Cynthia D. Trowbridge

Integrative species delimitation in photosynthetic sea slugs reveals twenty candidate species in three nominal taxa studied for drug discovery, plastid symbiosis or biological control

DNA barcoding can highlight taxa in which conventional taxonomy underestimates species richness, identifying mitochondrial lineages that may correspond to unrecognized species. However, key assumptions of barcoding remain untested for many groups of soft-bodied marine invertebrates with poorly resolved taxonomy. Here, we applied an integrative approach for species delimitation to herbivorous sea slugs in clade Sacoglossa, in which unrecognized diversity may complicate studies of drug discovery, plastid endosymbiosis, and biological control. Using the mitochondrial barcoding COI gene and the nuclear histone 3 gene, we tested the hypothesis that three widely distributed species each comprised a complex of independently evolving lineages. Morphological and reproductive characters were then used to evaluate whether each lineage was distinguishable as a candidate species. The circumtropical Elysia ornata comprised a Caribbean species and four Indo-Pacific candidate species that are potential sources of kahalalides, anti-cancer compounds. The monotypic and highly photosynthetic Plakobranchus ocellatus, used for over 60 years to study chloroplast symbiosis, comprised 10 candidate species. Finally, six candidate species were distinguished in the Elysia tomentosa complex, including potential biological control agents for invasive green algae (Caulerpa spp.). We show that a candidate species approach developed for vertebrates effectively categorizes cryptic diversity in marine invertebrates, and that integrating threshold COI distances with non-molecular character data can delimit species even when common assumptions of DNA barcoding are violated.

Poecilogony and Population Genetic Structure in Elysia pusilla (Heterobranchia: Sacoglossa), and Reproductive Data for Five Sacoglossans that Express Dimorphisms in Larval Development

Credible cases of poecilogony, the production of two distinct larval morphs within a species, are extremely rare in marine invertebrates, yet peculiarly common in a clade of herbivorous sea slugs, the Sacoglossa. Only five animal species have been reported to express dimorphic egg sizes that result in planktotrophic and lecithotrophic larvae: the spionid polychaete Streblospio benedicti and four sacoglossans distributed in temperate estuaries or the Caribbean. Here, we present developmental and genetic evidence for a fifth case of poecilogony via egg-size dimorphism in the Sacoglossa and the first example from the tropical Indo-Pacific. The sea slug Elysia pusilla produced both planktotrophic and lecithotrophic larvae in Guam and Japan. Levels of genetic divergence within populations were markedly low and rule out cryptic species. However, divergence among populations was exceptionally high (10-12% at the mitochondrial cytochrome c oxidase I locus), illustrating that extensive phylogeographic structure can persist in spite of the dispersal potential of planktotrophic larvae. We review reproductive, developmental, and ecological data for the five known cases of poecilogony in the Sacoglossa, including new data for Costasiella ocellifera from the Caribbean. We hypothesize that sacoglossans achieve lecithotrophy at smaller egg sizes than do related clades of marine heterobranchs, which may facilitate developmental plasticity that is otherwise vanishingly rare among animals. Insight into the environmental drivers and evolutionary results of shifts in larval type will continue to be gleaned from population-level studies of poecilogonous taxa like E. pusilla, and should inform life-history theory about the causes and consequences of alternative development modes in marine animals.

Effects of photosynthesis on the survival and weight retention of two kleptoplastic sacoglossan opisthobranchs

Many sacoglossan sea slugs utilize chloroplasts ingested from food algae for photosynthesis (functional kleptoplasty), and the extent and duration of kleptoplast retention differs greatly among sacoglossan species. Although most recent studies focus on the genetic, microscopic, or physiological mechanisms responsible for this unique phenomenon, its effects on the life history traits of sacoglossans have not been fully explored. To study the effects of light conditions on survival and weight retention, adult individuals of two sacoglossan species, Elysia trisinuata and Plakobranchus ocellatus (‘black type), were reared under light conditions (a 14-hour light: 10-hour dark photoperiod with an irradiance level of 28xa0µmolxa0m −2 s −1 ) or complete darkness for 21 days. There was no significant difference in the survival rate between the light and dark treatments for E. trisinuata , and its wet weight relative to the initial weight was smaller in the light than in the dark. However, both the survival and relative weights were greater in the light than dark for P. ocellatus . Based on the fluorescent yield measurement using pulse-amplitude-modulated fluorometry, the retention duration of functional chloroplasts was longer (>17 days) for P. ocellatus than E. trisinuata ( P. ocellatus benefits from photosynthesis for survival and growth, whereas E. trisinuata does not under starved conditions. This interspecific difference is likely related to the period of functional chloroplast retention.

Decadal-scale changes in the shallow subtidal benthos of an Irish marine reserve.

Abstract Long-term annual monitoring of rocky intertidal and shallow subtidal shores of an Irish sea lough (1990–2010) documented major regime shifts in the past decade. When population densities of the purple urchin Paracentrotus lividus plummeted in Lough Hyne Marine Reserve in southwestern Ireland, the warm-water fucalean alga Cystoseira foeniculacea and ephemeral algae proliferated shortly after. We discuss the possible influences of release from herbivory and climate change on this algal proliferation, which blanketed the benthos. Smothering of the benthos led to high levels of shallow subtidal anoxia. Furthermore, the invasive fucalean alga Sargassum muticum has made repeated incursions into the reserve. Although being reduced by persistent eradication efforts (2003–2011), Sargassum is spreading within the lough. Limited seawater flushing and propagule dispersal within the lough and eutrophication in coastal waters may have contributed to community-level changes. Whether the regime change is cyclical (contingent on re-establishment of urchins within the lough and continued eradication of S. muticum) or irreversible (the result of critical transitions) remains unclear.

Species Selection Favors Dispersive Life Histories in Sea Slugs, but Higher Per-Offspring Investment Drives Shifts to Short-Lived Larvae

For 40 years, paleontological studies of marine gastropods have suggested that species selection favors lineages with short-lived (lecithotrophic) larvae, which are less dispersive than long-lived (planktotrophic) larvae. Although lecithotrophs appeared to speciate more often and accumulate over time in some groups, lecithotrophy also increased extinction rates, and tests for state-dependent diversification were never performed. Molecular phylogenies of diverse groups instead suggested lecithotrophs accumulate without diversifying due to frequent, unidirectional character change. Although lecithotrophy has repeatedly originated in most phyla, no adult trait has been correlated with shifts in larval type. Thus, both the evolutionary origins of lecithotrophy and its consequences for patterns of species richness remain poorly understood. Here, we test hypothesized links between development mode and evolutionary rates using likelihood-based methods and a phylogeny of 202 species of gastropod molluscs in Sacoglossa, a clade of herbivorous sea slugs. Evolutionary quantitative genetics modeling and stochastic character mapping supported 27 origins of lecithotrophy. Tests for correlated evolution revealed lecithotrophy evolved more often in lineages investing in extra-embryonic yolk, the first adult trait associated with shifts in development mode across a group. However, contrary to predictions from paleontological studies, species selection actually favored planktotrophy; most extant lecithotrophs originated through recent character change, and did not subsequently diversify. Increased offspring provisioning in planktotrophs thus favored shifts to short-lived larvae, which led to short-lived lineages over macroevolutionary time scales. These findings challenge long-standing assumptions about the effects of alternative life histories in the sea. Species selection can explain the long-term persistence of planktotrophy, the ancestral state in most clades, despite frequent transitions to lecithotrophy.

Changes in brown seaweed distributions in Lough Hyne, SW Ireland: a long-term perspective

Abstract Distributions of brown seaweeds (Phaeophyceae: Fucales, Laminariales and Tilopteridales) were surveyed in 2011 in Europe’s first marine reserve, Lough Hyne in SW Ireland, and compared with distributions from three historical surveys (1930, 1955 and 1980). The most salient phycological differences were the incursion of the low intertidal and shallow subtidal species Fucus serratus, Himanthalia elongata and Saccharina latissima into the north basin of the marine reserve after the recent mass mortality of the purple urchin (Paracentrotus lividus). Monitoring surveys (1990–2012) at ten sites in the lough indicated that populations of S. latissima peaked in 1994, abruptly crashed in 1996 and then slowly recovered. As well as documenting the expansion of kelp, our annual, whole-lough snorkel surveys (2004–2012) demonstrated the recent proliferation of the introduced Sargassum muticum and native Cystoseira foeniculacea. Although the causal role of top-down factors (fishing ban and/or urchin population crash), bottom-up factors (nutrient enhancement), climatic variables (warming vs. episodic cooling) and pathogens (urchin and algal) is being widely debated, decadal-scale and inter-annual changes are clearly detectable and most consistent with a release from herbivory within the marine protected area.

Inventory of Japanese Sacoglossan Opisthobranchs: Historical Review, Current Records, and Unresolved Issues

Abstract: In the past ∼155 years, professional and amateur malacologists have recorded ca. 90 described species of sacoglossan opisthobranchs in ∼25 genera on Japanese shores. In addition, there are at least 20 to 40 undescribed or unrecognized sacoglossans also recorded. The extraordinary species richness has been a source of admiration as well as vexation. Worldwide scientific excitement in this group was largely due to two pivotal discoveries by Japanese researchers: (1) the acquisition and retention of functional chloroplasts by the sacoglossan Elysia atroviridis Baba, 1955 and (2) the existence of extant populations of bivalved sacoglossans (initially Tamanovalva limax Kawaguti and Baba, 1959 and then related taxa). Eight of the nine sacoglossan families recognized by Jensen (1996, 2007) are represented in Japan. All the recognized sacoglossan genera are represented in Japan except: Roburnella Marcus, 1982; Platyhedyle Salvini-Plawen, 1973; Gascoignella Jensen, 1985; Olea Agersborg, 1923; Limapontia Johnston, 1836; and the Australian genera Edenttellina Gatliff and Gabriel, 1911 and Midorigai Burn, 1960. Taxonomic uncertainty has been caused by the absence of vouchers, incomplete and/or questionable descriptions, photographic misidentifications (books and internet), chronically unstable classification, and other scientific challenges; in particular, the small size, cryptic coloration, and patchy distribution of sacoglossans have contributed to limited collections of many species. Since 2000, we have collected, photographed, and preserved unusually large numbers of Japanese sacoglossans, including species traditionally considered rare by malacologists. Although it is premature to produce a comprehensive inventory of the Japanese sacoglossan fauna, we consider it necessary to describe explicitly the strengths and weaknesses of current information. This assessment should assist professional and amateur malacologists with future sacoglossan study, particularly in the areas of biogeography, phylogeny, and ecology.

Subtropical Sacoglossans in Okinawa—At “Special Risk” or “Predictably Rare”?

On low intertidal and shallow subtidal shores on the west coast of Okinawa, Japan, we investigated the trophic associations of sacoglossan opisthobranchs associated with Bryopsidalean green algae. During 11 short research visits (55 days total) from 2002 to 2008, we recorded almost 500 specimens of 11 species. These sacoglossans include a new record for Japan (Caliphylla A. Costa, 1867), a recent record for Japan (Placida daguilarensis Jensen, 1990), two undescribed species (Placida Trinchese, 1876 and Elysia Risso, 1818), one unnamed (but well-described) species (Placida sp. sensu Baba 1986), and six other Indo-Pacific species. Not only did we record more sacoglossan species but also we found higher slug abundances than other colleagues in Okinawa or the Indo-Pacific region. Quantitative population attributes and feeding preferences are described for these sacoglossans. In contrast to temperate geographic regions, several of these Japanese sacoglossans specialized on a single algal genus rather than two or more genera in different families. This specificity is consistent with narrower host-plant associations in high-diversity communities; yet monophagy has not yet been demonstrated in this guild of Okinawan sacoglossans. Given the broad geographic ranges, restricted host ranges, often predictable populations, and high frequency of life cycles with planktotrophic larvae, western Pacific subtropical sacoglossans should be considered “predictably rare” (sensu Rabinowitz 1981) rather than at “special risk” (sensu Clark 1994).

Spatial variation in littoral Codium assemblages on Jersey, Channel Islands (southern English Channel)

Abstract Littoral surveys on Jersey shores in the Channel Islands (southern English Channel) revealed unexpected spatial variation in local-scale distributions of green macroalgae belonging to Codium species. Traditionally considered a low-shore alga, native C. vermilara inhabited high and/or mid-shore pools at nine of ten shores surveyed. This alga was frequent on semi-exposed shores. The introduced taxon C. fragile ssp. tomentosoides was patchily distributed with highest abundances in wave-sheltered areas. Zonation patterns of Codium spp. on Channel Island shores were more complex than those north of the English Channel.

Taxonomic Clarification in the Genus Elysia (Gastropoda: Sacoglossa): E. atroviridis and E. setoensis

Abstract. n Species of the genus Elysia Risso, 1818 have relatively few external characters useful for species identification, and features such as papillae and/or color markings on the body surface are often used to classify and identify species. Elysia atroviridis Baba, 1955 and E. setoensis Hamatani, 1968, for example, have been described as distinct based mainly on such characters. Close examinations of 46 specimens identifiable as either species collected from nine localities in Japan revealed that many specimens exhibited mixed features of the two species. Detailed comparisons of radular morphology could also not differentiate the two species. With phylogenetic analyses based on COI and 16S rRNA (1072 bp in total), all of the examined specimens formed a single clade with very little variation among specimens. The AMOVA of the specimen haplotypes showed there was no significant genetic differentiation in relation to differences in external morphology, geographic region, or algal host. Our results indicate that E. atroviridis and E. setoensis are conspecific. In contrast, cryptic species were found in two other clades of congeners, indicating that further taxonomic scrutiny is needed within the genus.