Adaptive maintenance of genetic polymorphism in an intertidal barnacle: habitat- and life-stage-specific survivorship of Mpi genotypes

In the northern acorn barnacle, Semibalanus balanoides, genotype frequencies of three genetic markers were tracked over time in four types of intertidal habitats. These habitats were selected to represent natural variation in several environmental parameters, specifically the degree of physical stress experienced by barnacles. Frequencies for one allozyme locus (Gpi) and a presumably neutral mtDNA marker were homogeneous among habitats in each temporal sample. Similarly, no temporal stratification in genotype frequencies was evident across the five sampling intervals: from planktonic larvae sampled in March to juveniles collected at the end of June. In contrast to the Gpi and mtDNA loci, Mpi genotypes significantly changed in frequency in two habitats in the high intertidal zone. On exposed substrate, the Mpi-FF homozygote increased in frequency, whereas the alternative homozygote, Mpi-SS, significantly decreased in frequency. Barnacles that were protected from environmental stress at high intertidal heights by the Ascophyllum nodosum algal canopy demonstrated the opposite pattern. In both habitats, the change in frequency of the heterozygote was intermediate to that of the homozygous genotypes. Furthermore, these patterns of genotype-by-environment association reflected a pulse of genotype-specific mortality that occurred over a two-week interval subsequent to metamorphosis from the larval to the adult form. These data indicate that each Mpi homozygote is the highest fitness genotype in some portion of the intertidal environment. Using the Levene (1953) model to evaluate the spatial variation in genotypic fitness, the stable maintenance of the Mpi polymorphism is predicted under certain subsets of conditions. Environmental heterogeneity in the intertidal zone translates to spatial variation in selection pressures, which may result in the active maintenance of the Mpi polymorphism in this species.     

The effects of diet and physiological stress on the evolutionary dynamics of an enzyme polymorphism

In the northern acorn barnacle Semibalanus balanoides, polymorphism at the mannose-6-phosphate isomerase (Mpi) locus appears to be maintained by distinct selection regimes that vary between intertidal microhabitats. The goal of the present experiment was to elucidate the mechanism of selection at the Mpi locus by examining the relationship between genotype and fitness-related life-history traits in laboratory manipulations. When barnacles were cultured on a mannose-supplemented diet and exposed to thermal stress, different Mpi genotypes exhibited differences in the rate of growth that predicted survivorship. In contrast, no such relationship was observed in control or fructose-supplemented dietary treatments either in the presence or in the absence of stress. Similarly, the phenotype and survivorship of genotypes at another allozyme locus and a presumably neutral mitochondrial DNA marker were homogeneous across all treatments and unaffected by experimental manipulations. These results suggest that the differential survivorship of Mpi genotypes in the field and laboratory results from a differential ability to process mannose-6-phosphate through glycolysis. The widespread polymorphism at Mpi observed in marine taxa may reflect the interaction between dietary composition and environmental heterogeneity in intertidal habitats.     

Environmental heterogeneity and balancing selection in the acorn barnacle Semibalanus balanoides

The northern acorn barnacle Semibalans banlanoides occupies several intertidal microhabitats which vary greatly in their degree of physical stress. This environmental heterogeneity creates distinct selection regimes which can maintain genetic variation in natural populations. Despite considerable attention placed on the link between spatial variation in fitness and balancing selection at specific loci, experimental manipulations and fitness estimates for molecular polymorphisms have rarely been conducted in the wild. The aim of this transplant experiment was to manipulate the level of physical stress experienced by a cohort of barnacles in the field and then investigate the spatial variation in fitness for genotypes at three loci: two candidate allozymes and the mitochondrial DNA control region. The viability of mannose-6-phosphate isomerase (Mpi) genotypes was dependent on the level of physical stress experienced in the various treatments; alternative homozygotes were favoured in alternative high stress-low stress environments. In contrast, the fitness of genotypes at other loci was equivalent among treatments and unaffected by the manipulation. Evaluated in the light of balancing selection models, these data indicate that the presence of multiple environmental niches is sufficient to promote a stable Mpi polymorphism in barnacle populations and that allelic variation at this locus reflects the process of adaptation to the heterogeneous intertidal landscape.     

GENE FLOW VERSUS LOCAL ADAPTATION IN THE NORTHERN ACORN BARNACLE, SEMIBALANUS BALANOIDES: INSIGHTS FROM MITOCHONDRIAL DNA VARIATION

Abstract In reciprocal transplant experiments, Bertness and Gaines (1993) found that Semibalanus balanoides juveniles that had settled in an upper Narragansett Bay estuary survived better in that estuary that did juveniles from coastal localities. The observed pattern of survivorship led to the claim that local adaptation may result from a combination of limited gene flow between and strong selection within these habitats. Here we test the hypothesis that limited gene flow has led to habitat‐specific population differentiation using sequence and restriction fragment length polymorphism analyses of the mitochondrial DNA D‐loop region of S. balanoides. Samples were analyzed from replicated coastal and estuary localities in both Narragansett Bay, Rhode Island, and Damariscotta River, Maine. The patterns of F_ST indicate that gene flow between coast and estuary is extensive (Nm > 100) and is not lower in the estuary with lower flushing rates (Narragansett Bay). Given the high estimate of genetic exchange, adaptations for unpredictable environments seem more likely than local adaptation in this species because loci that respond to selection in one generation are essentially homogenized by the next seasons' settlement. Nevertheless, these estimates of neutral gene flow can help identify the strength of selection necessary for local adaptation to accumulate in Semibalanus.     

Genetic variation in the acorn barnacle from allozymes to population genomics

Understanding the patterns of genetic variation within and among populations is a central problem in population and evolutionary genetics. We examine this question in the acorn barnacle, Semibalanus balanoides, in which the allozyme loci Mpi and Gpi have been implicated in balancing selection due to varying selective pressures at different spatial scales. We review the patterns of genetic variation at the Mpi locus, compare this to levels of population differentiation at mtDNA and microsatellites, and place these data in the context of genome-wide variation from high-throughput sequencing of population samples spanning the North Atlantic. Despite considerable geographic variation in the patterns of selection at the Mpi allozyme, this locus shows rather low levels of population differentiation at ecological and trans-oceanic scales (F(ST)?～?5%). Pooled population sequencing was performed on samples from Rhode Island (RI), Maine (ME), and Southwold, England (UK). Analysis of more than 650 million reads identified approximately 335,000 high-quality SNPs in 19 million base pairs of the S. balanoides genome. Much variation is shared across the Atlantic, but there are significant examples of strong population differentiation among samples from RI, ME, and UK. An F(ST) outlier screen of more than 22,000 contigs provided a genome-wide context for interpretation of earlier studies on allozymes, mtDNA, and microsatellites. F(ST) values for allozymes, mtDNA and microsatellites are close to the genome-wide average for random SNPs, with the exception of the trans-Atlantic F(ST) for mtDNA. The majority of F(ST) outliers were unique between individual pairs of populations, but some genes show shared patterns of excess differentiation. These data indicate that gene flow is high, that selection is strong on a subset of genes, and that a variety of genes are experiencing diversifying selection at large spatial scales. This survey of polymorphism in S. balanoides provides a number of genomic tools that promise to make this a powerful model for ecological genomics of the rocky intertidal.     

Seasonal species composition of barnacle larvae (Cirripedia: Thoracica) in Rhode Island waters, 1977-1978

Cirripede larvae can occur year-round in temperate and tropicalwaters, often in significant numbers, yet the species compositionof the ‘Balanus sp.’ component is rardy studied.Weekly plankton samples were analyzed qualitatively for larvalcirripede species and stage over a year (1977–1978) attwo Rhode Island stations. Six species of larvae were foundin Lower Narragansett Bay (30° salinity). Semibalanus balanoidesand Balanus balanus have a single winter brood. S. balanoidesis the predominant winter breeder with a minor release of naupliiin early December and major release in March followed by cypridsin mid-April.B. balanus populations release all larvae in Marchwith cyprids in mid-April. Balanus crenatus is mainly a winterbreeder, but has multiple broods; it does not breed in July-Septemberwhen the water temperature is above 18°C. Balanus venustusis the predominant summer breeder, and larvae were observedfrom May through December (water >8°C). Larvae of Chthamalusfragilis and Balanus eburneus occur in low numbers from May-October.At the Pettaquamscutt River site (12 salinity), Balanus improvisuslarvae predominate and early stage nauplii (I-II) occur in samplesyear round (0–27°C). Two peaks of later stage naupliiand cyprids occur in late spring (May) and early winter (Nov.-Jan.).Continued temperatures bdow 5°C or above 20°C appearto inhibit larval development. Comparison of results with existing literature reveals severalsignificant findings. The bimodal rdease of S. balanoides larvaeis unusual and may be in response to the phytoplankton dynamicsof the year; however, the existence of distinct races of S.balanoides may also be a factor. Larvae of B. venustus predominatein the lower bay during the summer, yet this species is unreportedin past studies. B. improvisus nauplii are more cold tolerantthan previously reported. Comparison of findings with reportedbreeding patterns in Florida indicate significant differencesin temperature responses between northern and southern populations. ¹Contribution No. 188 from EPA Environmental Research Laboratory,South Ferry Rd., Narragansett, RI 02882     

The seasonal distribution of the marine diatom genus Chaetoceros Ehr. in Narragansett Bay, Rhode Island (1981-1982)

The seasonal dynamics of the diatom genus Chaetoceros Ehrenbergwere investigated in Narragansett Bay, Rhode Island. Whole watersamples were collected weekly from July 1981 to October 1982at four locations in the lower Bay and were preserved, concentratedby settling and enumerated. Chaetoceros were most abundant inlate winter–early spring; a second peak occurred in thefall. As in many other temperate neritic areas, C. debilis,C. compressus and C. didymus were major components of die Chaetocerospopulation and could be found throughout the year. Species compositionand dynamics were quite similar at the Jerusalem, GSO Pier andWickford Harbor stations, but were markedly different in PettaquamscuttRiver, a reduced salinity tidal estuary.     

The Asian red seaweed <Emphasis Type="Italic">Grateloupia turuturu</Emphasis> (Rhodophyta) invades the Gulf of Maine

We report the invasion of the Gulf of Maine, in the northwest Atlantic Ocean, by the largest red seaweed in the world, the Asian Grateloupia turuturu. First detected in 1994 in Narragansett Bay, Rhode Island, south of Cape Cod, this alga had expanded its range in the following years only over to Long Island and into Long Island Sound. In July 2007 we found Grateloupia in the Cape Cod Canal and as far north (east) as Boston, Massachusetts, establishing its presence in the Gulf of Maine. Grateloupia can be invasive and may be capable of disrupting low intertidal and shallow subtidal seaweeds. The plant's broad physiological tolerances suggest that it will be able to expand possibly as far north as the Bay of Fundy. We predict its continued spread in North America and around the world, noting that its arrival in the major international port of Boston may now launch G. turuturu on to new global shipping corridors.     

INTERTIDAL MICROHABITAT AND SELECTION AT MPI: INTERLOCUS CONTRASTS IN THE NORTHERN ACORN BARNACLE,SEMIBALANUS BALANOIDES

Barnacles were sampled from various microhabitats in the rocky intertidal at multiple sites in two years. At sites in which there were large differences among microhabitats in temperature profiles, Mpi genotype frequencies were consistently and significantly different. Genotype frequencies for another allozyme locus (Gpi) as well as a DNA marker shown to be neutral (the mtDNA control region) were statistically homogeneous among thermal microhabitats at all sites in both years. The data indicate that temperature and/or desiccation mediated selection is operating at Mpi or a linked locus and that Mpi genotypes experience differential mortality in the various habitat types. If the relative fitness of genotypes is dependent on habitat type, the Mpi polymorphism may be actively maintained by a Levene model of balancing selection (Levene 1953). Because barnacle larvae are produced in abundance each year and spend five to eight weeks dispersing in the water column, there is little opportunity for the accumulation of adaptive divergence over the environmental grain size relevant in intertidal habitats. The Mpi polymorphism may be an important component of a suite of traits involved in the adaptation of barnacles to heterogeneous environments. Due to the relatively high concentration of mannose in a variety of algal groups, the metabolism of mannose may substantially affect individual performance and fitness in this and other species that feed on algae and phytoplankton. Because the Mpi locus is one of the most strongly polymorphic in marine organisms, these findings may be relevant for a diversity of other such species.     

Induction of embryonic dormancy in the calanoid copepod Acartia hudsonica: heritability and phenotypic plasticity in two geographically separated populations

The planktonic copepod Acartia hudsonica apparently requires summer diapause or dormancy to persist in Narragansett Bay, RI. In estuaries to the north, however, active A. hudsonica populations are present year-round and may not express dormancy. Using a full-sibling rearing design and analysis of variance (ANOVA), I assessed the importance of phenotypic and genetic sources of variation in two geographically separate populations, one from Maine and one from Rhode Island. Both populations showed phenotypic plasticity in the percentage of dormant eggs produced. Moreover, experiments revealed significant sibship-environment interactions in both populations, indicating that the phenotypic plasticity has a genetic component. Both populations also revealed a significant amount of genetic variation in the percentage of dormant eggs produced. For the Maine population, broad sense heritability was high (0.91, 1.10) in two short-day (12L:12D) treatments and near zero (0.08) in the long-day treatment (15L:9D). For a Rhode Island population, broad sense heritability was higher (0.95) in a low-temperature short-day treatment (13.5 °C, 12L:12D) than in a high-temperature short-day treatment (17.5 °C, 12L:12D; h²=0.25).     

Genetic heterogeneity among intertidal habitats in the flat periwinkle, Littorina obtusata

Comparisons among patterns exhibited by functionally distinct genetic markers have been widely used to infer the impacts of demography and selection in structuring genetic variation in natural populations. However, such multilocus comparisons remain an indirect evaluation of selection at particular candidate loci; ideally, the identification of a candidate gene by comparative genetic methodologies should be complemented by functional analyses and experimental manipulations of genotypes in the laboratory or field. We examined genotype frequency variation among replicated intertidal habitats at two spatial scales in the grazing snail Littorina obtusata. Both of the candidate allozyme markers varied predictably with environment, and these patterns were consistent at both spatial scales. Three of four reference loci were spatially homogeneous, but one microsatellite exhibited significant structure at both geographical and mesoscales. To initiate a direct examination of whether the observed genotype frequency variation at one of the candidate markers, mannose-6-phosphate isomerase (MPI), was impacted by differential survivorship of genotypes, we conducted a series of laboratory-based thermal stress assays using snails from two geographically disparate source populations. When snails were exposed to bouts of thermal/desiccation stress, patterns of mortality were nonrandom with respect to MPI genotype. Furthermore, patterns of mortality in the laboratory manipulation coincided with the observed distribution of genotypes in the field. The data suggest the operation of selection at the Mpi or a linked locus, but functional studies and further experimentation are required to establish the relationship between MPI genotype and fitness across heterogeneous intertidal environments.     

The role of habitat choice in micro‐evolutionary dynamics: An experimental study on the Mediterranean killifish Aphanius fasciatus (Cyprinodontidae)

Habitat choice is defined as a nonrandom distribution of genotypes in different microhabitats. Therefore, it could exert a great impact on the genetic variance of natural populations by promoting genetic divergence, local adaptation, and may even lead to sympatric speciation. Despite this potential role in micro‐ and macro‐evolutionary processes, there is little empirical evidence that the various genotypes within a population may differ in habitat choice‐related behaviors. Here, we tested whether habitat choice may have contributed to genetic divergence within a local population of the Mediterranean killifish Aphanius fasciatus, which emerged between groups inhabiting microhabitats with different oxygen concentrations during previous field studies. In a first experiment, we studied the distribution of individuals in conditions of hypoxia and normoxia to test whether they had a different ability to shy away from a hypoxic environment; in a second experiment, we analyzed the individual behavior of fish separately in the two conditions, to verify whether they showed peculiar behavioral responses linked to a possible differential distribution. We then analyzed the six allozyme loci, whose allelic and genotypic frequencies were significantly divergent in the previous studies. In the first test, we found that the distribution of the two homozygote genotypes of the glucose‐6‐phosphate isomerase‐1 locus (GPI‐1) was significantly different between the hypoxic and the normoxic conditions. During the second test, all individuals were more active in hypoxic conditions, but the two GPI‐1 homozygotes showed a significant difference in time spent performing surface breathing, which was consistent with their distribution observed in the first experiment. These results provide evidence that individual behavioral traits, related to genetic features, may lead to a nonrandom distribution of genotypes in heterogeneous although contiguous microhabitats and, consequently, that habitat choice can play a significant role in driving the micro‐evolutionary dynamics of this species.     

Genetic polymorphism of adenosine deaminase and mannose phosphate isomerase in blood of arctic foxes, Alopex lagopus

Blood samples of 70 foxes, including 10 families, have been investigated by horizontal starch gel electrophoresis for the enzymes adenosine deaminase (ADA) and mannose phosphate isomerase (MPI). The observed variation of the enzymes could be explained as a result of one locus with two codominant alleles and one with three respectively. The segregation in the families of the alleles assumed for the two loci is in accordance with this genetic model. The frequency of the two alleles at the Ada locus is about the same and the slowest anodic migrating allele at the Mpi locus is the most frequent.     

Effects of Selection for Dark Respiration Rate on Enzyme Genotypes in Lolium perenne

Two lines of perennial ryegrass (Lolium perenne) that had previouslybeen selected for ‘fast’ and ‘slow’rates of leaf dark respiration were examined to determine whetherselection had differentiated allelic or genotypic frequenciesof five polymorphic enzyme loci, and determine whether respirationrates differed among genotypes. The lines were differentiatedfor genes coding for phosphoglucose isomerase (PGI) and UDP-glucosepyrophosphorylase (UDP). Respiration rates were heterogeneousamong UDP and 6-phosphogluconate dehydrogenase (6PGD) genotypes.Founder effects may have produced the genetic differentiationof lines at the PGI locus as we detected no differences amonggenotypes for respiration rates. The genetic differentiationamong lines for UDP was consistent with differences in darkrespiration among genotypes. Although we detected large differences(75%) in dark respiration rate among 6PGD genotypes in the fastrespiring line, selection did not differentiate the lines atthis locus. Lolium perenne L., perennial ryegrass, enzyme polymorphisms, dark respiration rate     

Evolutionary consequences of introgression at M7 lysin, a gamete recognition locus, following secondary contact between blue mussel species

Hybridizing populations of blue mussels, Mytilus edulis and Mytilus trossulus, in Cobscook Bay (eastern Maine) have been used by our laboratory to study the evolution of gamete incompatibility and molecular evolution of the vitelline coat lysin proteins expressed in sperm. The M7 lysin locus has been the most studied of the three lysins, but evidence for positive selection necessary to help confirm its role in gamete recognition in western Atlantic hybrid zones is contradictory. We developed an alternative test, based on rates of introgression at M7 lysin. Contrary to expectations, introgression at this locus is much higher (instead of much lower) than is introgression at neutral markers. In this article, we present simulations, constructed using synthetic populations of combinations of admixed genotypes, representing various hybrid offspring categories. Simulations produced variation in introgression across loci, but did not generate the massive introgression at M7 lysin observed in natural populations in Cobscook Bay. We consider these results in the context of selection operating on gamete recognition loci, both within and between species, during the third stage of allopatric speciation in Mytilus.     

An ‘ecological trap’ for yellow warbler nest microhabitat selection

Contrary to assumptions of habitat selection theory, field studies frequently detect ‘ecological traps’, where animals prefer habitats conferring lower fitness than available alternatives. Evidence for traps includes cases where birds prefer breeding habitats associated with relatively high nest predation rates despite the importance of nest survival to avian fitness. Because birds select breeding habitat at multiple spatial scales, the processes underlying traps for birds are likely scale‐dependent. We studied a potential ecological trap for a population of yellow warblers Dendroica petechia while paying specific attention to spatial scale. We quantified nest microhabitat preference by comparing nest‐ versus random‐site microhabitat structure and related preferred microhabitat features with nest survival. Over a nine‐year study period and three study sites, we found a consistently negative relationship between preferred microhabitat patches and nest survival rates. Data from experimental nests described a similar relationship, corroborating the apparent positive relationship between preferred microhabitat and nest predation. As do other songbirds, yellow warblers select breeding habitat in at least two steps at two spatial scales; (1) they select territories at a coarser spatial scale and (2) nest microhabitats at a finer scale from within individual territories. By comparing nest versus random sites within territories, we showed that maladaptive nest microhabitat preferences arose during within‐territory nest site selection (step 2). Furthermore, nest predation rates varied at a fine enough scale to provide individual yellow warblers with lower‐predation alternatives to preferred microhabitats. Given these results, tradeoffs between nest survival and other fitness components are unlikely since fitness components other than nest survival are probably more relevant to territory‐scale habitat selection. Instead, exchanges of individuals among populations facing different predation regimes, the recent proliferation of the parasitic brown‐headed cowbird Molothrus ater, and/or anthropogenic changes to riparian vegetation structure are more likely explanations.     

The determination of fresh organic carbon weight from formaldehyde preserved macrofaunal samples

Methodologies are presented whereby the fresh organic carbon weight of formaldehyde preserved macrofaunal samples may be estimated. Length-organic carbon weight regressions were determined for the four numerically dominant bivalves in Narragansett Bay, Rhode Island (Nucula annulata, Yoldia limatula, Mulinia lateralis, and Pandora gouldiana) and one commercially important, but less abundant species (Mercenaria mercenaria). Constants were determined to convert the dry weight of preserved softbodied organisms (polychaetes, oligochaetes, amphipods, etc.) to fresh (unpreserved) organic carbon weight. These results can be used by investigators studying the energetics of benthic communities similar to those in Narragansett Bay.     

Effect of light and substratum structural complexity on microhabitat selection by the snail Helix aspersa muller

Terrestrial snails choose their microhabitat according to anumber of environmental factors. We evaluated the effect oflight intensity and substratum complexity on microhabitat preferenceof the terrestrial snail Helix aspersa using a multi-factorialdesign. The snails were offered two levels of light intensityand two types of structural complexity, hence 16 treatmentsin total were used: 12 in which choice was offered and 4 inwhich no choice was offered. The snails preferred ambient lightover dim light, regardless of substratum complexity, and complexover smooth substrata, regardless of light intensity. The levelof one factor did not affect the response to the other. Thus,the results revealed a preference for microhabitats with greaterlight intensity and that were structurally complex, and a rejectionof dimly lit microhabitats with smooth substrates. (Received 17 March 2006; accepted 21 September 2006)     

Changes in gene frequencies at the octanol dehydrogenase locus of Drosophila melanogaster imposed by environmental ethanol

Experiments have been performed to study the effect of selection at the Odh locus in Drosophila melanogaster populations using different alcohol concentrations in the medium. The data can be best interpreted by assuming frequency-dependent selection. When genotype frequencies are considered as independent variables and values of Wrightian fitness as dependent variables, it turns out that different functions describe the selection of the coexisting genotypes. A linear equation is used for the SF genotype and a hyperbolic function for the FF genotype. No function of good fit could be found for the SS genotype. Simulation experiments using these functions fit our data well.