Marine Isolates Research Articles

Insights into the planktonic to sessile transition in a marine biofilm-forming Pseudoalteromonas isolate using comparative proteomic analysis

Bacterial biofilms play an important role in marine biofouling. The formation of a biofilm starts when marine bacterial cells transition from a planktonic to an attached state. However, the molecular mechanisms involved in this transition are poorly understood. Here, 51 strains of marine bacteria were isolated from natural biofilms growing on submerged artificial surfaces (glass slides, epoxy panels, and bridge pillars) and evaluated for their biofilm-forming capacity. Eleven strains formed relatively strong biofilms and 16S rRNA gene sequence analysis indicated that they belonged to the genera Leisingera, Roseobacter, Pseudoalteromonas, Alteromonas, Tenacibaculum, Vibrio, Chryseobacterium, Aquimarina, and Acinetobacter. Strain Pseudoalteromonas sp. W-7 showed efficient and rapid attachment and was therefore chosen for further study. An iTRAQ-based comparative proteomic analysis of planktonic and attached strain W-7 cells was carried out. A total of 3468 proteins were identified, of which 163 showed significant differential expression (120 down-regulated and 43 up-regulated in attached cells relative to planktonic cells). KEGG (Kyoto encyclopedia of genes and genomes) analysis indicated that pyruvate metabolism, carbon fixation, and carbon metabolism were significantly affected in attached cells. Up-regulated proteins such as UTP-glucose-1-phosphate uridylyltransferase, acetyltransferase component of pyruvate dehydrogenase complex, OmpA-like protein, and acetyl-coenzyme A synthetase may be important during initial adhesion. Our findings provide a deeper understanding of the planktonic to sessile transition of marine fouling bacteria.

Biofilms of Pseudomonas and Lysinibacillus Marine Strains on High-Density Polyethylene.

Environmental pollution by plastic debris is estimated on a scale of 100 million metric tons, a portion of which is fragmented into micro- and nanoplastics. These fragments are often colonized by bacterial species in marine environments, possibly contributing to the biodegradation of such materials. However, further investigations are necessary to determine the impact of abiotic polymer weathering on biofilm adhesion, as well as the specific biofilm formation strategies employed by marine isolates. Here, we evaluate deep-sea sediment bacterial isolates for biofilm adhesion, extracellular matrix production, and polymer degradation ability. Our study focuses on high-density polyethylene (HDPE) fragments for their high durability and environmental persistence, subjecting fragments to abiotic weathering prior to bacterial colonization. Marine isolates identified as Pseudomonas sp. and Lysinibacillus sp. exhibited decreasing biofilm formation on weathered HDPE, especially over the first 24h of incubation. This effect was countered by increased extracellular matrix production, likely improving cell adhesion to surfaces roughened by abiotic degradation. These adhesion strategies were contrasted with a reference Pseudomonas aeruginosa strain, which displayed high levels of biofilm formation on non-weathered HDPE and lower extracellular matrix production over the first 24h of incubation. Furthermore, our results suggest that an increase in biofilm biomass correlated with changes to HDPE structure, indicating that these strains have a potential for biodegradation of plastic fragments. Therefore, this work provides a detailed account of biofilm formation strategies and bacteria-plastic interactions that represent crucial steps in the biodegradation of plastic fragments in marine environments.

SCREENING OF BACILLUS SP. ISOLATED FROM CORAL SIDERASTREA STELLATA FOR ANTIMICROBIAL ACTIVITY AGAINST ENTEROPATHOGENIC STRAINS OF SALMONELLA AND ESCHERICHIA COLI / SCREENING DE BACILLUS SP. ISOLADOS DO CORAL SIDERASTREA STELLATA PARA ATIVIDADE ANTIMICROBIANA CONTRA LINHAGENS ENTEROPATOGÊNICAS DE SALMONELLA E ESCHERICHIA COLI

Siderastrea stellata is a coral endemic to Brazilian waters and is widely distributed throughout the coast of Bahia state. Exposure of the coral to pathogens stimulates the production of important enzymes by associated symbiont microorganisms. Besides being natural sources of bioactive compounds, these microorganisms possess characteristics that help them survive under extreme conditions. Sixty-eight bacterial strains isolated from the S. stellata coral were analyzed for antimicrobial activity against ten pathogenic bacteria. The cultivation of marine isolates was carried out in liquid or solid seawater. Eight isolates produced antimicrobial compounds against at least two of the ten pathogenic bacteria tested. When isolates were grown in Mueller-Hinton broth, a distinct biocidal spectrum was observed, indicating that the culture medium directly influences the production of antimicrobial compounds. Through molecular characterization of activity-positive isolates, they were identified as belonging to the genus Bacillus . The bioactive compounds produced by the Bacillus stratosphericus SS85 and SS69 strains were stable after storage time for three months. This is the first paper reporting on the antimicrobial activity of microorganisms isolated from coral Siderastrea stellata .

Exopolysaccharide production from isolated Enterobacter sp. strain ACD2 from the northwest of Saudi Arabia

Substantial interest has emerged on the use of marine microorganisms as producers of important macromolecules such as exopolysaccharides (EPSs), to apply them in areas including the food, pharmaceutical, and cosmetic industries, as well as other important sectors. In this study, thirty seven marine bacterial isolates were isolated from Haqel Beach in Tabuk region, Saudi Arabia. We found that only nine isolates have the ability to produce extracellular polysaccharides in medium. We selected the best producer and characterized it morphologically, biochemically, and molecularly using 16S rRNA analysis, revealing it to be Enterobacter sp. We also optimized the culture fermentation conditions to obtain the highest EPS yield. Our results showed that the optimal medium conditions were as follows: 15% sucrose, 0.5% peptone, presence of all fermentation medium salts, pH 7.5–8.0, incubation temperature 37 °C, and shaking at 150 rpm for 72 h, which conferred the highest EPS yield of 8.6 g/l.

Genetic classification of two Vibrio species isolated from Florida Gulf Coast using multi-locus sequence analysis

Prokaryotic species classification has lagged dramatically behind eukaryotic species due to the dependence on the difficult DNA-DNA hybridization technique required for species designation. This is disadvantageous for the general understanding of the diversity of microbial life and unique microorganisms, along with their metabolic abilities, like bioremediation of hazardous waste. This applies to the secondary metabolites microorganisms produce that can be beneficial for humans and other living organisms, such as antibiotics or other industrially important compounds. In an effort to isolate and identify new microbial species, environmental samples were collected from marine sediment environments as marine habitats are under-sampled compared to non-marine habitats. Two marine isolates designated MI-1 and MI-2 were isolated from marine sediment environments off the Gulf Coast of Florida and chosen for further study because of their antibacterial production and unique colony pigmentation. They were genetically analyzed through sequencing of the 16s rRNA gene, shotgun cloning, and an eight-gene multi-locus sequence analysis comparison to 66 other species of the same genus. From these, MI-1 and MI-2 can be classified as members of the Vibrio genus most closely related to Vibrio ruber and a distinct strain from V. ruber type strain VR-1.

The Viral Hemorrhagic Septicemia Virus (VHSV) Markers of Virulence in Rainbow Trout (Oncorhynchus mykiss).

Viral hemorrhagic septicemia virus (VHSV) is a highly contagious virus leading to high mortality in a large panel of freshwater and marine fish species. VHSV isolates originating from marine fish show low pathogenicity in rainbow trout. The analysis of several nearly complete genome sequences from marine and freshwater isolates displaying varying levels of virulence in rainbow trout suggested that only a limited number of amino acid residues might be involved in regulating the level of virulence. Based on a recent analysis of 55 VHSV strains, which were entirely sequenced and phenotyped in vivo in rainbow trout, several amino acid changes putatively involved in virulence were identified. In the present study, these amino acid changes were introduced, alone or in combination, in a highly-virulent VHSV 23–75 genome backbone by reverse genetics. A total of 35 recombinant VHSV variants were recovered and characterized for virulence in trout by bath immersion. Results confirmed the important role of the NV protein (R116S) and highlighted a major contribution of the nucleoprotein N (K46G and A241E) in regulating virulence. Single amino acid changes in these two proteins drastically affect virus pathogenicity in rainbow trout. This is particularly intriguing for the N variant (K46G) which is unable to establish an active infection in the fins of infected trout, the main portal of entry of VHSV in this species, allowing further spread in its host. In addition, salmonid cell lines were selected to assess the kinetics of replication and cytopathic effect of recombinant VHSV and discriminate virulent and avirulent variants. In conclusion, three major virulence markers were identified in the NV and N proteins. These markers explain almost all phenotypes (92.7%) observed in trout for the 55 VHSV strains analyzed in the present study and herein used for the backward validation of virulence markers. The identification of VHSV specific virulence markers in this species is of importance both to predict the in vivo phenotype of viral isolates with targeted diagnostic tests and to improve prophylactic methods such as the development of safer live-attenuated vaccines.

Oceanipulchritudo coccoides gen. nov., sp. nov., isolated from marine sediment within the family Puniceicoccaceae.

A Gram-stain-negative, aerobic coccus, designated CK1056T, was isolated from coastal sediment of Xiaoshi Island, Weihai, PR China. Strain CK1056T was found to grow at 15-37 °C (optimum, 30 °C), with 0.5-6.5 % (w/v) NaCl (optimum, 3.5 %) and displayed alkaliphilic growth within the pH range of pH 6.5-10.0 (optimum, pH 8.0). The major fatty acids identified were iso-C15 : 0 and summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c). The main polar lipids consisted of aminophosphoglycolipid and phosphatidylethanolamine. The predominant respiratory quinone was MK-7. The G+C content of the genomic DNA was 54.0 mol%. The result of the 16S rRNA gene sequence analysis confirmed the affiliation of this micro-organism to the family Puniceicoccaceae, with Coraliomargarita akajimensis KCTC 12865T as its closest relative with only 88.0 % sequence similarity. From the taxonomic data obtained in this study, we propose that the new marine isolate be placed into a novel species within a novel genus in the family Puniceicoccaceae, phylum Verrucomicrobia, for which the name Oceanipulchritudo coccoides gen. nov., sp. nov. is proposed. The type strain is CK1056T (=KCTC 72798T=MCCC 1H00425T).

Polynuclear aromatic anthracene biodegradation by psychrophilic Sphingomonas sp., cultivated with tween-80

Anthracene is a low molecular weight polynuclear aromatic hydrocarbons (PAHs) being identified as a precedence toxic contaminant in the ecosystem. Thus, the present work was designed to evaluate anthracene biodegradation efficiency by selected marine bacteria. From the marine isolates, the most effective anthracene biodegrading strain was identified as Sphingomonas sp., KSU05. Time course batch growth results indicated that the isolate KSU05 was capable of surviving up to 500 mg/L of anthracene. The influence of various nutrient sources were screened for enhanced growth and pyrene degradation, based on results glucose and tween-80 were used for further optimization studies. Batch experimental analysis showed maximum biodegradation (70.5%) of anthracene (50 mg/L) with enhanced survival of Sphingomonas sp. KSU05 was observed at 96 h of cultivation. Box-Behnken design optimization results showed that the culture conditions enhanced the anthracene biodegradation (90.0%) at pH 7.0, 0.3 mM of tween-80 concentration, and 5.5% of glucose concentration. In addition, the isolate Sphingomonas sp. KSU05 was found to rapidly degrade anthracene within 96 h. The anthracene intermediates was analyzed using Gas chromatography mass spectrophotometer (GC-MS). Overall, this research shown that the Sphingomonas sp., cultivated with suggested optimum conditions could provide an effective prospective for the degradation of anthracene from contaminated environment.

Diversity and distribution of marine heterotrophic bacteria from a large culture collection

BackgroundIsolation of marine microorganisms is fundamental to gather information about their physiology, ecology and genomic content. To date, most of the bacterial isolation efforts have focused on the photic ocean leaving the deep ocean less explored. We have created a marine culture collection of heterotrophic bacteria (MARINHET) using a standard marine medium comprising a total of 1561 bacterial strains, and covering a variety of oceanographic regions from different seasons and years, from 2009 to 2015. Specifically, our marine collection contains isolates from both photic (817) and aphotic layers (744), including the mesopelagic (362) and the bathypelagic (382), from the North Western Mediterranean Sea, the North and South Atlantic Ocean, the Indian, the Pacific, and the Arctic Oceans. We described the taxonomy, the phylogenetic diversity and the biogeography of a fraction of the marine culturable microorganisms to enhance our knowledge about which heterotrophic marine isolates are recurrently retrieved across oceans and along different depths.ResultsThe partial sequencing of the 16S rRNA gene of all isolates revealed that they mainly affiliate with the classes Alphaproteobacteria (35.9%), Gammaproteobacteria (38.6%), and phylum Bacteroidetes (16.5%). In addition, Alteromonas and Erythrobacter genera were found the most common heterotrophic bacteria in the ocean growing in solid agar medium. When comparing all photic, mesopelagic, and bathypelagic isolates sequences retrieved from different stations, 37% of them were 100% identical. This percentage increased up to 59% when mesopelagic and bathypelagic strains were grouped as the aphotic dataset and compared to the photic dataset of isolates, indicating the ubiquity of some bacterial isolates along different ocean depths. Finally, we isolated three strains that represent a new species, and the genome comparison and phenotypic characterization of two of these strains (ISS653 and ISS1889) concluded that they belong to a new species within the genus Mesonia.ConclusionsOverall, this study highlights the relevance of culture-dependent studies, with focus on marine isolated bacteria from different oceanographic regions and depths, to provide a more comprehensive view of the culturable marine bacteria as part of the total marine microbial diversity.

A Novel Ethyl 3(R)-Acetamido-3-(4-Hydroxyphenyl)Propanoate from the Marine Isolate of the Fungus Penicillium thomii KMM 4680

A Novel Ethyl 3(R)-Acetamido-3-(4-Hydroxyphenyl)Propanoate from the Marine Isolate of the Fungus Penicillium thomii KMM 4680

Bacteria associated with marine macroorganisms as potential source of quorum-sensing antagonists.

Samples were collected from different undisturbed areas along the coast of Gujarat like Okha, Diu, Veraval, and Somnath. A total of 68 marine isolates were obtained out of which 53 were associated with various marine macroorganisms like sponges, gastropods, and algae, whereas 15 were free living. Quorum-quenching ability of all the isolates was tested against Chromobacterium violaceum MK by co-culture technique as a way to simultaneously detect signal-degrading as well as nondegrading quorum-sensing inhibitors. Nineteen macroorganism-associated bacteria and eight free-living bacteria were found to possess quorum-sensing inhibitory activity against C. violaceum MK without affecting its growth. Isolate OA22 from grape alga and OA10 from purple sponge (Haliclona sp.) were found to possess the highest C6-HSL degradation activity and extracellular non-N-acyl-homoserine lactone degrading QSI activity, respectively. OA22 was also found to degrade 3-oxo-C12 homoserine lactone. Acid recovery of both the C6- and C12-HSL after degradation by OA22 indicated the presence of lactonase enzyme in the isolate. Cell-free supernatant of OA10 was extracted with ethyl acetate to obtain the quorum-quenching compound. Pigment inhibition in C. violaceum MK treated with OA10 extract was demonstrated in various ways and was indicative of QSI activity of the extract without degradation of the quorum-sensing signaling molecule. The isolates OA22 and OA10 were identified as Desemzia incerta and Bacillus sp., respectively, by 16S ribosomal DNA sequence analysis.

A novel fibrinolytic enzyme from marine Pseudomonas aeruginosa KU1 and its rapid in vivo thrombolysis with little haemolysis

A direct acting, extracellular, fibrinolytic enzyme, ~50 KDa from marine Pseudomonas aeruginosa KU1 (PEKU1), was purified. It was found to be a metalloprotease. 60% of the total activity of the purified PEKU1 was retained at 70 °C and the enzyme was practically denatured at 80 °C, 2 h. Metal ions, such as Na+, K+ and Co2+, were found to enhance slightly the fibrinolytic activity, while Fe2+, Mn2+ and Zn2+ were found to be inhibiting. The enzyme showed only less than 5% haemolysis, suggesting its thrombolytic administration safe. Tryptic digestion revealed its similarity to serralysin like alkaline protease of P. aeruginosa. In silico studies showed its binding of protease substrates and fibrin D-dimer in its active site. High affinity binding of bradykinin to the active site of PEKU1, confirmed by in vitro cleaving, suggested its future use as an analgesic. The purified enzyme with Na+, K+ and Co2+, and without Fe2+, Mn2+ and Zn2+ showed thrombolysis in vivo in carrageenan induced murine tail thrombolytic model. The enzyme PEKU1, a novel protease from marine isolate Pseudomonas aeruginosa KU1 has great potential to be developed as a therapeutic agent to combat cardiovascular diseases, as well as analgesic and anti-inflammatory drug in appropriate sites.

Pyrone-derived Marine Natural Products: A Review on Isolation, Bio-activities and Synthesis

Marine natural products (MNPs) containing pyrone rings have been isolated from numerous marine organisms, and also produced by marine fungi and bacteria, particularly, actinomycetes. They constitute a versatile structure unit of bioactive natural products that exhibit various biological activities such as antibiotic, antifungal, cytotoxic, neurotoxic, phytotoxic and anti-tyrosinase. The two structure isomers of pyrone ring are γ- pyrone and α-pyrone. In terms of chemical motif, γ-pyrone is the vinologous form of α- pyrone which possesses a lactone ring. Actinomycete bacteria are responsible for the production of several α-pyrone compounds such as elijopyrones A-D, salinipyrones and violapyrones etc. to name a few. A class of pyrone metabolites, polypropionates which have fascinating carbon skeleton, is primarily produced by marine molluscs. Interestingly, some of the pyrone polytketides which are found in cone snails are actually synthesized by actinomycete bacteria. Several pyrone derivatives have been obtained from marine fungi such as Aspergillums flavus, Altenaria sp., etc. The γ-pyrone derivative namely, kojic acid obtained from Aspergillus fungus has high commercial demand and finds various applications. Kojic acid and its derivative displayed inhibition of tyrosinase activity and, it is also extensively used as a ligand in coordination chemistry. Owing to their commercial and biological significance, the synthesis of pyrone containing compounds has been given attention over the past years. Few reviews on the total synthesis of pyrone containing natural products namely, polypropionate metabolites have been reported. However, these reviews skipped other marine pyrone metabolites and also omitted discussion on isolation and detailed biological activities. This review presents a brief account of the isolation of marine metabolites containing a pyrone ring and their reported bio-activities. Further, the review covers the synthesis of marine pyrone metabolites such as cyercene-A, placidenes, onchitriol-I, onchitriol-II, crispatene, photodeoxytrichidione, (-) membrenone-C, lihualide-B, macrocyclic enol ethers and auripyrones-A & B.

Anthracycline Shunt Metabolites From Philippine Marine Sediment-Derived Streptomyces Destroy Cell Membrane Integrity of Multidrug-Resistant Staphylococcus aureus

The rise of antibiotic resistance (ABR) and the drying up of the pipeline for the development of new antibiotics demands an urgent search for new antibiotic leads. While the majority of clinically available antibiotics were discovered from terrestrial Streptomyces, related species from marine sediments as a source of antibiotics remain underexplored. Here, we utilized culture-dependent isolation of thirty-five marine sediment-derived actinobacterial isolates followed by a screening of their antibacterial activity against multidrug-resistant S. aureus ATCC BAA-44. Our results revealed that the crude extract of Streptomyces griseorubens strain DSD069 isolated from marine sediments collected in Romblon, Philippines displays the highest antibacterial activity, with 96.4% growth inhibition. The S. aureus ATCC BAA-44 cells treated with crude extract of Streptomyces griseorubens strain DSD069 showed cell membrane damage as demonstrated by (a) leakage and loss of vital cell constituents, including DNA and proteins, (b) irregular shrinkage of cells, and (c) increase membrane permeability. The antibiotic compounds were identified as Bisanhydroaklavinone and 1-Hydroxybisanhydroaklavinone with MIC value of 6.25 μg/mL and 50.00 μg/mL, respectively. Bisanhydroaklavinone and 1-Hydroxybisanhydroaklavinone are shunt metabolites in the biosynthesis of anticancer anthracycline derivatives namely doxorubicin, daunorubicin, and cinerubins. It is rare, however, that shunt metabolites are accumulated during fermentation of marine sediment-derived Streptomyces strain without genetic modification. Thus, our study provides evidence that natural bacterial strain can produce Bisanhydroaklavinone and 1-Hydroxybisanhydroaklavinone as antibiotic leads to combat ABR.

Marine pharmacology: an ocean to explore novel drugs

The rise in burden of diseases and treatment failure demands discovery of novel compounds. Exploiting natural sources which include territory of land and water in potential manner paves the way for significant future innovations in drug discovery. Marine environment has striking functionalities in its skeleton that has fascinated scientists to show enormous interest in investigation of new compounds. Marine sponges, algae, tunicates, sea whip etc. from the marine pipeline are the important sources for biological active compounds. Recent technology advancements further added to the domain of drug research in isolation and evaluation of marine derived products. To date, significant number of compounds have been isolated. Wide range of antibacterial, anti-inflammatory, antiparasitic, neuroprotective, antiviral, anticancer, analgesic, antimicrobial, antimalarial compounds have been pursued in control and management of diseases. These represent marine ecosystem as a hopeful resource in discovery of novel compounds with ideal starting point in scaffolding additional screening of natural marine products. This is review of abstracting on history, lead development process in identifying and comprehending basic nature of compounds that is promising initial step towards unique pharmacological design, the triumph of approved and ongoing trails, brief depiction on current status and challenges being faced in marine drug discovery field.

Morphotypes and molecular characterisation of pink pigmented bacterial symbiont of Turbinaria sp.

Turbinaria sp. is one of the species of the brown algae class that has extraordinary prospects for human health. The potential of the brown algae makes it a target for the search for sources of beneficial bioactive compounds in the pharmaceutical world. However, a serious problem in the development of bioactive compounds from these biotas is the problem of supply, because to obtain a relatively small number of active compounds requires a large number of marine organisms. It was reported that there are associations of microorganisms with brown algae which are thought to synthesize secondary metabolites such as the host organism. The presence of bacteria that is symbiotic with brown algae has made it possible to use these organisms as the main source of bioactive compounds from the marine. KRT-7 isolate is a pink bacterial isolate from the Turbinaria sp. collected from Menjangan Kecil, Karimunjawa Island, Indonesia. This isolate has been known to produce bio pigment and genes coding for its bioactive compounds. However, the physiological morpho character and its molecular identity are unknown. The study aims to determine the morphophysiological character and molecular identification of the prospective isolates. The research results obtained will contribute a scientific finding of the prospect of genetic diversity of Indonesian marine bacterial isolates and their corridor utilization as a source of bioactive compounds.

In vitro anti-biofilm activity of bacteriocin from a marine Bacillus sp. strain Sh10 against Proteus mirabilis

Biofilm formed by Proteus mirabilis strains is one of the most important medical problems especially in the case of device-related urinary tract infections. This study was conducted to evaluate the bacteriocin produced by a marine isolate of Bacillus sp. Sh10, for it's in vitro inhibitory activity against pre-formed biofilm and in interference with the biofilm-forming of two biofilm-producing bacteria (P. mirabilis UCa4 and P. mirabilis UCe1). Sensitivity of two biofilm-producing bacteria (P. mirabilis UCa4 and P. mirabilis UCe1) to bacteriocin, was investigated in planktonic and biofilm states by cell viability and crystal violet assay, respectively. Scanning electron microscopy (SEM) was also performed to determine the effect of bacteriocin on the morphology of the cells associated with biofilm. It was found that bacteriocin possessed bactericidal activity to biofilm-forming isolates in the planktonic state. However, bacteriocin interferes with the formation of biofilms and disrupts established biofilms. Bacteriocin reduced biofilm formation in the isolates of P. mirabilis UCa4 and P. mirabilis UCe1 with SMIC50 of 32 and 128 μg/mL, desirable SMIC50 of bacteriocin for biofilm disruption were 128 and 256 μg/mL, respectively. The SEM results indicated that bacteriocin affected the cell morphology of biofilm-associated cells. The present findings indicated that bacteriocin from Bacillus sp. Sh10 has bactericidal properties against biofilm-forming isolates of P. mirabilis UCa4 and P. mirabilis UCe1 and has the ability to inhibit the formation of biofilm and disrupt established biofilm.

Comparative analysis of marine and freshwater viral haemorrhagic septicaemia virus (VHSV) isolates antagonistic activity

Viral Haemorrhagic Septicaemia Virus (VHSV) isolates virulent to marine fish species can replicate in freshwater species, although producing little or no mortality. Conversely, isolates from freshwater fish do not cause disease in marine species. An inverse relationship between VHSV virulence and host mx gene up-regulation has been described for several fish species, suggesting that differences between the antagonistic activity exerted by these isolates might be involved in the outcome of infections. In this study, the antagonistic activity against the type I interferon system of two representative marine and freshwater VHSV isolates has been characterised using RTG-2 cells stably transfected with the luciferase gene under the control of the Senegalese sole mx (ssmx) promoter, RTG pssmx-luc cells. Both isolates exerted a dose-dependent negative effect on the activation of ssmx promoter, showing a notably different minimal viral dose to exert the antagonism. In particular, an inverse relationship between the minimal MOI required and the viral virulence to sole has been recorded, which suggests this parameter as a possible in vivo VHSV virulence marker. Furthermore, the quantification of the endogenous inf I, mx1 and mx3 mRNA has demonstrated differences between both isolates in their antagonistic activity. Besides, a different nv RNA kinetics, which seems to depend on specific cellular factors, has been recorded for both isolates. This knowledge could contribute to the development of efficient tools to fight against viral infections in fish farming. For that purpose, the RTG pssmx-luc cells may be a suitable in vitro tool to identify the molecular mechanisms underlying VHSV-host interactions.

Insight into the resilience and susceptibility of marine bacteria to T6SS attack by Vibrio cholerae and Vibrio coralliilyticus.

The type VI secretion system (T6SS) is a nanomachine capable of killing adjacent microbial cells in a contact-dependent manner. Due to limited studies, relatively little is known about the range of marine bacteria that are susceptible to T6SS attack. Here, 15 diverse marine bacterial isolates from the phyla Bacteroidetes and Ɣ-Proteobacteria were challenged against the marine bacterium and human pathogen, Vibrio cholerae, which has a well described T6SS. V. cholerae killed several of the tested Ɣ-Proteobacteria, including members of the orders Vibrionales, Alteromonadales, Oceanospirillales, and Pseudomonadales. In contrast, V. cholerae co-existed with multiple Bacteroidetes and Ɣ-Proteobacteria isolates, but was killed by Vibrio coralliilyticus. Follow-up experiments revealed that five V. coralliilyticus strains, including known coral and shellfish pathogens survived the T6SS challenge and killed V. cholerae. By using predicted protein comparisons and mutagenesis, we conclude that V. coralliilyticus protected itself in the challenge by using its own T6SS to kill V. cholerae. This study provides valuable insight into the resilience and susceptibility of marine bacteria to the V. cholerae T6SS, and provides the first evidence for a functional T6SS in V. coralliilyticus, both of which have implications for human and ocean health.

Kinetic models application on glycerol production from glucose by the marine yeast Candida orthopsilosis

This study aims to investigate the kinetic parameters of carbon source effect on biomass and glycerol production by Candida orthopsilosis HH52 isolated from marine sediment. The yeast isolate HH52 was selected among five tested marine isolates as the most efficient producer of glycerol (64.42 gl-1). Based on the phenotypic and genotypic identification, this isolate was identified as Candida orthopsilosis and was deposited in the GenBank database under the accession number MK156301. The maximum specific glycerol production rate (vmax) was 0.101 gg-1 (glycerol /biomass) at 225 gl-1glucose, whereas the maximum specific growth rate (μmax) was 0.211 hr-1 at 150 gl-1 glucose. The glycerol (YP/S) and biomass (YX/S) yield coefficients were 0.3576 and 0.1128 gg-1, respectively. These findings suggest the possibility of using Candida orthopsilosis HH52 as a promising source for glycerol production.