Ascidian Tunic Research Articles

Pax3/7 gene function in Oikopleura dioica supports a neuroepithelial-like origin for its house-making Fol territory

Larvacean tunicates feature a spectacular innovation not seen in other animals - the trunk oikoplastic epithelium (OE). This epithelium produces a house, a large and complex extracellular structure used for filtering and concentrating food particles. Previously we identified several homeobox transcription factor genes expressed during early OE patterning. Among these are two Pax3/7 copies that we named pax37A and pax37B. The vertebrate homologs, PAX3 and PAX7 are involved in developmental processes related to neural crest and muscles. In the ascidian tunicate Ciona intestinalis, Pax3/7 plays a role in the development of cells deriving from the neural plate border, including trunk epidermal sensory neurons and tail nerve cord neurons, as well as in the neural tube closure. Here we have investigated the roles of Oikopleura dioica pax37A and pax37B in the development of the OE, by using CRISPR-Cas9 mutant lines and analyzing scRNA-seq data from wild-type animals. We found that pax37B but not pax37A is essential for the differentiation of cell fields that produce the food concentrating filter of the house: the anterior Fol, giant Fol and Nasse cells. Trajectory analysis supported a neuroepithelial-like or a preplacodal ectoderm transcriptional signature in these cells. We propose that the highly specialized secretory epithelial cells of the Fol region either maintained or evolved neuroepithelial features. This is supported by a fragmented gene regulatory network involved in their development that also operates in ascidian epidermal neurons.

Extreme genome scrambling in marine planktonic Oikopleura dioica cryptic species.

Genome structural variations within species are rare. How selective constraints preserve gene order and chromosome structure is a central question in evolutionary biology that remains unsolved. Our sequencing of several genomes of the appendicularian tunicate Oikopleura dioica around the globe reveals extreme genome scrambling caused by thousands of chromosomal rearrangements, although showing no obvious morphological differences between these animals. The breakpoint accumulation rate is an order of magnitude higher than in ascidian tunicates, nematodes, Drosophila, or mammals. Chromosome arms and sex-specific regions appear to be the primary unit of macrosynteny conservation. At the microsyntenic level, scrambling did not preserve operon structures, suggesting an absence of selective pressure to maintain them. The uncoupling of the genome scrambling with morphological conservation in O. dioica suggests the presence of previously unnoticed cryptic species and provides a new biological system that challenges our previous vision of speciation in which similar animals always share similar genome structures.

Anti-Trichomonas vaginalis Activity of Marine Ascidians (Tunicates; Ascidiacea) from the Bushehr Province, Iran.

The aim of the current research is to evaluate the antiparasite effects of compounds isolated from marine ascidian tunicates on Trichomonas vaginalis. Ascidian tunicates after collection were cut into small pieces, freeze-dried, and powdered. The resulting material was subjected to extraction in double-distilled water, ethanol, n-hexane, and dichloromethane. To fractionate the extracts and identify the most bioactive compound, silica gel column chromatography and GC-M/S analysis were used. Fraction 18 of silica gel column chromatography of ethanol extract was the most effective against T. vaginalis. The respective IC50, CC50, and SI values for fraction 18 were 28.62 μg/mL, ˃800 μg/mL, and ˃27.95. GC-M/S analysis of this fraction identified a major phenolic compound (2, 4-bis (1, 1-dimethyl ethyl), whose toxicity against vero cells was only 10.15%. The ethanolic fraction containing phenol-2,4-bis (1,1-dimethylethyl), which has a potent lethality effect on T. vaginalis, may be considered as an antiparasite drug candidate.

Multiple forms of bioimmuration in a coral–crinoid–bryozoan association from the Middle Devonian Onondaga Formation of southeastern Ontario

A large favositid tabulate coral colony from the Middle Devonian Edgecliff Member of the Onondaga Formation, collected from Port Colborne, southernmost Ontario, is described herein because of several noteworthy paleontological features preserved via overgrowth by the coral. The initial substratum colonized and subsequently overgrown by the favositid was a large solitary rugose coral, and outward growth resulted in overgrowth of multiple macrofossils, including relatively long portions of large-diameter (probably camerate) crinoid columns. The crinoid columns, which likely represent portions of robust, recumbent dististelar attachment structures, are otherwise known almost entirely from isolated columnals at the collection locality and similar high-energy biostromal lithofacies in the Edgecliff Member elsewhere. One of the crinoid columns is encrusted by a stenolaemate bryozoan containing several examples of a parasitic bioclaustration structure similar to Catellocaula. Hence, the composite coral–crinoid–bryozoan association contains two forms of bioimmuration: preservation of the relatively intact crinoid columns via overgrowth and eventual incorporation into the base of the tabulate coral and preservation of the outline of the bioclaustrated organism (potentially an ascidian tunicate) via embedment in the bryozoan. This specimen serves as a testament to the diversity of biotic interactions and preservational mechanisms that occurred in Paleozoic carbonate build-ups.

On the occurrence of rare nannoliths (calcareous nannofossils) in the Early Jurassic and their implications for the end‐Triassic mass extinction

AbstractA peculiar record of previously undescribed and/or rarely described nannoliths from the Hettangian to lower Pliensbachian of the Llanbdr (Mochras Farm) core, Wales (UK) is documented here. Some of the observed morphotypes are assigned to didemnid ascidians, commonly observed in the upper Rhaetian, as evidence of the resilience of ascidian tunicates during the end‐Triassic mass extinction. One newly described nannolith present throughout the Hettangian and Sinemurian,Quadrifolium hesselboi, is similar to forms previously observed in the upper Rhaetian, and is therefore considered to be a new nannofossil survivor from the end‐Triassic mass extinction. High‐amplitude changes observed in the absolute and relative abundances of nannoliths, schizosphaerellids and coccoliths across the Hettangian suggest a recovery interval from the end‐Triassic crisis spanning theplanorbis–liasicusammonite zones in four distinct steps with high volatility in environmental conditions, followed by the establishment of more stable environmental conditions in theangulataammonite zone.

The cosmopolitan appendicularian Oikopleura dioica reveals hidden genetic diversity around the globe

Appendicularian tunicates are some of the most abundant mesozooplankton organisms with key roles in marine trophic webs and global carbon flux. Like most appendicularians with cosmopolitan distributions, Oikopleura dioica Fol, 1872 is considered a single species worldwide based on morphological features that distinguish them from other appendicularians. Despite their abundance, however, there are still only ~ 70 described appendicularian species, compared to over 2800 ascidian tunicates. Here we perform a molecular phylogenetic, morphological, and reproductive assessment of O. dioica specimens collected from the Ryukyu Archipelago, mainland Japan, and Europe. The specimens are morphologically very similar, with only detailed examination of the oikoplastic epithelium and quantitative measurements revealing minor distinguishing characteristics. Phylogenetic analyses of the ribosomal gene loci and mitochondrial cytochrome oxidase I (COI) gene strongly indicate that they form three separate genetic clades despite their morphological similarities. Finally, in vitro crosses between the Ryukyu and mainland Japanese specimens show total prezygotic reproductive isolation. Our results reveal that the current taxonomic O. dioica classification likely hides multiple cryptic species, highlighting the genetic diversity and complexity of their population structures. Cryptic organisms are often hidden under a single species name because their morphological similarities make them difficult to distinguish and their correct identification is fundamental to understanding Earth’s biodiversity. O. dioica is an attractive model to understand how morphological conservation can be maintained despite pronounced genetic divergence.

Trifunctional Rare‐Metal‐Free Electrocatalysts Prepared Entirely from Biomass

As part of ongoing efforts to realize a low‐carbon and hydrogen‐based society, there is an increasing demand for high‐performance electrocatalysts, which are key materials for fuel cells and water electrolysis systems. Although platinum group metals exhibit high catalytic activity, increasing production costs due to resource constraints, limited countries of production, and geopolitical supply issues are threatening their use. Therefore, it is crucial to make substantial changes in resource utilization for the sustainable production of electrochemical catalysts. Herein, the preparation of trifunctional biomass‐derived electrocatalysts for the oxygen reduction reaction and oxygen/hydrogen evolution reactions by the pyrolysis of cellulose nanofibers from ascidian tunicates, dried blood meal, and cyanocobalamin is reported. The combination of N, P, FeN4, and CoN4 structures incorporated in the carbon network leads to high electrochemical performance comparable with rare‐metal‐based electrocatalysts and other carbon alloy electrocatalysts. This work provides a possible strategy for fabricating high‐capacity energy storage devices from biomass resources such as industrial waste without the need for rare metals or other harmful heavy metals.

A novel symbiotic relationship between ascidians and a new tunic-boring polychaete (Annelida: Spionidae: Polydora).

Polydora tunicola Abe, Hoshino Yamada, sp. nov., a new spionid species currently considered an obligate symbiont of styelid ascidians, is described based on materials collected from Polycarpa cf. cryptocarpa kroboja (Oka, 1906) and Cnemidocarpa sp. in Izu-Oshima Island and Polycarpa sp. in Wakayama Prefecture, Japan. Polychaeteascidian symbiotic relationships are known only in two syllid species: Myrianida pinnigera (Montagu, 1808) and Proceraea exoryxae Martin, Nygren Cruz-Rivera, 2017. The latter has been the only polychaete known to bore into the tunic of an ascidian. Polydora tunicola sp. nov. is the second known example of a tunic-boring polychaete, which constructs U-shaped burrows in the tunic of the host ascidians. Worms were often concentrated near the host siphons and assumed to use water currents created by the filter-feeding host for suspension feeding. Although the boring mechanism into ascidian tunica is unknown, the plate assay and zymography results consistently detected cellulase activities, suggesting that cellulose digestion may enable the worms to bore into the cellulose-rich ascidian tunics. Polydora tunicola sp. nov. is morphologically similar to P. aura Sato-Okoshi, 1998, P. cornuta Bosc, 1802, P. fusca Radashevsky Hsieh, 2000, P. glycymerica Radashevsky, 1993, P. latispinosa Blake Kudenov, 1978, P. lingulicola Abe Sato-Okoshi, 2020, P. nanomon Orensky Williams, 2009, P. robi Williams, 2000, and P. vulgaris Mohammad, 1972 in having a single median antenna on the caruncle and chaetiger 5 without dorsal superior capillaries but with ventral capillaries. The new species is unique in having a black-rimmed pygidium, distinguishing it from these species. The phylogenetic analyses of the concatenated 18S, 28S, and 16S sequences recovered P. tunicola sp. nov. as the sister species to P. aura within a well-supported clade also including P. lingulicola and P. cf. glycymerica. The bright yellow body color of P. tunicola sp. nov. in life is similar to that of P. aura, however, these two species are distinguished by the former not having modified posterior notochaetae. The symbiotic nature of the association between P. tunicola sp. nov. and styelid ascidians is discussed.

Cultivation of gut microorganisms of the marine ascidian Halocynthia roretzi reveals their potential roles in the environmental adaptation of their host

It has long been known that abundant symbiotic bacteria exist in the tunic and gut of marine ascidians, and that these play crucial roles in host development, physiological metabolism, and environmental adaptation. However, the identity, roles and functions of these symbiotic bacteria are known for only a few strains. In this study, we isolated and cultivated 263 strains of microorganisms from the intestine of the marine ascidian Halocynthia roretzi through a combination of aerobic and anaerobic culture approaches. Most cultivated species, both aerobic and anaerobic, from ascidian stool samples belonged to the genus Bacillus based on 16S rDNA sequencing identification and phylogenetic assays. The distribution of cultured bacteria varied with seasonal changes in environmental conditions. To explore the functions of cultured bacteria, we screened out a strain of Serratia sp. whose extracts showed high antibacterial activity against aquatic pathogens. These findings revealed the potential roles of gut microorganisms in ascidian defense and environmental adaptation, thus providing insights into the interaction and co-evolution between gut bacteria and their hosts.

Massive Gene Loss and Function Shuffling in Appendicularians Stretch the Boundaries of Chordate Wnt Family Evolution.

Gene loss is a pervasive source of genetic variation that influences species evolvability, biodiversity and the innovation of evolutionary adaptations. To better understand the evolutionary patterns and impact of gene loss, here we investigate as a case study the evolution of the wingless (Wnt) family in the appendicularian tunicate Oikopleura dioica, an emergent EvoDevo model characterized by its proneness to lose genes among chordates. Genome survey and phylogenetic analyses reveal that only four of the thirteen Wnt subfamilies have survived in O. dioica—Wnt5, Wnt10, Wnt11, and Wnt16,—representing the minimal Wnt repertoire described in chordates. While the loss of Wnt4 and Wnt8 likely occurred in the last common ancestor of tunicates, representing therefore a synapomorphy of this subphylum, the rest of losses occurred during the evolution of appendicularians. This work provides the first complete Wnt developmental expression atlas in a tunicate and the first insights into the evolution of Wnt developmental functions in appendicularians. Our work highlights three main evolutionary patterns of gene loss: (1) conservation of ancestral Wnt expression domains not affected by gene losses; (2) function shuffling among Wnt paralogs accompanied by gene losses; and (3) extinction of Wnt expression in certain embryonic directly correlated with gene losses. Overall our work reveals that in contrast to “conservative” pattern of evolution of cephalochordates and vertebrates, O. dioica shows an even more radical “liberal” evolutionary pattern than that described ascidian tunicates, stretching the boundaries of the malleability of Wnt family evolution in chordates.

Glycoside hydrolases from the tunics of two Antarctic ascidians (Ascidia challengeri and Pyura bouvetensis) and the tropical species Phallusia nigra

Tunicates are the only metazoans capable of synthesising cellulose, which is present in the tunic. Cellulases (endo and exoglucanases) that can degrade or remodel cellulose might be necessary during the growth phase and during scarring after injury to the tunic. However, the occurrence of these enzymes in the tunic has not been reported. Hence, the aim of the study was to investigate the presence of cellulases as well as other glycoside hydrolases, such as xyloglucanases and mannanases, in the tunic of two Antarctic ascidian species, Ascidia challengeri and Pyura bouvetensis, and in the tropical species Phallusia nigra. Endoglucanases and xyloglucanase were found in all three species. Galactomannanases were detected in P. nigra and P. bouvetensis but were not found in A. challengeri. Cellulases that can degrade Avicel were found in P. bouvetensis. To the best of our knowledge, this is the first report detailing the presence of these enzymes in the tunic of ascidians. In A. challengeri, the optimal temperature for the endoglucanase activity was 9 °C, higher than their habitat temperature, but within the temperature tolerance of the species, while that of P. nigra was at 25–30 °C, which is close to the maximum temperature of its habitat. The temperature at which the endoglucanase has optimal activity is discussed as a proxy to the limit of thermal tolerance of the species that express this enzyme.

Feeding habits of the white-spotted boxfish Ostracion meleagris reveal a strong preference for colonial ascidians

Boxfishes (Ostraciidae) are known to be benthivores and mainly consume sessile organisms on reefs, such as tunicates. Ascidian tunicates contain various toxins and seldom suffer from predation by reef fishes, except for boxfishes. However, the importance of ascidian tunicates has not been sufficiently examined in the feeding ecology of boxfishes. We conducted field surveys to investigate the feeding habits of the white-spotted boxfish Ostracion meleagris on the reefs of southern Japan. Photosymbiotic ascidians predominantly contributed to the boxfish diet, and observed colonies of ascidians were bitten frequently by the boxfish during the survey. During the day, the feeding boxfish moved within their home ranges in a rocky, shallow zone of the studied reefs, which harbored photosymbiotic ascidian colonies of the family Didemnidae at higher densities than in the surrounding zones, including the spawning sites that were located hundreds of meters offshore. Boxfish home ranges overlapped considerably, but intraspecific agonistic interactions rarely occurred among the females, likely due to the abundance of ascidian colonies. No other types of fish were observed to make attempts to feed on the ascidian colonies on the reef throughout the study period, except for a congener Ostracion cubicus. These results suggest that the boxfish strongly rely on the photosymbiotic ascidians as a less-competitive food resource, and the limited distribution pattern of photosymbiotic ascidians in shallow water near shore can be an important factor in promoting a spawning migration in the boxfish.

The role of invasive tunicates as reservoirs of molluscan pathogens

Ascidian tunicates frequently display rapid expansion when introduced beyond their native range and are considered successful invaders. This invasive potential may be exacerbated by a warming climate, allowing for the occupation of environmental niches previously held by native species. Research into tunicate invasion ecology is prevalent, but less is known about their role in pathogen maintenance. This study investigated the impact of invasive tunicates on the maintenance of pathogens that affect commercial bivalves, including the cultured species Ostrea edulis (European flat oyster) and Crassostrea gigas (Pacific oyster), and the fished species Cerastoderma edule (Common cockle). Focal pathogens included ostreid herpesvirus OsHV-1 μVar, Vibrio aestuarianus, Bonamia ostreae and Minchinia spp. The range of pathogens in their molluscan hosts was determined and the tunicates Botrylloides violaceus, Didemnum vexillum and Styela clava were then screened for these same pathogens, using both field samples from oyster culture sites and marinas and a series of laboratory cohabitation trials. Sample sites reflected areas close to and further away from known pathogen sources. PCR, Sanger sequencing and histology confirmed the presence of B. ostreae and Minchinia mercenariae-like in S. clava, and V. aestuarianus was confirmed by qPCR in B. violaceus and D. vexillum. Furthermore, histology confirmed Minchinia mercenariae-like sporonts in S. clava suggesting that the tunicate can facilitate replication of this species. S. clava also maintained B. ostreae in tanks with no oysters present. The results indicate that tunicates can act as reservoirs of infection in areas where disease occurs and potentially transport diseases to uninfected sites.

Convergent evolution of the vertical transmission mode of the cyanobacterial obligate symbiont Prochloron distributed in the tunic of colonial ascidians

Convergent evolution of the vertical transmission mode of the cyanobacterial obligate symbiont <i>Prochloron</i> distributed in the tunic of colonial ascidians

A rare case of an evolutionary late and ephemeral biomineralization: tunicates with composite calcareous skeletons

AbstractIn contrast to almost all other invertebrate phyla that constructed biomineralized skeletons during the “Cambrian explosion” and maintained them during the entire fossil record, ascidian tunicates evolved this protective and stabilizing advantage only during the Permian, although soft-bodied representatives of this subphylum made their first appearance already in the early Cambrian. It remains enigmatic why these compound calcareous skeletons persisted only until the Late Triassic, subsequently followed by less-rigid internal skeletons from the Lower Jurassic onwards, which consist of scattered isolated spicules only. In addition to recently described aragonitic ascidian exoskeletons from the Permian and Triassic, new discoveries of similar, but colonial ascidian compound endoskeletons in the lower Carnian exhibit a short-living branch of this group, which moreover contain the first indubitable calcareous spicules. The latter are embedded in the solid endoskeleton, which is composed of polygonal aragonitic plates with smooth outer and zigzag lined inner boundaries. They consist of irregular, parallel (orthogonal), or fan-shaped (clinogonal) arrangements of acicular aragonite crystals. The following taxa are described as new: order Cassianomorpha new order with the family Cassianosomidae new family and the genusToscanisomanew genus with the speciesT.multipartitumnew species andT.triplicatumnew species.UUID:http://zoobank.org/03555353-cdab-42e8-8e99-9bfce15fa249

Homologues of p48 Protein from Morula Cells of Ascidian Styela rustica in Other Species of Stolidobranchia

Tunicata is an interesting phylogenetic group, at the base of the branch leading to Chordata. Ascidians (Tunicata, Ascidiacea) morula blood cells are involved in defense reactions and in the formation of the tunic extracellular matrix. The ascidian tunic is hardened as a result of sclerotization of matrix proteins due to the action of phenol oxidase enzyme, contained in morula cells. Morula cells of the ascidian Styela rustica contain two major proteins, the function of which is still unknown; one of them is a protein with molecular mass of 48 kDa – p48. The aim of present study was to search for possible homologues of p48 in ascidians belonging to the Stolidobranchia order: Styela coriacea (Styelidae), Molgula citrina (Molgulidae), Boltenia echinata, Halocynthia aurantium (Pyuridae) and to determine the tissue distribution of those homologues. In order to show the presence of p48 in the tissues of sea squirts we used indirect immunolabeling method on paraffin sections.It was shown that the antibodies bind with morula cells of Styela rustica, Styela coreacea, Boltenia echinata, with the tunic matrix in all studied species and with test cells of Styela rustica, Styela coreacea, Boltenia echinata, Molgula citrina. It gives us a ground to assume the existence of p48 homologues in all studied ascidians and to expect a common mechanism of their participation in the tunic formation, as possible substrates of the phenol oxidase system.

Nephromyces Represents a Diverse and Novel Lineage of the Apicomplexa That Has Retained Apicoplasts.

A most interesting exception within the parasitic Apicomplexa is Nephromyces, an extracellular, probably mutualistic, endosymbiont found living inside molgulid ascidian tunicates (i.e., sea squirts). Even though Nephromyces is now known to be an apicomplexan, many other questions about its nature remain unanswered. To gain further insights into the biology and evolutionary history of this unusual apicomplexan, we aimed to 1) find the precise phylogenetic position of Nephromyces within the Apicomplexa, 2) search for the apicoplast genome of Nephromyces, and 3) infer the major metabolic pathways in the apicoplast of Nephromyces. To do this, we sequenced a metagenome and a metatranscriptome from the molgulid renal sac, the specialized habitat where Nephromyces thrives. Our phylogenetic analyses of conserved nucleus-encoded genes robustly suggest that Nephromyces is a novel lineage sister to the Hematozoa, which comprises both the Haemosporidia (e.g., Plasmodium) and the Piroplasmida (e.g., Babesia and Theileria). Furthermore, a survey of the renal sac metagenome revealed 13 small contigs that closely resemble the genomes of the nonphotosynthetic reduced plastids, or apicoplasts, of other apicomplexans. We show that these apicoplast genomes correspond to a diverse set of most closely related but genetically divergent Nephromyces lineages that co-inhabit a single tunicate host. In addition, the apicoplast of Nephromyces appears to have retained all biosynthetic pathways inferred to have been ancestral to parasitic apicomplexans. Our results shed light on the evolutionary history of the only probably mutualistic apicomplexan known, Nephromyces, and provide context for a better understanding of its life style and intricate symbiosis.

Structural and Mass Spectrometric Imaging Analyses of Adhered Tunic and Adhesive Projections of Solitary Ascidians.

Most ascidian species settle on underwater substrates during a short free-swimming tadpole larval period. During this process, "rapid adhesion" occurs on adhesive papillae located at the anterior region of the cephalenteron. Settled and transformed ascidians subsequently expand the attachment area by "slow adhesion" with ampullae. In the present study, we attempted to identify the ultrastructures related to the adhesion process and adhesive materials in the ascidian tunic and to elucidate the biological function of vanadium in adhesion. We focused on an adhesive organ named the adhesive projection, which is newly generated by the adhered tunic to enlarge the bonding area between ascidian and substrate. Based on its structure and the presence of vanadiumcontaining blood cells, the adhesive projection was considered to be a large tunic vessel. At the adhered tunic, eosinophilic regions and migrated tunic cells were observed, but metal deposition was not detected. We speculate that the eosinophilic materials were components of the adhesive glue, and these are likey produced in epithelial cells, tunic cells, or both. Furthermore, using imaging mass spectrometry, we identified eight tunic-specific molecules as glue candidates.

The colonial ascidian Diplosoma listerianum enhances the occurrence of the hydrozoan Obelia sp. during early phases of succession

Recruitment patterns of sessile species often do not reflect the composition of the local propagule pool. This is, among other processes, attributed to the stimulation or inhibition of settlement by resident species. In an experimental study, we evaluated the effects of different densities of the ascidian Diplosoma listerianum on the settlement of the hydrozoan Obelia sp. For this, we monitored the cover of the dominant fouler Obelia sp. on vertically orientated PVC tiles, which were either bare or pre-seeded with two different densities (sparse or dense) of Diplosoma colonies, over the course of 8 weeks. The settlement tiles were deployed at two study sites in La Herradura Bay, Chile. The presence of D. listerianum enhanced the settlement or the growth or both of the colonial hydrozoan, but this effect disappeared within 4–8 weeks. Furthermore, we tested whether the initial enhancement of Obelia sp. by Diplosoma colonies goes back to the fact that larvae, which reject the ascidian tunic as a settlement substratum after a first contact, colonize nearby surfaces because of their limited mobility. However, we found no support for this assumption. We rather suggest that D. listerianum facilitated colonization indirectly by the accumulation of organic material in its vicinity and/or by its pumping activity. Initial resident-mediated enhancement of the hydrozoan was overridden by processes such as competition between later colonizers within the course of weeks and we could not detect any lasting effects of D. listerianum on the structure of the developing communities.

Measurement of refractive indices of tunicates\u2019 tunics: light reflection of the transparent integuments in an ascidian Rhopalaea sp. and a salp Thetys vagina

BackgroundTunic is a cellulosic, integumentary matrix found in tunicates (Subphylum Tunicata or Urochordata). The tunics of some ascidian species and pelagic tunicates, such as salps, are nearly transparent, which is useful in predator avoidance. Transparent materials can be detected visually using light reflected from their surfaces, with the different refractive indices between two media, i.e., tunic and seawater, being the measure of reflectance. A larger difference in refractive indices thus provides a larger measure of reflectance.ResultsWe measured the refractive indices of the transparent tunic of Thetys vagina (salp: Thaliacea) and Rhopalaea sp. (ascidian: Ascidiacea) using an Abbe refractometer and an ellipsometer to estimate the light reflection at the tunic surface and evaluate the anti-reflection effect of the nipple array structure on the tunic surface of T. vagina. At D-line light (λ = 589 nm), the refractive indices of the tunics were 0.002–0.004 greater than seawater in the measurements by Abbe refractometer, and 0.02–0.03 greater than seawater in the measurements by ellipsometer. The refractive indices of tunics were slightly higher than that of seawater. According to the simulation of light reflection based on rigorous coupled wave analysis (RCWA), light at a large angle of incidence will be completely reflected from a surface when its refractive indices are smaller than seawater. Therefore, the refractive index of integument is important for enabling transparent organisms to remain invisible in the water column.ConclusionIn order to minimize reflectance, the refractive index should be similar to, but never smaller than, that of the surrounding seawater. The simulation also indicated that the presence or absence of a nipple array does not cause significant difference in reflectance on the surface. The nipple array on the tunic of the diurnal salp may have another function, such as bubble repellence, other than anti-reflection.