Exploring the Diversity and Biotechnological Potential of Cultured and Uncultured Coral-Associated Bacteria

Coral-associated microbes are crucial for the biology of their hosts, contributing to nutrient cycling, adaptation, mitigation of toxic compounds, and biological control of pathogens. Natural products from coral-associated micro-organisms (CAM) may possess unique traits. Despite this, the use of CAM for biotechnological purposes has not yet been adequately explored. Here, we investigated the production of commercially important enzymes by 37 strains of bacteria isolated from the coral species Mussismilia braziliensis, Millepora alcicornis, and Porites astreoides. In-vitro enzymatic assays showed that up to 56% of the isolates produced at least one of the seven enzymes screened (lipase, caseinase, keratinase, cellulase, chitinase, amylase, and gelatinase); one strain, identified as Bacillus amyloliquefaciens produced all these enzymes. Additionally, coral species-specific cultured and uncultured microbial communities were identified. The phylum Firmicutes predominated among the isolates, including the genera Exiguobacterium, Bacillus, and Halomonas, among others. Next-generation sequencing and bacteria culturing produced similar but also complementary data, with certain genera detected only by one or the other method. Our results demonstrate the importance of exploring different coral species as sources of specific micro-organisms of biotechnological and industrial interest, at the same time reinforcing the economic and ecological importance of coral reefs as reservoirs of such diversity.

Plasticity of Porites astreoides Early Life History Stages Suggests Mesophotic Coral Ecosystems Act as Refugia in Bermuda

As the devastating impacts of global climate change and local anthropogenic stressors on shallow-water coral reefs are expected to rise, mesophotic coral ecosystems have increasingly been regarded as potential lifeboats for coral survival, providing a source of propagules to replenish shallower reefs. Yet, there is still limited knowledge of the capacity for coral larvae to adjust to light intensities that change with depth. This study elucidates the mechanisms underlying plasticity during early life stages of the coral Porites astreoides that enable survival across broad depth gradients. We examined physiological and morphological variations in larvae from shallow (8–10 m) and mesophotic (45 m) reefs in Bermuda, and evaluated differences in survival, settlement patterns and size among recruits depending on light conditions using a reciprocal ex situ transplantation experiment. Larvae released from mesophotic adults were found to have significantly lower respiration rates and were significantly larger than those derived from shallow adults, indicating higher content of energetic resources and suggesting a greater dispersal potential for mesophotic larvae compared to their shallow counterparts. Additionally, larvae released from mesophotic adults experienced higher settlement success and larger initial spat size compared to larvae from shallow adults, demonstrating a potential connection between parental origin, offspring quality, and recruitment success. Although both shallow and mesophotic larvae exhibited the capacity to survive and settle under reciprocal light conditions, all larvae had higher survival under mesophotic light conditions regardless of parental origin, suggesting that conditions experienced under low light may enable longer larval life, further extending the dispersal period. These results indicate that larvae from mesophotic Porites astreoides colonies are likely capable of reseeding shallow reefs in Bermuda, thereby supporting the Deep Reef Refugia Hypothesis.

Variation in Immune-Related Gene Expression Provides Evidence of Local Adaptation in Porites astreoides (Lamarck, 1816) between Inshore and Offshore Meta-Populations Inhabiting the Lower Florida Reef Tract, USA

Coral communities of the Florida Reef Tract (FRT) have changed dramatically over the past 30 years. Coral cover throughout the FRT is disproportionately distributed; >70% of total coral cover is found within the inshore patch reef zone (<2 km from shore) compared to 30% found within the offshore bank reef zone (>5 km from shore). Coral mortality from disease has been differentially observed between inshore and offshore reefs along the FRT. Therefore, differences between the response of inshore and offshore coral populations to bacterial challenge may contribute to differences in coral cover. We examined immune system activation in Porites astreoides (Lamarck, 1816), a species common in both inshore and offshore reef environments in the FRT. Colonies from a representative inshore and offshore site were reciprocally transplanted and the expression of three genes monitored biannually for two years (two summer and two winter periods). Variation in the expression of eukaryotic translation initiation factor 3, subunit H (eIF3H), an indicator of cellular stress in Porites astreoides, did not follow annual patterns of seawater temperatures (SWT) indicating the contribution of other stressors (e.g., irradiance). Greater expression of tumor necrosis factor (TNF) receptor associated factor 3 (TRAF3), a signaling protein of the inflammatory response, was observed among corals transplanted to, or located within the offshore environment indicating that an increased immune response is associated with offshore coral more so than the inshore coral (p < 0.001). Corals collected from the offshore site also upregulated the expression of adenylyl cyclase associated protein 2 (ACAP2), increases which are associated with decreasing innate immune system inflammatory responses, indicating a counteractive response to increased stimulation of the innate immune system. Activation of the innate immune system is a metabolically costly survival strategy. Among the two reefs studied, the offshore population had a smaller mean colony size and decreased colony abundance compared to the inshore site. This correlation suggests that tradeoffs may exist between the activation of the innate immune system and survival and growth. Consequently, immune system activation may contribute to coral community dynamics and declines along the FRT.

Comparison of Preservation and Extraction Methods on Five Taxonomically Disparate Coral Microbiomes

All animals are host to a multitude of microorganisms that are essential to the animal’s health. Host-associated microbes have been shown to defend against potential pathogens, provide essential nutrients, interact with the host’s immune system, and even regulate mood. However, it can be difficult to preserve and obtain nucleic acids from some host-associated microbiomes, making studying their microbial communities challenging. Corals are an example of this, in part due to their potentially remote, underwater locations, their thick surface mucopolysaccharide layer, and various inherent molecular inhibitors. This study examined three different preservatives (RNAlater, DNA/RNA Shield, and liquid nitrogen) and two extraction methods (the Qiagen PowerBiofilm kit and the Promega Maxwell RBC kit with modifications) to determine if there was an optimum combination for examining the coral microbiome. These methods were employed across taxonomically diverse coral species, including deep-sea/shallow, stony/soft, and zooxanthellate/azooxanthellate: Lophelia pertusa, Paragorgia johnsoni, Montastraea cavernosa, Porites astreoides, and Stephanocoenia intersepta. Although significant differences were found between preservative types and extraction methods, these differences were subtle, and varied in nature from coral species to coral species. Significant differences between coral species were far more profound than those detected between preservative or extraction method. We suggest that the preservative types presented here and extraction methods using a bead-beating step provide enough consistency to compare coral microbiomes across various studies, as long as subtle differences in microbial communities are attributed to dissimilar methodologies. Additionally, the inclusion of internal controls such as a mock community and extraction blanks can help provide context regarding data quality, improving downstream analyses.

Genetic structure and diversity of the mustard hill coral Porites astreoides along the Florida Keys reef tract

Increases in local and global stressors have led to major declines in coral populations throughout the western Atlantic. While abundances of other species have declined, however, the relative abundance of the mustard hill coral, Porites astreoides, has increased. Porites astreoides is relatively resilient to some stressors, and because of its mixed reproductive strategies, its populations often recover quickly following disturbances. The ability for P. astreoides to continue as a potential “winner” in western Atlantic reefs relies on maintaining sufficient genetic variation within populations to support acclimatization and adaptation to current and future environmental change. Without high genetic diversity and gene flow within the population, it would have limited capacity for adaptation and the species’ competitive advantages could be short-lived. In this study, we determined the genetic relatedness of 37 P. astreoides colonies at four shallow reefs along the offshore Florida Keys Reef Tract (FKRT), a region particularly hard-hit by recent disturbances. Using previously designed microsatellite markers, we determined the genetic diversity and connectivity of individuals among and between sites. Our results suggest that the FKRT likely contains a single, well-mixed genetic population of P. astreoides, with high levels of gene flow and evidence for larval migration throughout the region. This suggests that regional populations of P. astreoides likely have a higher chance of maintaining resilience than many other western Atlantic species as they face current and future disturbances.

Coverage Increases of Porites astreoides in Grenada Determined by Shifts in Size-Frequency Distribution

Despite coral community collapse, the mustard hill coral (Porites astreoides) is a species currently experiencing success throughout the Caribbean. The inshore reefs of Grenada were selected to study the influence of benthic factors on the abundance, size, and coverage of P. astreoides colonies. Surveys of reef communities along established 30 m transects were conducted at eight sites in 2014 and 2017 using a 0.5 m² quadrat. Coral Point Count was used to annotate the images, estimating the coverage of scleractinian corals, sponges, algae, and benthic substrates. Coverage, size, and abundance of P. astreoides colonies were quantified using the area measurement tool in ImageJ standardized against the quadrats. There were significant differences in benthic community assemblages between islands, selected sites, and between years. From 2014 to 2017 there was a significant decrease in the mean abundance of P. astreoides colonies and significant increases in mean colony size and coverage. The presence of P. astreoides colonies was significantly correlated with: rubble (−), sand (−); pavement (+); macroalgae (−); coralline algae (+); sponges (varying response); gorgonians (−); massive corals (+); and branching corals (−). P. astreoides follows similar recruitment patterns as other scleractinian corals. Observed changes in P. astreoides populations appear to indicate a recovery event following a disturbance, potentially tropical storm Chantal in 2013.

Physiological Differences in Bleaching Response of the Coral Porites astreoides Along the Florida Keys Reef Tract During High-Temperature Stress

The Florida Keys reef tract (FKRT) has a unique geological history wherein Holocene sea-level rise and bathymetry interacted, resulting in a reef-building system with notable spatial differences in reef development. Overprinted on this geologic history, recent global and local stressors have led to degraded reefs dominated by fleshy algae, soft corals, and sponges. Here, we assessed how coral physiology (calcification rate, tissue thickness, reproduction, symbiosis, and bleaching) varies seasonally (winter vs. summer) and geographically using 40 colonies of the mustard hill coral Porites astreoides from four sites across 350 km along the FKRT from 2015 to 2017. The study coincided with a high-temperature event in late summer 2015 that caused heterogeneous levels of coral bleaching across sites. Bleaching severity differed by site, with bleaching response more aligned with heat stress retroactively calculated from local degree heating weeks than those predicted by satellites. Despite differences in temperature profiles and bleaching severity, all colonies hosted Symbiodiniaceae of the same genus (formerly Clade A and subtypes). Overall, P. astreoides at Dry Tortugas National Park, the consistently coolest site, had the highest calcification rates, symbiont cell densities, and reproductive potential (all colonies were reproductive, with most planula larvae per polyp). Corals at Dry Tortugas and Fowey Rocks Light demonstrated strong seasonality in net calcification (higher in summer) and did not express visual or partial-mortality responses from the bleaching event; in contrast, colonies in the middle and southern part of the upper keys, Sombrero Key and Crocker Reef, demonstrated similar reduced fitness from bleaching, but differential recovery trajectories following the heat stress. Identifying reefs, such as Dry Tortugas and possibly Fowey Rocks Light that may serve as heat-stress refugia, is important in selecting candidate sites for adaptive reef-management strategies, such as selective propagation and assisted gene flow, to increase coral-species adaptation to ocean warming.