Crude Oil and Its Burnt Residues Induce Metamorphosis in Marine Invertebrates.

Metamorphosis is a critical process in the life cycle of most marine benthic invertebrates, determining their transition from plankton to benthos. It affects dispersal and settlement and therefore decisively influences the dynamics of marine invertebrate populations. An extended period of metamorphic competence is an adaptive feature of numerous invertebrate species that increases the likelihood of finding a habitat suitable for settlement and survival. We found that crude oil and residues of burnt oil rapidly induce metamorphosis in two different marine invertebrate larvae, a previously unknown sublethal effect of oil pollution. When exposed to environmentally realistic oil concentrations, up to 84% of tested echinoderm larvae responded by undergoing metamorphosis. Similarly, up to 87% of gastropod larvae metamorphosed in response to burnt oil residues. This study demonstrates that crude oil and its burned residues can act as metamorphic inducers in marine planktonic larvae, short-circuiting adaptive metamorphic delay. Future studies on molecular pathways and oil-bacteria-metamorphosis interactions are needed to fully understand the direct or indirect mechanisms of oil-induced metamorphosis in marine invertebrates. With 90% of chronic oiling occurring in coastal areas, this previously undescribed impact of crude oil on planktonic larvae may have global implications for marine invertebrate populations and biodiversity.

Dominance of Sulfurospirillum in Metagenomes Associated with the Methane Ice Worm (Sirsoe methanicola).

Sirsoe methanicola, commonly known as the methane ice worm, is the only macrofaunal species known to inhabit the Gulf of Mexico methane hydrates. Little is known about this elusive marine polychaete that can colonize rich carbon and energy reserves. Metagenomic analysis of gut contents and worm fragments predicted diverse metabolic capabilities with the ability to utilize a range of nitrogen, sulfur, and organic carbon compounds through microbial taxa affiliated with Campylobacterales, Desulfobacterales, Enterobacterales, SAR324, Alphaproteobacteria, and Mycoplasmatales. Entomoplasmatales and Chitinivibrionales were additionally identified from extracted full-length 16S rRNA sequences, and read analysis identified 196 bacterial families. Overall, the microbial community appeared dominated by uncultured Sulfurospirillum, a taxon previously considered free-living rather than host-associated. Metagenome-assembled genomes (MAGs) classified as uncultured Sulfurospirillum predicted thiosulfate disproportionation and the reduction of tetrathionate, sulfate, sulfide/polysulfide, and nitrate. Microbial amino acid and vitamin B12 biosynthesis genes were identified in multiple MAGs, suggesting nutritional value to the host. Reads assigned to aerobic or anaerobic methanotrophic taxa were rare. IMPORTANCE Methane hydrates represent vast reserves of natural gas with roles in global carbon cycling and climate change. This study provided the first analysis of metagenomes associated with Sirsoe methanicola, the only polychaete species known to colonize methane hydrates. Previously unrecognized participation of Sulfurospirillum in a gut microbiome is provided, and the role of sulfur compound redox reactions within this community is highlighted. The comparative biology of S. methanicola is of general interest given research into the adverse effects of sulfide production in human gut microbiomes. In addition, taxonomic assignments are provided for nearly 200 bacterial families, expanding our knowledge of microbiomes in the deep sea.

Distributions of microplastics and larger anthropogenic debris in Norfolk Canyon, Baltimore Canyon, and the adjacent continental slope (Western North Atlantic Margin, U.S.A.).

Anthropogenic debris has been reported in all studied marine environments, including the deepest parts of the sea. Finding areas of accumulation and methods of transport for debris are important to determine potential impacts on marine life. This study analyzed both sediment cores and Remotely Operated Vehicle video to determine the density and distribution of debris, including both micro- and macroplastics, in Norfolk and Baltimore canyons. The average microplastic density in Norfolk Canyon sediment was 37.30 plastic particles m-2 within the canyon and 21.03 particles m-2 on the adjacent slope, suggesting that microplastics could accumulate within submarine canyons. In video transects from both Norfolk and Baltimore canyons, the largest amounts of macroplastic were recorded near the canyon heads. Our findings contribute to a growing evidence base that canyons and their associated benthic invertebrate communities are important repositories and conduits for debris to the deep sea.

Megafauna of the UKSRL exploration contract area and eastern Clarion-Clipperton Zone in the Pacific Ocean: Annelida, Arthropoda, Bryozoa, Chordata, Ctenophora, Mollusca.

BACKGROUND: There is growing interest in mining polymetallic nodules from the abyssal Clarion-Clipperton Zone (CCZ) in the tropical Pacific Ocean. Despite having been the focus of environmental studies for decades, the benthic megafauna of the CCZ remain poorly known. To predict and manage the environmental impacts of mining in the CCZ, baseline knowledge of the megafauna is essential. The ABYSSLINE Project has conducted benthic biological baseline surveys in the UK Seabed Resources Ltd polymetallic-nodule exploration contract area (UK-1). Prior to ABYSSLINE research cruises in 2013 and 2015, no biological studies had been done in this area of the eastern CCZ. NEW INFORMATION: Using a Remotely Operated Vehicle and Autonomous Underwater Vehicle (as well as several other pieces of equipment), the megafauna within the UK Seabed Resources Ltd exploration contract area (UK-1) and at a site ~250 km east of the UK-1 area were surveyed, allowing us to make the first estimates of megafaunal morphospecies richness from the imagery collected. Here, we present an atlas of the abyssal annelid, arthropod, bryozoan, chordate, ctenophore and molluscan megafauna observed and collected during the ABYSSLINE cruises to the UK-1 polymetallic-nodule exploration contract area in the CCZ. There appear to be at least 55 distinct morphospecies (8 Annelida, 12 Arthropoda, 4 Bryozoa, 22 Chordata, 5 Ctenophora, and 4 Mollusca) identified mostly by morphology but also using molecular barcoding for a limited number of animals that were collected. This atlas will aid the synthesis of megafaunal presence/absence data collected by contractors, scientists and other stakeholders undertaking work in the CCZ, ultimately helping to decipher the biogeography of the megafauna in this threatened habitat.

Salinity and Temperature Tolerance of the Nemertean Worm Carcinonemertes errans, an Egg Predator of the Dungeness Crab.

Estuaries can be harsh habitats for the marine animals that enter them, but they may also provide these species with sub-saline refuges from their parasites. The nemertean egg predator Carcinonemertes errans is known to occur less frequently and in smaller numbers on its host, the Dungeness crab Metacarcinus magister, when the hosts are found within estuaries. We examined the temperature and salinity tolerances of C. errans to determine if this observed distribution represents a true salinity refuge. We monitored the survival of juvenile and larval worms exposed to ecologically relevant salinities (5-30) and temperatures (8-20 °C) over the course of several days under laboratory conditions. Juvenile worms were unaffected by the experimental temperature levels and exhibited robustness to salinity treatments 25 and 30. However, significant mortality was seen at salinity treatments 20 and below. Larvae were less tolerant than juveniles to lowered salinity and were also somewhat more susceptible to the higher temperatures tested. Given that the Dungeness crab can tolerate forays into mesohaline (salinity 5-18) waters for several days at a time, our findings suggest that salinity gradients play an important role in creating a parasite refuge for this species within the estuaries of the Pacific Northwest.

Larvae from deep-sea methane seeps disperse in surface waters.

Many species endemic to deep-sea methane seeps have broad geographical distributions, suggesting that they produce larvae with at least episodic long-distance dispersal. Cold-seep communities on both sides of the Atlantic share species or species complexes, yet larval dispersal across the Atlantic is expected to take prohibitively long at adult depths. Here, we provide direct evidence that the long-lived larvae of two cold-seep molluscs migrate hundreds of metres above the ocean floor, allowing them to take advantage of faster surface currents that may facilitate long-distance dispersal. We collected larvae of the ubiquitous seep mussel "Bathymodiolus" childressi and an associated gastropod, Bathynerita naticoidea, using remote-control plankton nets towed in the euphotic zone of the Gulf of Mexico. The timing of collections suggested that the larvae might disperse in the water column for more than a year, where they feed and grow to more than triple their original sizes. Ontogenetic vertical migration during a long larval life suggests teleplanic dispersal, a plausible explanation for the amphi-Atlantic distribution of "B." mauritanicus and the broad western Atlantic distribution of B. naticoidea. These are the first empirical data to demonstrate a biological mechanism that might explain the genetic similarities between eastern and western Atlantic seep fauna.

Impacts of an endoparasitic copepod, Ismaila belciki, on the reproduction, growth and survivorship of its nudibranch host, Janolus fuscus.

Copepods from the genus Ismaila are large endoparasites that inhabit the main body cavity and/or cerata of opisthobranch molluscs. These parasites exhibit many life history characteristics typically found in parasitic castrators, yet the actual impact of infection on reproduction, growth or survivorship of the hosts are unknown. On the Oregon (USA) coast, Ismaila belciki can infect over 80% of their hermaphroditic hosts, Janolus fuscus. In laboratory mating experiments, we compared the reproductive output (egg mass weight, number of egg capsules, number of viable embryos) and the gonadal somatic index of infected versus uninfected J. fuscus. Infected J. fuscus could produce viable sperm and copulate. Mating with an infected individual did not limit a sea slug's reproductive output. However, infected J. fuscus had significantly lower reproductive output (by 34-54%), producing smaller egg masses with fewer capsules and viable embryos. Infected hosts had significantly lower gonadal somatic index than their uninfected counterparts, although there was no significant difference in gonadal somatic index between hosts with single and double infections. By collecting the egg sacs produced by the copepod parasite during experiments, we estimated that 25-34% of the host's reproductive output is usurped by the parasite and re-directed to the parasite's own reproduction. In the laboratory, infection did not alter growth in J. fuscus. However, infection significantly decreased survivorship in mature (but not immature) nudibranch hosts. These results suggest that I. belciki is not a true castrator, but it does reduce the reproductive output of its host and may therefore limit the natural population size of J. fuscus.

Fixed, free, and fixed: the fickle phylogeny of extant Crinoidea (Echinodermata) and their Permian-Triassic origin.

Although the status of Crinoidea (sea lilies and featherstars) as sister group to all other living echinoderms is well-established, relationships among crinoids, particularly extant forms, are debated. All living species are currently placed in Articulata, which is generally accepted as the only crinoid group to survive the Permian-Triassic extinction event. Recent classifications have recognized five major extant taxa: Isocrinida, Hyocrinida, Bourgueticrinina, Comatulidina and Cyrtocrinida, plus several smaller groups with uncertain taxonomic status, e.g., Guillecrinus, Proisocrinus and Caledonicrinus. Here we infer the phylogeny of extant Crinoidea using three mitochondrial genes and two nuclear genes from 59 crinoid terminals that span the majority of extant crinoid diversity. Although there is poor support for some of the more basal nodes, and some tree topologies varied with the data used and mode of analysis, we obtain several robust results. Cyrtocrinida, Hyocrinida, Isocrinida are all recovered as clades, but two stalked crinoid groups, Bourgueticrinina and Guillecrinina, nest among the featherstars, lending support to an argument that they are paedomorphic forms. Hence, they are reduced to families within Comatulida. Proisocrinus is clearly shown to be part of Isocrinida, and Caledonicrinus may not be a bourgueticrinid. Among comatulids, tree topologies show little congruence with current taxonomy, indicating that much systematic revision is required. Relaxed molecular clock analyses with eight fossil calibration points recover Articulata with a median date to the most recent common ancestor at 231-252mya in the Middle to Upper Triassic. These analyses tend to support the hypothesis that the group is a radiation from a small clade that passed through the Permian-Triassic extinction event rather than several lineages that survived. Our tree topologies show various scenarios for the evolution of stalks and cirri in Articulata, so it is clear that further data and taxon sampling are needed to recover a more robust phylogeny of the group.

Deep sequencing of Myxilla (Ectyomyxilla) methanophila, an epibiotic sponge on cold-seep tubeworms, reveals methylotrophic, thiotrophic, and putative hydrocarbon-degrading microbial associations.

The encrusting sponge Myxilla (Ectyomyxilla) methanophila (Poecilosclerida: Myxillidae) is an epibiont on vestimentiferan tubeworms at hydrocarbon seeps on the upper Louisiana slope of the Gulf of Mexico. It has long been suggested that this sponge harbors methylotrophic bacteria due to its low δ(13)C value and high methanol dehydrogenase activity, yet the full community of microbial associations in M. methanophila remained uncharacterized. In this study, we sequenced 16S rRNA genes representing the microbial community in M. methanophila collected from two hydrocarbon-seep sites (GC234 and Bush Hill) using both Sanger sequencing and next-generation 454 pyrosequencing technologies. Additionally, we compared the microbial community in M. methanophila to that of the biofilm collected from the associated tubeworm. Our results revealed that the microbial diversity in the sponges from both sites was low but the community structure was largely similar, showing a high proportion of methylotrophic bacteria of the genus Methylohalomonas and polycyclic aromatic hydrocarbon (PAH)-degrading bacteria of the genera Cycloclasticus and Neptunomonas. Furthermore, the sponge microbial clone library revealed the dominance of thioautotrophic gammaproteobacterial symbionts in M. methanophila. In contrast, the biofilm communities on the tubeworms were more diverse and dominated by the chemoorganotrophic Moritella at GC234 and methylotrophic Methylomonas and Methylohalomonas at Bush Hill. Overall, our study provides evidence to support previous suggestion that M. methanophila harbors methylotrophic symbionts and also reveals the association of PAH-degrading and thioautotrophic microbes in the sponge.

Dispersal of deep-sea larvae from the intra-American seas: simulations of trajectories using ocean models.

Using data on ocean circulation with a Lagrangian larval transport model, we modeled the potential dispersal distances for seven species of bathyal invertebrates whose durations of larval life have been estimated from laboratory rearing, MOCNESS plankton sampling, spawning times, and recruitment. Species associated with methane seeps in the Gulf of Mexico and/or Barbados included the bivalve "Bathymodiolus" childressi, the gastropod Bathynerita naticoidea, the siboglinid polychaete tube worm Lamellibrachia luymesi, and the asteroid Sclerasterias tanneri. Non-seep species included the echinoids Cidaris blakei and Stylocidaris lineata from sedimented slopes in the Bahamas and the wood-dwelling sipunculan Phascolosoma turnerae, found in Barbados, the Bahamas, and the Gulf of Mexico. Durations of the planktonic larval stages ranged from 3 weeks in lecithotrophic tubeworms to more than 2 years in planktotrophic starfish. Planktotrophic sipunculan larvae from the northern Gulf of Mexico were capable of reaching the mid-Atlantic off Newfoundland, a distance of more than 3000 km, during a 7- to 14-month drifting period, but the proportion retained in the Gulf of Mexico varied significantly among years. Larvae drifting in the upper water column often had longer median dispersal distances than larvae drifting for the same amount of time below the permanent thermocline, although the shapes of the distance-frequency curves varied with depth only in the species with the longest larval trajectories. Even species drifting for >2 years did not cross the ocean in the North Atlantic Drift.

Larval development and metamorphosis of the deep-sea cidaroid urchin Cidaris blakei.

Cidaroids, one of the two major sister clades of sea urchins, first appeared during the lower Permian (ca. 270 mya) and are considered to represent the primitive form of all living echinoids. This study of Cidaris blakei, a deep-sea cidaroid urchin with planktotrophic larvae, provides a description of development from fertilization through early juvenile stages and is the first report of a deep-sea urchin reared through metamorphosis. C. blakei resembles other cidaroids in its lack of a cohesive hyaline layer, the absence of an amniotic invagination for juvenile rudiment formation, and the presence of spines with a single morphotype at metamorphosis. C. blakei differed from other cidaroids in the presence of an apical tuft, the extent of fenestration of postoral skeletal rods, the shape of juvenile spines, and an extended (14-day) lecithotrophic stage prior to development of a complete gut. The development of C. blakei, 120 days from fertilization to metamorphosis, was protracted relative to that of shallow-water cidaroids. Preliminary work on temperature tolerances suggests that C. blakei larvae would be unable to survive the warmer temperatures higher in the water column and are therefore unable to vertically migrate.

Complete development of the northeast Pacific arminacean nudibranch Janolus fuscus.

Early life-history stages of the nudibranch Janolus fuscus (superfamily Arminoidea) were reared in the laboratory and collected from the field to document embryology, larval morphology, and pre-and post-metamorphic growth. This species produces Type B egg masses containing an average of 23,063 viable embryos. The spiral holoblastic cleavage pattern resembled that of other nudibranchs, with early cell divisions occurring roughly at 4-h intervals at 11-13 °C. Embryos progress through a gastrula stage with cellular extensions covering the blastopore and a trochophore-like stage before hatching as veliger larvae after 10-18 days. Veligers ranged in size from 125-153.8 µm at hatching, with size being positively correlated with the duration of encapsulated development. After a 36-41-day larval period, some veligers ceased growing at 266 µm and showed signs of approaching competence. Four larvae settled and two metamorphosed into 280-µm juveniles on the bryozoan prey of the adults, Bugula pacifica, 46 and 54 days post-hatching. Average growth of pre-ovipositional (<19 mm) J. fuscus was more rapid (8.79% per day) than growth of ovipositional individuals (3.52% per day). Growth continued to a maximum of 57 mm in the laboratory, with an estimated total lifespan of about 5 months. These data, which agree with concurrent field observations, suggest a subannual life cycle.

New molluscan larval form: brooding and development in a hydrothermal vent gastropod, Ifremeria nautilei (Provannidae).

Despite extreme differences between some shallow and deep-sea habitats, the developmental modes and larval forms of deep-sea animals are typically similar to those of their shallow-water relatives. Here we report one of the first documented exceptions to this general rule. The hydrothermal vent snail Ifremeria nautilei displays two novel life-history traits: (1) an unusual uniformly ciliated larva that we here name Warén's larva, and (2) internal brood protection in a modified metapodial pedal gland. Warén's larva emerges from the internal brood pouch as a fully ciliated lecithotrophic larva with a unique external cuticle. The larvae swim with their posterior end forward and metamorphose into typical veliger larvae after 15 days at room temperature. Warén's larva is the only known example of a free-swimming pre-veliger larval stage in the higher gastropods and is the first new gastropod larval form to be described in more than 100 years.

Spawning, development, and the duration of larval life in a deep-sea cold-seep mussel.

We describe culturing techniques and development for the cold-seep mussel "Bathymodiolus" childressi, the only deep-sea bivalve for which development has been detailed. Spawning was induced in mature mussels by injection of 2 mmol l(-1) serotonin into the anterior adductor muscle. The mean egg diameter is 69.15 +/- 2.36 microm (+/-S.D.; n = 50) and eggs are negatively buoyant. Cleavages are spiral and at 7-8 degrees C occur at a rate of one per 3-9 h through hatching, with free-swimming blastulae hatching by 40 h and shells beginning to develop by day 12. When temperature was raised to 12-14 degrees C after hatching, larvae developed to D-shell veligers by day 8 without being fed. Egg size and larval shell morphology indicate that "B." childressi has a planktotrophic larva, but we did not observe feeding in culture. Wide distribution of this species throughout the Gulf of Mexico and amphi-Atlantic distributions of closely related congeners suggest that larvae may spend extended periods in the plankton. Duration of larval life was estimated for "B." childressi by comparing calculated settlement times to known spawning seasons. These estimates suggest variability in the larval duration, with individuals spending more than a year in the plankton.

Vailulu'u Seamount, Samoa: Life and death on an active submarine volcano.

Submersible exploration of the Samoan hotspot revealed a new, 300-m-tall, volcanic cone, named Nafanua, in the summit crater of Vailulu'u seamount. Nafanua grew from the 1,000-m-deep crater floor in <4 years and could reach the sea surface within decades. Vents fill Vailulu'u crater with a thick suspension of particulates and apparently toxic fluids that mix with seawater entering from the crater breaches. Low-temperature vents form Fe oxide chimneys in many locations and up to 1-m-thick layers of hydrothermal Fe floc on Nafanua. High-temperature (81 degrees C) hydrothermal vents in the northern moat (945-m water depth) produce acidic fluids (pH 2.7) with rising droplets of (probably) liquid CO(2). The Nafanua summit vent area is inhabited by a thriving population of eels (Dysommina rugosa) that feed on midwater shrimp probably concentrated by anticyclonic currents at the volcano summit and rim. The moat and crater floor around the new volcano are littered with dead metazoans that apparently died from exposure to hydrothermal emissions. Acid-tolerant polychaetes (Polynoidae) live in this environment, apparently feeding on bacteria from decaying fish carcasses. Vailulu'u is an unpredictable and very active underwater volcano presenting a potential long-term volcanic hazard. Although eels thrive in hydrothermal vents at the summit of Nafanua, venting elsewhere in the crater causes mass mortality. Paradoxically, the same anticyclonic currents that deliver food to the eels may also concentrate a wide variety of nektonic animals in a death trap of toxic hydrothermal fluids.

Influence of environmental conditions on early development of the hydrothermal vent polychaete Alvinella pompejana.

Dispersal and colonisation processes at deep-sea vents are still not fully understood, essentially because early life stages of vent species remain unknown. The polychaete worm Alvinella pompejana forms colonies on chimney walls at East Pacific Rise vent sites where the temperature can frequently exceed 20 degrees C. In vitro studies in pressure vessels showed that the early embryos tolerate temperatures in a lower range (10-14 degrees C), suggesting that they would have to escape the colony to develop. Pressure vessels offer the advantage that each parameter can be independently controlled, but they do not simulate the more complex and dynamic conditions naturally encountered at vent sites. Accordingly, in addition to incubations in pressure vessels, we incubated embryos directly at a vent site, in different habitats along a gradient of hydrothermal influence. Embryos incubated on an adult A. pompejana colony where temperature and H(2)S concentrations were relatively high showed a very low survival rate and did not develop, whereas embryos incubated in a Riftia pachyptila clump environment with a lower hydrothermal signature, or at the base of the chimney where the influence of the hydrothermal activity was very weak, survived well and developed. Although the average temperature recorded in the A. pompejana colony was within the range tolerated by embryos (13 degrees C), frequent peaks above 20 degrees C were recorded. Estimated sulphide concentration at this site reached 200 mumol l(-1). Punctuated exposure to both high temperature and elevated sulphide levels probably explain the low survival of embryos within the A. pompejana colony. The in situ experiments further support the idea that embryos require conditions with moderate hydrothermal influence not generally found within an adult colony. However, as much more benign physicochemical conditions can be found within a few tens of cm of adult colonies, embryos do not necessarily have to leave their vent of origin to develop. Further analyses are needed to pinpoint the specific factors that affect the survival and development of embryos at vents.

Sperm storage, internal fertilization, and embryonic dispersal in vent and seep tubeworms (Polychaeta: Siboglinidae: Vestimentifera).

Vestimentiferan tubeworms are ecologically important members of deep-sea chemosynthetic communities, including hydrothermal vents and cold seeps. Some are community dominants and others are primary colonists of new vent sites; they include some of the longest living and fastest growing marine invertebrates. Their mechanisms of propagation, dispersal, and genetic exchange have been widely discussed. Direct sperm transfer from males to females has been documented in one species, Ridgeia piscesae, but others are known to discharge what are apparently primary oocytes. Brooding of embryos has never been observed in any vestimentiferan. These observations have led to the supposition that fertilization might be external in most species. Here we report sperm storage at the posterior end of the oviduct in five species, including tubeworms from both vents and seeps. We show experimentally that most eggs are inseminated internally, that fertilization rate is typically lower than 100%, that meiosis is completed after eggs are released from the female, and that the dispersal phase includes the entire embryonic period.

Spermiogenesis and modified sperm morphology in the "seepworm" Methanoaricia dendrobranchiata (Polychaeta: Orbiniidae) from a methane seep environment in the Gulf of Mexico: implications for fertilization biology.

Spermatogenesis and mature sperm morphology have been described along with limited observations of the ovary in Methanoaricia dendrobranchiata, an orbiniid polychaete associated with dense populations of the mussel Bathymodiolus childressi at brine pools on the Louisiana slope, Gulf of Mexico. The species is gonochoric with gonads serially repeated in numerous segments and each associated with a nexus of blood vessels at the base of the parapodia. In the female, synchronous, intraovarian egg development occurs with the release from the ovary of large, yolky eggs into the coelom at first meiotic metaphase. Sperm develop in the coelom as free-floating, plasmodial clones interconnected via an anuclear cytophore. At the end of spermiogenesis, mature spermatozoa float freely in the coelom. The mature spermatozoon differs significantly from that of shallow-water orbiniid species by possessing an elongated nucleus and a greatly elongated and curved acrosome reaching 19.5 microm in length. The spermatozoon resembles an ent-aquasperm and may not fertilize the eggs directly in seawater in the classical manner. We hypothesize that the unusual spermatozoon morphology in this species has evolved due to the hypoxic environment in which the adults live and that fertilization biology is likely modified in some way to minimize sperm exposure to high levels of hydrogen sulfide. An analysis of life-history features in shallow-water orbiniids is used to infer reproductive features in M. dendrobranchiata that could not be directly documented.