Limited accessibility of nitrogen supplied as amino acids, amides, and amines as energy sources for marine Thaumarchaeota

Genomic and physiological evidence from some strains of ammonia-oxidizing Thaumarchaeota demonstrate their additional ability to oxidize nitrogen (N) supplied as urea or cyanate, fueling conjecture about their ability to conserve energy by directly oxidizing reduced N from other dissolved organic nitrogen (DON) compounds. Similarly, field studies have shown rapid oxidation of polyamine-N in the ocean, but it is unclear whether Thaumarchaeota oxidize polyamine-N directly or whether heterotrophic DON remineralization is required. We tested growth of two marine Nitrosopumilus isolates on DON compounds including polyamines, amino acids, primary amines, and amides as their sole energy source. Though axenic cultures only consumed N supplied as ammonium or urea, there was rapid but inconsistent oxidation of N from the polyamine putrescine when cultures included a heterotrophic bacterium. Surprisingly, axenic cultures oxidized 15N-putrescine during growth on ammonia, suggesting co-metabolism or accelerated breakdown of putrescine by reactive metabolic byproducts. Nitric oxide, hydrogen peroxide, or peroxynitrite did not oxidize putrescine in sterile seawater. These data suggest that the N in common DON molecules is not directly accessible to marine Thaumarchaeota, with thaumarchaeal oxidation (and presumably assimilation) of DON-N requiring initial heterotrophic remineralization. However, reactive byproducts or enzymatic co-metabolism may facilitate limited thaumarchaeal DON-N oxidation.

PRELIMINARY STUDIES ON ISOLATION AND SCREENING OF ANTIBIOTIC PRODUCING SYMBIOTIC BACTERIA FROM STARFISH (Protoreaster nodosus) COLLECTED FROM COASTAL AREA TAKALAR REGENCY, INDONESIA

Marine organisms are well known for the availability of bioactive compounds which have various biological activities including antibacterial activity. Likewise, their symbiotic bacteria can also produce compounds that have similar activities. The purpose of this study was to isolate and screen the symbiotic bacteria from starfish (Protoreaster nodosus) collected from coastal area Takalar Regency, South Sulawesi, Indonesia. Isolation was carried out by the pour plate method using nutrient agar medium dissolved in sterile seawater. The isolated symbiotic bacteria were purified by using the quadrant method. The pure isolate was culture through submerged fermentation using nutrient broth media enriched with 1% yeast extract and sterile seawater for 7 days. The selected symbiont bacterial isolates were tested for their antibacterial activity against gram-positive and gram-negative bacteria using disc diffusion assays. The results of fermentation were separated from the biomasses and tested for antibacterial activity against Staphylococcus aureus (S. aureus, ATCC 25923), Bacillus subtilis (B. subtilis, ATCC 6633), Salmonella typhi (S. typhi, NCTC 786), and Escherichia coli (E. coli, ATCC 25923). The results of study revealed that four symbiotic bacteria (SB 1T, SB 2T, SB 3T, and SB 4T) were successfully isolated. All the SB isolates have good antibacterial activity against all tested bacterial strains with an average diameter of inhibition zone larger than 11 mm. Among all isolates, isolate SB 4T showed a remarkable size of zones growth inhibition (> 15 mm) against all tested bacterial strains. Thus, the symbiotic bacteria isolated from P. nodosus in this study have a promising broad-spectrum antibacterial activity.

Short-Term Exposure to Sterile Seawater Reduces Bacterial Community Diversity in the Sea Anemone, Exaiptasia diaphana

The global decline of coral reefs heightens the need to understand how corals may persist under changing environmental conditions. Restructuring of the coral-associated bacterial community, either through natural or assisted strategies, has been suggested as a means of adaptation to climate change. A low complexity microbial system would facilitate testing the efficacy of microbial restructuring strategies. We used the model organism for corals, Exaiptasia diaphana, and determined that short-term (3 weeks) exposure to filter-sterilized seawater conditions alone reduced the complexity of the microbiome. Metabarcoding of the V5–V6 region of the bacterial 16S rRNA gene revealed that alpha diversity was approximately halved in anemones reared in filter-sterilized seawater compared to controls reared in unfiltered seawater and that the composition (beta diversity) differed significantly between the two. By reducing the complexity of the E. diaphana microbiome, the development of a system for testing assisted strategies such as probiotics, is more feasible.

Distributed Biomanufacturing of Liquefied Petroleum Gas

Liquefied Petroleum Gas (LPG) is a major domestic and transport fuel. Its combustion lessens NOx, greenhouse gas and particulates emissions compared to other fuels. Propane – the major constituent of LPG – is a clean, high value ‘drop-in’ fuel that can help governments develop integrated fuels and energy policies with low carbon burden, providing solutions to the multi-faceted challenges of future energy supply. We show that bio-LPG (bio-propane and bio-butane) can be produced by microbial conversion of waste volatile fatty acids that can be derived from anaerobic digestion, industrial waste, or CO2via photosynthesis. Bio-LPG production was achieved photo-catalytically, using biomass propagated from bioengineered bacteria includingE. coli, Halomonas(in non-sterile seawater), andSynechocystis(photosynthetic). These fuel generation routes could be implemented rapidly in advanced and developing nations of the world to meet energy needs, global carbon reduction targets and clean air directives.

IDENTIFICATION AND LIFE CYCLE OF MARINE LEECH ISOLATED FROM CULTURED HYBRID GROUPER IN THE NORTHERN BALI WATERS OF INDONESIA

The aims of this study were to identify and to determine life cycle of marine leech isolated from cultured hybrid grouper “cantik” (Epinephelus fuscoguttatus fx E. polyphekadion m) in the northern Bali waters of Indonesia under laboratory conditions. Observation of the life cycle of the marine leech was done using petri-dishes (9 cm in diameter) arranged into two groups. In group-1, a petri-dish was filled with sterile seawater (with water exchange of 50%-60% every two days) and in group-2, a petri-dish was filled with continuous running water. DNA sequence was aligned with the sequences from GenBank by BLAST program. Results of similarity index with GenBank sequence exhibited that the nucleic acid of the marine leech isolated from the hybrid grouper “cantik” showed high similarity (99%) with Zeylanicobdella arugamensis. One adult leech could produce 1-63 eggs. The eggs were developed into morula, blastula, and gastrula within five days. The early phase of the embryo with daily water exchange treatment started on day-6 and hatched into larvae on day-10. The eggs incubated with continuous running water had hatched faster (eight days). However, not all eggs hatched at the same time. Some of the eggs hatched 1-3 days after the first one. Hatching rate of eggs varied from 2.70% to 100%. The newly hatched Z. arugamensis larva has transparent color and length of 1.0-1.5 mm. On day-6, Z. arugamensis larvae were already seen attaching to the body of the fish. The size of the Z. arugamensis larvae ranged between 3-11 mm on day-9. In that stage, they were able to produce eggs. Therefore, we argue that Z. arugamensis only requires 17 to 22 days to develop into the adult stage.

Labyrinthulomycota from Brazilian mangrove swamps and coastal waters

The phylum Labyrinthulomycota is composed of three distinct groups of fungus-like organisms: the labyrinthulids, the thraustochytrids and the aplanochytrids. They are present in marine and brackish waters as saprobes and/or parasites. In recent years, there was an increased interest in studying these organisms due to the ability that some have to produce large amounts of lipids, particularly polyunsaturated fatty acids. However, in spite of their importance and diversity, in Brazil only four species were reported in the 1970s. Due to their ecological and economic importance globally and the limited knowledge of them in the country, this study aimed to evaluate the diversity of the Labyrinthulomycota in mangrove swamps and coastal waters of São Paulo State, Brazil. The specimens were isolated from water and leaf samples using different methodologies, and purified in an agar medium prepared with half-strength sterile seawater, peptone, yeast extract and glucose. After purification, they were identified by means of morphology and phylogenetic analysis of the SSU rDNA region. A total of 193 specimens representing five genera were observed, 26 specimens were incorporated into the culture collections and 29 sequences of the SSU rDNA region were deposited in GenBank.Labyrinthulasp.,Aurantiochytriumsp.,Parietichytrium sarkarianumandThraustochytrium striatumare new records for Brazil, contributing to our knowledge of the diversity of fungus-like organisms in the country.

Peeking under the Iron Curtain: Development of a Microcosm for Imaging the Colonization of Steel Surfaces by Mariprofundus sp. Strain DIS-1, an Oxygen-Tolerant Fe-Oxidizing Bacterium

ABSTRACTMicrobially influenced corrosion (MIC) is a major cause of damage to steel infrastructure in the marine environment. Despite their ability to grow directly on Fe(II) released from steel, comparatively little is known about the role played by neutrophilic iron-oxidizing bacteria (FeOB). Recent work has shown that FeOB grow readily on mild steel (1018 MS) incubatedin situor as a substrate for pure culturesin vitro; however, details of how they colonize steel surfaces are unknown yet are important for understanding their effects. In this study, we combine a novel continuously upwelling microcosm with confocal laser scanning microscopy (CLSM) to determine the degree of colonization of 1018 MS by the marine FeOB strain DIS-1. 1018 MS coupons were incubated with sterile seawater (pH 8) inoculated with strain DIS-1. Incubations were performed both under oxic conditions and in an anoxic-to-oxic gradient. Following incubations of 1 to 10 days, the slides were removed from the microcosms and stained to visualize both cells and stalk structures. Stained coupons were visualized by CLSM after being mounted in a custom frame to preserve the three-dimensional structure of the biofilm. The incubation of 1018 MS coupons with strain DIS-1 under oxic conditions resulted in initial attachment of cells within 2 days and nearly total coverage of the coupon with an ochre film within 5 days. CLSM imaging revealed a nonadherent biofilm composed primarily of the Fe-oxide stalks characteristic of strain DIS-1. When incubated with elevated concentrations of Fe(II), DIS-1 colonization of 1018 MS was inhibited.IMPORTANCEThese experiments describe the growth of a marine FeOB in a continuous culture system and represent direct visualizations of steel colonization by FeOB. We anticipate that these experiments will lay the groundwork for studying the mechanisms by which FeOB colonize steel and help to elucidate the role played by marine FeOB in MIC. These observations of the interaction between an FeOB, strain DIS-1, and steel suggest that this experimental system will provide a useful model for studying the interactions between microbes and solid substrates.

Specificity of Associations between Bacteria and the Coral Pocillopora meandrina during Early Development

ABSTRACTRelationships between corals and specific bacterial associates are thought to play an important role in coral health. In this study, the specificity of bacteria associating with the coralPocillopora meandrinawas investigated by exposing coral embryos to various strains of cultured marine bacteria, sterile seawater, or raw seawater and examining the identity, density, and location of incorporated cells. The isolates utilized in this experiment included members of the Roseobacter and SAR11 clades of theAlphaproteobacteria, aPseudoalteromonasspecies of theGammaproteobacteria, and aSynechococcusspecies of theCyanobacteriaphylum. Based on terminal restriction fragment length polymorphism analysis of small-subunit rRNA genes, similarities in bacterial communities associated with 170-h-old planulae were observed regardless of treatment, suggesting that bacteria may have been externally associated from the outset of the experiment. Microscopic examination ofP. meandrinaplanulae by fluorescencein situhybridization with bacterial and Roseobacter clade-specific oligonucleotide probes revealed differences in the densities and locations of planulae-associated cells. Planulae exposed to either raw seawater or strains ofPseudoalteromonasand Roseobacter harbored the highest densities of internally associated cells, of which 20 to 100% belonged to the Roseobacter clade. Planulae exposed to sterile seawater or strains of the SAR11 clade andSynechococcusdid not show evidence of prominent bacterial associations. Additional analysis of the raw-seawater-exposed planulae via electron microscopy confirmed the presence of internally associated prokaryotic cells, as well as virus-like particles. These results suggest that the availability of specific microorganisms may be an important factor in the establishment of coral-bacterial relationships.

Evaluation of the Petrifilm Aerobic Count Plate for Enumeration of Aerobic Marine Bacteria from Seawater and Caulerpa lentillifera

The enumeration and evaluation of the activity of marine bacteria are important in the food industry. However, detection of marine bacteria in seawater or seafood has not been easy. The Petrifilm aerobic count plate (ACP) is a ready-to-use alternative to the traditional enumeration media used for bacteria associated with food. The purpose of this study was to evaluate the usefulness of a simple detection and enumeration method utilizing the Petrifilm ACP for enumeration of aerobic marine bacteria from seawater and an edible seaweed, Caulerpa lentillifera. The efficiency of enumeration of total aerobic marine bacteria on Petrifilm ACP was compared with that using the spread plate method on marine agar with 80 seawater and 64 C. lentillifera samples. With sterile seawater as the diluent, a close correlation was observed between the method utilizing Petrifilm ACP and that utilizing the conventional marine agar (r = 0.98 for seawater and 0.91 for C. lentillifera). The Petrifilm ACP method was simpler and less time-consuming than the conventional method. These results indicate that Petrifilm ACP is a suitable alternative to conventional marine agar for enumeration of marine microorganisms in seawater and C. lentillifera samples.