes. Furthermore, the newly discovered and closely related species F. radicans showed a dominance of asexually produced thalli in two populations at the Swedish Baltic Sea coast (Bergstro¨m et al. 2005, Tatarenkov et al. 2005). F. radicans is endemic to the Baltic Sea and most likely originated only a few thousand years ago, or less, from F. vesiculosus inside the postglacial marine Baltic Sea (Pereyra et al. 2009). Both species have the capacity to reproduce asexually by means of small vegetative fragments that get loose from the thallus, drift and attach to the substrate by forming new rhizoids, and grow

into sexually mature individuals (Tatarenkov et al. 2005). These two species are the only fucoids of the northern Baltic Sea (Bergstro¨m et al. 2005), and they provide an important habitat for small invertebrate species, fish larvae, and epiphytic macroalgae (Ra˚berg and Kautsky 2007, Wikstro¨m and Kautsky 2007). The aim of the present study was to provide a comprehensive description of the occurrence of clonality in these two macroalgae species inside the Baltic Sea providing a platform for further investigations of its evolutionary and ecological implications. The Baltic fucoid system seems to be of particular interest for studies of evolution of asexual reproduction and its consequences because: (i) These species represent a systematic group that is rarely observed to have asexual reproduction of fully outgrown and attached plants (fucoid macroalgae), and clonality is hitherto only reported in one distinct (and environmentally marginal) area, the Baltic Sea. (ii) The Baltic Sea is a young postglacial environment, and the evolution of clonality is hence potentially a very recent event (20.0 6.7 5.0 5.0 3.8 3.5 4.0 4.3 5.2 5.5 5.6 5.8 5.2

Absent Absent 48 49 30 48 50 50 50 44 15 25 Absent

42 43 48 51 34 Absent Absent 16 Not sampled Not sampled 9 Absent 23

June salinities are averaged over 10 years (see text for details). PSU, practical salinity units.

A

15

B

25

65

H

D

E

I

C

K

D

J

B

M

K

A

F

G

E

H

60

C

L

0

km

200

FIG. 1. Distribution of unique and multiple (clonal) multilocus genotypes (MLGs) over populations of Fucus radicans (A) and F. vesiculosus (B) in the Baltic Sea. Gray segments indicate total proportion of unique MLGs in each site, black and white segments show MLGs found in two or more copies (clones) but unique to the particular site, and colored segments show MLGs distributed over more than one site. Sample sizes (number of thalli analyzed) are indicated by black and white frames.

using Viogene’s Plant Genomic DNA Extraction Miniprep System (Viogene, Sunnyvale, CA, USA), according to the manufacturer’s protocol. Samples were genotyped at nine microsatellite loci (L20, L38, L58, L85, L94: Engel et al. 2003; and Fsp1, Fsp2, Fsp3, Fsp 4: Perrin et al. 2007). These markers have proved reliable in earlier studies in the discrimination of Fucus clones (Tatarenkov et al. 2005, Pereyra et al. 2009). PCRs of L20, L38, L58, L94, and Fsp4 were performed in a 10 lL mixture containing 1.0 lL 10· PCR Buffer, 0.2 lL of 10 mM dNTP mix (2.5 mM of each nucleotide), 0.3 lL of 25 mM MgCl2, 0.5 lL of each 10 lM forward and reverse primer, 0.3 U Taq, and 1.6 lL of DNA template. The PCR mix for L85 had 0.01 lg BSA also added and DNA template was increased to 3.2 lL in an 11.5 lL PCR volume. The amplified fragments were separated in a capillary automated sequencer (CEQ8000, Beckman Coulter Inc., Fullerton, CA, USA) and sized using CEQ software (version 8.0.52; Beckman Coulter Inc.). All

individuals sampled were successfully genotyped in most sites, but total sample size varied owing to low densities of fucoids in some sites. In two Finnish sites (I and J), F. vesiculosus was not sampled, although it may have been present. The reason was that these samples were originally sampled for another purpose (Table 1). Data analysis. The software GENCLONE 2.0 (ArnaudHaond and Belkhir 2007) was used to assign each thallus into its putative genet (a group of thalli that originally arose by the same sexual event). GENCLONE estimates in a round-robin fashion the probability that sampling units share the same genotype as a product of different sexual events (by chance, contrary to a clonal event). It also estimates the probability of clonal identity taking into account the FIS to (i) check for possible scoring errors whenever individual pair-wise genetic distances are unusually low and (ii) compute the frequency distribution of these genetic distances that may screen for

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R ¼ ðG 1Þ=ðN 1Þ

ð1Þ

where G indicates the number of genets, and N the number of thalli sampled. Hence, R ranges between zero (a monoclonal population) and 1 (each individual is a genetically unique multilocus genotype [MLG]). Thus, clonal richness indicates the ratio between the number of multilocus genotypes and the number of sampled thalli in a given population. We also calculated allelic richness (the number of alleles per population) of each population as this estimate has a high relevance in conservation genetics and can be used to detect populations for conservation (Petit et al. 1998). However, as allelic richness is dependent on sample size, we calculated allelic richness using the rarefaction method (Hurlbert 1971) as implemented in HP-RARE (Kalinowski 2005) that compares populations at similar size.

RESULTS

The distribution of clonality. In total, 170 MLGs were found among the 10 populations of F. radicans, and 143 (84%) of these were unique MLGs (only one thallus of all thalli sampled carried a particular MLG), while 27 (16%) were multicopy MLGs. All multicopy MLGs were most likely the result of asexual reproduction. Indeed, probabilities that thalli of the same genotype were derived from different sexual events were low (psex < 0.05) for all genotypes in all populations that were not dominated by one or a few genotypes (see below). In populations dominated by one or a few clonal lineages, psex values increased dramatically (close to 1.0) for the common clones (while remained low for rare clones) as a consequence of low statistical power in samples dominated by a few clonal lineages (see ArnaudHaond et al. 2007). Hence, our conclusion is that new copies of the same clonal lineage have been formed by release of fragments (adventitious branches) from which new thalli are formed. Unique MLGs, on the other hand, may either be single representatives of rare clonal lineages, so rare that by chance only one thallus of each lineage was sampled, or they are the result of sexual reproduction. Three of the F. radicans clonal lineages were common with 11, 24, and 161 thalli each, while the rest were rare with two to six thalli each. In the eight populations of F. vesiculosus, we found 230 different MLGs where 213 (93%) of these were unique and 17 (7%) were clones. One clonal lineage was present as 12 thalli from two adjacent sites, but the other clones were relatively rare with only two to four thalli of each. The number of clonal lineages varied considerably among populations for both species (Fig. 1),

and clonal richness increased from north to south (correlations between clonal richness and latitude) in F. radicans (R2 = 0.39, P = 0.05), while in F. vesiculosus there was a tendency of a similar pattern (R2 = 0.46, P = 0.06). In addition, clonal richness increased with average surface salinity for June in F. radicans (R2 = 0.62, P = 0.007) while not in F. vesiculosus (R2 = 0.34, P = 0.17) (Fig. 2). However, both species also unveiled strong variation among adjacent localities or localities at similar salinities without any discernable pattern. For example, we found very few clonal lineages in Estonian populations (K–M), while extensive clonality was present in ¨ regrund (C) and Djursten (D) at both species at O similar salinities as in Estonia. Moreover, two populations of F. vesiculosus only 3 km apart (Djursten, D ¨ regrund, H) had very different clonal richness and O (0.88 and 0.47) despite similar salinities. Most multicopy clonal lineages, that is, 19 of 27 in F. radicans and 16 of 17 in F. vesiculosus, were only detected in one location. Four additional clonal lineages in F. radicans and one in F. vesiculosus had restricted distributions and occurred only in neighboring locations. Only two F. radicans clones were found in three or more sites. Most noteworthy was the distribution of one of these F. radicans clonal lineages that was extensively distributed and present in several localities at both sides of the northern Baltic Sea (Fig. 1). This clonal lineage was dominant in four of the samples (60%–90% of the thalli) and relatively common in two additional populations (12% and 39%). Thalli of this clone are all females, develop receptacles, produce eggs, and can contribute to the formation of new germlings in laboratory crosses (H. Forslund pers. obs.). Two other clonal lineages of F. radicans were common at the northernmost Finnish location (Ha¨llkalla, G), and together they dominated this site with frequencies of 22% and 48% of the individuals. Both these lineages were males with receptacles producing sperm.

1

0.8 Clonal richness

somatic mutations. The distribution of genetic distances indicates if a given sampling unit (although slightly genetically different) is likely to be part of the same clonal lineage or not. Observed and expected genotype frequencies and deviations from expected Hardy–Weinberg distributions were calculated using the software GENEPOP (Rousset 2007) for each loci and population separately. We further calculated clonal richness (R) according to Ellstrand and Roose (1987) but with the modification suggested by Dorken and Eckert (2001):

0.6

0.4

0.2

0 3

3.5

4

4.5

5

5.5

6

6.5

7

Salinity (psu)

FIG. 2. Relationship between salinity and clonal richness of populations of Fucus radicans (crosses, dotted trend line, R2 = 0.62, P = 0.007) and F. vesiculosus (diamonds, broken trend line, R2 = 0.34, P = 0.17).

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K E R S TI N J O H A N N E S S O N E T A L .

Table 2. Number of populations for each species and loci that showed significant heterozygote deficiency or excess. Microsatellite locus

Heterozygote Heterozygote Heterozygote Heterozygote

deficiency (F. radicans) deficiency (F. vesiculosus) excess (F. radicans) excess (F. vesiculosus)

L20

L38

L58

L85

L94

Fsp1

Fsp2

Fsp3

Fsp4

0 0 1 0

2 0 1 1

0 0 0 2

0 0 2 1

1 0 0 0

3 3 0 0

3 6 0 0

2 5 2 0

0 1 1 1

0.6

0.8

The total number of populations analyzed were 10 for Fucus radicans and eight for F. vesiculosus. 3.5

Table 3. Number of loci for each population that showed significant heterozygote deficiency or excess. Het. excess

2 2 0 1 0 0 2 2 1 1

3 4 0 0 0 0 0 0 0 0

2 2 1 3 2 0 2 3

0 0 0 2 0 3 0 0

The total number of loci analyzed were nine for both species.

Genetic variation and clonality. Observed genotype frequencies of genet populations were in general in accordance with Hardy–Weinberg expectation in both species, but we found significant heterozygote deficiencies in three of the loci (Fsp1, Fsp2, and Fsp3). These deviations occurred in both species and seemed randomly distributed over populations suggesting loci-specific effects such as null alleles, but the data are inconclusive (Tables 2 and 3). Observed heterozygosities calculated for the complete population of ramets were heavily affected by the presence or absence of the dominant female clone as this clone is heterozygote in seven out of nine loci, and it caused extensive excess of heterozygotes in populations where it was common. Clonal richness varied extensively among populations, providing a strong case for a test if there was a relationship between prevalence of sexual recruitment (indicated by clonal richness) and amount of genetic variation in a population (indicated by allelic richness). We tested separately the two species and observed a significant positive relationship between clonal richness and allelic richness in F. radicans (Fig. 3) indicating that clonality in this

3 Allelic richness

F. radicans ¨ regrund C O Djursten D Bo¨nhamn E Ja¨rna¨s F Finland G Finland H Finland I Finland J Estonia K Estonia L F. vesiculosus Kristineberg A ¨ land B O ¨ regrund C O Djursten D Bo¨nhamn E Finland H Estonia K Estonia M

Het. deficiency

2.5

2

1.5 0

0.2

0.4

1

Clonal richness

FIG. 3. Relationship between clonal richness (indicating prevalence of sexual recruitment) and allelic richness (indicating genetic variation) for populations of Fucus radicans (crosses, dotted trend line, R2 = 0.69, P = 0.003) and F. vesiculosus (diamonds, broken trend line, R2 = 0.09, P = 0.5).

species is negatively correlated with genetic variation of populations. Despite no significant relationship in F. vesiculosus, the variation of allelic richness in F. vesiculosus was similar to that of F. radicans over the corresponding range of clonal richness (Fig. 3). Almost identical results were obtained if allelic richness was replaced by observed or expected heterozygosities as alternative proxies of genetic variation (data not shown). DISCUSSION

Both prevalence and distribution of clonality varied greatly in both species in the Baltic Sea. The dominance of asexual recruitment of thalli in the northwest populations contrasted markedly with the sexually recruited eastern and southern populations in F. radicans. In F. vesiculosus, the most important observation was that asexual recruitment of attached thalli is widespread in the Baltic Sea, while hitherto not reported from populations outside the Baltic Sea (see Tatarenkov et al. 2005, Pereyra et al. 2009, although comprehensive screening for clonality is missing in many areas). Increased prevalence of asexual reproduction is known from other species entering the Baltic Sea, such as the red alga Ceramium tenuicorne and the seagrass Zostera marina (Reusch et al. 2000, Gabrielsen et al. 2002, Bergstro¨m et al. 2003), and similar to the situation in F. radicans, Z. marina also unveils local difference


in the prevalence of clonality (Olsen et al. 2004). An important difference, however, is that both C. tenuicorne and Z. marina have the capacity to reproduce asexually also outside the Baltic Sea, but clonality is prevalent inside this sea. If it is a general trend that populations become increasingly clonal inside the Baltic Sea, this fits earlier observations from other habitats and areas that clonality tends to be more common in marginal environments than in central areas of species’ distributions (Eckert 2002, Kearney et al. 2006, Silvertown 2008). Mechanisms of cloning. Tatarenkov et al. (2005) reported development of rhizoids in loose adventitious branches followed by reattachment to the substratum. Notably, adventitious branches are frequent also on thalli of F. vesiculosus living outside the Baltic Sea, in areas where asexually recruited thalli are not found. Possibly, poor rhizoid formation, poor survival of small individuals, or difficulties in reattachment of fragments in tidal areas (the Baltic Sea is atidal) may explain the absence of asexually recruited thalli in the North Sea outside the Baltic Sea. Fucoids have problems with sexual reproduction in low salinities; sperm motility and velocity decrease substantially below 10 PSU (although sperm of Baltic F. vesiculosus are more tolerant to low salinity), and fertilization success drops rapidly below 6–8 PSU (Serraõ et al. 1996). In low salinities, the initial block to polyspermy breaks down as the egg membrane repolarizes too rapidly after penetration of the first sperm (Brawley 1991), and as expected, polyspermy levels are high in fucoids in the northern Baltic (Serraõ et al. 1999). Partial disability of sexual reproduction is likely to promote asexual alternatives of reproduction, in particular, in those regions of the Baltic Sea where salinities are the lowest, and hence it seems likely that clonality is favored by natural selection in low-saline areas. However, our result indicates that factors other than ambient salinity may be important, as populations living at similar salinities sometimes had very different degrees of clonality. An important issue for future studies is to determine the reason for the absence of clones in F. vesiculosus outside the Baltic Sea by testing, for example, if secondary rhizoid formation in adventitious branches is to any extent an inherited trait, or if it is induced by extreme environmental conditions. The dominant clone. The presence of one extensively large clone of F. radicans is intriguing. This clonal lineage has an extensive geographic distribution (550 · 100 km), and in four of six of the populations where it was present, it was numerically dominant with 40%–90% of sampled thalli belonging to this clone (Fig. 1). In fucoids, parts of a thallus may survive while drifting long distances, and this may help to explain the extensive distribution. Nevertheless, it seems likely that such a spatial and numerical dominance can only have been established over a long period of time. Moreover, earlier

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observations of populations dominated by this clonal lineage indicate that at least the majority of individuals produce viable eggs (Serraõ et al. 1999, in this study F. radicans was considered a northern dwarf form of F. vesiculosus). Furthermore, eggs of this clone have resulted in viable offspring in laboratory crosses (H. Forslund pers. obs.), suggesting that at least in part this clonal lineage may also contribute to sexual recruitment. Among terrestrial angiosperms, clones are rarely observed to be old and widespread (Ellstrand and Roose 1987, Wilk et al. 2009), and an explanation put forward is that deleterious mutations accumulate and eventually eliminate old clones by natural selection (‘‘mutational melt-down,’’ Lynch and Gabriel 1990). Nevertheless, Bengtsson (2009) argues that such ‘‘mutational melt-down’’ in itself is not a major problem since asexually reproducing individuals are also under natural selection and clones with low fitness will be eradicated. More likely, perhaps, is that persistence of the same clones over long time periods will require high environmental stability, as clone evolution will be restricted by rate of somatic mutations. Indeed, the finding of a very old clonal lineage of eelgrass (Z. marina) in the northern Baltic Sea (Reusch et al. 2000) suggests that over the past thousands of years, the Baltic environment may have been stable enough to host long-lived clones. In addition, long persistence and large size (wide distribution) of individual clones are likely to be a result of high fitness (Weeks and Hoffman 1998, Gardner and Mangel 1999). Although a direct relationship between heterozygosity of microsatellite loci and fitness is highly unlikely, positive correlations between marker heterozygosity and fitness of individuals can be expected as, for example, heterozygosity is likely to indicate level of inbreeding ⁄ outbreeding (Hansson and Westerberg 2002, Ha¨mmerli and Reusch 2003, Szulkin et al. 2010). However, although the largest clonal lineage found in F. radicans was more heterozygote (seven of nine loci), than was the average clone (five of nine loci), we found no significant correlation between clone heterozygosity and size of clone (R = 0.10, P = 0.6). Indeed, the two next largest clonal lineages (present in the Finnish site D) were among the least heterozygote ones with none and one heterozygote locus out of nine, respectively. Effects of clonality. There was no dramatic difference on overall level of genetic variation in Baltic populations of F. radicans and F. vesiculosus when compared to populations of F. vesiculosus outside the Baltic Sea and to populations of closely related Atlantic fucoids (Table 4; note that F. spiralis is hermaphrodite and capable of self-fertilization, which likely explains the much lower variation in this species, Engel et al. 2005). However, a high prevalence of asexual recruitment in some populations of the two Baltic fucoids clearly affected the genetic

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Table 4. Level of overall genetic variation (expected heterozygosity) in Fucus radicans and Baltic populations of F. vesiculosus, and in closely related fucoid populations and species. Species

Mode of reproduction

Fucus radicans F. vesiculosus

Dioecious Dioecious

F. spiralis

Hermaphrodite

F. serratus

Dioecious

Area

Number of populations

Expected heterozygosity

Reference

Baltic Sea Baltic Sea Baltic Sea North Sea ME, USA SW Europe SW Europe ME, USA North Sea

10 8 16 8 4 7 7 4 21

0.51 0.58 0.52 0.67 0.57 0.50 0.21 0.39 0.54

This study This study Tatarenkov et al. (2007) Tatarenkov et al. (2007) Wallace et al. (2004) Perrin et al. (2007) Perrin et al. (2007) Wallace et al. (2004) Coyer et al. (2003)

structure and variation compared to sexually recruiting populations. Not least the formation of a widespread and dominant female clone in F. radicans has changed the genetic structure and pattern of variation substantially in this species. For example, low clonal richness of populations is likely to have negative consequence on the potential to survive rapid environmental changes; there may be simply too few different genotypes present in a population to survive a period of extreme environmental stress. Furthermore, populations with low clonal richness also have fewer alleles than other populations, and this restricts the potential number of new genotypes that can be created by recombination during sexual reproduction, in this way impeding long-term evolvability of populations. In contrast to selfing or inbreeding, asexual reproduction is not in itself the direct cause of the observed loss of genetic variation (Ellstrand and Roose 1987, Wideń et al. 1994). Indeed, loss of genetic variation in asexually and sexually recruiting populations alike is set by the effective population size (Bengtsson 2009), and theoretical and modeling analyses show that effective population size is not expected to decrease in populations dominated by asexual recruitment (Balloux et al. 2003, Bengtsson 2003, but see Orive 1993 for a different result). Still our result indicated that populations of F. radicans with low clonal richness (dominance of asexual recruitment) had lost roughly ¼ of the genetic variation compared to populations dominated by sexual recruitment (see data in Fig. 3). Our suggestion is that this loss is a consequence of the fact that populations with low clonal richness are dominated by one or a few clones and that these clones contribute a large number of identical gametes. In populations dominated by one or a few clonal lineages, the production of gametes from the clone (or clones) will skew the contribution of gametes in a way that effectively decreases the effective population size. For example, the F. radicans populations with low clonal richness in our study had either a dominance of the large female clone (populations C–F and H) or was dominated by two male clones (G) in both cases creating gamete pools strongly dominated by a few genetic individuals (and in both these cases also of

the same sex), while gametes of unique MLGs will all be strongly underrepresented in numbers. Hence, the effective population size will be largely determined by very few of the MLGs present in the population. Metapopulation structure and gene flow among local populations will also affect genetic variation and distribution of individual alleles (Charlesworth 2003), and these relationships need further investigation. In addition, somatic mutations may contribute with new genetic variation; in particular, fitness of specific clones may be changed through the addition of positive or negative mutations and in this way influencing local adaptation and selection among clones. Cloning combined with sexual recruitment adds the possibility of transferring new mutations to different genetic backgrounds. Hence both selection among clonal lineages and selection among genes with a somatic origin may provide contributions to local adaptation of F. radicans and F. vesiculosus inside the Baltic Sea, and this as well needs further investigation. For both species, but in particular for the endemic F. radicans, the situation inside the Baltic Sea seems critical over the coming few hundred years, as salinity is expected to decrease with increased runoff from land (Meier 2006). If, for example, the dominant female clone will not withstand further changes, a large part of the species’ distribution will be affected. However, fucoids colonized the Baltic Sea 8,000 years ago, and surface salinity has decreased during this period of time from 15& to 6&–8&. Evidently, Baltic fucoids have evolved local adaptation in important traits such as sperm motility, freezing, desiccation tolerance, and thermal tolerance during this relatively short period of time (Serraõ et al. 1996, Pearson et al. 2000, Lago-Lestoń et al. 2010) indicating a capacity for evolutionary change during the recent past. Whether further adaptations are at all possible, and at what rates, will be critical to the species’ survival in the Baltic Sea, but the more genetic variation that is present, the larger the chance that further adaptation is possible. For this reason, management of Baltic fucoids should aim to optimize genetic variation by protecting populations of high clonal richness and


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