Outlook
GENOME ANALYSIS
Archaeal and bacterial hyperthermophiles
Archaeal and bacterial
hyperthermophiles
horizontal gene exchange or common ancestry?
ravind and colleagues1 recently concluded that massive
gene transfer has occurred from Archaea to the bacterial ancestors of the hyperthermophile Aquifex aeolicus.
Their analyses were based primarily on similarity searches
of all complete bacterial genomes against the nonredundant protein sequence database, showing that the
genome of A. aeolicus2 has a much larger fraction of proteins with best hits to archaeal proteins than any other
bacterium. In particular, they reported that 246 Aquifex
proteins are most similar to archaeal proteins, with 26 of
these proteins belonging to families found only in Archaea
and Aquifex. Thus, the authors suggested that at least
10% of the Aquifex genes have been horizontally transferred from Archaea. Although we agree that gene transfer
has played an important role in the history of life3,4, we do
not agree with the conclusions that Aravind et al.1 reach.
In particular, limitations imposed by their assumptions
and flaws in the analyses and conclusions drawn will be
discussed.
The two most fundamental problems with the conclusions of the work cited of Aravind et al.1 are that the
authors (1) ignore the evidence that Aquifex is the most
deeply branching eubacterium with a complete genome
sequence5 (Fig. 1) and (2) assume that hyperthermophilicity in Bacteria and Archaea are independent inventions.
Firstly, ancestral genes passed vertically through the bacterial lineage could be transmitted to Aquifex (and possibly to other early diverging bacterial lineages) but are
lost in the common ancestor of the more recently diverging bacterial lineages for which genome sequences are
available. Neither the data nor the discussion of Aravind
et al.1 deals with this simple explanation of genes that are
shared by Aquifex and the Archaea but are absent in other
bacterial genomes.
Secondly, hyperthermophiles are represented among all
of the deepest and least diverged lineages both in Bacteria
and Archaea6 (Fig. 1), leading many workers6,7 but not
all8 to argue that the last universal common ancestor
(cenancestor) was a hyperthermophile. Aravind et al.1
ignored this possibility when they interpreted their data.
Regardless of whether the root of the tree of life is placed
in the bacterial branch9,10 or in the eukaryotic branch11,
and regardless of whether life originated at a hot environment or started cool and later adapted to high temperatures12, it is possible – even likely – that hyperthermophilicity was invented once (prior to the last prokaryotic
common ancestor stage; Fig. 1), and not independently at
two or more later times (as explicitly assumed by Aravind
et al.1). If the cenancestor was a hyperthermophile, it
would be natural for it to pass genes contributing to
A
*Nikos C. Kyrpides
nikos@darwin.life.
uiuc.edu
Gary J. Olsen
gary@phylo.life.
uiuc.edu
Department of
Microbiology, University
of Illinois at UrbanaChampaign, IL 61801,
USA.
*Also at the Mathematics
and Computer Science
Division, Argonne
National Laboratory,
IL 60439, USA.
298
TIG August 1999, volume 15, No. 8
thermal tolerance to the archaeal hyperthermophiles and
Aquifex by vertical inheritance. Consistent with this, we
have identified presumptive homologs of at least a third of
these genes in the incomplete genome of another deeply
diverging bacterial hyperthermophile, Thermotoga
maritima (see below).
We also find methodological problems in the analysis
of Aravind et al.1 The 246 Aquifex proteins reported as
‘reliable best hits’ with their archaeal homologs were
defined by having an E-value (expected number of
matches at least this good in random data) of at least 100
times lower than that obtained with any bacterial or
eukaryotic protein, which is not a particularly stringent
criterion. In reality there is no simple relationship between
differences in expectation and being significantly more
related. Nor is there any translation of this measure into
relative phylogenetic distances (amino acid replacements
per position), or even into a difference in percentage
amino acid identity. Because they did not compare all
pairs of sequences within a family, these data are not even
sufficient for a cluster analysis, but the authors have
drawn conclusions about the histories of genes (phylogenetic analyses). For statements about the histories of
these genes, it would be more appropriate to use explicit
phylogenetic analysis, supported by bootstrap analysis of
confidence. Of the 60 protein families (27% of the 220
families that go beyond Archaea and Aquifex) for which
Aravind et al.1 report such analyses, they find bootstrap
support for an Aquifex–Archaea grouping in only 26
families. Thus, only 43% of the cases they examined
(12% of these 220 ‘reliable best hits’ with the Archaea) are
actually demonstrated to support their suggestion.
Finally, because of our own interest in the set of proteins uniquely shared between Aquifex and Archaea, we
repeated this analysis comparing our results with those of
Aravind et al.1 Although there were a number of differences in the genes identified, the real importance of this
analysis lies in the fact that the majority of these genes are
found in only one or two of the four complete archaeal
genomes. Thus, even if these genes have been horizontally
transferred, we cannot possibly infer whether the transfer
occurred from Archaea to Aquifex (as the authors suggested) or vice versa. In addition, we identified homologs
of at least a third of these genes in the partial genome of
Thermotoga maritima, another bacterial hyperthermophile, suggesting that vertical inheritance via a thermophilic lineage from the archaeal–bacterial common ancestor (Fig. 1) will be a more parsimonious explanation
than independent lateral transfers as suggested by
Aravind et al.1
0168-9525/99/$ – see front matter © 1999 Elsevier Science All rights reserved. PII: S0168-9525(99)01811-9
GENOME ANALYSIS
Archaeal and bacterial hyperthermophiles
FIGURE 1. A rooted phylogenetic tree of Bacteria, Eukarya and Archaea
Escherichia
Agrobacterium
Planctomyces
Flavobacterium
Chlamydia
Leptonema
Synechocystis
Bacillus
Thermomicrobium
Thermus
Thermotoga
Aquifex
Outlook
In summary, we find the ideas
expressed by Aravind et al.1 to be very
interesting, but we also argue that these
authors have made assumptions (without offering justification) that led them
to conclusions that do not follow from
the data per se. In particular, alternative
hypotheses on the history of extreme
thermophilicity would suggest (regardless of the rooting of the tree) that substantial numbers of the genes discussed
could be vertically inherited from the
cenancestor.
Giardia
Tritrichomonas
Physarum
Entamoeba
Dictyostelium
Trypanosoma
Paramecium
Zea
Coprinus
Homo
Desulfurococcus
Sulfolobus
Pyrococcus
Thermoproteus
Thermophilum
Methanopyrus
Methanobacterium formicicum
Methanothermus
Thermococcus
Methanococcus vannielii
0.10
Methanococcus jannaschii
Archaeoglobus
Thermoplasma
Haloferax
Methanospirillum
This Maximum Likelihood tree was inferred essentially as described in Ref. 5, and rooted as in Ref. 9.
The heavy lines trace the evolution of extreme thermophilicity under the assumption that it originated
only once. Scale bar: 0.10 amino acid substitutions per site.
References
1 Aravind, L. et al. (1998) Evidence for massive gene
exchange between archaeal and bacterial
hyperthermophiles. Trends Genet. 14, 442–444
2 Deckert, G. et al. (1998) The complete genome of the
hyperthermophilic bacterium Aquifex aeolicus. Nature
392, 353–358
3 Médigue, C. et al. (1991) Evidence for horizontal gene
transfer in Escherichia coli speciation. J. Mol. Biol.
222, 851–856
4 Woese, C.R. (1998) The universal ancestor. Proc. Natl.
Acad. Sci. U. S. A. 95, 6854–6859
5 Burggraf, S. et al. (1992) A phylogenetic analysis of
Aquifex pyrophilus. Syst. Appl. Microbiol. 15, 352–356
6 Stetter, K.O. (1996) Hyperthermophilic prokaryotes.
FEMS Microbiol. Rev. 18, 149–158
7 Pace, N.R. (1991) Origin of life-facing up to the
physical setting. Cell 65, 531–533
8 Galtier, N. et al. (1999) A nonhyperthermophilic common
ancestor to extant life forms. Science 283, 220–221
9 Iwabe, N. et al. (1989) Evolutionary relationship of
archaebacteria, eubacteria and eukaryotes inferred
from phylogenetic trees of duplicated genes. Proc.
Natl. Acad. Sci. U. S. A. 86, 9355–9359
10 Brown, J.R. and Doolittle, W.F. (1995) Root of the
universal tree of life based on ancient aminoacyl-tRNA
synthetase gene duplications. Proc. Natl. Acad. Sci.
U. S. A. 92, 2441–2445
11 Forterre, P. (1995) Thermoreduction, a hypothesis for
the origin of prokaryotes. C. R. Acad. Sci. 318,
415–422
12 Forterre, P. (1996) A hot topic: the origin of
hyperthermophiles. Cell 85, 789–792
Reply
e welcome the discussion of the evolutionary mechanism(s) underlying the special relationship between
archaeal and bacterial hyperthermophiles. First of all, to
our satisfaction, we find that Kyrpides and Olsen1 agree
with us on the critically important issue: such a special
relationship does exist – something that was not at all
obvious from the original publication on the Aquifex
genome sequence2. In fact, this was the principal point
that we tried to convey, as convincingly as we could, in the
article that is discussed3. Perhaps we should have been
more explicit about distinguishing between these basic
observations and their evolutionary interpretation, which
necessarily remains hypothetical. In that sense, it might be
prudent to accept the criticism.
However, we still believe that the hypothesis we preferred, namely massive horizontal gene exchange, is a better
explanation for what is observed than the alternative
hypothesis favored by Kyrpides and Olsen1, namely ances-
W
0168-9525/99/$ – see front matter © 1999 Elsevier Science All rights reserved. PII: S0168-9525(99)01786-2
tral origin of the archaeal genes in Aquifex. The main
reasons for this are simple and have little to do with the
details of the phylogenetic methods used by us, or others,
but rather stem directly from the nature of the special
relationship. We discuss these reasons briefly below.
With respect to the majority of its genes, Aquifex looks
like a ‘garden-variety’ bacterium and does not show any
specific affinity with the Archaea. A significant subset of
the Aquifex genes, however, appears to be very different in
that they show a much greater similarity to archaeal
orthologs than to bacterial ones, and some are (so far)
simply unique for Archaea and Aquifex (or, in several
cases, shared with other thermophilic Bacteria). Is this the
pattern of sequence conservation one would expect under
the ‘conservation of ancestral features due to common
lifestyle’ hypothesis favored by Kyrpides and Olsen1?
Hardly so. The simplest form of this hypothesis would
suggest that all (or perhaps most, allowing for some
TIG August 1999, volume 15, No. 8
L. Aravind
aravind@ncbi.nlm.nih.gov
Roman L. Tatusov
tatusov@
ncbi.nlm.nih.gov
Yuri I. Wolf
wolf@ncbi.nlm.nih.gov
D. Roland Walker
walker@
ncbi.nlm.nih.gov
Eugene V. Koonin
koonin@ncbi.nlm.nih.gov
National Center for
Biotechnology Information,
National Library of Medicine,
National Institutes of Health,
Bethesda, MD 20894, USA.
299