Search Results (29)

In this study, the distribution patterns of the nitrifying and denitrifying microbiome in a large-scale biofilter (587.24 m³) in a cold freshwater recirculating aquaculture system (RAS) was investigated. Previous studies have revealed that the water quality, nitrification, and denitrification rates in the front (BFF), middle (BFM), and back (BFB) of this biofilter are different. The results showed the highest diversity of the denitrifying microbiome in the BFB, followed by BFF and BFM, whereas nitrifying microbiome diversity remained consistent across different positions. Two genera, Nitrosomonas and Nitrosospira, dominated the nitrifying microbiome, while Pseudomonas, Thauera, Cupriavidus, Dechloromonas, Azoarcus, and Paracoccus comprised the top six denitrifying genera. Principal coordinate analysis indicated a distinct spatial distribution pattern of the denitrifying microbiome but not the nitrifying microbiome. The genera Pseudomonas and Dechloromonas were the biomarkers of the BFF and BFB, respectively. Redundancy analysis showed that nitrite, nitrate, dissolved oxygen, and soluble reactive phosphorus influenced the functional microbiome distribution pattern. Network correlation analysis identified one nitrifying hub (Nitrosospira) in the BFF, five denitrifying hubs (Aromatoleum, Dechloromonas, Paracoccus, Ruegeria, and Thauera) in the BFM, and three denitrifying hubs (Azoarcus, Magnetospirillum, and Thauera) in the BFB. Exclusively negative correlations were found between hubs and its adjacent nodes in the BFF and BFB. This study demonstrates that habitat can shape the distribution patterns of the nitrifying and denitrifying microbiome in the biofilter of the RAS, with the BFF exhibiting greater benefits for the nitrification process. Full article

The mevalonate pathway is crucial for synthesizing isopentenyl pyrophosphate (IPP), the universal precursor of terpenoids, with 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) serving as the rate-determining enzyme that catalyzes the reduction of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) to mevalonate, requiring NAD(P)H as an electron donor. Improving the cofactor promiscuity of HMGR can facilitate substrate utilization and terpenoid production by overcoming cofactor specificity limitations. In this study, we heterologously expressed rpHMGR from Ruegeria pomeroyi in Escherichia coli BL21(DE3) for the first time and established that it predominantly utilizes NADH. To broaden its cofactor usage, we employed Molecular Operating Environment (MOE)-assisted design to engineer the cofactor binding site, creating a dual-cofactor-utilizing mutant, D154K (the substitution of aspartic acid with lysine at residue 154). This mutant exhibited a significant 53.7-fold increase in activity toward NADPH, without compromising protein stability at physiological temperatures. The D154K mutant displayed an optimal pH of 6, maintaining over 80% of its catalytic activity across the pH range of 6–8, regardless of whether NADH or NADPH was the cofactor. These findings highlight the value of rational design, enhance our understanding of HMGR-cofactor recognition mechanisms, and provide a foundation for future efforts to optimize and engineer HMGR for broader cofactor flexibility. Full article

Pathogenic infections can reshape the intestinal microbiota of aquatic animals, thereby impacting their health status. In this study, we aimed to investigate whether Vibrio parahaemolyticus infection induces dysbiosis in the intestinal bacterial community of Penaeus vannamei and to assess the associated ecological risks. Our findings revealed the deterministic processes in intestinal bacterial community assembly during bacterial infections, indicating that host selection, i.e., host immune response post-infection, has a significant influence on intestinal microbes. More importantly, we found that bacterial infection reshaped the intestinal community by reducing the relative abundance of probiotic Ruegeria species (e.g., R. atlantica, R. lacuscaerulensis, R. conchae, R. profundi, R. arenilitoris, R. pomeroyi) and increasing the relative abundance of Vibrio species (V. harveyi, V. sinaloensis, V. coralliilyticus, and V. brasiliensis). Significant negative correlations were observed between the relative abundance of these Ruegeria species and the relative abundance of Vibrio species. Moreover, the control P. vannamei contained a substantially higher number of keystone species belonging to Ruegeria in the bacterial community network, whereas bacterial infection individuals had few or no keystone species belonging to Ruegeria, with keystone species belonging to Vibrio becoming more prominent. Thus, the significant increase in Vibrio species abundance in the P. vannamei intestine following bacterial infection was associated with the marked reduction in Ruegeria species. Our findings will provide valuable insights into the complex interactions among bacterial infection, intestinal microbiota, and host health, and they provide guidance for the development of probiotics in promoting the healthy culture of P. vannamei. Full article

Mariculture wastewater is an intractable wastewater, owing to its high salinity inhibiting microbial metabolism. The biocarrier bacterial–microbial consortium (BBM) and bacterial–microbial consortium (BM) were developed to investigate the mechanism of pollutant degradation and microbial community evolution. The BBM exhibited excellent mariculture wastewater treatment, with the highest removal for TOC (91.78%), NH₄⁺-N (79.33%) and PO₄³⁻-P (61.27%). Biocarriers accelerated anaerobic region formation, with the levels of denitrifying bacteria accumulation improving nitrogen degradation in the BBM. Moreover, the biocarrier enhanced the production of soluble microbial products (SMPs) (11.53 mg/L) and extracellular polymeric substances (EPSs) (370.88 mg/L), which accelerated the formation of bacterial and microalgal flocs in the BBM. The fluorescence excitation–emission matrix (EEM) results demonstrated that the addition of biocarriers successfully decreased the production of aromatic-like components in anoxic and aerobic supernatants. Additionally, the biocarrier shifted the bacterial community constitutions significantly. Biocarriers provided an anoxic microenvironment, which enhanced enrichments of Rhodobacteraceae (66%) and Ruegeria (70%), with a satisfying denitrification in the BBM. This study provided a novel biocarrier addition to the BBM system for actual mariculture wastewater treatment. Full article

The microbiome of the shrimp’s digestive tract shows differences between healthy and acute hepatopancreatic necrosis disease (AHPND)-affected shrimp. The present study aimed to evaluate the impact of probiotic consumption on the microbial community in experimentally AHPND-infected shrimp. Effective probiotics (EPs) Vibrio alginolyticus (Va32A), V. campbellii (VcHA), and Bacillus pumilus (BPY100) and non-effective probiotics (NEPs) B. pumilus (Bp43, and BpY119), were employed in bioassays with Penaeus vannamei and challenged with AHPND-causing V. parahaemolyticus (Vp_AHPND). Stomach (Sto), intestine (Int), and hepatopancreas (Hep) were analyzed by metabarcoding (16S rRNA gene) to characterize the microbiome and biomarkers. Hep-VcHA showed the highest alpha diversity (Shannon index = 5.88; 166 ASVs), whereas the lowest was for Hep-Bp43 (2.33; 7 ASVs). Proteobacteria, Actinobacteria, Bacteroidetes, and Saccharibacteria were the most abundant phyla. The relative abundance of Vibrio sp. was the highest in the Hep and Int of Bp43, BPY119 and the positive control, followed by Rhodobacteraceae in the EP group. Principle coordinate analysis (PCoA) showed a cluster grouped negative (Sto and Hep) control with almost all organs in the EP group causing 28.79% of the variation. The core microbiome of EP was mainly represented by Rhodobacteraceae, Caldilineaceae, Celeribacter indicus, Illumatobacter, Microbacterium, Ruegeria atlantica, Saccharibacteria sp., Shimia biformata, and Thalassobius mediterraneus, whose relative abundance was enriched by probiotics, which may explain their protective roles against Vp_AHPND, whereas the low survival in the NEP group was associated with a higher diversity of Vibrio spp. Our results present an ecosystem-friendly alternative based on beneficial microorganisms to prevent and control AHPND and probably other bacterial diseases in shrimp farming. Full article

Biofloc technology (BFT) is an advanced aquaculture method that uses microbial communities to enhance water quality and support aquatic species cultivation. Our research aims to delve into the pivotal role of aeration intensity within BFT systems, revealing its influence on microbial community structures, water quality, and nutrient cycling for L. vannamei culture. Three aeration levels were set with intensities of V75 (75 L/min), V35 (35 L/min), and V10 (10 L/min). The results showed that the lowest aeration intensity (V10) resulted in larger floc sizes and a reduction in the 2D-fractal dimensions, indicating a decreased overall structural complexity of the bioflocs. In addition, water quality parameters, including total ammonia nitrogen and nitrite, remained low across all treatments, highlighting the water-purifying capacity of biofloc. While protein and lipid contents in biofloc did not differ significantly among treatments, docosahexaenoic acid (DHA) levels were highest in the V75 treatment, suggesting that higher aeration promotes the accumulation of essential fatty acids. RDA analysis revealed that microorganisms like Ruegeria sp. and Sulfitobacter mediterraneus negatively correlated with ammonia and nitrite levels, suggesting their key role in converting ammonia to nitrite and nitrate in marine nitrogen cycles. The functional annotation of metagenomes across different aeration levels showed the similarly active roles of microorganisms in nitrogen metabolism and protein synthesis. In conclusion, while variations in aeration intensity affect floc size and the accumulation of essential fatty acids in biofloc, they do not significantly impact overall water quality or core microbial functions in L. vannamei aquaculture. Future research should focus on the effects of aeration strategies on microbial community dynamics and the integration of these data with performance metrics in L. vannamei. These insights can help optimize biofloc cultivation and enhance environmental sustainability in the aquaculture industry. Full article

The symbiotic relationship between corals and their associated microorganisms is crucial for the health of coral reef eco-environmental systems. Recently, there has been a growing interest in unraveling how the manipulation of symbiont nutrient cycling affects the stress tolerance in the holobiont of coral reefs. However, most studies have primarily focused on coral–Symbiodiniaceae–bacterial interactions as a whole, neglecting the interactions between Symbiodiniaceae and bacteria, which remain largely unexplored. In this study, we proposed a hypothesis that there exists an inner symbiotic loop of Symbiodiniaceae and bacteria within the coral symbiotic loop. We conducted experiments to demonstrate how metabolic exchanges between Symbiodiniaceae and bacteria facilitate the nutritional supply necessary for cellular growth. It was seen that the beneficial bacterium, Ruegeria sp., supplied a nitrogen source to the Symbiodiniaceae strain Durusdinium sp., allowing this dinoflagellate to thrive in a nitrogen-free medium. The Ruegeria sp.–Durusdinium sp. interaction was confirmed through ¹⁵N-stable isotope probing–single cell Raman spectroscopy, in which ¹⁵N infiltrated into the bacterial cells for intracellular metabolism, and eventually the labeled nitrogen source was traced within the macromolecules of Symbiodiniaceae cells. The investigation into Symbiodiniaceae loop interactions validates our hypothesis and contributes to a comprehensive understanding of the intricate coral holobiont. These findings have the potential to enhance the health of coral reefs in the face of global climate change. Full article

Immobilisation plays an important role in the industrial application of enzymes. The stabilisation and reusability of immobilised enzymes reduce the cost of the catalyst and facilitate their use in continuously operated reactors. For this purpose, an applicable type of immobilisation needs to be identified. In this study, we investigate the conversion of CDP and PolyP to CTP by NDP polyphosphate phosphotransferase 3 from Ruegeria pomeroyi (RpPPK2-3) and describe the covalent immobilisation of RpPPK2-3. In order to select a suitable carrier for the immobilisation of RpPPK2-3, a screening with different amino methacrylate (glutaraldehyde-pre-activated) and epoxy methacrylate carriers was carried out. The epoxy methacrylate carrier ECR8209M (Purolite^®) was found to be the most suitable. With a half-life of 462 d when stored at 6 °C and a 50-fold reusability with a 93% residual activity, the immobilised enzyme showed a higher stability compared to the soluble enzyme with a half-life of 0.04 d. Although the half-life of the soluble enzyme could be increased to 32 d by adding PP_i, it could not reach the stability of the immobilisate. Due to the resilience of the immobilisate, it is suitable for application in continuous reactor set-ups, e.g., packed-bed reactors. Full article

A 90 d feeding experiment was conducted to investigate the effects of vitamin E (VE) on growth, intestinal microbiota, immune response, and related gene expression of juvenile sea urchin (Strongylocentrotus intermedius). Six dry feeds were made to contain graded levels of VE (78, 105, 152, 235, 302, and 390 mg/kg); these were named E78, E105, E152, E235, E302, and E390, respectively. Dry feed E50 and fresh kelp (HD) were used as the control diets. There were six replicates of cages in each dietary group, and each cage held 20 sea urchins with an initial body weight of approximately 1.50 g. Results exhibited that weight gain rate and gonadosomatic index (GSI) of the sea urchins were not significantly affected by dietary VE ranging from 78 to 390 mg/kg. Sea urchins in the dry feed groups showed poorer growth performance, but significantly higher GSI than those in the fresh kelp groups. The pepsin and lipase activities were not significantly promoted by low or moderate VE, but were inhibited by a high level of VE (302–390 mg/kg), while amylase and cellulase activities were significantly increased by low or moderate VE, with the highest values observed in the E105 and E235 groups, respectively. VE addition at a low dosage (105–152 mg/kg) showed inhibitory effects on immune and antioxidant enzyme activities and expression of inflammation-related genes, but showed no beneficial effects at moderate or high dosage (235–390 mg/kg), while a moderate or relatively higher level of VE (235–302 mg/kg) significantly increased the expression of several immune-related genes. The relative abundance of Proteobacteria, Actinobacteria, Ruegeria, and Maliponia in the intestine of the sea urchins increased with the increase in VE in the dry feeds. On the contrary, the relative abundance of the Firmicutes, Bacteroidetes, Escherichia-Shigella, Bacteroides, and Clostridium sensu stricto 1 gradually decreased as VE content increased. These results indicated that a moderate level of VE (172.5–262.4) can achieve ideal digestive enzyme activities and growth performance, but a relatively higher level of VE (235–302 mg/kg) was beneficial for maintaining the immune and antioxidant capacity of juvenile S. intermedius by regulating the expression of inflammation- and immune-related genes and abundance of some bacteria to a healthy state. Full article

As the intensive development of aquaculture persists, the demand for fishmeal continues to grow; however, since fishery resources are limited, the price of fishmeal remains high. Therefore, there is an urgent need to develop new sources of protein. They are rich in proteins, fatty acids, amino acids, chitin, vitamins, minerals, and antibacterial substances. Maggot meal-based diet is an ideal source of high-quality animal protein and a new type of protein-based immune enhancer with good application prospects in animal husbandry and aquaculture. In the present study, we investigated the effects of three different diets containing maggot protein on the growth and intestinal microflora of Litopenaeus vannamei. The shrimp were fed either a control feed (no fly maggot protein added), FM feed (compound feed with 30% fresh fly maggot protein added), FF feed (fermented fly maggot protein), or HT feed (high-temperature pelleted fly maggot protein) for eight weeks. The results showed that fresh fly maggot protein in the feed was detrimental to shrimp growth, whereas fermented and high-temperature-pelleted fly maggot protein improved shrimp growth and survival. The effects of different fly maggot protein treatments on the intestinal microbiota of L. vannamei also varied. Fermented fly maggot protein feed and high-temperature-pelleted fly maggot protein feed increased the relative abundance of Ruegeria and Pseudomonas, which increased the abundance of beneficial bacteria and thus inhibited the growth of harmful bacteria. In contrast, fresh fly maggot proteins alter the intestinal microbiome, disrupting symbiotic relationships between bacteria, and causing invasion by Vibrio and antibiotic-resistant bacteria. These results suggest that fresh fly maggot proteins affect the composition of intestinal microorganisms, which is detrimental to the intestinal tract of L. vannamei, whereas fermented fly maggot protein feed affected the growth of L. vannamei positively by improving the composition of intestinal microorganisms. Full article

Bacterial communities associated with fish larvae are highly influenced by the microbiota of live prey used as feed (rotifers or Artemia), generally dominated by bacterial strains with a low degree of specialization and high growth rates, (e.g., Vibrionaceae), which can be detrimental to larvae. Co-cultivation of microalgae used in the enrichment of Artemia (e.g., Phaeodactylum tricornutum, or Chlorella minutissima) with Vibrio-antagonistic probiotics belonging to the Roseobacter clade bacteria (e.g., Phaeobacter spp. or Ruegeria spp.) was studied. The introduction of the probiotics did not affect microalgae growth or significantly modify the composition of bacterial communities associated with both microalgae, as revealed by DGGE analysis. The inoculation of P. tricornutum with Ruegeria ALR6 allowed the maintenance of the probiotic in the scale-up of the microalgae cultures, both in axenic and non-axenic conditions. Using Ruegeria-inoculated P. tricornutum cultures in the enrichment of Artemia reduced the total Vibrionaceae count in Artemia by 2 Log units, therefore preventing the introduction of opportunistic or pathogenic bacteria to fish larvae fed with them. Full article

Greater amberjack (Seriola dumerili) is a new species in marine aquaculture with high mortalities at the larval stages. The microbiota of amberjack larvae was analyzed using 16S rDNA sequencing in two groups, one added copepod nauplii (Acartia tonsa) in the diet, and one without copepods (control). In addition, antagonistic bacteria were isolated from amberjack larvae and live food cultures. Proteobacteria was the most abundant phylum followed by Bacteroidota in amberjack larvae. The composition and diversity of the microbiota were influenced by age, but not by diet. Microbial community richness and diversity significantly increased over time. Rhodobacteraceae was the most dominant family followed by Vibrionaceae, which showed the highest relative abundance in larvae from the control group 31 days after hatching. Alcaligenes and Thalassobius genera exhibited a significantly higher relative abundance in the copepod group. Sixty-two antagonistic bacterial strains were isolated and screened for their ability to inhibit four fish pathogens (Aeromonas veronii, Vibrio harveyi, V. anguillarum, V. alginolyticus) using a double-layer test. Phaeobacter gallaeciensis, Phaeobacter sp., Ruegeria sp., and Rhodobacter sp. isolated from larvae and Artemia sp. inhibited the fish pathogens. These antagonistic bacteria could be used as host-derived probiotics to improve the growth and survival of the greater amberjack larvae. Full article

Copiotrophic bacteria that respond rapidly to nutrient availability, particularly high concentrations of carbon sources, play indispensable roles in marine carbon cycling. However, the molecular and metabolic mechanisms governing their response to carbon concentration gradients are not well understood. Here, we focused on a new member of the family Roseobacteraceae isolated from coastal marine biofilms and explored the growth strategy at different carbon concentrations. When cultured in a carbon-rich medium, the bacterium grew to significantly higher cell densities than Ruegeria pomeroyi DSS-3, although there was no difference when cultured in media with reduced carbon. Genomic analysis showed that the bacterium utilized various pathways involved in biofilm formation, amino acid metabolism, and energy production via the oxidation of inorganic sulfur compounds. Transcriptomic analysis indicated that 28.4% of genes were regulated by carbon concentration, with increased carbon concentration inducing the expression of key enzymes in the EMP, ED, PP, and TCA cycles, genes responsible for the transformation of amino acids into TCA intermediates, as well as the sox genes for thiosulfate oxidation. Metabolomics showed that amino acid metabolism was enhanced and preferred in the presence of a high carbon concentration. Mutation of the sox genes decreased cell proton motive force when grown with amino acids and thiosulfate. In conclusion, we propose that copiotrophy in this Roseobacteraceae bacterium can be supported by amino acid metabolism and thiosulfate oxidation. Full article

Mediterranean mussels (Mytilus galloprovincialis), due to their nutritional mechanisms which involve filtering huge amounts of water, are affected by seawater pollution and can host microbial diversity of environmental origin, as well as pathogenic bacteria that must be constantly monitored. Herein, we applied a Next Generation Sequencing (NGS) metabarcoding approach in order to study the M. galloprovincialis microbiota. Collection of samples was conducted during winter and summer months from various mussel farm zones located in specific farm regions in the Thermaikos gulf, the northern Aegean Sea, Greece. A microbiological test was performed for the enumeration of Escherichia coli and the presence of Salmonella sp. DNA extraction and amplification of the whole bacterial 16S rRNA gene, followed by NGS amplicon sequencing and taxonomic classification, were carried out. Statistically significant differences (p < 0.05) in the abundance of the most dominant bacterial phyla, families and genera between winter and summer time periods, regions, as well as zones within each region of sampling, were evaluated with z-score computation. According to the obtained results, the most prevalent taxa at the genus level were Mycoplasma (12.2%), Anaplasma (5.8%), Ruegeria (5.2%) and Mariniblastus (2.1%). Significant differences in the abundance of the most dominant genera were found at all levels of comparison (seasons, regions and zones within each region), highlighting the dynamic character of microorganisms, which might be affected by microenvironmental, temporal and spatial changes. The present research contributes to the characterization of M. galloprovincialis microbiome in areas that have not been studied previously, setting the baseline for future, more thorough investigations of the specific bivalve species and its bacterial profile in the above geographic regions. Full article

Loss of algal production from the crashes of algal mass cultivation systems represents a significant barrier to the economic production of microalgal-based biofuels. Current strategies for crash prevention can be too costly to apply broadly as prophylaxis. Bacteria are ubiquitous in microalgal mass production cultures, however few studies investigate their role and possible significance in this particular environment. Previously, we demonstrated the success of selected protective bacterial communities to save Microchloropsis salina cultures from grazing by the rotifer Brachionus plicatilis. In the current study, these protective bacterial communities were further characterized by fractionation into rotifer-associated, algal-associated, and free-floating bacterial fractions. Small subunit ribosomal RNA amplicon sequencing was used to identify the bacterial genera present in each of the fractions. Here, we show that Marinobacter, Ruegeria, and Boseongicola in algae and rotifer fractions from rotifer-infected cultures likely play key roles in protecting algae from rotifers. Several other identified taxa likely play lesser roles in protective capability. The identification of bacterial community members demonstrating protective qualities will allow for the rational design of microbial communities grown in stable co-cultures with algal production strains in mass cultivation systems. Such a system would reduce the frequency of culture crashes and represent an essentially zero-cost form of algal crop protection. Full article