CHARACTERIZATION OF ATTACHED BACTERIAL-POPULATIONS IN DEEP GRANITIC GROUNDWATER FROM THE STRIPA RESEARCH MINE BY 16S RIBOSOMAL-RNA GENE SEQUENCING AND SCANNING ELECTRON-MICROSCOPY

This paper presents the molecular characterization of attached bacteri al populations growing in slowly flowing artesian groundwater from dee p crystalline bed-rock of the Stripa mine, south central Sweden. Bacte ria grew on glass slides in laminar flow readers connected to the anox ic groundwater flowing up through tubing from two levels of a borehole , 812-820 m and 970-1240 m. The glass slides were collected, the bacte rial DNA was extracted and the 16S rRNA genes were amplified by PCR us ing primers matching universally conserved positions 519-536 and 1392- 1405. The resulting PCR fragments were subsequently cloned and sequenc ed. The sequences were compared with each other and with 16S rRNA gene sequences in the EMBL database. Three major groups of bacteria were f ound. Signature bases placed the clones in the appropriate systematic groups. All belonged to the proteobacterial groups beta and gamma. One group was found only at the 812-820 m level, where it constituted 63% of the sequenced clones, whereas the second group existed almost excl usively at the 970-1240 m level, where it constituted 83% of the seque nced clones. The third group was equally distributed between the level s. A few other bacteria were also found. None of the 16S rRNA genes fr om the dominant bacteria showed more than 88% similarity to any of the others, and none of them resembled anything in the database by more t han 96%. Temperature did not seem to have any effect on species compos ition at the deeper level. SEM images showed rods appearing in microco lonies. The conditions at the levels differ in ph. temperature, redox and flow rate, and in content of sulphate, iron and sulphide. The pres ence of one dominant species in the laminar-flow reactors at each leve l indicates that the environments might have offered restrictive physi cal or physiological conditions difficult to adapt to.