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2006 Laboratory 2 Background Information
I. Basic Facts About Microbes: Microbes are believed to be the common ancestors of all
organisms (see Figure below; instruct.uwo.ca/ biology/284/intro.html). Microbes not only
grow virtually everywhere but also are present in abundance. In contrast to the relatively
small number of humans (6 x 109), populations of terrestrial and marine bacteria are immense, 5
x 1030 and 1.2 x 1029, respectively. In fact, the human body contains ten times more bacterial
cells than human cells. Microbes carry out innumerable transformations of matter that are
essential to life and thus have an enormous effect on climate and the geosphere. Much of our
knowledge base in biology and molecular biology has derived from microbial studies.
II. The Ubiquitous SAR 11 Clade: Previously Uncultivated Oceanic Microbes:
When scientists realized in the early 1990’s that that only a small fraction (1%) of microbes in
natural communities was known, these communities became the focus of many studies. Several
years later, however, it is still shocking to realize the depth of our ignorance, which is well
illustrated by the story of the SAR11 clade. When the technique of ribotyping (cloning and
sequencing 16S rRNA genes) was first used to survey natural ecosystems, SAR 11 was one of
the first groups of novel microbes to be discovered (Giovannoni et. al., 1990). We now know that
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2006 Laboratory 2 Background Information
the highest percentage of bacterial 16S ribosomal genes present in all oceanic and coastal waters
are from members of the SAR11 clade, making this group “one of the most successful clades of
organisms on the planet” (Morris et al, 2002; Giovannoni et. al., 2005). In some waters, SAR11
comprise up to 50% of the total surface bacterial community and approximately 25% of the subeuphotic zones. On average, members of the SAR11 clade account for about one third of all cells
in surface waters. Both the great abundance and global distribution of SAR11 suggests that its
members are instrumental in metabolizing oceanic dissolved organic matter (DOM), but the
specific roles of these bacteria in biogeochemical cycles is still unknown (Malmstrom et al.,
2004; Malmstrom et al, 2005).
Although most members of the SAR11 clade remain uncultured, a few have recently been
cultivated in the laboratory in seawater, though growth in synthetic medium remains elusive. As
a result of several subsequent studies, SAR 11 was placed into the -subclass of Proteobacteria.
However, members of the SAR11 group show less than 82% sequence similarity to cultivated Proteobacteria (Rappe et al., 2002.). One SAR11 isolate, Pelagibacter ubique. has the smallest
genome (1.3 x 106 base pairs) of any known free-living cell in nature capable of independent
replication (Rappe et al., 2002; Giovannoni et al. 2005). Although surprisingly the small P.
ubique genome encodes almost all basic functions characteristic of -Proteobacteria, this
genome contains little, if any, nonfunctional or redundant DNA and very short intergenic DNA
regions, averaging only three bases in length (Giovannoni et al. 2005). It seems certain that many
more surprises await from future studies of SAR 11.
III. Nucleotide Sequence of 16S rRNA Genes: The basis for classification by
ribotyping is the sequence of the 16S ribosomal RNA gene in prokaryotes and the 18S gene in
eukaryotes. The 16S and 18S rRNA genes were selected for classification and identification of
microbes because these genes are universal and essential; all living organisms must synthesize
proteins to survive (Woese and Fox, 1977). These genes are also well suited for this purpose
because they contain both conserved and variable regions, as is evident in the nucleotide
sequence of the 16S gene shown in the Figure on the following pages. Sequences that are highly
conserved are shown in brown, conserved regions are red, variable regions are black, highly
variable sequences are blue and sequences that are > 75% variable are green. The map locations
of some common PCR primers are also shown in the Figure, which was adapted from Baker et
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2006 Laboratory 2 Background Information
al, 2003. Conserved sequences are found in the 16S genes of all members of a domain, bacteria
or archaea, and are used as universal PCR primers. In contrast, variable sequences are
characteristic of certain genera, species or strains and are useful as more specific PCR primers.
IV. References
Baker, G.C., Smith, J.J., and D.A. Cowan. 2003. “Review and analysis of domain-specific 16S
primers.” J. Microbiol. Meth. 55: 541 – 555.
Giovannoni, S.J., Britschgi, T.B., Moyer, C.L. and K.G. Field. 1990. “Genetic diversity in
Sargasso Sea bacterioplankton.” Nature 345: 60 – 63.
Giovannoni, S.J., Tripp, H.J., Givan, S., Podar, M., Vergin, K.L., Baptista, D., Bibbs, L., Eads,
J., Richardson, T.H., Noordewier, M., Rappe, M.S., Short, J.M., Carrington, J.C., and E.J.
Mathur. 2005. “Genome streamlining in a cosmopolitan oceanic bacterium.” Science: 309:
1242 – 1245.
Malmstrom, R.R., Kiene, R.P., Cottrell, M. T., and D. L. Kirchman. 2004. “Contribution of
SAR11 bacteria to dissolved dimethysulfonioproprionate and amino acid uptake in the North
Atlantic Ocean.” Appl. Environ. Microbiol. 70: 4129 - 4135.
Malmstrom, R.R., Cottrell, M. T., Elifantz, H., and D. L. Kirchman. 2005. “Biomass production
and assimilation of dissolved organic matter by SAR11 bacteria in the northwest Atlantic
Ocean.” Appl. Environ. Microbiol. 71: 2979 - 2986.
Morris, R.M., Rappe, M.S., Connn, S.A., Vergin, K.L., Siebold, W.A., Carlson, C.A., and S.J.
Giovannoni. 2002. “SAR11 clase domoinates ocean surface bacterioplankton communities.”
Nature 420: 806 – 810.
Rappe, M.S. Connon, S.A., Vergin, K.L., and S.J. Giovannoni. 2002. “Cultivation of the
ubiquitous SAR11 marine bacterioplankton clade.” Nature 418: 630 – 633
Woese, C. R. and G. E. Fox. 1977. “Phylogenetic Structure of the Prokaryotic Domain: The
Primary Kingdoms.” Proc. Natl. Acad. Sci. USA. 74: 5088 - 5090.
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2006 Laboratory 2 Background Information
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2006 Laboratory 2 Background Information
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