Deep-sea Cold Seeps Research Articles

A virulent Bacillus cereus strain from deep-sea cold seep induces pyroptosis in a manner that involves NLRP3 inflammasome, JNK pathway, and lysosomal rupture

ABSTRACT Recent studies indicate that the Bacillus species is distributed in deep-sea environments. However, no specific studies on deep-sea Bacillus cereus have been documented. In the present work, we isolated a B. cereus strain, H2, from the deep-sea cold seep in South China Sea. We characterized the pathogenic potential of H2 and investigated H2-induced death of different types of cells. We found that H2 was capable of tissue dissemination and causing acute mortality in mice and fish following intraperitoneal/intramuscular injection. In vitro studies revealed that H2 infection of macrophages induced pyroptosis and activation of the NLRP3 inflammasome pathway that contributed partly to cell death. H2 infection activated p38, JNK, and ERK, but only JNK proved to participate in H2-triggered cell death. Reactive oxygen species (ROS) and intracellular Ca2+ were essential to H2-induced activation of JNK and NLRP3 inflammasome. In contrast, lysosomal rupture and cathepsins were required for H2-induced NLRP3 inflammasome activation but not for JNK activation. This study revealed for the first time the virulence characteristics of deep-sea B. cereus and provided new insights into the mechanism of B. cereus infection.

The first two complete mitogenomes of the order Apodida from deep-sea chemoautotrophic environments: New insights into the gene rearrangement, origin and evolution of the deep-sea sea cucumbers

The deep-sea ecosystem is considered as the largest and most remote biome of the world. It is meaningful and important to elucidate the life origins by exploring the origin and adaptive genetic mechanisms of the large deep-sea organisms. Sea cucumbers (Holothuroidea) are abundant and economically important group of echinoderms, living from the shallow-waters to deep-sea. In this study, we present the mitochondrial genomes of the sea cucumber Chiridota heheva and Chiridota sp. collected from the deep-sea cold seep and hydrothermal vent, respectively. This is the first reported mitochondrial genomes from the order Apodida. The mitochondrial genomes of C. heheva (17,200 bp) and Chiridota sp. (17,199 bp) display novel gene arrangements with the first protein-coding gene rearrangements in the class Holothuroidea. Bases composition analysis showed that the A + T content of deep-sea holothurians were significantly higher than that of the shallow-water groups. We compared the arrangement of genes from the 24 available holothurian mitogenomes and found that the transposition, reverse transposition and tandem-duplication-random-losses (TDRL) may be involved in the evolution of mitochondrial gene arrangements in Holothuroidea. Phylogenetic analysis revealed that the Apodida clustered with Elasipodida, forming two basal deep-sea holothurian clades. The divergence between the deep-sea and shallow-water holothurians was located at 386.93 Mya, during the Late Devonian. Mitochondrial protein-coding genes of deep-sea holothurians underwent relaxed purifying selection. There are 57 positive selected amino acids sites for some mitochondrial genes of the three deep-sea clades, implying they may involve in the adaption of deep-sea sea cucumbers.

A glimpse of deep-sea adaptation in chemosynthetic holobionts: Depressurization causes DNA fragmentation and cell death of methanotrophic endosymbionts rather than their deep-sea Bathymodiolinae host.

Bathymodiolinae mussels are typical species in deep-sea cold seeps and hydrothermal vents and an ideal model for investigating chemosynthetic symbiosis and the influence of high hydrostatic pressure on deep-sea organisms. Herein, the potential influence of depressurization on DNA fragmentation and cell death in Bathymodiolinae hosts and their methanotrophic symbionts were surveyed using isobaric and unpressurized samples. As a hallmark of cell death, massive DNA fragmentation was observed in methanotrophic symbionts from unpressurized Bathymodiolinae while several endonucleases and restriction enzymes were upregulated. Additionally, genes involved in DNA repair, glucose/methane metabolism as well as two-component regulatory system were also differentially expressed in depressurized symbionts. DNA fragmentation and programmed cell death, however, were rarely detected in the host bacteriocytes owing to the orchestrated upregulation of inhibitor of apoptosis genes and downregulation of caspase genes. Meanwhile, diverse host immune recognition receptors were promoted during depressurization, probably enabling the regain of symbionts. When the holobionts were subjected to a prolonged acclimation at atmospheric pressure, alternations in both the DNA fragmentation and the expression atlas of aforesaid genes were continuously observed in symbionts, demonstrating the persistent influence of depressurization. Contrarily, the host cells demonstrated certain tolerance against depressurization stress as expression level of some immune-related genes returned to the basal level in isobaric samples. Altogether, the present study illustrates the distinct stress responses of Bathymodiolinae hosts and their methanotrophic symbionts against depressurization, which could provide further insight into the deep-sea adaptation of Bathymodiolinae holobionts while highlighting the necessity of using isobaric sampling methods in deep-sea research.

Characterization of Two Unique Cold-Active Lipases Derived from a Novel Deep-Sea Cold Seep Bacterium.

The deep ocean microbiota has unexplored potential to provide enzymes with unique characteristics. In order to obtain cold-active lipases, bacterial strains isolated from the sediment of the deep-sea cold seep were screened, and a novel strain gcc21 exhibited a high lipase catalytic activity, even at the low temperature of 4 °C. The strain gcc21 was identified and proposed to represent a new species of Pseudomonas according to its physiological, biochemical, and genomic characteristics; it was named Pseudomonas marinensis. Two novel encoding genes for cold-active lipases (Lipase 1 and Lipase 2) were identified in the genome of strain gcc21. Genes encoding Lipase 1 and Lipase 2 were respectively cloned and overexpressed in E. coli cells, and corresponding lipases were further purified and characterized. Both Lipase 1 and Lipase 2 showed an optimal catalytic temperature at 4 °C, which is much lower than those of most reported cold-active lipases, but the activity and stability of Lipase 2 were much higher than those of Lipase 1 under different tested pHs and temperatures. In addition, Lipase 2 was more stable than Lipase 1 when treated with different metal ions, detergents, potential inhibitors, and organic solvents. In a combination of mutation and activity assays, catalytic triads of Ser, Asp, and His in Lipase 1 and Lipase 2 were demonstrated to be essential for maintaining enzyme activity. Phylogenetic analysis showed that both Lipase 1 and Lipase 2 belonged to lipase family III. Overall, our results indicate that deep-sea cold seep is a rich source for novel bacterial species that produce potentially unique cold-active enzymes.

EPS364, a Novel Deep-Sea Bacterial Exopolysaccharide, Inhibits Liver Cancer Cell Growth and Adhesion.

The prognosis of liver cancer was inferior among tumors. New medicine treatments are urgently needed. In this study, a novel exopolysaccharide EPS364 was purified from Vibrio alginolyticus 364, which was isolated from a deep-sea cold seep of the South China Sea. Further research showed that EPS364 consisted of mannose, glucosamine, gluconic acid, galactosamine and arabinose with a molar ratio of 5:9:3.4:0.5:0.8. The relative molecular weight of EPS364 was 14.8 kDa. Our results further revealed that EPS364 was a β-linked and phosphorylated polysaccharide. Notably, EPS364 exhibited a significant antitumor activity, with inducing apoptosis, dissipation of the mitochondrial membrane potential (MMP) and generation of reactive oxygen species (ROS) in Huh7.5 liver cancer cells. Proteomic and quantitative real-time PCR analyses indicated that EPS364 inhibited cancer cell growth and adhesion via targeting the FGF19-FGFR4 signaling pathway. These findings suggest that EPS364 is a promising antitumor agent for pharmacotherapy.

Pseudodesulfovibrio cashew sp. Nov., a Novel Deep-Sea Sulfate-Reducing Bacterium, Linking Heavy Metal Resistance and Sulfur Cycle.

Sulfur cycling is primarily driven by sulfate reduction mediated by sulfate-reducing bacteria (SRB) in marine sediments. The dissimilatory sulfate reduction drives the production of enormous quantities of reduced sulfide and thereby the formation of highly insoluble metal sulfides in marine sediments. Here, a novel sulfate-reducing bacterium designated Pseudodesulfovibrio cashew SRB007 was isolated and purified from the deep-sea cold seep and proposed to represent a novel species in the genus of Pseudodesulfovibrio. A detailed description of the phenotypic traits, phylogenetic status and central metabolisms of strain SRB007 allowed the reconstruction of the metabolic potential and lifestyle of a novel member of deep-sea SRB. Notably, P. cashew SRB007 showed a strong ability to resist and remove different heavy metal ions including Co2+, Ni2+, Cd2+ and Hg2+. The dissimilatory sulfate reduction was demonstrated to contribute to the prominent removal capability of P. cashew SRB007 against different heavy metals via the formation of insoluble metal sulfides.

Phenylhydrazone and Quinazoline Derivatives from the Cold-Seep-Derived Fungus Penicillium oxalicum

Three new phenylhydrazones, penoxahydrazones A–C (compounds 1–3), and two new quinazolines, penoxazolones A (compound 4) and B (compound 5), with unique linkages were isolated from the fungus Penicillium oxalicum obtained from the deep sea cold seep. Their structures and relative configurations were assigned by analysis of 1D/2D NMR and mass spectroscopic data, and the absolute configurations of 1, 4, and 5 were established on the basis of X-ray crystallography or ECD calculations. Compound 1 represents the first natural phenylhydrazone-bearing steroid, while compounds 2 and 3 are rarely occurring phenylhydrazone tautomers. Compounds 4 and 5 are enantiomers that feature quinazoline and cinnamic acid units. Some isolates exhibited inhibition of several marine phytoplankton species and marine-derived bacteria.

Artificial construction of the biocoenosis of deep-sea ecosystem via seeping methane.

Deep-sea ecosystems, such as cold seeps and hydrothermal vents, have high biomass, even though they are located in the benthic zone, where no sunlight is present to provide energy for organism proliferation. Based on the coexistence of the reduced gases and chemoautotrophic microbes, it is inferred that the energy from the reduced gases supports the biocoenosis of deep-sea ecosystems. However, there is no direct evidence to support this deduction. Here, we developed and placed a biocoenosis generator, a device that continuously seeped methane, on the 1000-m deep-sea floor of the South China Sea to artificially construct a deep-sea ecosystem biocoenosis. The results showed that microorganisms, including bacteria and archaea, appeared in the biocoenosis generator first, followed by jellyfish and Gammaridea arthropods, indicating that a biocoenosis had been successfully constructed in the deep sea. Anaerobic methane-oxidizing archaea, which shared characteristics with the archaea of natural deep-sea cold seeps, acted as the first electron acceptors of the emitted methane; then, the energy in the electrons was transferred to downstream symbiotic archaea and bacteria and finally to animals. Nitrate-reducing bacteria served as partners to complete anaerobic oxidation of methane process. Further analysis revealed that viruses coexisted with these organisms during the origin of the deep-sea biocoenosis. Therefore, our study mimics a natural deep-sea ecosystem and provides the direct evidence to show that the chemical energy of reduced organic molecules, such as methane, supports the biocoenosis of deep-sea ecosystems.

Anaerobic methane oxidation coupled to denitrification is an important potential methane sink in deep-sea cold seeps

Microbes play a crucial role in mediating the methane flux in deep-sea cold seep ecosystems, where only methane-related microbes have been well studied, while the whole microbial community and their ecological functions were still largely unknown. Here, we utilized metagenomic data to investigate how the structure and metabolism of microbial community shift in the reduced sediment habitats along the spatial scales. Microbial communities in cold seeps and troughs formed two distinct clades likely driven by environmental factors, such as total sulfur, total phosphate and NO3−, rather than geographical proximity. The predominance of Methanosarcinales reflected a high potential for methane production. In addition to the already well-reported ANME-1/SRB consortia, prevalence of bacterial Methylomirabilis and archaeal Methanoperedens as important performers in the n-damo process with respective of nitrite and nitrate as respective electron acceptor was observed in deep-sea hydrate-bearing regions as well. Aerobic methane oxidization was conducted mainly by type I methanotrophs at Site F (Formosa Ridge), but also via the n-damo process by Methanoperedens and Methylomirabilis in the Haima seep and Xisha Trough, respectively. Based on the high abundance of those denitrifying-dependent methane oxidizers and their related functional genes, we concluded that the previously overlooked n-damo process might be a major methane sink in cold seeps or in gas hydrate-bearing sediments if nitrate is available in the anoxic zones. The signature of isotopic labeling would be essential to confirm the contribution of different anaerobic methane oxidizing pathways in deep-sea cold seep ecosystems.

Sphingosinithalassobacter tenebrarum sp. nov., isolated from a deep-sea cold seep.

A Gram-stain-negative, facultatively anaerobic, yellow-pigmented, non-motile, rod-shaped bacterium, designated zrk23T, was isolated from a deep-sea cold seep. The strain was characterized by a polyphasic approach to clarify its taxonomic position. Phylogenetic analysis based on 16S rRNA gene sequences placed zrk23T within the genus Sphingosinithalassobacter and showed the highest similarity to Sphingosinithalassobacter portus FM6T (97.93 %). Growth occurs at temperatures from 16 to 45 °C (optimum, 30 °C), at pH values between pH 6.0 and 8.5 (optimum, pH 7.0) and in 0-5.0 % (w/v) NaCl (optimum, 1.5 %). The major fatty acids were C16 : 0, C14 : 0 2-OH and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c). The major isoprenoid quinone was ubiquinone-10. Predominant polar lipids were diphosphatidylglycerol, phosphatidylglycerol, one unidentified phosphoglycolipid, three unidentified glycolipids and three unidentified phospholipids. The G+C content of the genomic DNA was 64.69 %. The average nucleotide identity values between zrk23T and the most closely related available genome, of Sphingosinithalassobacter portus FM6T, was 82.21 %, indicating that zrk23T was clearly distinguished from S. portus. The analysis of genome sequence of zrk23T revealed that there were many genes associated with degradation of aromatic compounds existing in the genome of zrk23T. As a result of the combination of the results of phylogenetic analysis and phenotypic and chemotaxonomic data, zrk23T was considered to represent a novel species of the genus Sphingosinithalassobacter, for which the name Sphingosinithalassobacter tenebrarum sp. nov. is proposed. The type strain is zrk23T (=KCTC 72896T=MCCC 1K04416T).

Metabolic diversification of anaerobic methanotrophic archaea in a deep-sea cold seep

Anaerobic methanotrophic archaea (ANME) can assimilate methane and govern the greenhouse effect of deep-sea cold seeps. In this study, a total of 13 ANME draft genomes representing five ANME types (ANME-1a, ANME-1b, ANME-2a, ANME-2b and ANME-2c), in size between 0.8 and 1.8 Mbp, were obtained from the Jiaolong cold seep in the South China Sea. The small metagenome-assembled genomes (MAGs) contained all the essential pathways for methane oxidization and carbon dioxide fixation. All genes related to nitrate and sulfate reduction were absent from the MAGs, indicating their syntrophic dependence on partner organisms. Aside from acetate secretion and sugar storage, propanoate synthesis pathway, as an alternative novel carbon flow, was identified in all the MAGs and transcriptionally active. Regarding type-specific features of the MAGs, the genes encoding archaellum and bacteria-derived chemotaxis were specific to ANME-2, perhaps for fitness under fluctuation of methane and sulfate concentration flux. Our genomic and transcriptomic results strongly suggested that ANME could carry out simple carbon metabolism from C1 assimilation to C3 biosynthesis in the SCS cold seep, which casts light on a novel approach for synthetic biology.

Phosphorus Species in Deep-Sea Carbonate Deposits: Implications for Phosphorus Cycling in Cold Seep Environments

Phosphorus (P) is an important nutrient for biological communities in cold seeps. However, our knowledge on the source, species, and cycling of P in cold seep environments is limited. In this study, the concentration, species, and micro to nanometer scale distribution of P in seep carbonates were examined at three deep-sea cold seeps in the South China Sea and East China Sea. The Ca-P accounts for the largest proportion of P—followed by detrital-P, Fe-P, organic-P, and exchangeable-P. The distribution patterns of Ca-P, detrital-P, and organic-P in the seep carbonates differ from one another, as shown by elemental mapping with NanoSIMS and scanning electron microscopy. The covariation of P with Ca and C reveals that Ca-P co-precipitates with Ca-carbonate, which is linked to the process of sulfate-driven anaerobic oxidation of methane. Organic-P is also observed within biofilm-like organic carbon aggregates, revealing the microbial enrichment of P by fluids in the process of anaerobic oxidation of methane. P with a granulated morphology was identified as detrital-P derived from deep sediments. Most importantly, it is evident that Ca-P is positively correlated to the Fe content in all the seep carbonates. This indicates the likelihood that the dissolved P in cold-seep fluids is released primarily from Fe oxides through Fe-driven anaerobic oxidation of methane in deep sediments. These processes associated with different species of P may have significant implications for P geochemical cycling and anaerobic oxidation of methane impelled by Fe and sulfate reduction in cold seep environments.

A novel bacterial thiosulfate oxidation pathway provides a new clue about the formation of zero-valent sulfur in deep sea

Zero-valent sulfur (ZVS) has been shown to be a major sulfur intermediate in the deep-sea cold seep of the South China Sea based on our previous work, however, the microbial contribution to the formation of ZVS in cold seep has remained unclear. Here, we describe a novel thiosulfate oxidation pathway discovered in the deep-sea cold seep bacterium Erythrobacter flavus 21–3, which provides a new clue about the formation of ZVS. Electronic microscopy, energy-dispersive, and Raman spectra were used to confirm that E. flavus 21–3 effectively converts thiosulfate to ZVS. We next used a combined proteomic and genetic method to identify thiosulfate dehydrogenase (TsdA) and thiosulfohydrolase (SoxB) playing key roles in the conversion of thiosulfate to ZVS. Stoichiometric results of different sulfur intermediates further clarify the function of TsdA in converting thiosulfate to tetrathionate (−O3S–S–S–SO3−), SoxB in liberating sulfone from tetrathionate to form ZVS and sulfur dioxygenases (SdoA/SdoB) in oxidizing ZVS to sulfite under some conditions. Notably, homologs of TsdA, SoxB, and SdoA/SdoB widely exist across the bacteria including in Erythrobacter species derived from different environments. This strongly indicates that this novel thiosulfate oxidation pathway might be frequently used by microbes and plays an important role in the biogeochemical sulfur cycle in nature.

Eyes of differing colors in Alvinocaris longirostris from deep-sea chemosynthetic ecosystems: genetic and molecular evidence of its formation mechanism

Coloration is an important phenotypic trait for multiple adaptive functions. It is interesting to find white-eye (AW) and orange-eye (AO) phenotypes in the shrimp Alvinocaris longirostris inhabiting the deep-sea cold seep and hydrothermal vent areas of the northwestern Pacific. By comparative transcriptome analyses, 1 491 differentially expressed genes (DEGs) were identified between AW and AO. Among them, many DEGs were associated with immunity, antioxidation, and detoxification. Two significant enzyme encoding genes, xanthine dehydrogenase, and tryptophan oxidase involved in pigment biosynthesis pathways were up-regulated in AW and AO, respectively, which might be related to the differences of white and orange eye phenotypes. Moreover, single nucleotide polymorphism (SNP) calling detected that genotypes of 28 SNP distributing in 14 unigenes were completely different between AW and AO. Particularly, there were three and two non-synonymous mutations in immune genes crustin Pm5 and antimicrobial peptide, respectively. Results indicate that the difference in eye color is probably resulted from immune response to variable micro-environmental stressors encountered in the dispersal process of the shrimps, such as symbiotic microbes, pathogens, and toxic substances, and might be genetically fixed at last. The suggested pathway preliminarily explained the formation mechanism of different eye phenotypes in Alvinocaridid shrimps, providing a basis for further study on adaptive evolution of eyes in deep-sea chemosynthetic faunas.

High-Throughput Sequencing Reveals a Potentially Novel Sulfurovum Species Dominating the Microbial Communities of the Seawater-Sediment Interface of a Deep-Sea Cold Seep in South China Sea.

In the Formosa cold seep of the South China Sea (SCS), large amounts of methane and sulfide hydrogen are released from the subseafloor. In this study, we systematically investigated the microbial communities in the seawater–sediment interface of Formosa cold seep using high-throughput sequencing techniques including amplicon sequencing based on next-generation sequencing and Pacbio amplicon sequencing platforms, and metagenomics. We found that Sulfurovum dominated the microbial communities in the sediment–seawater interface, including the seawater close to the seepage, the surface sediments, and the gills of the dominant animal inhabitant (Shinkaia crosnieri). A nearly complete 16S rRNA gene sequence of the dominant operational taxonomic units (OTUs) was obtained from the Pacbio sequencing platforms and classified as OTU-L1, which belonged to Sulfurovum. This OTU was potentially novel as it shared relatively low similarity percentages (<97%) of the gene sequence with its close phylogenetic species. Further, a draft genome of Sulfurovum was assembled using the binning technique based on metagenomic data. Genome analysis suggested that Sulfurovum sp. in this region may fix carbon by the reductive tricarboxylic acid (rTCA) pathway, obtain energy by oxidizing reduced sulfur through sulfur oxidizing (Sox) pathway, and utilize nitrate as electron acceptors. These results demonstrated that Sulfurovum probably plays an important role in the carbon, sulfur, and nitrogen cycles of the Formosa cold seep of the SCS. This study improves our understanding of the diversity, distribution, and function of sulfur-oxidizing bacteria in deep-sea cold seep.

Vertical shifts of particle-attached and free-living prokaryotes in the water column above the cold seeps of the South China Sea

Marine particle-attached (PA) and free-living (FL) microbes play important roles in the biogeochemical cycling of organic matter along the water column. Deep-sea cold seeps are highly productive and chemosynthetic ecosystems, their continuous emission of CH4, CO2, and H2S can reach up to 100 m in the above water, therefore would influence the distribution and potential metabolic functions of deep-sea prokaryotes. In this study, the vertical distribution profiles of both PA and FL microbes in the water column above two cold seeps of the South China Sea were investigated using Illumina sequencing and quantitative PCR (qPCR) based on 16S rRNA gene. Photosynthetic and heterotrophic prokaryotes were predominant in respective surface and deep layers below the photic zone. The typical cold seep chemosynthetic microbes, such as methanotrophs and sulfate-reducing bacteria were observed with low proportions in the two cold seeps as well. Distinct PA and FL microbial fractions were found in terms of abundance and diversity. FL fraction exposed to the bulk water was significantly affected by temperature and inorganic nutrients, whereas PA fraction relied more on the organic matter of the particles and less susceptible to the environmental variability. Our study highlights the importance of vertical geochemical gradients on the distribution and potential metabolic choice of marine microbes and extends our current knowledge of depth-associated microbial distribution patterns.

Acoustic characteristics of cold-seep methane bubble behavior in the water column and its potential environmental impact

The amount of methane leaked from deep sea cold seeps is enormous and potentially affects the global warming, ocean acidification and global carbon cycle. It is of great significance to study the methane bubble movement and dissolution process in the water column and its output to the atmosphere. Methane bubbles produce strong acoustic impedance in water bodies, and bubble strings released from deep sea cold seeps are called “gas flares” which expressed as flame-like strong backscatter in the water column. We characterized the morphology and movement of methane bubbles released into the water using multibeam water column data at two cold seeps. The result shows that methane at site I reached 920 m water depth without passing through the top of the gas hydrate stability zone (GHSZ, 850 m), while methane bubbles at site II passed through the top of the GHSZ (597 m) and entered the non-GHSZ (above 550 m). By applying two methods on the multibeam data, the bubble rising velocity in the water column at sites I and II were estimated to be 9.6 cm/s and 24 cm/s, respectively. Bubble velocity is positively associated with water depth which is inferred to be resulted from decrease of bubble size during methane ascending in the water. Combined with numerical simulation, we concluded that formation of gas hydrate shells plays an important role in helping methane bubbles entering the upper water bodies, while other factors, including water depth, bubble velocity, initial kinetic energy and bubble size, also influence the bubble residence time in the water and the possibility of methane entering the atmosphere. We estimate that methane gas flux at these two sites is 0.4×106−87.6×106 mol/a which is extremely small compared to the total amount of methane in the ocean body, however, methane leakage might exert significant impact on the ocean acidification considering the widespread distributed cold seeps. In addition, although methane entering the atmosphere is not observed, further research is still needed to understand its potential impact on increasing methane concentration in the surface seawater and gas-water interface methane exchange rate, which consequently increase the greenhouse effect.

Characterization of Antifungal Lipopeptide Biosurfactants Produced by Marine Bacterium Bacillus sp. CS30.

This study was initiated to screen for marine bacterial agents to biocontrol Magnaporthe grisea, a serious fungal pathogen of cereal crops. A bacterial strain, isolated from the cold seep in deep sea, exhibited strong growth inhibition against M. grisea, and the strain was identified and designated as Bacillus sp. CS30. The corresponding antifungal agents were purified by acidic precipitation, sequential methanol extraction, Sephadex LH-20 chromatography, and reversed phase high-performance liquid chromatography (RP-HPLC), and two antifungal peaks were obtained at the final purification step. After analysis by mass spectrometry (MS) and tandem MS, two purified antifungal agents were deduced to belong to the surfactin family, and designated as surfactin CS30-1 and surfactin CS30-2. Further investigation showed that although the antifungal activity of surfactin CS30-1 is higher than that of surfactin CS30-2, both of them induced the increased generation of reactive oxygen species (ROS) and caused serious damage to the cell wall and cytoplasm, thus leading to the cell death of M. grisea. Our results also show the differences of the antifungal activity and antifungal mechanism of the different surfactin homologs surfactin CS30-1 and surfactin CS30-2, and highlight them as potential promising agents to biocontrol plant diseases caused by M. grisea.

Complex gas hydrate system in a gas chimney, South China Sea

Abstract Gas hydrates and deep-sea cold seeps are of interest to industry and academia for several reasons, including their implications for offshore geohazards and unconventional resources, and for global climate change and carbon cycling. Guangzhou Marine Geological Survey (GMGS) implemented its fifth gas hydrate drilling expedition in the QiongDongNan Sea Area (QDN-SA) of the northern South China Sea (SCS) from June to September 2018. Multidisciplinary investigations, including Logging While Drilling (LWD), pressure and non-pressure coring, gas composition analyses, in situ temperature and permeability measurements, pore water geochemical analyses, and a regional seismic survey, reveal a complex distribution of gas hydrate within a vertical chimney structure. LWD profiles record high resistivity anomalies and high P-wave velocity variations at 9–174 mbsf, suggesting the coexistence of free gas and gas hydrate in the sediment column throughout the investigated gas chimney. Hydrate-bound and void gas is an admixture of methane (70.1%–98.3%) and heavier hydrocarbons (C2H6: 0.16%–17.59%, C3H8: 0.05%–9.88%, i-C4H10:

Diverging processes of symbiotic bacteria on vent mussels in the eastern Pacific Ocean

Chemoautolithotrophic bacteria thriving in deep-sea hydrothermal vents and cold seeps play an important role as primary producers in these remote, extreme ecosystems, and some of them have evolved symbiotic relationships with diverse marine invertebrate animals. These symbiotic bacteria harness energy through oxidation of reduced gases such as hydrogen sulfide, methane, hydrogen emissions, and eventually, their organic materials are transferred to their host invertebrates. Using symbiont-specific, multi-loci Next Generation Sequencing (NGS) data, geographical population genetics for symbiotic bacteria of Bathymodiolinae vent mussels were examined, with samples from nine vent localities. This is one of the most dominant and widespread taxa in global chemosynthetic environments on the sea floor, along the East Pacific Rise, Galapagos Rift, and the Pacific-Antarctic Ridge. The hope of this study is to gain insights into the evolution of bacteria in space and time. Particularly, the symbiotic bacteria of these Bathymodiolinae species have free-living and intracellular symbiotic stages, which allowed examination of their population dynamics and evolution in benthic environments. Per the results, it appears that two sister parapatric Bathymodiolinae species harbor the same symbiont species, thiotrophic Gamma-proteobacteria, regarding 16S rRNA encoding gene, but completely divergent lineages living in the north and south of the Easter Microplate regarding six protein-coding genes. Such divergence of bacteria seems to be initiated by orogeny of the Easter Microplate dated 2.5–5.3 million years ago, and equivalent to synonymous substitution rates of 0.77–1.62% per nucleotide per million years of protein-coding genes.