Computational Methods Reveal a Series of Cyclic and Linear Lichenysins and Surfactins from the Vietnamese Marine Sediment-Derived Streptomyces Strain G222.

The Streptomyces strain G222, isolated from a Vietnamese marine sediment, was confidently identified by 16S rRNA gene sequencing. Its AcOEt crude extract was successfully analyzed using non-targeted LC-MS/MS analysis, and molecular networking, leading to a putative annotation of its chemical diversity thanks to spectral libraries from GNPS and in silico metabolite structure prediction obtained from SIRIUS combined with the bioinformatics tool conCISE (Consensus Annotation Propagation of in silico Elucidations). This dereplication strategy allowed the identification of an interesting cluster of a series of putative cyclic and linear lipopeptides of the lichenysin and surfactin families. Lichenysins (3-7) were isolated from the sub-fraction, which showed significant anti-biofilm activity against Pseudomonas aeruginosa MUC-N1. Their structures were confirmed by detailed 1D and 2D NMR spectroscopy (COSY, HSQC, HMBC, TOCSY, ROESY) recorded in CD3OH, and their absolute configurations were determined using the modified Marfey's method. The isolated lichenysins showed anti-biofilm activity at a minimum concentration of 100 µM. When evaluated for antibacterial activity against a panel of Gram-positive and Gram-negative strains, two isolated lichenysins exhibited selective activity against the MRSA strain without affecting its growth curve and without membranotropic activity. This study highlights the power of the MS/MS spectral similarity strategy using computational methods to obtain a cross-validation of the annotated molecules from the complex metabolic profile of a marine sediment-derived Streptomyces extract. This work provides the first report from a Streptomyces strain of combined cyclic and linear lichenysins and surfactins, known to be characteristic compounds of the genus Bacillus.

Antimicrobial Indole-3-Carboxamido-Polyamine Conjugates Target Bacterial Membranes and Are Antibiotic Potentiators.

Small molecules that can restore the action of legacy antibiotics toward drug-resistant bacteria represent an area of ongoing research interest. We have previously reported indole-3-glyoxylamido and indole-3-acetamido-polyamine conjugates that exhibit intrinsic activity toward bacterial and fungal species, and the ability to enhance the action of doxycycline toward the Gram-negative bacteria Pseudomonas aeruginosa; however, these desirable activities were commonly associated with unfavorable cytotoxicity and/or red blood cell hemolytic properties. In this paper, we report the synthesis and biological investigation of a new class of α,ω-di(indole-3-carboxamido)polyamine derivatives, leading to the identification of several analogues that exhibit antimicrobial- and antibiotic-potentiating activities without detectable cytotoxic or hemolytic properties. 5-Bromo-substituted indole analogues 3 and 12-18 were generally more broad-spectrum in their activity than others in the set, with 13b (polyamine PA-3-6-3) being particularly notable for its anti-Staphylococcus aureus, Acinetobacter baumannii, and Cryptococcus neoformans activities (MIC ≤ 0.28 µM). The same analogue also restored the action of doxycycline toward P. aeruginosa with a 21-fold enhancement, while the corresponding 5-bromo-indole-3-carboxamide-PA3-7-3 analogue was able to enhance the action of both doxycycline and erythromycin toward P. aeruginosa and Escherichia coli, respectively. The analogue 13b was capable of disrupting the bacterial membrane of both S. aureus and methicillin-resistant S. aureus (MRSA) and the outer membrane of P. aeruginosa, suggesting that membrane perturbation could be a mechanism of action of both intrinsic antimicrobial activities and antibiotic potentiation.

Genomes of nine biofilm-forming filamentous strains of Cyanobacteria (genera Jaaginema, Scytonema, and Karukerafilum gen. nov.) isolated from mangrove habitats of Guadeloupe (Lesser Antilles).

Biofilm-forming cyanobacteria are abundant in mangrove ecosystems, colonizing various niches including sediment surface and periphyton where they can cover large areas, yet have received limited attention. Several filamentous isolates were recently isolated from Guadeloupe, illustrating the diversity and novelty present in these biofilms. In this study, nine strains belonging to three novel lineages found abundantly in Guadeloupe biofilms are characterized by genome sequencing, morphological and ultrastructural examination, metabolome fingerprinting and searched for secondary metabolites biosynthesis pathways. Assignation of two lineages to known genera is confirmed, namely Scytonema and Jaaginema. The third lineage corresponds to a new Coleofasciculales genus herein described as Karukerafilum gen. nov. The four strains belonging to this genus group into two subclades, one of which displays genes necessary for nitrogen fixation as well as the complete pathway for geosmin production. This study gives new insights into the diversity of mangrove biofilm-forming cyanobacteria, including genome-based description of a new genus and the first genome sequence available for the genus Jaaginema.

Preliminary SAR of Novel Pleuromutilin-Polyamine Conjugates.

While pleuromutilin (1) and its clinically available derivatives (2-6) are highly effective against Gram-positive bacteria, they remain inactive against many pathogenic Gram-negative bacteria due to the efflux pump AcrAB-TolC. In an effort to broaden the spectrum of activity of pleuromutilin (1), we developed a series of novel pleuromutilin-polyamine conjugates (9a-f) which exhibited promising intrinsic antimicrobial properties, targeting both Gram-positive and Gram-negative bacteria, including Staphylococcus aureus, methicillin-resistant S. aureus (MRSA), and Escherichia coli, along with the fungal strain Cryptococcus neoformans, and were devoid of cytotoxic and hemolytic properties with the exception of one conjugate. Furthermore, this series displayed moderate to low antibiotic potentiation of legacy antibiotics doxycycline and erythromycin, with three conjugates enhancing the activity four-fold in combination with doxycycline. In comparison to pleuromutilin (1) and tiamulin (2), one of the conjugates exhibited an expanded spectrum of activity, including Gram-negative bacteria and fungi, making it a promising option for combating microbial infections.

Indole-3-Acetamido-Polyamines as Antimicrobial Agents and Antibiotic Adjuvants.

The widespread incidence of antimicrobial resistance necessitates the discovery of new classes of antimicrobials as well as adjuvant molecules that can restore the action of ineffective antibiotics. Herein, we report the synthesis of a new class of indole-3-acetamido-polyamine conjugates that were evaluated for antimicrobial activities against a panel of bacteria and two fungi, and for the ability to enhance the action of doxycycline against Pseudomonas aeruginosa and erythromycin against Escherichia coli. Compounds 14b, 15b, 17c, 18a, 18b, 18d, 19b, 19e, 20c and 20d exhibited strong growth inhibition of methicillin-resistant Staphylococcus aureus (MRSA) and Cryptococcus neoformans, with minimum inhibitory concentrations (MIC) typically less than 0.2 µM. Four analogues, including a 5-bromo 15c and three 5-methoxyls 16d-f, also exhibited intrinsic activity towards E. coli. Antibiotic kill curve analysis of 15c identified it to be a bactericide. While only one derivative was found to (weakly) enhance the action of erythromycin against E. coli, three examples, including 15c, were found to be strong enhancers of the antibiotic action of doxycycline against P. aeruginosa. Collectively, these results highlight the promising potential of α,ω-disubstituted indole-3-acetamido polyamine conjugates as antimicrobials and antibiotic adjuvants.

Exploration of Bis-Cinnamido-Polyamines as Intrinsic Antimicrobial Agents and Antibiotic Enhancers.

The marine natural product ianthelliformisamine C is a bis-cinnamido substituted spermine derivative that exhibits intrinsic antimicrobial properties and can enhance the action of doxycycline towards the Gram-negative bacterium Pseudomonas aeruginosa. As part of a study to explore the structure-activity requirements of these activities, we have synthesized a set of analogues that vary in the presence/absence of methoxyl group and bromine atoms and in the polyamine chain length. Intrinsic antimicrobial activity towards Staphylococcus aureus, methicillin-resistant S. aureus (MRSA) and the fungus Cryptococcus neoformans was observed for only the longest polyamine chain examples of non-brominated analogues while all examples bearing either one or two bromine atoms were active. Weak to no activity was typically observed towards Gram-negative bacteria, with exceptions being the longest polyamine chain examples 13f, 14f and 16f against Escherichia coli (MIC 1.56, 7.2 and 5.3 µM, respectively). Many of these longer polyamine-chain analogues also exhibited cytotoxic and/or red blood cell hemolytic properties, diminishing their potential as antimicrobial lead compounds. Two of the non-toxic, non-halogenated analogues, 13b and 13d, exhibited a strong ability to enhance the action of doxycycline against P. aeruginosa, with >64-fold and >32-fold enhancement, respectively. These results suggest that any future efforts to optimize the antibiotic-enhancing properties of cinnamido-polyamines should explore a wider range of aromatic ring substituents that do not include bromine or methoxyl groups.

Structure-Activity Relationship Studies of Indolglyoxyl-Polyamine Conjugates as Antimicrobials and Antibiotic Potentiators.

Antibiotic resistance is a growing global health threat, requiring urgent attention. One approach to overcome antibiotic resistance is to discover and develop new antibiotic enhancers, molecules that work with legacy antibiotics to enhance their efficacy against resistant bacteria. Our previous screening of a library of purified marine natural products and their synthetic analogues led to the discovery of an indolglyoxyl-spermine derivative that exhibited intrinsic antimicrobial properties and was also able to potentiate the action of doxycycline towards the difficult to treat, Gram-negative bacterium Pseudomonas aeruginosa. A set of analogues have now been prepared, exploring the influence of indole substitution at the 5- and 7- positions and length of the polyamine chain on biological activity. While limiting cytotoxicity and/or hemolytic activities were observed for many analogues, two 7-methyl substituted analogues (23b and 23c) were found to exhibit strong activity towards Gram-positive bacteria with no detectable cytotoxicity or hemolytic properties. Different molecular attributes were required for antibiotic enhancing properties, with one example identified, a 5-methoxy-substitiuted analogue (19a), as being a non-toxic, non-hemolytic enhancer of the action of two tetracycline antibiotics, doxycycline and minocycline, towards P. aeruginosa. These results provide further stimulation for the search for novel antimicrobials and antibiotic enhancers amongst marine natural products and related synthetic analogues.

Investigation of Naphthyl-Polyamine Conjugates as Antimicrobials and Antibiotic Enhancers.

As part of our search for new antimicrobials and antibiotic enhancers, a series of naphthyl- and biphenyl-substituted polyamine conjugates have been synthesized. The structurally-diverse library of compounds incorporated variation in the capping end groups and in the length of the polyamine (PA) core. Longer chain (PA-3-12-3) variants containing both 1-naphthyl and 2-naphthyl capping groups exhibited more pronounced intrinsic antimicrobial properties against methicillin-resistant Staphylococcus aureus (MRSA) (MIC ≤ 0.29 µM) and the fungus Cryptococcus neoformans (MIC ≤ 0.29 µM). Closer mechanistic study of one of these analogues, 20f, identified it as a bactericide. In contrast to previously reported diarylacyl-substituted polyamines, several examples in the current set were able to enhance the antibiotic action of doxycycline and/or erythromycin towards the Gram-negative bacteria Pseudomonas aeruginosa and Escherichia coli. Two analogues (19a and 20c) were of note, exhibiting greater than 32-fold enhancement in activity. This latter result suggests that α,ω-disubstituted polyamines bearing 1-naphthyl- and 2-naphthyl-capping groups are worthy of further investigation and optimization as non-toxic antibiotic enhancers.

α,ω-Diacyl-Substituted Analogues of Natural and Unnatural Polyamines: Identification of Potent Bactericides That Selectively Target Bacterial Membranes.

In this study, α-ω-disubstituted polyamines exhibit a range of potentially useful biological activities, including antimicrobial and antibiotic potentiation properties. We have prepared an expanded set of diarylbis(thioureido)polyamines that vary in central polyamine core length, identifying analogues with potent methicillin-resistant Staphylococcus aureus (MRSA), Escherichia coli, Acinetobacter baumannii and Candida albicans growth inhibition properties, in addition to the ability to enhance action of doxycycline towards Gram-negative bacterium Pseudomonas aeruginosa. The observation of associated cytotoxicity/hemolytic properties prompted synthesis of an alternative series of diacylpolyamines that explored aromatic head groups of varying lipophilicity. Examples bearing terminal groups each containing two phenyl rings (15a-f, 16a-f) were found to have optimal intrinsic antimicrobial properties, with MRSA being the most susceptible organism. A lack of observed cytotoxicity or hemolytic properties for all but the longest polyamine chain variants identified these as non-toxic Gram-positive antimicrobials worthy of further study. Analogues bearing either one or three aromatic-ring-containing head groups were either generally devoid of antimicrobial properties (one ring) or cytotoxic/hemolytic (three rings), defining a rather narrow range of head group lipophilicity that affords selectivity for Gram-positive bacterial membranes versus mammalian. Analogue 15d is bactericidal and targets the Gram-positive bacterial membrane.

Investigation of α,ω-Disubstituted Polyamine-Cholic Acid Conjugates Identifies Hyodeoxycholic and Chenodeoxycholic Scaffolds as Non-Toxic, Potent Antimicrobials.

With the increased incidence of antibiotic resistance, the discovery and development of new antibacterials is of increasing importance and urgency. The report of the natural product antibiotic squalamine in 1993 has stimulated a lot of interest in the study of structurally simplified cholic acid-polyamine derivatives. We report the synthesis of a focused set of deoxycholic acid-polyamine conjugates and the identification of hyodeoxycholic acid derivatives as being potently active towards S. aureus MRSA and some fungal strains, but with no attendant cytotoxicity or hemolytic properties. Analogue 7e exhibited bactericidal activity towards a range of Gram-positive bacteria, while preliminary investigation of its mechanism of action ruled out the bacterial membrane as being a primary cellular target as determined using an ATP-release bioluminescence assay.

Marine life as a source for breast cancer treatment: A comprehensive review.

Breast cancer, one of the most significant tumors among all cancer cells, still has deficiencies for effective treatment. Moreover, substitute treatments employing natural products as bioactive metabolites has been seriously considered. The source of bioactive metabolites are not only the most numerous but also represent the richest source. A unique source is from the oceans or marine species which demonstrated intriguing chemical and biological diversity which represents an astonishing reserve for discovering novel anticancer drugs. Notably, marine sponges produce the largest amount of diverse bioactive peptides, alkaloids, terpenoids, polyketides along with many secondary metabolites whose potential is mostly therapeutic. In this review, our main focus is on the marine derived secondary metabolites which demonstrated cytotoxic effects towards numerous breast cancer cells and have been isolated from the marine sources such as marine sponges, cyanobacteria, fungi, algae, tunicates, actinomycetes, ascidians, and other sources of marine organisms.

Computational Metabolomics Tools Reveal Subarmigerides, Unprecedented Linear Peptides from the Marine Sponge Holobiont Callyspongia subarmigera.

A detailed examination of a unique molecular family, restricted to the Callyspongia genus, in a molecular network obtained from an in-house Haplosclerida marine sponge collection (including Haliclona, Callyspongia, Xestospongia, and Petrosia species) led to the discovery of subarmigerides, a series of rare linear peptides from Callyspongia subarmigera, a genus mainly known for polyacetylenes and lipids. The structure of the sole isolated peptide, subarmigeride A (1) was elucidated through extensive 1D and 2D NMR spectroscopy, HRMS/MS, and Marfey's method to assign its absolute configuration. The putative structures of seven additional linear peptides were proposed by an analysis of their respective MS/MS spectra and a comparison of their fragmentation patterns with the heptapeptide 1. Surprisingly, several structurally related analogues of subarmigeride A (1) occurred in one distinct cluster from the molecular network of the cyanobacteria strains of the Guadeloupe mangroves, suggesting that the true producer of this peptide family might be the microbial sponge-associated community, i.e., the sponge-associated cyanobacteria.

Discovery and Preliminary Structure-Activity Investigation of 3-Substituted-1H-imidazol-5-yl-1H-indoles with In Vitro Activity towards Methicillin-Resistant Staphylococcus aureus.

Antibiotics have been the cornerstone of modern medicine saving lives by virtue of being able to cure infectious diseases and to prevent infections in those who are immune compromised. Their intense use has led to a surging increase in the incidence of antibiotic-resistant bacteria resulting in a desperate need for antibiotics with new mechanisms of action. As part of our search for new antimicrobials we have screened an in-house library of compounds and identified two 3-substituted-1H-imidazol-5-yl-1H-indoles as weak growth inhibitors (MIC 16 µg/mL) against methicillin-resistant Staphylococcus aureus (MRSA). An extensive library of analogues was prepared using the Van Leusen three-component reaction, biological evaluation of which led to the identification of two analogues (26 and 32) with favorable anti-MRSA activity (MIC ≤ 0.25 µg/mL) which also lacked cytotoxic or hemolytic properties. The screening campaign also identified two derivatives, a phenethyl-indole-imidazole 57 and a 5-phenyl-1H-imidazole 111 that were non-toxic selective antifungals towards Cryptococcus neoformans. These results have identified 3-substituted-1H-imidazol-5-yl-1H-indoles and 5-phenyl-1H-imidazoles as new structural scaffolds for further investigation as anti-MRSA and anti-C. neoformans agents, respectively.

Total Synthesis of the Four Stereoisomers of Cyclo(l-Trp-l-Arg) Raises Uncertainty of the Structures of the Natural Products and Invalidates Their Promising Antimicrobial Activities.

New therapeutic options to combat the growing incidence of antimicrobial resistance are urgently needed. A 2015 publication reported the isolation and biological evaluation of two diketopiperazine natural products, cyclo(l-Trp-l-Arg) (CDP 2) and cyclo(d-Trp-d-Arg) (CDP 3), from an Achromobacter sp. bacterium, finding that the latter metabolite in particular exhibited strong antibacterial activity towards a range of wound-related microorganisms and could synergize the action of ampicillin. Intrigued by these biological activities and noting inconsistencies in the structural characterization of the natural products, we synthesized the four diastereomers of cyclo(Trp-Arg) and evaluated them for antimicrobial and antibiotic enhancement properties. The detailed comparison of spectroscopic data raises uncertainty regarding the structure of CDP 2 and disproves the structure of CDP 3. In our hands, none of the four stereoisomers of cyclo(Trp-Arg) exhibited detectable intrinsic antimicrobial properties towards a range of Gram-positive and Gram-negative bacteria or fungi nor could they potentiate the action of antibiotics. These discrepancies in biological properties, compared with the activities reported in the literature, reveal that these specific cyclic dipeptides do not represent viable templates for the development of new treatments for microbial infections.

Metabolomics with multi-block modelling of mass spectrometry and nuclear magnetic resonance in order to discriminate Haplosclerida marine sponges.

A comprehensive metabolomic strategy, integrating 1H NMR and MS-based multi-block modelling in conjunction with multi-informational molecular networking, has been developed to discriminate sponges of the order Haplosclerida, well known for being taxonomically contentious. An in-house collection of 33 marine sponge samples belonging to three families (Callyspongiidae, Chalinidae, Petrosiidae) and four different genera (Callyspongia, Haliclona, Petrosia, Xestospongia) was investigated using LC-MS/MS, molecular networking, and the annotations processes combined with NMR data and multivariate statistical modelling. The combination of MS and NMR data into supervised multivariate models led to the discrimination of, out of the four genera, three groups based on the presence of metabolites, not necessarily previously described in the Haplosclerida order. Although these metabolomic methods have already been applied separately, it is the first time that a multi-block untargeted approach using MS and NMR has been combined with molecular networking and statistically analyzed, pointing out the pros and cons of this strategy.

Valorisation of the diterpene podocarpic acid - Antibiotic and antibiotic enhancing activities of polyamine conjugates.

As part of our search for new antimicrobials and antibiotic adjuvants, a series of podocarpic acid-polyamine conjugates have been synthesized. The library of compounds made use of the phenolic and carboxylic acid moieties of the diterpene allowing attachment of polyamines (PA) of different lengths to afford a structurally-diverse set of analogues. Evaluation of the conjugates for intrinsic antimicrobial properties identified two derivatives of interest: a PA3-4-3 (spermine) amide-bonded variant 7a that was a non-cytotoxic, non-hemolytic potent growth inhibitor of Gram-positive Staphylococcus aureus (MRSA) and 9d, a PA3-8-3 carbamate derivative that was a non-toxic selective antifungal towards Cryptococcus neoformans. Of the compound set, only one example exhibited activity towards Gram-negative bacteria. However, in the presence of sub-therapeutic amounts of either doxycycline (4.5 µM) or erythromycin (2.7 µM) several analogues were observed to exhibit weak to modest antibiotic adjuvant properties against Pseudomonas aeruginosa and/or Escherichia coli. The observation of strong cytotoxicity and/or hemolytic properties for subsets of the library, in particular those analogues bearing methyl ester or n-pentylamide functionality, highlighted the fine balance of structural requirements and lipophilicity for antimicrobial activity as opposed to mammalian cell toxicity.

The potential role of dietary plant ingredients against mammary cancer: a comprehensive review.

Breast cancer is known as the most devastating cancer in the global female community and is considered as one of the severe health care burdens in both developed and developing countries. In many cases, breast cancer has shown resistance to chemotherapy, radiotherapy and hormonal therapy. Keeping in view these limitations, there is an urgent need to develop safe, readily available and effective breast anticancer treatments. Therefore, the scientists are keen in the extraction of plant-based phytochemicals (organosulfur compounds, betalains, capsaicinoids, terpenes, terpenoids, polyphenols, and flavonoids) and using them as breast anticancer agents. Results of numerous epidemiological investigations have revealed the promising role of phytochemicals in the prevention and treatment of breast cancer. The diverse classes of plant bioactive metabolites regulate different metabolic and molecular processes, which can delay the proliferation of cancers. These phytochemicals possess chemo-preventive properties as they down-regulate the expression of estrogen receptor-α, inhibit the proliferation of cancer cells, and cause cell cycle arrest by inducing apoptotic conditions in tumor cells. This review article discusses the potent role of various plant-based phytochemicals as potential therapeutic agents in the treatment or prevention of breast cancer along with the proposed mechanisms of action.

Repurposing primaquine as a polyamine conjugate to become an antibiotic adjuvant.

In our search for new antibiotic adjuvants as a novel strategy to deal with the emergence of multi-drug resistant (MDR) bacteria, a series of succinylprimaquine-polyamine (SPQ-PA) conjugates and derivatives of a cationic amphiphilic nature have been prepared. Evaluation of these primaquine conjugates for intrinsic antimicrobial properties and the ability to restore the antibiotic activity of doxycycline identified two derivatives, SPQ-PA3-8-3 and SPQ-PA3-10-3 that exhibited intrinsic activity against the Gram-positive bacteria Staphylococcus aureus and the yeast Cryptococcus neoformans. None of the analogues were active against the Gram-negative bacterium Pseudomonas aeruginosa. However, in the presence of a sub-therapeutic amount of doxycycline (4.5 µM), both SPQ-PA3-4-3 and SPQ-PA3-10-3 compounds displayed potent antibiotic adjuvant properties against P. aeruginosa, with MIC's of 6.25 µM. A series of derivatives were prepared to investigate the structure-activity relationship that explored the influence of both a simplified aryl lipophilic substituent and variation of the length of the polyamine scaffold on observed intrinsic antimicrobial properties and the ability to potentiate the action of doxycycline against P. aeruginosa.

Spermine Derivatives of Indole-3-carboxylic Acid, Indole-3-acetic Acid and Indole-3-acrylic Acid as Gram-Negative Antibiotic Adjuvants.

The discovery of new antibiotic adjuvants is an attractive option for overcoming antimicrobial resistance. We have previously reported the discovery of a bis-6-bromoindolglyoxylamide derivative of spermine as being able to enhance the action of antibiotics against Gram-negative bacteria but suffers from being cytotoxic and red-blood cell haemolytic. A series of analogues was prepared exploring variation of the indolglyoxylamide unit, to include indole-3-acrylic, indole-3-acetic and indole-3-carboxylate units, and evaluated for antibiotic enhancing properties against a range of Gram-negative bacteria, and for intrinsic antimicrobial, cytotoxic and haemolytic properties. Two spermine derivatives, bearing 5-bromo-indole-3-acetic acid (17) and 5-methoxy-indole-3-acrylic acid (14) end groups were found to exhibit good to moderate antibiotic adjuvant activities for doxycycline towards the Gram-negative bacteria Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumoniae, but with more modest intrinsic antimicrobial activity and greatly reduced cytotoxic and haemolytic properties. The mechanism of action of the latter derivative identified its ability to disrupt the outer membranes of bacteria and to inhibit the AcrAB-TolC efflux pump directly or by inhibiting the proton gradient.

Functional diversity of microboring Ostreobium algae isolated from corals.

The filamentous chlorophyte Ostreobium sp. dominates shallow marine carbonate microboring communities, and is one of the major agents of reef bioerosion. While its large genetic diversity has emerged, its physiology remains little known, with unexplored relationship between genotypes and phenotypes (endolithic versus free-living growth forms). Here, we isolated nine strains affiliated to two lineages of Ostreobium (>8% sequence divergence of the plastid gene rbcL), one of which was assigned to the family Odoaceae, from the fast-growing coral host Pocillopora acuta Lamarck 1816. Free-living isolates maintained their bioerosive potential, colonizing pre-bleached coral carbonate skeletons. We compared phenotypes, highlighting shifts in pigment and fatty acid compositions, carbon to nitrogen ratios and stable isotope compositions (δ13 C and δ15 N). Our data show a pattern of higher chlorophyll b and lower arachidonic acid (20:4ω6) content in endolithic versus free-living Ostreobium. Photosynthetic carbon fixation and nitrate uptake, quantified via 8 h pulse-labeling with 13 C-bicarbonate and 15 N-nitrate, showed lower isotopic enrichment in endolithic compared to free-living filaments. Our results highlight the functional plasticity of Ostreobium phenotypes. The isotope tracer approach opens the way to further study the biogeochemical cycling and trophic ecology of these cryptic algae at coral holobiont and reef scales.