Abstract
The application of beneficial microorganisms for corals (BMC) decreases the bleaching susceptibility and mortality rate of corals. BMC selection is typically performed via molecular and biochemical assays, followed by genomic screening for BMC traits. Herein, we present a comprehensive in silico framework to explore a set of six putative BMC strains. We extracted high-quality DNA from coral samples collected from the Red Sea and performed PacBio sequencing. We identified BMC traits and mechanisms associated with each strain as well as proposed new traits and mechanisms, such as chemotaxis and the presence of phages and bioactive secondary metabolites. The presence of prophages in two of the six studied BMC strains suggests their possible distribution within beneficial bacteria. We also detected various secondary metabolites, such as terpenes, ectoines, lanthipeptides, and lasso peptides. These metabolites possess antimicrobial, antifungal, antiviral, anti-inflammatory, and antioxidant activities and play key roles in coral health by reducing the effects of heat stress, high salinity, reactive oxygen species, and radiation. Corals are currently facing unprecedented challenges, and our revised framework can help select more efficient BMC for use in studies on coral microbiome rehabilitation, coral resilience, and coral restoration.
Similar content being viewed by others
Introduction
Coral reefs are one of the most diverse and productive ecosystems on Earth. However, they are currently facing unprecedented challenges due to overfishing, pollution, and climate change1,2,3,4. Increased greenhouse gas concentrations leading to global warming5, pollution and ocean acidification have emerged as key stressors, threatening the survival of coral reefs worldwide3,6,7,8. Some of these stressors cause periods of high seawater temperatures, leading to the loss of Symbiodiniaceae, the algal symbionts that reside within the holobiont and fulfill up to 90% of the corals’ nutritional requirements9 except when they become nutrient competitors and can be expelled10,11,12. This loss, especially over a long-term, can lead to devastating bleaching events worldwide, in which corals are deprived of vital nutrients, leading to increased disease susceptibility and reduced reproductive success and skeletal growth2,13,14,15,16. This disruption extends beyond the loss of algal symbionts because it is followed by shifts in the composition and function of the entire coral-associated microbiome17. This causes post-heat stress disorder18 that compromises the coral’s ability to grow, resist pathogens, modulate nutrient cycling, and maintain holobiont homeostasis18,19,20,21,22,23,24,25,26,27.
The symbiotic relationship between corals and their bacterial symbionts28,29 plays a pivotal role in maintaining the health of these organisms and consequently the functioning of reef ecosystems. Although some corals and their microbiomes have shown the capacity to recover from acute stress events30,31, the coral microbiome may not fully restore its original composition and functionality following a severe disturbance32.
Rehabilitation of the coral microbiome33 has recently emerged as a promising strategy to enhance coral health and resilience18,24, 26, 33,34,35,36,37,38,39,40,41,42. Coral-associated bacteria possess potential beneficial traits that enhance the fitness and resilience of their coral host by maintaining homeostasis through their involvement in various essential processes, such as nutrient cycling, production of antibiotics and antimicrobial compounds, and mitigation of toxic compounds33,43. Harnessing the symbiotic interactions between corals and their associated bacteria28 allows researchers to selectively introduce these beneficial microorganisms for corals (BMC)33 into the coral holobiont for use as customized probiotics. This can help maintain or even enrich the proportion of native beneficial microorganisms to enhance coral health and resilience under stress33,35.
In this study, we performed an in silico investigation of the probiotic potential of six putative BMC (pBMC) strains isolated from corals in the Red Sea with the aim of developing the first Red Sea BMC consortium. This was achieved by analyzing the complete genomes of all six strains and screening for the presence of potential beneficial mechanisms for corals. Our research provides a framework for the selection of novel, customized BMC consortia based on the presence of particular BMC characteristics that support host health and survival under stress conditions.
Materials and methods
pBMC isolation and genome sequencing
pBMC strains were isolated from coral fragments (± 5 cm long) of Pocillopora verrucosa, Galaxea fascicularis, and Stylophora pistillata Clade IV. The fragments were collected between February and April 2021 from Al Fahal reef (22°18′18.4′′N, 38°57′52.5′′E), Aquarium reef (22°23′15.6′′N, 38°55′07.2′′E), and Tahala reef (22°15′46.9′′N, 39°03′05.9′′E) in the eastern central Red Sea, Saudi Arabia, as described by Delgadillo-Ordoñez and colleagues44.
Briefly, coral fragments were macerated in 50 mL of 3.5% sterile saline solution using a sterile mortar and pestle. Then, the macerated paste was serially diluted up to 10−6 with 3.5% saline solution, and 100 µL of each dilution was plated on Zobel Marine Agar 2216 (MA; HiMedia Laboratories, Mumbai, India), diluted marine agar (DMA; 2 × dilution of MA with 3.5% NaCl and adjusted agar content) and Lennox Broth (Sigma-Aldrich) and cultured overnight at 25 °C. The macerate was also incubated in parallel at 25 °C overnight and 130 rpm in a 250-mL sterile Erlenmeyer flask containing crystal beads. After this first incubation, 100 µL triplicate subsamples of 10−4, 10−5, and 10−6 dilutions were plated on approximately 20 mL of MA and DMA. Alternatively, 0.5-cm long coral fragments were plated on both media. All plates were incubated at 25 °C for at least 48 h or until visible bacterial colonies were observed. Based on colony morphology, > 350 isolates were obtained, which were preserved at −80 °C in sterilized 20% glycerol. To select pBMC strains, the isolates were screened for various beneficial traits using polymerase chain reaction (PCR) assays targeting nitrogen cycle-related genes involved in nitrogen fixation (nifH) and denitrification (nirK) as well as biochemical analyses of catalase activity, siderophore production45 and phosphate assimilation46. Finally, the antagonistic effect of the isolates against the coral pathogen Vibrio coralliilyticus47,48 was tested following the methodology described by Rosado and colleagues26 and Santoro and colleagues18. Based on the positive results of these tests, six strains were selected as pBMC and stored in glycerol stocks (20%) at −80 °C.
We extracted genomic DNA (gDNA) from these six pBMC strains using Wizard Genomic DNA Purification Kit (Promega, Madison, WI, USA) according to the manufacturer’s protocol. The resultant gDNA was purified with AMPure beads (1:8 ratio; Beckman Coulter) and quantified using Qubit dsDNA Assay Kit (Invitrogen). Then, the quality and integrity of the extracted gDNA were confirmed via spectrometry using a NanoDrop 8000 spectrophotometer (Thermo Fisher Scientific) and via electrophoresis using the 4200 TapeStation System (Agilent Technologies). Next, we prepared SMRTbell libraries using SMRTbell Express Template Prep Kit 2.0 (Pacific Biosciences) and Barcoded Overhang Adapter Kit 8A (Pacific Biosciences) according to the PacBio multiplexed microbial libraries protocol. The quality of the libraries was assessed using Qubit dsDNA High Sensitivity Assay Kit (Thermo Fisher Scientific) and FEMTO Pulse pulsed-field capillary electrophoresis system (Agilent Technologies). For sequencing, the libraries were first annealed using Primer v4 via Sequel Binding Kit 3.0 (Pacific Biosciences) and then sequenced using SMRT Cell 1 M v3 LR Tray (Pacific Biosciences) via the Sequel system at the Bioscience Core Lab of King Abdullah University of Science and Technology, Saudi Arabia.
pBMC genome assembly and taxonomic classification
The quality of raw reads was assessed using LongQC tool49, and the reads were trimmed according to the default parameters. Next, the reads were assembled using Canu50, Flye51, Raven52, Miniasm/Minipolish53, and Redbean54. The generated assemblies were then evaluated and compared to determine the quality of general genome assembly via Quast55, genomic completeness/contamination via CheckM software56, chimerism via GUNC57, and indel errors/pseudogenes via ideel (https://github.com/phiweger/ideel). The assemblies from Canu were considered the best based on assembly quality, genomic completeness, and lower number of indels/interrupted open reading frames. FastANI58, and Genome-to-Genome Distance Calculator59 were used to assess the phylogenetic relatedness between the study genomes and the publicly available closest reference strain genomes.
Structural and functional annotation of pBMC genomes
The six pBMCs genomes were annotated using Prokka60 and the PATRIC Bacterial and Viral Bioinformatics Resource Center (BV-BRC)61, which uses the RASTtk online server62, according to the default parameters. The output data were visualized on the BV-BRC interface. We screened for protein-coding genes that might provide beneficial characteristics for host health and resilience33,43 by searching the subsystems generated by the RASTtk platform in BV-BRC. Additionally, the six pBMC genomes were searched for putative prophage sequences using the PHASTEST and VirSorter2 online analysis tools63,66. PHASTEST analyzes the proportion of phage proteins within a bacterial genomic region and scores the potential phages based on their length and inclusion of several phage-related proteins, such as “capsid,” “head,” “integrase,” “plate,” “tail,” “fiber,” “coat,” “transposase,” “portal,” “terminase,” “protease,” or “lysin,” in the input file. VirSorter2 uses a set of viral HMM databases created from various large-scale viral metagenome projects and trained viral classifiers to identify viral sequences within predominantly metagenomic samples. Prophages detected by both tools were further analyzed using CheckV67. Only the consensus prophages characterized as high quality by both identification tools were used in this analysis. Only phages considered intact based on the abovementioned criteria were used for further analysis. Intact phages were queried using the IMG/VR v4 online BLAST database, and similarity was assessed based on the highest reported bit score with high-quality reference phages68. AntiSMASH v769 was also used to identify, annotate, and analyze secondary metabolite biosynthetic gene clusters (BGCs) present in the genomes of all six pBMC strains.
Phylogenomic trees
A phylogenomic assemblage was constructed using the codon tree method via the BV-BRC platform. This method randomly selects up to 10 proteins to represent each genus-level family and merges them to create a single set of representatives, preventing cluster formation, which is based on genus similarity rather than protein similarity (PATRIC global protein families [PGFams])61,70. Amino acids and nucleotide sequences were used for analyzing each selected gene from the PGFams. Protein sequences were aligned using MUSCLE71, and encoding gene sequences were aligned using the Codon_align function in BioPython72. The output was a concatenated alignment of all proteins and nucleotides as well as a partitioned file describing the alignment of proteins for analysis using RAxML73. Support values were generated using 100 rounds of the “Rapid” bootstrap option74 in RAxML. The phylogenomic trees were constructed by comparing and analyzing protein-coding genes (n = 1000, Figs. 1, 2, 3, 4 and 5; n = 500, Fig. S1) shared among all genomes using the best protein model method identified by RAxML75. The output, in Newick format, was then visualized via iTOL version 676,77. The trees generated are separated by genus (Pseudoalteromonas, Cobetia, Halomonas, and Sutcliffiella), and the genomes used to construct these trees were selected from the BV-BRC platform database based on various criteria, such as being reference genomes, complete genomes, and isolated from corals and/or aquatic environments, as per their availability.
Phylogenomic inference of publicly available genomes from 14 Pseudoalteromonas sp. strains and the genomes of our two Pseudoalteromonas galatheae strains (pBMC1 and pBMC2; highlighted in bold), totaling 16 genomes. The tree was assembled by comparing 1000 proteins using the codon tree method of the PATRIC platform. This method selects global protein families as homology groups and analyzes aligned proteins and DNA encoding single-copy genes using the RAxML program. Based on RAxML, the Jones–Taylor–Thornton protein model was considered the best model to construct this tree. Gray dots on the branches correspond to a bootstrap value of 100. The isolate source is indicated on the right side of each strain. The blue-shaded area represents the monophyletic clade comprising pBMC1 and pBMC2.
Phylogenomic inference of publicly available genomes from 26 Cobetia sp. strains, the genome of Cobetia sp. BMC6 from Rosado and colleagues172, and the genome of our two Cobetia sp. strains (pBMC3 and pBMC4; highlighted in bold) totaling 29 genomes. The tree was assembled by comparing 1000 proteins using the codon tree method of the PATRIC platform. This method selects global protein families as homology groups and analyzes aligned proteins and DNA encoding single-copy genes using the RAxML program. Based on RAxML, the Jones–Taylor–Thornton protein model was considered the best model to construct this tree. The isolate source is indicated on the right side of each strain. Gray dots on the branches correspond to a bootstrap value of 100. The blue-shaded area represents the monophyletic clade comprising pBMC3 and pBMC4. The Cobetia sp. BMC strain from the study by Rosado and colleagues172 is shaded in green.
Phylogenomic inference of publicly available genomes from 24 Halomonas sp. strains, the genome of Halomonas sp. BMC7 from Rosado and colleagues172 and the genome of our Halomonas sp. strain pBMC5 (highlighted in bold), totaling 26 genomes. The tree was assembled by comparing 1000 proteins using the codon tree method of the PATRIC platform. This method selects global protein families as homology groups and analyzes aligned proteins and DNA encoding single-copy genes using the RAxML program. Based on RAxML, the LG model was considered the best model to construct this tree. Gray dots on the branches correspond to a bootstrap value of 100. The isolate source is indicated on the right side of each strain. The blue-shaded area represents the strains that form a monophyletic clade with pBMC5. The Halomonas sp. BMC strain from Rosado and colleagues26,172 is shaded in green.
Phylogenomic inference of publicly available genomes from 1 Sutcliffiella sp. strain and ten Bacillus sp. strains and the genome of our Sutcliffiella sp. strain pBMC6 (highlighted in bold), totaling 12 genomes. The tree was assembled by comparing 1000 proteins using the codon tree method of the PATRIC platform. This method selects global protein families as homology groups and analyzes aligned proteins and DNA encoding single-copy genes using the RAxML program. Based on RAxML, the LG model was considered the best model to construct this tree. Gray dots on the branches correspond to a bootstrap value of 100. The isolate source is indicated on the right side of each strain. The blue-shaded area represents the strains that form a monophyletic clade with pBMC6.
Genome maps of the six putative BMC strains. The rings (from outside to inside) indicate the nucleotide position in the genome, predicted genes on the forward strand, predicted genes on the reverse strand, location of putative prophages (as predicted by PHASTEST and VirSorter2), and percent GC content (higher GC content = yellow, lower GC content = purple). Genes predicted to encode beneficial proteins are color-coded in the outer rings. The circular genomes were visualized using DNAplotter 177.
Pangenomic analysis and gene categorization
We used the Prokka annotations to generate a separate pangenome for each pBMC genus (Pseudoalteromonas, Cobetia, Halomonas, and Sutcliffiella) using the Roary tool77, with the default settings. For each pangenome, sequences from the closest genomes to our pBMC strains, which were identified based on the phylogenomic tree results, were added to the analysis. For the pangenomic analysis of Pseudoalteromonas sp., in addition to our two Pseudoalteromonas galatheae genomes (pBMC1 and pBMC2), 19 additional Pseudoalteromonas sp. genomes were included (from Fig. 1). For the pangenomic analysis of Cobetia sp., in addition to our two Cobetia amphilecti genomes (pBMC3 and pBMC4), three Cobetia sp. genomes were added (from Fig. 2). The pangenome of Halomonas sp. included our single Halomonas sp. genome (pBMC5) and three other Halomonas sp. genomes (from Fig. 3). For Sutcliffiella sp., its pangenome consisted of the genome of Sutcliffiella sp. pBMC6, one other Sutcliffiella sp. genome, and two Bacillus sp. genomes (from Fig. 4). All genomes used for comparison with the pBMC genomes were obtained from the BV-BRC genome database61. The pangenomes were created and visualized using the Anvi’o platform78, and the genes were categorized according to these results. Singleton genes were defined as genes that were only present in one or more of the pBMC genomes and not in the other genomes that were part of the same pangenome. These singleton genes were identified for each pBMC and screened for their potential to enhance the health status and resilience of the host using UniProt (https://www.uniprot.org/)79.
Results
Genomic features of pBMC
The six pBMC genomes were sequenced using PacBio, resulting in several cleaned and trimmed high-quality reads of 137,126–345,950, with an average read length of 5827 bp. The genomes were assembled using Canu50, resulting in 2–9 contigs per strain. CheckM analysis revealed a high level of genome completeness (> 99% for all genomes except Sutcliffiella sp. pBMC6, which was > 97.5%) and a low level of contamination (< 2.5%) for all genomes (Table S1).
The genomes of P. galatheae pBMC1 and pBMC2 had very similar characteristics, with a GC content of 42.94% and 42.98% and a total length of 5,528,212 and 5,474,806 bp, respectively. The genomes of Cobetia sp. pBMC3 and pBMC4 also had very similar characteristics, with a GC content of 62.32% and 62.31% and a total length of 4,046,628 and 4,041,826 bp, respectively (Table S1).
Taxonomic attribution
pBMC1 and pBMC2 represented the same species, sharing an average nucleotide identity (ANI) of > 99.5% at both genome and 16S rRNA gene levels. The DNA–DNA hybridization (DDH) and ANI values with the strain Pseudoalteromonas galatheae were 92% and 99.17% for pBMC1 and 92.4% and 99.19% for pBMC2, respectively, suggesting that they belonged to this species (Table S2). Notably, the species-level thresholds are 70% for DDH80 and 95–96% for ANI81. Similarly, pBMC3 and pBMC4 showed an ANI of > 99.5% at the genome and 16S rRNA gene levels. Their DDH and ANI values with other Cobetia sp. were also high, and pBMC3 and pBMC4 were classified as Cobetia sp. in this study (Table S2). pBMC5 showed 97.94% ANI with the genomes of two strains of Halomonas meridiana (Table S2); however, compared with the type strain of H. meridiana (H. meridiana DSM5425), pBMC5 showed an ANI of < 90% and DDH of < 70%, suggesting that it is a novel species. pBMC6 also potentially represents a novel species, with the closest species genome being that of Bacillus horikoshii at an ANI of 85.96% (Table S2). The systematic classification of the family Bacillaceae has undergone several modifications due to the implementation of new taxonomic polyphasic techniques82,83, which have led to the creation of new genera from the previously classified genus Bacillus, including the new genus Sutcliffiella83. Considering this, the overall taxonomic classification of the six pBMC strains investigated in our study should be as follows: P. galatheae (n = 2), Cobetia sp. (n = 2), Halomonas sp. (n = 1), and Sutcliffiella sp. (n = 1).
Phylogenomic trees
The phylogenomic relationships between the six pBMC genomes and the closest publicly available genomes were evaluated. We used 115 genomes to assemble the phylogenomic tree of P. galatheae pBMC1 and pBMC2 (Fig. S1). A well-supported clade, formed by pBMC1, pBMC2, and 14 other Pseudoalteromonas sp. strains, was established in this phylogenomic tree. Consequently, another phylogenomic tree was assembled using the genomes of this clade (Fig. 1). All 16 strains included in this new tree were host-associated. For the phylogenomic tree of Cobetia sp. pBMC3 and pBMC4, the genomes of 27 other Cobetia sp. were added, and pBMC3 and pBMC4 formed a clade (Fig. 2). The phylogenomic tree of Halomonas sp. pBMC5 included 25 other Halomonas sp. genomes (Fig. 3). The Halomonas sp. pBMC5 formed a small clade with one Halomonas sp. strain and one H. meridiana strain, creating a coral-associated clade. In the phylogenomic tree of Sutcliffiella sp. pBMC6, all 11 publicly available genomes were included (Fig. 4). There was a small clade between Sutcliffiella sp. pBMC6 and two other Sutcliffiella sp. strains (Sutcliffiella horikoshii and B. horikoshii), one of which was isolated from soil and the other from a clean room.
Potentially beneficial gene functions
We used the BV-BRC platform to screen for genes encoding proteins that are potentially beneficial for corals33,43, 84 and their endosymbiotic algae85. Subsystems that are known to play a role in the health and resilience of corals, as described by Peixoto and colleagues33,43, and Matthews and colleagues85, were selected, and genes associated with oxidative stress, nitrogen cycle, ectoine and B-complex vitamin biosynthesis, iron and sulfur metabolism, and dimethylsulfoniopropionate (DMSP) degradation were identified (Table S3).
Within the subsystems of interest, all pBMC harbored genes related to protection from reactive oxygen species (ROS) and biosynthesis of vitamins B2, B7, and B9. pBMC1, pBMC2, pBMC5, and pBMC6 shared genes related to catalase and peroxidase activities and vitamin B12 biosynthesis. All pBMC, except for pBMC6, harbored genes involved in vitamin B6 biosynthesis. pBMC6 was the only pBMC harboring genes involved in iron acquisition. Genes related to ectoine biosynthesis were only detected in pBMC3, pBMC4, and pBMC5, whereas genes related to the nitrogen cycle were found in pBMC3, pBMC4, and pBMC5. DMSP degradation-related genes were only detected in pBMC3 and pBMC4, whereas genes related to sulfur metabolism were only identified in pBMC1 and pBMC2 (Table S3).
Putative prophages
Of the six described pBMC strains herein, two contained prophages that were determined to be high quality by both PHASTEST and VirSorter2. Every high-quality prophage also had a 100% completeness score. One prophage was predicted in the genome of P. galatheae pBMC1 (Fig. 5). This prophage was most similar to the tailed phage that infects P. piscicida DE2-A. No putative prophages were detected in P. galatheae strain pBMC2, either of the Cobetia sp. strains (pBMC3 and pBMC4), nor Halomonas sp. pBMC5. However, two prophages were found in Sutcliffiella sp. pBMC6, and they were most similar to the tailed phages infecting S. horikoshii FJAT-14233 and S. horikoshii DSM 8719.
Pangenomes
A pangenomic analysis was conducted for each pBMC clade to screen for genes unique to pBMC strains (Figs. 6, 7, 8 and 9). For the pangenome of pBMC1 and pBMC2, 19 Pseudoalteromonas sp. strains were added, including BMC from the study by Rosado et al. (2018) (Fig. 6). We identified 714 unique genes (i.e., 714 genes were present in the genomes of pBMC1 or pBMC2 but absent in the genomes of other 19 Pseudoalteromonas sp. strains). Of these 714 unique genes (450 of which were shared between pBMC1 and pBMC2), 604 were classified as hypothetical proteins, and the remaining 110 had known functions. Of the 110 genes with known functions, 49 were shared by pBMC1 and pBMC2. Furthermore, 3 of these 110 unique pBMC genes encoded proteins related to previously described beneficial traits for corals (Table 1).
Pangenomic analysis of Pseudoalteromonas galatheae pBMC1 and pBMC2, other Pseudoalteromonas sp. strains from their phylogenomic clade in Fig. 1, and the Pseudoalteromonas sp. BMC1–5 strains from Rosado and colleagues26,172. The outermost circle displays single-copy genes (SCG cluster) shared by all genomes. The presence and absence of coding sequences in the genomes are represented in dark blue and faded dark blue, respectively, for each strain. The average nucleotide identity (95%–100%) comparison between the 21 genomes is shown in the heat map on the upper right. The dendrogram above the heat map shows the hierarchical clustering of genomes based on the occurrence of gene clusters (dark blue, below the heat map), number of singleton gene clusters (turquoise blue, below the heat map), and total length (gray, below the heat map) of each genome.
Pangenomic analysis of Cobetia sp. pBMC3 and pBMC4, the closest Cobetia sp. strains from the phylogenomic tree shown in Fig. 2, and the Cobetia sp. BMC6 strain from Rosado and colleagues172. The outermost circle displays single-copy genes (SCG cluster) shared by all genomes. The presence and absence of coding sequences in the genomes are represented in dark blue and faded dark blue, respectively, for each strain. The average nucleotide identity (95%–100%) comparison between the five genomes is shown in the heat map on the upper right. The dendrogram above the heat map shows the hierarchical clustering of genomes based on the occurrence of gene clusters (dark blue, below the heat map), number of singleton gene clusters (turquoise blue, below the heat map), and total length (gray, below the heat map) of each genome.
Pangenomic analysis of Halomonas sp. pBMC5 and other Halomonas sp. strains from its phylogenomic clade in Fig. 3 and Halomonas sp. BMC7 from the study by Rosado and colleagues172. The outermost circle displays single-copy genes (SCG cluster) shared by all genomes. The presence and absence of coding sequences in the genomes are represented in dark blue and faded dark blue, respectively, for each strain. The average nucleotide identity (95%–100%) comparison between the four genomes is shown in the heat map on the upper right. The dendrogram above the heat map shows the hierarchical clustering of genomes based on the occurrence of gene clusters (dark blue, below the heat map), number of singleton gene clusters (turquoise blue, below the heat map), and total length (gray, below the heat map) of each genome.
Pangenomic analysis of Sutcliffiella sp. pBMC6 and other Sutcliffiella sp. and Bacillus sp. strains from its phylogenomic clade in Fig. 4. The outermost circle displays single-copy genes (SCG cluster) shared by all genomes. The presence and absence of coding sequences in the genomes are represented in dark blue and faded dark blue, respectively, for each strain. The average nucleotide identity (95–100%) comparison between the four genomes is displayed in the heat map on the upper right. The dendrogram above the heat map shows the hierarchical clustering of genomes based on the occurrence of gene clusters (dark blue, below the heat map), number of singleton gene clusters (turquoise blue, below the heat map), and total length (gray, below the heat map) of each genome.
There were 32 singleton genes identified in the pangenome of pBMC3 and pBMC4, which included 3 other Cobetia sp. strains (Fig. 7). Of these 32 singleton genes (with no genes being shared between pBMC3 and pBMC4), 5 had known functions, and the remaining were classified as hypothetical proteins. Of these 5 unique pBMC genes, none encoded proteins related to previously assigned beneficial traits for corals (Table 1).
Halomonas sp. pBMC5 formed a clade with two other Halomonas sp. strains, and a pangenome involving these three strains and Halomonas sp. BMC7 from the study by Rosado and colleagues26 was established (Fig. 8). A total of 740 genes were identified as singletons, which were present in the pBMC5 genome alone. Among these 740 genes, 536 were classified as hypothetical proteins, and 204 had known functions. Of these 204 unique pBMC5 genes, 4 encoded proteins related to previously assigned and potentially novel beneficial traits for corals related to antibiotic synthesis and transport and sulfur assimilation (Table 1).
Sutcliffiella sp. pBMC6 formed a clade with another Sutcliffiella sp. and one Bacillus sp. strain (Fig. 4). For a more comprehensive pangenome, B. horikoshii strain FJAT-14233 was added to the above-mentioned strains (Fig. 9). This pangenome harbored the highest number of unique pBMC genes, with 3088 genes present only in the pBMC6 genome. Of these, 1755 were classified as hypothetical proteins, and the remaining 1333 had known functions. Among these 1333 genes, 27 encoded proteins related to previously assigned and potentially new beneficial traits for corals related to siderophore, carotenoid, vitamin, and antibiotic synthesis, nitrogen, sulfur and iron metabolism, chemotaxis, degradation of toxic compounds and stress resistance (Table 1).
Biosynthesis of natural product
Mining of the six pBMC genomes on antiSMASH to screen for secondary metabolite BGCs revealed 49 BGCs across the six pBMC genomes. These included terpene (n = 2), type I and III polyketide synthase (PKS; n = 9 and n = 1, respectively), ectoine (n = 3), nonribosomal peptide synthetases (NRPSs; n = 8), nonribosomal peptide metallophores (n = 4), opine-like zincophores (n = 1), NRPS-independent (NI) IucA/IucC-like siderophores (n = 3), beta-lactone (n = 1), aryl polyene (n = 2), class I and IV lanthipeptides (both, n = 2), and lasso peptide (n = 1) as well as some putative clusters.
The NI siderophore cluster found in pBMC3 and pBMC4 showed 75% homology with known BGCs encoding desferrioxamine E (BGC0001572, BGC0001478, NCBI: MH015039.1). The ectoine cluster from pBMC5 showed 75% homology with several known ectoine-coding BGCs (BGC0000855, BGC0000856, BGC0000859, NCBI: DQ238213.1). The ectoine cluster of pBMC3 and pBMC4 showed 66% homology with the ectoine BGC000852 (NCBI: AF316874.1). The opine-like metallophore BGC found in pBMC6 showed 100% similarity with a known BGC encoding bacillopaline (BGC0002488, NCBI: CP002869.1), and the lasso peptide cluster showed 80% homology with the known BGC encoding paeninodin (BGC0001356, NCBI: AHKH01000064.1). Two terpenoid clusters were found in pBMC6. The first showed low homology with known clusters, highlighting the importance of continuing to study the biosynthesis of this potentially novel natural product in marine organisms. The other terpenoid gene cluster showed higher (66%) homology with a known BGC encoding a carotenoid (BGC0000645, NCBI: FJ040212.1). The pBMC6 genome contained a type III PKS cluster, and all other strains contained a type I-PKS cluster.
A protein family (Pfam)-based analysis was performed using antiSMASH, revealing squalene and phytoene synthase in Sutcliffiella sp. pBMC6 and a carotenoid biosynthesis-encoding gene (Table S4). The genomes of P. galatheae pBMC1 and pBMC2 contained lanthionine synthetase and lantibiotic biosynthesis-encoding genes as well as thioesterase synthase, which is necessary for peptide antibiotic production (Table S4). Cobetia sp. pBMC3 and pBMC4 harbored Pfam domains related to siderophore production and vitamin B12 and ectoine biosynthesis (Table S4), and ectoine synthase-encoding genes were present in Halomonas sp. pBMC5.
Discussion
Halomonas sp. pBMC5 and Sutcliffiella sp. pBMC6 as candidate novel species
Our results, particularly those based on the ANI and DDH values, suggest that both pBMC5 and pBMC6 are novel species, and further characterization is planned. The systematic classification of the family Bacillaceae has undergone numerous modifications in recent years due to the implementation of new taxonomic polyphasic techniques82,83. This has resulted in the creation of new genera from the previously classified Bacillus genus, such as the new genus Sutcliffiella83, and justifies the presence of B. horikoshii strains in the phylogenomic tree of pBMC6—likely an older classification of the current S. horikoshii—and the dominance of Bacillus sp. genomes in this tree. This is further corroborated by the presence of two putative prophages in pBMC6 that were predicted to infect S. horikoshii hosts.
Genome screening reveals previously proposed beneficial traits of pBMC
Following the classification of each pBMC for identification and phylogenomic analysis, we screened the pBMC genomes for genes encoding proteins that are potentially beneficial for corals. We screened for genes related to catalase, urease, and siderophore production; phosphate assimilation; and nitrogen cycle and DMSP degradation through biochemical tests and PCR assays44, which are typically employed for BMC selection. We also detected genes involved in other potential beneficial traits (Table S3), including those related to oxidative stress, such as superoxide dismutases (all pBMC genomes), which exert an antioxidant effect by catalyzing the dismutation of superoxide (an ROS molecule that causes cell damage)86; catalase KatE (all pBMC genomes) and catalase-peroxidase KatG (pBMC1, pBMC2, pBMC5, and pBMC6 genomes), both of which protect cells from the toxic effects of H2O2 and aerated growth conditions87,86,87,88,91; manganese catalase (pBMC5 and pBMC6 genomes), which is also involved in the protection of cells from H2O292,93; and glutathione synthetase (all pBMC genomes except for pBMC6), which produces glutathione that can subsequently be used by glutathione peroxidase (all pBMC genomes except for pBMC3 and pBMC4) to scavenge ROS, such as H2O294,95. When seawater temperatures rise, the coral holobiont produce ROS, resulting in cell damage in both host and its symbionts96,95,98. A direct correlation between bleaching and ROS production has been previously reported3, and ROS-scavenging pBMC strains were hypothesized to mitigate coral bleaching33, making this a crucial trait when selecting pBMC.
Several genes involved in vitamin B-complex biosynthesis were found in our pBMC genomes, such as riboflavin synthase (all pBMC), pyridoxine 5’-phosphate synthase (all pBMC except for pBMC6), biotin synthase (all pBMC), dihydrofolate synthase and thymidylate synthase (all pBMC), and cobalamin synthase (all pBMC except for pBMC3 and pBMC4) for the biosynthesis of vitamins B2, B6, B7, B9, and B12, respectively. Vitamin B2 is necessary for glutathione reductase activity, which is involved in stress reduction by increasing antioxidant potential, and B2 deficiency increases lipid peroxidation99. Vitamin B6 catalyzes approximately 2% of all prokaryotic functions100, but it has not been widely studied in the marine environment101,102. It acts as an antioxidant during light exposure and against oxidative stress103,104. Vitamin B7 is a cofactor in several metabolic pathways, such as fatty acid biosynthesis, amino acid metabolism, and gluconeogenesis105. Vitamin B12 is involved in several metabolic pathways, including the production of the antioxidants glutathione and DMSP106, which are important for neutralizing high concentrations of ROS generated from heat stress events33,43. Bacteria that exist in association with corals possess genes encoding for proteins related to the biosynthesis of essential vitamins, such as B1, B2, and B7, whereas their coral host does not have the capacity to produce them107. This suggests that the coral holobiont can only take up these essential vitamins through heterotrophic feeding and/or from its bacterial symbionts. Furthermore, coral symbionts from the family Symbiodiniaceae are auxotrophs for various B-complex vitamins, which they acquire from exogenous sources such as bacteria85,106, 108,109,110. This highlights the important role of bacterial symbionts in ensuring coral health. For these reasons and because of the close interaction between several B-complex vitamins, the presence of genes encoding proteins related to B-complex vitamin biosynthesis is suggested as a BMC trait.
We also screened for other genes related to metabolism. Siderophore synthase was present in pBMC6. This enzyme produces siderophores that can scavenge iron from the environment, a trait that is beneficial for corals33,84 and other organisms111. In general, the bioavailability of iron in oceans is extremely low; consequently, the growth and survival of organisms that use iron for essential physiological processes, such as photosynthesis and nitrogen fixation, cannot be guaranteed112,113. Thus, bacteria that exist in association with other marine organisms, such as corals and microalgae, produce siderophores to capture and concentrate iron into a bioavailable form that can be used by other organisms114,115 Apart from siderosphere production, we also found that some of our pBMC produced ectoine (pBMC3, pBMC4, and pBMC5) and betaine (pBMC1, pBMC2, pBMC3, pBMC4, and pBMC5), which have been previously described in BMC genomes and proposed as compounds that plays a role in beneficial mechanisms in corals84. Ectoines and betaines are important for osmoregulation and act as protective agents under thermal stress and high irradiance116,115,118. They also contribute to the nitrogen biomass of corals in reefs119. In marine microalgae, the ectoine content was found to increase in the presence of bacteria, highlighting the crucial role of these microorganisms in host health120. Betaine and ectoine production significantly improves environmental stress tolerance, including pH stress121 and heat stress122 in aquatic organisms123, such as corals and their symbionts118. Ectoines can help mitigate the harmful effects of heat stress, high salinity, ROS, and radiation124. Pei et al. identified betaine lipids as leading metabolite drivers for differentiating heat-bleached corals from healthy ones, revealing new tools to screen for heat-resistant corals and their symbionts, such as BMC118,125.
We also found genes involved in the nitrogen cycle, including nitrate reductase (pBMC3, pBMC4, and pBMC5) and cyanate hydratase (pBMC5). The presence of these genes was previously proposed as a BMC trait33,43 because balancing this nutrient’s availability contributes to maintaining desirable levels of bioavailable nitrogen, limiting algal growth and leading to an accumulation of photosynthates in algal cells that, when released, feed the coral host and promote its growth. Additionally, increased coral catabolic activity due to an environmental stressor leads to host starvation and increased nitrogen availability to Symbiodiniaceae members of the holobiont, potentially causing destabilization of the host’s nutrient cycle and of the Symbiodiniaceae–coral interaction10,126.
Screening for genes related to DMSP degradation and sulfur metabolism revealed the presence of DMSP CoA transferase/lyase DddD (pBMC3 and pBMC4) and acryloyl-CoA reductase AcuI/YhdH (BMC1 and pBMC2). DMSP is found in several marine organisms, including Symbiodiniaceae109,127, and is a ROS scavenger in marine algae128, an attribute that has been proposed as a BMC trait due to its antioxidant activity33. Mechanisms of DMSP breakdown have also been hypothesized as BMC traits because a high DMSP concentration can lead to dysbiosis and signal the location of more vulnerable coral to pathogens through chemotaxis18,33, 129.
Discovery of the presence of BMC-associated prophages
Prophages are DNA from bacteriophages (or bacteria-infecting viruses) that are integrated into the genomes of their bacterial host upon infection. They are mainly studied as contributors to the virulence of pathogenic reef bacteria, such as V. coralliilyticus, because they encode virulence factors130. The detection of putative prophages in two of our six pBMC strains revealed the potentially presence of prophages across marine bacteria taxa, and also implies a positive role of prophages associated with beneficial bacteria. As prophages also protect the bacterial host against virulent phage infections via superinfection exclusion131, pBMC containing prophages may have a competitive advantage against other bacteria, including pathogens. Thus, prophages might both expand the metabolic capabilities and protection of beneficial bacteria, and further research is warranted.
Pangenomes exhibit novel pBMC beneficial traits and other applicable functions
We conducted pangenomic analyses that included our pBMC strains and strains closely related to them in the phylogenomic trees to screen for genes unique to our pBMC strains. In the Pseudoalteromonas pangenome, 110 genes with known functions were only present in the pBMC genomes (i.e., they were present in one or both of our P. galatheae pBMC strains). This included a protein involved in chemotaxis and some proteins involved in iron/sulfur metabolism. Chemotaxis may be an important BMC trait because it was previously suggested to play a crucial role in defining patterns of microbial diversity, coral metabolism, coral infection dynamics, and chemical cycling processes, thereby influencing coral holobiont health132. Despite the absence of DMSP-related genes in the genomes of the Pseudoalteromonas sp. pBMC strains compared with the other Pseudoalteromonas sp. strains, the metabolism and production of sulfur compounds has been proposed as a BMC trait because they inhibit the growth of coral pathogens and also play a role in the structure of bacterial communities of the coral holobiont33.
In the Cobetia pangenome, we found five genes with known functions that were only present in the pBMC genomes, but none of them showed a clear benefit to the host. In the Halomonas pangenome, we found 204 genes with known functions that were only present in the pBMC5 genome, inclunding genes involved in carotenoid biosynthesis, antibiotic biosynthesis, and sulfur metabolism. Although not directly related to host health but more to the environment as a whole, the presence of mercury resistance-related genes in the pBMC5 genome is of crucial interest. Heavy metals are known to be environmental toxins due to their bioaccumulation in the food chain, becoming increasingly hazardous for the higher trophic levels133,134 The use of bioremediation for the removal of toxic metals has been studied, but only a few studies were performed in a marine environment135. The trend is similar for mercury-resistant marine bacteria, and the use of mercury-resistant marine bacteria for bioremediation of mercury contamination has received little attention136. However, their use is associated with certain advantages, such as simple process, lower amount of secondary metabolites, and lower cost than commonly used chemical technologies as well as better adaptability and higher resistance to adverse environmental conditions compared with terrestrial bacteria136,137. Some studies have reported that mercury-resistant marine bacteria show a higher capability for mercury bioremediation and can reduce the toxic effects of mercury in contaminated environments136,138.
We found that the Sutcliffiella genome had the highest amount of singleton genes of all the pangenomes generated in this study, with 1333 genes with known functions present only in the pBMC6 genome, including genes encoding proteins related to the biosynthesis of siderophores, carotenoids, antibiotics, vitamins B1, B2 and B12; nitrogen, iron, and sulfur metabolism; chemotaxis; and oxidative stress resistance. Other than the abovementioned genes, we identified a gene that was involved in the detoxification of reactive aldehydes, which are highly reactive organic chemical compounds that mostly arise due to oxidative stress139. We also identified proteins involved in the degradation of aromatic compounds, such as phenols, cresols, catechols, and diphenyl phosphate (DPHP). Phenols and cresols are harmful to the environment140, and catechol bioaccumulation negatively affects the entire ecosystem141. DPHP is used as a chemical additive in numerous industrial products, and because it does not bind with other chemicals, it is easily spread to the environment, where it has been widely detected142,141,144. When in the environment, DPHP has a long half-life and is immunotoxic and neurotoxic to other organisms145. Despite not being comprehensively studied in marine organisms, recent studies have reported its negative effects on fish growth, energy metabolism, and reproduction146,145,146,149. Further investigations are necessary to understand the toxic and negative effects of DPHP on corals, and it may be beneficial to possess the necessary genetic machinery to degrade such harmful compounds. Lastly, we identified a regulatory system that functions under specific stress conditions, such as hypoxia and starvation, and appears to be beneficial to host health and resilience.
pBMC and the importance of their secondary metabolites
Secondary metabolites are important for defense against microorganisms, toxic compounds, and UV radiation as well as essential for symbiotic relationships150; they are abundantly found within the coral holobiont151. Accordingly, we assessed the BGCs and Pfams of our six pBMC strains using the antiSMASH platform, revealing 11 secondary metabolite production clusters: ectoine, beta-lactone, terpene, aryl polyene, lasso peptide, NRPS, nonribosomal peptide metallophores, NI siderophores, opine-like zincophores, class I and IV lanthipeptides, and type I and III PKS. Ectoines are important because they reduce the effects of heat stress, high salinity, ROS, and radiation. Beta-lactone derivatives are extremely diverse, comprising 30 distinct families, many of which have antimicrobial activity152, while others are important elements for antibiotic production150. Terpenes are diverse organic compounds that play a role in defense mechanisms in plants and fungi153,154, serving as antioxidants and protecting cells from oxidative stress155, and their presence indicates production of diverse bioactive compounds156. Some terpenes and carotenoids, such as squalenes, are pigments involved in photosynthesis and signaling. However, they also possess antioxidant activity and can neutralize oxidative stress157,156,159. Aryl polyenes are pigments that are structurally and functionally related to carotenoids and confer protection against photo-oxidative damage and lipid peroxidation160,161. Lasso peptides have antimicrobial and antiviral properties and also show thermal and chemical resistance150,162. NRPSs are sources of newly discovered antibacterial agents that have also been widely studied for their antiviral and anti-inflammatory properties150,163. Microorganisms scavenge metal ions from the environment via metallophores164. Some of these metallophores, such as siderophores like desferrioxamine E, play important roles in biocontrol and bioremediation165,164,167, and are considered beneficial when selecting for BMC33. Zinc is also an essential nutrient for several cellular processes and is taken from the surrounding environment by bacteria168. Similar to iron availability, zinc is also present at low concentrations in the environment and is captured by zincophores (also called opine-like zincophores in bacteria), such as bacillopaline, produced by microorganisms169. Lanthipeptides that possess antimicrobial activity are known as lantibiotics170, but their functions are not limited to this; they may also possess antifungal and antiviral properties177,171. The presence of at least one PKS BGC in each pBMC genome suggests that these bacterial strains have the necessary genomic tools to synthesize various polyketides that are likely to have beneficial properties163.
Red Sea pBMC and their Indo-Pacific Ocean counterparts
Rosado and colleagues26 successfully manipulated the coral microbiome of P. damicornis by adding BMC to coral fragments in a mesocosm setting, and the genomes of these BMC were screened to identify potential beneficial mechanisms and/or traits based on previous literature84. Some of the BMC from the study by Rosado and colleagues26,172 share several gene functions related to putative beneficial traits for corals with some of our pBMC (e.g., superoxide dismutase, glutathione synthetase, catalase-peroxidase, adenosylcobinamide-phosphate synthase, adenosylcobinamide kinase, betaine aldehyde dehydrogenase, choline dehydrogenase, ectoine hydroxylase, L-ectoine synthase, nitrite reductase, and CoA transferase) (Table S3)84. However, when examining the pangenomes of each genus, we noted that our pBMC had a unique set of genes that were absent in the genomes of the BMC from the study by Rosado and colleagues26,172. Moreover, some of these genes represent potential beneficial traits or mechanisms (Figs. 6, 7, 8 and Table 1)26,172.
Our results revealed potential insights into how bacteria help corals during periods of stress. Although some of the abovementioned beneficial traits are hypothetical, others have already been validated in previous studies on the differential expression of genes during heat stress experiments in corals18. Moreover, the identification and classification of these beneficial features depend on and is limited to existing databases. Additionally, relying on cultivation-dependent methods to obtain candidate pBMC introduces an element of chance in discovering and identifying potential candidates. This can be mitigated by obtaining a large number of isolates and meticulously screening and testing them for beneficial traits. Notably, all pBMC examined in this study, even those from the same genus, were distinct and demonstrated potential to contribute to the health and resilience of corals, indicating the need for continued efforts to isolate64,171, explore175,174,178,172,173,174,177 and test173novel pBMC. We also highlight the discovery of prophages associated with two of the BMCs and their potential role in providing competitive advantage to coral probiotics against other bacteria.
Data availability
Scripts that were used in the assembly and evaluation of the study isolate genomes are available at https://zenodo.org/records/10774634. The raw reads and assemblies from this study are publicly available in the European Nucleotide Archive (study accession number PRJEB62849; https://www.ebi.ac.uk/ena/browser/view/PRJEB62849), and the respective sample accession numbers are as follows: (i) Pseudoalteomonas galatheae pBMC1, SAMEA114261728; (ii) Pseudoalteomonas galatheae pBMC2, SAMEA114261729; (iii) Cobetia sp. pBMC3, SAMEA114261730; (iv) Cobetia sp. pBMC4, SAMEA114261731; (v) Halomonas sp. pBMC5, SAMEA114261732; (vi) Sutcliffiella sp. pBMC6, SAMEA114261733. The genome assemblies have also been made available at https://zenodo.org/records/11204102.
References
Hoegh-Guldberg, O. et al. Coral reefs under rapid climate change and ocean acidification. Science 318, 1737–1742 (2007).
Heron, S. F., Maynard, J. A., Van Hooidonk, R. & Eakin, C. M. Warming trends and bleaching stress of the world’s coral reefs 1985–2012. Sci. Rep. 6(1), 38402 (2016).
Hughes, T. P. et al. Coral reefs in the anthropocene. Nature 546(7656), 82–90 (2017).
Knowlton, N. et al. Rebuilding coral reefs: a decadal grand challenge. (2021) https://doi.org/10.53642/NRKY9386.
McCulloch, M. T., Winter, A., Sherman, C. E. & Trotter, J. A. 300 years of sclerosponge thermometry shows global warming has exceeded 1.5 °C. Nat. Clim. Change 14, 171–177 (2024).
Lough, J. M. Small change, big difference: Sea surface temperature distributions for tropical coral reef ecosystems. J. Geophys. Res 117, 9018 (2012).
Duarte, G. A. S. et al. Heat waves are a major threat to turbid coral reefs in Brazil. Front. Mar. Sci. 7, 515393 (2020).
Voolstra, C. R., Peixoto, R. S. & Ferrier‐Pagès, C. Mitigating the ecological collapse of coral reef ecosystems. EMBO Rep. 24(4). (2023) https://doi.org/10.15252/EMBR.202356826.
Muscatine, L. The role of symbiotic algae in carbon and energy flux in reef corals. Coral Reefs: Ecosyst. World 25, 75–87 (1990).
Rädecker, N. et al. Heat stress destabilizes symbiotic nutrient cycling in corals. Proc. Natl. Acad. Sci. USA 118, e2022653118 (2021).
Cui, G. et al. A carbon-nitrogen negative feedback loop underlies the repeated evolution of cnidarian–Symbiodiniaceae symbioses. Nat. Commun. 14, 1–12 (2023).
Rädecker, N., Escrig, S., Spangenberg, J. E., Voolstra, C. R. & Meibom, A. Coupled carbon and nitrogen cycling regulates the cnidarian–algal symbiosis. Nat. Commun. 14, 1–10 (2023).
Baker, A. C., Glynn, P. W. & Riegl, B. Climate change and coral reef bleaching: an ecological assessment of long-term impacts, recovery trends and future outlook. Estuar. Coast Shelf Sci. 80, 435–471 (2008).
Oliver, J. Coral bleaching in space and time climate change, ocean warming and coral bleaching view project representative areas project, great barrier reef marine park authority view project. Coral Bleach.: Patterns, Process., Causes Conseq. 27–49 (2018) https://doi.org/10.1007/978-3-319-75393-5_3.
Burge, C. A. et al. Climate change influences on marine infectious diseases: implications for management and society. Ann. Rev. Mar. Sci. 6, 249–277 (2014).
Cantin, N. E. & Lough, J. M. Surviving coral bleaching events: Porites growth anomalies on the Great Barrier Reef. PLoS One 9, (2014).
Peixoto, R. S. & Voolstra, C. R. The baseline is already shifted: marine microbiome restoration and rehabilitation as essential tools to mitigate ecosystem decline. Front. Mar. Sci. 10, 1218531 (2023).
Santoro, E. P. et al. Coral microbiome manipulation elicits metabolic and genetic restructuring to mitigate heat stress and evade mortality. Sci. Adv. 7 (2021).
Alagely, A., Krediet, C. J., Ritchie, K. B. & Teplitski, M. Signaling-mediated cross-talk modulates swarming and biofilm formation in a coral pathogen Serratia marcescens. ISME J. 5, 1609–1620 (2011).
Sharp, K. H. & Ritchie, K. B. Multi-partner interactions in corals in the face of climate change. Biol. Bull. 223, 66–77 (2012).
Krediet, C. J., Ritchie, K. B., Paul, V. J. & Teplitski, M. Coral-associated micro-organisms and their roles in promoting coral health and thwarting diseases. P. Roy. Soc. B-Biol. Sci. 280, (2013).
Williams, A. D., Brown, B. E., Putchim, L. & Sweet, M. J. Age-related shifts in bacterial diversity in a reef coral. PLoS One 10, e0144902 (2015).
Hernandez-Agreda, A., Leggat, W., Bongaerts, P. & Ainsworth, T. D. The microbial signature provides insight into the mechanistic basis of coral success across reef habitats. mBio 7, (2016).
Welsh, R. M. et al. Alien vs. predator: bacterial challenge alters coral microbiomes unless controlled by Halobacteriovorax predators. PeerJ, e3315 (2017).
Nair, S., Abraham, J., Nair, S. & Abraham, J. Coral reef microbiota and its role in marine ecosystem sustainability. Microb. Interv. Agricult. Environ.: Volume 1: Res. Trends, Prior. Prospects. 453–478 (2019) https://doi.org/10.1007/978-981-13-8391-5_17.
Rosado, P. M. et al. Marine probiotics: Increasing coral resistance to bleaching through microbiome manipulation. ISME J. 13, 921–936 (2019).
Mohamed, A. R., Ochsenkühn, M. A., Kazlak, A. M., Moustafa, A. & Amin, S. A. The coral microbiome: towards an understanding of the molecular mechanisms of coral–microbiota interactions. FEMS Microbiol. Rev. 47, (2023).
Reshef, L., Koren, O., Loya, Y., Zilber-Rosenberg, I. & Rosenberg, E. The coral probiotic hypothesis. Environ. Microbiol. 8, 2068–2073 (2006).
Apprill, A., Marlow, H. Q., Martindale, M. Q. & Rappé, M. S. Specificity of associations between bacteria and the coral Pocillopora meandrina during early development. Appl. Environ. Microbiol. 78, 7467 (2012).
Bourne, D., Iida, Y., Uthicke, S. & Smith-Keune, C. Changes in coral-associated microbial communities during a bleaching event. ISME J. 2, 350–363 (2007).
Ziegler, M. et al. Coral bacterial community structure responds to environmental change in a host-specific manner. Nat. Commun. 10, 1–11 (2019).
Tracy, A. M., Koren, O., Douglas, N., Weil, E. & Harvell, D. C. Persistent shifts in Caribbean coral microbiota are linked to the 2010 warm thermal anomaly. Environ. Microbiol. Rep. 7, 471–479 (2015).
Peixoto, R. S., Rosado, P. M., Leite, D. C. D. A., Rosado, A. S. & Bourne, D. G. Beneficial microorganisms for corals (BMC): Proposed mechanisms for coral health and resilience. Front. Microbiol. 8, 236713 (2017).
Fragoso dos Santos, H. et al. Impact of oil spills on coral reefs can be reduced by bioremediation using probiotic microbiota. Sci. Rep. 5, 1–11 (2015).
Peixoto, R. S., Sweet, M. & Bourne, D. G. Customized medicine for corals. Front. Mar. Sci. 6, 487855 (2019).
Damjanovic, K., Van Oppen, M. J. H., Menéndez, P. & Blackall, L. L. Experimental inoculation of coral recruits with marine bacteria indicates scope for microbiome manipulation in Acropora tenuis and Platygyra daedalea. Front. Microbiol. 10, 452611 (2019).
Morgans, C. A., Hung, J. Y., Bourne, D. G. & Quigley, K. M. Symbiodiniaceae probiotics for use in bleaching recovery. Restor. Ecol. 28, 282–288 (2020).
Doering, T. et al. Towards enhancing coral heat tolerance: A “microbiome transplantation” treatment using inoculations of homogenized coral tissues. Microbiome 9, 1–16 (2021).
Silva, D. P. et al. Multi-domain probiotic consortium as an alternative to chemical remediation of oil spills at coral reefs and adjacent sites. Microbiome 9, 1–19 (2021).
Zhang, Y. et al. Shifting the microbiome of a coral holobiont and improving host physiology by inoculation with a potentially beneficial bacterial consortium. BMC Microbiol. 21, 1–14 (2021).
Li, J. et al. A coral-associated actinobacterium mitigates coral bleaching under heat stress. Environ. Microbiome 18, 1–14 (2023).
Moradi, M. et al. Probiotics mitigate thermal stress- and pathogen-driven impacts on coral skeleton. Front. Mar. Sci. 10, 1212690 (2023).
Peixoto, R. S. et al. Coral probiotics: Premise, promise, Prospects. Annu. Rev. Anim. Biosci. 9, 265–288 (2021).
Delgadillo Ordonez, N. et al. Probiotics reshape the coral microbiome in situ without affecting the surrounding environment. Commun. Biol. 7(1), 434 (2024).
Schwyn, B. & Neilands, J. B. Universal chemical assay for the detection and determination of siderophores. Anal. Biochem. 160, 47–56 (1987).
Nautiyal, C. S. An efficient microbiological growth medium for screening phosphate solubilizing microorganisms. FEMS Microbiol. Lett. 170, 265–270 (1999).
Ben-Haim, Y., Zicherman-Keren, M. & Rosenberg, E. Temperature-regulated bleaching and lysis of the coral Pocillopora damicomis by the novel pathogen Vibrio coralliilyticus. Appl. Environ. Microbiol. 69, 4236–4242 (2003).
Giambiagi-Marval, M., Mafra, M. A., Penido, E. G. C. & Bastos, M. C. F. Distinct groups of plasmids correlated with bacteriocin production in Staphylococcus aureus. J. Gen. Microbiol. 136, 1591–1599 (1990).
Fukasawa, Y., Ermini, L., Wang, H., Carty, K. & Cheung, M. S. LongQC: a quality control tool for third generation sequencing long read data. G3 Genes/Genomes/Genetics 10, 1193–1196 (2020).
Koren, S. et al. Canu: scalable and accurate long-read assembly via adaptive k-mer weighting and repeat separation. Genome Res. 27, 722–736 (2017).
Kolmogorov, M., Yuan, J., Lin, Y. & Pevzner, P. A. Assembly of long, error-prone reads using repeat graphs. Nat. Biotechnol. 37, 540–546 (2019).
Vaser, R. & Šikić, M. Time- and memory-efficient genome assembly with Raven. Nat. Comput. Sci. 1, 332–336 (2021).
Wick, R. R. & Holt, K. E. Benchmarking of long-read assemblers for prokaryote whole genome sequencing. F1000Research 8, 2138 (2019).
Ruan, J. & Li, H. Fast and accurate long-read assembly with wtdbg2. Nat. Methods 17, 155–158 (2020).
Gurevich, A., Saveliev, V., Vyahhi, N. & Tesler, G. QUAST: Quality assessment tool for genome assemblies. Bioinformatics 29, 1072–1075 (2013).
Parks, D. H., Imelfort, M., Skennerton, C. T., Hugenholtz, P. & Tyson, G. W. CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res. 25, 1043–1055 (2015).
Orakov, A. et al. GUNC: Detection of chimerism and contamination in prokaryotic genomes. Genome Biol. 22, 1–19 (2021).
Yoon, S. H., Ha, S. M., Lim, J., Kwon, S. & Chun, J. A large-scale evaluation of algorithms to calculate average nucleotide identity. Antonie Van Leeuwenhoek 110, 1281–1286 (2017).
Meier-Kolthoff, J. P., Carbasse, J. S., Peinado-Olarte, R. L. & Göker, M. TYGS and LPSN: a database tandem for fast and reliable genome-based classification and nomenclature of prokaryotes. Nucleic Acids Res. 50, D801–D807 (2022).
Seemann, T. Prokka: rapid prokaryotic genome annotation. Bioinformatics 30, 2068–2069 (2014).
Olson, R. D. et al. Introducing the Bacterial and Viral Bioinformatics Resource Center (BV-BRC): A resource combining PATRIC IRD and ViPR. Nucleic Acids Res. 51, D678–D689 (2023).
Brettin, T. et al. RASTtk: A modular and extensible implementation of the RAST algorithm for building custom annotation pipelines and annotating batches of genomes. Sci. Rep. 5, 1–6 (2015).
Wishart, D. S. et al. PHASTEST: Faster than PHASTER, better than PHAST. Nucleic Acids Res. 51, W443–W450 (2023).
Guo, J. et al. VirSorter2: a multi-classifier, expert-guided approach to detect diverse DNA and RNA viruses. Microbiome 9, 1–13 (2021).
Nayfach, S. et al. CheckV assesses the quality and completeness of metagenome-assembled viral genomes. Nature Biotechnology 39(5), 578–585 (2020).
Camargo, A. P. et al. IMG/VR v4: an expanded database of uncultivated virus genomes within a framework of extensive functional, taxonomic, and ecological metadata. Nucleic Acids Res. 51, D733–D743 (2023).
Blin, K. et al. antiSMASH 60: Improving cluster detection and comparison capabilities. Nucleic Acids Res. 49, W29–W35 (2021).
Davis, J. J. et al. PATtyFams: Protein families for the microbial genomes in the PATRIC database. Front. Microbiol. 7, 118 (2016).
Edgar, R. C. MUSCLE: A multiple sequence alignment method with reduced time and space complexity. BMC Bioinformatics 5, 1–19 (2004).
Cock, P. J. A. et al. Biopython: freely available Python tools for computational molecular biology and bioinformatics. Bioinformatics 25, 1422–1423 (2009).
Stamatakis, A. RAxML version 8: A tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30, 1312–1313 (2014).
Stamatakis, A., Hoover, P. & Rougemont, J. A rapid bootstrap algorithm for the RAxML web servers. Syst. Biol. 57, 758–771 (2008).
Le, S. Q. & Gascuel, O. An improved general amino acid replacement matrix. Mol. Biol. Evol. 25, 1307–1320 (2008).
Letunic, I. & Bork, P. Interactive tree of life (iTOL) v5: An online tool for phylogenetic tree display and annotation. Nucleic Acids Res. 49, W293–W296 (2021).
Page, A. J. et al. Roary: Rapid large-scale prokaryote pangenome analysis. Bioinformatics 31, 3691–3693 (2015).
Eren, A. M. et al. Community-led, integrated, reproducible multi-omics with anvio. Nat. Microbiol. 6, 3–6 (2020).
Bateman, A. et al. UniProt: The universal protein knowledgebase in 2023. Nucleic Acids Res. 51, D523–D531 (2023).
Meier-Kolthoff, J. P., Göker, M., Spröer, C. & Klenk, H. P. When should a DDH experiment be mandatory in microbial taxonomy?. Arch. Microbiol. 195, 413–418 (2013).
Kim, M., Oh, H. S., Park, S. C. & Chun, J. Towards a taxonomic coherence between average nucleotide identity and 16S rRNA gene sequence similarity for species demarcation of prokaryotes. Int. J. Syst. Evol. Microbiol. 64, 346–351 (2014).
Woo, P. C. Y., Lau, S. K. P., Teng, J. L. L., Tse, H. & Yuen, K.-Y. Then and now: use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories. Clin. Microbiol. Infec. 14, 908–934 (2008).
Gupta, R. S., Patel, S., Saini, N. & Chen, S. Robust demarcation of 17 distinct Bacillus species clades, proposed as novel bacillaceae genera, by phylogenomics and comparative genomic analyses: Description of robertmurraya kyonggiensis sp. nov. and proposal for an emended genus bacillus limiting it only to the members of the subtilis and cereus clades of species. Int. J. Syst. Evol. Microbiol. 70, 5753–5798 (2020).
Rosado, P. M. et al. Exploring the potential molecular mechanisms of interactions between a probiotic consortium and its coral host. mSystems 8(1), e00921-e922 (2023).
Matthews, J. L. et al. Symbiodiniaceae-bacteria interactions: rethinking metabolite exchange in reef-building corals as multi-partner metabolic networks. Env. Microbiol. 22, 1675–1687 (2020).
Hayyan, M., Hashim, M. A. & Alnashef, I. M. Superoxide ion: generation and chemical implications. Chem. Rev. 116, 3029–3085 (2016).
Rochat, T. et al. High-level resistance to oxidative stress in Lactococcus lactis conferred by Bacillus subtilis catalase KatE. Microbiology (NY) 151, 3011–3018 (2005).
Boháčová, V., Zámocký, M., Godočíková, J., Bučková, M. & Polek, B. The expression and diversity of catalases in isolates of genus Comamonas in response to the oxidative stress of a polluted environment. Curr. Microbiol. 53, 430–434 (2006).
Jittawuttipoka, T., Buranajitpakorn, S., Vattanaviboon, P. & Mongkolsuk, S. The catalase-peroxidase KatG is required for virulence of Xanthomonas campestris pv. campestris in a host plant by providing protection against low levels of H2O2. J. Bacteriol. 191, 7372–7377 (2009).
Godobíková, J., Zámocký, M., Bubková, M., Obinger, C. & Polek, B. Molecular diversity of katG genes in the soil bacteria Comamonas. Arch. Microbiol. 192, 175–184 (2010).
Bernroitner, M., Zamocky, M., Furtmüller, P. G., Peschek, G. A. & Obinger, C. Occurrence, phylogeny, structure, and function of catalases and peroxidases in cyanobacteria. J. Exp. Bot. 60, 423–440 (2009).
Kono, Y. & Fridovich, I. Functional significance of manganese catalase in Lactobacillus plantarum. J. Bacteriol. 155, 742 (1983).
Whittaker, J. W. Non-heme manganese catalase—The ‘other’ catalase. Arch. Biochem. Biophys. 525, 111–120 (2012).
Njå Lsson, R. & Norgren, S. Physiological and pathological aspects of GSH metabolism. Acta Paediatr. 94, 132–137 (2005).
Barycki, J. J. et al. Antioxidant molecules and redox cofactors. Redox Biochem. https://doi.org/10.1002/9780470177334.CH2 (2007).
Weis, V. M., Davy, S. K., Hoegh-Guldberg, O., Rodriguez-Lanetty, M. & Pringle, J. R. Cell biology in model systems as the key to understanding corals. Trends Ecol. Evol. 23, 369–376 (2008).
Roberty, S., Furla, P. & Plumier, J. C. Differential antioxidant response between two Symbiodinium species from contrasting environments. Plant Cell Environ. 39, 2713–2724 (2016).
Szabó, M., Larkum, A. W. D. & Vass, I. A review: The role of reactive oxygen species in mass coral bleaching. Adv. Photosynth. Respir. 45, 459–488 (2020).
Ashoori, M. & Saedisomeolia, A. Riboflavin (vitamin B2) and oxidative stress: a review. Brit. J. Nutr. 111(11), 1985–1991 (2014).
Percudani, R. & Peracchi, A. A genomic overview of pyridoxal-phosphate-dependent enzymes. EMBO Rep. 4, 850–854 (2003).
Sañudo-Wilhelmy, S. A. et al. Multiple B-vitamin depletion in large areas of the. Proc. Natl. Acad. Sci. USA 109, 14041–14045 (2012).
Gómez-Consarnau, L. et al. Mosaic patterns of B-vitamin synthesis and utilization in a natural marine microbial community. Environ. Microbiol. 20, 2809–2823 (2018).
Havaux, M. et al. Vitamin B6 deficient plants display increased sensitivity to high light and photo-oxidative stress. BMC Plant Biol. 9, 1–22 (2009).
Vanderschuren, H. et al. Strategies for vitamin B6 biofortification of plants: A dual role as a micronutrient and a stress protectant. Front. Plant Sci. 4, 48827 (2013).
Zempleni, J., Wijeratne, S. S. K. & Hassan, Y. I. Biotin. Biofactors 35, 36–46 (2009).
Croft, M. T., Lawrence, A. D., Raux-Deery, E., Warren, M. J. & Smith, A. G. Algae acquire vitamin B12 through a symbiotic relationship with bacteria. Nature 438, 90–93 (2005).
Hochart, C. et al. Ecology of Endozoicomonadaceae in three coral genera across the Pacific Ocean. Nat. Commun. 14, 1–16 (2023).
Tang, Y. Z., Koch, F. & Gobler, C. J. Most harmful algal bloom species are vitamin B1 and B12 auxotrophs. Proc. Natl. Acad. Sci. USA 107, 20756–20761 (2010).
Robbins, S. J. et al. A genomic view of the reef-building coral Porites lutea and its microbial symbionts. Nat. Microbiol. 4, 2090–2100 (2019).
Pogoreutz, C. et al. Coral holobiont cues prime Endozoicomonas for a symbiotic lifestyle. ISME J. 16, 1883–1895 (2022).
Do Carmo, F. L. et al. Bacterial structure and characterization of plant growth promoting and oil degrading bacteria from the rhizospheres of mangrove plants. J. Microbiol. 49, 535–543 (2011).
Behrenfeld, M. J. et al. Satellite-detected fluorescence reveals global physiology of ocean phytoplankton. Biogeosci. Discuss. 5, 4235–4270 (2008).
Hopkinson, B. M. & Morel, F. M. M. The role of siderophores in iron acquisition by photosynthetic marine microorganisms. BioMetals 22, 659–669 (2009).
Amin, S. A. et al. Photolysis of iron-siderophore chelates promotes bacterial-algal mutualism. Proc. Natl. Acad. Sci. USA 106, 17071–17076 (2009).
Lawson, C. A., Raina, J. B., Kahlke, T., Seymour, J. R. & Suggett, D. J. Defining the core microbiome of the symbiotic dinoflagellate, Symbiodinium. Environ. Microbiol. Rep. 10, 7–11 (2018).
Jebbar, M., von Blohn, C. & Bremer, E. Ectoine functions as an osmoprotectant in Bacillus subtilis and is accumulated via the ABC-transport system OpuC. FEMS Microbiol. Lett. 154, 325–330 (1997).
Shivanand, P. & Mugeraya, G. Halophilic bacteria and their compatible solutes—osmoregulation and potential applications. Curr. Sci. 100, 1516–1521 (2011).
Pei, J. Y. et al. Mass spectrometry–based metabolomic signatures of coral bleaching under thermal stress. Anal. Bioanal. Chem. 414, 7635–7646 (2022).
Ngugi, D. K., Ziegler, M., Duarte, C. M. & Voolstra, C. R. Genomic blueprint of glycine betaine metabolism in coral metaorganisms and their contribution to reef nitrogen budgets. iScience 23, 101120 (2020).
Fenizia, S., Thume, K., Wirgenings, M. & Pohnert, G. Ectoine from bacterial and algal origin is a compatible solute in microalgae. Mar. Drugs 18, 42 (2020).
Van-Thuoc, D., Hashim, S. O., Hatti-Kaul, R. & Mamo, G. Ectoine-mediated protection of enzyme from the effect of pH and temperature stress: A study using Bacillus halodurans xylanase as a model. Appl. Microbiol. Biotechnol. 97, 6271–6278 (2013).
Teixidó, N. et al. Accumulation of the compatible solutes, glycine–betaine and ectoine, in osmotic stress adaptation and heat shock cross-protection in the biocontrol agent Pantoea agglomerans CPA-2. Lett. Appl. Microbiol. 41, 248–252 (2005).
Bownik, A., Stępniewska, Z. & Skowroński, T. Protective effects of ectoine on heat-stressed Daphnia magna. J. Comp. Physiol. B 184, 961–976 (2014).
Pastor, J. M. et al. Ectoines in cell stress protection: uses and biotechnological production. Biotechnol. Adv. 28, 782–801 (2010).
Roach, T. N. F. et al. Metabolomic signatures of coral bleaching history. Nat. Ecol. Evol. 5, 495–503 (2021).
Rädecker, N. et al. Heat stress reduces the contribution of diazotrophs to coral holobiont nitrogen cycling. ISME J. 16, 1110–1118 (2021).
Yost, D. M., Jones, R. J. & Mitchelmore, C. L. Alterations in dimethylsulfoniopropionate (DMSP) levels in the coral Montastraea franksi in response to copper exposure. Aquat. Toxicol. 98, 367–373 (2010).
Sunda, W., Kieber, D. J., Kiene, R. P. & Huntsman, S. An antioxidant function for DMSP and DMS in marine algae. Nature 418, 317–320 (2002).
Garren, M. et al. A bacterial pathogen uses dimethylsulfoniopropionate as a cue to target heat-stressed corals. ISME J. 8, 999–1007 (2013).
Weynberg, K. D., Voolstra, C. R., Neave, M. J., Buerger, P. & Van Oppen, M. J. H. From cholera to corals: viruses as drivers of virulence in a major coral bacterial pathogen. Sci. Rep. 5, 1–9 (2015).
Bondy-Denomy, J. et al. Prophages mediate defense against phage infection through diverse mechanisms. ISME J. 10, 2854–2866 (2016).
Tout, J. et al. Chemotaxis by natural populations of coral reef bacteria. ISME 9, 1764–1777 (2015).
Monperrus, M. et al. Determination of metal and organometal trophic bioaccumulation in the benthic macrofauna of the Adour estuary coastal zone (SW France, Bay of Biscay). J. Environ. Monitor. 7, 693–700 (2005).
Fang, T. H., Hwang, J. S., Hsiao, S. H. & Chen, H. Y. Trace metals in seawater and copepods in the ocean outfall area off the northern Taiwan coast. Mar. Environ. Res. 61, 224–243 (2006).
Das, N., Vimala, R. & Karthika, P. Biosorption of heavy metals - an overview. Indian J. Biotechnol. 7, 159–169 (2008).
Dash, H. R., Mangwani, N. & Das, S. Characterization and potential application in mercury bioremediation of highly mercury-resistant marine bacterium Bacillus thuringiensis PW-05. Nviron. Sci. Pollut. R. 21, 2642–2653 (2014).
Singh, S., Kang, S. H., Mulchandani, A. & Chen, W. Bioremediation: Environmental clean-up through pathway engineering. Curr. Opin. Biotechnol. 19, 437–444 (2008).
Zhang, W., Chen, L. & Liu, D. Characterization of a marine-isolated mercury-resistant Pseudomonas putida strain SP1 and its potential application in marine mercury reduction. Appl. Microbiol. Biotechnol. 93, 1305–1314 (2012).
Fritz, K. S. & Petersen, D. R. An overview of the chemistry and biology of reactive aldehydes. Free Radic. Biol. Med. 59, 85–91 (2013).
Duan, W. et al. Ecotoxicity of phenol and cresols to aquatic organisms: a review. Ecotoxicol. Environ. Saf. 157, 441–456 (2018).
Paasivirta, J. et al. Polychlorinated phenols, guaiacols and catechols in environment. Chemosphere 14, 469–491 (1985).
Hoffman, K., Garantziotis, S., Birnbaum, L. S. & Stapleton, H. M. Monitoring indoor exposure to organophosphate flame retardants: hand wipes and house dust. Environ. Health Perspect. 123, 160–165 (2014).
Zhao, F. et al. Levels of blood organophosphorus flame retardants and association with changes in human sphingolipid homeostasis. Environ. Sci. Technol. 50(16), 8896–8903 (2016).
Yan, Z. et al. In vitro metabolic kinetics of cresyl diphenyl phosphate (CDP) in liver microsomes of crucian carp (Carassius carassius). Environ. Pollut. 274, 116586 (2021).
Ren, R. et al. Identification of a novel glycerophosphodiester phosphodiesterase from Bacillus altitudinis W3 and its application in degradation of diphenyl phosphate. 3 Biotech 11, 1–9 (2021).
Zhu, Y. et al. Environmentally relevant concentrations of the flame retardant tris(1,3-dichloro-2-propyl) phosphate inhibit growth of female zebrafish and decrease fecundity. Environ. Sci. Technol. 49, 14579–14587 (2015).
Li, Y. et al. Environmentally relevant concentrations of the organophosphorus flame retardant triphenyl phosphate impaired testicular development and reproductive behaviors in japanese medaka (Oryzias latipes). Environ. Sci. Technol. Lett. 5, 649–654 (2018).
Chen, Q. et al. Life Cycle exposure to environmentally relevant concentrations of diphenyl phosphate (DPhP) inhibits growth and energy metabolism of zebrafish in a sex-specific manner. Environ. Sci. Technol. 55, 13122–13131 (2021).
Yang, R. et al. Insights into the sex-dependent reproductive toxicity of 2-ethylhexyl diphenyl phosphate on zebrafish (Danio rerio). Environ. Int. 158, 106928 (2022).
Cho, S. H. et al. Elucidation of the biosynthetic pathway of vitamin b groups and potential secondary metabolite gene clusters via genome analysis of a marine bacterium Pseudoruegeria sp. M32A2M. J. Microbiol. Biotechnol. 30, 505–514 (2020).
Modolon, F., Barno, A. R., Villela, H. D. M. & Peixoto, R. S. Ecological and biotechnological importance of secondary metabolites produced by coral-associated bacteria. J. Appl. Microbiol. 129, 1441–1457 (2020).
Robinson, S. L., Christenson, J. K. & Wackett, L. P. Biosynthesis and chemical diversity of β-lactone natural products. Nat. Prod. Rep. 36, 458–475 (2019).
Yamada, Y. et al. Terpene synthases are widely distributed in bacteria. Proc. Natl. Acad. Sci. USA 112, 857–862 (2015).
Oldfield, E. & Lin, F. Y. Terpene biosynthesis: modularity rules. Angew. Chem. Int. Ed. Engl. 51, 1124–1137 (2012).
Baccouri, B. & Rajhi, I. Potential antioxidant activity of terpenes. Terpenes Terpenoids-Recent Adv. 53–62 (2021).
Cheng, A. X. et al. Plant terpenoids: Biosynthesis and ecological functions. J. Integr. Plant Biol. 49, 179–186 (2007).
Stahl, W. & Sies, H. Antioxidant activity of carotenoids. Mol. Aspects Med. 24, 345–351 (2003).
Spanova, M. & Daum, G. Squalene—biochemistry, molecular biology, process biotechnology, and applications. Eur. J. Lipid Sci. Tech. 113, 1299–1320 (2011).
Saini, R. K. et al. Bioactive compounds of citrus fruits: a review of composition and health benefits of carotenoids, flavonoids, limonoids, and terpenes. Antioxidants 11, 239 (2022).
Rajagopal, L., Sundari, C. S., Balasubramanian, D. & Sonti, R. V. The bacterial pigment xanthomonadin offers protection against photodamage. FEBS Lett. 415, 125–128 (1997).
Schöner, T. A. et al. Aryl polyenes, a highly abundant class of bacterial natural products, are functionally related to antioxidative carotenoids. ChemBioChem 17, 247–253 (2016).
Zhu, S. et al. Insights into the unique phosphorylation of the lasso peptide paeninodin. J. Biol. Chem. 291, 13662–13678 (2016).
Raimundo, I., Silva, S. G., Costa, R. & Keller-Costa, T. Bioactive secondary metabolites from octocoral-associated microbes-new chances for blue growth. Mar. Drugs 16, (2018).
Reitz, Z. L., Butler, A. & Medema, M. H. Automated genome mining predicts combinatorial diversity and taxonomic distribution of peptide metallophore structures. bioRxiv 2022.12.14.519525 (2022).
Tam, T. et al. Iron Chelator research: Past, present, and future. Curr. Med. Chem. 10, 983–995 (2005).
Johnstone, T. C. & Nolan, E. M. Beyond iron: non-classical biological functions of bacterial siderophores. Dalton T 44, 6320–6339 (2015).
Soares, E. V. Perspective on the biotechnological production of bacterial siderophores and their use. Appl. Microbiol. Biot. 106, 3985–4004 (2022).
Andreini, C., Bertini, I., Cavallaro, G., Holliday, G. L. & Thornton, J. M. Metal ions in biological catalysis: From enzyme databases to general principles. J. Biol. Inorg. Chem. 13, 1205–1218 (2008).
Morey, J. R. & Kehl-Fie, T. E. Bioinformatic mapping of opine-like zincophore biosynthesis in bacteria. mSystems 5, (2020).
Willey, J. M. & Van Der Donk, W. A. Lantibiotics: peptides of diverse structure and function. Annu. Rev. Microbiol. 61, 477–501 (2007).
Férir, G. et al. The lantibiotic peptide labyrinthopeptin a1 demonstrates broad anti-HIV and anti-HSV activity with potential for microbicidal applications. PLoS One 8, e64010 (2013).
Mohr, K. I. et al. Pinensins: the first antifungal lantibiotics. Angew. Chem. Int. Edit. 54, 11254–11258 (2015).
Schultz, J. et al. Methods and strategies to uncover coral-associated microbial dark matter. mSystems 7, (2022).
(2023) In situ devices can culture the microbial dark matter of corals iScience 26(12) 108374-10.1016/j.isci.2023.108374
Sweet, M. et al. Insights into the cultured bacterial fraction of corals. mSystems 6, (2021).
Doering, T. et al. Genomic exploration of coral-associated bacteria: identifying probiotic candidates to increase coral bleaching resilience in Galaxea fascicularis. Microbiome 11, (2023).
Maire, J. et al. Functional potential and evolutionary response to long-term heat selection of bacterial associates of coral photosymbionts. mSystems 8, (2023).
Peixoto, R. S. et al. Harnessing the microbiome to prevent global biodiversity loss. Nat. Microbiol. 7, 1726–1735 (2022).
Garcias-Bonet, N. et al. Horizon scanning the application of probiotics for wildlife. Trends Microbiol. 0, (2023).
Carver, T., Thomson, N., Bleasby, A., Berriman, M. & Parkhill, J. DNAPlotter: circular and linear interactive genome visualization. Bioinformatics 25, 119–120 (2009).
Acknowledgements
The authors would like to thank the King Abdullah University of Science and Technology (KAUST) Bioscience Core Lab (BCL), Thuwal, Saudi Arabia, for allowing sequencing of genomes and use of the resources at the KAUST Advanced Computing Core Lab for performing some of the computational analyses.
Funding
R.S.P. was supported through funding provided by KAUST grant numbers BAS/1/1095–01-01 and FCC/1/1976–40-01.
Author information
Authors and Affiliations
Contributions
R.S.P. and I.R. conceived the study. IR performed the molecular work. C.P.A. performed the initial processing of the sequencing data. I.R., P.M.R., and A.R.B. performed the subsequent analyses and interpreted the data. R.S.P. also discussed the data interpretation and provided financial support. I.R. and R.S.P. wrote the manuscript. All the authors contributed to the final manuscript and have read and accepted the submission.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing interests.
Additional information
Publisher's note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Rights and permissions
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.
About this article
Cite this article
Raimundo, I., Rosado, P.M., Barno, A.R. et al. Unlocking the genomic potential of Red Sea coral probiotics. Sci Rep 14, 14514 (2024). https://doi.org/10.1038/s41598-024-65152-8
Received:
Accepted:
Published:
DOI: https://doi.org/10.1038/s41598-024-65152-8
- Springer Nature Limited