Evidence for transporter-mediated uptake of environmental L-glutamate in a freshwater sponge, Ephydatia muelleri.

The freshwater sponge, Ephydatia muelleri, lacks a nervous or endocrine system and yet it exhibits a coordinated whole-body action known as a "sneeze" that can be triggered by exposure to L-glutamate. It is not known how L-glutamate is obtained by E. muelleri in sufficient quantities (i.e., 70 µM) to mediate this response endogenously. The present study tested the hypothesis that L-glutamate can be directly acquired from the environment across the body surface of E. muelleri. We demonstrate carrier mediated uptake of two distinct saturable systems with maximal transport rates (Jmax) of 64.27 ± 4.98 and 25.12 ± 1.87 pmols mg-1 min-1, respectively. The latter system has a higher calculated substrate affinity (Km) of 2.87 ± 0.38 µM compared to the former (8.75 ± 1.00 µM), indicative of distinct systems that can acquire L-glutamate at variable environmental concentrations. Further characterization revealed potential shared pathways of L-glutamate uptake with other negatively charged amino acids, namely D-glutamate and L-aspartate, as well as the neutral amino acid L-alanine. We demonstrate that L-glutamate uptake does not appear to rely on exogenous sodium or proton concentrations as removal of these ions from the bathing media did not significantly alter uptake. Likewise, L-glutamate uptake does not seem to rely on internal proton motive forces driven by VHA as application of 100 nM of the VHA inhibitor bafilomycin did not alter uptake rates within E. muelleri tissues. Whether the acquired amino acid is used to supplement feeding or is stored and accumulated to mediate the sneeze response remains to be determined.

The genome of the reef-building glass sponge Aphrocallistes vastus provides insights into silica biomineralization.

Well-annotated and contiguous genomes are an indispensable resource for understanding the evolution, development, and metabolic capacities of organisms. Sponges, an ecologically important non-bilaterian group of primarily filter-feeding sessile aquatic organisms, are underrepresented with respect to available genomic resources. Here we provide a high-quality and well-annotated genome of Aphrocallistes vastus, a glass sponge (Porifera: Hexactinellida) that forms large reef structures off the coast of British Columbia (Canada). We show that its genome is approximately 80 Mb, small compared to most other metazoans, and contains nearly 2500 nested genes, more than other genomes. Hexactinellida is characterized by a unique skeletal architecture made of amorphous silicon dioxide (SiO2), and we identified 419 differentially expressed genes between the osculum, i.e. the vertical growth zone of the sponge, and the main body. Among the upregulated ones, mineralization-related genes such as glassin, as well as collagens and actins, dominate the expression profile during growth. Silicateins, suggested being involved in silica mineralization, especially in demosponges, were not found at all in the A. vastus genome and suggests that the underlying mechanisms of SiO2 deposition in the Silicea sensu stricto (Hexactinellida + Demospongiae) may not be homologous.

The compact genome of the sponge Oopsacas minuta (Hexactinellida) is lacking key metazoan core genes.

BACKGROUND: Explaining the emergence of the hallmarks of bilaterians is a central focus of evolutionary developmental biology-evodevo-and evolutionary genomics. For this purpose, we must both expand and also refine our knowledge of non-bilaterian genomes, especially by studying early branching animals, in particular those in the metazoan phylum Porifera. RESULTS: We present a comprehensive analysis of the first whole genome of a glass sponge, Oopsacas minuta, a member of the Hexactinellida. Studying this class of sponge is evolutionary relevant because it differs from the three other Porifera classes in terms of development, tissue organization, ecology, and physiology. Although O. minuta does not exhibit drastic body simplifications, its genome is among the smallest of animal genomes sequenced so far, and surprisingly lacks several metazoan core genes (including Wnt and several key transcription factors). Our study also provides the complete genome of a symbiotic Archaea dominating the associated microbial community: a new Thaumarchaeota species. CONCLUSIONS: The genome of the glass sponge O. minuta differs from all other available sponge genomes by its compactness and smaller number of encoded proteins. The unexpected loss of numerous genes previously considered ancestral and pivotal for metazoan morphogenetic processes most likely reflects the peculiar syncytial tissue organization in this group. Our work further documents the importance of convergence during animal evolution, with multiple convergent evolution of septate-like junctions, electrical-signaling and multiciliated cells in metazoans.

Molecular machineries of ciliogenesis, cell survival, and vasculogenesis are differentially expressed during regeneration in explants of the demosponge Halichondria panicea.

Sponges are interesting animal models for regeneration studies, since even from dissociated cells, they are able to regenerate completely. In particular, explants are model systems that can be applied to many sponge species, since small fragments of sponges can regenerate all elements of the adult, including the oscula and the ability to pump water. The morphological aspects of regeneration in sponges are relatively well known, but the molecular machinery is only now starting to be elucidated for some sponge species. Here, we have used an explant system of the demosponge Halichondria panicea to understand the molecular machinery deployed during regeneration of the aquiferous system. We sequenced the transcriptomes of four replicates of the 5-day explant without an osculum (NOE), four replicates of the 17-18-day explant with a single osculum and pumping activity (PE) and also four replicates of field-collected individuals with regular pumping activity (PA), and performed differential gene expression analysis. We also described the morphology of NOE and PE samples using light and electron microscopy. Our results showed a highly disorganised mesohyl and disarranged aquiferous system in NOE that is coupled with upregulated pathways of ciliogenesis, organisation of the ECM, and cell proliferation and survival. Once the osculum is formed, genes involved in "response to stimulus in other organisms" were upregulated. Interestingly, the main molecular machinery of vasculogenesis described in vertebrates was activated during the regeneration of the aquiferous system. Notably, vasculogenesis markers were upregulated when the tissue was disorganised and about to start forming canals (NOE) and angiogenic stimulators and ECM remodelling machineries were differentially expressed once the aquiferous system was in place (PE and PA). Our results are fundamental to better understanding the molecular mechanisms involved in the formation of the aquiferous system in sponges, and its similarities with the early onset of blood-vessel formation in animal evolution.

Sponges sneeze mucus to shed particulate waste from their seawater inlet pores.

Sponges, among the oldest extant multicellular organisms on Earth,1 play a key role in the cycling of nutrients in many aquatic ecosystems.2-5 They need to employ strategies to prevent clogging of their internal filter system by solid wastes,6-8 but self-cleaning mechanisms are largely unknown. It is commonly assumed that sponges remove solid waste with the outflowing water through distinct outflow openings (oscula).3,9 Here, we present time-lapse video footage and analyses of sponge waste revealing a completely different mechanism of particle removal in the Caribbean tube sponge Aplysina archeri. This sponge actively moves particle-trapping mucus against the direction of its internal water flow and ejects it into the surrounding water from its seawater inlet pores (ostia) through periodic surface contractions that have been described earlier as "sneezing."10,11 Visually, it appears as if the sponge is continuously streaming mucus-embedded particles and sneezes to shed this particulate waste, resulting in a notable flux of detritus that is actively consumed by sponge-associated fauna. The new data are used to estimate production of detritus for this abundant sponge on Caribbean coral reefs. Last, we discuss why waste removal from the sponge inhalant pores may be a common feature among sponges and compare the process in sponges to equivalent mechanisms of mucus transport in other animals, including humans.

Microscopy Studies of Placozoans.

Trichoplax adhaerens is an enigmatic animal with an extraordinarily simple morphology and a cellular organization, which are the focus of current research. Protocols outlined here provide detailed descriptions of advanced techniques for light and electron microscopic studies of Trichoplax. Studies using these techniques have enhanced our understanding of cell type diversity and function in placozoans and have provided insight into the evolution, development, and physiology of this little understood group.

Hydrodynamics of sponge pumps and evolution of the sponge body plan.

Sponges are suspension feeders that filter vast amounts of water. Pumping is carried out by flagellated chambers that are connected to an inhalant and exhalant canal system. In 'leucon' sponges with relatively high-pressure resistance due to a complex and narrow canal system, pumping and filtering are only possible owing to the presence of a gasket-like structure (forming a canopy above the collar filters). Here, we combine numerical and experimental work and demonstrate how sponges that lack such sealing elements are able to efficiently pump and force the flagella-driven flow through their collar filter, thanks to the formation of a 'hydrodynamic gasket' above the collar. Our findings link the architecture of flagellated chambers to that of the canal system, and lend support to the current view that the sponge aquiferous system evolved from an open-type filtration system, and that the first metazoans were filter feeders.

Tracing animal genomic evolution with the chromosomal-level assembly of the freshwater sponge Ephydatia muelleri.

The genomes of non-bilaterian metazoans are key to understanding the molecular basis of early animal evolution. However, a full comprehension of how animal-specific traits, such as nervous systems, arose is hindered by the scarcity and fragmented nature of genomes from key taxa, such as Porifera. Ephydatia muelleri is a freshwater sponge found across the northern hemisphere. Here, we present its 326 Mb genome, assembled to high contiguity (N50: 9.88 Mb) with 23 chromosomes on 24 scaffolds. Our analyses reveal a metazoan-typical genome architecture, with highly shared synteny across Metazoa, and suggest that adaptation to the extreme temperatures and conditions found in freshwater often involves gene duplication. The pancontinental distribution and ready laboratory culture of E. muelleri make this a highly practical model system which, with RNAseq, DNA methylation and bacterial amplicon data spanning its development and range, allows exploration of genomic changes both within sponges and in early animal evolution.

Geographic range extensions of stalked, flabelliform sponges (Porifera) from eastern Canada with a new combination of a species of Plicatellopsis in the North Atlantic.

Three new species records, Axinella arctica (Vosmaer, 1885), Semisuberites cribrosa (Miklucho-Maclay, 1870), and Cladocroce spatula (Lundbeck, 1902), and one new combination, Plicatellopsis bowerbanki (Vosmaer, 1885) comb. nov. from eastern Canada are described. The four species have similar growth forms which are either fan or cup-shaped with obvious stalks. This is the first description of a member of the genus Plicatellopsis in the North Atlantic, and the second record of the genus in the northern hemisphere. The four species described here have a history of misidentification in eastern Canada and this work aims to guide future identifications in the region.

Sponge (Porifera) fauna of Frobisher Bay, Baffin Island, Canada with the description of an Iophon rich sponge garden.

The deep-water sponge fauna of the Canadian Arctic remains to be fully described, particularly in areas that are not sampled by fisheries stock-assessment trawl surveys such as the major bays and fjords of the northern Labrador Sea and Baffin Bay. Frobisher Bay is a large inlet located on the southeast of Baffin Island. We used a remotely operated vehicle, as well as box cores and Agassiz trawls to study the sponge fauna of this bay. Over three years, from 2015 to 2017, sponge specimens were collected representing 24 distinct sponge taxa. Dense gardens of Iophon koltuni Morozov, Sabirov, Zimina, 2019 were discovered at a site near Hill Island in inner Frobisher Bay. The species has a unique finger-like growth form and provides complex habitat in the inner bay. Other sponge species are new to the Northern Labrador marine ecoregion. In particular, we report geographic range extensions of Tetilla sibirica (Fristedt, 1887) and Craniella polyura (Schmidt, 1870), and provide spicule measurements and descriptions of Iophon piceum (Vosmaer, 1882) and Mycale lingua (Bowerbank, 1866). These species identifications, geographic range extensions, and an expanded description of a species synonym represent the first inventory of the sponge fauna of Frobisher Bay.

Animal Evolution: The Hidden Biology of the Ctenophore Cilium.

A new study uncovers the function of a completely novel protein in the eye-catchingly iridescent cilia of the ctenophore comb plate.

A problematic animal fossil from the early Cambrian Hetang Formation, South China.

The lower-middle Hetang Formation (Cambrian Stage 2-3) deposited in slope-basinal facies in South China is well-known for its preservation of the earliest articulated sponge fossils, providing an important taphonomic window into the Cambrian explosion. However, the Hetang Formation also hosts a number of problematic animal fossils that have not been systematically described. This omission results in an incomplete picture of the Hetang biota and limits its contribution to the understanding of the early evolution of animals. Here we describe a new animal taxon, Cambrowania ovata Tang and Xiao, new genus new species, from the middle Hetang Formation in the Lantian area of southern Anhui Province, South China. Specimens are preserved as carbonaceous compressions, although some are secondarily mineralized. A comprehensive analysis using reflected light microscopy, scanning electron microscopy, energy dispersive X-ray spectroscopy, and micro-CT reveals that the new species is characterized by a spheroidal to fusoidal truss-like structure consisting of rafter-like crossbars, some of which are secondarily baritized and may have been internally hollow. Some specimens have aperture-like structures that are broadly similar to oscula of sponges, whereas others show evidence of a medial split reminiscent of gaping carapaces. While the phylogenetic affinity of Cambrowania ovata Tang and Xiao, new genus new species remains problematic, we propose that it may represent carapaces of bivalved arthropods or more likely sponges in early life stages. Along with other problematic metazoan fossils such as hyolithids and sphenothallids, Cambrowania ovata Tang and Xiao, new genus new species adds to the diversity of the sponge-dominated Hetang biota in an early Cambrian deep-water slope-basinal environment.

Sponge Behavior and the Chemical Basis of Responses: A Post-Genomic View.

Sponges perceive and respond to a range of stimuli. How they do this is still difficult to pin down despite now having transcriptomes and genomes of an array of species. Here we evaluate the current understanding of sponge behavior and present new observations on sponge activity in situ. We also explore biosynthesis pathways available to sponges from data in genomes/transcriptomes of sponges and other non-bilaterians with a focus on exploring the role of chemical signaling pathways mediating sponge behavior and how such chemical signal pathways may have evolved. Sponge larvae respond to light but opsins are not used, nor is there a common photoreceptor molecule or mechanism used across sponge groups. Other cues are gravity and chemicals. In situ recordings of behavior show that both shallow and deep-water sponges move a lot over minutes and hours, and correlation of behavior with temperature, pressure, oxygen, and water movement suggests that at least one sponge responds to changes in atmospheric pressure. The sensors for these cues as far as we know are individual cells and, except in the case of electrical signaling in Hexactinellida, these most likely act as independent effectors, generating a whole-body reaction by the global reach of the stimulus to all parts of the animal. We found no evidence for use of conventional neurotransmitters such as serotonin and dopamine. Intriguingly, some chemicals synthesized by symbiont microbes could mean other more complex signaling occurs, but how that interplay might happen is not understood. Our review suggests chemical signaling pathways found in sponges do not reflect loss of a more complex set.

Understanding Animal Evolution: The Added Value of Sponge Transcriptomics and Genomics: The disconnect between gene content and body plan evolution.

Sponges are important but often-neglected organisms. The absence of classical animal traits (nerves, digestive tract, and muscles) makes sponges challenging for non-specialists to work with and has delayed getting high quality genomic data compared to other invertebrates. Yet analyses of sponge genomes and transcriptomes currently available have radically changed our understanding of animal evolution. Sponges are of prime evolutionary importance as one of the best candidates to form the sister group of all other animals, and genomic data are essential to understand the mechanisms that control animal evolution and diversity. Here we review the most significant outcomes of current genomic and transcriptomic analyses of sponges, and discuss limitations and future directions of sponge transcriptomic and genomic studies.

Oxygen and the Energetic Requirements of the First Multicellular Animals.

The appearance of multicellular animals during the Neoproterozoic Era is thought to have coincided with oxygenation of the oceans; however, we know little about the physiological needs of early animals or about the environment they lived in. Approaches using biomarkers, fossils, and phylogenomics have provided some hints of the types of animals that may have been present during the Neoproterozoic, but extant animals are our best modern links to the theoretical ancestors of animals. Neoproterozoic oceans were low energy habitats, with low oxygen concentrations and sparse food availability for the first animals. We examined tolerance of extant ctenophores and sponges-as representatives of extant lineages of the earliest known metazoan groups-to feeding and oxygen use. A review of respiration rates in species across several phyla suggests that suspension feeders in general have a wide range of metabolic rates, but sponges have some of the highest of invertebrates and ctenophores some of the lowest. Our own studies on the metabolism of two groups of deep water sponges show that sponges have different approaches to deal with the cost of filtration and low food availability. We also confirmed that deep water sponges tolerate periods of hypoxia, but at the cost of filtration, indicating that normal feeding is energetically expensive. Predictions of oxygen levels in the Neoproterozoic suggest the last common ancestor of multicellular animals was unlikely to have filtered like modern sponges. Getting enough food at low oxygen would have been a more important driver of the evolution of early body plans.

Suspended sediment causes feeding current arrests in situ in the glass sponge Aphrocallistes vastus.

Bottom-contact trawling generates large, moving clouds of suspended sediments that can alter the behaviour of organisms adjacent to trawl paths. While increased suspended sediment concentrations (SSCs) are known to cause glass sponges to arrest filtration in lab studies, the response of sponges to sediment in situ is not yet known. Here we describe arrest behaviours in response to increased SSCs recorded from the glass sponge Aphrocallistes vastus at the Fraser Ridge sponge reef in the Strait of Georgia, British Columbia, Canada. We identified 23 arrests of the sponges' feeding current during experimental disturbances that raised SSC to between 10 and 80 mg l-1. Single arrests lasted 4.25 ±â€¯1.3 min (±SD) and were characterized by a 2 cm s-1 reduction in feeding current lasting 0.5-3 min (mean 1.91 ±â€¯0.97 min, n = 19). In comparison, coughing arrests varied in length (31 ±â€¯22.89 min) with arrest phases lasting 4-15 min (10.46 ±â€¯5.26 min, n = 4). Coughing arrests showed a distinctive on/off pattern as sponge filtration returned to normal excurrent velocities, distinguishing them from single arrests. The onset of both arrest types was correlated with elevated SSCs (r = -0.83 to -0.92). Natural SSCs at the reef averaged 4.4 mg l-1 and were correlated with tidal flow (r = 0.86 to 0.89). The combined data provide evidence that suspended sediments can stop glass sponge feeding in situ even at SSCs below those known to be generated by trawling.

Wnt signaling and polarity in freshwater sponges.

BACKGROUND: The Wnt signaling pathway is uniquely metazoan and used in many processes during development, including the formation of polarity and body axes. In sponges, one of the earliest diverging animal groups, Wnt pathway genes have diverse expression patterns in different groups including along the anterior-posterior axis of two sponge larvae, and in the osculum and ostia of others. We studied the function of Wnt signaling and body polarity formation through expression, knockdown, and larval manipulation in several freshwater sponge species. RESULTS: Sponge Wnts fall into sponge-specific and sponge-class specific subfamilies of Wnt proteins. Notably Wnt genes were not found in transcriptomes of the glass sponge Aphrocallistes vastus. Wnt and its signaling genes were expressed in archaeocytes of the mesohyl throughout developing freshwater sponges. Osculum formation was enhanced by GSK3 knockdown, and Wnt antagonists inhibited both osculum development and regeneration. Using dye tracking we found that the posterior poles of freshwater sponge larvae give rise to tissue that will form the osculum following metamorphosis. CONCLUSIONS: Together the data indicate that while components of canonical Wnt signaling may be used in development and maintenance of osculum tissue, it is likely that Wnt signaling itself occurs between individual cells rather than whole tissues or structures in freshwater sponges.

Trophic ecology of glass sponge reefs in the Strait of Georgia, British Columbia.

Sponges link the microbial loop with benthic communities by feeding on bacteria. Glass sponge reefs on the continental shelf of western Canada have extremely high grazing rates, consuming seven times more particulate carbon than can be supplied by vertical flux alone. Unlike many sponges, the reef building species Aphrocallistes vastus has no microbial symbionts and removes little dissolved organic carbon. To determine how reef sponges therefore get enough food to sustain such substantial grazing we measured stable carbon and nitrogen isotope signatures of water, sediment and sponge tissues. To ensure samples were temporally associated, we also studied the duration particles were retained in tissues in controlled feeding studies using microscopic beads and 13C-labeled bacteria. Although fecal pellets were expelled from sponges within 24 hours of feeding, intact bacteria were still found in tissues and sponge tissues retained elevated 13C levels for at least 14 days. These independent lines of evidence suggest that carbon in reef sponge tissues may reflect food consumed from days to weeks earlier. Stable isotope analysis suggests that heterotrophic bacteria ingested by the sponges comes from a confluence of trophic subsidies: from terrestrial and oceanic sources, and also potentially on sediment-borne bacteria resuspended by tidal currents.