A Novel Archaeal Lineage from an East African Alkaline Saltern
Susan GRANT*a, William GRANTa, Brian JONESb, Chiaki KATOc, and Lina LIc
a Department of Microbiology and Immunology, Univers. ity of Leicester, PO Box 138, Leicester LE1 9HN, UK
b Genencor International BV, PO Box 642, 2600 AP Delft, The Netherlands
c The DEEPSTAR group, Japan Marine Science and Technology Centre, 2-15 Natsushima-cho, Yokohoma 236, Japan
Soda lakes are characterised by the presence of large amounts of Na2CO3 and are significantly depleted in Mg2+ and Ca2+. The East African soda lakes are immensely productive, supporting dense populations of prokaryotes (1) and equally diverse ranging from relatively dilute examples such as lakes Bogoria and Elmenteita at pH 10.5 - 11.0 with total dissolved salts of around 5% (w/v) to pH 11.5 - 12.00 at lake Magadi where the brines are saturated with roughly equal proportions of Na2CO3 and NaC1. At lake Magadi, C1 is crystallized from the alkaline brines in a series of salterns. These salterns are populated by haloalkaliphilic archaea of the genera Natronobacterium and Natronococcus which have been repeatedly isolated particularly from the final, most concentrated salt-making ponds (2). In December 1996 we filtered brines from these ponds on site through 8 μm cellulose nitrate filters, transferred the filters to ice cold TE pH 9.0 containing 15% (w/v) NaC1, until we were able to extract DNA from these samples (usually within 3-4 h) using the guanidine thiocyanate-based Pharmacia Rapid Prep Microgenomic Isolation Kit. Extracts were transported back to the UK in a small volume of isopropanol. Attempts were made to amplify 16S rRNA genes using both bacterial and archaeal primers, but only archaeal primers were successful in generating 16S rRNA gene product. The amplified 16S rRNA gene product was cloned using the Stratagene PCR-Script AMP SK(+) Cloning Kit, and cloned 16S rRNA genes reamplified using M13 primers. Clones were selected for sequence determination by RFLP restriction analysis.
Phylogenetic analysis of ten clones indicate that nine of these cloned 16S rRNA genes are distinct from those of cultivated haloalkaliphiles and other halobacteria, forming three groups on the periphery of the halobacterial branch of the Euryarchaeota. One cloned gene shows only low similarity to known archaeal sequences, representing a distinct archaeal line only distantly related to the haloarchaeal branch of the Euryarchaeaota.
1. Duckworth, A.W., Grant, W.D., Jones, B.E., and van Steenbergen, R. (1996) FEMS Microbial Ecology, 19, 181 - 191.
2. Tindall, B.J., Ross, H.N.M., and Grant, W.D. (1984) System. Appl. Microbiol., 5, 41-47.
