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isolate 492, was considered too small a fragment for confident identification. Isolate 494
produced the most AFP active protein extract of the 19 isolates identified by SPLAT
analysis as being AFP active (Chapter 4). Therefore it was considered important to
identify at least the genus of the isolate. Isolate 494 showed an identical relationship with
two other bacterial species using multiple restriction enzymes for ARDRA analysis (Fig
5.5). These two bacterial strains M3C203B-B (AF395031) and SIA181- IAI (AF395032)
(Christner et al., 2001) were also sequenced and shown to be members of the genus
Sphingomonas. This suggested that isolate 494 was probably also a member of the
Sphingomonas genus. This genus is characterised as Gram negative, non-spore forming
rods, with yellow pigmentation (Yabuuchi et al., 1990) which correlates with the
phenotypic characteristics of isolate 494. The genus Sphingomonas. which belongs to the
a-Proteobacteria, has been readily identified in certain Antarctic environments,
specifically oil contaminated soil (Panicker et al., 2002) and the accretion ice above Lake
Vostok (Christner et al., 2001). Bowman et al. (2000b) have also shown that a-
Proteobacteria are one of the most common groups of eubacteria isolated from the saline
lake sediments of the Vestfold Hills, Antarctica, although they did not isolate any
examples of the genus Sphingomonas. Strains of the genus Sphingomonas have been
isolated from a wide variety of sources including soil, marine and fresh water, marine
organisms, and from plants (http: //www. jgi. doe. gov/JGI_microbial). Isolate 494, being
the only AFP active isolate from the current study which did not belong to the y-
Proteobacteria, was isolated from Pendant Lake, which is brackish and marine derived
and therefore the presence of this isolate in such a lacustrine environment is not
remarkable. Isolate 494 showed 99.02% sequence similarity with a Sphingomonas sp
(AF39503 1) which was isolated from samples of melt water from the accretion ice above
Lake Vostok, this ice originated in the lake, indicating that the bacterial isolate may also
have originated in the lake (Christner et al., 2001), which further supports the isolation of
494 from a lacustrine system. In the phylogenetic tree analysis (Fig 5.4) the
Sphingomonas species, which was entered to propose the positioning of isolate 494, was
a distantly related outlier to the main group. This is as expected because Sphingomonas is
a genus belonging to the a-Proteobacteria, whereas the rest of the sequences in the
phylogenetic alignment belong to the y-Proteobacteria as already mentioned. This was
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