Diseased Corals Research Articles

Disease prevalence and bacterial isolates associated with Acropora palmata in the Colombian Caribbean

The decline in Acropora palmata populations in Colombian reefs has been mainly attributed to diseases outbreaks. The population size structure and prevalence of white pox and white band disease were evaluated in six localities of the Colombian Caribbean. Here, we aimed to isolate enteric bacteria and Vibrios from healthy and diseased coral mucus to relate its presence to the health status of Acropora palmata. The isolated bacteria were identified using molecular analyses with the 16S rRNA gene. Larger colonies had the highest percentage of the prevalence of both diseases. The strains that were identified as Vibrio sp. and Bacillus sp. were common in the healthy and diseased mucus of the holobiont. The Exiguobacterium sp. and Cobetia sp. strains isolated from diseased mucus may indicate maintenance and resilience mechanisms in the coral. Enterococcus sp. and other bacteria of the Enterobacteriaceae family were isolated from some localities, suggesting that probably contamination due to poor treatment of domestic wastewater and contributions from river discharges can affect coral health. The spatial heterogeneity of Colombian coral reefs exhibited variability in the bacteria, wherein environmental alterations can trigger signs of disease.

Temporal patterns in coral disease prevalences at Thudufushi Island, Maldives, 2010-2022.

Coral reefs are lately suffering a fast decline in biodiversity due to the coupled effect of climate change and disease outbreaks, which in recent decades have been reported with higher frequency and shorter intervals. Limited studies have been conducted on coral diseases in the Maldives resulting in the impossibility of assessing the temporal trend in their dynamics. In this context, we evaluated the change in the distribution, prevalence, and host range of 4 diseases, namely black band disease (BBD), brown band disease (BrB), skeletal eroding band (SEB) and white syndrome (WS), in the reef system around Thudufushi Island after an interval of 12 yr since the last assessment. In this period, the overall disease prevalence increased, except for BrB, with SEB showing the most severe increase in 2022 in comparison to 2010. The overall average prevalence of coral diseases is approximately 2%, indicating an increase of about 0.7% since 2010. Diseased coral colonies were found in all the investigated sites, with the east site being the most affected and SEB emerging as the most prevalent disease across all the investigated sites. The affected colonies belong to 13 genera, with Psammocora genus showing the highest overall mean disease prevalence. This study depicted a basic temporal trend in disease prevalence that confirms an increase in coral diseases in the region and calls for a dedicated national monitoring protocol to better understand and predict future coral disease dynamics at regional scales.

Characterization of the defense properties of healthy and diseased coral mucus

The surface mucus layer of corals aids in feeding, silt removal, and defense against pathogens. However, first line of defense of secreted mucus of corals against tissue damage generated through pathogen or environmental factors is poorly understood. Hence, we used various methods such as a well diffusion assay and tests for quorum quenching, free radical scavenging, antioxidant enzyme expression and phenoloxidase (PO) activity to determine the mucus defense properties using mucus of healthy and diseased Porites sp. and Acropora sp. Interestingly the coral mucus showed antimicrobial activity against coral pathogens such as bacteria and protozoan ciliates. Inhibition of the N-Acyl homoserine lactone (AHL) molecule suggests quorum quenching. Free radical scavenging of mucus was screened using hydrogen peroxide, hydroxyl radical, and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) methods, which was found significantly more in diseased corals (p < 0.05). Antioxidant enzymes superoxide dismutase (SOD), catalase, and peroxidase activity were observed in both the diseased and healthy coral mucus. The presence of serine and metalloproteases was also detected in coral mucus. Further, phenoloxidase (PO) activity was highest in diseased coral mucus affected by pink line syndrome and white patch Acropora sp. disease than the healthy coral mucus. Thus, the present study of antimicrobial properties, antioxidant enzymes, and quorum quenching properties in coral mucus may aid in understanding the corals defense and survival against pathogens and any abiotic stress.

Molecular Cartography of a Hawaiian Coral Assemblage

Coral reefs are declining due to multiple factors including overfishing, anthropogenic pollution, and ocean acidification. Diseases affecting corals have increased in recent decades, yet the etiology of nearly all diseases remains poorly understood. Here, we investigated three-dimensionally mapped molecules and microbes from healthy and diseased coral tissue sampled across the landscape of a coral assemblage at the Wai‘ōpae tide pools, Southeast Hawai‘i Island. A 3D molecular cartographic platform was used in combination with molecular networking tools to characterize healthy coral tissue and tissue affected by the disease growth anomaly (GA). Tissues of healthy Montipora flabellata and Montipora capitata exhibited higher microbial diversity compared to Porites lobata and GA-affected M. capitata corals. Increases in relative abundance of Ulvophyceae and sterols were observed in GA lesions, while chlorophyll decreased. Conversely, healthy coral tissues were characterized by the presence of cyanobacteria in the order of Stramenopiles, in addition to higher relative chlorophyll levels. Leveraging innovative molecular cartography provides new insight into molecular characteristics of coral colonies, and helps to better understand how diseases affect the molecular landscape of corals.

Bacterial Communities Associated with Healthy and Diseased Corals during a Heatwave Event in the Northern Red Sea, Egypt

Bacterial Communities Associated with Healthy and Diseased Corals during a Heatwave Event in the Northern Red Sea, Egypt

Quantitative detection of Vibrio alginolyticus strain XSBZ14 by a newly developed RT-PCR method

PurposeCoral degradation is a worldwide ecological problem. Bacterial diseases are a great danger to coral health. The pathogenic bacterium Vibrio alginolyticus XSBZ14 isolated from diseased coral had been identified as the pathogenic bacterium of Porites andrewsi White syndrome (PAWS) in Xisha Archipelago on transmission experiment. To date, the molecular mechanism by which this pathogen causes disease is unknown, and molecular diagnostics for diseases caused by this bacterium have not been developed. In an effort to restore damaged coral ecosystems in the South China Sea, efforts are underway to transplant flat-branch shore corals. There is therefore an urgent need to further develop specific and rapid detection methods for V. alginolyticus XSBZ14 in order to prevent this epidemic and ensure the successful implementation of compilation transplants.MethodsAt first, a low sequence identity single-copy sequence S2 was selected from the genome by in-house Perl script. Using the designed specific primers, four different types of standard curves were subsequently plotted for the accurate quantification of the strain XSBZ14 in four different samples (DNA, bacterial suspension, coral tissue, seawater). Then, use the strain to infect the Galaxea fascicularis and test the strain in the coral culture water during the week.ResultsThe rapid detection method of pathogenic bacteria by RT-PCR was established. The limit of detection (LOD) of the RT-PCR was 0.88 pg/reaction (0.44 pg/μL) in DNA, 2 CFU/reaction (1000 CFU/mL) in bacterial suspension, 2 CFU/reaction in coral tissue, and 20 CFU/reaction in seawater for the strain XSBZ14, respectively. In addition, according to the detection results of the RT-PCR, the strain XSBZ14 could survive in Galaxea fascicularis for a week, and the strain could also be detected from its reared seawater.ConclusionThese results indicated that the RT-PCR detection method of a coral pathogenic strain XSBZ14 was established. The method is a robust tool for the rapid detection and quantification of the coral pathogen, XSBZ14, and is very useful for PAWS epidemiological survey and specific pathogen-free coral transplantation in the South China Sea. And other coral species and their habitats might act as an important reservoir for the strain XSBZ14 and mediated its horizontal transmission in coral reefs.

Concordance of microbial and visual health indicators of white-band disease in nursery reared Caribbean coral Acropora cervicornis.

Coral diseases are one of the leading causes of declines in coral populations. In the Caribbean, white band disease (WBD) has led to a substantial loss of Acropora corals. Although the etiologies of this disease have not been well described, characterizing the coral microbiome during the transition from a healthy to diseased state is critical for understanding disease progression. Coral nurseries provide unique opportunities to further understand the microbial changes associated with diseased and healthy corals, because corals are monitored over time. We characterized the microbiomes before and during an outbreak of WBD in Acropora cervicornis reared in an ocean nursery in Little Cayman, CI. We asked (1) do healthy corals show the same microbiome over time (before and during a disease outbreak) and (2) are there disease signatures on both lesioned and apparently healthy tissues on diseased coral colonies? Microbial mucus-tissue slurries were collected from healthy coral colonies in 2017 (before the disease) and 2019 (during the disease onset). Diseased colonies were sampled at two separate locations on an individual coral colony: at the interface of Disease and ∼10 cm away on Apparently Healthy coral tissue. We sequenced the V4 region of the 16S rRNA gene to characterize bacterial and archaeal community composition in nursery-reared A. cervicornis. We assessed alpha diversity, beta diversity, and compositional differences to determine differences in microbial assemblages across health states (2019) and healthy corals between years (2017 and 2019). Microbial communities from healthy A. cervicornis from 2017 (before disease) and 2019 (after disease) did not differ significantly. Additionally, microbial communities from Apparently Healthy samples on an otherwise diseased coral colony were more similar to Healthy colonies than to the diseased portion on the same colony for both alpha diversity and community composition. Microbial communities from Diseased tissues had significantly higher alpha diversity than both Healthy and Apparently Healthy tissues but showed no significant difference in beta-diversity dispersion. Our results show that at the population scale, Healthy and Apparently Healthy coral tissues are distinct from microbial communities associated with Diseased tissues. Furthermore, our results suggest stability in Little Cayman nursery coral microbiomes over time. We show healthy Caymanian nursery corals had a stable microbiome over a two-year period, an important benchmark for evaluating coral health via their microbiome.

Latent Dirichlet Allocation modeling of environmental microbiomes.

Interactions between stressed organisms and their microbiome environments may provide new routes for understanding and controlling biological systems. However, microbiomes are a form of high-dimensional data, with thousands of taxa present in any given sample, which makes untangling the interaction between an organism and its microbial environment a challenge. Here we apply Latent Dirichlet Allocation (LDA), a technique for language modeling, which decomposes the microbial communities into a set of topics (non-mutually-exclusive sub-communities) that compactly represent the distribution of full communities. LDA provides a lens into the microbiome at broad and fine-grained taxonomic levels, which we show on two datasets. In the first dataset, from the literature, we show how LDA topics succinctly recapitulate many results from a previous study on diseased coral species. We then apply LDA to a new dataset of maize soil microbiomes under drought, and find a large number of significant associations between the microbiome topics and plant traits as well as associations between the microbiome and the experimental factors, e.g. watering level. This yields new information on the plant-microbial interactions in maize and shows that LDA technique is useful for studying the coupling between microbiomes and stressed organisms.

Photo-physiology of healthy-looking and diseased/health-compromised hard corals from Mauritius Island, Western Indian Ocean

Abstract. Jogee SY, Jeetun S, Ricot M, Taleb-Hossenkhan N, Mattan-Moorgawa S, Kaullysing D, Riemann P, Blanc L, Casareto BE, Suzuki Y, Bhagooli R. 2023. Photo-physiology of healthy-looking and diseased/health-compromised hard corals from Mauritius Island, Western Indian Ocean. Indo Pac J Ocean Life 7: 27-37. The spatial photo-physiological responses of in hospite zooxanthellae in hard corals, including coenosarc and polyps, healthy-looking and affected parts in four coral diseases, namely Brown Band, Black Band, Skeletal Eroding Band and White Band on the coral Acropora muricata, and two health-compromised conditions such as the Pink Pigmentation Response and its differentiated morphology, the Pink Line Syndrome, on the coral Porites were investigated using the Imaging-PAM fluorometry. A significantly lower Fv/Fm was observed in case of Black Band, White Band, Brown Band and Pink Pigmentation Response affected parts compared to the healthy-looking parts. The Fv/Fm had the highest decline in Brown Band disease. Both the polyps and coenosarc had significantly lower Fv/Fm in White Band and Brown Band diseased parts compared to their healthy-looking parts. The rETRmax did not change significantly between diseased/health-comprised parts and healthy-looking parts. NPQmax declined significantly in White Band, Black Band and Pink Pigmentation Response cases. ? and ? generally did not tend to be affected in diseased/health-compromised conditions. The photo-physiology of in hospite zooxanthellae was least affected in Pink Line Syndrome. These findings suggest that diseased/health-compromised parts of corals behave differently in terms of their photo-physiology in different diseased and health-compromised coral conditions in important reef-building corals species such as A. muricata and Porites species, with important implications for the productivity and thus adaptive management of coral reefs in a globally warming ocean.

Identification of Vibrio ponticus as a bacterial pathogen of coral trout Plectropomus leopardus.

Vibrio ponticus is a vital pathogen with potential danger for aquaculture animals. Yet V. ponticus pathogenic to the coral trout Plectropomus leopardus is still unknown. In this study, a virulent bacterial strain, temporarily named DX2, was isolated from diseased coral trout suffering liver necrosis with cell vacuolar degeneration, and was identified molecularly and phenotypically as V. ponticus. Besides, the DX2 isolate showed an LD50 value of 6.64×105 CFU mL-1, developed multiple resistances to cephalosporins, macrolides, penicillins, peptides, and sulfonamides antimicrobials, and was highly susceptible to doxycycline and florfenicol in aquaculture use. To the best of our knowledge, this is the first report of the pathogenicity of V. ponticus to the coral trout, and the findings provide a reference for the control of pathogenic V. ponticus in the coral trout.

Biofilms as potential reservoirs of stony coral tissue loss disease

Since 2014, corals throughout Florida’s Coral Reef have been plagued by an epizootic of unknown etiology, colloquially termed stony coral tissue loss disease (SCTLD). Although in Florida the movement of this waterborne coral disease has been consistent with natural transport via water currents, outbreaks in the Caribbean have been more sporadic, with infections occurring in locations inconsistent with spread via natural means. Often Caribbean outbreaks have been clustered near ports, potentially implicating ships as mediators of SCTLD into new regions. Biofilms attached to ship hulls, ballast tank walls, or other surfaces could represent a possible vector for the disease. We investigated whether bacteria shed by healthy and SCTLD-diseased corals would form distinct biofilms, and whether a SCTLD signal would be detectable within biofilm bacterial communities. Stainless steel plates serving as proxies for ship hulls, ballast tank walls, and other colonizable surfaces were incubated for three days in filtered seawater mesocosms containing healthy or SCTLD-infected corals. Resulting biofilm bacterial communities were characterized through sequencing of the V4 region of the 16S rRNA gene. We determined that bacteria shed by healthy and diseased corals formed significantly different biofilms consisting of highly diverse taxa. Comparison with 16S data from previous SCTLD investigations spanning different coral species, collection locations, years, and source material revealed the presence of numerous genetically identical sequences within the biofilm bacterial communities formed during exposure to SCTLD-infected corals, including several previously identified as possible SCTLD bioindicators. These results suggest ship-associated biofilms may have the potential to be vectors for the transmission of SCTLD into new regions.

Transmission of stony coral tissue loss disease (SCTLD) in simulated ballast water confirms the potential for ship-born spread

Stony coral tissue loss disease (SCTLD) remains an unprecedented epizootic disease, representing a substantial threat to the persistence and health of coral reef ecosystems in the Tropical Western Atlantic since its first observation near Miami, Florida in 2014. In addition to transport between adjacent reefs indicative of waterborne pathogen(s) dispersing on ocean currents, it has spread throughout the Caribbean to geographically- and oceanographically-isolated reefs, in a manner suggestive of ship and ballast water transmission. Here we evaluate the potential for waterborne transmission of SCTLD including via simulated ballast water, and test the efficacy of commonly-used UV radiation treatment of ballast water. Two species of reef-building corals (Orbicella faveolata and Pseudodiploria strigosa) were subjected to (1) disease-exposed or UV-treated disease-exposed water, and (2) a ballast hold time series of disease-exposed water in two carefully-controlled experiments to evaluate transmission. Our experiments demonstrated transmission of SCTLD through water, rather than direct contact between diseased and healthy corals. While UV treatment of disease-exposed water led to a 50% reduction in the number of corals exhibiting disease signs in both species, the statistical risk of transmission and volume of water needed to elicit SCTLD lesions remained similar to untreated disease-exposed water. The ballast hold time (24 h vs. 120 h) did not have a significant effect on the onset of visible disease signs for either species, though there appeared to be some evidence of a concentration effect for P. strigosa as lesions were only observed after the 120 h ballast hold time. Results from both experiments suggest that the SCTLD pathogens can persist in both untreated and UV-treated ballast water and remain pathogenic. Ballast water may indeed pose a threat to the continued spread and persistence of SCTLD, warranting further investigation of additional ballast water treatments and pathogen detection methods.

Prevalence of differential microbiome in healthy, diseased and nipped colonies of corals, Porites lutea in the Gulf of Kachchh, north-west coast of India

Coral reefs are constantly subjected to multiple stresses like diseases and fish predation, which can profoundly influence the coral microbiome. This study investigated the differences in bacterial community structure of healthy, white syndrome affected and blenny nipped coral colonies of Porites lutea, collected from the coral reefs of Gulf of Kachchh, north-west coast of India. Present study observed that the stressed coral colonies harbored more OTUs and contained higher diversity values compared to healthy corals colonies. Similarly, beta diversity analysis indicated the dissimilarities among the three coral samples analyzed. Though the taxonomy analysis indicated bacterial phyla like Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria among the entire coral samples studied, there was a variation in their relative abundances. Huge variations were observed in the relative dominance at the bacterial genera level. About 13phyla and 11 genera was identified in healthy coral. The PBN sample was found to contain Proteobacteria, Cyanobacteria, Verrucomicrobia, and Lentisphaerae as dominant phyla and Endozoicomonas, Dyella, Woeseia, and Winogradskyella as dominant genera. The PWS sample contained Proteobacteria, Lentisphaerae, Spirochaetes, and Tenericutes as dominant phyla and Endozoicomonas, Arcobacter, Sunxiuqinia, and Carboxylicivirgia as dominant genera. Among the healthy samples, sequences belonging to Uncultured Rhodospirillaceae were dominant, while Woeseia and sequences belonging to Uncultured Rhodovibrionaceae were dominant among the blenny nipped white syndrome infected corals. Although any previously established pathogen was not identified, present study revealed the presence of a potentially pathogenic bacterium, Arcobacter, among the diseased corals. It also demonstrated a dynamic microbiome among the Porites lutea colonies on subjecting to various stresses.

The coral pathogen Vibrio coralliilyticus kills non-pathogenic holobiont competitors by triggering prophage induction

The coral reef microbiome is central to reef health and resilience. Competitive interactions between opportunistic coral pathogens and other commensal microbes affect the health of coral. Despite great advances over the years in sequencing-based microbial profiling of healthy and diseased coral, the molecular mechanism underlying colonization competition has been much less explored. In this study, by examining the culturable bacteria inhabiting the gastric cavity of healthy Galaxea fascicularis, a scleractinian coral, we found that temperate phages played a major role in mediating colonization competition in the coral microbiota. Specifically, the non-toxigenic Vibrio sp. inhabiting the healthy coral had a much higher colonization capacity than the coral pathogen Vibrio coralliilyticus, yet this advantage was diminished by the latter killing the former. Pathogen-encoded LodAB, which produces hydrogen peroxide, triggers the lytic cycle of prophage in the non-toxicogenic Vibrio sp. Importantly, V. coralliilyticus could outcompete other coral symbiotic bacteria (for example, Endozoicomonas sp.) through LodAB-dependent prophage induction. Overall, we reveal that LodAB can be used by pathogens as an important weapon to gain a competitive advantage over lysogenic competitors when colonizing corals.

Experimental transmission of Stony Coral Tissue Loss Disease results in differential microbial responses within coral mucus and tissue

Stony coral tissue loss disease (SCTLD) is a widespread and deadly disease that affects nearly half of Caribbean coral species. To understand the microbial community response to this disease, we performed a disease transmission experiment on US Virgin Island (USVI) corals, exposing six species of coral with varying susceptibility to SCTLD. The microbial community of the surface mucus and tissue layers were examined separately using a small subunit ribosomal RNA gene-based sequencing approach, and data were analyzed to identify microbial community shifts following disease acquisition, potential causative pathogens, as well as compare microbiota composition to field-based corals from the USVI and Florida outbreaks. While all species displayed similar microbiome composition with disease acquisition, microbiome similarity patterns differed by both species and mucus or tissue microhabitat. Further, disease exposed but not lesioned corals harbored a mucus microbial community similar to those showing disease signs, suggesting that mucus may serve as an early warning detection for the onset of SCTLD. Like other SCTLD studies in Florida, Rhodobacteraceae, Arcobacteraceae, Desulfovibrionaceae, Peptostreptococcaceae, Fusibacter, Marinifilaceae, and Vibrionaceae dominated diseased corals. This study demonstrates the differential response of the mucus and tissue microorganisms to SCTLD and suggests that mucus microorganisms may be diagnostic for early disease exposure.

The Influence of Foureye Butterflyfish (Chaetodon capistratus) and Symbiodiniaceae on the Transmission of Stony Coral Tissue Loss Disease

Marine diseases have caused large scale decreases in coral cover across the Caribbean and are unfortunately projected to increase as sea surface temperatures rise. Identifying the main drivers of disease transmission is essential for our understanding and response to diseases in the future. This study investigates the effects of direct-contact, waterborne, Symbiodiniaceae composition, and butterflyfish foraging on the transmission rates of stony coral tissue loss disease (SCTLD) across Montastraea cavernosa (MCAV) coral fragments. M. cavernosa fragments were placed in direct contact with diseased Orbicella and Montastraea colonies with the purpose of creating disease MCAV fragments for experimental trials with butterflyfish. Three treatments were used to investigate waterborne transmission (control), mechanically injury to a healthy coral (limited), and direct contact feeding across a diseased and healthy coral (unlimited). After the experimental trials, the composition of Symbiodiniaceae was analyzed for every MCAV fragment. Direct contact transmission took on average 3.9 days in 2019 and 11.9 days in 2020 with significantly quicker rates of transmission between donor diseased Orbicella and Montastraea than Montastraea to Montastraea. The composition of Symbiodiniaceae differed significantly between the fragments used in 2019 (dominated by Durusdinium) than in 2020 (dominated by Cladocopium spp.). The limited treatment had the quickest time to transmission compared to the unlimited and control treatments. Symbiodiniaceae differences between 2019 and 2020 might explain differences in transmission rates and overall susceptibility between the years. The species of Symbiodiniaceae may play a role in the susceptibility of corals to the transmission of SCTLD. Additionally, we also have some suggestive evidence that butterflyfish do not directly increase infection rates, but instead might be increasing infection recovery.

Metabolomics Approaches to Dereplicate Natural Products from Coral-Derived Bioactive Bacteria

Stony corals (Scleractinia) are invertebrates that form symbiotic relationships with eukaryotic algal endosymbionts and the prokaryotic microbiome. The microbiome has the potential to produce bioactive natural products providing defense and resilience to the coral host against pathogenic microorganisms, but this potential has not been extensively explored. Bacterial pathogens can pose a significant threat to corals, with some species implicated in primary and opportunistic infections of various corals. In response, probiotics have been proposed as a potential strategy to protect corals in the face of increased incidence of disease outbreaks. In this study, we screened bacterial isolates from healthy and diseased corals for antibacterial activity. The bioactive extracts were analyzed using untargeted metabolomics. Herein, an UpSet plot and hierarchical clustering analyses were performed to identify isolates with the largest number of unique metabolites. These isolates also displayed different antibacterial activities. Through application of in silico and experimental approaches coupled with genome analysis, we dereplicated natural products from these coral-derived bacteria from Florida's coral reef environments. The metabolomics approach highlighted in this study serves as a useful resource to select probiotic candidates and enables insights into natural product-mediated chemical ecology in holobiont symbiosis.

Reef Sediments Can Act As a Stony Coral Tissue Loss Disease Vector

Stony coral tissue loss disease (SCTLD) was first observed in 2014 near Virginia Key in Miami-Dade County, Florida. Field sampling, lab experiments, and modeling approaches have suggested that reef sediments may play a role in SCTLD transmission, though a positive link has not been tested experimentally. We conducted an ex situ transmission assay using a statistically-independent disease apparatus to test whether reef sediments can transmit SCTLD in the absence of direct contact between diseased and healthy coral tissue. We evaluated two methods of sediment inoculation: batch inoculation of sediments collected from southeast Florida using whole colonies of diseased Montastraea cavernosa, and individual inoculations of sediments following independent, secondary infections of ∼5 cm2 coral fragments. Healthy fragments of the coral species Orbicella faveolata and M. cavernosa were exposed to these diseased sediment treatments, as well as direct disease contact and healthy sediment controls. SCTLD transmission was observed for both batch and individual diseased sediment inoculation treatments, albeit with lower proportions of infected individuals as compared to disease contact controls. The time to onset of lesions was significantly different between species and among disease treatments, with the most striking infections occurring in the individual diseased sediment treatment in under 24 h. Following infection, tissue samples were confirmed for the presence of SCTLD signs via histological examination, and sediment subsamples were analyzed for microbial community variation between treatments, identifying 16 SCTLD indicator taxa in sediments associated with corals experiencing tissue loss. This study demonstrated that reef sediments can indeed transmit SCTLD through indirect exposure between diseased and healthy corals, and adds credence to the assertion that SCTLD transmission occurs via an infectious agent or agents. This study emphasizes the critical need to understand the roles that sediment microbial communities and coastal development activities may have on the persistence of SCTLD throughout the endemic zone, especially in the context of management and conservation strategies in Florida and the wider Caribbean.

Combining tangential flow filtration and size fractionation of mesocosm water as a method for the investigation of waterborne coral diseases.

The causative agents of most coral diseases today remain unknown, complicating disease response and restoration efforts. Pathogen identifications can be hampered by complex microbial communities naturally associated with corals and seawater, which create complicating "background noise" that can potentially obscure a pathogen's signal. Here, we outline an approach to investigate waterborne coral diseases that use a combination of coral mesocosms, tangential flow filtration, and size fractionation to reduce the impact of this background microbial diversity, compensate for unknown infectious dose, and further narrow the suspect pool of potential pathogens. As proof of concept, we use this method to compare the bacterial communities shed into six Montastraea cavernosa coral mesocosms and demonstrate this method effectively detects differences between diseased and healthy coral colonies. We found several amplicon sequence variants (ASVs) in the diseased mesocosms that represented 100% matches with ASVs identified in prior studies of diseased coral tissue, further illustrating the effectiveness of our approach. Our described method is an effective alternative to using coral tissue or mucus to investigate waterborne coral diseases of unknown etiology and can help more quickly narrow the pool of possible pathogens to better aid in disease response efforts. Additionally, this versatile method can be easily adapted to characterize either the entire microbial community associated with a coral or target-specific microbial groups, making it a beneficial approach regardless of whether a causative agent is suspected or is completely unknown.

Bacterial Metabolic Potential and Micro-Eukaryotes Enriched in Stony Coral Tissue Loss Disease Lesions

The epizootic disease outbreak known as stony coral tissue loss disease (SCTLD) is arguably the most devastating coral disease in recorded history. SCTLD emerged off the coast of South Florida in 2014 and has since moved into the Caribbean, resulting in coral mortality rates that have changed reef structure and function. Currently, the cause of SCTLD is unknown, but there is evidence from 16S rRNA gene sequencing and bacterial culture studies that the microbial community plays a role in the progression of SCTLD lesions. In this study, we applied shotgun metagenomics to characterize the potential function of bacteria, as well as the composition of the micro-eukaryotic community, associated with SCTLD lesions. We re-examined samples that were previously analyzed using 16S rRNA gene high-throughput sequencing from four coral species: Stephanocoenia intersepta, Diploria labyrinthiformis, Dichocoenia stokesii, and Meandrina meandrites. For each species, tissue from apparently healthy (AH) corals, and unaffected tissue (DU) and lesion tissue (DL) on diseased corals, were collected from sites within the epidemic zone of SCTLD in the Florida Keys. Within the micro-eukaryotic community, the taxa most prominently enriched in DL compared to AH and DU tissue were members of Ciliophora. We also found that DL samples were relatively more abundant in less energy-efficient pathways like the pentose phosphate pathways. While less energy-efficient processes were identified, there were also relatively higher abundances of nucleotide biosynthesis and peptidoglycan maturation pathways in diseased corals compared to AH, which suggests there was more bacteria growth in diseased colonies. In addition, we generated 16 metagenome-assembled genomes (MAGs) belonging to the orders Pseudomonadales, Beggiatoales, Rhodobacterales, Rhizobiales, Rs-D84, Flavobacteriales, and Campylobacterales, and all MAGs were enriched in DL samples compared to AH samples. Across all MAGs there were antibiotic resistance genes that may have implications for the treatment of SCTLD with antibiotics. We also identified genes and pathways linked to virulence, such as nucleotide biosynthesis, succinate dehydrogenase, ureases, nickel/iron transporters, Type-1 secretion system, and metalloproteases. Some of these enzymes/pathways have been previously targeted in the treatment of other bacterial diseases and they may be of interest to mitigate SCTLD lesion progression.