Characterization of the organellar genomes of Mazzaella laminarioides and Mazzaella membranacea (Gigartinaceae, Rhodophyta).

Mazzaella, a genus with no genomic resources available, has extensive distribution in the cold waters of the Pacific, where they represent ecologically and economically important species. In this study, we aimed to sequence, assemble, and annotate the complete mitochondrial and chloroplast genomes from two Mazzaella spp. and characterize the intraspecific variation among them. We report for the first time seven whole organellar genomes (mitochondria: OR915856, OR947465, OR947466, OR947467, OR947468, OR947469, OR947470; chloroplast: OR881974, OR909680, OR909681, OR909682, OR909683, OR909684, OR909685) obtained through high-throughput sequencing for six M. laminarioides sampled from three Chilean regions and one M. membranacea. Sequenced Mazzaella mitogenomes have identical gene number, gene order, and genome structure. The same results were observed for assembled plastomes. A total of 52 genes were identified in mitogenomes, and a total of 235 genes were identified in plastomes. Although the M. membranacea plastome included a full-length pbsA gene, in all M. laminarioides samples, the pbsA gene was split in three open reading frames (ORFs). Within M. laminarioides, we observed important plastome lineage-specific variations, such as the pseudogenization of the two hypothetical protein-coding genes, ycf23 and ycf45. Nonsense mutations in the ycf23 and ycf45 genes were only detected in the northern lineage. These results are consistent with phylogenetic reconstructions and divergence time estimation using concatenated coding sequences that not only support the monophyly of M. laminarioides but also underscore that the three M. laminarioides lineages are in an advanced stage of divergence. These new results open the question of the existence of still undisclosed species in M. laminarioides.

Impact of persistent barrier to gene flow and catastrophic events on red algae evolutionary history along the Chilean coast.

Historical vicariance events, linked to the existence of stable physical barriers to gene flow, generate concordant genetic breaks in co-distributed species while stochastic processes (e.g., costal uplift) could cause species-specific genetic breaks as a result of local strong demographic bottlenecks or extinction. In Chile, previous studies show that the area of the 30°S-33°S could correspond to a stable barrier to gene flow that have affected the genetic structure of various algae and marine invertebrates. Here we sequenced two organellar genes (COI and rbcL) in four taxonomically accepted co-distributed red seaweeds species characterized by a low dispersal potential: Mazzaella laminarioides, M. membranacea, Asterfilopsis disciplinalis, and Ahnfeltiopsis vermicularis. Our results revealed the existence of ten strongly differentiated linages in the taxa studied. Strong genetic breaks, concordant in both space and time (divergence estimated to have occurred some 2.9-12.4 million years ago), were observed between taxa distributed across the 33°S. Conversely, in the Central/South part of the Chilean coast, the localization of the genetic breaks/sub-structure observed varied widely (36°S, 38°S, 39°S, and 40°S). These results suggest that a major historical vicariance event has modeled the genetic structure of several Chilean marine organisms in the north of the Chilean coast during the mid-Miocene, while more recent stochastic events and genetic drift could be the driving forces of genetic divergence/structuration in the central-southern part of the coast.

The Rhodoexplorer Platform for Red Algal Genomics and Whole-Genome Assemblies for Several Gracilaria Species.

Macroalgal (seaweed) genomic resources are generally lacking as compared with other eukaryotic taxa, and this is particularly true in the red algae (Rhodophyta). Understanding red algal genomes is critical to understanding eukaryotic evolution given that red algal genes are spread across eukaryotic lineages from secondary endosymbiosis and red algae diverged early in the Archaeplastids. The Gracilariales is a highly diverse and widely distributed order including species that can serve as ecosystem engineers in intertidal habitats and several notorious introduced species. The genus Gracilaria is cultivated worldwide, in part for its production of agar and other bioactive compounds with downstream pharmaceutical and industrial applications. This genus is also emerging as a model for algal evolutionary ecology. Here, we report new whole-genome assemblies for two species (Gracilaria chilensis and Gracilaria gracilis), a draft genome assembly of Gracilaria caudata, and genome annotation of the previously published Gracilaria vermiculophylla genome. To facilitate accessibility and comparative analysis, we integrated these data in a newly created web-based portal dedicated to red algal genomics (https://rhodoexplorer.sb-roscoff.fr). These genomes will provide a resource for understanding algal biology and, more broadly, eukaryotic evolution.

A temporal and spatial study of genetic structure in four species of bladed Bangiales (Rhodophyta) from the southeastern Pacific coast of Chile.

The coastline is a heterogeneous and highly dynamic environment influenced by abiotic and biotic variables affecting the temporal stability of genetic diversity and structure of marine organisms. The aim of this study was to determine how much the genetic structure of four species of marine Bangiales vary in time and space. Partial sequences of the cytochrome oxidase I (COI) gene obtained from two Pyropia (Py. sp. CHJ and Py. orbicularis) and two Porphyra (P. mumfordii and P. sp. FIH) species were used to compare the effect of the 40° S/41° S biogeographic break (spatial-regional scale) and the one of the Valdivia River discharges (spatial-local scale) and determine their temporal stability. Four seasonal samplings were taken during 1 year at five sites, one site located in Melinka (Magallanes province) and four sites along the coast of Valdivia (Intermediate area), on both sides of the river mouth. Results showed a strong genetic spatial structure at regional scale (ΦST > 0.4) in Py. sp. CHJ, Py. orbicularis, and P. mumfordii, congruent with the 41° S/42° S biogeographic break. A potential barrier to gene flow, related to the Valdivia River discharge, was detected only in P. mumfordii. In P. sp. FIH, spatial genetic structure was not detected at any scale. The genetic structure of all four species is stable throughout the year. The potential effect of main currents and river discharge in limiting the transport of Bangiales spores are discussed. We propose that both a restricted propagule dispersal and the formation potential for persistent banks of microscopic stages could lead to a temporally stable spatial partitioning of genetic variation in bladed Bangiales.

The impact of Pleistocene glaciations and environmental gradients on the genetic structure of Embothrium coccineum.

The South American temperate forests were subjected to drastic topographic and climatic changes during the Pliocene-Pleistocene as a consequence of the Andean orogeny and glacial cycles. Such changes are common drivers of genetic structure and adaptation. Embothrium coccineum (Proteaceae) is an emblematic tree of the South American temperate forest (around 20°S of latitude) that has strongly been affected by topographic and climatic events. Previous studies have shown a marked genetic structure in this species, and distinct ecotypes have been described. Yet, little is known about their adaptive genetic responses. The main goal of this study was to investigate the effects of historical and contemporary landscape features affecting the genetic diversity and connectivity of E. coccineum throughout its current natural distribution. Using over 2000 single nucleotide polymorphisms (SNPs), we identified two genetic groups (a Northern and a Central-Southern group) that diverged around 2.8 million years ago. The level of genetic structure was higher among populations within the Northern genetic group than within the Central-Southern group. We propose that these differences in genetic structure may be due to differences in the assemblages of pollinators and in the evolutionary histories of the two genetic groups. Moreover, the data displayed a strong pattern of isolation by the environment in E. coccineum, suggesting that selection could have led to adaptive divergence among localities. We propose that in the Chilean temperate forest, the patterns of genetic variation in E. coccineum reflect both a Quaternary phylogenetic imprint and signatures of selection as a consequence of a strong environmental gradient.

Pre-domestication bottlenecks of the cultivated seaweed Gracilaria chilensis.

Gracilaria chilensis is the main cultivated seaweed in Chile. The low genetic diversity observed in the Chilean populations has been associated with the over-exploitation of natural beds and/or the founder effect that occurred during post-glacial colonization from New Zealand. How these processes have affected its evolutionary trajectory before farming and incipient domestication is poorly understood. In this study, we used 2232 single nucleotide polymorphisms (SNPs) to assess how the species' evolutionary history in New Zealand (its region of origin), the founder effect linked to transoceanic dispersion and colonization of South America, and the recent over-exploitation of natural populations have influenced the genetic architecture of G. chilensis in Chile. The contrasting patterns of genetic diversity and structure observed between the two main islands in New Zealand attest to the important effects of Quaternary glacial cycles on G. chilensis. Approximate Bayesian Computation (ABC) analyses indicated that Chatham Island and South America were colonized independently near the end of the Last Glacial Maximum and emphasized the importance of coastal and oceanic currents during that period. Furthermore, ABC analyses inferred the existence of a recent and strong genetic bottleneck in Chile, matching the period of over-exploitation of the natural beds during the 1970s, followed by rapid demographic expansion linked to active clonal propagation used in farming. Recurrent genetic bottlenecks strongly eroded the genetic diversity of G. chilensis prior to its cultivation, raising important challenges for the management of genetic resources in this incipiently domesticated species.

More than What Meets the Eye: Differential Spatiotemporal Distribution of Cryptic Intertidal Bangiales.

Morphologically similar but genetically distinct species have been termed cryptic and most have been assumed to be ecologically similar. However, if these species co-occur at a certain spatial scale, some niche differences at finer scales should be expected to allow for coexistence. Here, we demonstrate the existence of a disjointed distribution of cryptic bladed Bangiales along spatial (intertidal elevations) and temporal (seasons) environmental gradients. Bladed Bangiales were identified and quantified across four intertidal elevations and four seasons for one year, at five rocky intertidal sites (between 39° S and 43° S) in southern Chile. Species determination was based on partial sequences of the mitochondrial cytochrome c oxidase 1 (COI) gene amplification. To assess species gross morphology, thallus shape, color, and maximum length and width were recorded. Hundreds of organisms were classified into nine Bangiales species belonging to three genera (i.e., Fuscifolium, Porphyra, and Pyropia), including five frequent (>97% of specimens) and four infrequent species. All species, except for Pyropia saldanhae, had been previously reported along the coasts of Chile. The thallus shape and color were very variable, and a large overlap of the maximum width and length supported the cryptic status of these species. Multivariate analyses showed that the main variable affecting species composition was intertidal elevation. Species such as Py. orbicularis were more abundant in low and mid intertidal zones, while others, such as Po. mumfordii and Po. sp. FIH, were principally observed in high and spray elevations. Despite all numerically dominant species being present all year long, a slight effect of seasonal variation on species composition was also detected. These results strongly support the existence of spatial niche partitioning in cryptic Bangiales along the Chilean rocky intertidal zone.

Whole genome genotyping reveals discrete genetic diversity in north-east Atlantic maerl beds.

Maerl beds are vital habitats for a diverse array of marine species across trophic levels, but they are increasingly threatened by human activities and climate change. Furthermore, little is known about the genetic diversity of maerl-forming species and the population structure of maerl beds, both of which are important for understanding the ability of these species to adapt to changing environments and for informing marine reserve planning. In this study, we used a whole genome genotyping approach to explore the population genomics of Phymatolithon calcareum, a maerl-forming red algal species, whose geographical distribution spans the north-east Atlantic, from Norway to Portugal. Our results, using 14,150 genome-wide SNPs (single nucleotide polymorphisms), showed that P. calcareum maerl beds across the north-east Atlantic are generally structured geographically, a pattern likely explained by low dispersal potential and limited connectivity between regions. Additionally, we found that P. calcareum from the Fal Estuary, south-west England, is genetically distinct from all other P. calcareum sampled, even from The Manacles, a site located only 13 km away. Further analysis revealed that this finding is not the result of introgression from two closely related species, Phymatolithon purpureum or Lithothamnion corallioides. Instead, this unique diversity may have been shaped over time by geographical isolation of the Fal Estuary maerl bed and a lack of gene flow with other P. calcareum populations. The genomic data presented in this study suggest that P. calcareum genetic diversity has accumulated over large temporal and spatial scales, the preservation of which will be important for maximizing the resilience of this species to changes in climate and the environment. Moreover, our findings underline the importance of managing the conservation of maerl beds across western Europe as distinct units, at a site-by-site level.

Exploring the Genetic Consequences of Clonality in Haplodiplontic Taxa.

Partially clonality is an incredibly common reproductive mode found across all the major eukaryotic lineages. Yet, population genetic theory is based on exclusive sexuality or exclusive asexuality, and partial clonality is often ignored. This is particularly true in haplodiplontic eukaryotes, including algae, ferns, mosses, and fungi, where somatic development occurs in both the haploid and diploid stages. Haplodiplontic life cycles are predicted to be correlated with asexuality, but tests of this prediction are rare. Moreover, there are unique consequences of having long-lived haploid and diploid stages in the same life cycle. For example, clonal processes uncouple the life cycle such that the repetition of the diploid stage via clonality leads to the loss of the haploid stage. Here, we surveyed the literature to find studies that had genotyped both haploid and diploid stages and recalculated population genetic summary metrics for seven red algae, one green alga, three brown algae, and three mosses. We compared these data to recent simulations that explicitly addressed the population genetic consequences of partial clonality in haplodiplontic life cycles. Not only was partial clonality found to act as a homogenizing force, but the combined effects of proportion of haploids, rate of clonality, and the relative strength of mutation versus genetic drift impacts the distributions of population genetic indices. We found remarkably similar patterns across commonly used population genetic metrics between our empirical and recent theoretical expectations. To facilitate future studies, we provide some recommendations for sampling and analyzing population genetic parameters for haplodiplontic taxa.

Differential Frond Growth in the Isomorphic Haploid-diploid Red Seaweed Agarophyton chilense by Long-term In Situ Monitoring.

Conditional differentiation between haploids and diploids has been proposed to drive the evolutionary stability of isomorphic biphasic life cycles. The cost of producing and maintaining genetic information has been posed as a possible driver of this conditional differentiation. Under this hypothesis, haploids benefit over diploids in resource-limited environments by halving the costs of producing and maintaining DNA. Spared resources can be allocated to enhance survival, growth or fertility. Here we test in the field whether indeed haploids have higher growth rates than diploids. Individuals of the red seaweed Agarophyton chilense, were mapped and followed during 2 years with 4-month census intervals across different stands within the Valdivia River estuary, Chile. As hypothesized, haploids grew larger and faster than diploids, but this was sex-dependent. Haploid (gametophyte) females grew twice as large and 15% faster than diploids (tetrasporophytes), whereas haploid males only grew as large and as fast as the maximum obtained by diploids in summer. However, haploid males maintained their maximum sizes and growth rates constant year-round, while diploids were smaller and had lower growth rates during the winter. In conclusion, our results confirm the conditional differentiation in size and growth between haploids and diploids but also identified important differences between males and females. Besides understanding life cycle evolution, the dynamics of A. chilense frond growth reported informs algal farmers regarding production optimization and should help in determining best planting and harvesting strategies.

Evaluating the effects of ocean warming and freshening on the physiological energetics and transcriptomic response of the Antarctic limpet Nacella concinna.

In the Southern Ocean, warming and freshening are expected to be prominent signals of climate change and the reduced ability of Antarctic marine organisms to cope with changing environmental conditions could challenge their future survival. The Antarctic limpet Nacella concinna is a macroinvertebrate of rocky ecosystems, which occurs in high densities in the shallow subtidal zone. Subtidal individuals were exposed to a combination of temperatures (1, 4, 8, 11, 14 °C) and salinities (20 and 30 psu) for a 60-day period. A drastic increment in mortality was observed with seawater warming, showing that N. concinna is highly stenothermal, with limited ability to survive at temperatures warmer than 4 °C, although there was some degree of acclimation at 4 °C and ambient salinity (30 psu). This study confirmed the stenohaline characteristic of this species, with mortality reaching 50% and lower scope for growth at low salinity (20 psu) even at the control temperature (1 °C). At the sub-cellular level, limpets' low tolerance to out-of range salinity is illustrated by the activation of cell remodelling processes whereas the down-regulation of chaperones proteins and plasma membrane ATPase suggest that under the combination of warming and freshening N. concinna experiences a severe level of stress and devote much of its energy to somatic maintenance and survival. The drastic effect observed can be explained by its subtidal origin, an environment with more stable conditions. The surviving individuals at 1 °C and lowered salinity (20 psu) were either more tolerant or showing signs of acclimation after 60 days, but the combination of warming and freshening have a greater combined stress. Projections of climate change for end of the century for this part of the Antarctic can, therefore, result in a significant diminution of the subtidal population of N. concinna, affecting ecological interactions and diversity of the food web.

After a catastrophe, a little bit of sex is better than nothing: Genetic consequences of a major earthquake on asexual and sexual populations.

Catastrophic events can have profound effects on the demography of a population and consequently on genetic diversity. The dynamics of postcatastrophic recovery and the role of sexual versus asexual reproduction in buffering the effects of massive perturbations remain poorly understood, in part because the opportunity to document genetic diversity before and after such events is rare. Six natural (purely sexual) and seven cultivated (mainly clonal due to farming practices) populations of the red alga Agarophyton chilense were surveyed along the Chilean coast before, in the days after and 2 years after the 8.8 magnitude earthquake in 2010. The genetic diversity of sexual populations appeared sensitive to this massive perturbation, notably through the loss of rare alleles immediately after the earthquake. By 2012, the levels of diversity returned to those observed before the catastrophe, probably due to migration. In contrast, enhanced rates of clonality in cultivated populations conferred a surprising ability to buffer the instantaneous loss of diversity. After the earthquake, farmers increased the already high rate of clonality to maintain the few surviving beds, but most of them collapsed rapidly. Contrasting fates between sexual and clonal populations suggest that betting on strict clonality to sustain production is risky, probably because this extreme strategy hampered adaptation to the brutal environmental perturbation induced by the catastrophe.

Comparative phylogeography of two Agarophyton species in the New Zealand archipelago.

Molecular studies have reported the coexistence of two species of Agarophyton in New Zealand: the newly described A.transtasmanicum with an apparently restricted distribution to some sites in the North Island, and the more widespread A.chilense. Here, we compared the distribution, genetic diversity, and structure of both Agarophyton species throughout the archipelago using sequences of the nuclear Internal Transcribed Spacer 2 (ITS2) marker. Agarophyton chilense's distribution was continuous and extensive along the North and South Islands, Stewart Island, and Chatham Island, and the genetic clusters were mostly concordant with boundaries between biogeographic regions. In contrast, specimens of A.transtasmanicum were collected in four sites broadly distributed in both the North and South Islands, with no clear spatial structure of the genetic diversity. Populations, where the species co-occurred, tended to display similar levels in genetic diversity for the two species. Demographic inferences supported a postglacial demographic expansion for two A.chilense genetic clusters, one present in the South Island and the eastern coast of the North Island, and the other present in northern South Island. A third genetic cluster located on the western coast of the North Island had a signature of long-term demographic stability. For A.transtasmanicum, the skyline plot also suggested a postglacial demographic expansion. Last, we developed a new molecular tool to quickly and easily distinguish between the two Agarophyton species, which could be used to ease future fine-scale population studies, especially in areas where the two species coexist.

Better off alone? Compared performance of monoclonal and polyclonal stands of a cultivated red alga growth.

The objective of this study was to test, using a field experiment, the effect of genotypic diversity on productivity of farmed populations (Ancud and Chaica, Chile) of the domesticated red alga Agarophyton chilense (formerly known as Gracilaria chilensis), a species considered as economically important in Chile. Monoclonal and polyclonal (4 and 8 genotypes) subplots were outplanted into the mid intertidal in Metri Bay (Puerto Montt, Chile) during summer, a season in which A. chilense face higher temperatures (>18°C) and low nitrogen availability (<4.00 µmol). Ancud farm genotypes show higher growth rates in the monoclonal rather than the two polyclonal subplots. A similar tendency, yet not significant, was discernible in Chaica. In addition, whatever the population of origin of the thalli, no effect of genotypic diversity was detected neither on the agar yield and its quality, nor on the epiphyte load. Such unexpected results of a higher performance in plots with a lower genotypic diversity could be explained (a) by human-assisted selection for dominant-best-performing genotypes that could counterbalance the negative effect caused by the low genotypic diversity in farms and (b) by the fact that the organisms inhabiting the algal mats do not impact the fitness of their host. Overall, the results obtained here suggest that despite farm induced selection lead to impoverished pools of genotypes, they may also have a positive effect of on the resistance of farmed populations to seasonal stressors. However, whether this may have a secondary negative effect on the longer term in a fluctuating environment remains to be determined, but may be avoided by adopting strategy of selection favoring different genotypes in space and time, as implemented in forestry.

Post-Disturbance Genetic Changes: The Impact of the 2010 Mega-Earthquake and Tsunami on Chilean Sandy Beach Fauna.

Earthquake/tsunamis can have profound impacts on species and their genetic patterns. It is expected that the magnitude of this impact might depend on the species and the time since the disturbance occurs, nevertheless these assumptions remain mostly unexplored. Here we studied the genetic responses of the crustacean species Emerita analoga, Excirolana hirsuticauda, and Orchestoidea tuberculata to the 27F mega-earthquake/tsunami that occurred in Chile in February 2010. mtDNA sequence analyses revealed a lower haplotype diversity for E. analoga and E. hirsuticauda in impacted areas one month after the 27F, and the opposite for O. tuberculata. Three years after the 27F we observed a recovery in the genetic diversity of E. analoga and E. hirsuticauda and decrease in the genetic diversity in O. tuberculata in 2/3 of sampled areas. Emerita analoga displayed decrease of genetic differentiation and increase in gene flow explained by long-range population expansion. The other two species revealed slight increase in the number of genetic groups, little change in gene flow and no signal of population expansion associated to adult survival, rapid colonization, and capacity to burrow in the sand. Our results reveal that species response to a same disturbance event could be extremely diverse and depending on life-history traits and the magnitude of the effect.

Dual influence of terrestrial and marine historical processes on the phylogeography of the Brazilian intertidal red alga Gracilaria caudata.

In this study, we explored how past terrestrial and marine climate changes have interacted to shape the phylogeographic patterns of the intertidal red seaweed Gracilaria caudata, an economically important species exploited for agar production in the Brazilian north-east. Seven sites were sampled along the north-east tropical and south-east sub-tropical Brazilian coast. The genetic diversity and structure of G. caudata was inferred using a combination of mitochondrial (COI and cox2-3), chloroplast (rbcL) and 15 nuclear microsatellite markers. A remarkable congruence between nuclear, mitochondrial and chloroplast data revealed clear separation between the north-east (from 03° S to 08° S) and the south-east (from 20° S to 23° S) coast of Brazil. These two clades differ in their demographic histories, with signatures of recent demographic expansions in the north-east and divergent populations in the south-east, suggesting the maintenance of several refugia during the last glacial maximum due to sea-level rise and fall. The Bahia region (around 12° S) occupies an intermediate position between both clades. Microsatellites and mtDNA markers showed additional levels of genetic structure within each sampled site located south of Bahia. The separation between the two main groups in G. caudata is likely recent, probably occurring during the Quaternary glacial cycles. The genetic breaks are concordant with (i) those separating terrestrial refugia, (ii) major river outflows and (iii) frontiers between tropical and subtropical regions. Taken together with previously published eco-physiological studies that showed differences in the physiological performance of the strains from distinct locations, these results suggest that the divergent clades in G. caudata correspond to distinct ecotypes in the process of incipient speciation and thus should be considered for the management policy of this commercially important species.

Genetic and morphological differentiation of Porphyra and Pyropia species (Bangiales, Rhodophyta) coexisting in a rocky intertidal in Central Chile.

A recent molecular taxonomic study along the Chilean coast (18° S-53° S) described 18 candidate species of bladed Bangiales of which only two were formally described. Few studies focused on local genetic and morphological diversity of bladed Bangiales and attempted to determine their intertidal distribution in contrasting habitats, and none were performed in Chile. To delimit intertidal distributions of genetic species, 66 samples of bladed Bangiales were collected at Maitencillo (32° S) in four zones: a rocky platform, a rocky wall, and two boulders zones surrounded by sandy and rocky bottoms, respectively. These samples were identified based on sequences of the mitochondrial COI and chloroplast rbcL markers. We also collected 87 specimens for morphological characterization of the most common species, rapidly assessing their putative species identity using newly developed species-diagnostic (PCR-RFLP) markers. Eight microscopic and two macroscopic morphological traits were measured. We described and named three of four species that predominate in Maitencillo (including Pyropia orbicularis): Pyropia variabilis Zapata, Meynard, Ramírez, Contreras-Porcia, sp. nov., Porphyra luchea Meynard, Ramírez, Contreras-Porcia sp. nov., and Porphyra longissima Meynard, Ramírez, Contreras-Porcia, sp. nov. With the exception of Po. longissima restricted to boulders surrounded by sandy bottom, and a morphotype of Py. variabilis restricted to rocky walls, the other species/morphotypes have overlapping intertidal distributions. Except for Po. longissima, which is clearly differentiated morphologically (longest and thinnest blades), we conclude that morphology is not sufficient to differentiate bladed Bangiales. Our findings underscore the importance of refining our knowledge of intrinsic and environmental determinants on the distribution of bladed Bangiales.

Differentiation of haploid and diploid fertilities in Gracilaria chilensis affect ploidy ratio.

BACKGROUND: Algal isomorphic biphasic life cycles alternate between free-living diploid (tetrasporophytes) and haploid (dioicious gametophytes) phases and the hypotheses explaining their maintenance are still debated. Classic models state that conditional differentiation between phases is required for the evolutionary stability of biphasic life cycles while other authors proposed that the uneven ploidy abundances observed in the field are explained by their cytological differences in spore production. RESULTS: We monitored the state and fate of individuals of the red seaweed Gracilaria chilensis periodically for 3 years in five intertidal pools from two sites with distinct conditions. We tested for differentiation in fecundity and spore survival among the gametophyte males and females (haploids) and the tetrasporophytes (diploids). We tested for the influence of fecundity and spore survival on the observed uneven ploidy abundances in recruits. The probability of a frond becoming fecund was size-dependent, highest for the haploid males and lowest for the haploid females, with the diploids displaying intermediate probabilities. Fecund diploids released more tetraspores than carpospores released by the haploid females. Spore survival depended on ploidy and on the local density of co-habiting adult fronds. An advantage of diploid over haploid germlings was observed at very low and very high adult fronds densities. CONCLUSIONS: Neither spore production nor spore survival determined the highly variable ploidy ratio within G. chilensis recruits. This result invalidates the hypothesis of natural cytological differences in spore production as the only driver of uneven field ploidy abundances in this species. Diploid spores (carpospores) survived better than haploid spores (tetraspores), especially in locations and time periods that were associated with the occurrence of strong biotic and abiotic stressors. We hypothesise that carpospore survival is higher due to support by their haploid female progenitors passing-on nutrients and chemical compounds improving survival under stressful conditions.

Haploid females in the isomorphic biphasic life-cycle of Gracilaria chilensis excel in survival.

BACKGROUND: Conditional differentiation is one of the most fundamental drivers of biodiversity. Competitive entities (usually species) differ in environmental or ecological niche enabling them to co-exist. Conditional differentiation of haploid and diploid generations is considered to be a requirement for the evolutionary stability of isomorphic biphasic life-cycles and the cause for the natural occurrence of both phases at uneven abundances. Theoretically, stage dependent survival rates are the most efficient way to explain conditional differentiation. RESULTS: We tested for conditional differentiation in survival rates among life stages (haploid males, haploid females, and diploids) of Gracilaria chilensis, an intertidal red alga occurring along the Chilean shores. Therefore, the fate of individuals was followed periodically for 3 years in five intertidal pools and, for the first time in isomorphic red algae, a composite model of the instantaneous survival rates was applied. The results showed the survival dependency on density (both competition and Allee effects), fertility, age, size, season and location, as well as the differentiation among stages for the survival dependencies of these factors. The young haploid females survived more than the young of the other stages under Allee effects during the environmentally stressful season at the more exposed locations, and under self-thinning during the active growth season. Furthermore, fertile haploid females had a higher survival than fertile haploid males or fertile diploids. CONCLUSIONS: Here, we show a survival advantage of haploids over diploids. The haploid females probably optimize their resource management targeting structural and physiological adaptations that significantly enhance survival under harsher conditions. In a companion paper we demonstrate a fertility advantage of diploids over haploids. Together, the survival and fertility differentiation support the evolution and prevalence of biphasic life-cycles.

Nutrients, but not genetic diversity, affect Gracilaria chilensis (Rhodophyta) farming productivity and physiological responses.

In terrestrial plants, it is well known that genetic diversity can affect responses to abiotic and biotic stress and have important consequences on farming. However, very little is known about the interactive effects of genetic and environmental factors on seaweed crops. We conducted a field experiment on Gracilaria chilensis to determine the effect of heterozygosity and nutrient addition on two southern Chilean farms: Ancud and Chaica. In addition to growth rate and productivity, we measured photosynthetic responses, photosynthetic pigment concentration (chlorophyll a and phycobiliproteins), C:N ratio (C:N), and epiphytic load. Nutrient addition affected the growth rate, productivity, phycobilin, and C:N content, but not the epiphytic load. These results were independent of the heterozygosity of the strains used in the experiments. Interestingly, depending on the sampled sites, distinct photosynthetic responses (i.e., maximal quantum yield, Fv /Fm , and maximal electron transport rate, ETRmax ) to nutrient addition were observed. We propose that thallus selection over the past few decades may have led to ecological differentiation between G. chilensis from Chaica and Ancud. The lack of effect of heterozygosity on growth and physiological responses could be related to the species domestication history in which there is a limited range of genetic variation in farms. We suggest that the existing levels of heterozygosity among our thalli is not sufficient to detect any significant effect of genetic diversity on growth or productivity in Metri bay, our experimental site located close to the city of Puerto Montt, during summer under nitrogen limiting conditions.