Evolutionary genetics of the hydroid allodeterminant alr2.

We surveyed genetic variation in alr2, an allodeterminant of the colonial hydroid Hydractinia symbiolongicarpus. We generated cDNA from a sample of 239 Hydractinia colonies collected at Lighthouse Point, Connecticut, and identified 473 alr2 alleles, 198 of which were unique. Rarefaction analysis suggested that the sample was near saturation. Most alleles were rare, with 86% occurring at frequencies of 1% or less. Alleles were highly variable, diverging on average by 18% of the amino acids in a predicted extracellular domain of the molecule. Analysis of 152 full-length alleles confirmed the existence of two structural types, defined by exons 4-8 of the gene. Several residues of the predicted immunoglobulin superfamily-like domains display signatures of positive selection. We also identified 77 unique alr2 pseudogene sequences from 85 colonies. Twenty-seven of these sequences matched expressed alr2 sequences from other colonies. This observation is consistent with pseudogenes contributing to alr2 diversification through sequence donation. A more limited collection of animals was made from a distant, relict population of H. symbiolongicarpus. Sixty percent of the unique sequences identified in this sample were found to match sequences from the Lighthouse Point population. The large number of alr2 alleles, their degree of divergence, the predominance of rare alleles in the population, their persistence over broad spatial and temporal scales, and the signatures of positive selection in multiple residues of the putative recognition domain paint a consistent picture of negative-frequency-dependent selection operating in this system. The genetic diversity observed at alr2 is comparable to that of the most highly polymorphic genetic systems known to date.

Comparison of gamete compatibility between two blue mussel species in sympatry and in allopatry.

Recent demonstrations of positive selection on genes controlling gamete compatibility have resulted in a proliferation of hypotheses concerning the sources of selection. We tested a prediction of one prominent hypothesis, selection to avoid hybridization (i.e., reinforcement), by comparing heterospecific gamete compatibility in two Mytilus edulis populations: one population in Cobscook Bay, Maine, in which the close congener, M. trossulus, is abundant (a region of sympatry), and one population in Kittery, Maine, in which M. trossulus is absent (a region of allopatry). Three diagnostic nuclear DNA markers were used to identify mussels to species and to estimate the frequency of both species and their hybrids in the two populations. Controlled crosses were then conducted by combining eggs of M. edulis females with a range of M.edulis and M. trossulus sperm concentrations. Results were not consistent with the reinforcement hypothesis. M. edulis females collected from the region of sympatry were no more incompatible with M. trossulus males than were M. edulis females collected from the region of allopatry. A trend in the opposite direction, toward greater compatibility in sympatry, suggests that introgression of M. trossulus genes that control egg compatibility, such as those encoding receptors for sperm, may influence evolution of gametic isolation in hybridizing populations.

Evidence of a Native Northwest Atlantic COI Haplotype Clade in the Cryptogenic Colonial Ascidian Botryllus schlosseri.

The colonial ascidian Botryllus schlosseri should be considered cryptogenic (i.e., not definitively classified as either native or introduced) in the Northwest Atlantic. Although all the evidence is quite circumstantial, over the last 15 years most research groups have accepted the scenario of human-mediated dispersal and classified B. schlosseri as introduced; others have continued to consider it native or cryptogenic. We address the invasion status of this species by adding 174 sequences to the growing worldwide database for the mitochondrial gene cytochrome c oxidase subunit I (COI) and analyzing 1077 sequences to compare genetic diversity of one clade of haplotypes in the Northwest Atlantic with two hypothesized source regions (the Northeast Atlantic and Mediterranean). Our results lead us to reject the prevailing view of the directionality of transport across the Atlantic. We argue that the genetic diversity patterns at COI are far more consistent with the existence of at least one haplotype clade in the Northwest Atlantic (and possibly a second) that substantially pre-dates human colonization from Europe, with this native North American clade subsequently introduced to three sites in Northeast Atlantic and Mediterranean waters. However, we agree with past researchers that some sites in the Northwest Atlantic have more recently been invaded by alien haplotypes, so that some populations are currently composed of a mixture of native and invader haplotypes.

Across-shelf distribution of blue mussel larvae in the northern Gulf of Maine: consequences for population connectivity and a species range boundary.

Studies of population connectivity have largely focused on along-shelf, as opposed to across-shelf, processes. We hypothesized that a discontinuity in across-shelf mixing caused by the divergence of the Eastern Maine Coastal Current (EMCC) from shore acts as an ecological barrier to the supply of mussel larvae to the coast. Existing data on the relative abundance of two congeneric blue mussels, Mytilus edulis and M. trossulus, were analysed to quantify the association of M. trossulus with the colder temperature signal of the EMCC and generate larval distribution predictions. We then sampled the across-shelf distribution of larvae along two transects during 2011. Larvae were identified using restriction digests of PCR amplicons from the mitochondrial 16S rDNA. Mytilus edulis larvae were consistently abundant on either the inshore and offshore transect ends, but not homogeneously distributed across the shelf, while M. trossulus larvae were less common throughout the study area. The divergence of the EMCC from shore appears to create a break in the connectivity of M. edulis populations by isolating those inshore of the EMCC from upstream larval sources. Across-shelf transport processes can thus produce connectivity patterns that would not be predicted solely on the basis of along-shelf processes.

The effects of precipitation, river discharge, land use and coastal circulation on water quality in coastal Maine.

Faecal pollution in stormwater, wastewater and direct run-off can carry zoonotic pathogens to streams, rivers and the ocean, reduce water quality, and affect both recreational and commercial fishing areas of the coastal ocean. Typically, the closure of beaches and commercial fishing areas is governed by the testing for the presence of faecal bacteria, which requires an 18-24 h period for sample incubation. As water quality can change during this testing period, the need for accurate and timely predictions of coastal water quality has become acute. In this study, we: (i) examine the relationship between water quality, precipitation and river discharge at several locations within the Gulf of Maine, and (ii) use multiple linear regression models based on readily obtainable hydrometeorological measurements to predict water quality events at five coastal locations. Analysis of a 12 year dataset revealed that high river discharge and/or precipitation events can lead to reduced water quality; however, the use of only these two parameters to predict water quality can result in a number of errors. Analysis of a higher frequency, 2 year study using multiple linear regression models revealed that precipitation, salinity, river discharge, winds, seasonality and coastal circulation correlate with variations in water quality. Although there has been extensive development of regression models for freshwater, this is one of the first attempts to create a mechanistic model to predict water quality in coastal marine waters. Model performance is similar to that of efforts in other regions, which have incorporated models into water resource managers' decisions, indicating that the use of a mechanistic model in coastal Maine is feasible.

Remarkable longevity of dilute sperm in a free-spawning colonial ascidian.

Many benthic marine invertebrates reproduce by releasing sperm into the sea (free-spawning), but the amount of time that sperm are viable after spawning may have different consequences for fertilization, depending on the type of free-spawner. In egg-broadcasting marine organisms, gamete age is usually assumed to be irrelevant because of the low probability of contact between dilute sperm and egg. However, direct dilution effects might be reduced in egg-brooding free-spawners that filter dilute sperm out of the water column, and sperm longevity may play a role in facilitating fertilization in these taxa. We investigated the effects of time, temperature, and mixing on the viability of naturally released sperm of the colonial ascidian Botryllus schlosseri. Our data indicate that B. schlosseri sperm have a functional life span that is considerably longer than those of the sperm of many other marine invertebrate taxa (half-life of approximately 16 to 26 h), are able to fertilize eggs at extremely low external sperm concentrations (ca. 10(1) sperm ml(-1)), and have a longevity that varies with temperature. It is possible that such prolonged sperm longevity may be achieved by reductions in motility, reactivation of quiescent sperm by chemical cues, or intermittent swimming.

Fertilization in an egg-brooding colonial ascidian does not vary with population density.

The possibility that free-spawning marine organisms may be subject to fertilization failure at low population density (due to the effects of sperm dilution) has sparked much interest, but these effects have been demonstrated only in a few species that broadcast their eggs. Some egg-brooding species may overcome dilution effects by filtering low concentrations of sperm from seawater and fertilizing eggs throughout an extended period of time. We examined the effects of population density and size on fertilization in Botryllus schlosseri, a hermaphroditic colonial ascidian that free-spawns sperm, but broods eggs. We experimentally manipulated the size and density of mating groups and surveyed fertilization levels in natural populations that varied in density. Fertilization was not affected by variation in population size or density in either the experimental or natural populations. Near the end of the reproductive season, some eggs may have been fertilized too late to complete development, suggesting a temporal form of sperm limitation that has not been considered in other systems. We also detected greater variability in fertilization levels at lower population density. Nevertheless, these results suggest that caution must be used in extrapolating reported density effects on fertilization to all taxa of free-spawners; density effects may be reduced in brooders that have efficient sperm collection mechanisms.

Relative effects of gamete compatibility and hydrodynamics on fertilization in the green sea urchin Strongylocentrotus droebachiensis.

Intraspecific variation in gamete compatibility among male/female pairs causes variation in the concentration of sperm required to achieve equivalent fertilization levels. Gamete compatibility is therefore potentially an important factor controlling mating success. Many broadcast-spawning marine invertebrates, however, also live in a dynamic environment where hydrodynamic conditions can affect the concentration of sperm reaching eggs during spawning. Thus flow conditions may moderate the effects of gamete compatibility on fertilization. Using the green sea urchin Strongylocentrotus droebachiensis as a model system, we assessed the relative effects of gamete compatibility (the concentration of sperm required to fertilize 50% of the eggs in specific male/female pairs; F50) and the root-mean-square of total velocity (urms; 0.01-0.11 m s(-1)) on fertilization in four locations near a spawning female (water column, wake eddy, substratum, and aboral surface) in both unidirectional and oscillatory flows. Percent fertilization decreased significantly with increasing urms at all locations and both flow regimes. However, although gamete compatibility varied by almost 1.5 orders of magnitude, it was not a significant predictor of fertilization for most combinations of position and flow. The notable exception was a significant effect of gamete compatibility on fertilization on the aboral surface under unidirectional flow. Our results suggest that selection on variation in gamete compatibility may be strongest in eggs fertilized on the aboral surface of sea urchins and that hydrodynamic conditions may add environmental noise to selection outcomes.

Increase in antimicrobial resistance in bacteria isolated from stranded marine mammals of the Northwest Atlantic.

Studies on marine mammals can inform our understanding of the environmental health of the ocean. To evaluate the potential for changes in antimicrobial resistance, we analyzed a database spanning 2004-2010 that consisted of bacterial isolate identity and antimicrobial sensitivity for stranded pinnipeds in the Northwest Atlantic. Samples (n = 170) from treated animals yielded 310 bacterial isolates representing 24 taxa. We evaluated changes in antimicrobial class resistance from 2004 to 2010 for eight taxa. Escherichia coli displayed a significant increase in resistance to several antimicrobial classes. Other taxa displayed significant increases in resistance to aminoglycosides, and/or fluoroquinolones. In addition, we observed a significant increase in multiple antimicrobial resistance in cultures from untreated animals. These results demonstrate an increase in resistance among common bacterial pathogens of marine mammals over a time span of 6 years.

Effects of oscillatory flow on fertilization in the green sea urchin Strongylocentrotus droebachiensis.

Broadcast spawning invertebrates that live in shallow, high-energy coastal habitats are subjected to oscillatory water motion that creates unsteady flow fields above the surface of animals. The frequency of the oscillatory fluctuations is driven by the wave period, which will influence the stability of local flow structures and may affect fertilization processes. Using an oscillatory water tunnel, we quantified the percentage of eggs fertilized on or near spawning green sea urchins, Strongylocentrotus droebachiensis. Eggs were sampled in the water column, wake eddy, substratum and aboral surface under a range of different periods (T = 4.5-12.7 s) and velocities of oscillatory flow. The root-mean-square wave velocity (rms(u(w))) was a good predictor of fertilization in oscillatory flow, although the root-mean-square of total velocity (rms(u)), which incorporates all the components of flow (current, wave and turbulence), also provided significant predictions. The percentage of eggs fertilized varied between 50-85% at low flows (rms(u(w)) <0.02 m s(-1)), depending on the location sampled, but declined to below 10% for most locations at higher rms(u(w)). The water column was an important location for fertilization with a relative contribution greater than that of the aboral surface, especially at medium and high rms(u(w)) categories. We conclude that gametes can be successfully fertilized on or near the parent under a range of oscillatory flow conditions.

Across-shelf transport of bivalve larvae: can the interface between a coastal current and inshore waters act as an ecological barrier to larval dispersal?

Using an integrated physical and biological approach, we examined across-shelf advection and exchange and the associated transport of bivalve larvae in the presence of a strong coastal current separated from the coast by a stratified inshore environment. We tested the hypothesis that the interface of the coastal current and inshore waters can act as an ecological barrier to across-shelf transport of larvae but can be overcome by wind- or tidally-induced transport. Our study region in the Gulf of Maine encompasses a coastal current that diverges from the coast as it moves downshelf. The region inshore of this current is home to several species that exhibit limited recruitment in spite of extensive upshelf larval sources. Analysis of surface water temperatures and wind velocities revealed episodic decreases in temperature along the coast correlated with alongshelf (but not upwelling) winds, indicating wind-forced onshore movement of the cold coastal current. Such wind-driven onshore migrations are more common along the northern portion of the study region where the coastal current is near the coast, tidal currents are strong, and wind directions are more conducive to onshore migration, but rarer further south where the interface between inshore waters and the coastal current is further offshore and suitable wind events are less common. The distribution of bivalve larvae was consistent with the physical measurements. There was little across-shelf variation in larval abundance where the current abuts the coast, indicating strong across-shelf exchange of larvae, but strong across-shelf variation in larval density where the stratified inshore waters separate the current from the coast, indicating weak across-shelf transport of larvae. Our results suggest that the interface between the coastal current and inshore waters may constitute a major ecological barrier to larval dispersal in the southern part of the region that may only be overcome by rare, strong wind-forced events.

Use of herring bait to farm lobsters in the Gulf of Maine.

BACKGROUND: Ecologists, fisheries scientists, and coastal managers have all called for an ecosystem approach to fisheries management, yet many species such as the American lobster (Homarus americanus) are still largely managed individually. One hypothesis that has yet to be tested suggests that human augmentation of lobster diets via the use of Atlantic herring (Clupea harengus) as bait may contribute to recent increases in lobster landings. Currently 70% of Atlantic herring landings in the Gulf of Maine are used as bait to catch lobsters in traps throughout coastal New England. METHODOLOGY/PRINCIPAL FINDINGS: We examined the effects of this herring bait on the diet composition and growth rate of lobsters at heavily baited vs. seasonally closed (i.e., bait free) sites in coastal Maine. Our results suggest that human use of herring bait may be subsidizing juvenile lobster diets, thereby enhancing lobster growth and the overall economic value and yield of one of the most valuable fisheries in the U.S. CONCLUSIONS/SIGNIFICANCE: Our study illustrates that shifting to an ecosystem approach to fisheries management should require consideration of cross-fishery interactions.

Effects of fertilization distance on male gain curves in a free-spawning marine invertebrate: a combined empirical and theoretical approach.

Male gain curves describe the relationship between allocation to sperm production and male reproductive success and are central to models of sex allocation in hermaphrodites. Sperm competition is expected to result in more linear gains and select for increased allocation. We hypothesized that high sperm production in passively mating systems may also be the result of selection to enhance the ability to fertilize distant ova. Consequently, we explored the effect of distance on male gain curves in a free-spawning colonial ascidian. The performance of focal males that varied in sperm production was assayed at three distances via microsatellite markers. An advection-diffusion model was used to estimate sperm concentration gradients, to predict male reproductive gain integrated across multiple downstream females, and explore effects of hydrodynamic conditions. As distance increased, male reproductive success decreased and empirical gain curves became increasingly linear. Our model predicted that the expected net gain curve is relatively insensitive to variation in flow regime and will saturate much more slowly than if only a single, nearby distance is considered. Thus, high levels of sperm production may enhance fitness both in competitive situations and with increasing fertilization distance, highlighting the need to consider distance effects when evaluating gain curves.

Variation in multiple paternity in natural populations of a free-spawning marine invertebrate.

For free-spawning marine invertebrates, fertilization processes control the genetic diversity of offspring. Each egg can potentially be fertilized by a sperm from a different male, and hence genetic diversity within a brood varies with levels of multiple paternity. Yet, few studies have characterized the frequency of multiple paternity in natural spawns. We analysed patterns of multiple paternity in two populations of the colonial ascidian Botryllus schlosseri using microsatellites. Because previous studies have shown that at moderate to high population densities, competition among male-phase B. schlosseri colonies results in the nearest male dominating the paternity of a brood, we specifically tested the effect of population density on patterns of paternity. Paternity was estimated using three multilocus indices: minimum number of fathers, counts of sperm haplotypes, and effective paternity (K(E)). Multiple paternity was evident in more than 92% of the broods analysed, but highly variable, with a few broods displaying unequal contributions of different males. We found no effect of population density on multiple paternity, suggesting that other factors may control paternity levels. Indirect benefits from increasing the genetic diversity of broods are a possible explanation for the high level of multiple paternity in this species.

Comment on "Divergent induced responses to an invasive predator in marine mussel populations".

Freeman and Byers (Reports, 11 August 2006, p. 831) presented evidence for the rapid evolution of antipredator defenses in the mussel Mytilus edulis. However, their analysis is confounded by three issues. Samples from some sites are likely to have included a second species, M. trossulus; their manipulation of chemical cues does not preclude other interpretations; and they failed to establish an adaptive significance to shell thickening.

NATURAL SELECTION ON HYDROID COLONY MORPHOLOGY BY INTRASPECIFIC COMPETITION.

Previous work on colonial hydroids in the genus Hydractinia has demonstrated that colony morphology is highly variable and determines intraspecific competitive ability. Competitive encounters are known to be common in nature, suggesting that intraspecific competition may be a major selective force acting on morphological variation. A replicated common garden experiment demonstrated a genetic basis to morphological variation and two data sets provided correlative support for the hypothesis of selection by intraspecific competition. First, morphologies inferior in competitive ability were less abundant in two adult, postcompetition, samples than in juvenile, precompetition, samples from the same populations. Second, among eight populations, the relative frequency of different morphologies was correlated with the frequency of intraspecific competition observed in each population. The direction of selection by competition on the morphological variation present in this species conflicts with recent predictions based on surveys across diverse taxa, suggesting limitations to the inference of competition as a past selective agent on the basis of present day correlations among species.

Phenotypic plasticity of reproductive effort in a colonial ascidian, Botryllus schlosseri.

Phenotypic plasticity is the capability of a genotype to produce different phenotypes in different environments. Previous studies have indicated phenotypic variability in asexual, male, and female reproduction in Botryllus schlosseri, a hermaphroditic, colonial ascidian, but not explicitly tested for genotype by environment interactions that indicate genetic variation in plastic responses. Consequently, clones derived from an estuarine population were deployed at their native site and a warmer, higher productivity site 10 km up-river. Male reproduction was assayed by testis size, female reproduction by the number of eggs produced, and asexual reproduction by colony growth rate. To test for ontogenetic effects, data were collected from two different generations of zooids born in the field. Analyses of variance indicated plasticity in asexual and female reproduction during the first zooid generation and plasticity in all three traits during the third zooid generation. Reaction norms varied significantly among genotypes in direction and magnitude for asexual reproduction at both times, implying that selection on asexual reproduction is weak. Sperm production during the third zooid generation was significantly lower at the nonnative site, but there was no genotype by environment interaction. The reaction norms for female reproduction varied significantly among genotypes in direction and magnitude during the first zooid generation, but only varied in magnitude during the third generation, with egg production being higher in all genotypes at the nonnative site. Comparisons of weighted frequency distributions between sites demonstrated that differences in egg production in the third generation were due to increases in the proportion of reproductive zooids within a colony. The greater emphasis on female reproduction at a site associated with higher food availability and temperature, and the greater emphasis on male reproduction at a colder, food-limited site, supports predictions from sex allocation theory.