Identification of genes subject to positive selection in uropathogenic strains of Escherichia coli: a comparative genomics approach

Abstract

Escherichia coli is a model laboratory bacterium, a species that is widely distributed in the environment, as well as a mutualist and pathogen in its human hosts. As such, E. coli represents an attractive organism to study how environment impacts microbial genome structure and function. Uropathogenic E. coli (UPEC) must adapt to life in several microbial communities in the human body, and has a complex life cycle in the bladder when it causes acute or recurrent urinary tract infection (UTI). Several studies designed to identify virulence factors have focused on genes that are uniquely represented in UPEC strains, whereas the role of genes that are common to all E. coli has received much less attention. Here we describe the complete 5,065,741-bp genome sequence of a UPEC strain recovered from a patient with an acute bladder infection and compare it with six other finished E. coli genome sequences. We searched 3,470 ortholog sets for genes that are under positive selection only in UPEC strains. Our maximum likelihood-based analysis yielded 29 genes involved in various aspects of cell surface structure, DNA metabolism, nutrient acquisition, and UTI. These results were validated by resequencing a subset of the 29 genes in a panel of 50 urinary, periurethral, and rectal E. coli isolates from patients with UTI. These studies outline a computational approach that may be broadly applicable for studying strain-specific adaptation and pathogenesis in other bacteria.

Fig. 1.

Overview of the analysis and specification of foreground branches. (A) Analysis scheme. Size of each dataset (boxes) is indicated in parentheses. Programs used are indicated next to the arrows. See text for more details about how datasets were generated. (B and C) Hypothetical phylogenetic tree to indicate branch specification. UPEC strains are boxed. Evidence for positive selection was evaluated in specific lineages (termed foreground branches). B shows the sets of UPEC-specific foreground branches used: (i) UTI89 only (green), (ii) CFT073 only (red), (iii) UPEC leaves only (green + red), (iv) common UPEC branch only (cyan), (v) all UPEC (green + red + cyan). C shows that when the common UPEC branch was not present due to the tree topology, only three sets of UPEC foreground branches were used (green, red, and green + red).

Fig. 2.

amiA, fepE, and ompC are under selection in clinical UPEC isolates. Phylogenetic trees of unique sequences for each gene are shown. Red branches/labels indicate foreground branches that show evidence for positive selection (see Table 2 and text). Red numbers to the right of the tree indicate the number of urine and fecal isolates represented by the red (foreground) labels in the phylogenetic tree. Black numbers to the right of the tree indicate the number of urine and fecal isolates represented by black labels in the phylogenetic tree. The sites of isolation of strains represented by sequence labels are summarized in Table 4. Scale bar for phylogenetic trees is shown at the bottom right.

Fig. 3.

UPEC-selected genes are enriched for genes in two COG functional categories. COG category codes are indicated on the y axis. The fraction of genes in each COG category is shown on the x axis. Black bars indicate genes under positive selection in UPEC strains (n = 29). Open bars are for all genes annotated in UTI89 (n = 5,066). COG categories that are significantly enriched (P < 0.05, binomial test) in the set of UPEC-selected genes relative to all UTI89 genes are indicated by an asterisk. COG category codes are as follows: U, intracellular trafficking and secretion; G, carbohydrate transport and metabolism; I, lipid transport and metabolism; R, general function prediction only; D, cell cycle control, mitosis and meiosis; H, coenzyme transport and metabolism; B, chromatin structure and dynamics; P, inorganic ion transport and metabolism; W, extracellular structures; O, posttranslational modification, protein turnover, chaperones; J, translation; A, RNA processing and modification; L, replication, recombination and repair; C, energy production and conversion; M, cell wall/membrane biogenesis; Q, secondary metabolites biosynthesis, transport and catabolism; Z, cytoskeleton; V, defense mechanisms; Y, nuclear structure; E, amino acid transport and metabolism; K, transcription; N, cell motility; T, signal transduction mechanisms; F, nucleotide transport and metabolism; and S, function unknown.