Unusual genotype of a Uropathogenic Escherichia coli strain assigned to the B2 phylogenetic group.
ABSTRACT Extraintestinal pathogenic Escherichia coli (ExPEC) cause infections such as urinary tract infections, septicemia and meningitis.…
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ABSTRACT: The characterization of population structures plays a main role for understanding outbreaks and the dynamics of bacterial spreading. In Escherichia coli, the widely used combination of multiplex-PCR scheme together with goeBURST has some limitations. The purpose of this study is to show that the combination of different phylogenetic approaches based on concatenated sequences of MLST genes results in a more precise assignment of E. coli phylogenetic groups, complete understanding of population structure and reconstruction of ancestral clones. A collection of 80 Escherichia coli strains of different origins was analyzed following the Clermont and Doumith's multiplex-PCR schemes. Doumith's multiplex-PCR showed only 1.7% of misassignment, whereas Clermont's-2000 protocol reached 14.0%, although the discrepancies reached 30% and 38.7% respectively when recombinant C, F and E phylogroups were considered. Therefore, correct phylogroup attribution is highly variable and depends on the clonal composition of the sample. As far as population structure of these E. coli strains, including 48 E. coli genomes from GenBank, goeBURST provides a quite dispersed population structure; whereas NeighborNet approach reveals a complex population structure. MLST-based eBURST can infer different founder genotypes, for instance ST23/ST88 could be detected as the founder genotypes for STC23; however, phylogenetic reconstructions might suggest ST410 as the ancestor clone and several evolutionary trajectories with different founders. To improve our routine understanding of E. coli molecular epidemiology, we propose a strategy based on three successive steps; first, to discriminate three main groups A/B1/C, D/F/E and B2 following Doumith's protocol; second, visualization of population structure based on MLST genes according to goeBURST, using NeighborNet to establish more complex relationships among STs; and third, to perform, a cost-free characterization of evolutionary trajectories in variants emerging along the clonal expansion using parsimony methods of phylogenetic analysis.PLoS ONE 01/2014; 9(8):e105395. · 3.53 Impact Factor
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ABSTRACT: Using data from whole-genome projects, an updated multiplex PCR strategy was developed to assign Escherichia coli isolates rapidly to major phylogenetic groups. This assay accommodates sequence variations detected within target sequences, thereby increasing sensitivity and reliability. It was validated using 185 isolates of known sequence types and showed improved congruence with multilocus sequence typing data.Journal of clinical microbiology 07/2012; 50(9):3108-10. · 4.16 Impact Factor
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ABSTRACT: The four major phylogenetic groups of Escherichia coli (A, B1, B2, D) can be determined using a multiplex PCR targeting three DNA fragments (ChuA, YjaA, TspE4.C2) published by Clermont et al. (AEM 2000, 66:4555). Isolates are assigned to one of the phylogroups based on the presence or absence of these three fragments, i.e. B1 isolates are defined by the absence of ChuA and the presence of TspE4.C2. Recently, Mendonca et al. (JCM 2011, 49:3105) proposed that an extraintestinal pathogenic E. coli positive for YjaA and TspE4.C2 was a new pathogenic genotype that should be assigned to phylogroup B2. We investigated the occurrence of this genotype among 1355 E. coli collected in Denmark from animals, meat, healthy humans, and patients. A total of four isolates were identified from pig, broiler chicken meat and patients. Investigations of multilocus sequence types, virulence genes and in vivo virulence indicated pathogenicity. Although B2 isolates are often pathogenic, some remain non-pathogenic. Until further investigations of larger collections of this genotype we do not find the evidence compelling to assign the YjaA/TspE4.C2 genotype to the B2 phylogroup.Journal of Medical Microbiology 05/2013; · 2.30 Impact Factor
JOURNAL OF CLINICAL MICROBIOLOGY, Aug. 2011, p. 3105–3106
Copyright © 2011, American Society for Microbiology. All Rights Reserved.
Vol. 49, No. 8
Unusual Genotype of a Uropathogenic Escherichia coli Strain
Assigned to the B2 Phylogenetic Group?
strains cause infections such as urinary tract infections,
septicemia, and meningitis. Knowledge of the phylogeny of
the strains contributes to the recognition of their virulence
potential and clinical outcome. ExPEC virulence factors are
often clustered in pathogenicity-associated islands (PAIs)
(2). Pathogenic ExPEC strains belong to the B2 and, to a
lesser extent, D phylogenetic groups, whereas commensal
isolates are assigned to groups A and B1. The triplex PCR
method is often used to assess phylogenetic groups (1). The
accuracy with which this method assigns strains to their
correct multilocus sequence typing (MLST)-based phylo-
group is good (4).
As part of a project investigating antimicrobial resistance,
virulence, and phylogeny, we used this method (1) to deter-
mine the phylogroup of E. coli isolates collected at a university
hospital. E. coli strain HUC270 was isolated in 2007 from the
urine of a 52-year-old man hospitalized in a medical ward. The
antimicrobial susceptibility testing was performed with the Vi-
tek 2 Advanced Expert System (AES) (bioMe ´rieux, France).
Eight PAIs were screened by a multiplex PCR (6). The ampli-
fication products were sequenced (Macrogen). MLST was used
to determine the sequence type (7).
E. coli HUC270 was resistant only to ampicillin and ceph-
alothin, being susceptible to amoxicillin-clavulanic acid, pip-
eracillin-tazobactam, cefotaxime, ceftazidime, levofloxacin,
ciprofloxacin, gentamicin, trimethoprim-sulfamethoxazole,
and nitrofurantoin. An inverted relationship between the
presence of virulence factors and susceptibility to quino-
lones has been reported in uropathogenic E. coli, which may
suggest a possible association of E. coli HUC270 with a
virulent phylogenetic group like B2 (5). The majority of
uropathogenic strains belonging to the B2 phylogroup con-
tain three or more PAIs (6). Indeed, E. coli HUC270 en-
coded different virulence determinants in PAIs: in PAI
III536, S-fimbriae and an iron siderophore system; in PAI
IV536, the yersiniabactin siderophore system; and in
PAI IICFT073, P-fimbriae and iron-regulated genes. How-
ever, it was not possible to cluster the strain in the major
phylogenetic groups or subgroups (1, 3). The chuA gene was
not amplified, but the yjaA and DNA fragment tspE4.C2
genes were present in the genome (Fig. 1). The result was
reproducible and consistent for six colonies submitted to
triplex PCR to avoid the hypothesis of a mix of strains
belonging to the A0or A1(3) and B1 phylogroups. Accord-
ing to the MLST scheme (MLST databases at the Environ-
mental Research Institute [ERI], University College Cork),
E. coli HUC270 revealed a new sequence type (ST2084),
which was related to the B2 reference E. coli strain (ST127)
by five housekeeping genes (adk, gyrB, icd, mdh, and recA).
These findings suggested that this unusual genotype profile
may be included in phylogroup B2 and not considered as a
“hybrid” strain like E. coli reference strain 70 (ECOR 70), in
which some housekeeping genes exhibit nucleotide se-
quences shared by group A ECOR strains and some show
sequences from B1 ECOR strains (1).
To our knowledge, the combination of yjaA and DNA
fragment tspE4.C2 genes reported here has not been de-
scribed previously. A study correlating phylogroup with
MSLT results for 600 E. coli strains did not report this
phylogenetic profile (4), suggesting that it is uncommon.
Also, it could not be assigned to one of the seven groups
defined by Escobar-Pa ´ramo et al. (3). On the basis of the
results of PAI determination, MLST profile, and suscepti-
bility to quinolones, we suggest that E. coli strains showing
this combination of genes by the triplex PCR method should
be assigned to the major phylogenetic group B2 and consid-
ered potentially pathogenic.
N. Mendonc ¸a is supported by grant SFRH/BPD/45815/2008 from
Fundac ¸a ˜o para a Cie ˆncia e a Tecnologia, Lisboa, Portugal. This
work was supported financially by the ESCMID 2010 Research
Grant from the European Society of Clinical Microbiology and
Infectious Diseases and by the Center for Pharmaceutical Studies,
University of Coimbra.
The authors of this letter declare they have no conflicts of inter-
1. Clermont, O., S. Bonacorsi, and E. Bingen. 2000. Rapid and simple determi-
nation of the Escherichia coli phylogenetic group. Appl. Environ. Microbiol.
2. Diard, M., et al. 2010. Pathogenicity-associated islands in extraintestinal
pathogenic Escherichia coli are fitness elements involved in intestinal coloni-
zation. J. Bacteriol. 192:4885–4893.
3. Escobar-Pa ´ramo, P., et al. 2004. Large-scale population structure of hu-
man commensal Escherichia coli isolates. Appl. Environ. Microbiol. 70:
4. Gordon, D. M., O. Clermont, H. Tolley, and E. Denamur. 2008. Assigning
Escherichia coli strains to phylogenetic groups: multi-locus sequence typing
versus the PCR triplex method. Environ. Microbiol. 10:2484–2496.
5. Moreno, E., et al. 2006. Quinolone, fluoroquinolone and trimethoprim/sulfa-
methoxazole resistance in relation to virulence determinants and phylogenetic
background among uropathogenic Escherichia coli. J. Antimicrob. Che-
6. Sabate ´, M., E. Moreno, T. Pe ´rez, A. Andreu, and G. Prats. 2006. Pathogenicity
FIG. 1. Triplex PCR profiles specific for E. coli phylogenetic
groups. Lanes 1 and 2, group A; lane 3, group B1; lanes 4 and 5, group
D; lanes 6 and 7, group B2; lane 8, E. coli HUC270 profile. Lane M
contains a 100-bp marker.
island markers in commensal and uropathogenic Escherichia coli isolates.
Clin. Microbiol. Infect. 12:880–886.
7. Wirth, T., et al. 2006. Sex and virulence in Escherichia coli: an evolutionary
perspective. Mol. Microbiol. 60:1136–1151.
Nuno Mendonc ¸a
Center of Pharmaceutical Sciences
Faculty of Pharmacy
University of Coimbra
College of Pharmacy
University of Michigan
Ann Arbor, Michigan
Grac ¸a Ribeiro
Clinical Pathology Service
University Hospitals of Coimbra
Gabriela Jorge Da Silva*
Laboratory of Microbiology
Center of Pharmaceutical Sciences
Faculty of Pharmacy
University of Coimbra
Health Sciences Campus
Azinhaga de Santa Comba
3000-548 Coimbra, Portugal
*Phone: 351 239488460
?Published ahead of print on 1 June 2011.
3106 LETTERS TO THE EDITORJ. CLIN. MICROBIOL.