Transgressive gene expression and expression plasticity under thermal stress in a stable hybrid zone.

Interspecific hybridization can lead to myriad outcomes, including transgressive phenotypes in which the hybrids are more fit than either parent species. Such hybrids may display important traits in the context of climate change, able to respond to novel environmental conditions not previously experienced by the parent populations. While this has been evaluated in an agricultural context, the role of transgressive hybrids under changing conditions in the wild remains largely unexplored; this is especially true regarding transgressive gene expression. Using the blue mussel species complex (genus Mytilus) as a model system, we investigated the effects of hybridization on temperature induced gene expression plasticity by comparing expression profiles in parental species and their hybrids following a 2-week thermal challenge. Hybrid expression plasticity was most often like one parent or the other (50%). However, a large fraction of genes (26%) showed transgressive expression plasticity (i.e. the change in gene expression was either greater or lesser than that of both parent species), while only 2% were intermediately plastic in hybrids. Despite their close phylogenetic relationship, there was limited overlap in the differentially expressed genes responding to temperature, indicating interspecific differences in the responses to high temperature in which responses from hybrids are distinct from both parent species. We also identified differentially expressed long non-coding RNAs (lncRNAs), which we suggest may contribute to species-specific differences in thermal tolerance. Our findings provide important insight into the impact of hybridization on gene expression under warming. We propose transgressive hybrids may play an important role in population persistence under future warming conditions.

Integration of natural selection across the life cycle stabilizes a marine mussel hybrid zone.

Hybridization among related species is now recognized as common but it remains unclear how hybrid zones persist for prolonged periods. Here, we test the hypothesis that selection in different components of the life cycle may stabilize a hybrid zone. A hybrid zone occurs in southwest England between the marine mussels Mytilus edulis and M. galloprovincialis. Previous studies have found strong directional selection against alleles from M. edulis occurs among hybrids in the adult stage. Traditional hybrid zone models argue that alleles that are selected within the hybrid zone are replaced by migration from neighboring parental population into the hybrid zone. In this system, however, migration occurs out of this hybrid zone into neighboring parental populations. This hybrid zone should therefore be unstable and dissipate, yet this zone has persisted for more than 30 years. We tested and rejected the hypothesis that differences in fecundity may select for M. edulis alleles within this hybrid zone and thus counter the selection observed against these alleles among adults. We also tested the hypothesis that selection during the larval stage may counter selection against M. edulis alleles in the adult stage. We found that selection favors M. edulis alleles during the veliger stage of larval development. The direction and strength of selection during the larval stage are sufficient to counter strong selection during the adult portion of the life cycle. This hybrid zone is stabilized by opposing forms of directional selection operating in different portions of the life cycle.

Long-term, high frequency in situ measurements of intertidal mussel bed temperatures using biomimetic sensors.

At a proximal level, the physiological impacts of global climate change on ectothermic organisms are manifest as changes in body temperatures. Especially for plants and animals exposed to direct solar radiation, body temperatures can be substantially different from air temperatures. We deployed biomimetic sensors that approximate the thermal characteristics of intertidal mussels at 71 sites worldwide, from 1998-present. Loggers recorded temperatures at 10-30 min intervals nearly continuously at multiple intertidal elevations. Comparisons against direct measurements of mussel tissue temperature indicated errors of ~2.0-2.5 °C, during daily fluctuations that often exceeded 15°-20 °C. Geographic patterns in thermal stress based on biomimetic logger measurements were generally far more complex than anticipated based only on 'habitat-level' measurements of air or sea surface temperature. This unique data set provides an opportunity to link physiological measurements with spatially- and temporally-explicit field observations of body temperature.

Loss of thermal refugia near equatorial range limits.

This study examines the importance of thermal refugia along the majority of the geographical range of a key intertidal species (Patella vulgata Linnaeus, 1758) on the Atlantic coast of Europe. We asked whether differences between sun-exposed and shaded microhabitats were responsible for differences in physiological stress and ecological performance and examined the availability of refugia near equatorial range limits. Thermal differences between sun-exposed and shaded microhabitats are consistently associated with differences in physiological performance, and the frequency of occurrence of high temperatures is most probably limiting the maximum population densities supported at any given place. Topographical complexity provides thermal refugia throughout most of the distribution range, although towards the equatorial edges the magnitude of the amelioration provided by shaded microhabitats is largely reduced. Importantly, the limiting effects of temperature, rather than being related to latitude, seem to be tightly associated with microsite variability, which therefore is likely to have profound effects on the way local populations (and consequently species) respond to climatic changes.

Climate change, species distribution models, and physiological performance metrics: predicting when biogeographic models are likely to fail.

Modeling the biogeographic consequences of climate change requires confidence in model predictions under novel conditions. However, models often fail when extended to new locales, and such instances have been used as evidence of a change in physiological tolerance, that is, a fundamental niche shift. We explore an alternative explanation and propose a method for predicting the likelihood of failure based on physiological performance curves and environmental variance in the original and new environments. We define the transient event margin (TEM) as the gap between energetic performance failure, defined as CTmax, and the upper lethal limit, defined as LTmax. If TEM is large relative to environmental fluctuations, models will likely fail in new locales. If TEM is small relative to environmental fluctuations, models are likely to be robust for new locales, even when mechanism is unknown. Using temperature, we predict when biogeographic models are likely to fail and illustrate this with a case study. We suggest that failure is predictable from an understanding of how climate drives nonlethal physiological responses, but for many species such data have not been collected. Successful biogeographic forecasting thus depends on understanding when the mechanisms limiting distribution of a species will differ among geographic regions, or at different times, resulting in realized niche shifts. TEM allows prediction of the likelihood of such model failure.

Physiological energetics and biogeographic range limits of three congeneric mussel species.

Closely related species with different physiological tolerances and distributions make ideal systems for documenting range shifts in response to a changing climate. Mytilus edulis, M. trossulus, and M. galloprovincialis are sibling species of marine mussels with distinct biogeographical ranges that are correlated with sea surface temperatures. We determined the scope for growth of these three species at a range of temperatures to determine if energetics could predict their distributions. Scope for growth (SFG) represents energy available for growth and/or reproduction above that necessary for maintenance requirements. The SFG of M. galloprovincialis, the species known to inhabit the warmest habitats, was shifted towards warmer temperatures compared to the other two species, remaining positive until nearly 30 °C. M. edulis, a cold-temperate species, maintained a positive SFG up to 23 °C. M. trossulus, a boreal species, generally was not able to maintain a positive SFG above 17 °C. The warm end of each species' range correlated strongly with the point at which that species' SFG became negative in summer and fall. Energetics at cold temperatures did not predict the cold end of the species' ranges, as there was no clear SFG advantage to explain the dominance of M. trossulus in cold habitats. As sea surface temperatures continue to warm with climate change, the energetics of these three species provide a basis for developing mechanistic models predicting future distribution and productivity changes in mussel populations.

Ecological genetics in the North Atlantic: environmental gradients and adaptation at specific loci.

The North Atlantic intertidal community provides a rich set of organismal and environmental material for the study of ecological genetics. Clearly defined environmental gradients exist at multiple spatial scales: there are broad latitudinal trends in temperature, meso-scale changes in salinity along estuaries, and smaller scale gradients in desiccation and temperature spanning the intertidal range. The geology and geography of the American and European coasts provide natural replication of these gradients, allowing for population genetic analyses of parallel adaptation to environmental stress and heterogeneity. Statistical methods have been developed that provide genomic neutrality tests of population differentiation and aid in the process of candidate gene identification. In this paper, we review studies of marine organisms that illustrate associations between an environmental gradient and specific genetic markers. Such highly differentiated markers become candidate genes for adaptation to the environmental factors in question, but the functional significance of genetic variants must be comprehensively evaluated. We present a set of predictions about locus-specific selection across latitudinal, estuarine, and intertidal gradients that are likely to exist in the North Atlantic. We further present new data and analyses that support and contradict these simple selection models. Some taxa show pronounced clinal variation at certain loci against a background of mild clinal variation at many loci. These cases illustrate the procedures necessary for distinguishing selection driven by internal genomic vs. external environmental factors. We suggest that the North Atlantic intertidal community provides a model system for identifying genes that matter in ecology due to the clarity of the environmental stresses and an extensive experimental literature on ecological function. While these organisms are typically poor genetic and genomic models, advances in comparative genomics have provided access to molecular tools that can now be applied to taxa with well-defined ecologies. As many of the organisms we discuss have tight physiological limits driven by climatic factors, this synthesis of molecular population genetics with marine ecology could provide a sensitive means of assessing evolutionary responses to climate change.

Patterns of larval dispersal and their effect on the maintenance of a blue mussel hybrid zone in southwestern England.

The blue mussels Mytilus edulis and M. galloprovincialis hybridize in southwestern England. Within this hybrid zone environmentally based directional selection favors individuals with alleles specific to M. galloprovincialis. What forces are countering this directional selection and allowing for the maintenance of a stable hybrid population are unknown. We used both the genetics of recently settled larvae and a fine-scale model of the physical oceanography of the region to determine the patterns of larval dispersal throughout the hybrid zone and the bordering parental populations. Evidence from both the model and the genetics suggests that the hybrid zone lies between two barriers to dispersal. Start Point separates the M. edulis population from the hybrid zone and allows minimal dispersal from the hybrid zone into the M. edulis population, but none in the other direction. Likewise, the M. galloprovincialis populations along the northern coast of Cornwall regularly receive immigrating larvae from the hybrid zone, but larvae from the M. galloprovincialis population do not enter the hybrid zone. However, larvae settling at hybrid zone sites have high frequencies of alleles specific to M. edulis, suggesting that reproductive barriers, selection in the larval stage, or gene flow from an undetermined source is effectively balancing the directional selection observed in the adults.

ASYMMETRIC INTROGRESSION OF MITOCHONDRIAL DNA AMONG EUROPEAN POPULATIONS OF BLUE MUSSELS (MYTILUS SPP.).

-Mytilus edulis and M. galloprovincialis are two blue mussel species that coexist in western Europe. Previously, we reported that M. galloprovincialis populations contain female and male haplotypes that are fixed in M. edulis populations as well as unique haplotypes. This study assesses whether paraphyly for these species is due to introgression or incomplete lineage extinction. The lineage extinction hypothesis predicts that the shared mtDNA haplotypes in M. galloprovincialis will be significantly diverged from those in M. edulis and form distinct sequence clades. In contrast, the introgression hypothesis proposes that M. edulis haplotypes have only recently been introduced into M. galloprovincialis through hybridization with relatively little divergence accumulating between the shared RFLP haplotypes. We examined 80 mtl6S gene sequences for both the maternal and paternal mtDNA lineages from mussels sampled from various European populations and found strong support for the introgression hypothesis. In addition, we found that M. edulis mtDNA haplotypes appear to be introgressing into mussel populations in the Baltic Sea, which have predominantly M. trossulus nuclear genotypes, indicating that introgressive hybridization is prevalent among European mussel populations.

GENETICS OF PHYSIOLOGICAL DIFFERENTIATION WITHIN THE MARINE MUSSEL GENUS MYTILUS.

Two divergent taxa in the marine mussel genus Mytilus are largely isolated geographically and are routinely exposed to distinctly different thermal environments. We tested the hypothesis that the two taxa are physiologically differentiated with respect to temperature and examined the evolved adaptations allowing one of the taxa to exploit habitats where warm-temperate conditions prevail for prolonged periods. We first analyzed the physiological response to high temperature of mussels collected from a hybrid population containing members of both pure taxa, F, hybrids, and a variety of introgressed genotypes. The experimental temperature of 23°C was chosen to be permissive to the taxon that occurs in warm-temperate regions (Mytilus galloprovincialis) and restrictive to the cold-water taxon (Mytilus edulis). The results show that the two taxa are physiologically differentiated. Under the experimental conditions, M. galloprovincialis exhibited a threefold higher feeding rate and a slightly elevated metabolic rate compared with M. edulis. These differences did not result in a significant difference in net energy balance between the two taxa, probably because of an interaction between physiological response and food availability. However, M. galloprovincialis grew significantly faster in the field, indicating that the physiological differences observed in the laboratory also occur in nature. Numerous introgressed genotypes provided the opportunity to test for cosegregation between the physiological differences and four highly differentiated genetic markers. Two of the markers (esterase and octopine dehydrogenase) cosegregate with variation in feeding rate and shell growth and explained most of the physiological differences observed between taxa. A strong concordance existed between these two loci, suggesting that they may be linked and may mark segregation of the same linkage group. The results suggest that the physiological differentiation between these taxa may be controlled by a few genes (perhaps only one) each with large effect.

GENOTYPE-ENVIRONMENT INTERACTION FOR JUVENILE GROWTH IN THE HARD CLAM MERCENARIA MERCENARIA (L.).

Offspring from half-sib and full-sib families of the hard clam, Mercenaria mercenaria were reared in five locations along the Atlantic Coast to test for the presence of genotype-environment interaction for juvenile growth rate. Location effects upon growth rate variation were prevalent; of the genetic effects, the additive genetic by location variance was predominant with the nonadditive genetic by location component contributing to a lesser degree to the interaction variance. The additive and nonadditive variation over all environments was negligible. Genotype-environment interaction was found to be at least partially due to a change in the amount of genetic variation expressed at each location; with significant additive variation detected at Charleston and Georgetown, SC sites and significant nonadditive variation at Millsboro, DE. Genetic covariance/correlation analysis indicated that reversals in relative family performance across locations were prevalent, implying the possibility of habitat specialization among genotypes. In addition, graphical analysis produced no evidence of a ubiquitously superior genotype. These analyses suggest that genotype-environment interaction should act to constrain the evolution of juvenile growth rate in Mercenaria, preserve any heritable variation associated with this trait and may lead to the development of phenotypic plasticity for growth.

QUANTITATIVE GENETICS OF JUVENILE GROWTH AND SHAPE IN THE MUD CRAB EURYPANOPEUS DEPRESSUS 1.

Rates of growth and development were measured for the first six molts following the crab 1 stage in the mud crab Eurypanopeus depressus. The genetic contribution to variation in growth rate, development rate, and shape was determined for each molt interval. Genetic variation in growth rate, measured as increases in both width and length, was evident at most molt intervals. There were also significant genetic effects upon the intermolt interval. Growth rates for each molt interval, calculated on a daily basis to remove the interaction between growth rate and development rate also showed genetic variation. There was no evidence that genetic variation in these parameters changed during early juvenile development; there were substantial levels of genetic variation in growth rate at most ontological stages. Despite high levels of genetic variation for growth rate in dimensions of the carapace, there was no evidence of genetic variation in shape. This analysis does not provide a quantitative estimate of the levels of genetic variance for these traits but does indicate that the magnitude of this source of variance must be very significant.

EXCLUSION OF THE ROLE OF SECONDARY CONTACT IN AN ALLELE FREQUENCY CLINE IN THE MUSSEL MYTILUS EDULIS.

We examined the hypothesis that secondary contact generates an allele-frequency cline at the aminopeptidase-I locus (Lap) in the marine mussel, Mytilus edulis. It has been proposed that variation at the Lap locus is neutral and that the cline results from secondary contact between differentiated oceanic and estuarine populations (Levinton, 1980). We tested this hypothesis by comparing the genotypic distributions in samples from the cline to distributions that incorporate mixing effects. We employed a statistical model that determines the degree of contact using a maximum likelihood estimator and then incorporates the mixing estimates into an expected distribution of genotypes. Wahlund effects resulting from possible admixture are thereby incorporated into the expected distribution. Failure of the model to reconcile the observed with the expected distribution of genotypes indicates that the observed population structure does not result from admixture. The null hypothesis of mixing was unable to explain about 33% of the samples. Combined tests demonstrated the general departure from the mixing model to be highly significant. The distribution of heterozygote discrepancies across the cline was inconsistent with the expectations of a mixing model. Therefore we reject explanations for the structure of the Lap cline that involve secondary contact. Selection directed at the Lap locus appears necessary to explain the genotypic structure of clinal populations.

THE PHYSIOLOGICAL BASIS OF NATURAL SELECTION AT THE LAP LOCUS.

An extensive research program was undertaken to evaluate the contribution of genetic variation at the Lap locus to variation in physiological traits under natural conditions. Rates of carbon and nitrogen metabolism were monitored in a population of the mussel Mytilus edulis near the center of the Lap allele frequency cline on the north shore of Long Island. The goal of this research was to establish whether the previously described genotype-dependent differences in physiological phenotype are meaningful in ecologically relevant circumstances. It was predicted from laboratory studies that, in nature, genotype-dependent differences will exist for rates of nitrogen excretion and that other aspects of the animal's physiology, particularly rates of carbon metabolism, will be unaffected by Lap genotype. Rates of amino acid and ammonia excretion were significantly dependent upon Lap genotype; individuals with the Lap94 allele exhibited greater rates of nitrogen loss. These differences among genotypes were most evident in the fall, between September and December. The genotype-dependent component of rates of nitrogen loss were also largest relative to the total rate of excretion during the fall period. As predicted, other aspects of the nitrogen metabolism (acquisition) and rates of carbon metabolism were independent of Lap genotype. There was a striking congruity among a variety of observations that all indicate that phenotypic differences in nitrogen metabolism are the basis of natural selection at the Lap locus in Long Island Sound. Rates of growth were minimal during the fall months (Hilbish, 1985) and mussels are known to lose weight in a genotype-specific manner during this period (Koehn et al., 1980). Rates of elemental gain and loss were summed to produce carbon and nitrogen budgets; these data show the fall to be a period of extended deficit in carbon and nitrogen balance. Genotype-dependent losses of ammonia and amino acids were greatest during the fall months. Finally, selection against the Lap94 allele occurs predominantly in the fall (Hilbish, 1985). The data indicate that the depletion of nitrogen resources provides the basis for selection against Lap94 genotypes during the fall months.