Isolation and identification of two extremely halophilic archaea from sebkhas in the Algerian Sahara.

In Algeria, many salt lakes are to be found spread from southern Tunisia up to the Atlas Mountains in northern Algeria. Oum Eraneb and Ain El beida sebkhas (salt lakes), are located in the Algerian Sahara. The aim of this study was to explore the diversity of the halobacteria in this type of habitats. The physicochemical properties of these shallow saline environments were examined and compared with other hypersaline and marine ecosystems. Both sites were relatively alkaline with a pH around 8.57- 8.74 and rich in salt at 13% and 16% (w/v) salinity for Oum Eraneb and Ain El beida, respectively, with dominant ions of sodium and chloride. The microbial approach revealed the presence of two halophilic archaea, strains JCM13561 and A33T in both explored sebkhas. Growth occurred between 10 and 25% (w/v) NaCl and the isolates grow optimally at 20% (w/v) NaCl. The pH range for growth was 6 to 9.5 with an optimum at pH 7.5 for the first strain and 7 to 9.5 with an optimum pH at 8.5-9 for the second strain. On the basis of 16S rRNA gene sequence analysis, strains JCM13561 and A33T were most closely related to Halorubrum litoreum and Natronorubrum bangense (99% and 96% similarity, respectively).

Developing a genetic manipulation system for the Antarctic archaeon, Halorubrum lacusprofundi: investigating acetamidase gene function.

No systems have been reported for genetic manipulation of cold-adapted Archaea. Halorubrum lacusprofundi is an important member of Deep Lake, Antarctica (~10% of the population), and is amendable to laboratory cultivation. Here we report the development of a shuttle-vector and targeted gene-knockout system for this species. To investigate the function of acetamidase/formamidase genes, a class of genes not experimentally studied in Archaea, the acetamidase gene, amd3, was disrupted. The wild-type grew on acetamide as a sole source of carbon and nitrogen, but the mutant did not. Acetamidase/formamidase genes were found to form three distinct clades within a broad distribution of Archaea and Bacteria. Genes were present within lineages characterized by aerobic growth in low nutrient environments (e.g. haloarchaea, Starkeya) but absent from lineages containing anaerobes or facultative anaerobes (e.g. methanogens, Epsilonproteobacteria) or parasites of animals and plants (e.g. Chlamydiae). While acetamide is not a well characterized natural substrate, the build-up of plastic pollutants in the environment provides a potential source of introduced acetamide. In view of the extent and pattern of distribution of acetamidase/formamidase sequences within Archaea and Bacteria, we speculate that acetamide from plastics may promote the selection of amd/fmd genes in an increasing number of environmental microorganisms.

Halorubrum salinum sp. nov., isolated from a marine solar saltern.

The halophilic archaeal strain GX71(T) was isolated from the Gangxi marine solar saltern near the Weihai city of Shandong Province, China. Cells of the strain were pleomorphic and lysed in distilled water, stained Gram-negative and formed red-pigmented colonies. Strain GX71(T) was able to grow at 25-45 °C (optimum 30 °C), in the presence of 1.7-4.8 M NaCl (optimum 2.6 M NaCl), with 0.005-0.7 M MgCl2 (optimum 0.05 M MgCl2) and at pH 5.5-9.5 (optimum pH 7.0-7.5). Cells lysed in distilled water and the minimal NaCl concentration to prevent cell lysis was 10 % (w/v). The major polar lipids of the strain were phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, phosphatidylglycerol sulfate, one major glycolipid chromatographically identical to sulfated mannosyl glucosyl diether (S-DGD-3) and an unidentified lipid was also detected. The 16S rRNA gene sequence of strain GX71(T) showed 94.0-97.0 % similarity to members of the genus Halorubrum of the family Halobacteriaceae. The rpoB' gene sequence of strain GX71(T) was 87.3-93.4 % similarity to current members of the genus Halorubrum. The DNA G+C content of GX71(T) was 67.1 mol%. Strain GX71(T) showed low DNA-DNA relatedness with Halorubrum lipolyticum CGMCC 1.5332(T), Halorubrum saccharovorum CGMCC 1.2147(T), Halorubrum kocurii CGMCC 1.7018(T) and Halorubrum arcis CGMCC 1.5343(T), the most closely related members of the genus Halorubrum. The phenotypic, chemotaxonomic and phylogenetic properties suggest that strain GX71(T) represents a novel species of the genus Halorubrum, for which the name Halorubrum salinum sp. nov. is proposed. The type strain is GX71(T) (= CGMCC 1.10458(T) = JCM 17093(T)).