Impacts of eutrophication on microbial community structure in sediment, seawater, and phyllosphere of seagrass ecosystems.

Introduction: Seagrass-associated microbial communities play a crucial role in the growth and health of seagrasses. However, like seagrass meadows, seagrass-associated microbial communities are often affected by eutrophication. It remains unclear how eutrophication influences the composition and function of microbial communities associated with different parts of seagrass. Methods: We employed prokaryotic 16S rRNA gene high-throughput sequencing combining microbial community structure analysis and co-occurrence network analysis to investigate variances in microbial community compositions, potential functions and complexities across sediment, seagrass leaves, and seawater within different eutrophic areas of two adjacent seagrass meadows on Hainan Island, China. Results: Our results indicated that microbial diversity on seagrass leaves was significantly lower than in sediment but significantly higher than in seawater. Both sediment and phyllosphere microbial diversity showed no significant difference between the highly eutrophic and less eutrophic sites in each lagoon. However, sediment microbial diversity was higher in the more eutrophic lagoon, while phyllosphere microbial diversity was higher in the less eutrophic lagoon. Heavy eutrophication increased the relative abundance of phyllosphere microorganisms potentially involved in anaerobic metabolic processes, while reducing those responsible for beneficial functions like denitrification. The main factor affecting microbial diversity was organic carbon in seawater and sediment, with high organic carbon levels leading to decreased microbial diversity. The co-occurrence network analysis revealed that heavy eutrophication notably reduced the complexity and internal connections of the phyllosphere microbial community in comparison to the sediment and seawater microbial communities. Furthermore, ternary analysis demonstrated that heavy eutrophication diminished the external connections of the phyllosphere microbial community with the sediment and seawater microbial communities. Conclusion: The pronounced decrease in biodiversity and complexity of the phyllosphere microbial community under eutrophic conditions can lead to greater microbial functional loss, exacerbating seagrass decline. This study emphasizes the significance of phyllosphere microbial communities compared to sediment microbial communities in the conservation and restoration of seagrass meadows under eutrophic conditions.

Neoagaro-Oligosaccharides Ameliorate Chronic Restraint Stress-Induced Depression by Increasing 5-HT and BDNF in the Brain and Remodeling the Gut Microbiota of Mice.

Neoagaro-oligosaccharides (NAOs) belong to the algae oligosaccharides. NAOs have been found to have diverse biological activities. However, the effects of NAOs on depression and their underlying mechanism have not been thoroughly studied. A chronic restraint stress (CRS)-induced C57BL/6J mouse model was used to assess the antidepressant effects of NAOs. Anxiety and depression behaviors were assessed by open field tests (OFT) and forced swimming tests (FST), while interleukin 18 (IL-18), 5-hydroxytryptamine (5-HT) and brain-derived neurotrophic factor (BDNF) were the molecular biomarkers of depression. Fecal microbiota transplantation (FMT) was performed. The results showed that NAO treatment significantly improved the body weight of depressed mice and reduced the central area time in the OFT and immobility time in the FST. NAO treatment decreased the levels of IL-18 in the serum and increased the levels of 5-HT in the serum and whole brain and of BDNF in the whole brain. NAO treatment mitigated the gut microbiota dysbiosis in the depressed mice and reversed the decreased levels of short-chain fatty acids (SCFAs) in the cecum of the depressed mice. FMT indicated that the gut microbiota is, indeed, linked to depression, which was reflected in the changes in weight gain and behaviors. In a word, NAOs effectively reversed the CRS-induced mice model of depression, which depended on the changes in the gut microbiota and SCFAs, as well as its modulation of 5-HT and BDNF.

The first complete genome sequence of Microbulbifer celer KCTC12973T, a type strain with multiple polysaccharide degradation genes.

Genus Microbulbifer plays important roles in element cycling process in marine environments, and the first type strain KCTC 12973T (=ISL-39T = CCUG 54356T) of M. celer was isolated and identified in 2007. However, the genome sequence of M. celer KCTC 12973T is still unclear, which complicates the functional exploration and new species identification of other species belonged to this genus. This study reported the complete genome sequence of M. celer KCTC 12973T with a genome size of 4,346,001 bp. A total of 3601 protein-coding genes were annotated in the genome. The potential genes involved in the polysaccharide degradation, including cellulose, chitin, xylan, and pectate, were found in the protein-coding genes. Besides, the reductase genes of nitrate and nitrite were also annotated in the genome. These findings indicated the potential crucial ecological functions of M. celer KCTC 12973T for carbon and nitrogen cycles in marine ecosystems.

Vibrio ziniensis sp. nov., isolated from mangrove sediments.

A novel Gram-staining-negative, catalase- and oxidase-positive, facultatively anaerobic and rod-shaped motile bacterial strain, designated as ZWAL4003T, was isolated from mangrove sediments of the Zini Mangrove Forest, Zhangzhou City, PR China. Phylogenetic analysis based on its 16S rRNA gene sequence indicated that ZWAL4003T was grouped into a separated branch with Vibrio plantisponsor MSSRF60T (97.38% nucleotide sequence identity) and Vibrio diazotrophicus NBRC 103148T (97.27%). The major cellular fatty acids were C14 : 0 (12.6%), C16 : 0 (17.6%), and summed feature 3 (C16 : 1ω6c /C16 : 1 ω7c, 45.6%). Its genome had a length of 4650556 bp with 42.8% DNA G+C content, and contained genes involved in the biosynthesis of bacteriocin, ß-lactone, resorcinol, N-acyl amino acid, and arylpolyene. The in silico DNA-DNA hybridization and average nucleotide identity values for whole-genome sequence comparisons between ZWAL4003T and V. plantisponsor LMG 24470T were clearly below the thresholds used for the delineation of a novel species. The morphological and chemotaxonomic characteristics and the genotypic data of ZWAL4003T indicated that it represented a novel species of the genus Vibrio. Its proposed name is Vibrio ziniensis sp. nov., and the type strain is ZWAL4003T (=KCTC 72971T=MCCC 1A17474T).

Amelioration of Androgenetic Alopecia by Algal Oligosaccharides Prepared by Deep-Sea Bacterium Biodegradation.

Androgenetic alopecia (AGA) is a dihydrotestosterone (DHT)-mediated hair loss disorder characterized by shortened anagen hair cycle. Oligosaccharides derived from seaweeds possess diverse biological functions. However, little is known about their effects on AGA. In this study, algal oligosaccharide (AOS) was characterized for its mitigation effects on key features involved in AGA pathogenesis, such as DHT- mediated cellular signaling and shortened anagen hair cycle. AOS with varying degrees of polymerization (DP), namely, AOS (DP2), AOS (DP4-6), and AOS (DP8-12), were prepared by agar biodegradation with Flammeovirga pacifica WPAGA1, an agarolytic bacterium isolated from deep-sea sediments. In vitro results showed that AOS with varying DPs significantly ameliorated the DHT-induced alterations of regulatory factors in human hair follicle dermal papilla cells in a dose- and DP-dependent manner, as revealed by the normalization of several hair-growth-stimulating or inhibitory factors. In vivo studies showed that AOS (DP2) extended the anagen phase and thereby delayed catagen progression in mice. Furthermore, AOS (DP2) stimulated dorsal hair growth in mice by increasing hair length, density, and thickness. Therefore, our findings indicated that AOS antagonized key factors involved in AGA pathogenesis, suggesting the potential application of AOS in the prevention and the treatment of AGA.

Production of Neoagaro-Oligosaccharides With Various Degrees of Polymerization by Using a Truncated Marine Agarase.

Bioactivities, such as freshness maintenance, whitening, and prebiotics, of marine neoagaro-oligosaccharides (NAOS) with 4-12 degrees of polymerization (DPs) have been proven. However, NAOS produced by most marine ß-agarases always possess low DPs (≤6) and limited categories; thus, a strategy that can efficiently produce NAOS especially with various DPs ≥8 must be developed. In this study, 60 amino acid residues with no functional annotation result were removed from the C-terminal of agarase AgaM1, and truncated recombinant AgaM1 (trAgaM1) was found to have the ability to produce NAOS with various DPs (4-12) under certain conditions. The catalytic efficiency and stability of trAgaM1 were obviously lower than the wild type (rAgaM1), which probably endowed trAgaM1 with the ability to produce NAOS with various DPs. The optimum conditions for various NAOS production included mixing 1% agarose (w/v) with 10.26 U/ml trAgaM1 and incubating the mixture at 50°C in deionized water for 100 min. This strategy produced neoagarotetraose (NA4), neoagarohexaose (NA6), neoagarooctaose (NA8), neoagarodecaose (NA10), and neoagarododecaose (NA12) at final concentrations of 0.15, 1.53, 1.53, 3.02, and 3.02 g/L, respectively. The NAOS served as end-products of the reaction. The conditions for trAgaM1 expression in a shake flask and 5 L fermentation tank were optimized, and the yields of trAgaM1 increased by 56- and 842-fold compared with those before optimization, respectively. This study provides numerous substrate sources for production and activity tests of NAOS with high DPs and offers a foundation for large-scale production of NAOS with various DPs at a low cost.

Effects of Supplement of Marichromatium gracile YL28 on Water Quality and Microbial Structures in Shrimp Mariculture Ecosystems.

The elevated NH3-N and NO2-N pollution problems in mariculture have raised concerns because they pose threats to animal health and coastal and offshore environments. Supplement of Marichromatium gracile YL28 (YL28) into polluted shrimp rearing water and sediment significantly decreased ammonia and nitrite concentrations, showing that YL28 functioned as a novel safe marine probiotic in the shrimp culture industry. The diversity of aquatic bacteria in the shrimp mariculture ecosystems was studied by sequencing the V4 region of 16S rRNA genes, with respect to additions of YL28 at the low and high concentrations. It was revealed by 16S rRNA sequencing analysis that Proteobacteria, Planctomycete and Bacteroidetes dominated the community (>80% of operational taxonomic units (OTUs)). Up to 41.6% of the predominant bacterial members were placed in the classes Gammaproteobacteria (14%), Deltaproteobacteria (14%), Planctomycetacia (8%) and Alphaproteobacteria (5.6%) while 40% of OTUs belonged to unclassified ones or others, indicating that the considerable bacterial populations were novel in our shrimp mariculture. Bacterial communities were similar between YL28 supplements and control groups (without addition of YL28) revealed by the ß-diversity using PCoA, demonstrating that the additions of YL28 did not disturb the microbiota in shrimp mariculture ecosystems. Instead, the addition of YL28 increased the relative abundance of ammonia-oxidizing and denitrifying bacteria. The quantitative PCR analysis further showed that key genes including nifH and amoA involved in nitrification and nitrate or nitrite reduction significantly increased with YL28 supplementation (p < 0.05). The supplement of YL28 decreased the relative abundance of potential pathogen Vibrio. Together, our studies showed that supplement of YL28 improved the water quality by increasing the relative abundance of ammonia-oxidizing and denitrifying bacteria while the microbial community structure persisted in shrimp mariculture ecosystems.

One-Step Process for Environment-Friendly Preparation of Agar Oligosaccharides From Gracilaria lemaneiformis by the Action of Flammeovirga sp. OC4.

Oligosaccharides extracted from agar Gracilaria lemaneiformis (G. lemaneiformis) have stronger physiological activities and a higher value than agar itself, but the pollution caused by the extraction process greatly restricts the sustainable use of agar. In this study, four bacterial strains with a high ability to degrade G. lemaneiformis were isolated from seawater by in situ enrichment in the deep sea. Among them, Flammeovirga sp. OC4, identified by morphological observation and its 16S rRNA sequencing (98.07% similarity to type strain JL-4 of Flammeovirga aprica), was selected. The optimum temperature and pH of crude enzyme produced by Flammeovirga sp. OC4 were 50°C and 8, respectively. More than 60% of the maximum enzyme activity remained after storage at pH 5.0-10.0 for 60 min. Both Mn2+ and Ba2+ could enhance the enzyme activity. A "one-step process" for preparation of oligosaccharides from G. lemaneiformis was established using Flammeovirga sp. OC4. After optimization of the Plackett-Burman (PB) design and response surface methodology (RSM), the yield of oligosaccharides was increased by 36.1% from 2.71 to 3.09 g L-1 in a 250-mL fermenter with optimized parameters: 30 g L-1 G. lemaneiformis powder, 4.84 g L-1 (NH4)2SO4, 44.8-mL working medium volume at 36.7°C, and a shaking speed of 200 × g for 42 h. The extracted oligosaccharides were identified by thin layer chromatography (TLC) and ion chromatography, which consisted of neoagarobiose, agarotriose, neoagarotetraose, agaropentaose, and neoagarohexaose. These results provided an alternative approach for environment-friendly and sustainable utilization of algae.

Discovery, gene modification, and optimization of fermentation of an enduracidin-producing strain.

Enduracidin significantly inhibits Gram-positive bacteria and had been widely used in many fields. However, as the poor technology for production of enduracidin and its scarcity, identification of novel strategies for production of enduracidin is important. Our group developed two methods to improve the yield of the production of enduracidin. The yield of enduracidin was increased by three- to fivefold. The highest yields of enduracidin A and enduracidin B achieved were 63.7 and 82.13 mg/ml. Thus, our results might provide a new reference method for the industrial production of enduracidin.

Preparation, characterization and alcoholic liver injury protective effects of algal oligosaccharides from Gracilaria lemaneiformis.

Oligosaccharides derived from seaweeds possess diverse biological functions. However, little is known about their effects on liver damage. In this study, algal oligosaccharides (AOS) were prepared from Gracilaria lemaneiformis by biodegradation approaches. HPLC analysis showed AOS were composed of agarooligosaccharides and neoagarooligosaccharides. In vivo animal studies showed AOS significantly attenuated alcohol-induced hepatopathy in mice to some extent, as revealed by the normalization of several serum liver-damage markers. Besides, AOS increased antioxidant levels of hepatic glutathione (GSH) and superoxide dismutase (SOD), and ameliorated the elevated formation of malonaldehyde (MDA), suggesting AOS attenuated hepatopathy mainly through their antioxidant activities. Interestingly, AOS could also enhance the activities of hepatic alcohol dehydrogenase (ADH). Histological examination of liver tissues showed AOS reduced the alcohol-induced liver injury in a dose-dependent manner. Moreover, the comparison of different administration strategies suggested AOS were best taken 2h before alcohol consumption. Therefore, our study provided a novel nutritional strategy for reducing alcohol-induced hepatotoxicity.

Insights into arsenic multi-operons expression and resistance mechanisms in Rhodopseudomonas palustris CGA009.

Arsenic (As) is widespread in the environment and causes numerous health problems. Rhodopseudomonas palustris has been regarded as a good model organism for studying arsenic detoxification since it was first demonstrated to methylate environmental arsenic by conversion to soluble or gaseous methylated species. However, the detailed arsenic resistance mechanisms remain unknown though there are at least three arsenic-resistance operons (ars1, ars2, and ars3) in R. palustris. In this study, we investigated how arsenic multi-operons contributed to arsenic detoxification in R. palustris. The expression of ars2 or ars3 operons increased with increasing environmental arsenite (As(III)) concentrations (up to 1.0 mM) while transcript of ars1 operon was not detected in the middle log-phase (55 h). ars2 operon was actively expressed even at the low concentration of As(III) (0.01 µM), whereas the ars3 operon was expressed at 1.0 µM of As(III), indicating that there was a differential regulation mechanism for the three arsenic operons. Furthermore, ars2 and ars3 operons were maximally transcribed in the early log-phase where ars2 operon was 5.4-fold higher than that of ars3 operon. A low level of ars1 transcript was only detected at 43 h (early log-phase). Arsenic speciation analysis demonstrated that R. palustris could reduce As(V) to As(III). Collectively, strain CGA009 detoxified arsenic by using arsenic reduction and methylating arsenic mechanism, while the latter might occur with the presence of higher concentrations of arsenic.

Draft genome sequence of an agar-degrading marine bacterium Flammeovirga pacifica WPAGA1.

Flammeovirga pacifica WPAGA1(T), which was isolated from sediment of the west Pacific Ocean in 2009 has the ability to produce agar-oligosaccharides from Gracilaria lemaneiformis directly by enzyme-degradation. The draft genome sequence of this strain was sequenced and annotated. Its draft genome contained 6,507,364 bp with a G+C content of 33.8%. Genome sequence information provided a basis for analyzing the digestion of G. lemaneiformis.