SUPPLEMENT ARTICLE

The Genome of Salmonella enterica Serovar Typhi

Stephen Baker and Gordon Dougan
The Wellcome Trust Sanger Institute, Hinxton, Cambridge, United Kingdom

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The generation of complete genome sequences provides a blueprint that facilitates the genetic characterization
of pathogens and their hosts. The genome of Salmonella enterica serovar Typhi (S. Typhi) harbors ∼5 million
base pairs encoding some 4000 genes, of which 1200 are functionally inactive. Comparison of S. Typhi isolates
from around the world indicates that they are highly related (clonal) and that they emerged from a single
point of origin ∼30,000–50,000 years ago. Evidence suggests that, as well as undergoing gene degradation, S.
Typhi has also recently acquired genes, such as those encoding the Vi antigen, by horizontal transfer events.

The species Salmonella enterica is made up of patho-
genic bacteria that have the ability to infect a wide range
of animals and cause a variety of disease syndromes.
Since the early days of microbiology, Salmonella has
fascinated researchers and clinicians alike, in part be-
cause of the antigenic diversity within the genus, leading
to the assignment of isolates to 12500 different serovars.
Serologic typing has dominated the classification of Sal-
monella, and the methodologies have proved useful in
both epidemiologic control and the clinical treatment
of infections. Taxonomists have struggled to define Sal-
monella as a species group. We now work with the
definition of the S. enterica species divided into several
subspecies and many serovars. Salmonella bongori has
been classified as a distinct species. The new science of
genomics and the ability to sequence the entire genome
of individual bacteria will allow us to improve our un-
derstanding of the organization and evolution of the
species S. enterica and allows us to look in detail at
specialized members, such as S. enterica serovar Typhi
(S. Typhi) [1, 2]. We should also recognize that species
such as S. enterica are still evolving, and we are currently
observing only a small snapshot as it evolves through
time. Examination of the organization of genomes
should allow us to gain a better understanding of the
Reprints or correspondence: Dr. Gordon Dougan, The Wellcome Trust Sanger
Institute, The Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SA,
UK (gd1@sanger.ac.uk).
Clinical Infectious Diseases 2007; 45:S29–33
 2007 by the Infectious Diseases Society of America. All rights reserved.
1058-4838/2007/4502S1-0008$15.00
DOI: 10.1086/518143
mechanisms by which species evolve and may also help
us predict the type of organisms that might appear in
the future.
BASIC FEATURES OF THE SALMONELLAE
Salmonella is a genus within the Enterobacteriacae and,
as such, falls into the general group of enteric bacteria
that includes Escherichia coli and Shigella species. The
basic life cycle of these microorganisms involves, al-
lowing for a few exceptions, colonization of the lumen
of the intestine of animals and transmission, via the
external environment, between hosts. Most members
of the Enterobacteriaceae are commensal microorgan-
isms that exist as components of the normal intestinal
microflora, although the pathogenic potential of some
is enhanced. A comparison of the genomes of several
sequenced enteric bacteria immediately highlights some
important common traits. All have a single chromo-
some, normally 4.3–5.0 Mb in size [1, 3–8]. Different
strains may also harbor extrachromosomal DNA in the
form of plasmids. Plasmids often carry genes associated
with virulence or antibiotic resistance and can be con-
sidered to be a rapidly evolving gene pool. Comparison
of the chromosomes of different enteric bacteria iden-
tifies a common set of so called “core genes” that are,
in general, shared among enteric species [6, 9]. These
core genes can be regarded as genes that perform
“household” functions associated with the common
shared lifestyle of intestinal colonization and transmis-
sion (environmental survival). The full definition of the
core gene set is difficult, but the shared genome can be
identified by comparing DNA sequences and examining

The S. Typhi Genome • CID 2007:45 (Suppl 1) • S29

shared gene function. Such core genes may play a role in central
metabolism or polysaccharide biosynthesis or encode common
structural proteins. However, perhaps a more interesting feature
is that the core genome is mainly organized with these genes
aligned in the same conserved order along the single chro-
mosome, a characteristic referred to as “synteny.” Genomic
synteny can be highlighted using a simple genome comparison
tool, such as the one illustrated in figure 1. The core genome
may be compared to the “chassis” of a car, in that it is a
conserved design feature that works successfully and aids the
in the core genome of different enteric bacteria are compared,
E. coli and S. enterica are found to differ by ∼10%, and Sal-
monella serovars within S. enterica differ by ∼1%. This 10%
divergence between the core sequences of E. coli and S. enterica
most likely represents evolutionary drift over the ∼100 million
years since the 2 species separated from a common ancestor.
The genomes of enteric bacteria are under intensive selective
pressure because of factors that include competition within the
normal flora, coping with fluctuating nutrient sources in the
host and the environment, and pressure from the host immune

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lifestyle of the enteric bacteria. If the DNA sequences of genes system, to name but a few. Thus, we may expect the genomes

Figure 1. An Artemis comparison tool (ACT) alignment of the genomes of Salmonella enterica serovar Typhi (S. Typhi) CT18, S. enterica serovar
Typhimurium (S. Typhimurium) LT2, and Escherichia coli K12. This image was generated using the ACT [10]. ACT permits the alignment of DNA
sequences and demonstrates regions of homology (calculated by BLAST) with a red line or block between similar sequences in different genomes.
The image depicts the alignment of 3 linear genome sequences (beginning at the origin of replication). E. coli K12 is represented at the top of diagram,
S. Typhi CT18 in the middle, and S. Typhimurium LT2 at the base. The red lines represent regions of significant DNA homology between the chromosomes
of the 3 different organisms. The figure demonstrates a high level of sequence conservation between S. Typhi and E. coli and, to a greater extent,
between S. Typhi and S. Typhimurium. There are exceptions: “a” indicates regions of inversion between strains due to recombination of repeating
sequences, and “b” indicates obvious breaks in the sequence homology due to an insertion of DNA sequence in one strain but not the other; these
are most noticeable when comparing S. Typhi and E. coli.

S30 • CID 2007:45 (Suppl 1) • Baker and Dougan

of enteric bacteria to show signatures of these evolutionary
pressures, and this appears to be the case. Scattered along the
core genome are blocks of genes—or, in some cases, single genes
or gene remnants—that have limited or no homology with the
core genome. Furthermore, these genes can be unrelated or
show significant divergence between different enteric species or
even between members of the same species. These novel genes
can, however, share some related features and common func-
tions. For example, they might work together to enhance the
virulence potential of a species. Such virulence-associated gene
combinations are often referred to as “pathogenicity islands”
[11]. Examples include Salmonella pathogenicity islands 1 and
2 (SPI-1 and SPI-2, respectively), which contribute to inva-
siveness and the ability of Salmonella organisms to survive in-
side eukaryotic cells. These gene clusters often have GC content
that differs from that of the core genome, suggesting that they
have been acquired more recently and independently by a hor-
izontal gene-transfer event. Indeed, these genes are frequently
associated with genes from plasmids and phage and are some-
times integrated into redundant transfer RNA genes that can
act as receptor sites for foreign DNA [11]. Prophages or phage
remnants are signature genes of the diverse, noncore regions
of the enteric bacterial genome. Bacteriophages have the ability
to mobilize DNA and appear to be one of critical drivers of
diversity in these microorganisms [12]. This is not necessarily
surprising, because we can imagine frequent and multiple in-
teractions between phage and bacteria in both the intestine and
the environment.
FEATURES OF THE GENOME OF S. TYPHI
Genome sequencing allows us to look in some detail at the
genomes of individual species, subspecies, serovars, and even
different isolates within the same serovar [1, 2, 6–9]. Thus, we
can examine the genetic blueprint of such bacteria and make
simple comparisons with the genomes of bacteria that share
certain phenotypic characteristics (such as host restriction or
biotype). We may also compare the genome of S. Typhi with
the genomes of other bacteria that are limited in their path-
ogenic potential for particular hosts (i.e., host promiscuity or
restriction). S. Typhi is a particular Salmonella serovar and is
recognized as the cause of typhoid fever [13, 14]. S. Typhi
isolates frequently share common antigenic determinants (e.g.,
O9, Hd, and Vi), and, indeed, serologic analysis is routinely
used in the confirmation of the identity of clinical isolates. S.
Typhi causes an invasive form of salmonellosis in humans that
is characterized by a prolonged incubation period, fever, and
systemic bacterial dissemination, which can be detected by iso-
lation of S. Typhi from the blood and bone marrow. Two key
phenotypic signatures of S. Typhi include the ability to express
the Vi polysaccharide antigen at the surface of the bacteria and
host restriction to humans. S. Typhi can infect some higher
primates but is poorly infectious in mice and other mammals
[15]. The human host restriction phenotype of S. Typhi has
inhibited direct studies of the pathogenicity of this serovar.
Most of our understanding of the pathogenicity of S. Typhi
has been gleaned from comparative studies with more pro-
miscuous S. enterica serovars, such as S. enterica serovar Ty-
phimurium (S. Typhimurium), in the mouse. At the time of
writing, the complete DNA sequences of 2 different S. Typhi
isolates are available in public databases [1, 2]. These genome
sequences are of the classic S. Typhi type strain Ty2 (originally
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isolated in the early 1900s), frequently used in laboratory stud-
ies around the world, and S. Typhi CT18, a multidrug-resistant
isolate collected in Vietnam in 1992. S. Typhi CT18 harbors 2
large plasmids, one of which encodes multidrug resistance, that
are not found in Ty2.
A comparison of the chromosomes of S. Typhi CT18 and
Ty2 demonstrates a remarkable degree of conservation. Almost
all of the genes are shared between the 2 isolates. Some dif-
ferences include an additional cluster of a few genes in Ty2 that
might be a novel pathogenicity island and a P4-like phage
determinant (N. R. Thomson, unpublished data). However, the
overriding feature is conservation of sequence. This comparison
immediately begs the question of how diverse the different S.
Typhi isolated around the world are. One simple method to
gauge bacterial diversity is to use multilocus sequence typing
(MLST), an approach that compares the sequences of portions
of 7 household genes from the chromosomal DNA of different
isolates [16]. MLST can be used in some species, such as S.
enterica itself, to classify isolates into related MLST types or
clusters [17]. Interestingly, MLST sometimes clusters isolates
of a particular serovar into a single cluster, but this is not always
the case, and some serovars may be polyphyletic, following
different evolutionary lineages. However, MLST and other
forms of analysis of S. Typhi isolates suggest a remarkable de-
gree of conservation, with a very limited number of highly
related MLST patterns [18–20]. These data in themselves (but
also taken together with information gleaned from other, in-
dependent approaches) suggest that S. Typhi belongs to a single
clonal type that has evolved from the same progenitor. Indeed,
examination of the DNA sequence and the rate of change of
single-nucleotide polymorphisms suggest that S. Typhi may be
as young as 30,000 years old [18], indicating that its divergence
occurred significantly later than the estimated divergence of S.
enterica and E. coli. Thus, S. Typhi has had a limited time frame
in which to accumulate diversity. The fact that S. Typhi isolates
are so highly related means that it is often difficult to differ-
entiate between isolates in a local area and even more difficult
to define evolutionary lineages within the population. Genomic
methodologies can be applied to help. Relatively simple meth-
ods, such as PFGE, have proved valuable to distinguish between
isolates [21]. This method has been helped by the unusual
The S. Typhi Genome • CID 2007:45 (Suppl 1) • S31
characteristic of S. Typhi to undergo recombination, between
different ribosomal RNA operons, that rearranges blocks of the
genome in a distinct manner [22]. However, PFGE has limited
potential for identifying evolutionary lineages at a global or
local level. We await the development of new single-nucleotide
polymorphism–based genomewide typing methods to address
this need.
One of the main characteristics of most S. Typhi isolates is
the ability to express the Vi polysaccharide capsule. Vi capsule
production is associated with a set of genes that are situated
within a novel gene island that has been designated as SPI-7
[23, 24]. This island is 134 kb in length and encodes a variety
of putative virulence-associated gene clusters, including the Vi
locus, a phage encoding the sopE effector protein of SPI-1, a
type IV pilus, and a putative type IV secretion system. SPI-7
has many typical features that are associated with horizontally
acquired DNA, and the structure is indicative of several in-
dependent integration events. There is also some evidence that
SPI-7 may be able to act in a fashion similar to that of a
conjugative transposon. Indeed, an SPI-7–like element that en-
codes Vi is present in some S. enterica serovar Paratyphi C and
some S. enterica serovar Dublin isolates. The fact that the Vi
genes are encoded on a mobile and potentially unstable genetic
region is of interest because the Vi antigen is the target of a
promising Vi-based typhoid vaccine. Surveillance methods will
have to be introduced to ensure that no capsule replacement
events occur as a result of selection driven by such Vi-based
mass-vaccination campaigns [25].
WHY IS S. TYPHI INVASIVE AND HOST
RESTRICTED?
The question of what the genetic basis of the invasive nature
and the human host–restricted phenotype of S. Typhi is has
fascinated researchers for many years. S. Typhi yielded few clues
prior to genome sequencing. However, examination of the S.
Typhi genome and comparison with other serovars, such as S.
enterica serovar Paratyphi A, that also have the ability to cause
invasive typhoid-like disease in humans has provided some
intriguing clues. The S. Typhi CT18 genome was the first to
be completely determined, and annotation of the gene reper-
toire of this isolate indicated that 1200 of the genes present in
the genome showed evidence of being disrupted or inactivated.
We refer to such genes as “pseudogenes.” Pseudogenes have
been described in human DNA and some fastidious bacteria
but were not expected to be present in such high numbers in
a free-living bacteria such as S. Typhi [1]. Comparison with
the genome of S. Typhimurium LT2 (the first publicly available
S. Typhimurium genome) showed that the majority of the genes
appeared to be intact and most likely fully functional in this
serovar [5]. Hence, for some reason, S. Typhi had accumulated
mutations that inactivated ∼5% of its gene repertoire.
S32 • CID 2007:45 (Suppl 1) • Baker and Dougan
Further analysis indicated that different S. Typhi isolates har-
bor an almost identical complement of pseudogenes with es-
sentially the same mutations, again supporting the hypothesis
that S. Typhi had evolved once, relatively recently, and had
spread in the human population from that one source. Inter-
estingly, if tables are drawn linking related genes, some inter-
esting gene types are found to have been inactivated in S. Typhi.
Seven of the 12 fimbrial systems are inactivated, along with
other genes that encode putative fimbrial-like genes [26]. Genes
such as shdA and ratB that have been associated with intestinal
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persistence in S. Typhimurium have been inactivated [27]. Sev-
eral genes associated with known pathogenicity islands such as
SPI-1, SPI-2, SPI-3, SPI-4, and SPI-5 have also become inac-
tivated. This suggests that S. Typhi has lost the ability to express
several virulence-associated traits, a factor that could begin to
explain the loss of host range of this serovar.
Interestingly, a number of the genes that are present as pseu-
dogenes in S. Typhi are also pseudogenes in S. enterica serovar
Paratyphi A, and some of these are inactivated by identical
mutations, suggesting a common evolutionary origin. Loss of
function of genes associated with intestinal attachment and
persistence would fit into a lifestyle associated with invasion of
systemic tissue and transmission by excretion via the gall blad-
der rather than luminal gut colonization. This trait may be
apparent from data provided by human challenge studies (both
virulent and live vaccine formulations), in which S. Typhi ap-
pears to be shed at a lower level after oral challenge, compared
with less-invasive serovars, such as S. Typhimurium [14, 28].
Although this evidence is, on the whole, circumstantial, it does
allow for the formulation of testable hypotheses by which to
address these questions. More recently, studies of Vi antigen
[2] have indicated that Vi may have immunomodulatory ac-
tivities that limit the ability of neutrophils to be recruited to
the intestine during the early phases of S. Typhi infection, lim-
iting diarrhea but promoting the invasive potential of the col-
onization process [29]. Thus, a combination of acquisition and
loss of virulence-associated traits may contribute to the path-
ogenic potential of S. Typhi.
CONCLUSIONS
The sequencing of the genomes of S. Typhi isolates and the
ability to compare these genetic blueprints with those of related
microorganisms has provided new insights into how this path-
ogen has evolved to cause invasive disease in humans. Pro-
ceeding from these leads, we can expect new tools to emerge
that will improve our abilities to interrogate the structure and
evolution of S. Typhi, and these developments are likely to be
followed by improved diagnostic tools. We might also expect
more efforts to be made to understand the basic pathogenic
mechanisms employed by this pathogen in the human host.
Acknowledgments
Financial support. Wellcome Trust.
Supplement sponsorship. This article was published as part of a sup-
plement entitled “Tribute to Ted Woodward,” sponsored by an unrestricted
grant from Cubist Pharmaceuticals and a donation from John G. McCor-
mick of McCormick & Company, Hunt Valley, Maryland.
Potential conflicts of interest. S.B. and G.D.: no conflicts.
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