The Bryozoan Cauloramphus magnus (Cheilostomata: Calloporidae) in Northern Japan Includes Multiple, Co-occurring Cryptic Species.

The cheilostome bryozoan Cauloramphus magnus is common in the rocky intertidal habitat from southeastern Alaska to northern Japan. We examined its phylogeography by analyzing 576 bp of the mitochondrial COI (cox1) gene sequenced for 298 colonies from 16 localities in northern Japan. A maximum-likelihood phylogeny detected three main clades (A, B, C). Clades A and B occurred throughout the study area but differed in frequency, haplotype diversity, and haplotype distribution; each resolved into three divergent subclades (AI-III, BI-III). Clade A shared none among 15 haplotypes between the Pacific and Sea of Japan sides of Hokkaido. In contrast, Clade B (29 haplotypes) was thrice as common as Clade A among samples, with haplotype B28 common on both sides. Divergent Clade C (nine haplotypes) was detected only at Rumoi. K2P divergences of 12.3-28.3% among Clades A-C suggest these are distinct biological species, a conclusion supported by different inferred evolutionary histories. A bPTP species delimitation analysis indicated nine phylogenetic species among the sequences included in our phylogeny (AI-III, BI-III, C, and one specimen each from Alaska and the Commander Islands), with K2P divergences of 3.9-6.5% among subclades in A or B. Statistical and principal components analyses suggested weak morphological differentiation between Clades A + B and C, although overlapping ranges of measurements preclude identification to clade; these three clades are morphologically cryptic. For taxonomy, we suggest retaining the name C. magnus for lineages within this species complex across its range, followed by a clade designation, if known.

Two new steroid sulfates from a cheilostome bryozoan, Calyptotheca sp.

Two new steroid sulfates 1 and 2 were obtained from a lipophilic extract of an undescribed bryozoan species in the genus Calyptotheca. The structures of compounds 1 and 2 were elucidated by spectroscopic methods and chemical modifications. Steroids 1 and 2 exhibited moderate cytotoxicity at IC50 54 and 30 µM, respectively, against NBT-T2 cells.

The Cribrilina annulata" problem and new species of Juxtacribrilina (Bryozoa: Cheilostomata: Cribrilinidae) from the North Pacific.

Originally described from Greenland, Juxtacribrilina annulata (Fabricius, 1780) (previously known as Cribrilina annulata) has long been regarded as having a circumpolar, Arctic-boreal distribution. The genus Juxtacribrilina Yang, Seo, Min, Grischenko Gordon, 2018 accommodated J. annulata and three related North Pacific species formerly in Cribrilina Gray, 1848 that lack avicularia, have a reduced (hood-like, cap-like, or vestigial) ooecium closely associated with modified latero-oral spines to form an ooecial complex, and produce frontally or marginally positioned dwarf ovicellate zooids. While the recently described NW Pacific species J. mutabilis and J. flavomaris, which have a vestigial ooecium like a short, flattened spine, clearly differ from J. annulata, the differences between J. annulata and other Pacific populations remained unclear. Here we provide descriptions for five species from the North Pacific region. We identified a specimen from the Sea of Okhotsk as J. annulata. Among the other four species, J. ezoensis n. sp. has a trans-Pacific distribution (abundant at Akkeshi, Hokkaido, Japan; also detected in the Commander Islands and at Ketchikan, Southeast Alaska); J. pushkini n. sp. was found only at Ketchikan; J. dobrovolskii n. sp. was found only at Shikotan Island in the Lesser Kuril Chain; and J. tumida n. sp. was found only at Kodiak, Gulf of Alaska. These four species all differ from J. annulata in having one or two frontal pore chambers on the proximal gymnocyst of most zooids; in budding frontal dwarf ovicellate zooids from these chambers rather than from basal pore chambers; in producing dwarf zooids more abundantly; and in having ooecia that are somewhat to markedly more reduced (cap-like rather than hood-like) and more closely integrated with the modified latero-oral spines. Furthermore, in the Pacific species, the ooecium in basal zooids arises from the roof of the distal pore chamber of the maternal zooid; ovicellate zooids can thus also bud a distal autozooid and are often arranged in columnar series with other zooids. In J. annulata, the hood-like kenozooidal ooecium budded from the maternal zooid replaces the distal autozooid, and ovicellate zooids are thus usually not embedded in a columnar series.

Global Distribution and Variation of the Invasive Cheilostome Bryozoan Cribrilina mutabilis.

Viable populations of the cheilostome bryozoan Cribrilina mutabilis Ito, Onishi & Dick exist in the NW Pacific (Russian Far East and northern Japan), NE Atlantic (Scandinavia and Scotland), and NW Atlantic (Maine, USA). The first NE and NW Atlantic records are from Norway (2008) and Casco Bay, Maine, USA (2018), respectively, indicating a relatively recent introduction to the region. Mitochondrial COI gene sequences from North Atlantic populations (Sweden, Norway, and Maine) showed two haplotypes differing by one substitution, but differed from two haplotypes from Akkeshi, northern Japan, by 6-8 substitutions. North Atlantic populations differed morphologically from the Akkeshi population in that some zooids formed a suboral projection, and frontal zooids were more common. While C. mutabilis in northern Japan has been found only on natural or artificial eelgrass (Zostera marina), across its range it has been found on several species of algae, plastic panels and strips, several species of Zostera, and mollusc shells. Similar frequencies of heteromorphic zooids with differing degree of frontal wall calcification, i.e., R (rib)-, I (intermediate)-, and S (shield)-type zooids, in colonies on eelgrass at comparable times of the season and across populations suggest an innate response to seasonal environmental fluctuations, although zooid frequencies were different on non-eelgrass substrates. The increase in trans-Arctic shipping along the Northern Sea Route in recent decades, and previous documentation of C. mutabilis on ship hulls in the Sea of Japan, indicate a clear mechanism for anthropogenic introduction from the Far East to Europe in recent decades.

Taxonomy and diversity of coelobite bryozoans from drift coral cobbles on Co To Island, northern Vietnam.

There has been no previous report detailing the taxonomy of marine bryozoans along the coasts of Vietnam. Here we report on the taxonomy and diversity of bryozoans collected among drift coral cobbles from a beach in the tropical Co To archipelago, Gulf of Tonkin, northern Vietnam. We detected 27 bryozoan species (23 cheilostomes, four cyclostomes) in a coelobite assemblage inhabiting crevices in the cobbles, and holes made by boring molluscs. The degree of bryozoan preservation varied greatly, suggesting that the cobbles had accumulated on the beach over a period of months to years, or even decades. Coral reefs in the Co To archipelago underwent a catastrophic decline in 2003-2008, and it is unclear whether the bryozoan assemblage reflects past diversity, present diversity remaining in the coral rubble, or both. We describe six new species: Parasmittina acondylata n. sp., Metroperiella cotoensis n. sp., Microporella tonkinensis n. sp., Rhynchozoon setiavicularium n. sp., R. latiavicularium n. sp., and Disporella phaohoa n. sp. All but two of the previously described species were already known from the Central Indo-Pacific coastal biogeographical realm of Spalding et al. (2007), which includes Vietnam. We report the third Recent record of the thalamoporellid Dibunostoma reversum (Harmer, 1926), which is quite similar to and might be conspecific with the lower Miocene species Thalamoporella transversa Guha Krishna, 2004; while it is premature to synonymize the two, we transfer T. transversa to Dibunostoma, as D. transversum. The calcareous, sheet-like, encrusting foraminiferan Planorbulina larvata was prominent in the coelobite assemblage and was often observed in substrate competition with bryozoans. A limited analysis of competitive interactions indicated that the encrusting bryozoans in the coelobite assemblage encountered P. larvata more often than they encountered other bryozoans, and that P. larvata out-competed bryozoans for substrate, reinforcing a growing sense of the importance of encrusting foraminifera in tropical and subtropical hard-substrate communities.

Cribrilina mutabilis n. sp., an Eelgrass-Associated Bryozoan (Gymnolaemata: Cheilostomata) with Large Variationin Zooid Morphology Related to Life History.

We describe the cribrimorph cheilostome bryozoan Cribrilina mutabilis n. sp., which we detected as an epibiont on eelgrass (Zostera marina) at Akkeshi, Hokkaido, northern Japan. This species shows three distinct zooid types during summer: the R (rib), I (intermediate), and S (shield) types. Evidence indicates that zooids commit to development as a given type, rather than transform from one type to another with age. Differences in the frontal spinocyst among the types appear to be mediated by a simple developmental mechanism, acceleration or retardation in the production of lateral costal fusions as the costae elongate during ontogeny. Colonies of all three types were identical, or nearly so, in partial nucleotide sequences of the mitochondrial COI gene (555-631 bp), suggesting that they represent a single species. Zooid types varied temporally in overall frequency in the population: colonies contained nearly exclusively R-type zooids in mid-June; predominantly I-type, or both R- and I-type, zooids in mid-July; and I-type, S-type, or both I- and S-type zooids (interspersed or in discrete bands) in mid- to late August. Reproduction occurred throughout the season, but peaked in July, with only R- and I-type zooids reproducing. Reproductive zooids bear a vestigial compound (tripartite) ooecium and brood internally; S-type zooids, first appearing in August, were non-reproductive, which suggests that they may serve as an overwintering stage. As this species is easily accessible, common, and simple in form, it is potentially useful as a model system for studying polyphenism at multiple levels (zooid, colony, and population) in the context of life-history adaptations.

Two kinorhynch species (Cyclorhagida, Echinoderidae, Echinoderes) show different distribution patterns across Tsugaru Strait, Northern Japan.

We investigated the geographic population structures of two intertidal kinorhynch species, Echinoderes sensibilis and Echinoderes sp. A, in the vicinity of Tsugaru Strait between Honshu and Hokkaido Islands, Japan, to examine whether the distribution or connectivity of populations of either species has been constrained by the strait. For each species, we examined the geographic distribution of COI haplotypes, constructed a median-joining haplotype network, and calculated statistics of genetic variation and connectivity. Tsugaru Strait is the northern range limit for E. sensibilis, which comprises a large, evolutionarily stable metapopulation that appears to have undergone a reduction in size followed by expansion; connectivity is low among most local populations, including across Tsugaru Strait. A divergent haplotype lineage showing no variation occurred only at Horozuki, suggesting recent immigration there from outside the study area. Echinoderes sp. A underwent a severe population bottleneck followed by rapid expansion. It occurred at all sampling sites on both sides of the strait, with high connectivity between populations across the strait. There is a zone of secondary contact between moderately divergent, presumably previously allopatric lineages in eastern Hokkaido. Present-day conditions in the strait have existed only for the past 8000 years, and differences in these species' distributions and apparent connectivity across the strait may relate to conditions existing in the strait when the species underwent population expansions or shifts in range; these historical events were not necessarily concurrent between the species, and occurred more than 8000 years ago. We discuss dispersal mechanisms for kinorhynchs, which could include suspension transport or rafting.

Limited phylogenetic distribution of a long tandem-repeat cluster in the mitochondrial control region in Bubo (Aves, Strigidae) and cluster variation in Blakiston's fish owl (Bubo blakistoni).

To investigate the phylogenetic position of Blakiston's fish owl (Bubo blakistoni), we sequenced the mitochondrial (mt) DNA control region and cytochrome b (cyt b) for nine Bubo species. Maximum-likelihood analyses of combined control region and cyt b sequences, and cyt b sequences alone, showed that species formerly placed in genus Ketupa comprise a monophyletic group. Unexpectedly, we discovered a long cluster of 20-25 tandem repeat units 77 or 78bp long in the third control region domain in four of the nine Bubo species for which the control region was sequenced (B. blakistoni, B. flavipes, and B. ketupu in the Ketupa clade; B. lacteus), leading to overall control region lengths of 3.0-3.8kpb estimated from agarose gel electrophoresis. The control region in B. lacteus is the longest (3.8kbp) reported to date in vertebrates. Sequencing of eight repeat units at each end of the cluster in 20 B. blakistoni individuals detected several types of repeat units 77 or 78bp long, and six patterns in the order of unit types. The occurrence of a repeat cluster in all three species examined in the Ketupa clade suggests their common ancestor also had a cluster, whereas a maximum parsimony tree showed repeat-unit types grouping by species, rather than by paralog groups, suggesting independent origins of the clusters. We reconcile these results with a turnover model, in which the range in cluster-length variation and unit types at the 5' end are hypothetically functionally constrained by the protein-binding function of the control region, but otherwise there is a continual turnover of units in evolutionary time, with new unit types arising through mutations, proliferating by duplication of single and double repeat blocks, and being lost through deletion. Estimated free energies for reconstructed secondary structures of single and especially pairs of repeat units were higher than for homologous single-unit blocks in species lacking a repeat cluster, supporting slipped-strand mispairing as the mechanism of cluster turnover.

Are plumatellid statoblasts in freshwater bryozoans phylogenetically informative?

Morphological characters of statoblasts (including floatoblasts and sessoblasts) in freshwater bryozoans have been important in phylactolaemate systematics and identification in that older phylogenetic hypotheses relied heavily on statoblast morphology. To assess the reliability of statoblast characters in drawing conclusions about phylogeny, we examined the phylogenetic distribution of metric and proportional floatoblast characters, floatoblast symmetry, and floatoblast and sessoblast microsculpture in Plumatellidae, the largest family of phylactolaemates, in the context of molecular phylogenetic reconstructions based on nucleotide sequences of parts of the mitochondrial 12S rRNA, 16S rRNA, and cytochrome b (cytb) genes. Adding cytb sequences to a previous phylogeny based on 12S and 16S increased support only for a node including Plumatella vaihiriae in the P. repens clade. Characters of gross floatoblast morphology were generally not phylogenetically informative individually, but collectively discriminated among members of three relatively well-supported clades that were analyzed as pre-defined groups in a discriminant analysis. Two characters of floatoblast microsculpture (reticulation and hypertubercles) were restricted to particular clades; other characters (e.g., villi) were clearly convergent. In nine of 11 cases, fenestral microsculpture was identical or partly correlated between the floatoblast and sessoblast of a species. Overall, our results indicate that statoblast morphology is not highly phylogenetically constrained.

Cribrimorph and other Cauloramphus species (Bryozoa: Cheilostomata) from the Northwestern Pacific.

We provide original descriptions for nine new species in the cheilostome bryozoan genus Cauloramphus (C. gracilis, C. Ordinarius, C. amphidisjunctus, C. cheliferoides, C. oshurkovi, C. infensus, C. parvus, C. peltatus, and C. ascofer) and a redescription of C. disjunctus Canu and Bassler, 1929 . We delineate a group of eight species, here termed the 'C. disjunctus clade,' that have the opesial spine joints calcified to a greater or lesser extent in mature zooids; most also have paired, hypertrophied avicularia. This group includes C. amphidisjunctus, C. cheliferoides, C. infensus, C. parvus, C. peltatus, and C. ascofer in the Aleutian Islands, Alaska; C. oshurkovi in the Commander Islands; and C. disjunctus in Japan. High levels of apparent endemism in two unrelated bryozoan genera (Cauloramphus and Monoporella), and geographical population differentiation in C. ascofer indicating ongoing allopatric speciation, suggest high speciation rates for deep benthic bryozoans in the western Aleutians. A phylogenetic hypothesis for the C. disjunctus clade indicates that populations of Cauloramphus dispersed between the Aleutians and Asia on at least three separate occasions, and that the polarity of at least two of these dispersal events was from the Aleutians to Asia.

The origin of ascophoran bryozoans was historically contingent but likely.

The degree to which evolutionary outcomes are historically contingent remains unresolved, with studies at different levels of the biological hierarchy reaching different conclusions. Here we examine historical contingency in the origin of two evolutionary novelties in bryozoans, a phylum of colonial animals whose fossil record is as complete as that of any major group. In cheilostomes, the dominant living bryozoans, key innovations were the costal shield and ascus, which first appeared in the Cretaceous 85-95 Myr ago. We establish the parallel origin of these structures less than 12 Myr ago in an extant bryozoan genus, Cauloramphus, with transitional stages remarkably similar to those inferred for a Cretaceous clade. By one measure, long lag times in the first origins of costal shield and ascus suggest a high degree of historical contingency. This, however, does not equate with dependence on a narrow set of initial conditions or a low probability of evolution. More than one set of initial conditions may lead to an evolutionary outcome, and alternative sets are not entirely independent. We argue that, although historically contingent, the origin of ascus and costal shield was highly likely with sufficient possibilities afforded by time.

Unexpectedly high diversity of Monoporella (Bryozoa: Cheilostomata) in the Aleutian Islands, Alaska: taxonomy and distribution of six new species.

The cheilostome bryozoan genus Monoporella is poorly resolved taxonomically; only four Recent species have been formally described, though several undescribed species have been reported in the literature. The literature indicates no more than five species in the genus occurring in any local region of the world, with one to three species in most regions where the genus has been reported. I examined bryozoans from 52 trawl catches in the western and western-central Aleutian Islands, Alaska, and found specimens of Monoporella in 12 of these samples. Study of these specimens by scanning electron microscopy (SEM) revealed six new species that are described herein: M. flexibila, M. elongata, M. gigantea, M. ellefsoni, M. seastormi, and M. aleutica. Two of the species have erect colony morphologies, a condition not previously reported in Monoporella. The species diversity of Monoporella appears to be greater in the Aleutians than in any other part of the world adequately surveyed. I discuss whether this apparent high diversity is an artifact due to insufficient sampling in the deep shelf zone, and present two hypotheses to explain this high diversity should it prove not to be an artifact: 1) the present high local diversity represents a relict of past high diversity occurring broadly around the North Pacific rim; and 2) a local radiation of Monoporella occurred in the Aleutian archipelago.

The internal-brooding apparatus in the bryozoan genus Cauloramphus (Cheilostomata: Calloporidae) and its inferred homology to ovicells.

We studied by SEM the external morphology of the ooecium in eight bryozoans of the genus Cauloramphus (Cheilostomata, Calloporidae): C. spinifer, C. variegatus, C. magnus, C. multiavicularia, C. tortilis, C. cryptoarmatus, C. niger, and C. multispinosus, and by sectioning and light microscopy the anatomy of the brooding apparatus of C. spinifer, C. cryptoarmatus, and C. niger. These species all have a brood sac, formed by invagination of the non-calcified distal body wall of the maternal zooid, located in the distal half of the maternal (egg-producing) autozooid, and a vestigial, maternally budded kenozooidal ooecium. The brood sac comprises a main chamber and a long passage (neck) opening externally independently of the introvert. The non-calcified portion of the maternal distal wall between the neck and tip of the zooidal operculum is involved in closing and opening the brood sac, and contains both musculature and a reduced sclerite that suggest homology with the ooecial vesicle of a hyperstomial ovicell. We interpret the brooding apparatus in Cauloramphus as a highly modified form of cheilostome hyperstomial ovicell, as both types share 1) a brood chamber bounded by 2) the ooecium and 3) a component of the distal wall of the maternal zooid. We discuss Cauloramphus as a hypothetical penultimate stage in ovicell reduction in calloporid bryozoans. We suggest that the internal-brooding genus Gontarella, of uncertain taxonomic affinities, is actually a calloporid and represents the ultimate stage in which no trace of the ooecium remains. Internal brooding apparently evolved several times independently within the Calloporidae.

Round-trip catadromous migration in a Japanese amphipod, Sternomoera rhyaca (Gammaridea: Eusiridae).

We conducted a field study of the life cycle of the eusirid gammaridean amphipod Sternomoera rhyaca Kuribayashi, Mawatari, and Ishimaru, 1996 in a stream at Gokibiru, Hokkaido, Japan over the course of two non-consecutive years. This species is biennial; it spends most of its life in freshwater, but undertakes a short catadromous migration to the sea for reproduction. Reproduction occurs from March-June. Mature adults drift downstream to the sea singly and in precopulating pairs. Copulation and oviposition in the marsupium occur in mixed water at the stream mouth. Males die after copulation; ovigerous females return upstream by walking or swimming, where their eggs develop and hatch, after which the females also die. Juveniles remain in the stream, growing until they reach sexual maturity. Laboratory experiments showed that survivorship of all stages was lowest in seawater and highest in freshwater, though juveniles survived equally well in mixed water (50% seawater) and freshwater. Eggs developed to hatching only in freshwater; hatchlings in seawater and mixed water died within one and 21 days, respectively. Thus, S. rhyaca is well adapted to freshwater. Indeed, the only stages that required elevated salinity were copulation and subsequent oviposition, and we speculate that freshwater inhibits the female pre-reproductive molt. Because the life cycle of S. rhyaca has the most ontogenetically and temporally restricted saltwater phase known in any catadromous animal, its origin and maintenance are of evolutionary interest. We discuss two alternative hypotheses for the origin of the migratory life cycle, and discuss its maintenance in terms of fitness costs and benefits.

Molecular phylogeny and phylogeography of free-living Bryozoa (Cupuladriidae) from both sides of the Isthmus of Panama.

Genetic data were used to identify Recent species of free-living bryozoans (Cupuladriidae) from both sides of the Isthmus of Panama, and to examine their phylogenetic relationships, species richness, and population structures. An approximately 480bp fragment of the 16S mitochondrial rRNA gene was sequenced from 182 individuals from Panama, the Gulf of Mexico, and El Salvador. Ten haplotype groups (Cupuladria 4, 5, and 6; Discoporella 1, 2, 3A, 3B, 3C, 7, and 8) were identified. Genetic distances between haplotype groups (3.2-26.5%; K2P+Gamma) were 1-2 orders of magnitude greater than within groups (0.1-1.4%). Seven of the haplotype groups represent morphologically distinct species; Discoporellas 3A-C appear to be cryptic species. Phylogenetic analyses identified two pairs of transisthmian sister clades. An average divergence rate derived from other taxa suggests that Cupuladrias 4 and 5 diverged approximately 7Ma, a Discoporella 7 clade diverged from a 3A-C clade approximately 11Ma, and the 3A-C clade radiated approximately 6-4Ma; these events all predated final closure of the isthmus? 3Ma. The Caribbean side of the isthmus, with 5 species, is only marginally richer in cupuladriids than the Pacific side, with 4, but has greater phylogenetic depth. The Caribbean retains lineages stemming from a New World Miocene radiation that are not represented in the eastern Pacific; extant eastern Pacific cupuladriids share most recent common ancestry with only two of the Caribbean lineages. Species in the eastern Pacific tend to show shallow population structures, with high levels of gene flow between geographically separate populations, whereas Caribbean species tend to show deeper populations structures, with indications of restricted gene flow between Bocas del Toro/Gulf of Mosquitos and Costa Arriba/San Blas. The population structures derive from Pleistocene histories and may be of limited value in interpreting the macroevolutionary pattern, as our results provide no evidence of speciation on either side of the isthmus following closure in the late Pliocene.