The visual pigment xenopsin is widespread in protostome eyes and impacts the view on eye evolution.

Photoreceptor cells in the eyes of Bilateria are often classified into microvillar cells with rhabdomeric opsin and ciliary cells with ciliary opsin, each type having specialized molecular components and physiology. First data on the recently discovered xenopsin point towards a more complex situation in protostomes. In this study, we provide clear evidence that xenopsin enters cilia in the eye of the larval bryozoan Tricellaria inopinata and triggers phototaxis. As reported from a mollusc, we find xenopsin coexpressed with rhabdomeric-opsin in eye photoreceptor cells bearing both microvilli and cilia in larva of the annelid Malacoceros fuliginosus. This is the first organism known to have both xenopsin and ciliary opsin, showing that these opsins are not necessarily mutually exclusive. Compiling existing data, we propose that xenopsin may play an important role in many protostome eyes and provides new insights into the function, evolution, and possible plasticity of animal eye photoreceptor cells.

Transcriptome Analysis Elucidates the Key Responses of Bryozoan Fredericella sultana during the Development of Tetracapsuloides bryosalmonae (Myxozoa).

Bryozoans are sessile, filter-feeding, and colony-building invertebrate organisms. Fredericella sultana is a well known primary host of the myxozoan parasite Tetracapsuloides bryosalmonae. There have been no attempts to identify the cellular responses induced in F. sultana during the T. bryosalmonae development. We therefore performed transcriptome analysis with the aim of identifying candidate genes and biological pathways of F. sultana involved in the response to T. bryosalmonae. A total of 1166 differentially up- and downregulated genes were identified in the infected F. sultana. Gene ontology of biological processes of upregulated genes pointed to the involvement of the innate immune response, establishment of protein localization, and ribosome biogenesis, while the downregulated genes were involved in mitotic spindle assembly, viral entry into the host cell, and response to nitric oxide. Eukaryotic Initiation Factor 2 signaling was identified as a top canonical pathway and MYCN as a top upstream regulator in the differentially expressed genes. Our study provides the first transcriptional profiling data on the F. sultana zooid's response to T. bryosalmonae. Pathways and upstream regulators help us to understand the complex interplay in the infected F. sultana. The results will facilitate the elucidation of innate immune mechanisms of bryozoan and will lay a foundation for further analyses on bryozoan-responsive candidate genes, which will be an important resource for the comparative analysis of gene expression in bryozoans.

The Phylum Bryozoa: From Biology to Biomedical Potential.

Less than one percent of marine natural products characterized since 1963 have been obtained from the phylum Bryozoa which, therefore, still represents a huge reservoir for the discovery of bioactive metabolites with its ~6000 described species. The current review is designed to highlight how bryozoans use sophisticated chemical defenses against their numerous predators and competitors, and which can be harbored for medicinal uses. This review collates all currently available chemoecological data about bryozoans and lists potential applications/benefits for human health. The core of the current review relates to the potential of bryozoan metabolites in human diseases with particular attention to viral, brain, and parasitic diseases. It additionally weighs the pros and cons of total syntheses of some bryozoan metabolites versus the synthesis of non-natural analogues, and explores the hopes put into the development of biotechnological approaches to provide sustainable amounts of bryozoan metabolites without harming the natural environment.

Young species of cupuladriid bryozoans occupied new Caribbean habitats faster than old species.

The breadth of habitat occupied by a species, and the rate at which a species can expand into new habitats has important ecological and evolutionary consequences. Here we explore when extant species of free-living cupuladriid bryozoans expanded into new benthic Caribbean habitats that emerged during the final stages of formation of the Isthmus of Panama. Habitat breadth was estimated using the abundances of over 90,000 colonies in ten cupuladriid species, along with the ecological and sedimentary characteristics of the samples in which they occurred. Data reveal that all species expanded their habitat breadths during the last 6 Myr, but did so at a different tempo. 'Young' species - those that originated after 5 Ma - expanded relatively quickly, whereas 'old' species - those that originated before 9 Ma - took a further 2 Myr to achieve a comparable level of expansion. We propose that, like invasive species, young species are less restrained when expanding their habitat breadths compared to older well-established species. Understanding the mechanism causing this restraint requires further research.

Chemoenzymatic Dissection of Polyketide ß-Branching in the Bryostatin Pathway.

ß-Branching is an expansion upon canonical polyketide synthase extension that allows for the installation of diverse chemical moieties in several natural products. Several of these moieties are unique among natural products, including the two vinyl methylesters found in the core structure of bryostatins. This family of molecules is derived from an obligate bacterial symbiont of a sessile marine bryozoan, Bugula neritina. Within this family, bryostatin 1 has been investigated as an anticancer, neuroprotective, and immunomodulatory compound. We have turned to the biosynthetic gene cluster within the bacterial symbiont to investigate the biosynthesis of bryostatins. Recent sequencing efforts resulted in the annotation of two missing genes: bryT and bryU. Using novel chemoenzymatic techniques, we have validated these as the missing enoyl-CoA hydratase and donor acyl carrier protein, essential components of the ß-branching cassette of the bryostatin pathway. Together, this cassette installs the vinyl methylester moieties essential to the activity of bryostatins.

Review of bioactive secondary metabolites from marine bryozoans in the progress of new drugs discovery.

Marine bryozoans play an important role for the discovery of novel bioactive compounds among marine organisms. In this review, we summarize 164 new secondary metabolites including macrocyclic lactones, sterols, alkaloids, sphingolipids and so forth from 24 marine bryozoans in the last two decades. The structural features, bioactivity, structure-activity relationship, mechanism and strategies to address the resupply of these scarce secondary metabolites are discussed. The structural and bioactive diversity of the secondary metabolites from marine bryozoans indicated the possibility of using these compounds, especially bryostatin 1 (1), bryostatin analog (BA1), alkaloids (50, 53, 127-128 and 134-139), sphingolipids sulfates (148 and 149) and sulfur-containing aromatic compound (160), as the starting points for new drug discovery.

Exploring the regulatory role of nitric oxide (NO) and the NO-p38MAPK/cGMP pathway in larval settlement of the bryozoan Bugula neritina.

The bryozoan Bugula neritina is a cosmopolitan marine fouling species that causes major fouling problems in sub-tropical waters. Settlement of B. neritina larvae can be triggered without an obvious external cue. Here, the negative regulatory role of nitric oxide (NO) during larval settlement of B. neritina was demonstrated to be mediated by cyclic guanosine monophosphate (cGMP). Although the regulatory role of the NO-p38 MAPK signaling axis in larval settlement was not evident, inhibition of nitric oxide synthase (NOS) led to the deactivation of p38 MAPK. Exclusive localization of NO and NO signaling components in sensory-related organs of the larvae is consistent with its signal transduction function in metamorphosis. Overall, this study provides new insights into the regulatory roles of the NO-p38MAPK/cGMP pathway in B. neritina settlement.

Parallel lives of symbionts and hosts: chemical mutualism in marine animals.

Covering: up to 2018 Symbiotic microbes interact with animals, often by producing natural products (specialized metabolites; secondary metabolites) that exert a biological role. A major goal is to determine which microbes produce biologically important compounds, a deceptively challenging task that often rests on correlative results, rather than hypothesis testing. Here, we examine the challenges and successes from the perspective of marine animal-bacterial mutualisms. These animals have historically provided a useful model because of their technical accessibility. By comparing biological systems, we suggest a common framework for establishing chemical interactions between animals and microbes.

New Cytotoxic Secondary Metabolites from Marine Bryozoan Cryptosula pallasiana.

A new sterol, (23R)-methoxycholest-5,24-dien-3ß-ol (1), two new ceramides, (2S,3R,4E,8E)-2-(tetradecanoylamino)-4,8-octadecadien-l,3-diol (6) and (2S,3R,2'R,4E,8E)-2-(tetradecanoylamino)-4,8-octadecadien-l,3,2'-triol (7), together with three known sterols (2-4), a lactone (5) and two ceramides (8,9), were isolated from the marine bryozoan Cryptosula pallasiana, collected at Huang Island of China. The structures of the new compounds were elucidated by extensive spectroscopic analyses, chemical methods and quantum electronic circular dichroism (ECD) calculations. Among the isolated compounds, sterol 1 possessed a rare side chain with a methoxy group at C-23, and a double bond between C-24 and C-25. Ceramides 6 and 7 possessed 14 carbons in their long-chain fatty acid base (FAB), which were different from the normal ceramides with 16 carbons in the FAB. Moreover, compounds 5 and 8 were isolated for the first time from marine bryozoans. Compounds 1-9 were evaluated for their cytotoxicity against human tumor cell lines HL-60, Hep-G2 and SGC-7901. The results showed that lactone 5 appears to have strong cytotoxicity against the test tumor cell lines, with IC50 values from 4.12 µM to 7.32 µM, and sterol 1 displayed moderate cytotoxicity with IC50 values between 12.34 µM and 18.37 µM, while ceramides 6-9 showed weak cytotoxicity with IC50 ranging from 21.13 µM to 58.15 µM.

Estimating monotonic rates from biological data using local linear regression.

Accessing many fundamental questions in biology begins with empirical estimation of simple monotonic rates of underlying biological processes. Across a variety of disciplines, ranging from physiology to biogeochemistry, these rates are routinely estimated from non-linear and noisy time series data using linear regression and ad hoc manual truncation of non-linearities. Here, we introduce the R package LoLinR, a flexible toolkit to implement local linear regression techniques to objectively and reproducibly estimate monotonic biological rates from non-linear time series data, and demonstrate possible applications using metabolic rate data. LoLinR provides methods to easily and reliably estimate monotonic rates from time series data in a way that is statistically robust, facilitates reproducible research and is applicable to a wide variety of research disciplines in the biological sciences.

Novel brominated metabolites from Bryozoa: a functional analysis.

Marine invertebrates are a promising source of novel natural products with biological activities. The phylum Bryozoa is relatively under-investigated in this context, although a number of compounds with medical potential has been discovered in recent years. Here, we report on the novel group of brominated metabolites from the bryozoan Terminoflustra membranaceatruncata, including analysis of biological activities of the tribrominated terminoflustrindole A (Cm-1) and the structures of the related dibrominated variants terminoflustrindoles B and C. Terminoflustrindole A was previously shown to have fungicidal properties. Although they vary by just one bromine group in each case from terminoflustrindole A, in this study, we report that terminoflustrindoles B and C exhibit no antimicrobial activity in the same assays. In addition to displaying antifungal activity, Terminoflustrindole A was also found to exhibit potent cytotoxic activity when tested against tumour cell lines. The gradient distribution of this compound within the bryozoan colony was demonstrated using LC-MS-analysis.

Inferring metabolic pathway activity levels from RNA-Seq data.

BACKGROUND: Assessing pathway activity levels is a plausible way to quantify metabolic differences between various conditions. This is usually inferred from microarray expression data. Wide availability of NGS technology has triggered a demand for bioinformatics tools capable of analyzing pathway activity directly from RNA-Seq data. In this paper we introduce XPathway, a set of tools that compares pathway activity analyzing mapping of contigs assembled from RNA-Seq reads to KEGG pathways. The XPathway analysis of pathway activity is based on expectation maximization and topological properties of pathway graphs. RESULTS: XPathway tools have been applied to RNA-Seq data from the marine bryozoan Bugula neritina with and without its symbiotic bacterium "Candidatus Endobugula sertula". We successfully identified several metabolic pathways with differential activity levels. The expression of enzymes from the identified pathways has been further validated through quantitative PCR (qPCR). CONCLUSIONS: Our results show that XPathway is able to detect and quantify the metabolic difference in two samples. The software is implemented in C, Python and shell scripting and is capable of running on Linux/Unix platforms. The source code and installation instructions are available at http://alan.cs.gsu.edu/NGS/?q=content/xpathway .

Recyclable plastics as substrata for settlement and growth of bryozoans Bugula neritina and barnacles Amphibalanus amphitrite.

Plastics are common and pervasive anthropogenic debris in marine environments. Floating plastics provide opportunities to alter the abundance, distribution and invasion potential of sessile organisms that colonize them. We selected plastics from seven recycle categories and quantified settlement of (i) bryozoans Bugula neritina (Linnaeus, 1758) in the lab and in the field, and of (ii) barnacles Amphibalanus (= Balanus) amphitrite (Darwin, 1854) in the field. In the laboratory we cultured barnacles on the plastics for 8 weeks and quantified growth, mortality, and breaking strength of the side plates. In the field all recyclable plastics were settlement substrata for bryozoans and barnacles. Settlement depended on the type of plastic. Fewer barnacles settled on plastic surfaces compared to glass. In the lab and in the field, bryozoan settlement was higher on plastics than on glass. In static laboratory rearing, barnacles growing on plastics were initially significantly smaller than on glass. This suggested juvenile barnacles were adversely impacted by materials leaching from the plastics. Barnacle mortality was not significantly different between plastic and glass surfaces, but breaking strength of side plates of barnacles on polyvinyl chloride (PVC) and polycarbonate (PC) were significantly lower than breakage strength on glass. Plastics impact marine ecosystems directly by providing new surfaces for colonization with fouling organisms and by contaminants shown previously to leach out of plastics and impact biological processes.

[FUNCTIONAL DIFFERENTIATION IN BRYOZOAN COLONY: A PROTEOMIC ANALYSIS].

Bryozoans are typical modular organisms. They consist of repetitive structural units, the zooids. Bryozoan colonies grow by zooidal budding, with the distribution pattern of the budding loci underlying the diversity of colony forms. Budding is usually restricted to the zooids at the periphery of the colony, which form a "growing edge" or local terminal growth zones. Non-budding parts of the colony can be functionally subdivided, too. In many species colonies consists of regular, often repetitive zones of feeding and non-feeding modules, associated with a periodical degeneration and regeneration of the polypide, retractile tentacle crown with a gut and the accompanying musculature. So, there is functional differentiation in bryozoan colonies but its mechanisms are unknown. Presumably, budding and/or polypide recycling in different colony parts are induced or inhibited by certain determinants of functional specialization. An effective tool of their identification is the comparison of proteomes of functionally different zones. Here we report the results of proteomic analysis of three bryozoan species from the White Sea, which have a different colony form: Flustrellidra hispida, Terminoflustra membranaceotruncata and Securiflustra securifrons. Using differential two-dimensional electrophoresis (2D-DIGE), we compared proteomes of the growing edge and the zones consisting of feeding and non-feeding zooids in these species. We estimated the overall proteome variability, revealed proteins whose relative abundance gradually changed along the proximal-distal colony axis and suggested that they might be involved in the functional differentiation of the colony.

Biofilm history and oxygen availability interact to affect habitat selection in a marine invertebrate.

In marine systems, oxygen availability varies at small temporal and spatial scales, such that current oxygen levels may not reflect conditions of the past. Different studies have shown that marine invertebrate larvae can select settlement sites based on local oxygen levels and oxygenation history of the biofilm, but no study has examined the interaction of both. The influence of normoxic and hypoxic water and oxygenation history of biofilms on pre-settlement behavior and settlement of the bryozoan Bugula neritina was tested. Larvae used cues in a hierarchical way: the oxygen levels in the water prime larvae to respond, the response to different biofilms is contingent on oxygen levels in the water. When oxygen levels varied throughout biofilm formation, larvae responded differently depending on the history of the biofilm. It appears that B. neritina larvae integrate cues about current and historical oxygen levels to select the appropriate microhabitat and maximize their fitness.

Distribution of serotonin and FMRF-amide in the nervous system of different zooidal types of cheilostome bryozoa: A case study of Arctonula arctica.

In the White Sea bryozoans Arctonula arctica, the structure of the nervous system and distribution of 5-hydroxytryptamine (5-HT) and FMRF-amide were studied for the first time using immunohistochemical methods and confocal scanning microscopy. The neurotransmitters studied have been actively involved into the integrative processes, gut functioning, and regulation of motion activity. In avicularia, 5-HT and FMRF-amide receptors are capable of performing the same functions, except for participation in the gut functioning, because they have no digestive system.

First chitin extraction from Plumatella repens (Bryozoa) with comparison to chitins of insect and fungal origin.

Chitin immediately suggests the representatives of the kingdom Fungi, as well as such phyla as Annelida, Mollusca, Porifera, Cnidaria and, mostly, Arthropoda. Although Bryozoa also represents a chitin-containing phylum, no study has been developed yet on the isolation or characterization of the chitin from it. In this study, physiochemical properties of the chitin isolated from Plumatella repens belonging to the phylum Bryozoa was determined for the first time. The chitin structure was also studied comparatively by isolating chitin from an insect species (Palomena prasina) of the phylum Arthropoda, and Fomes fomentarius belonging to the kingdom Fungi. It was observed that the bryozoan chitin was in the α form, as in the arthropod and fungal chitins. The chitin contents in the dry weight of the bryozoan, fungal and insect species were observed to be 13.3%, 2.4%, and 10.8%, respectively. The insect chitin exhibited the highest thermal stability followed by that of the bryozoan and then the fungal chitins. Surface morphologies reveal that the insect and bryozoan chitins were composed of nano fibre and pore structures, whereas the fungal chitin had no pores or fibres. The crystallinity of the insect chitin (CrI=84.9%) was higher than the bryozoan (CrI=60.1%) and fungal chitins (CrI=58.5%).

Antimicrobial activity of Antarctic bryozoans: an ecological perspective with potential for clinical applications.

The antimicrobial activity of Antarctic bryozoans and the ecological functions of the chemical compounds involved remain largely unknown. To determine the significant ecological and applied antimicrobial effects, 16 ether and 16 butanol extracts obtained from 13 different bryozoan species were tested against six Antarctic (including Psychrobacter luti, Shewanella livingstonensis and 4 new isolated strains) and two bacterial strains from culture collections (Escherichia coli and Bacillus cereus). Results from the bioassays reveal that all ether extracts exhibited antimicrobial activity against some bacteria. Only one butanol extract produced inhibition, indicating that antimicrobial compounds are mainly lipophilic. Ether extracts of the genus Camptoplites inhibited the majority of bacterial strains, thus indicating a broad-spectrum of antimicrobial activity. Moreover, most ether extracts presented activities against bacterial strains from culture collections, suggesting the potential use of these extracts as antimicrobial drugs against pathogenic bacteria.

Host differentially expressed genes during association with its defensive endosymbiont.

Mutualism, a beneficial relationship between two species, often requires intimate interaction between the host and symbiont to establish and maintain the partnership. The colonial marine bryozoan Bugula neritina harbors an as yet uncultured endosymbiont, "Candidatus Endobugula sertula," throughout its life stages. The bacterial symbiont is the putative source of bioactive complex polyketide metabolites, the bryostatins, which chemically defend B. neritina larvae from predation. Despite the presence of "Ca. Endobugula sertula" in all life stages of the host, deterrent bryostatins appear to be concentrated in reproductive portions of the host colony, suggesting an interaction between the two partners to coordinate production and distribution of the metabolites within the colony. In this study, we identified host genes that were differentially expressed in control colonies and in colonies cured of the symbiont. Genes that code for products similar to glycosyl hydrolase family 9 and family 20 proteins, actin, and a Rho-GDP dissociation inhibitor were significantly downregulated (more than twice) in antibiotic-cured non-reproductive zooids compared to control symbiotic ones. Differential expression of these genes leads us to hypothesize that the host B. neritina may regulate the distribution of the symbiont within the colony via mechanisms of biofilm degradation and actin rearrangement, and consequently, influences bryostatin localization to bestow symbiont-associated protection to larvae developing in the reproductive zooids.

Neritinaceramides A-E, new ceramides from the marine bryozoan Bugula neritina inhabiting South China Sea and their cytotoxicity.

Five new ceramides, neritinaceramides A (1), B (2), C (3), D (4) and E (5), together with six known ceramides (6-11), two known alkyl glycerylethers (12 and 13) and a known nucleoside (14), were isolated from marine bryozoan Bugula neritina, which inhabits the South China Sea. The structures of the new compounds were elucidated as (2S,3R,3'S,4E,8E,10E)-2-(hexadecanoylamino)-4,8,10-octadecatriene-l,3,3'-triol (1), (2S,3R,2'R,4E,8E,10E)-2-(hexadecanoylamino)-4,8,10-octadecatriene-l,3,2'-triol (2), (2S,3R,2'R,4E,8E,10E)-2-(octadecanoylamino)-4,8,10-octadecatriene-l,3,2'-triol (3), (2S,3R,3'S,4E,8E)-2-(hexadecanoylamino)-4,8-octadecadiene-l,3,3'-triol (4) and (2S,3R,3'S,4E)-2-(hexadecanoylamino)-4-octadecene-l,3,3'-triol (5) on the basis of extensive spectral analysis and chemical evidences. The characteristic C-3'S hydroxyl group in the fatty acid moiety in compounds 1, 4 and 5, was a novel structural feature of ceramides. The rare 4E,8E,10E-triene structure in the sphingoid base of compounds 1-3, was found from marine bryozoans for the first time. The new ceramides 1-5 were evaluated for their cytotoxicity against HepG2, NCI-H460 and SGC7901 tumor cell lines, and all of them exhibited selective cytotoxicity against HepG2 and SGC7901 cells with a range of IC50 values from 47.3 µM to 58.1 µM. These chemical and cytotoxic studies on the new neritinaceramides A-E (1-5) added to the chemical diversity of B. neritina and expanded our knowledge of the chemical modifications and biological activity of ceramides.