Development of a CRISPR/Cas9-mediated gene-editing method to isolate a mutant of the unicellular green alga Parachlorella kessleri strain NIES-2152 with improved lipid productivity.

Background: Previously, we isolated a mutant of Parachlorella kessleri named strain PK4 that accumulated higher concentrations of lipids than the wild-type strain. Resequencing of the PK4 genome identified mutations in three genes which may be associated with the high-lipid phenotype. The first gene, named CDMT1, encodes a protein with a calcium-dependent membrane association domain; the second gene, named DMAN1, encodes endo-1,4-β-mannanase, while the third gene, named AATPL1, encodes a plastidic ATP/ADP antiporter-like protein.

Nitrite as a causal factor for nitrate-dependent anaerobic corrosion of metallic iron induced by Prolixibacter strains.

Microbially influenced corrosion (MIC) may contribute significantly to overall corrosion risks, especially in the gas and petroleum industries. In this study, we isolated four Prolixibacter strains, which belong to the phylum Bacteroidetes, and examined their nitrate respiration- and Fe -corroding activities, together with two previously isolated Prolixibacter strains. Four of the six Prolixibacter strains reduced nitrate under anaerobic conditions, while the other two strains did not.

Cas9-guide RNA ribonucleoprotein-induced genome editing in the industrial green alga sp. strain KJ.

Background: Oxygen-evolving photosynthetic microorganisms, collectively termed as microalgae, are gaining attention as alternative fuel sources. The unicellular alga sp. strain KJ that belongs to the class Trebouxiophyceae can grow rapidly in minimal mineral media and accumulate triacylglycerols at levels > 60% (w/w) of its dry weight under nitrogen depletion conditions.

An extracellular [NiFe] hydrogenase mediating iron corrosion is encoded in a genetically unstable genomic island in Methanococcus maripaludis.

Certain methanogens deteriorate steel surfaces through a process called microbiologically influenced corrosion (MIC). However, the mechanisms of MIC, whereby methanogens oxidize zerovalent iron (Fe), are largely unknown. In this study, Fe-corroding Methanococcus maripaludis strain OS7 and its derivative (strain OS7mut1) defective in Fe-corroding activity were isolated.

Metabolic engineering using iterative self-cloning to improve lipid productivity in Coccomyxa.

We previously developed a self-cloning system that introduces cDNA of the uridine monophosphate synthase gene (cUMPS) of Coccomyxa sp. strain Obi as a selectable marker into uracil-auxotrophic mutants (Ura) of the same alga. Here, we developed a Cre/loxP-based system for the removal of cUMPS flanked by directly repeated loxP sites from the Coccomyxa genome using the intracellular delivery of purified Cre recombinase to generate an Ura strain that was used as a host for second-round transformation using cUMPS as the selection marker.

Biofilm-based photobioreactor absorbing water and nutrients by capillary action.

Cells of the unicellular green alga, "Pseudochoricystis ellipsoidea", were uniformly spread on a cellulosic sheet or on a polytetrafluoroethylene (PTFE) membrane sheet superimposed on a cellulosic sheet at a density of 3.5-5.0gdry weight per m, and the sheet was adhered to an inverted V-shaped acrylic plate of 10cm in height.

Construction of Marker-Free Transgenic Strains of Chlamydomonas reinhardtii Using a Cre/loxP-Mediated Recombinase System.

The Escherichia coli bacteriophage P1 encodes a site-specific recombinase called Cre and two 34-bp target sites of Cre recombinase called loxP. The Cre/loxP system has been used to achieve targeted insertion and precise deletion in many animal and plant genomes. The Cre/loxP system has particularly been used for the removal of selectable marker genes to create marker-free transgenic organisms.

Construction of a self-cloning system in the unicellular green alga Pseudochoricystis ellipsoidea.

Background: Microalgae have received considerable interest as a source of biofuel production. The unicellular green alga Pseudochoricystis ellipsoidea (non-validated scientific name) strain Obi appears to be suitable for large-scale cultivation in outdoor open ponds for biodiesel production because it accumulates lipids to more than 30 % of dry cell weight under nitrogen-depleted conditions. It also grows rapidly under acidic conditions at which most protozoan grazers of microalgae may not be tolerant.

Iron corrosion induced by nonhydrogenotrophic nitrate-reducing Prolixibacter sp. strain MIC1-1.

Microbiologically influenced corrosion (MIC) of metallic materials imposes a heavy economic burden. The mechanism of MIC of metallic iron (Fe(0)) under anaerobic conditions is usually explained as the consumption of cathodic hydrogen by hydrogenotrophic microorganisms that accelerates anodic Fe(0) oxidation. In this study, we describe Fe(0) corrosion induced by a nonhydrogenotrophic nitrate-reducing bacterium called MIC1-1, which was isolated from a crude-oil sample collected at an oil well in Akita, Japan.

Corrosion of iron by iodide-oxidizing bacteria isolated from brine in an iodine production facility.

Elemental iodine is produced in Japan from underground brine (fossil salt water). Carbon steel pipes in an iodine production facility at Chiba, Japan, for brine conveyance were found to corrode more rapidly than those in other facilities. The corroding activity of iodide-containing brine from the facility was examined by immersing carbon steel coupons in "native" and "filter-sterilized" brine samples.

Influence of extracellular polysaccharides (EPS) produced by two different green unicellular algae on membrane filtration in an algae-based biofuel production process.

In the present study, two strains of green algae named S1 and S2, categorized as the same species of Pseudo-coccomyxa ellipsoidea but showing 99% homology, were cultivated under the same conditions and filtrated with a microfiltration membrane. On the basis of the results of the extracellular polysaccharides (EPS) characteristics of these two green algae and the degree of fouling, the influence of these characteristics on the performance of membrane filtration was investigated. There was no difference in the specific growth rate between the S1 and S2 strains; however, large differences were seen in the amount and quality of EPS between S1 and S2.

Identification of novel non-metal haloperoxidases from the marine metagenome.

Haloperoxidase (HPO, E.C.1.

Genetic transformation of Pseudochoricystis ellipsoidea, an aliphatic hydrocarbon-producing green alga.

Pseudochoricystis ellipsoidea is a recently isolated unicellular green alga, which is classified within the family Trebouxiophyceae. This alga has a unique ability to synthesize and accumulate intracellularly a significant amount of aliphatic hydrocarbons. To elucidate molecular mechanisms of the hydrocarbon production in this organism, the development of genetic methods including DNA transformation methods are important.

Mobilicoccus pelagius gen. nov., sp. nov. and Piscicoccus intestinalis gen. nov., sp. nov., two new members of the family Dermatophilaceae, and reclassification of Dermatophilus chelonae (Masters et al. 1995) as Austwickia chelonae gen. nov., comb. nov.

Two Gram-positive bacteria, designated strains Aji5-31(T) and Ngc37-23(T), were isolated from the intestinal tracts of fishes. 16S rRNA gene sequence analysis indicated that both strains were related to the members of the family Dermatophilaceae, with 95.6-96.

Advances in the field of high-molecular-weight polycyclic aromatic hydrocarbon biodegradation by bacteria.

Interest in understanding prokaryotic biotransformation of high-molecular-weight polycyclic aromatic hydrocarbons (HMW PAHs) has continued to grow and the scientific literature shows that studies in this field are originating from research groups from many different locations throughout the world. In the last 10 years, research in regard to HMW PAH biodegradation by bacteria has been further advanced through the documentation of new isolates that represent diverse bacterial types that have been isolated from different environments and that possess different metabolic capabilities. This has occurred in addition to the continuation of in-depth comprehensive characterizations of previously isolated organisms, such as Mycobacterium vanbaalenii PYR-1.

A simple and rapid GC/MS method for the simultaneous determination of gaseous metabolites.

We modified and tuned a commercial model of a gas chromatography/mass spectrometry (GC/MS) instrument to develop a simple and rapid method for the simultaneous quantification of a variety of gas species. Using the developed method with the newly modified instrument, gas species such as H(2), N(2), O(2), CO, NO, CH(4), CO(2), and N(2)O, which are common components of microbial metabolism, were accurately identified based on their retention times and/or mass-to-charge ratios (m/z) in less than 2.5 min.

Iron corrosion activity of anaerobic hydrogen-consuming microorganisms isolated from oil facilities.

The purpose of the present study was to test the hypothesis that anaerobic hydrogen-consuming microorganisms generally promote iron corrosion. We isolated 26 hydrogen-consuming microorganisms (acetogens, sulfate-reducing bacteria, and methanogens) from oil facilities in Japan using hydrogen as an electron donor. The iron corrosion activities of these microorganisms were examined using iron (Fe0) granules as the sole electron donor.

The potential of Cycloclasticus and Altererythrobacter strains for use in bioremediation of petroleum-aromatic-contaminated tropical marine environments.

Cycloclasticus sp. A5, which has been suggested to be a major degrader of petroleum aromatics spilled in temperate seas, showed higher degrading activities for petroleum aromatics, at both 25 degrees C and tropical sea temperature 30 degrees C, than the novel aromatic-degrading isolates, related to Altererythrobacter epoxidivorans (97.5% similarity in the almost full-length 16S rRNA gene sequence) and Rhodovulum iodosum (96.

Oleibacter marinus gen. nov., sp. nov., a bacterium that degrades petroleum aliphatic hydrocarbons in a tropical marine environment.

Three Gram-negative, motile, mesophilic, aerobic, rod-shaped bacterial strains, designated 2O1(T), 1O14 and 1O18, were isolated from Indonesian seawater after enrichment with crude oil and a continuous supply of supplemented seawater. The strains exhibited high n-alkane-degrading activity, which indicated that the strains were important degraders of petroleum aliphatic hydrocarbons in tropical marine environments. Phylogenetic analyses based on 16S rRNA gene sequences of members of the Gammaproteobacteria showed that the isolates formed a coherent and distinct cluster in a stable lineage containing Oceanobacter kriegii IFO 15467(T) (96.

Methanobacterium petrolearium sp. nov. and Methanobacterium ferruginis sp. nov., mesophilic methanogens isolated from salty environments.

Two methane-producing archaea, designated Mic5c12(T) and Mic6c05(T), were isolated from sludge deposited in a crude oil storage tank and a tubercle on the interior of a pipe transporting natural gas-containing brine, respectively. The isolates were Gram-staining-variable, non-motile rods and grew only on H(2)/CO(2). Strain Mic6c05(T) produced methane from some alcohols without showing any growth; strain Mic5c12(T) did not utilize alcohols.

Iron-corroding methanogen isolated from a crude-oil storage tank.

Microbiologically influenced corrosion of steel in anaerobic environments has been attributed to hydrogenotrophic microorganisms. A sludge sample collected from the bottom plate of a crude-oil storage tank was used to inoculate a medium containing iron (Fe(0)) granules, which was then incubated anaerobically at 37 degrees C under an N(2)-CO(2) atmosphere to enrich for microorganisms capable of using iron as the sole source of electrons. A methanogen, designated strain KA1, was isolated from the enrichment culture.

Euzebya tangerina gen. nov., sp. nov., a deeply branching marine actinobacterium isolated from the sea cucumber Holothuria edulis, and proposal of Euzebyaceae fam. nov., Euzebyales ord. nov. and Nitriliruptoridae subclassis nov.

A tangerine-coloured, Gram-positive actinobacterial strain, designated F10(T), was isolated from the abdominal epidermis of a sea cucumber, Holothuria edulis, collected in seawater off the coast of Japan. A 16S rRNA gene sequence analysis indicated that strain F10(T) was a member of the class Actinobacteria and was most closely related to Nitriliruptor alkaliphilus ANL-iso2(T) (87.4 % sequence similarity).

Phycisphaera mikurensis gen. nov., sp. nov., isolated from a marine alga, and proposal of Phycisphaeraceae fam. nov., Phycisphaerales ord. nov. and Phycisphaerae classis nov. in the phylum Planctomycetes.

Three strains, FYK2301M01(T), FYK2301M18 and FYK2301M52, all being Gram-negative, spherical, motile and facultatively anaerobic, were isolated from a marine alga (Porphyra sp.) collected on Mikura Island, Japan. Colonies of the strains were circular and pink-pigmented on Marine Agar 2216 (Difco) at 25 degrees C.

Ignavibacterium album gen. nov., sp. nov., a moderately thermophilic anaerobic bacterium isolated from microbial mats at a terrestrial hot spring and proposal of Ignavibacteria classis nov., for a novel lineage at the periphery of green sulfur bacteria.

A moderately thermophilic chemoheterotrophic bacterium, strain Mat9-16(T), was isolated from microbial mats developed in hot spring water streams from Yumata, Nagano, Japan. Cells of strain Mat9-16(T) were strictly anaerobic, Gram-stain-negative, non-sporulating, non-motile and short to long rods (2.0-15.

Serinibacter salmoneus gen. nov., sp. nov., an actinobacterium isolated from the intestinal tract of a fish, and emended descriptions of the families Beutenbergiaceae and Bogoriellaceae.

A novel Gram-positive bacterium, designated Kis4-28(T), was isolated from the intestinal tract of a fish, and its taxonomic position was investigated by a polyphasic approach. The sample was collected from the coast of Tokyo Bay, Japan. Cells of strain Kis4-28(T) were rod-shaped, non-motile and non-sporulating.