Co-occurring nitrifying symbiont lineages are vertically inherited and widespread in marine sponges.

Ammonia-oxidising archaea and nitrite-oxidising bacteria are common members of marine sponge microbiomes. They derive energy for carbon fixation and growth from nitrification - the aerobic oxidation of ammonia to nitrite and further to nitrate - and are proposed to play essential roles in the carbon and nitrogen cycling of sponge holobionts. In this study, we characterise two novel nitrifying symbiont lineages, Candidatus Nitrosokoinonia and Candidatus Nitrosymbion in the marine sponge Coscinoderma matthewsi using a combination of molecular tools, in situ visualisation, and physiological rate measurements. Both represent a new genus in the ammonia-oxidising archaeal class Nitrososphaeria and the nitrite-oxidising bacterial order Nitrospirales, respectively. Furthermore, we show that larvae of this viviparous sponge are densely colonised by representatives of Ca. Nitrosokoinonia and Ca. Nitrosymbion indicating vertical transmission. In adults, the representatives of both symbiont genera are located extracellularly in the mesohyl. Comparative metagenome analyses and physiological data suggest that ammonia-oxidising archaeal symbionts of the genus Ca. Nitrosokoinonia strongly rely on endogenously produced nitrogenous compounds (i.e., ammonium, urea, nitriles/cyanides, and creatinine) rather than on exogenous ammonium sources taken up by the sponge. Additionally, the nitrite-oxidising bacterial symbionts of the genus Ca. Nitrosymbion may reciprocally support the ammonia-oxidisers with ammonia via the utilisation of sponge-derived urea and cyanate. Comparative analyses of published environmental 16S rRNA gene amplicon data revealed that Ca. Nitrosokoinonia and Ca. Nitrosymbion are widely distributed and predominantly associated with marine sponges and corals, suggesting a broad relevance of our findings.

The road forward to incorporate seawater microbes in predictive reef monitoring.

Marine bacterioplankton underpin the health and function of coral reefs and respond in a rapid and sensitive manner to environmental changes that affect reef ecosystem stability. Numerous meta-omics surveys over recent years have documented persistent associations of opportunistic seawater microbial taxa, and their associated functions, with metrics of environmental stress and poor reef health (e.g. elevated temperature, nutrient loads and macroalgae cover). Through positive feedback mechanisms, disturbance-triggered heterotrophic activity of seawater microbes is hypothesised to drive keystone benthic organisms towards the limit of their resilience and translate into shifts in biogeochemical cycles which influence marine food webs, ultimately affecting entire reef ecosystems. However, despite nearly two decades of work in this space, a major limitation to using seawater microbes in reef monitoring is a lack of a unified and focused approach that would move beyond the indicator discovery phase and towards the development of rapid microbial indicator assays for (near) real-time reef management and decision-making. By reviewing the current state of knowledge, we provide a comprehensive framework (defined as five phases of research and innovation) to catalyse a shift from fundamental to applied research, allowing us to move from descriptive to predictive reef monitoring, and from reactive to proactive reef management.

Taurine as a key intermediate for host-symbiont interaction in the tropical sponge Ianthella basta.

Marine sponges are critical components of marine benthic fauna assemblages, where their filter-feeding and reef-building capabilities provide bentho-pelagic coupling and crucial habitat. As potentially the oldest representation of a metazoan-microbe symbiosis, they also harbor dense, diverse, and species-specific communities of microbes, which are increasingly recognized for their contributions to dissolved organic matter (DOM) processing. Recent omics-based studies of marine sponge microbiomes have proposed numerous pathways of dissolved metabolite exchange between the host and symbionts within the context of the surrounding environment, but few studies have sought to experimentally interrogate these pathways. By using a combination of metaproteogenomics and laboratory incubations coupled with isotope-based functional assays, we showed that the dominant gammaproteobacterial symbiont, 'Candidatus Taurinisymbion ianthellae', residing in the marine sponge, Ianthella basta, expresses a pathway for the import and dissimilation of taurine, a ubiquitously occurring sulfonate metabolite in marine sponges. 'Candidatus Taurinisymbion ianthellae' incorporates taurine-derived carbon and nitrogen while, at the same time, oxidizing the dissimilated sulfite into sulfate for export. Furthermore, we found that taurine-derived ammonia is exported by the symbiont for immediate oxidation by the dominant ammonia-oxidizing thaumarchaeal symbiont, 'Candidatus Nitrosospongia ianthellae'. Metaproteogenomic analyses also suggest that 'Candidatus Taurinisymbion ianthellae' imports DMSP and possesses both pathways for DMSP demethylation and cleavage, enabling it to use this compound as a carbon and sulfur source for biomass, as well as for energy conservation. These results highlight the important role of biogenic sulfur compounds in the interplay between Ianthella basta and its microbial symbionts.

Microbial Surface Biofilm Responds to the Growth-Reproduction-Senescence Cycle of the Dominant Coral Reef Macroalgae Sargassum spp.

Macroalgae play an intricate role in microbial-mediated coral reef degradation processes due to the release of dissolved nutrients. However, temporal variabilities of macroalgal surface biofilms and their implication on the wider reef system remain poorly characterized. Here, we study the microbial biofilm of the dominant reef macroalgae Sargassum over a period of one year at an inshore Great Barrier Reef site (Magnetic Island, Australia). Monthly sampling of the Sargassum biofilm links the temporal taxonomic and putative functional metabolic microbiome changes, examined using 16S rRNA gene amplicon and metagenomic sequencing, to the pronounced growth-reproduction-senescence cycle of the host. Overall, the macroalgal biofilm was dominated by the heterotrophic phyla Firmicutes (35% ± 5.9% SD) and Bacteroidetes (12% ± 0.6% SD); their relative abundance ratio shifted significantly along the annual growth-reproduction-senescence cycle of Sargassum. For example, Firmicutes were 1.7 to 3.9 times more abundant during host growth and reproduction cycles than Bacteroidetes. Both phyla varied in their carbohydrate degradation capabilities; hence, temporal fluctuations in the carbohydrate availability are potentially linked to the observed shift. Dominant heterotrophic macroalgal biofilm members, such as Firmicutes and Bacteroidetes, are implicated in exacerbating or ameliorating the release of dissolved nutrients into the ambient environment, though their contribution to microbial-mediated reef degradation processes remains to be determined.

Microbiome dynamics in the tissue and mucus of acroporid corals differ in relation to host and environmental parameters.

Corals are associated with diverse microbial assemblages; however, the spatial-temporal dynamics of intra-species microbial interactions are poorly understood. The coral-associated microbial community varies substantially between tissue and mucus microhabitats; however, the factors controlling the occurrence, abundance, and distribution of microbial taxa over time have rarely been explored for different coral compartments simultaneously. Here, we test (1) differentiation in microbiome diversity and composition between coral compartments (surface mucus and tissue) of two Acropora hosts (A. tenuis and A. millepora) common along inshore reefs of the Great Barrier Reef, as well as (2) the potential linkage between shifts in individual coral microbiome families and underlying host and environmental parameters. Amplicon based 16S ribosomal RNA gene sequencing of 136 samples collected over 14 months, revealed significant differences in bacterial richness, diversity and community structure among mucus, tissue and the surrounding seawater. Seawater samples were dominated by members of the Synechococcaceae and Pelagibacteraceae bacterial families. The mucus microbiome of Acropora spp. was dominated by members of Flavobacteriaceae, Synechococcaceae and Rhodobacteraceae and the tissue was dominated by Endozoicimonaceae. Mucus microbiome in both Acropora species was primarily correlated with seawater parameters including levels of chlorophyll a, ammonium, particulate organic carbon and the sum of nitrate and nitrite. In contrast, the correlation of the tissue microbiome to the measured environmental (i.e., seawater parameters) and host health physiological factors differed between host species, suggesting host-specific modulation of the tissue-associated microbiome to intrinsic and extrinsic factors. Furthermore, the correlation between individual coral microbiome members and environmental factors provides novel insights into coral microbiome-by-environment dynamics and hence has potential implications for current reef restoration and management efforts (e.g. microbial monitoring and observatory programs).

Spatial patterns of microbial communities across surface waters of the Great Barrier Reef.

Microorganisms are fundamental drivers of biogeochemical cycling, though their contribution to coral reef ecosystem functioning is poorly understood. Here, we infer predictors of bacterioplankton community dynamics across surface-waters of the Great Barrier Reef (GBR) through a meta-analysis, combining microbial with environmental data from the eReefs platform. Nutrient dynamics and temperature explained 41.4% of inter-seasonal and cross-shelf variation in bacterial assemblages. Bacterial families OCS155, Cryomorphaceae, Flavobacteriaceae, Synechococcaceae and Rhodobacteraceae dominated inshore reefs and their relative abundances positively correlated with nutrient loads. In contrast, Prochlorococcaceae negatively correlated with nutrients and became increasingly dominant towards outershelf reefs. Cyanobacteria in Prochlorococcaceae and Synechococcaceae families occupy complementary cross-shelf biogeochemical niches; their abundance ratios representing a potential indicator of GBR nutrient levels. One Flavobacteriaceae-affiliated taxa was putatively identified as diagnostic for ecosystem degradation. Establishing microbial observatories along GBR environmental gradients will facilitate robust assessments of microbial contributions to reef health and inform tipping-points in reef condition.

Comparative genome-centric analysis reveals seasonal variation in the function of coral reef microbiomes.

Microbially mediated processes contribute to coral reef resilience yet, despite extensive characterisation of microbial community variation following environmental perturbation, the effect on microbiome function is poorly understood. We undertook metagenomic sequencing of sponge, macroalgae and seawater microbiomes from a macroalgae-dominated inshore coral reef to define their functional potential and evaluate seasonal shifts in microbially mediated processes. In total, 125 high-quality metagenome-assembled genomes were reconstructed, spanning 15 bacterial and 3 archaeal phyla. Multivariate analysis of the genomes relative abundance revealed changes in the functional potential of reef microbiomes in relation to seasonal environmental fluctuations (e.g. macroalgae biomass, temperature). For example, a shift from Alphaproteobacteria to Bacteroidota-dominated seawater microbiomes occurred during summer, resulting in an increased genomic potential to degrade macroalgal-derived polysaccharides. An 85% reduction of Chloroflexota was observed in the sponge microbiome during summer, with potential consequences for nutrition, waste product removal, and detoxification in the sponge holobiont. A shift in the Firmicutes:Bacteroidota ratio was detected on macroalgae over summer with potential implications for polysaccharide degradation in macroalgal microbiomes. These results highlight that seasonal shifts in the dominant microbial taxa alter the functional repertoire of host-associated and seawater microbiomes, and highlight how environmental perturbation can affect microbially mediated processes in coral reef ecosystems.

Microbial indicators of environmental perturbations in coral reef ecosystems.

BACKGROUND: Coral reefs are facing unprecedented pressure on local and global scales. Sensitive and rapid markers for ecosystem stress are urgently needed to underpin effective management and restoration strategies. Although the fundamental contribution of microbes to the stability and functioning of coral reefs is widely recognised, it remains unclear how different reef microbiomes respond to environmental perturbations and whether microbiomes are sensitive enough to predict environmental anomalies that can lead to ecosystem stress. However, the lack of coral reef microbial baselines hinders our ability to study the link between shifts in microbiomes and ecosystem stress. In this study, we established a comprehensive microbial reference database for selected Great Barrier Reef sites to assess the diagnostic value of multiple free-living and host-associated reef microbiomes to infer the environmental state of coral reef ecosystems. RESULTS: A comprehensive microbial reference database, originating from multiple coral reef microbiomes (i.e. seawater, sediment, corals, sponges and macroalgae), was generated by 16S rRNA gene sequencing for 381 samples collected over the course of 16 months. By coupling this database to environmental parameters, we showed that the seawater microbiome has the greatest diagnostic value to infer shifts in the surrounding reef environment. In fact, 56% of the observed compositional variation in the microbiome was explained by environmental parameters, and temporal successions in the seawater microbiome were characterised by uniform community assembly patterns. Host-associated microbiomes, in contrast, were five-times less responsive to the environment and their community assembly patterns were generally less uniform. By applying a suite of indicator value and machine learning approaches, we further showed that seawater microbial community data provide an accurate prediction of temperature and eutrophication state (i.e. chlorophyll concentration and turbidity). CONCLUSION: Our results reveal that free-living microbial communities have a high potential to infer environmental parameters due to their environmental sensitivity and predictability. This highlights the diagnostic value of microorganisms and illustrates how long-term coral reef monitoring initiatives could be enhanced by incorporating assessments of microbial communities in seawater. We therefore recommend timely integration of microbial sampling into current coral reef monitoring initiatives.

Disentangling the effect of host-genotype and environment on the microbiome of the coral Acropora tenuis.

Genotype-specific contributions to the environmental tolerance and disease susceptibility of corals are widely accepted. Yet our understanding of how host genotype influences the composition and stability of the coral microbiome subjected to environmental fluctuations is limited. To gain insight into the community dynamics and environmental stability of microbiomes associated with distinct coral genotypes, we assessed the microbial community associated with Acropora tenuis under single and cumulative pressure experiments. Experimental treatments comprised either a single pulse of reduced salinity (minimum of 28 psu) or exposure to the cumulative pressures of reduced salinity (minimum of 28 psu), elevated seawater temperature (+2 °C), elevated pCO2 (900 ppm), and the presence of macroalgae. Analysis of 16S rRNA gene amplicon sequence data revealed that A. tenuis microbiomes were highly host-genotype specific and maintained high compositional stability irrespective of experimental treatment. On average, 48% of the A. tenuis microbiome was dominated by Endozoicomonas. Amplicon sequence variants (ASVs) belonging to this genus were significantly different between host individuals. Although no signs of stress were evident in the coral holobiont and the vast majority of ASVs remained stable across treatments, a microbial indicator approach identified 26 ASVs belonging to Vibrionaceae, Rhodobacteraceae, Hahellaceae, Planctomycetes, Phylobacteriaceae, Flavobacteriaceae, and Cryomorphaceae that were significantly enriched in corals exposed to single and cumulative stressors. While several recent studies have highlighted the efficacy of microbial indicators as sensitive markers for environmental disturbance, the high host-genotype specificity of coral microbiomes may limit their utility and we therefore recommend meticulous control of host-genotype effects in coral microbiome research.

Exploring the diversity-stability paradigm using sponge microbial communities.

A key concept in theoretical ecology is the positive correlation between biodiversity and ecosystem stability. When applying this diversity-stability concept to host-associated microbiomes, the following questions emerge: (1) Does microbial diversity influence the stability of microbiomes upon environmental fluctuations? (2) Do hosts that harbor high versus low microbial diversity differ in their stress response? To test the diversity-stability concept in host-associated microbiomes, we exposed six marine sponge species with varying levels of microbial diversity to non-lethal salinity disturbances and followed their microbial composition over time using 16S rRNA gene amplicon sequencing. No signs of sponge stress were evident following salinity amendment and microbiomes exhibited compositional resistance irrespective of their microbial diversity. Compositional stability of the sponge microbiome manifests itself at distinct host taxonomic and host microbial diversity groups, with (1) stable host genotype-specific microbiomes at oligotype-level; (2) stable host species-specific microbiomes at genus-level; and (3) stable and specific microbiomes at phylum-level for hosts with high versus low microbial diversity. The resistance of sponge microbiomes together with the overall stability of sponge holobionts upon salinity fluctuations suggest that the stability-diversity concept does not appear to hold for sponge microbiomes and provides further evidence for the widely recognized environmental tolerance of sponges.

Microbiome variation in corals with distinct depth distribution ranges across a shallow-mesophotic gradient (15-85 m).

Mesophotic coral ecosystems (MCEs) are generally poorly studied, and our knowledge of lower MCEs (below 60 m depth) is largely limited to visual surveys. Here, we provide a first detailed assessment of the prokaryotic community associated with scleractinian corals over a depth gradient to the lower mesophotic realm (15-85 m). Specimens of three Caribbean coral species exhibiting differences in their depth distribution ranges (Agaricia grahamae, Madracis pharensis and Stephanocoenia intersepta) were collected with a manned submersible on the island of Curaçao, and their prokaryotic communities assessed using 16S rRNA gene sequencing analysis. Corals with narrower depth distribution ranges (depth-specialists) were associated with a stable prokaryotic community, whereas corals with a broader niche range (depth-generalists) revealed a higher variability in their prokaryotic community. The observed depth effects match previously described patterns in Symbiodinium depth zonation. This highlights the contribution of structured microbial communities over depth to the coral's ability to colonize a broader depth range.

Evaluating the core microbiota in complex communities: A systematic investigation.

The study of complex microbial communities poses unique conceptual and analytical challenges, with microbial species potentially numbering in the thousands. With transient or allochthonous microorganisms often adding to this complexity, a 'core' microbiota approach, focusing only on the stable and permanent members of the community, is becoming increasingly popular. Given the various ways of defining a core microbiota, it is prudent to examine whether the definition of the core impacts upon the results obtained. Here we used complex marine sponge microbiotas and undertook a systematic evaluation of the degree to which different factors used to define the core influenced the conclusions. Significant differences in alpha- and beta-diversity were detected using some but not all core definitions. However, findings related to host specificity and environmental quality were largely insensitive to major changes in the core microbiota definition. Furthermore, none of the applied definitions altered our perception of the ecological networks summarising interactions among bacteria within the sponges. These results suggest that, while care should still be taken in interpretation, the core microbiota approach is surprisingly robust, at least for comparing microbiotas of closely related samples.

The microbiome of coral surface mucus has a key role in mediating holobiont health and survival upon disturbance.

Microbes are well-recognized members of the coral holobiont. However, little is known about the short-term dynamics of mucus-associated microbial communities under natural conditions and after disturbances, and how these dynamics relate to the host's health. Here we examined the natural variability of prokaryotic communities (based on 16S ribosomal RNA gene amplicon sequencing) associating with the surface mucus layer (SML) of Porites astreoides, a species exhibiting cyclical mucus aging and shedding. Shifts in the prokaryotic community composition during mucus aging led to the prevalence of opportunistic and potentially pathogenic bacteria (Verrucomicrobiaceae and Vibrionaceae) in aged mucus and to a twofold increase in prokaryotic abundance. After the release of aged mucus sheets, the community reverted to its original state, dominated by Endozoicimonaceae and Oxalobacteraceae. Furthermore, we followed the fate of the coral holobiont upon depletion of its natural mucus microbiome through antibiotics treatment. After re-introduction to the reef, healthy-looking microbe-depleted corals started exhibiting clear signs of bleaching and necrosis. Recovery versus mortality of the P. astreoides holobiont was related to the degree of change in abundance distribution of the mucus microbiome. We conclude that the natural prokaryotic community inhabiting the coral SML contributes to coral health and that cyclical mucus shedding has a key role in coral microbiome dynamics.

Quantification of patient compliance with Hawley retainers and removable functional appliances during the retention phase.

INTRODUCTION: The success of retention with removable retainers is highly dependent on efficient patient compliance. The aim of this study was to quantify patient compliance with removable retainers using microelectronic wear-time documentation during the retention phase. METHODS: One hundred patients, between 13 and 20 years of age, were retained with removable Hawley retainers and functional appliance retainers after successful multibracket treatment at the University Hospital of Tübingen, Germany, and in 4 private practices in Germany. Microsensors were incorporated into the orthodontic retainers by polymerization, and daily wear time was documented in 15-minute intervals during the retention phase for up to 15 months. Patient compliance was quantified with wear-time documentation. Additionally, the influences of age, sex, place of treatment, device type, and health insurance status on compliance were determined and statistically evaluated. RESULTS: Most study participants complied with the prescribed wear time of 8 hours or more per day. Combined patient data indicated a median wear time of 7.0 hours per day over the evaluation period. Wear-time documentation showed either regular or irregular patterns of compliance. Initial compliance did not usually alter over the retention phase. Compliance was not influenced by device type, but age, sex, place of treatment, and insurance status produced changes in the median wear time of up to 50%. CONCLUSIONS: Electronic wear-time documentation of patients' compliance is an easily comprehensible measurement that allows orthodontists to examine the patient's contribution to the success of retention and personalize treatment accordingly. Place of treatment and health insurance status are more closely associated with compliance than are basic patient demographics.

Application of a new viscoelastic finite element method model and analysis of miniscrew-supported hybrid hyrax treatment.

INTRODUCTION: In this study, we aimed to assess the ability of a new viscoelastic finite element method model to accurately simulate rapid palatal expansion with a miniscrew-supported hybrid hyrax appliance. METHODS: A female patient received 3-dimensional craniofacial imaging with computed tomography at 2 times: before expansion and immediately after expansion, with the latter serving as a reference model for the analysis. A novel approach was applied to the finite element method model to improve simulation of the viscoelastic properties of osseous tissue. RESULTS: The resulting finite element method model was a suitable approximation of the clinical situation and adequately simulated the forced expansion of the midpalatal suture. Specifically, it demonstrated that the hybrid hyrax appliance delivered a force via the 2 mini-implants at the center of resistance of the nasomaxillary complex. CONCLUSIONS: The newly developed model provided a suitable simulation of the clinical effects of the hybrid hyrax appliance, which proved to be a suitable device for rapid palatal expansion.

Changes in soft-tissue profiles after treatment of class II/1 patients with bite-jumping appliances.

We performed this study to describe changes in the soft-tissue profile after orthodontic treatment with Tränkmann's bite-jumping appliances. A total of 69 patients thus treated were compared to a control group of 36 age-matched patients based on cephalograms. Statistical analysis included mean values, standard deviations, t-tests, and Pearson's correlation testing. Highly significant (p<0.001) changes in total profile angle (N'-Ns-Pog') were observed over the course of treatment. Furthermore, a mildly significant (p<0.05) correlation with SNA angles was noted. Changes in the soft-tissue profile angle (N'-Sn-Pog') were moderately significant (p<0.001). The profile angles of the upper lip (Sn-SS-Ls) and lower lip (Pog'-Sm-Li) did not reveal significant changes. We observed highly significant (p<0.001) findings in the esthetic line (NsPog') advancement relative to the Ls and Li landmarks and in increases in lower-face height. Midface heights remained unchanged. Our results indicate that treatment with bite-jumping appliances results in increased facial convexity, advancement of the esthetic line, and increased lower-face height. The cumulative effects of growth and treatment do not, however, appear pronounced enough to result in a preference for or against treatment with a bite-jumping appliance.

Effects of fixed appliances in correcting Angle Class II on the depth of the posterior airway space: FMA vs. Herbst appliance--a retrospective cephalometric study.

OBJECTIVE: The objective of this retrospective study based on the metric evaluation of lateral cephalograms was to investigate the extent to which treatment with two different fixed appliances for the correction of Angle Class II influenced the morphology of the extrathoracic airway space (the posterior airway space, PAS). PATIENTS AND METHODS: A total of 43 patients with Angle Class II malocclusion were classified into two groups according to the appliance used for treatment: the functional mandibular advancer (FMA; n = 18) or the Herbst appliance (n = 25). Lateral cephalograms were taken of each patient at the start of functional jaw orthopedic treatment (time point T1) and at its completion (time point T2). Specific distances and angles were measured and analyzed in a cephalometric analysis. RESULTS: We observed major differences among the 43 patients in the depth of the posterior airway space during treatment with fixed appliances for Angle Class II correction. Regression analysis revealed that changes in sagittal and vertical positions had different effects on the depth of specific PAS sections: increases in anterior facial height are associated proportionately with increases in PAS width, particularly in the upper region. On the other hand, increases in posterior facial height and in the mandible's forward displacement correlated inversely to the decreases in depth, particularly in the central and lower PAS regions. The two treatment appliances (FMA, Herbst appliance) had the same effects on extrathoracic airway depth. CONCLUSIONS: Analyses of lateral cephalograms indicate that Angle Class II treatment with fixed appliances does not prevent sleep apnea in patients at risk. Nevertheless, this study does not permit absolutely reliable conclusions about the dimensions of the pharyngeal airway space. As the lateral cephalogram provides good images of structures in the midsagittal plane but is incapable of imaging the transverse dimension, there is an automatic lack of information concerning the precise width and volume of the extrathoracic airway space.