Bacteria associated with the in hospite Symbiodiniaceae's phycosphere.

Symbiotic interactions between Symbiodiniaceae and bacteria are still poorly explored, especially those in hospite. Here, we adapted a technique that allows for the enrichment of intact and metabolically active in hospite Symbiodiniaceae cells (ihSC) and their associated bacteria from the tissue of the model coral Pocillopora damicornis, using a discontinuous gradient of solution of isotonic Percoll (SIP). The ihSC were concentrated in the 50% SIP fraction, as determined by microscopy. The presence of bacteria associated with ihSC was confirmed by fluorescence in situ hybridization, while microbiome analysis indicated that bacteria of the families Halieaceae, Flavobacteriaceae, and Alcanivoraceae are significantly associated with ihSC. Extracellular vesicles that could be exuding molecules were detected on the symbiosome membranes. Our technique and data contribute to elucidate ihSC-bacteria interactions.

Disinfection and Shaping of Vertucci Class II Root Canals after Preparation with Two Instrument Systems and Supplementary Ultrasonic Activation of Sodium Hypochlorite.

INTRODUCTION: This study compared disinfection and shaping after root canal preparation with either XP-endo Shaper or TruNatomy instrument systems, supplemented by ultrasonic activation of sodium hypochlorite (NaOCl) with either stainless-steel (SS) or nickel-titanium (NiTi) inserts. METHODS: Mesial roots from mandibular molars with Vertucci class II configuration were divided into 2 groups (n = 24) based on anatomically paired micro-computed tomography (micro-CT) analyses. Pre and postpreparation micro-CT scans were obtained to evaluate the shaping performance. The canals were contaminated with a mixed bacterial culture for 30 days and then subjected to preparation with either XP-endo Shaper or TruNatomy instruments using NaOCl irrigation. Supplementary ultrasonic activation of NaOCl was conducted using either an SS (TruNatomy group) or NiTi (XP-endo Shaper group) insert. Bacteriological samples were taken from the canals before preparation (S1), after preparation (S2), and after the supplementary approach (S3). Bacterial reduction was evaluated using a quantitative real-time polymerase chain reaction. RESULTS: Preparation with both instrument systems significantly reduced bacterial counts (P < .01). After preparation, 36% (TruNatomy) and 35% (XP-endo Shaper) were negative for bacteria. These values increased to 59% and 65% after ultrasonic activation with the SS and NiTi inserts, respectively. The quantitative data in S2 showed that XP-endo Shaper promoted a significantly higher bacterial reduction than TruNatomy (P < .05). No significant intragroup differences were observed after ultrasonic activation (P > .05), probably because the SS insert promoted a significantly higher S2-to-S3 reduction than the NiTi insert (P < .01). Micro-CT analysis revealed no significant differences in the unprepared areas between the groups (P > .05). CONCLUSIONS: The XP-endo Shaper caused a significantly higher bacterial reduction than TruNatomy in Vertucci class II canals. Better antibacterial results after ultrasonic activation were observed for the SS ultrasonic inserts than for the NiTi inserts.

Species-specific relationships between deep sea sponges and their symbiotic Nitrosopumilaceae.

Sponges thrive in the deep, dark and nutrient-depleted ocean and may rely on microbial symbionts for carbon acquisition and energy generation. However, these symbiotic relationships remain largely unexplored. In this study, we analyze the microbiome of deep-sea sponges and show that ammonia-oxidizing archaea (AOA) of the family Nitrosopumilaceae make up at least 75% of the microbial communities of the sponges Aphrocallistes sp., Farrea sp. and Paratimea sp.. Given the known autotrophic metabolism of AOAs, this implies that these sponge holobionts can have the capacity for primary production in the deep-sea. We also show that specific AOA lineages are highly specific towards their hosts, hinting towards an unprecedent vertical transmission of these symbionts in deep-sea sponges. Our results show that the ecology and evolution of symbiotic relationships in deep-sea sponge is distinct from that of their shallow-water counterparts.

In situ devices can culture the microbial dark matter of corals.

Most microorganisms found in environmental samples have never been cultured and can often only be explored through molecular or microscopic approaches. Here, we adapt the use of in situ diffusion-based devices to culture "yet-to-be-cultured" microorganisms associated with coral mucus and compare this with a traditional culturing method. The culturability of microorganisms associated with mucus of the coral Pocillopora damicornis increased by 420% and 570% with diffusion growth chambers and microwell chip devices, respectively, compared with the traditional method tested. The obtained cultures represent up to 64.4% of the total diversity of amplicon sequence variants (ASVs) found in the mucus of the coral P. damicornis. In addition, some previously uncultured microorganisms, such as members of the family Nitrosopumilaceae and halophilic/halotolerant bacteria were cultured. Our results validate alternative microbial culturing strategies to culture coral-associated microorganisms, while significantly increasing the culturability of previous microbial dark matter.

Global qualitative and quantitative distribution of micropollutants in the deep sea.

Micropollutants (MPs) include a wide range of biological disruptors that can be toxic to wildlife and humans at very low concentrations (<1 µg/L). These mainly anthropogenic pollutants have been widely detected in different areas of the planet, including the deep sea, and have impacts on marine life. Because of this potential toxicity, the global distribution, quantity, incidence, and potential impacts of deep-sea MPs were investigated in a systematic review of the literature. The results showed that MPs have reached different zones of the ocean and are more frequently reported in the Northern Hemisphere, where higher concentrations are found. MPs are also concentrated in depths up to 3000 m, where they are also more frequently studied, but also extend deeper than 10,000 m. Potentially toxic metals (PTMs), polychlorinated biphenyls (PCBs), dichlorodiphenyltrichloroethane (DDTs), organotins, and polycyclic aromatic hydrocarbons (PAHs) were identified as the most prevalent and widely distributed MPs at ≥200 m depth. PTMs are widely distributed in the deep sea in high concentrations; aluminum is the most prevalent up to 3000 m depth, followed by zinc and copper. PCBs, organotins, hexachlorocyclohexanes (HCHs), PAHs, and phenols were detected accumulated in both organisms and environmental samples above legislated thresholds or known toxicity levels. Our assessment indicated that the deep sea can be considered a sink for MPs.

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.

Coral microbiome manipulation elicits metabolic and genetic restructuring to mitigate heat stress and evade mortality.

Beneficial microorganisms for corals (BMCs) ameliorate environmental stress, but whether they can prevent mortality and the underlying host response mechanisms remains elusive. Here, we conducted omics analyses on the coral Mussismilia hispida exposed to bleaching conditions in a long-term mesocosm experiment and inoculated with a selected BMC consortium or a saline solution placebo. All corals were affected by heat stress, but the observed "post-heat stress disorder" was mitigated by BMCs, signified by patterns of dimethylsulfoniopropionate degradation, lipid maintenance, and coral host transcriptional reprogramming of cellular restructuration, repair, stress protection, and immune genes, concomitant with a 40% survival rate increase and stable photosynthetic performance by the endosymbiotic algae. This study provides insights into the responses that underlie probiotic host manipulation. We demonstrate that BMCs trigger a dynamic microbiome restructuring process that instigates genetic and metabolic alterations in the coral host that eventually mitigate coral bleaching and mortality.

Multi-domain probiotic consortium as an alternative to chemical remediation of oil spills at coral reefs and adjacent sites.

BACKGROUND: Beginning in the last century, coral reefs have suffered the consequences of anthropogenic activities, including oil contamination. Chemical remediation methods, such as dispersants, can cause substantial harm to corals and reduce their resilience to stressors. To evaluate the impacts of oil contamination and find potential alternative solutions to chemical dispersants, we conducted a mesocosm experiment with the fire coral Millepora alcicornis, which is sensitive to environmental changes. We exposed M. alcicornis to a realistic oil-spill scenario in which we applied an innovative multi-domain bioremediator consortium (bacteria, filamentous fungi, and yeast) and a chemical dispersant (Corexit® 9500, one of the most widely used dispersants), to assess the effects on host health and host-associated microbial communities. RESULTS: The selected multi-domain microbial consortium helped to mitigate the impacts of the oil, substantially degrading the polycyclic aromatic and n-alkane fractions and maintaining the physiological integrity of the corals. Exposure to Corexit 9500 negatively impacted the host physiology and altered the coral-associated microbial community. After exposure, the abundances of certain bacterial genera such as Rugeria and Roseovarius increased, as previously reported in stressed or diseased corals. We also identified several bioindicators of Corexit 9500 in the microbiome. The impact of Corexit 9500 on the coral health and microbial community was far greater than oil alone, killing corals after only 4 days of exposure in the flow-through system. In the treatments with Corexit 9500, the action of the bioremediator consortium could not be observed directly because of the extreme toxicity of the dispersant to M. alcicornis and its associated microbiome. CONCLUSIONS: Our results emphasize the importance of investigating the host-associated microbiome in order to detect and mitigate the effects of oil contamination on corals and the potential role of microbial mitigation and bioindicators as conservation tools. Chemical dispersants were far more damaging to corals and their associated microbiome than oil, and should not be used close to coral reefs. This study can aid in decision-making to minimize the negative effects of oil and dispersants on coral reefs. Video abstract.

Estrogen induces shift in abundances of specific groups of the coral microbiome.

Synthetic estrogens such as ethinylestradiol (EE2) are persistent micropollutants that are not effectively removed from wastewater by conventional treatments. These contaminants are released into waterbodies, where they disrupt endocrine systems of organisms and cause harmful effects such as feminization, infertility, reproduction problems and genital malformations. The consequences of this pollution for key marine ecosystems such as coral reefs and their associated microbiomes are underexplored. We evaluated the effects of EE2 concentrations of 100 ng L-1 and 100 µg L-1 on the coral metaorganism Mussismilia harttii. The results indicated no effects on visible bleaching or Fv/Fm ratios in the corals during a 17-day microcosm experiment. However, next-generation sequencing of 16S rDNA revealed a statistically significant effect of high EE2 concentrations on OTU richness, and shifts in specific microbial groups after treatments with or without EE2. These groups might be bioindicators of early shifts in the metaorganism composition caused by EE2 contamination.

Assessing the impact of synthetic estrogen on the microbiome of aerated submerged fixed-film reactors simulating tertiary sewage treatment and isolation of estrogen-degrading consortium.

17α-ethinylestradiol (EE2) is a synthetic estrogen that can cause harmful effects on animals, such as male feminization and infertility. However, the impact of the EE2 contamination on microbial communities and the potential role of bacterial strains as bioremediation agents are underexplored. The aim of this work was to evaluate the impact of EE2 on the microbial community dynamics of aerated submerged fixed-film reactors (ASFFR) simulating a polishing step downstream of a secondary sewage treatment. For this purpose, the reactors were fed with a synthetic medium with low COD content (around 50 mg l-1), supplemented (reactor H) or not (reactor C) with 1 µg l-1 of EE2. Sludge samples were periodically collected during the bioreactors operation to assess the bacterial profile over time by 16S rRNA gene amplicon sequencing or by bacterial isolation using culture-dependent approach. The results revealed that the most abundant phyla in both reactors were Proteobacteria and Bacteroidetes. At genus level, Chitinophagaceae, Nitrosomonas and Bdellovibrio predominated. Significant effects caused by EE2 treatment and bioreactors operating time were observed by non-metric multidimensional scaling. Therefore, even at low concentrations as 1 µg l-1, EE2 is capable of influencing the bioreactor microbiome. Culture-dependent methods showed that six bacterial isolates, closely related to Pseudomonas and Acinetobacter genera, could grow on EE2 as the sole carbon source under aerobic conditions. These organisms may potentially be used for the assembly of an EE2-degrading bacterial consortium and further exploited for bioremediation applications, including tertiary sewage treatment to remove hormone-related compounds not metabolized in secondary depuration stages.

Prospecting Microbial Strains for Bioremediation and Probiotics Development for Metaorganism Research and Preservation.

Pollution affects all biomes. Marine environments have been particularly impacted, especially coral reefs, one of the most sensitive ecosystems on Earth. Globally, 4.5 billion people are economically dependent on the sea, where most of their livelihood is provided by coral reefs. Corals are of great importance and therefore their extinction leads to catastrophic consequences. There are several possible solutions to remediate marine pollutants and local contamination, including bioremediation. Bioremediation is the capacity of organisms to degrade contaminants. The approach presents several advantages, such as sustainability, relatively low cost, and the fact that it can be applied in different ecosystems, causing minimal impacts to the environment. As an extra advantage, the manipulation of endogenous microbiomes, including putative beneficial microorganisms for corals (pBMCs), may have probiotic effects for marine animals. In this context, the use of the two approaches, bioremediation and pBMC inoculation combined, could be promising. This strategy would promote the degradation of specific pollutants that can be harmful to corals and other metaorganisms while also increasing host resistance and resilience to deal with pollution and other threats. This method focuses on the selection of pBMCs to degrade two contaminants: the synthetic estrogen 17a-ethinylestradiol (EE2) and crude oil. Both have been reported to negatively impact marine animals, including corals, and humans. The protocol describes how to isolate and test bacteria capable of degrading the specific contaminants, followed by a description of how to detect some putative beneficial characteristics of these associated microbes to their coral host. The methodologies described here are relatively cheap, easy to perform, and highly adaptable. Almost any kind of soluble target compound can be used instead of EE2 and oil.

Water contamination by endocrine disruptors: Impacts, microbiological aspects and trends for environmental protection.

Hormone active agents constitute a dangerous class of pollutants. Among them, those agents that mimic the action of estrogens on target cells and are part of the group of endocrine-disruptor compounds (EDCs) are termed estrogenic EDCs, the main focus of this review. Exposure to these compounds causes a number of negative effects, including breast cancer, infertility and animal hermaphroditism. However, especially in underdeveloped countries, limited efforts have been made to warn people about this serious issue, explain the methods of minimizing exposure, and develop feasible and efficient mitigation strategies at different levels and in various environments. For instance, the use of bioremediation processes capable of transforming EDCs into environmentally friendly compounds has been little explored. A wide diversity of estrogen-degrading microorganisms could be used to develop such technologies, which include bioremediation processes for EDCs that could be implemented in biological filters for the post-treatment of wastewater effluent. This review describes problems associated with EDCs, primarily estrogenic EDCs, including exposure as well as the present status of understanding and the effects of natural and synthetic hormones and estrogenic EDCs on living organisms. We also describe potential biotechnological strategies for EDC biodegradation, and suggest novel treatment approaches for minimizing the persistence of EDCs in the environment.