Abundance and diversity of lipid compounds in Exaiptasia diaphana is altered as a function of symbiotic state and life stage

Abundance and diversity of lipid compounds in Exaiptasia diaphana is altered as a function of symbiotic state and life stage | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Abundance and diversity of lipid compounds in Exaiptasia diaphana is altered as a function of symbiotic state and life stage Erick White, Luke Marney, Mark Phillips, Jenna Scott, Aster Parkin, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9023951/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 4 You are reading this latest preprint version Abstract Scleractinian corals engage in an endosymbiotic relationship with dinoflagellate algae to meet their nutritional demands in oligotrophic waters. Algae transfer photosynthetically-derived compounds to the host and host is structured to support and sustain the symbionts. The composition and abundance of these compounds as a function of host life stage and symbiotic state have yet to be described. Therefore, we aimed to characterize the effect of symbiosis and host life stage on the lipidomic profiles of the sea anemone Exaiptasia diaphana (commonly referred to as Aiptasia), a model system for the study of coral symbiosis. We sampled aposymbiotic and symbiotic Aiptasia adults, pedal lacerates, and larvae and isolated lipid fractions for liquid-chromatography mass-spectrometry (LC-MS) analysis. Over 200 lipid compounds were identified across all sample groups. There were more compounds in symbiotic than in aposymbiotic at every life stage. Lipidomic profiles separated based on both symbiotic state and life stage. Of the major lipids classes detected, triglycerides and ceramides were more abundant in symbiotic animals, phosphatidylcholines (PCs) were more abundant in pedal lacerates in both symbiotic states, and abundances of phosphatidylethanolamines (PEs) varied based on symbiotic state. Our results highlight the major influence of symbiotic state on lipid profiles in Aiptasia at different life stages and provide insight into how the lipidome changes in early life-stage cnidarians. Lipidomics LC-MS Aiptasia symbiosis pedal lacerates larvae Figures Figure 1 Figure 2 Figure 3 Introduction Reef-building coals form the foundation of coral reef ecosystems. Corals are comprised of a mutualistic endosymbiosis between the cnidarian host and photosynthetic dinoflagellate algae (family Symbiodiniaceae) that reside within host gastrodermal cells (Muscatine and Porter 1977 ; Falkowski et al. 1984 ; LaJeunesse et al. 2018 ). Maintenance of the symbiosis is based on a reciprocal exchange of nutrients between the partners. The symbiont supplies photosynthetically-fixed carbon that meets most of the metabolic needs of the host (Muscatine et al. 1984 ; Yellowlees et al. 2008 ). In return, the host provides the symbiont with waste products in the form of inorganic carbon and nitrogen, the raw materials for photosynthesis and growth, and a stable habitat with access to sunlight (Falkowski et al. 1993 ; Cui et al. 2022 ). Corals thrive in the oligotrophic waters of the tropics and sub-tropics due in large part to this efficient exchange of nutrients between symbiotic partners. The majority (approximately 80%) of coral species acquire symbionts from the environment with each host generation (i.e. horizontal transmission) (Baird et al. 2009a , b ; Hartmann et al. 2019 ). The remaining species pass their symbionts directly from the host parent to offspring (i.e. vertical transmission). For corals that engage in horizontal transmission, algae are acquired either at the larval or juvenile polyp stage (Schwarz et al. 1999; Elder et al. 2022 ). Prior to the onset of symbiosis, aposymbiotic larvae must rely on other sources of nutrition to fuel their metabolic needs. They can acquire nutrients either from heterotrophic feeding (Schwarz et al. 1999; Brusca and Brusca 2003 ) or maternally derived nutrients in the form of lipid droplets (Thorson 1950 ; Marlow and Martindale 2007 ). Studies of the compounds present in coral eggs and larvae across species have revealed that nutritionally-rich lipids, such as wax esters and triacylglycerols, are the predominant metabolites present, and vary in abundance based on larval settlement period and dispersal range (Harii et al. 2007 ; Roper et al. 2024 ). In addition, the metabolites produced vary depending on mode of symbiont acquisition (horizontal versus vertical) and developmental stage. Studies of larvae have shown that the presence of symbionts can increase lipid droplets compared to aposymbiotic larvae (Voss et al. 2023 ) and that lipid sources shift from being maternally-derived to symbiont-derived as symbiosis becomes established (Huffmyer et al. 2025 ). Thus, symbiosis is an important factor in determining the lipidomic profiles of symbiotic and aposymbiotic offspring at early life stages. In addition to sexual reproduction, many corals and sea anemones engage in asexual reproduction by fission, budding, or pedal laceration. Asexual reproduction results in the formation of large colonies of corals, which in turn form the structure of the reef ecosystem (Reitzel et al. 2013). Since asexual offspring are formed directly from parental tissue, symbionts are vertically transmitted and already present as offspring progress through development. The presence of symbionts could therefore influence metabolism and development at these early life-stages. Indeed, recent studies in the sea anemone Exaiptasia diaphana (from here on referred to as Aiptasia) have shown that the presence or absence of heterotrophic feeding and symbiosis can greatly influence the developmental patterning and survival of pedal lacerates (Presnell et al. 2022 ; Bedgood et al. 2024 ). While the lipidome of coral adults and larvae have been described (Garret et al. 2013; Imbs et al. 2021 ; Roper et al. 2024 ; Huffmyer et al. 2025 ), less attention has been given to the lipidome of asexual offspring and the influence of symbiosis on its composition. Aiptasia is an ideal model system for the study of coral-dinoflagellate symbiosis. Like corals, it harbors species of Symbiodiniaceae in its tissues. However, unlike corals, it reproduces both sexually and asexually in the laboratory and can be cultured in both symbiotic and aposymbiotic states (Grawunder et al. 2015 ; Roberty et al. 2024 ). In this study, we used liquid chromatography-mass spectrometry (LC-MS) to characterize the lipidome of Aiptasia from different life stages. We compared lipid profiles from symbiotic and aposymbiotic adults, sexually-produced larvae, and asexually-produced pedal lacerates to evaluate the effects of symbiosis and life stage on lipid composition. Methods Animal husbandry Symbiotic and aposymbiotic adults and pedal lacerates were sampled from the H2 clonal line of Aiptasia with its native symbiont Breviolum minutum (Xiang et al. 2013; LaJeunesse et al. 2018). Aposymbiotic adults were generated via menthol bleaching (Matthews et al. 2016) and maintained for at least three months prior to the start of the experiment. Adults used for adult and pedal lacerate samples were maintained in clear (for symbiotic animals) or black (for aposymbiotic animals) polycarbonate tubs (Cambro) in 300 mL 0.45 µm filtered artificial sea water (FASW, Instant Ocean; 32 ppt) in a Percival incubator set to 25°C on a 12-hour/12-hour light/dark cycle, with a full spectrum irradiance of 10-20 µmol quanta m -2 s -1 . Adult populations were fed with S.presso (INVE Aquaculture, Belgium) gut-infused brine shrimp ( Artemia nauplii ) three times a week with subsequent cleaning and changing of water prior to the start of the experiment. Symbiotic and aposymbiotic adults were randomly selected and used as individual replicate samples for the experiment (n=3 each). For generation of sexually-produced larvae, separate tubs of symbiotic Aiptasia (H2 females, CC7 males, and VWA males) meant for reproductive purposes were kept in a separate Percival incubator set to 29°C on a 12-hour/12-hour light/dark cycle, with a full spectrum irradiance of 10-20 µmol quanta m -2 s -1 . These populations were fed gut-infused brine shrimp five times a week to attain a mature body size followed by cleaning and water changes with FASW pre-warmed to 29°C. Once adults had reached maturity with visible and viable gonads, reproductive animals were maintained at these conditions for at least three months prior to the experiment. Larvae destined for addition of symbionts were inoculated with Breviolum minutum algae (SSB01 strain). B. minutum was cultured in 250 mL suspension culture flasks (Greiner 658190) in F/2 medium (Bigelow MKf250L) under the same temperature and lighting conditions as non-reproductive adult Aiptasia anemones described above. Generation of pedal lacerates and larvae Pedal lacerates were generated as previously described in White et al. (2026) and in Presnell et al. (2022). A total of 90 symbiotic and aposymbiotic lacerates each (n=180 total) were generated and divided between three replicate 96-well polystyrene plates. Plates were cleaned with cotton swabs and FASW. Lacerates were cut one week prior to the sampling timepoint to ensure larvae and lacerates were approximately the same age when sampled. To induce spawning and generate larvae, fluorescent blue light was used in the reproductive animal incubator to simulate the lunar cycle that triggers spawning. This blue light was turned on during the night phase of the 12:12 hr light cycle for five consecutive days. Seven to eight days after the blue light was turned on, gravid adults released eggs and sperm. Eggs were pipetted into six glass dishes. Sperm was first filtered through a 50 µm filter with FASW to remove any tissue debris or symbiont clumps and then decanted into the glass dishes with the eggs to generate H2 x CC7/VWA crosses. Three dishes were designated for later addition of symbionts to generate symbiotic larvae while the other three were untouched, with the larvae left to develop into aposymbiotic larvae. Complete FASW water changes were completed everyday by filtering larvae from each dish through a 50 µm filter and placing larvae back into their dishes with fresh FASW. For larvae designated to become symbiotic, larvae first underwent a water change as described above and added back into glass dishes with minimal FASW on day three post-fertilization. Then, B. minutum cultures were resuspended in FASW and added to larval dishes to reach a concentration of 1 x 10 5 cells/mL. Incubation of larvae with B. minutum lasted for 24 hours and underwent a complete water change afterwards. This process was then repeated on day four post-fertilization to ensure symbiont uptake into host tissue with a complete water change on day five post-fertilization. Lipid isolation Adults, lacerates, and symbiotic larvae were all sampled one week (day seven post-laceration; day seven post-fertilization) after the start of the experiment. Aposymbiotic larvae were sampled at five days post-fertilization to ensure there would be a robust sample size due to a decrease in live and active larvae as more time passed. Animals were sampled and divided into three biological replicates for adults (n=1), larvae (n=300), and lacerates (n=30) in 1.5 mL microfuge tubes. For all life stages, host tissue samples were homogenized with a motorized mortar and pestle in 300 µL of extraction buffer (extraction buffer [100 mM 7.33 pH Tris, 10 mM 8.04 pH EDTA, 100 mM NaCl], protease inhibitor [Roche 04693159001], and 1% Triton-X). Glass beads (450 microns) were added to each tube and topped up to 1 mL with extraction buffer and then material was further lysed on a Qiagen Tissue Lyser for one minute at 30 Hz to dissociate any remaining cells and release metabolites into the extract. Samples were centrifuged at 3000 rpm for five minutes to pellet glass beads, host debris, and algae. The resulting extract was aliquoted for lipid extraction (600 µL). Prior to lipid extraction, 6 µL of EquiSPLASH Quantitative Mass Spec Internal Standard (Avanti Research 330731), containing 13 deuterated lipid standards at a concentration of 100 µg/mL each, was added to each tube at a final concentration of 0.5-1 µg/mL. Lipids were extracted using a modified Folch method (Folch et al. 1956) described in Baumann et al. (2021) and frozen at -20°C until further use. Lipid fractions were dried under vacuum and resuspended in 100 µL of chloroform and prepped for LC-MS analysis. Lipidomic analysis Ultra-performance liquid-chromatography was performed using a 1.7 μm particle, 2.1 ×100 mm, CSH C18 Column (Waters, Milford, MA, United States) coupled to a quadrupole time-of-flight mass spectrometer (SCIEX, ZenoTOF 5600) housed at the Oregon State University Mass Spectrometry center. The SCIEX, ZenoTOF 5600 was operated in information-dependent MS/MS acquisition mode (IDA) with the following parameters: IDA survey accumulation time of 0.1 s, IDA survey TOF mass range of 100-1200 m/z, a IDA survey collision energy of 10 V with no collision energy spread, 30 candidates for data dependent acquisition with a collision energy of 30, without excluding former candidate ions, and scanning a TOF range of 50-1200. A declustering potential of 50 was used throughout the analysis with a Zeno threshold of 20,000. The electrospray ionization source (Turbo Ion Spray) was operated at 5500 V with the following parameters: curtain gas of 35. CAD gas of 7, Ion source gas 1 of 50 psi, Ion source gas 2 of 40 psi, and temperature of 500°C. The chemical composition of lipid compounds determines their ability to be ionized to a cation (positively charged) or an anion (negatively charged) and detected. Therefore, samples were ionized in both positive ion mode by the addition of a proton (H+) to molecules and negative mode by the loss of a proton from molecules to ensure the greatest number of compounds were identified by their mass-to-charge ratio (m/z+1) within a sample. For positive ion mode, the mobile phases consisted of (A) 60:40 (v/v) acetonitrile: water with ammonium formate (10 mM) and formic acid (0.1%) and (B) 90:10 (v/v) isopropanol: acetonitrile with ammonium formate (10 mM) and formic acid (0.1% formic acid). For analyses run in the negative ion mode, ammonium acetate (10 mM) was used as the modifier. The chromatographic gradient has been described previously (Cajka and Fiehn, doi: 10.1007/978-1-4939-6996-8_14). Annotation of metabolite signals was completed in MSDial (5.1.230517) using the LipidMaps database. Statistical analysis Statistical analysis of the relative abundance of detected lipids across life stages was conducted using MetaboAnalyst (version 6; Pang et al. 2024). Relative abundances from each sample were normalized to the peak areas of the internal standard values of 13 deuterated lipids (EquiSPLASH A83731, Avanti Research) and host protein content (Bradford 1976) prior to analysis in MetaboAnalyst. Normalized data were then subjected to log 2 transformation and mean-centering and pareto scaling, resulting in a clear relation between rank of metabolites and relative abundance observed. Data normality was visually confirmed. Lipidomic profiles between all treatment groups were compared using principal component analysis (PCA) from Euclidean distances and statistically analyzed via permutational multivariate analysis of variance (PERMANOVA) based on 999 permutations. Due to a recovery of protein below the limits of detection of the Bradford assay, larval data were removed from the main dataset and analyzed separately without indexing to protein but still underwent log 2 transformation and mean center-pareto scaling (Supplemental Figures and Tables). A one-way ANOVA with an FDR correction was used to test for significant differences in lipid abundance among adult and lacerate test groups. The top 75 most significantly different compounds between adult and lacerate groups were visualized with a heatmap using the Euclidean distance and Ward clustering algorithm. UpSet plots were used to illustrate intersections of lipids specific to each sample type using the UpSetR package in RStudio version 4.0.2. M/z+1 values were deemed relevant if present in at least two replicates. Log 2 -fold changes between aposymbiotic and symbiotic larvae were visualized using a volcano plot with the ggplot2 package in R. Statistical significance for all tests was set with an FDR correction p <0.05. Results A total of 220 lipids were detected and assigned putative identities across all sample groups (Table S1). The lipidomes of adults and lacerates formed separate clusters with no overlap based on both symbiotic state and life stage (Figure 1; PERMANOVA, F=170.82, R 2 =0.98463, FDR P=0.001). Symbiotic state (PC1) accounted for 58.7% of the variance detected in lipid profiles between samples, while life stage (PC2) accounted for 18.5% of the variance between samples. A total of 164 of the 220 lipids that were detected between lacerates and adults were significantly different based on life stage, symbiotic state, or both (Table S1; Tukey’s HSD, FDR P< 0.05). Larval samples were also separated and clustered based on symbiotic state, though this was not statistically significant (Figure S1; PERMANOVA, F-value=25.883, R 2 =0.86615, FDR P=0.1). We observed differences in the total number of shared and unique lipids detected across sample groups based on symbiotic state and life stage (Figure 2). Symbiotic sample groups had more lipids present than their aposymbiotic counterparts, measured across both adults and pedal lacerates. Within symbiotic states, adult replicates had more lipids detected (sym=172; apo=112) followed by lacerates (sym=135; apo=81) (Figure 2A). A total of 71 lipids were shared across all symbiotic states and life stages. There were 42 lipids that were unique to symbiotic samples, 26 unique to symbiotic adults, and only one that was unique to symbiotic lacerates. There were four lipids that were unique to aposymbiotic samples at both life stages, four lipids that were only present in aposymbiotic adults, while aposymbiotic lacerates had no unique lipids. Adults had six lipids that were unique to that life stage, while lacerates had no unique lipids corresponding to that life stage (Figure 2B). A heatmap of the top 75 significantly different lipid compounds across sample groups clearly depicts differences across symbiotic states and life stages (Figure 3; One-way ANOVA, FDR P<0.05). Symbiotic treatment groups had higher concentrations of nearly all ceramide and triglyceride compounds detected. Aposymbiotic treatment groups had higher concentrations of a subset of ether-linked phosphatidylethanolamine (PE-O) compounds, while a different subset of PE-O compounds was significantly upregulated in adult groups. All phosphatidylcholines (PCs) detected were significantly upregulated in lacerate sample groups only. Lastly, there were eight compounds, mostly ceramides, that were unique to the symbiotic adult sample group alone. For larvae, a total of 43 out of 80 lipids were significantly different between symbiotic and aposymbiotic samples, which was comprised of entirely phospholipids (PCs and PE O-s); only one of the significantly different lipids detected in larval samples was upregulated in aposymbiotic larvae (PC 27.0; Figure S2 and Table S2; Student’s T-test, FDR P<0.05). Discussion Symbiosis alters the lipidome in Aiptasia Numerous studies have shown that symbiotic state exerts a major influence on symbiotic cnidarians at the genomic (Matthews et al. 2017), transcriptomic (Lehnert et al. 2014), and proteomic (Valadez-Ingersoll et al. 2025) level. Our study extends these findings by showing that the presence of symbionts changes the lipidomic profiles of Aiptasia (Figure 1) in relation to the total amounts and types of lipids detected, as well as their abundances at early (pedal lacerate) and mature (adult) life stages (Figures 2-3). The higher number of lipids in symbiotic compared to aposymbiotic animals illustrates the dominant effect that symbiosis has on the lipidomic profiles in Aiptasia. Differences in the lipidomes of adult Aiptasia as a function of symbiotic state have been described. Garrett et al. (2013) found that non-polar lipids ( i.e . energy-rich compounds and shorter chain length phospholipids) were higher in symbiotic compared to aposymbiotic Aiptasia. The authors hypothesized that symbiosis leads to a remodeling of the chain lengths in phospholipids, and that the released fatty acids are used as fuel for the synthesis of other lipids. This points to the high enery demand required to maintain symbiosis. Hambleton et al. (2019) expanded on this research and found that the abundances of different sterol compounds in Aiptasia are not only influenced by symbiotic state, but also by host genetic line, species of algae present, and heterotrophic feeding. In addition, lipids can have specific spatial distribution between symbiotic states that is involved in regulation of symbionts, as seen with ceramides being localized in the gastrodermis around symbiont-containing host cells and symbiont-derived betaine lipids concentrated in light-exposed tentacles (Chan et al. 2023). The lack of differences between symbiotic and aposymbiotic larvae suggests that, within the first week of symbiont incorporation into host tissue, the presence of symbionts does not drive a major change in the types and amounts of lipids detected in larvae. The low algal density in larvae and young age of larvae at the sampling time could explain these results, and as symbionts proliferate in larvae, lipid profiles between symbiotic and aposymbiotic larvae could begin to differentiate. Cnidarian larvae are lecithotrophic, which means that they are provisioned with maternally-derived nutrients, typically in the form of lipids, that are used to fuel development and growth until symbionts are acquired from the environment (Marlow and Martindale 2007; Huffmyer et al. 2025) or the larvae begin to feed. Expression profiling of Aiptasia larvae has shown over 300 differentially expressed genes between aposymbiotic and symbiotic larvae, many of which are hypothesized to be involved in symbiosis establishment (Wolfowicz et al. 2016). Together with our data, this suggests that during the early days post-fertilization and post-colonization when the algal density remains low, symbiotic larvae are using maternally-derived lipids to fuel their energetic needs while prioritizing development and symbiosis establishment, thereby resulting in a lipidomic profile that is similar to aposymbiotic larvae. Future studies could conduct a time-series experiment examining lipidomic profiles of larvae sampled through time post-fertilization and post-inoculation to determine how lipid profiles change as larvae transition from being aposymbiotic to symbiotic and as they develop into juvenile polyps. Pedal lacerates are distinct from adult Aiptasia The separation of lipidomic profiles and abundances by life stage in Aiptasia in this study suggests that lipidomic profiles are altered in an age-dependent manner. For symbiotic Aiptasia, this could be due to the difference in symbiont density between life stages and/or the symbiosis being at different stages of establishment. The symbiosis is fully established in adults, while it is becoming re-established with vertically-transmitted symbionts in lacerates. Studies in Montipora capitata larvae showed that there are metabolic changes as larvae develop, switching from a maternally-derived nutrition source to a symbiont-derived one (Huffmyer et al. 2025). Therefore, as symbionts proliferate in lacerates, host metabolism will likely shift to accommodate inter-partner nutrient exchange, and in turn will lead to differential production and usage of lipids within the holobiont. Future studies can examine if lipid profiles in lacerates change based on symbiont density and how profiles change as lacerates progress into mature adults. The separation of lipidomic profiles by life stage in our study also suggests that, even though pedal lacerates are formed from adult tissue, the process of laceration and clonal propagation alters the lipidomic profile in these organisms. Cnidarians have a variety of life history strategies, even within asexual reproduction alone, that emerge from discrete patterns of developmental patterning (Reitzel et al. 2013; summarized in Caruso et al. 2022). Other studies have shown that Aiptasia pedal lacerates have specific patterns of tentacle formation, development, and growth that are affected by symbiotic state and feeding (Presnell et al. 2022; Bedgood et al. 2024). Taken together with our data, this suggests that pedal lacerates represent a distinct and unique stage in the Aiptasia life cycle and are not simply pieces of adult tissue. Our study highlights the importance of life stage in the cellular makeup of Aiptasia and is something to consider in future studies examining cellular mechanisms of cnidarian symbiosis. Symbiosis and life change influence levels of triglycerides, ceramides, and phospholipids The differential abundance of different lipid classes across symbiotic states we found in our study illustrates that the interpartner dynamics between symbiotic partners greatly alters the lipidomic landscape. Our study shows that triglycerides and ceramides, heavily associated with symbiosis, are upregulated even at early life stages in pedal lacerates (Yamashiro et al. 2005; Imbs et al. 2010). Triglycerides are a major food source for the host and support host health and metabolism (Hamoutene et al. 2008; Imbs 2013). These storage molecules in pedal lacerates be in the mesenteries, which originate in the parental polyp and are replaced with new mesenteries as lacerates develop (Cary 1911; Steinmetz et al. 2017; Presnell et al. 2022). Ceramides are precursors to the sphingolipids sphingomyelin and glycosphingolipids that are used in the plasma membrane. They also play a pivotal role in cell fate determination where they can be converted into sphingosine, a pro-apoptotic signal, or sphingosine-1-phosphate, a pro-survival signal (Kitchen et al. 2017; Rosset et al. 2021). Ceramides are heavily concentrated in symbiotic Aiptasia compared to aposymbiotic anemones and are primarily located in the gastrodermis (Chan et al. 2023). The authors concluded that ceramide-based sphingolipids are involved in the regulation of symbionts in the cnidarian-dinoflagellate symbiosis, and that symbiosis triggers de-novo synthesis or salvaging of sphingolipids. The span of upregulated PE-O lipids as a function of symbiotic state and life stage in our study is an indication of the diversity of functions that they play in cellular processes. Ether lipids, such as PE-O, make up the typical classes of cnidarian lipids and are markers for cnidarian host tissue (Imbs et al. 2010; Sikorskaya et al. 2024, 2025). In addition to being a primary component for phospholipid cellular membranes in eukaryotes, PE-O can act as an antioxidant by scavenging reactive oxygen species (ROS), which are produced as a byproduct of cellular respiration and photosynthesis (Magnusson and Haraldsso 2011; Sikorskaya et al., 2025). The high levels of PE-Os in symbiotic animals could be indicative of the use of PE-O for protective measures against the photosynthetic activity of the algae, although future studies are needed to confirm this hypothesis. PE-O compounds are also known to play a role in membrane fusion and cellular division. A study in Aiptasia found that aposymbiotic anemones had a higher cell density and more dividing cells in both the epidermis and gastrodermis than did symbiotic anemones (Tivey 2019). In addition, PE has been linked to autophagy pathways in the formation of the autophagosome in eukaryotic cells (Calzada et al. 2016). A study in Aiptasia found that autophagy was upregulated in aposymbiotic anemones compared to symbiotic anemones (Gorman et al. 2025). The authors suggested that the aposymbiotic state is sub-optimal and that the elevated levels of autophagy are a stress response to starvation. Therefore, the increase in PE-O compounds in our aposymbiotic samples could be an indicator of higher cellular turnover compared to symbiotic animals. Overall, our study shows that PE compounds are a major component of cellular physiology irrespective of symbiotic state and life stage. The upregulation of the phosphatidylcholine group (PCs) only in pedal lacerates in this study contradicts other studies on lipid composition in symbiotic cnidarians. PC lipids are primarily structural and are one of the chief components for cell membranes, assisting in membrane fluidity (Sikorskaya 2023). Therefore, adults would also be expected to have a high levels of PCs. Upregulation of PCs in lacerates could be due to the developmental progression and growth of the lacerates. Lacerates for this study were generated one week prior to sample processing, to allow for lacerates and larvae, which required a few days post-fertilization to develop a mouth to acquire symbionts, to be similar in age. During this time, lacerate development progressed past wound healing and mesentery formation to early stages of tentacle growth (Presnell et al. 2022). These processes involved the rapid remodeling and turnover of host tissue and production of new cell membranes in lacerates, which could explain the increase in PC compounds compared to adults. In conclusion, lipids play central roles in the varied life history stages of Aiptasia, as a food source, signaling molecules, and building blocks of cell membranes. Our results show that both symbiosis and life stage are major influences on Aiptasia lipidomic profiles, and that pedal lacerates have a lipid signature that is distinct from the adult parent accentuating their distinct life history stage. Declarations Acknowledgements We are grateful to lab members (Olivia Burleigh, Jun Cai, Maria Ruggeri) and the anemone husbandry staff of the Weis lab for feedback and assistance during the execution of our experiment. We are also grateful to the Burke and Blouin Labs in the Department of Integrative Biology for access to equipment for the extraction process, as well as the Mass Spectrometry Center at Oregon State University for the preparation and running of samples for LC-MS. Funding : This research was made possible through funding provided by the National Science Foundation (IOS: 2124119 to Virginia Weis), as well as the Integrative Biology Research Award (IBRA; to Erick White) and the Launching Undergraduate Research Experiences (LURE; to Jenna Scott) award at Oregon State University designed to promote undergraduate and graduate student research. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Conflict of interest : The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Author contributions : Erick White and Virginia Weis contributed to the conception and design of the study. Material preparation and data collection were performed by Erick White, Aster Parkin, and Jenna Scott. Lipidomics data was performed by Luke Marney. Statistical analysis was performed by Erick White and Mark Phillips. The first draft of the manuscript was written by Erick White and Virginia Weis commented and made edits on previous versions of the manuscript. All authors read and approved the final manuscript. Data availability The dataset generated and analyzed for the current study as well as the supplemental tables are available in the Zenodo depository as “Data_of_MABI_manuscript” (https://doi.org/10.5281/zenodo.18853484). Ethics approval All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. References Baird AH, Bhagoolli R, Ralph PJ, Takahashi S (2009a) Coral bleaching: The role of the host. Trends Ecol Evol 24:16–20. https://doi.org/10.1016/j.tree.2008.09.005 Baird AH, Guest JR, Willis BL (2009b) Systematic and biogeographical patterns in the reproductive biology of scleractinian corals. Annu Rev Ecol Evol Syst 40:551–571. https://doi.org/10.1146/annurev.ecolsys.110308.120220 Baumann JH, Bove CB, Carne L, Gutierrez I, Castillo KD (2021) Two offshore coral species show greater acclimatization capacity to environmental variation than nearshore counterparts in southern Belize. Coral Reefs 40:1181-1194. https://doi.org/10.1007/s00338. Bedgood SA, Plichon K, Weis VM (2024) Symbiosis collapses during development of asexual offspring in the absence of heterotrophic feeding in a model cnidarian-algal symbiosis. bioRxiv 2024.11.15.623823. https://doi.org/10.1101/2024.11.15.623823 Bradford MM (1976) A rapid and sensitive method for the quantita- tion of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72(1–2):248–254. Brusca RC, Brusca GJ (2003) in: Invertebrates (2 nd ed) Sinauer Associates, pp. 253–260. Calzada E, Onguka O, Claypool SM (2016). Phosphatidylethanolamine Metabolism in Health and Disease. Int Rev Cell Mol Biol 321:29-88. doi: 10.1016/bs.ircmb.2015.10.001. Caruso C, Rocha de Souza M, Ruiz-Jones L, Conetta D, Hancock J, Hobbs C, Hobbs C, Kahkejian V, Kitchen R, Marin C, Monismith S, Madin J, Gates R, Drury C (2022) Genetic patterns in Montipora capitata across an environmental mosaic in Kāne’ohe Bay, O’ahu, Hawai’i. Mol Ecol 31:5201–5213. https://doi.org/10.1111/mec.16655. Cary LR (1911) A study ofpedal laceration in actinians. Biol Bull 20:81–108. doi: 10.2307/1536038. Chan WY, Rudd D, van Oppen MJ (2023) Spatial metabolomics for symbiotic marine invertebrates. Life Sci alliance 6:1–15. https://doi.org/10.26508/lsa.202301900. Cui G, Liew YJ, Konciute MK, Zhan Y, Hung S-H, Thistle J, Gastoldi L, Schmidt-Roach S, Dekker J, Aranda M (2022). Nutritional control regulates symbiont proliferation and life history in coral-dinoflagellate symbiosis. BMC Biol 20:103. https://doi.org/10.1186/s12915-022-01306-2. Elder H, Million WC, Bartels E, Krediet CJ, Muller EM, Kenkel CD (2022) Long-term maintenance of a heterologous symbiont association in Acropora palmata on natural reefs. ISME J 17:486-489. https://doi.org/10.1038/s41396-022-01349-x. Falkowski P, Dubinsky Z, Muscatine L, McCloskey L (1993) Population control in symbiotic corals—Ammonium ions and organic materials maintain the density of zooxanthellae. Bioscience 43:606–611. https://doi.org/10.2307/1312147 Falkowski P, Dubinsky Z, Muscatine L, Porter JW (1984) Light and the bioenergetics of a symbiotic coral. Bioscience 34:705–709. https://doi.org/10.2307/1309663 Folch J, Lees M, Sloane-Stanley G (1956) A simple method for isolation and purification of total lipids from animal tissue. J Biol Chem 226:497–509. doi: 10.1016/S0021-9258(18)64849-5 Garrett TA, Schmeitzel JL, Klein JA, Hwang JJ, Schwarz JA (2013) Comparative Lipid Profiling of the Cnidarian Aiptasia pallida and Its Dinoflagellate Symbiont. PLoS One 8:e57975. https://doi.org/10.1371/JOURNAL.PONE.0057975. Gorman LM, Tivey TR, Raymond EH, Ashley IA, Oakley CA, Grossman AR, Weis VM, Davy SK (2025) Stability of the cnidarian-dinoflagellate dymbiosis is primarily determined by symbiont cell-cycle arrest. PNAS 122:1–10. https://doi.org/10.1073/pnas. Grawunder D, Hambleton EA, Bucher M, Wolfowicz I, Bechtoldt N, Guse A (2015) Induction of Gametogenesis in the Cnidarian Endosymbiosis Model Aiptasia sp. Sci Rep 5:1–11. https://doi.org/10.1038/srep15677. Hambleton EA, Arnold V, Jones S, Maegele I, Kvaskoff D, Sachsenheimer T, Guse A (2019) Sterol transfer by atypical cholesterol-binding NPC2 proteins in coral-algal symbiosis. eLife 8:e43923. https://doi.org/10.7554/eLife.43923.001. Hamoutene D, Puestow T, Miller-Banoub J, Wareham V (2008) Main lipid classes in some species of deep-sea corals in the Newfoundland and Labrador region (Northwest Atlantic Ocean). Coral Reefs 27:237–246. https://doi.org/10.1007/s00338-007-0318-7. Harii S, Nadaoka K, Yamamoto M, Iwao K (2007) Temporal changes in settlement, lipid content and lipid composition of larvae of the spawning hermatypic coral Acropora tenuis. Mar Ecol Prog Ser 346:89–96. https://doi.org/10.3354/meps07114. Harrison PL (2011) Sexual Reproduction of Scleractinian Corals, in: Coral Reefs: An Ecosystem in Transition, pp. 59–85. Hartmann AC, Marhaver KL, Klueter A, Lovci MT, Closek CJ, Diaz E, Chamberland VF, Archer FI, Deheyn DD, Vermeij MJA, Medina M (2019) Acquisition of obligate mutualist symbionts during the larval stage is not beneficial for a coral host. Mol Ecol 28:141–155. https://doi.org/10.1111/mec.14967. Huffmyer AS, Wong KH, Becker DM, Strand E, Mass T, Putnam HM (2025) Shifts and critical periods in coral metabolism reveal energetic vulnerability during development. Curr Biol 35:2858–2871. doi: 10.1016/j.cub.2025.05.013. Imbs AB, Latyshev NA, Dautova TN, Latypov YY (2010) Distribution of lipids and fatty acids in corals by their taxonomic position and presence of zooxanthellae. Mar Ecol Prog Ser 409:65–75. https://doi.org/10.3354/meps08622. Imbs AB (2013) Fatty acids and other lipids of corals: Composition, distribution, and biosynthesis. Russ J Mar Biol 39:153–168. https://doi.org/10.1134/S1063074013030061. Imbs AB, Ermolenko EV, Grigorchuk VP, Sikorskaya TV, Velansky PV (2021) Current progress in lipidomics of marine invertebrates. Mar Drugs 19: 1–18. https://doi.org/10.3390/md19120660. Kitchen SA, Poole AZ, Weis VM (2017) Sphingolipid Metabolism of a Sea Anemone Is Altered by the Presence of Dinoflagellate Symbionts. Biol Bull 233:242–254. doi: 10.1086/695846. LaJeunesse TC, Parkinson JE, Gabrielson PW, Jeong HJ, Reimer JD, Voolstra CR, Santos SR (2018) Systematic Revision of Symbiodiniaceae Highlights the Antiquity and Diversity of Coral Endosymbionts. Curr Biol 28:2570-2580.e6. https://doi.org/10.1016/J.CUB.2018.07.008. Lehnert EM, Mouchka ME, Burriesci MS, Gallo ND, Schwarz JA, Pringle JR (2014) Extensive differences in gene expression between symbiotic and aposymbiotic cnidarians. G3 Genes, Genomes, Genet 4:277–295. https://doi.org/10.1534/g3.113.009084 Magnusson CD, Haraldsson GG (2011) Ether lipids. Chem Phys Lipids 164:315–340. Marlow HQ, Martindale MQ (2007). Embryonic development in two species of scleractinian coral embryos: Symbiodinium localization and mode of gastrulation. Evol Dev 9:355–367. https://doi.org/10.1111/j.1525-142X.2007.00173.x. Matthews JL, Sproles AE, Oakley CA, Grossman AR, Weis VM, Davy SK (2016). Menthol-induced bleaching rapidly and effectively provides experimental aposymbiotic sea anemones (Aiptasia sp.) for symbiosis investigations. J Exp Biol 219:306-310. https://doi.org/10.1242/jeb.128934. Matthews JL, Crowder CM, Oakley CA, Lutz A, Roessner U, Meyer E, Grossman AR, Weis VM, Davy SK (2017). Optimal nutrient exchange and immune responses operate in partner specificity in the cnidarian-dinoflagellate sym-biosis. PNAS 114:13194-13199. https://doi.org/10.1073/pnas.1710733114. Muscatine L, Falkowski PG, Porter JW, Dubinsky Z (1984) Fate of photosynthetic fixed carbon in light-and shade-adapted colonies of the symbiotic coral Stylophora pistillata. Proc R Soc Land B 222:181-202. https://doi.org/10.1098/rspb.1984.0058 Muscatine L, Porter JW (1977) Reef corals: mutualistic symbioses adapted to nutrient-poor environments. Bioscience 27:454–460. https://doi.org/10.2307/1297526 Pang Z, Lu Y, Zhou G, Hui F, Xu L, Viau C, Spigelman AF, MacDonald PE, Wishart DS, Li S, Xia J (2024). MetaboAnalyst 6.0: towards a unified platform for metabolomics data processing, analysis and interpretation. Nucleic Acids Res 52:W398-W406. https://doi.org/10.1093/nar/gkae253 Presnell JS, Wirsching E, Weis VM (2022) Tentacle patterning during Exaiptasia diaphana pedal lacerate development differs between symbiotic and aposymbiotic animals. PeerJ 10:e12770. https://doi.org/10.7717/peerj.12770. Reitzel A, Stefanik D, Finnerty JR (2011) Asexual reproduction in Cnidaria: comparative developmental processes and candidate mechanisms. In: Mechanisms of Life History Evolution: The Genetics and Physiology of Life History Traits and Tradeoffs, pp. 101–113. Roberty S, Weis VM, Davy SK, Voolstra CR (2024). Editorial: Aiptasia: a model system in coral symbiosis research. Front Mar Sci 11:1370814. doi: 10.3389/fmars.2024.1370814. Roper CD, Matthews JL, Camp EF, Padula MP, Kuzhiumparambil U, Edmondson J, Howlett L, Suggett DJ (2024) Lipid composition of coral propagules and reproductive material in coral restoration nurseries. Coral Reefs 43:1483–1496. https://doi.org/10.1007/s00338-024-02553-1. Rosset SL, Oakley CA, Ferrier-Pagès C, Suggett DJ, Weis VM, Davy SK (2021) The Molecular Language of the Cnidarian–Dinoflagellate Symbiosis. Trends Microbiol 29. https://doi.org/10.1016/j.tim.2020.08.005. Sikorskaya TV (2023) Coral Lipidome: Molecular Species of Phospholipids, Glycolipids, Betaine Lipids, and Sphingophosphonolipids. Mar Drugs 21:335. doi: 10.3390/md21060335. Sikorskaya TV, Ermolenko EV, Ginanova TT, Boroda AV, Efimova KV, Bogdanov M (2024) Membrane vectorial lipidomic features of coral host cells’ plasma membrane and lipid profiles of their endosymbionts Cladocopium. Commun Biol 7:1–13. https://doi.org/10.1038/s42003-024-06578-8. Sikorskaya TV, Ginanova TT, Ermolenko, EV, Boroda AV (2025). Lipidomic and physiological changes in the coral Acropora aspera during bleaching and recovery. Sci Rep 15:1–16. https://doi.org/10.1038/s41598-025-90484-4 1. Simona F, Zhang H, Voolstra CR (2019). Evidence for a role of protein phosphorylation in the maintenance of the cnidarian–algal symbiosis. Mol Ecol mec15298. https://doi.org/10.1111/mec.15298. Steinmetz PRH, Aman A, Kraus JEM, Technau U (2017). Gut-like ectodermal tissue in a sea anemone challenges germ layer homology. Nat Ecol Evol 1:1535–1542. doi: 10.1038/s41559-017-0285-5. Thorson G (1950). Reproductive and larval ecology of marine bottom invertebrates. Biol Rev Camb Philos Soc 25:1-45. doi: 10.1111/j.1469-185x.1950.tb00585.x. PMID: 24537188. Tivey TR (2019) Cellular Recognition, Division, and Proliferation In the Cnidarian-Dinoflagellate Symbiosis. Dissertation, Oregon State University. Valadez-Ingersoll M, Rugerio MIM, Gilmore TD, Rivera HE, Kanke MR, Gomez-campo K, Metz S, Sweet M, Kwan J, Hekman R, Emili A, Davies SW (2025) Cell type-specific immune regulation under symbiosis in a facultatively symbiotic coral. ISME J 19: wraf132. doi: 10.1093/ismejo/wraf132. Voss PA, Gornik SG, Jacobovitz MR, Rupp S, Dörr M, Maegele I, Guse A (2023) Host nutrient sensing is mediated by mTOR signaling in cnidarian-dinoflagellate symbiosis. Curr Biol 33:3634-3647. https://doi.org/10.1016/j.cub.2023.07.038. Weis VM, Davy SK, Hoegh-Guldberg O, Rodriguez-Lanetty M, Pringle JR (2008) Cell biology in model systems as the key to understanding corals. Trends Ecol Evol 23:369–376. https://doi.org/10.1016/j.tree.2008.03.004. White E, Ruggeri M, Weis VM. 2026. Heterotrophy and symbiosis affect energy reserves for pedal lacerates in the sea anemone Exaiptasia diaphana. PeerJ 14:e20851 https://doi.org/10.7717/peerj.20851 Wolfowicz I, Baumgarten S, Voss PA, Hambleton EA, Voolstra CR, Hatta M, Guse A (2016) Aiptasia sp. larvae as a model to reveal mechanisms of symbiont selection in cnidarians. Sci Rep 6:32366. https://doi.org/10.1038/srep32366 Xiang T, Hambleton EA, DeNofrio JC, Pringle JR, Grossman AR (2013) Isolation of clonal axenic strains of the symbiotic dinoflagellate Symbiodinium and their growth and host specificity. J Phycol 49:447–458. https://doi.org/10.1111/jpy.12055. Yamashiro H, Oku H, Onaga K (2005) Effect of bleaching on lipid content and composition of Okinawan corals. Fish Sci 71:448–453. https://doi.org/10.1111/j.1444-2906.2005.00983.x. Yellowlees D, Rees TAV, Leggat W (2008). Metabolic interactions between algal symbionts and invertebrate hosts. Plant, Cell Environ 31:679–694. https://doi.org/10.1111/j.1365-3040.2008.01802.x. Supplementary Files MABIFigS1.docx MABIFigS2.docx MarineBiologySupplementalTables.csv Cite Share Download PDF Status: Under Review Version 1 posted Reviewers agreed at journal 16 Apr, 2026 Reviewers invited by journal 09 Mar, 2026 Editor assigned by journal 04 Mar, 2026 First submitted to journal 03 Mar, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9023951","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":602882597,"identity":"c48db90e-32dd-46bc-9141-1ab2b7673ec8","order_by":0,"name":"Erick White","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABAElEQVRIiWNgGAWjYDACCcYGIFkjhybMhldLI1DPMWM4H6IarxYGkDXMiQ1EazG43dz+4EMFW3q/9OHDLz7uqKvjn998gOFD2WHcWu4cbGyccUYmd2ZfWprlzDOHJSSOsSUwzjiHW4vZjcTGZt42ttwNZ3jMjHnbDkgwHOMxYOZtI6Dl7z/mdIMz/N+AWuok5I/xf2D+S0gLYwNzgsEZHubHvG3MEgbHeBiYGfFosQdqmdlz7JjhzB42M8aZbYclNx5LMzjYcy4dpxbJGekPPvyoqZHnB1ry4WNbHb/c4cMPH/wos8apBRmwScBYB4hSDwTMH4hVOQpGwSgYBSMLAAAFIFqvN7WLFwAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0003-2029-3149","institution":"Oregon State University","correspondingAuthor":true,"prefix":"","firstName":"Erick","middleName":"","lastName":"White","suffix":""},{"id":602882598,"identity":"777020fa-ca8b-4737-ac6e-1bb3303e1b18","order_by":1,"name":"Luke Marney","email":"","orcid":"","institution":"Oregon State University","correspondingAuthor":false,"prefix":"","firstName":"Luke","middleName":"","lastName":"Marney","suffix":""},{"id":602882599,"identity":"46cc158f-7bef-4cc0-a836-d5e30207e53c","order_by":2,"name":"Mark Phillips","email":"","orcid":"","institution":"Oregon State University","correspondingAuthor":false,"prefix":"","firstName":"Mark","middleName":"","lastName":"Phillips","suffix":""},{"id":602882600,"identity":"0c50230e-0dad-41d6-91f3-8619a85a4ee4","order_by":3,"name":"Jenna Scott","email":"","orcid":"","institution":"Oregon State University","correspondingAuthor":false,"prefix":"","firstName":"Jenna","middleName":"","lastName":"Scott","suffix":""},{"id":602882601,"identity":"d29425fc-ec95-4159-b4b5-c9cdd4e1342d","order_by":4,"name":"Aster Parkin","email":"","orcid":"","institution":"Oregon State University","correspondingAuthor":false,"prefix":"","firstName":"Aster","middleName":"","lastName":"Parkin","suffix":""},{"id":602882602,"identity":"9bbb4e13-3ed8-4822-9bbb-cac3779aeab2","order_by":5,"name":"Virginia M. Weis","email":"","orcid":"","institution":"Oregon State University","correspondingAuthor":false,"prefix":"","firstName":"Virginia","middleName":"M.","lastName":"Weis","suffix":""}],"badges":[],"createdAt":"2026-03-03 22:08:49","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-9023951/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9023951/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":104496081,"identity":"d4047fe2-a1e1-4ba9-ab3c-bb7aeaa608af","added_by":"auto","created_at":"2026-03-12 12:46:50","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":109179,"visible":true,"origin":"","legend":"\u003cp\u003eDistribution of lipid profiles based on symbiotic state and life stage. Principal component analysis with dots representing lacerates and triangles representing adults. Individual points represent a single replicate within that sample group. Gray-colored shapes are aposymbiotic samples and orange-colored shapes are symbiotic samples. Ellipses are the 95% confidence intervals for each sample group. Figure made in MetaboAnalyst\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/66d82b9fbcb56892481a9777.png"},{"id":104780512,"identity":"19d5d6f4-7fcc-449e-8b90-a9127081aaf9","added_by":"auto","created_at":"2026-03-17 07:53:14","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":63947,"visible":true,"origin":"","legend":"\u003cp\u003eComparison of the total amount of and communal \u003cem\u003evs\u003c/em\u003e shared lipids based on symbiotic state and life stage. A: the total number of lipids present in each sample group (detected in at least two replicates). B: the intersection of lipids unique to (single dot) or shared between (dots connected by lines) each sample group. Figure made in R\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/3cd88d838d0581be360f65b1.png"},{"id":104496082,"identity":"63c15d6b-42d4-4154-9374-6f1f059161c6","added_by":"auto","created_at":"2026-03-12 12:46:50","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":268436,"visible":true,"origin":"","legend":"\u003cp\u003eDifferential abundance of the top 75 significantly different lipids based on symbiotic state and life stage.\u003cstrong\u003e \u003c/strong\u003eColumns are samples grouped by replicates and rows are different lipids detected across sample types. Color scale of the heatmap represents the log\u003csub\u003e2\u003c/sub\u003e-fold change in relative abundance with red and blue colors indicating higher and lower abundance, respectively. Legend abbreviations and colors: AA=aposymbiotic adults (purple); AL=aposymbiotic lacerates (gold); SA=symbiotic adults (teal); SL=symbiotic lacerates (magenta). Figure made in MetaboAnalyst\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/f5c0a36fb78b94b92c78d3a5.png"},{"id":104784354,"identity":"9fabc647-6f38-403c-b735-2d8fdc4f5d78","added_by":"auto","created_at":"2026-03-17 08:06:56","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":808925,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/1cec6fea-2b6d-48dc-9937-5bf2df45f52b.pdf"},{"id":104496085,"identity":"28a65d49-2707-48ce-aae3-3ca01feba111","added_by":"auto","created_at":"2026-03-12 12:46:50","extension":"docx","order_by":7,"title":"","display":"","copyAsset":false,"role":"supplement","size":89711,"visible":true,"origin":"","legend":"","description":"","filename":"MABIFigS1.docx","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/748d3c5ae4540582d30c844c.docx"},{"id":104496084,"identity":"220b556e-a74a-48cc-966e-01f71f93c910","added_by":"auto","created_at":"2026-03-12 12:46:50","extension":"docx","order_by":8,"title":"","display":"","copyAsset":false,"role":"supplement","size":28517,"visible":true,"origin":"","legend":"","description":"","filename":"MABIFigS2.docx","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/49def139bdb3299fa08f595b.docx"},{"id":104780611,"identity":"9ddc6bc3-17e0-431e-a62c-fb81d2568eb5","added_by":"auto","created_at":"2026-03-17 07:53:22","extension":"csv","order_by":9,"title":"","display":"","copyAsset":false,"role":"supplement","size":4187,"visible":true,"origin":"","legend":"","description":"","filename":"MarineBiologySupplementalTables.csv","url":"https://assets-eu.researchsquare.com/files/rs-9023951/v1/c99e2832788ad1bea5365b7a.csv"}],"financialInterests":"","formattedTitle":"\u003cp\u003eAbundance and diversity of lipid compounds in Exaiptasia diaphana is altered as a function of symbiotic state and life stage\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eReef-building coals form the foundation of coral reef ecosystems. Corals are comprised of a mutualistic endosymbiosis between the cnidarian host and photosynthetic dinoflagellate algae (family Symbiodiniaceae) that reside within host gastrodermal cells (Muscatine and Porter \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e1977\u003c/span\u003e; Falkowski et al. \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; LaJeunesse et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Maintenance of the symbiosis is based on a reciprocal exchange of nutrients between the partners. The symbiont supplies photosynthetically-fixed carbon that meets most of the metabolic needs of the host (Muscatine et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Yellowlees et al. \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). In return, the host provides the symbiont with waste products in the form of inorganic carbon and nitrogen, the raw materials for photosynthesis and growth, and a stable habitat with access to sunlight (Falkowski et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e1993\u003c/span\u003e; Cui et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Corals thrive in the oligotrophic waters of the tropics and sub-tropics due in large part to this efficient exchange of nutrients between symbiotic partners.\u003c/p\u003e \u003cp\u003eThe majority (approximately 80%) of coral species acquire symbionts from the environment with each host generation (i.e. horizontal transmission) (Baird et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2009a\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003eb\u003c/span\u003e; Hartmann et al. \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). The remaining species pass their symbionts directly from the host parent to offspring (i.e. vertical transmission). For corals that engage in horizontal transmission, algae are acquired either at the larval or juvenile polyp stage (Schwarz et al. 1999; Elder et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Prior to the onset of symbiosis, aposymbiotic larvae must rely on other sources of nutrition to fuel their metabolic needs. They can acquire nutrients either from heterotrophic feeding (Schwarz et al. 1999; Brusca and Brusca \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2003\u003c/span\u003e) or maternally derived nutrients in the form of lipid droplets (Thorson \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e1950\u003c/span\u003e; Marlow and Martindale \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Studies of the compounds present in coral eggs and larvae across species have revealed that nutritionally-rich lipids, such as wax esters and triacylglycerols, are the predominant metabolites present, and vary in abundance based on larval settlement period and dispersal range (Harii et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Roper et al. \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). In addition, the metabolites produced vary depending on mode of symbiont acquisition (horizontal versus vertical) and developmental stage. Studies of larvae have shown that the presence of symbionts can increase lipid droplets compared to aposymbiotic larvae (Voss et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2023\u003c/span\u003e) and that lipid sources shift from being maternally-derived to symbiont-derived as symbiosis becomes established (Huffmyer et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2025\u003c/span\u003e). Thus, symbiosis is an important factor in determining the lipidomic profiles of symbiotic and aposymbiotic offspring at early life stages.\u003c/p\u003e \u003cp\u003eIn addition to sexual reproduction, many corals and sea anemones engage in asexual reproduction by fission, budding, or pedal laceration. Asexual reproduction results in the formation of large colonies of corals, which in turn form the structure of the reef ecosystem (Reitzel et al. 2013). Since asexual offspring are formed directly from parental tissue, symbionts are vertically transmitted and already present as offspring progress through development. The presence of symbionts could therefore influence metabolism and development at these early life-stages. Indeed, recent studies in the sea anemone \u003cem\u003eExaiptasia diaphana\u003c/em\u003e (from here on referred to as Aiptasia) have shown that the presence or absence of heterotrophic feeding and symbiosis can greatly influence the developmental patterning and survival of pedal lacerates (Presnell et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Bedgood et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). While the lipidome of coral adults and larvae have been described (Garret et al. 2013; Imbs et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Roper et al. \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Huffmyer et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2025\u003c/span\u003e), less attention has been given to the lipidome of asexual offspring and the influence of symbiosis on its composition.\u003c/p\u003e \u003cp\u003eAiptasia is an ideal model system for the study of coral-dinoflagellate symbiosis. Like corals, it harbors species of Symbiodiniaceae in its tissues. However, unlike corals, it reproduces both sexually and asexually in the laboratory and can be cultured in both symbiotic and aposymbiotic states (Grawunder et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Roberty et al. \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). In this study, we used liquid chromatography-mass spectrometry (LC-MS) to characterize the lipidome of Aiptasia from different life stages. We compared lipid profiles from symbiotic and aposymbiotic adults, sexually-produced larvae, and asexually-produced pedal lacerates to evaluate the effects of symbiosis and life stage on lipid composition.\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003e\u003cem\u003eAnimal husbandry\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eSymbiotic and aposymbiotic adults and pedal lacerates were sampled from the H2 clonal line of Aiptasia with its native symbiont \u003cem\u003eBreviolum minutum\u0026nbsp;\u003c/em\u003e(Xiang et al. 2013; LaJeunesse et al. 2018). Aposymbiotic adults were generated via menthol bleaching (Matthews et al. 2016) and maintained for at least three months prior to the start of the experiment. Adults used for adult and pedal lacerate samples were maintained in clear (for symbiotic animals) or black (for aposymbiotic animals) polycarbonate tubs (Cambro) in 300 mL 0.45 \u0026micro;m filtered artificial sea water (FASW, Instant Ocean; 32 ppt) in a Percival incubator set to 25\u0026deg;C on a 12-hour/12-hour light/dark cycle, with a full spectrum irradiance of 10-20 \u0026micro;mol quanta m\u003csup\u003e-2\u003c/sup\u003es\u003csup\u003e-1\u003c/sup\u003e. Adult populations were fed with S.presso\u003cem\u003e\u0026nbsp;\u003c/em\u003e(INVE Aquaculture, Belgium) gut-infused brine shrimp (\u003cem\u003eArtemia nauplii\u003c/em\u003e) three times a week with subsequent cleaning and changing of water prior to the start of the experiment. Symbiotic and aposymbiotic adults were randomly selected and used as individual replicate samples for the experiment (n=3 each).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFor generation of sexually-produced larvae, separate tubs of symbiotic Aiptasia (H2 females, CC7 males, and VWA males) meant for reproductive purposes were kept in a separate Percival incubator set to 29\u0026deg;C on a 12-hour/12-hour light/dark cycle, with a full spectrum irradiance of 10-20 \u0026micro;mol quanta m\u003csup\u003e-2\u003c/sup\u003es\u003csup\u003e-1\u003c/sup\u003e. These populations were fed gut-infused brine shrimp five times a week to attain a mature body size followed by cleaning and water changes with FASW pre-warmed to 29\u0026deg;C. Once adults had reached maturity with visible and viable gonads, reproductive animals were maintained at these conditions for at least three months prior to the experiment. Larvae destined for addition of symbionts were inoculated with \u003cem\u003eBreviolum minutum\u0026nbsp;\u003c/em\u003ealgae (SSB01 strain). \u003cem\u003eB. minutum\u003c/em\u003e was cultured in 250 mL suspension culture flasks (Greiner 658190) in F/2 medium (Bigelow MKf250L) under the same temperature and lighting conditions as non-reproductive adult Aiptasia anemones described above.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eGeneration of pedal lacerates and larvae\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003ePedal lacerates were generated as previously described in White et al. (2026) and in Presnell et al. (2022). A total of 90 symbiotic and aposymbiotic lacerates each (n=180 total) were generated and divided between three replicate 96-well polystyrene plates. Plates were cleaned with cotton swabs and FASW. Lacerates were cut one week prior to the sampling timepoint to ensure larvae and lacerates were approximately the same age when sampled.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTo induce spawning and generate larvae, fluorescent blue light was used in the reproductive animal incubator to simulate the lunar cycle that triggers spawning. This blue light was turned on during the night phase of the 12:12 hr light cycle for five consecutive days. Seven to eight days after the blue light was turned on, gravid adults released eggs and sperm. Eggs were pipetted into six glass dishes. Sperm was first filtered through a 50 \u0026micro;m filter with FASW to remove any tissue debris or symbiont clumps and then decanted into the glass dishes with the eggs to generate H2 x CC7/VWA crosses. Three dishes were designated for later addition of symbionts to generate symbiotic larvae while the other three were untouched, with the larvae left to develop into aposymbiotic larvae. Complete FASW water changes were completed everyday by filtering larvae from each dish through a 50 \u0026micro;m filter and placing larvae back into their dishes with fresh FASW. For larvae designated to become symbiotic, larvae first underwent a water change as described above and added back into glass dishes with minimal FASW on day three post-fertilization. Then, \u003cem\u003eB. minutum\u0026nbsp;\u003c/em\u003ecultures were resuspended in FASW and added to larval dishes to reach a concentration of 1 x 10\u003csup\u003e5\u003c/sup\u003e cells/mL. Incubation of larvae with \u003cem\u003eB. minutum\u003c/em\u003e lasted for 24 hours and underwent a complete water change afterwards. This process was then repeated on day four post-fertilization to ensure symbiont uptake into host tissue with a complete water change on day five post-fertilization.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eLipid isolation\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eAdults, lacerates, and symbiotic larvae were all sampled one week (day seven post-laceration; day seven post-fertilization) after the start of the experiment. Aposymbiotic larvae were sampled at five days post-fertilization to ensure there would be a robust sample size due to a decrease in live and active larvae as more time passed. Animals were sampled and divided into three biological replicates for adults (n=1), larvae (n=300), and lacerates (n=30) in 1.5 mL microfuge tubes.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFor all life stages, host tissue samples were homogenized with a motorized mortar and pestle in 300 \u0026micro;L of extraction buffer (extraction buffer [100 mM 7.33 pH Tris, 10 mM 8.04 pH EDTA, 100 mM NaCl], protease inhibitor [Roche 04693159001], and 1% Triton-X). Glass beads (450 microns) were added to each tube and topped up to 1 mL with extraction buffer and then material was further lysed on a Qiagen Tissue Lyser for one minute at 30 Hz to dissociate any remaining cells and release metabolites into the extract. Samples were centrifuged at 3000 rpm for five minutes to pellet glass beads, host debris, and algae. The resulting extract was aliquoted for lipid extraction (600 \u0026micro;L). Prior to lipid extraction, 6 \u0026micro;L of EquiSPLASH Quantitative Mass Spec Internal Standard (Avanti Research 330731), containing 13 deuterated lipid standards at a concentration of 100 \u0026micro;g/mL each, was added to each tube at a final concentration of 0.5-1 \u0026micro;g/mL. Lipids were extracted using a modified Folch method (Folch et al. 1956) described in Baumann et al. (2021) and frozen at -20\u0026deg;C until further use. Lipid fractions were dried under vacuum and resuspended in 100 \u0026micro;L of chloroform and prepped for LC-MS analysis.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eLipidomic analysis\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eUltra-performance liquid-chromatography was performed using a 1.7 \u0026mu;m particle, 2.1 \u0026times;100 mm, CSH C18 Column (Waters, Milford, MA, United States) coupled to a quadrupole time-of-flight mass spectrometer (SCIEX, ZenoTOF 5600) housed at the Oregon State University Mass Spectrometry center. The SCIEX, ZenoTOF 5600 was operated in information-dependent MS/MS acquisition mode (IDA) with the following parameters: IDA survey accumulation time of 0.1 s, IDA survey TOF mass range of 100-1200 m/z, a IDA survey collision energy of 10 V with no collision energy spread, 30 candidates for data dependent acquisition with a collision energy of 30, without excluding former candidate ions, and scanning a TOF range of 50-1200. A declustering potential of 50 was used throughout the analysis with a Zeno threshold of 20,000. The electrospray ionization source (Turbo Ion Spray) was operated at 5500 V with the following parameters: curtain gas of 35. CAD gas of 7, Ion source gas 1 of 50 psi, Ion source gas 2 of 40 psi, and temperature of 500\u0026deg;C. The chemical composition of lipid compounds determines their ability to be ionized to a cation (positively charged) or an anion (negatively charged) and detected. Therefore, samples were ionized in both positive ion mode by the addition of a proton (H+) to molecules and negative mode by the loss of a proton from molecules to ensure the greatest number of compounds were identified by their mass-to-charge ratio (m/z+1) within a sample. For positive ion mode, the mobile phases consisted of (A) 60:40 (v/v) acetonitrile: water with ammonium formate (10 mM) and formic acid (0.1%) and (B) 90:10 (v/v) isopropanol: acetonitrile with ammonium formate (10 mM) and formic acid (0.1% formic acid). For analyses run in the negative ion mode, ammonium acetate (10 mM) was used as the modifier. The chromatographic gradient has been described previously (Cajka and Fiehn, doi: 10.1007/978-1-4939-6996-8_14). Annotation of metabolite signals was completed in MSDial (5.1.230517) using the LipidMaps database.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eStatistical analysis\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eStatistical analysis of the relative abundance of detected lipids across life stages was conducted using MetaboAnalyst (version 6; Pang et al. 2024). Relative abundances from each sample were normalized to the peak areas of the internal standard values of 13 deuterated lipids (EquiSPLASH A83731, Avanti Research) and host protein content (Bradford 1976) prior to analysis in MetaboAnalyst. Normalized data were then subjected to log\u003csub\u003e2\u003c/sub\u003e transformation and mean-centering and pareto scaling, resulting in a clear relation between rank of metabolites and relative abundance observed. Data normality was visually confirmed. Lipidomic profiles between all treatment groups were compared using principal component analysis (PCA) from Euclidean distances and statistically analyzed via permutational multivariate analysis of variance (PERMANOVA) based on 999 permutations. Due to a recovery of protein below the limits of detection of the Bradford assay, larval data were removed from the main dataset and analyzed separately without indexing to protein but still underwent log\u003csub\u003e2\u003c/sub\u003e transformation and mean center-pareto scaling (Supplemental Figures and Tables). A one-way ANOVA with an FDR correction was used to test for significant differences in lipid abundance among adult and lacerate test groups. The top 75 most significantly different compounds between adult and lacerate groups were visualized with a heatmap using the Euclidean distance and Ward clustering algorithm. UpSet plots were used to illustrate intersections of lipids specific to each sample type using the \u003cem\u003eUpSetR\u0026nbsp;\u003c/em\u003epackage in RStudio version 4.0.2. M/z+1 values were deemed relevant if present in at least two replicates. Log\u003csub\u003e2\u003c/sub\u003e-fold changes between aposymbiotic and symbiotic larvae were visualized using a volcano plot with the \u003cem\u003eggplot2\u0026nbsp;\u003c/em\u003epackage in R. Statistical significance for all tests was set with an FDR correction\u003cem\u003e\u0026nbsp;p\u003c/em\u003e\u0026lt;0.05.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eA total of 220 lipids were detected and assigned putative identities across all sample groups (Table S1). The lipidomes of adults and lacerates formed separate clusters with no overlap based on both symbiotic state and life stage (Figure 1; PERMANOVA, F=170.82, R\u003csup\u003e2\u003c/sup\u003e=0.98463, FDR P=0.001). Symbiotic state (PC1) accounted for 58.7% of the variance detected in lipid profiles between samples, while life stage (PC2) accounted for 18.5% of the variance between samples. A total of 164 of the 220 lipids that were detected between lacerates and adults were significantly different based on life stage, symbiotic state, or both (Table S1; Tukey\u0026rsquo;s HSD, FDR P\u0026lt; 0.05). Larval samples were also separated and clustered based on symbiotic state, though this was not statistically significant (Figure S1; PERMANOVA, F-value=25.883, R\u003csup\u003e2\u003c/sup\u003e=0.86615, FDR P=0.1).\u003c/p\u003e\n\u003cp\u003eWe observed differences in the total number of shared and unique lipids detected across sample groups based on symbiotic state and life stage (Figure 2). Symbiotic sample groups had more lipids present than their aposymbiotic counterparts, measured across both adults and pedal lacerates. Within symbiotic states, adult replicates had more lipids detected (sym=172; apo=112) followed by lacerates (sym=135; apo=81) (Figure 2A). A total of 71 lipids were shared across all symbiotic states and life stages. There were 42 lipids that were unique to symbiotic samples, 26 unique to symbiotic adults, and only one that was unique to symbiotic lacerates. There were four lipids that were unique to aposymbiotic samples at both life stages, four lipids that were only present in aposymbiotic adults, while aposymbiotic lacerates had no unique lipids. Adults had six lipids that were unique to that life stage, while lacerates had no unique lipids corresponding to that life stage (Figure 2B).\u003c/p\u003e\n\u003cp\u003eA heatmap of the top 75 significantly different lipid compounds across sample groups clearly depicts differences across symbiotic states and life stages (Figure 3; One-way ANOVA, FDR P\u0026lt;0.05). Symbiotic treatment groups had higher concentrations of nearly all ceramide and triglyceride compounds detected. Aposymbiotic treatment groups had higher concentrations of a subset of ether-linked phosphatidylethanolamine (PE-O) compounds, while a different subset of PE-O compounds was significantly upregulated in adult groups. All phosphatidylcholines (PCs) detected were significantly upregulated in lacerate sample groups only. Lastly, there were eight compounds, mostly ceramides, that were unique to the symbiotic adult sample group alone. For larvae, a total of 43 out of 80 lipids were significantly different between symbiotic and aposymbiotic samples, which was comprised of entirely phospholipids (PCs and PE O-s); only one of the significantly different lipids detected in larval samples was upregulated in aposymbiotic larvae (PC 27.0; Figure S2 and Table S2; Student\u0026rsquo;s T-test, FDR P\u0026lt;0.05).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003e\u003cem\u003eSymbiosis alters the lipidome in Aiptasia\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eNumerous studies have shown that symbiotic state exerts a major influence on symbiotic cnidarians at the genomic (Matthews et al. 2017), transcriptomic (Lehnert et al. 2014), and proteomic (Valadez-Ingersoll et al. 2025) level. Our study extends these findings by showing that the presence of symbionts changes the lipidomic profiles of Aiptasia (Figure 1) in relation to the total amounts and types of lipids detected, as well as their abundances at early (pedal lacerate) and mature (adult) life stages (Figures 2-3). The higher number of lipids in symbiotic compared to aposymbiotic animals illustrates the dominant effect that symbiosis has on the lipidomic profiles in Aiptasia. Differences in the lipidomes of adult Aiptasia as a function of symbiotic state have been described. Garrett et al. (2013) found that non-polar lipids (\u003cem\u003ei.e\u003c/em\u003e. energy-rich compounds and shorter chain length phospholipids) were higher in symbiotic compared to aposymbiotic Aiptasia. The authors hypothesized that symbiosis leads to a remodeling of the chain lengths in phospholipids, and that the released fatty acids are used as fuel for the synthesis of other lipids. This points to the high enery demand required to maintain symbiosis. Hambleton et al. (2019) expanded on this research and found that the abundances of different sterol compounds in Aiptasia are not only influenced by symbiotic state, but also by host genetic line, species of algae present, and heterotrophic feeding. In addition, lipids can have specific spatial distribution between symbiotic states that is involved in regulation of symbionts, as seen with ceramides being localized in the gastrodermis around symbiont-containing host cells and symbiont-derived betaine lipids concentrated in light-exposed tentacles (Chan et al. 2023).\u003c/p\u003e\n\u003cp\u003eThe lack of differences between symbiotic and aposymbiotic larvae suggests that, within the first week of symbiont incorporation into host tissue, the presence of symbionts does not drive a major change in the types and amounts of lipids detected in larvae. The low algal density in larvae and young age of larvae at the sampling time could explain these results, and as symbionts proliferate in larvae, lipid profiles between symbiotic and aposymbiotic larvae could begin to differentiate. Cnidarian larvae are lecithotrophic, which means that they are provisioned with maternally-derived nutrients, typically in the form of lipids, that are used to fuel development and growth until symbionts are acquired from the environment (Marlow and Martindale 2007; Huffmyer et al. 2025) or the larvae begin to feed. Expression profiling of Aiptasia larvae has shown over 300 differentially expressed genes between aposymbiotic and symbiotic larvae, many of which are hypothesized to be involved in symbiosis establishment (Wolfowicz et al. 2016). Together with our data, this suggests that during the early days post-fertilization and post-colonization when the algal density remains low, symbiotic larvae are using maternally-derived lipids to fuel their energetic needs while prioritizing development and symbiosis establishment, thereby resulting in a lipidomic profile that is similar to aposymbiotic larvae. Future studies could conduct a time-series experiment examining lipidomic profiles of larvae sampled through time post-fertilization and post-inoculation to determine how lipid profiles change as larvae transition from being aposymbiotic to symbiotic and as they develop into juvenile polyps.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePedal lacerates are distinct from adult Aiptasia\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe separation of lipidomic profiles and abundances by life stage in Aiptasia in this study suggests that lipidomic profiles are altered in an age-dependent manner. For symbiotic Aiptasia, this could be due to the difference in symbiont density between life stages and/or the symbiosis being at different stages of establishment. The symbiosis is fully established in adults, while it is becoming re-established with vertically-transmitted symbionts in lacerates. Studies in \u003cem\u003eMontipora capitata\u0026nbsp;\u003c/em\u003elarvae showed that there are metabolic changes as larvae develop, switching from a maternally-derived nutrition source to a symbiont-derived one (Huffmyer et al. 2025). Therefore, as symbionts proliferate in lacerates, host metabolism will likely shift to accommodate inter-partner nutrient exchange, and in turn will lead to differential production and usage of lipids within the holobiont. Future studies can examine if lipid profiles in lacerates change based on symbiont density and how profiles change as lacerates progress into mature adults.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe separation of lipidomic profiles by life stage in our study also suggests that, even though pedal lacerates are formed from adult tissue, the process of laceration and clonal propagation alters the lipidomic profile in these organisms. Cnidarians have a variety of life history strategies, even within asexual reproduction alone, that emerge from discrete patterns of developmental patterning (Reitzel et al. 2013; summarized in Caruso et al. 2022). Other studies have shown that Aiptasia pedal lacerates have specific patterns of tentacle formation, development, and growth that are affected by symbiotic state and feeding (Presnell et al. 2022; Bedgood et al. 2024). Taken together with our data, this suggests that pedal lacerates represent a distinct and unique stage in the Aiptasia life cycle and are not simply pieces of adult tissue. Our study highlights the importance of life stage in the cellular makeup of Aiptasia and is something to consider in future studies examining cellular mechanisms of cnidarian symbiosis.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eSymbiosis and life change influence levels of triglycerides, ceramides, and phospholipids\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe differential abundance of different lipid classes across symbiotic states we found in our study illustrates that the interpartner dynamics between symbiotic partners greatly alters the lipidomic landscape. Our study shows that triglycerides and ceramides, heavily associated with symbiosis, are upregulated even at early life stages in pedal lacerates (Yamashiro et al. 2005; Imbs et al. 2010). Triglycerides are a major food source for the host and support host health and metabolism (Hamoutene et al. 2008; Imbs 2013). These storage molecules in pedal lacerates \u0026nbsp;be in the mesenteries, which originate in the parental polyp and are replaced with new mesenteries as lacerates develop (Cary 1911; Steinmetz et al. 2017; Presnell et al. 2022). Ceramides are precursors to the sphingolipids sphingomyelin and glycosphingolipids that are used in the plasma membrane. They also play a pivotal role in cell fate determination where they can be converted into sphingosine, a pro-apoptotic signal, or sphingosine-1-phosphate, a pro-survival signal (Kitchen et al. 2017; Rosset et al. 2021). Ceramides are heavily concentrated in symbiotic Aiptasia compared to aposymbiotic anemones and are primarily located in the gastrodermis (Chan et al. 2023). The authors concluded that ceramide-based sphingolipids are involved in the regulation of symbionts in the cnidarian-dinoflagellate symbiosis, and that symbiosis triggers de-novo synthesis or salvaging of sphingolipids.\u003c/p\u003e\n\u003cp\u003eThe span of upregulated PE-O lipids as a function of symbiotic state and life stage in our study is an indication of the diversity of functions that they play in cellular processes. Ether lipids, such as PE-O, make up the typical classes of cnidarian lipids and are markers for cnidarian host tissue (Imbs et al. 2010; Sikorskaya et al. 2024, 2025). In addition to being a primary component for phospholipid cellular membranes in eukaryotes, PE-O can act as an antioxidant by scavenging reactive oxygen species (ROS), which are produced as a byproduct of cellular respiration and photosynthesis (Magnusson and Haraldsso 2011; Sikorskaya et al., 2025). The high levels of PE-Os in symbiotic animals could be indicative of the use of PE-O for protective measures against the photosynthetic activity of the algae, although future studies are needed to confirm this hypothesis. PE-O compounds are also known to play a role in membrane fusion and cellular division. A study in Aiptasia found that aposymbiotic anemones had a higher cell density and more dividing cells in both the epidermis and gastrodermis than did symbiotic anemones (Tivey 2019). In addition, PE has been linked to autophagy pathways in the formation of the autophagosome in eukaryotic cells (Calzada et al. 2016). A study in Aiptasia found that autophagy was upregulated in aposymbiotic anemones compared to symbiotic anemones (Gorman et al. 2025). The authors suggested that the aposymbiotic state is sub-optimal and that the elevated levels of autophagy are a stress response to starvation. Therefore, the increase in PE-O compounds in our aposymbiotic samples could be an indicator of higher cellular turnover compared to symbiotic animals. Overall, our study shows that PE compounds are a major component of cellular physiology irrespective of symbiotic state and life stage.\u003c/p\u003e\n\u003cp\u003eThe upregulation of the phosphatidylcholine group (PCs) only in pedal lacerates in this study contradicts other studies on lipid composition in symbiotic cnidarians. PC lipids are primarily structural and are one of the chief components for cell membranes, assisting in membrane fluidity (Sikorskaya 2023). Therefore, adults would also be expected to have a high levels of PCs. Upregulation of PCs in lacerates could be due to the developmental progression and growth of the lacerates. Lacerates for this study were generated one week prior to sample processing, to allow for lacerates and larvae, which required a few days post-fertilization to develop a mouth to acquire symbionts, to be similar in age. During this time, lacerate development progressed past wound healing and mesentery formation to early stages of tentacle growth (Presnell et al. 2022). These processes involved the rapid remodeling and turnover of host tissue and production of new cell membranes in lacerates, which could explain the increase in PC compounds compared to adults.\u003c/p\u003e\n\u003cp\u003eIn conclusion, lipids play central roles in the varied life history stages of Aiptasia, as a food source, signaling molecules, and building blocks of cell membranes. Our results show that both symbiosis and life stage are major influences on Aiptasia lipidomic profiles, and that pedal lacerates have a lipid signature that is distinct from the adult parent accentuating their distinct life history stage.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe are grateful to lab members (Olivia Burleigh, Jun Cai, Maria Ruggeri) and the anemone husbandry staff of the Weis lab for feedback and assistance during the execution of our experiment. We are also grateful to the Burke and Blouin Labs in the Department of Integrative Biology for access to equipment for the extraction process, as well as the Mass Spectrometry Center at Oregon State University for the preparation and running of samples for LC-MS.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/em\u003e: This research was made possible through funding provided by the National Science Foundation (IOS: 2124119 to Virginia Weis), as well as the Integrative Biology Research Award (IBRA; to Erick White) and the Launching Undergraduate Research Experiences (LURE; to Jenna Scott) award at Oregon State University designed to promote undergraduate and graduate student research. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u003cstrong\u003eConflict of interest\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e:\u003c/strong\u003e The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e:\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eErick White and Virginia Weis contributed to the conception and design of the study. Material preparation and data collection were performed by Erick White, Aster Parkin, and Jenna Scott. Lipidomics data was performed by Luke Marney. Statistical analysis was performed by Erick White and Mark Phillips. The first draft of the manuscript was written by Erick White and Virginia Weis commented and made edits on previous versions of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eThe dataset generated and analyzed for the current study as well as the supplemental tables are available in the Zenodo depository as \u0026ldquo;Data_of_MABI_manuscript\u0026rdquo; (https://doi.org/10.5281/zenodo.18853484).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u003cstrong\u003eEthics approval\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eAll applicable international, national, and/or institutional guidelines for the care and use of animals were followed.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eBaird AH, Bhagoolli R, Ralph PJ, Takahashi S (2009a) Coral bleaching: The role of the host. Trends Ecol Evol 24:16\u0026ndash;20. https://doi.org/10.1016/j.tree.2008.09.005\u003c/li\u003e\n\u003cli\u003eBaird AH, Guest JR, Willis BL (2009b) Systematic and biogeographical patterns in the reproductive biology of scleractinian corals. Annu Rev Ecol Evol Syst 40:551\u0026ndash;571. https://doi.org/10.1146/annurev.ecolsys.110308.120220\u003c/li\u003e\n\u003cli\u003eBaumann JH, Bove CB, Carne L, Gutierrez I, Castillo KD (2021) Two offshore coral species show greater acclimatization capacity to environmental variation than nearshore counterparts in southern Belize. Coral Reefs 40:1181-1194. https://doi.org/10.1007/s00338.\u003c/li\u003e\n\u003cli\u003eBedgood SA, Plichon K, Weis VM (2024) Symbiosis collapses during development of asexual offspring in the absence of heterotrophic feeding in a model cnidarian-algal symbiosis. bioRxiv 2024.11.15.623823. https://doi.org/10.1101/2024.11.15.623823\u003c/li\u003e\n\u003cli\u003eBradford MM (1976) A rapid and sensitive method for the quantita- tion of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72(1\u0026ndash;2):248\u0026ndash;254.\u003c/li\u003e\n\u003cli\u003eBrusca RC, Brusca GJ (2003) in: Invertebrates (2\u003csup\u003end\u003c/sup\u003e ed) Sinauer Associates, pp. 253\u0026ndash;260.\u003c/li\u003e\n\u003cli\u003eCalzada E, Onguka O, Claypool SM (2016). Phosphatidylethanolamine Metabolism in Health and Disease. Int Rev Cell Mol Biol 321:29-88. doi: 10.1016/bs.ircmb.2015.10.001.\u003c/li\u003e\n\u003cli\u003eCaruso C, Rocha de Souza M, Ruiz-Jones L, Conetta D, Hancock J, Hobbs C, Hobbs C, Kahkejian V, Kitchen R, Marin C, Monismith S, Madin J, Gates R, Drury C (2022) Genetic patterns in Montipora capitata across an environmental mosaic in Kāne\u0026rsquo;ohe Bay, O\u0026rsquo;ahu, Hawai\u0026rsquo;i. Mol Ecol 31:5201\u0026ndash;5213. https://doi.org/10.1111/mec.16655.\u003c/li\u003e\n\u003cli\u003eCary LR (1911) A study ofpedal laceration in actinians. Biol Bull 20:81\u0026ndash;108. doi: 10.2307/1536038.\u003c/li\u003e\n\u003cli\u003eChan WY, Rudd D, van Oppen MJ (2023) Spatial metabolomics for symbiotic marine invertebrates. Life Sci alliance 6:1\u0026ndash;15. https://doi.org/10.26508/lsa.202301900.\u003c/li\u003e\n\u003cli\u003eCui G, Liew YJ, Konciute MK, Zhan Y, Hung S-H, Thistle J, Gastoldi L, Schmidt-Roach S, Dekker J, Aranda M (2022). Nutritional control regulates symbiont proliferation and life history in coral-dinoflagellate symbiosis. BMC Biol 20:103. https://doi.org/10.1186/s12915-022-01306-2.\u003c/li\u003e\n\u003cli\u003eElder H, Million WC, Bartels E, Krediet CJ, Muller EM, Kenkel CD (2022) Long-term maintenance of a heterologous symbiont association in Acropora palmata on natural reefs. ISME J 17:486-489. https://doi.org/10.1038/s41396-022-01349-x.\u003c/li\u003e\n\u003cli\u003eFalkowski P, Dubinsky Z, Muscatine L, McCloskey L (1993) Population control in symbiotic corals\u0026mdash;Ammonium ions and organic materials maintain the density of zooxanthellae. Bioscience 43:606\u0026ndash;611. https://doi.org/10.2307/1312147\u003c/li\u003e\n\u003cli\u003eFalkowski P, Dubinsky Z, Muscatine L, Porter JW (1984) Light and the bioenergetics of a symbiotic coral. Bioscience 34:705\u0026ndash;709. https://doi.org/10.2307/1309663\u003c/li\u003e\n\u003cli\u003eFolch J, Lees M, Sloane-Stanley G (1956) A simple method for isolation and purification of total lipids from animal tissue. J Biol Chem 226:497\u0026ndash;509. doi: 10.1016/S0021-9258(18)64849-5\u003c/li\u003e\n\u003cli\u003eGarrett TA, Schmeitzel JL, Klein JA, Hwang JJ, Schwarz JA (2013) Comparative Lipid Profiling of the Cnidarian Aiptasia pallida and Its Dinoflagellate Symbiont. PLoS One 8:e57975. https://doi.org/10.1371/JOURNAL.PONE.0057975.\u003c/li\u003e\n\u003cli\u003eGorman LM, Tivey TR, Raymond EH, Ashley IA, Oakley CA, Grossman AR, Weis VM, Davy SK (2025) Stability of the cnidarian-dinoflagellate dymbiosis is primarily determined by symbiont cell-cycle arrest. PNAS 122:1\u0026ndash;10. https://doi.org/10.1073/pnas.\u003c/li\u003e\n\u003cli\u003eGrawunder D, Hambleton EA, Bucher M, Wolfowicz I, Bechtoldt N, Guse A (2015) Induction of Gametogenesis in the Cnidarian Endosymbiosis Model Aiptasia sp. Sci Rep 5:1\u0026ndash;11. https://doi.org/10.1038/srep15677.\u003c/li\u003e\n\u003cli\u003eHambleton EA, Arnold V, Jones S, Maegele I, Kvaskoff D, Sachsenheimer T, Guse A (2019) Sterol transfer by atypical cholesterol-binding NPC2 proteins in coral-algal symbiosis. eLife 8:e43923. https://doi.org/10.7554/eLife.43923.001.\u003c/li\u003e\n\u003cli\u003eHamoutene D, Puestow T, Miller-Banoub J, Wareham V (2008) Main lipid classes in some species of deep-sea corals in the Newfoundland and Labrador region (Northwest Atlantic Ocean). Coral Reefs 27:237\u0026ndash;246. https://doi.org/10.1007/s00338-007-0318-7.\u003c/li\u003e\n\u003cli\u003eHarii S, Nadaoka K, Yamamoto M, Iwao K (2007) Temporal changes in settlement, lipid content and lipid composition of larvae of the spawning hermatypic coral Acropora tenuis. Mar Ecol Prog Ser 346:89\u0026ndash;96. https://doi.org/10.3354/meps07114.\u003c/li\u003e\n\u003cli\u003eHarrison PL (2011) Sexual Reproduction of Scleractinian Corals, in: Coral Reefs: An Ecosystem in Transition, pp. 59\u0026ndash;85.\u003c/li\u003e\n\u003cli\u003eHartmann AC, Marhaver KL, Klueter A, Lovci MT, Closek CJ, Diaz E, Chamberland VF, Archer FI, Deheyn DD, Vermeij MJA, Medina M (2019) Acquisition of obligate mutualist symbionts during the larval stage is not beneficial for a coral host. Mol Ecol 28:141\u0026ndash;155. https://doi.org/10.1111/mec.14967.\u003c/li\u003e\n\u003cli\u003eHuffmyer AS, Wong KH, Becker DM, Strand E, Mass T, Putnam HM (2025) Shifts and critical periods in coral metabolism reveal energetic vulnerability during development. Curr Biol 35:2858\u0026ndash;2871. doi: 10.1016/j.cub.2025.05.013.\u003c/li\u003e\n\u003cli\u003eImbs AB, Latyshev NA, Dautova TN, Latypov YY (2010) Distribution of lipids and fatty acids in corals by their taxonomic position and presence of zooxanthellae. Mar Ecol Prog Ser 409:65\u0026ndash;75. https://doi.org/10.3354/meps08622.\u003c/li\u003e\n\u003cli\u003eImbs AB (2013) Fatty acids and other lipids of corals: Composition, distribution, and biosynthesis. Russ J Mar Biol 39:153\u0026ndash;168. https://doi.org/10.1134/S1063074013030061.\u003c/li\u003e\n\u003cli\u003eImbs AB, Ermolenko EV, Grigorchuk VP, Sikorskaya TV, Velansky PV (2021) Current progress in lipidomics of marine invertebrates. Mar Drugs 19: 1\u0026ndash;18. https://doi.org/10.3390/md19120660.\u003c/li\u003e\n\u003cli\u003eKitchen SA, Poole AZ, Weis VM (2017) Sphingolipid Metabolism of a Sea Anemone Is Altered by the Presence of Dinoflagellate Symbionts. Biol Bull 233:242\u0026ndash;254. doi: 10.1086/695846.\u003c/li\u003e\n\u003cli\u003eLaJeunesse TC, Parkinson JE, Gabrielson PW, Jeong HJ, Reimer JD, Voolstra CR, Santos SR (2018) Systematic Revision of Symbiodiniaceae Highlights the Antiquity and Diversity of Coral Endosymbionts. Curr Biol 28:2570-2580.e6. https://doi.org/10.1016/J.CUB.2018.07.008.\u003c/li\u003e\n\u003cli\u003eLehnert EM, Mouchka ME, Burriesci MS, Gallo ND, Schwarz JA, Pringle JR (2014) Extensive differences in gene expression between symbiotic and aposymbiotic cnidarians. G3 Genes, Genomes, Genet 4:277\u0026ndash;295. https://doi.org/10.1534/g3.113.009084\u003c/li\u003e\n\u003cli\u003eMagnusson CD, Haraldsson GG (2011) Ether lipids. Chem Phys Lipids 164:315\u0026ndash;340.\u003c/li\u003e\n\u003cli\u003eMarlow HQ, Martindale MQ (2007). Embryonic development in two species of scleractinian coral embryos: Symbiodinium localization and mode of gastrulation. Evol Dev 9:355\u0026ndash;367. https://doi.org/10.1111/j.1525-142X.2007.00173.x.\u003c/li\u003e\n\u003cli\u003eMatthews JL, Sproles AE, Oakley CA, Grossman AR, Weis VM, Davy SK (2016). Menthol-induced bleaching rapidly and effectively provides experimental aposymbiotic sea anemones (Aiptasia sp.) for symbiosis investigations. J Exp Biol 219:306-310. https://doi.org/10.1242/jeb.128934.\u003c/li\u003e\n\u003cli\u003eMatthews JL, Crowder CM, Oakley CA, Lutz A, Roessner U, Meyer E, Grossman AR, Weis VM, Davy SK (2017). Optimal nutrient exchange and immune responses operate in partner specificity in the cnidarian-dinoflagellate sym-biosis. PNAS 114:13194-13199. https://doi.org/10.1073/pnas.1710733114.\u003c/li\u003e\n\u003cli\u003eMuscatine L, Falkowski PG, Porter JW, Dubinsky Z (1984) Fate of photosynthetic fixed carbon in light-and shade-adapted colonies of the symbiotic coral Stylophora pistillata. Proc R Soc Land B 222:181-202. https://doi.org/10.1098/rspb.1984.0058\u003c/li\u003e\n\u003cli\u003eMuscatine L, Porter JW (1977) Reef corals: mutualistic symbioses adapted to nutrient-poor environments. Bioscience 27:454\u0026ndash;460. https://doi.org/10.2307/1297526\u003c/li\u003e\n\u003cli\u003ePang Z, Lu Y, Zhou G, Hui F, Xu L, Viau C, Spigelman AF, MacDonald PE, Wishart DS, Li S, Xia J (2024). MetaboAnalyst 6.0: towards a unified platform for metabolomics data processing, analysis and interpretation. Nucleic Acids Res 52:W398-W406. https://doi.org/10.1093/nar/gkae253\u003c/li\u003e\n\u003cli\u003ePresnell JS, Wirsching E, Weis VM (2022) Tentacle patterning during Exaiptasia diaphana pedal lacerate development differs between symbiotic and aposymbiotic animals. PeerJ 10:e12770. https://doi.org/10.7717/peerj.12770.\u003c/li\u003e\n\u003cli\u003eReitzel A, Stefanik D, Finnerty JR (2011) Asexual reproduction in Cnidaria: comparative developmental processes and candidate mechanisms. In: Mechanisms of Life History Evolution: The Genetics and Physiology of Life History Traits and Tradeoffs, pp. 101\u0026ndash;113.\u003c/li\u003e\n\u003cli\u003eRoberty S, Weis VM, Davy SK, Voolstra CR (2024). Editorial: Aiptasia: a model system in coral symbiosis research. Front Mar Sci 11:1370814. doi: 10.3389/fmars.2024.1370814.\u003c/li\u003e\n\u003cli\u003eRoper CD, Matthews JL, Camp EF, Padula MP, Kuzhiumparambil U, Edmondson J, Howlett L, Suggett DJ (2024) Lipid composition of coral propagules and reproductive material in coral restoration nurseries. Coral Reefs 43:1483\u0026ndash;1496. https://doi.org/10.1007/s00338-024-02553-1.\u003c/li\u003e\n\u003cli\u003eRosset SL, Oakley CA, Ferrier-Pag\u0026egrave;s C, Suggett DJ, Weis VM, Davy SK (2021) The Molecular Language of the Cnidarian\u0026ndash;Dinoflagellate Symbiosis. Trends Microbiol 29. https://doi.org/10.1016/j.tim.2020.08.005.\u003c/li\u003e\n\u003cli\u003eSikorskaya TV (2023) Coral Lipidome: Molecular Species of Phospholipids, Glycolipids, Betaine Lipids, and Sphingophosphonolipids. Mar Drugs 21:335. doi: 10.3390/md21060335.\u003c/li\u003e\n\u003cli\u003eSikorskaya TV, Ermolenko EV, Ginanova TT, Boroda AV, Efimova KV, Bogdanov M (2024) Membrane vectorial lipidomic features of coral host cells\u0026rsquo; plasma membrane and lipid profiles of their endosymbionts Cladocopium. Commun Biol 7:1\u0026ndash;13. https://doi.org/10.1038/s42003-024-06578-8.\u003c/li\u003e\n\u003cli\u003eSikorskaya TV, Ginanova TT, Ermolenko, EV, Boroda AV (2025). Lipidomic and physiological changes in the coral Acropora aspera during bleaching and recovery. Sci Rep 15:1\u0026ndash;16. https://doi.org/10.1038/s41598-025-90484-4 1.\u003c/li\u003e\n\u003cli\u003eSimona F, Zhang H, Voolstra CR (2019). Evidence for a role of protein phosphorylation in the maintenance of the cnidarian\u0026ndash;algal symbiosis. Mol Ecol mec15298. https://doi.org/10.1111/mec.15298.\u003c/li\u003e\n\u003cli\u003eSteinmetz PRH, Aman A, Kraus JEM, Technau U (2017). Gut-like ectodermal tissue in a sea anemone challenges germ layer homology. Nat Ecol Evol 1:1535\u0026ndash;1542. doi: 10.1038/s41559-017-0285-5.\u003c/li\u003e\n\u003cli\u003eThorson G (1950). Reproductive and larval ecology of marine bottom invertebrates. Biol Rev Camb Philos Soc 25:1-45. doi: 10.1111/j.1469-185x.1950.tb00585.x. PMID: 24537188. \u003c/li\u003e\n\u003cli\u003eTivey TR (2019) Cellular Recognition, Division, and Proliferation In the Cnidarian-Dinoflagellate Symbiosis. Dissertation, Oregon State University.\u003c/li\u003e\n\u003cli\u003eValadez-Ingersoll M, Rugerio MIM, Gilmore TD, Rivera HE, Kanke MR, Gomez-campo K, Metz S, Sweet M, Kwan J, Hekman R, Emili A, Davies SW (2025) Cell type-specific immune regulation under symbiosis in a facultatively symbiotic coral. ISME J 19: wraf132. doi: 10.1093/ismejo/wraf132.\u003c/li\u003e\n\u003cli\u003eVoss PA, Gornik SG, Jacobovitz MR, Rupp S, D\u0026ouml;rr M, Maegele I, Guse A (2023) Host nutrient sensing is mediated by mTOR signaling in cnidarian-dinoflagellate symbiosis. Curr Biol 33:3634-3647. https://doi.org/10.1016/j.cub.2023.07.038.\u003c/li\u003e\n\u003cli\u003eWeis VM, Davy SK, Hoegh-Guldberg O, Rodriguez-Lanetty M, Pringle JR (2008) Cell biology in model systems as the key to understanding corals. Trends Ecol Evol 23:369\u0026ndash;376. https://doi.org/10.1016/j.tree.2008.03.004.\u003c/li\u003e\n\u003cli\u003eWhite E, Ruggeri M, Weis VM. 2026. Heterotrophy and symbiosis affect energy reserves for pedal lacerates in the sea anemone Exaiptasia diaphana. PeerJ 14:e20851 https://doi.org/10.7717/peerj.20851\u003c/li\u003e\n\u003cli\u003eWolfowicz I, Baumgarten S, Voss PA, Hambleton EA, Voolstra CR, Hatta M, Guse A (2016) Aiptasia sp. larvae as a model to reveal mechanisms of symbiont selection in cnidarians. Sci Rep 6:32366. https://doi.org/10.1038/srep32366\u003c/li\u003e\n\u003cli\u003eXiang T, Hambleton EA, DeNofrio JC, Pringle JR, Grossman AR (2013) Isolation of clonal axenic strains of the symbiotic dinoflagellate Symbiodinium and their growth and host specificity. J Phycol 49:447\u0026ndash;458. https://doi.org/10.1111/jpy.12055.\u003c/li\u003e\n\u003cli\u003eYamashiro H, Oku H, Onaga K (2005) Effect of bleaching on lipid content and composition of Okinawan corals. Fish Sci 71:448\u0026ndash;453. https://doi.org/10.1111/j.1444-2906.2005.00983.x.\u003c/li\u003e\n\u003cli\u003eYellowlees D, Rees TAV, Leggat W (2008). Metabolic interactions between algal symbionts and invertebrate hosts. Plant, Cell Environ 31:679\u0026ndash;694. https://doi.org/10.1111/j.1365-3040.2008.01802.x.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"marine-biology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mabi","sideBox":"Learn more about [Marine Biology](https://www.springer.com/journal/227)","snPcode":"227","submissionUrl":"https://submission.nature.com/new-submission/227/3","title":"Marine Biology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Lipidomics, LC-MS, Aiptasia, symbiosis, pedal lacerates, larvae","lastPublishedDoi":"10.21203/rs.3.rs-9023951/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9023951/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eScleractinian corals engage in an endosymbiotic relationship with dinoflagellate algae to meet their nutritional demands in oligotrophic waters. Algae transfer photosynthetically-derived compounds to the host and host is structured to support and sustain the symbionts. The composition and abundance of these compounds as a function of host life stage and symbiotic state have yet to be described. Therefore, we aimed to characterize the effect of symbiosis and host life stage on the lipidomic profiles of the sea anemone \u003cem\u003eExaiptasia diaphana\u003c/em\u003e (commonly referred to as Aiptasia), a model system for the study of coral symbiosis. We sampled aposymbiotic and symbiotic Aiptasia adults, pedal lacerates, and larvae and isolated lipid fractions for liquid-chromatography mass-spectrometry (LC-MS) analysis. Over 200 lipid compounds were identified across all sample groups. There were more compounds in symbiotic than in aposymbiotic at every life stage. Lipidomic profiles separated based on both symbiotic state and life stage. Of the major lipids classes detected, triglycerides and ceramides were more abundant in symbiotic animals, phosphatidylcholines (PCs) were more abundant in pedal lacerates in both symbiotic states, and abundances of phosphatidylethanolamines (PEs) varied based on symbiotic state. Our results highlight the major influence of symbiotic state on lipid profiles in Aiptasia at different life stages and provide insight into how the lipidome changes in early life-stage cnidarians.\u003c/p\u003e","manuscriptTitle":"Abundance and diversity of lipid compounds in Exaiptasia diaphana is altered as a function of symbiotic state and life stage","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-03-12 12:46:46","doi":"10.21203/rs.3.rs-9023951/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewerAgreed","content":"","date":"2026-04-16T21:44:47+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-03-09T06:40:01+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-03-04T13:53:52+00:00","index":"","fulltext":""},{"type":"submitted","content":"Marine Biology","date":"2026-03-03T17:08:10+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"marine-biology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mabi","sideBox":"Learn more about [Marine Biology](https://www.springer.com/journal/227)","snPcode":"227","submissionUrl":"https://submission.nature.com/new-submission/227/3","title":"Marine Biology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"a74b0955-0bde-4d1b-ad5c-951e5124309c","owner":[],"postedDate":"March 12th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-03-12T12:46:46+00:00","versionOfRecord":[],"versionCreatedAt":"2026-03-12 12:46:46","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9023951","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9023951","identity":"rs-9023951","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}