Halorubrum glutamatedens sp. nov., a Halophilic Archaeon Isolated from a Rock Salt.

An extremely halophilic archaeon, strain ZY8T, was isolated from a rock salt of Yunnan salt mine. It was able to grow at 12-30% (w/v) NaCl (optimum, 15-20%), pH 7.0-9.0 (optimum, pH 8.5), and 20-45 °C (optimum, 42 °C). Sequence similarity search of its 16S rRNA gene showed that strain ZY8T belonged to the genus Halorubrum, and it is closely related to species of H. aethiopicum SAH-A6T (98.6%), H. aquaticum EN-2T (98.6%), and H. halodurans Cb34T (98.5%), respectively. Strain ZY8T contained phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester and phosphatidylglycerol sulfate as its major phospholipids, and a sulfated diglycosyl diether as its major glycolipid. The DNA G+C content was 66.7 mol%. DNA-DNA relatedness between strains ZY8T and closely related species were far below 70%. Based on the phenotypic and phylogenetic analyses, it is proposed that strain ZY8T represents a novel species of the genus Halorubrum, for which the name Halorubrum glutamatedens sp. nov. is proposed. The type strain is ZY8T (=CGMCC 1.16026T=NBRC 112866T).

An Antarctic Extreme Halophile and Its Polyextremophilic Enzyme: Effects of Perchlorate Salts.

Effects of perchlorate salts prevalent on the surface of Mars are of significant interest to astrobiology from the perspective of potential life on the Red Planet. Halorubrum lacusprofundi, a cold-adapted halophilic Antarctic archaeon, was able to grow anaerobically on 0.04 M concentration of perchlorate. With increasing concentrations of perchlorate, growth was inhibited, with half-maximal growth rate in ca. 0.3 M NaClO4 and 0.1 M Mg(ClO4)2 under aerobic conditions. Magnesium ions were also inhibitory for growth, but at considerably higher concentrations, with half-maximal growth rate above 1 M. For a purified halophilic ß-galactosidase enzyme of H. lacusprofundi expressed in Halobacterium sp. NRC-1, 50% inhibition of catalytic activity was observed at 0.88 M NaClO4 and 0.13 M Mg(ClO4)2. Magnesium ions were a more potent inhibitor of the enzyme than of cell growth. Steady-state kinetic analysis showed that Mg(ClO4)2 acts as a mixed inhibitor (KI = 0.04 M), with magnesium alone being a competitive inhibitor (KI = 0.3 M) and perchlorate alone acting as a very weak noncompetitive inhibitor (KI = 2 M). Based on the estimated concentrations of perchlorate salts on the surface of Mars, our results show that neither sodium nor magnesium perchlorates would significantly inhibit growth and enzyme activity of halophiles. This is the first study of perchlorate effects on a purified enzyme. Key Words: Halophilic archaea-Perchlorate-Enzyme inhibition-Magnesium. Astrobiology 18, 412-418.

Characterization of a bacterioruberin-producing Haloarchaea isolated from the marshlands of the Odiel river in the southwest of Spain.

In this work, we describe the isolation, identification, pigment characterization, and optimization of the culture conditions for a haloarchaea strain isolated from salt evaporation ponds in the Odiel river, at Southwest of Spain. The haloarchaea belongs to the genus Halorobrum, as deduced from the analysis of its 16S rRNA encoding gene and has been designated as Halorubrum sp. SH1. The growth conditions for the new strain were optimized studying temperature, NaCl concentration, agitation rate and light intensity. The C50-carotenoids, bacterioruberin, and its derivatives bisanhydrobacterioruberin and trisanhydrobacterioruberin, were found to be the predominant pigments produced by this strain of Halorubrum, as determined using HPLC-DAD and UHPLC-ESI-MS/MS techniques. This extremely halophilic archaeon could be a good candidate for the production of bacterioruberins of high added-value due to their coloring, antioxidant, and possible anticancer properties. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:592-600, 2016.

Agar-supported cultivation of Halorubrum sp. SSR, and production of halocin C8 on the scale-up prototype Platotex.

Halorubrum sp. SSR was isolated from a solar saltern in Algeria. The strain exhibited a high antibiotic activity against the indicator strain Natronorubrum aibiense G23, and the bioactive compound showed thermal, acid and alkali stability. SSR was grown on agar-supported cultivation (AgSF) to compare yields and applicability with traditional submerged cultivation. AgSF scale-up was implemented taking benefit from the solid-state cultivation prototype Platotex. This technology leads to high amounts of the target Halocin and facilitate the downstream steps. The antibiotic compound was purified according to a fast efficient procedure including ion exchange chromatography followed by a fractionation on C18 Sep-Pack cartridge. The compound was identified as Halocin C8 according to N-terminal amino acid sequencing and high-resolution mass spectrometry.

Hyperhalophilic archaeal biofilms: growth kinetics, structure, and antagonistic interaction in continuous culture.

Biofilms by the hyperhalophilic archaea Halorubrum sp. and Halobacterium sp. were analyzed, and for the first time the progression of structural features and the developmental parameters of these sessile populations are described. Optical slicing and digital analysis of sequential micrographs showed that their three dimensional structure was microorganism dependent. Biofilms of Halobacterium sp. developed in clusters that covered about 30% of the supporting surface at the interface level and expanded over about 86 ± 4 µm in thickness, while Halorubrum sp. biofilms covered less than 20% of the surface and reached a thickness of 41 ± 1 µm. The kinetics of growth was lower in biofilms, with generation times of 27 ± 1 and 36 ± 2 h for Halobacterium sp. and Halorubrum sp., respectively, as compared to 8.4 ± 0.3 and 14 ± 1 h in planktonic cultures. Differences between microorganisms were also observed at the cell morphology level. The interaction between the two microorganisms was also evaluated, showing that Halobacterium sp. can outcompete already established Halorubrum sp. biofilms by a mechanism that might include the combined action of tunnelling swimmers and antimicrobial compounds.

[Experimental interaction of halophilic prokaryotes and opportunistic bacteria in brine].

AIM: Study the effect of extremely halophilic archaea and moderately halophilic bacteria on preservation of opportunistic bacteria in brine. MATERIALS AND METHODS: 17 strains of moderately halophilic bacteria and 2 strains of extremely halophilic archaea were isolated from continental hypersaline lake Razval of Sol-Iletsk area of Orenburg Region. Identification of pure cultures of prokaryotes was carried out taking into account their phenotype properties and based on determination of 16S RNA gene sequence. The effect of halophilic prokaryote on elimination of Escherichia coli from brine was evaluated during co-cultivation. Antagonistic activity of cell extracts of the studied microorganisms was evaluated by photometric method. RESULTS: A more prolonged preservation of an E. coli strain in brine in the presence of live cells of extremely halophilic archaea Halorubrum tebenquichense and moderately halophilic bacteria Marinococcus halophilus was established. Extracts of cells of extremely halophilic archaea and moderately halophilic bacteria on the contrary displayed antagonistic activity. CONCLUSION: The protective effect of live cells of halophilic prokaryotes and antagonistic activity of their cell extracts change the period of conservation of opportunistic bacteria in brine that regulates inter-microbial interactions and changes the period of self-purification that reflects the sanitary condition of a hypersaline water body.