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Antimicrobial Agents and Chemotherapy, November 1999, p. 2657-2662, Vol. 43, No. 11
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Insertion of Mini-IS605 and Deletion of
Adjacent Sequences in the Nitroreductase (rdxA) Gene Cause
Metronidazole Resistance in Helicobacter pylori
NCTC11637
Yvette J.
Debets-Ossenkopp,1
Raymond G. J.
Pot,1
David J.
van
Westerloo,1
Avery
Goodwin,2
Christina M. J. E.
Vandenbroucke-Grauls,1
Douglas E.
Berg,3
Paul S.
Hoffman,2 and
Johannes
G.
Kusters1,*
Department of Medical Microbiology and
Infection Control, Faculty of Medicine, Vrije Universiteit, Amsterdam,
The Netherlands1; Department of
Molecular Microbiology, Washington University, St. Louis,
Missouri3; and Department of
Microbiology and Immunology, Faculty of Medicine, Dalhousie University,
Halifax, Canada2
Received 19 January 1999/Returned for modification 19 April
1999/Accepted 27 August 1999
 |
ABSTRACT |
We found that NCTC11637, the type strain of Helicobacter
pylori, the causative agent of peptic ulcer disease and an early risk factor for gastric cancer, is metronidazole resistant. DNA transformation, PCR-based restriction analysis, and DNA sequencing collectively showed that the metronidazole resistance of this strain
was due to mutation in rdxA (gene HP0954 in the full genome sequence of H. pylori 26695) and that resistance did not
depend on mutation in any of the other genes that had previously been suggested: catalase (katA), ferredoxin (fdx),
flavodoxin (fldA), pyruvate:flavodoxin oxidoreductase
(por
), RecA (recA), or superoxide
dismutase (sodB). This is in accord with another recent study that attributed metronidazole resistance to point mutations in
rdxA. However, the mechanism of rdxA
inactivation that we found in NCTC11637 is itself also novel: insertion
of mini-IS605, one of the endogenous transposable elements
of H. pylori, and deletion of adjacent DNA sequences
including 462 bp of the 851-bp-long rdxA gene.
| |
INTRODUCTION |
Helicobacter pylori is a
microaerophilic gram-negative bacterium that chronically infects the
gastric mucosa of more than half of all persons worldwide and is a
major cause of chronic active gastritis and peptic ulcer disease
(23) and an early risk factor for gastric cancer
(17). Antimicrobial therapy that leads to eradication of
infection and ulcer healing is often achieved with metronidazole, a
nitroheterocyclic compound
[1-(2-hydroxyethyl)-2-methyl-5-nitroimidazole], in combination with
other drugs (30). Resistance to metronidazole is common,
however, and is the major factor in many cases of failure to eradicate
H. pylori and cure the disease (6, 20, 24).
For metronidazole to kill H. pylori, it must be taken up by
an energized membrane (22) and then reduced to form a toxic metabolite. Several mechanisms of metronidazole resistance have been
proposed in recent years, including enhanced scavenging of toxic oxygen
radicals by an altered catalase or superoxide dismutase (7, 11,
26), a more efficient DNA damage repair mechanism (9,
27), and loss of function of a critical reductase (15, 16). Recent studies of fresh clinical isolates by several of us
(12) indicated that metronidazole resistance often results from point mutation in rdxA (HP0954, in the fully sequenced
genome of strain 26695 [28]), a gene that encodes an
oxygen-insensitive NADPH nitroreductase. However, some researchers have
questioned the generality of this interpretation (13, 21).
Here we report that NCTC11637, the type strain of H. pylori
that Barry Marshall had isolated in Australia and that we obtained from
the American Type Culture Collection, displays a high level of
resistance to metronidazole and that this resistance is attributable to
mutational inactivation of rdxA. Our experiments further
suggest that mutation in other genes, previously invoked to explain the metronidazole resistance phenomenon, does not contribute to the high-level metronidazole resistance of NCTC11637 (katA
[catalase], fdx [ferredoxin], fldA
[flavodoxin], por [pyruvate:flavodoxin oxidoreductase], recA [RecA], and sodB
[superoxide dismutase]).
| |
MATERIALS AND METHODS |
Bacterial strains and growth conditions.
H. pylori
strains used in this study were NCTC11637 (MIC of >256 µg/ml)
(obtained from the American Type Culture Collection as ATCC 43504),
which was found here to be metronidazole resistant (Mtzr)
(3), and the unrelated, metronidazole-susceptible
(Mtzs) strain NCTC11638 (MIC of <0.032 µg/ml). Bacteria
were routinely grown on Dent plates (Columbia agar plates supplemented
with 7% lysed horse blood and H. pylori selective
supplement [Oxoid, Basingstoke, United Kingdom]). Plates were
incubated at 37°C in a microaerophilic atmosphere of 5%
O2-10% CO2-85% N2.
Determination of MIC.
MICs were determined with the Etest as
described before (10).
Natural transformation.
Metronidazole-resistant NCTC11638
transformants were created by natural transformation of NCTC11638
(Mtzs) with DNA from the NCTC11637 (Mtzr)
strain, essentially as described by Wang et al. (31). As a control, bacteria were transformed with TE (10 mM Tris-HCl [pH 8.0],
1 mM EDTA) instead of DNA. Transformants (selected on Dent plates
containing 32 µg of metronidazole per ml) were generated at a
frequency of 10
5 per viable cell. From one such
experiment, 12 individual colonies were selected at random, and the
metronidazole MICs were determined.
PCR.
Unless noted otherwise, standard techniques were used
for all DNA manipulations (5). Primers (Table
1) for PCR amplification were based on
the full genome sequence of strain 26695 (28) plus other
entries in the public database. PCR was performed in an automated
thermal cycler (Mastercycler 5330; Eppendorf, Hamburg, Germany), in a
final volume of 100 µl by using the Primezym DNA polymerase kit
(Biometra, Göttingen, Germany), with approximately 2.5 ng of
template genomic DNA and 100 pmol of each primer. Amplification conditions are listed in Table 1. The size of the PCR products was
evaluated by electrophoresis on a 1.5% agarose-0.5× Tris-borate-EDTA gel containing 0.5 µg of ethidium bromide per ml.
Cloning and sequencing of the PCR products.
PCR products
were ligated into the pGEM-T vector (Promega, Madison, Wis.) according
to the manufacturer's recommendations. The ligation products were then
transformed into Escherichia coli ER1793 (18),
and transformants were selected on Luria-Bertani plates supplemented
with 100 µg of ampicillin per ml. Plasmid DNA was isolated from
transformants with the QIAprep spin plasmid (Qiagen, Hilden, Germany).
These plasmids were then used as templates for DNA sequencing with the
Thermo-Sequenase premixed cycle sequencing kit (Amersham, Little
Chalfont, Buckinghamshire, United Kingdom) with the M13 forward and
reverse primers. Sequencing was performed on an Amersham Vistra 725 DNA
sequencer, and data were analyzed with Lasergene software (DNASTAR,
Madison, Wis.).
Restriction fragment length polymorphism (RFLP).
Approximately 300 ng of PCR product was digested with restriction
endonucleases (New England Biolabs, Beverly, Mass.). The endonucleases
used for RFLP analysis of the various genes are indicated in Table 1.
The restriction enzyme digestion profile was subsequently analyzed by
electrophoresis on a 1.5% agarose gel.
| |
RESULTS |
Metronidazole-resistant transformant derivatives of NCTC11638.
The type strain, NCTC11637, is resistant to metronidazole (MIC of >256
µg/ml). We used a DNA transformation and PCR product characterization
strategy to determine the basis of its resistance. This was based on
findings that Mtzr determinants can be transferred to
Mtzs strains by transformation (31) and that the
differences in distributions of restriction sites in a given gene from
unrelated strains usually allow those strains to be distinguished
(1). Mtzr (32 µg/ml) transformant derivatives
of NCTC11638 were obtained at a frequency of approximately
105 per recipient cell with NCTC11637 genomic DNA, in
contrast to a frequency of <107 per recipient cell in
DNA-free mock-transformation controls. Each of the 12 transformants
tested, although selected at 32 µg/ml, was resistant to more than 256 µg/ml, as was its Mtzr NCTC11637 parent.
RFLP analysis of the selected genes.
A number of different
genes (fdx [ferredoxin], fldA [flavodoxin],
por [pyruvate:flavodoxin oxidoreductase],
recA [RecA], katA [catalase], and
sodB [superoxide dismutase]) had been postulated by
various other groups to contribute to high-level
Mtzr. The possible involvement of any of them was tested by
scoring whether the Mtzr transformants contained
NCTC11637-type or NCTC11638-type alleles at these loci in an RFLP
test. First, primers for amplification of these genes were designed
(Table 1), based on the sequences in the public databases. PCR products
of the expected size were obtained from the NCTC11637 donor DNA, from
the NCTC11638 recipient, and from each transformant.
To maximize the power of RFLP analysis and to get a better sense of the
sequence divergence of these two much-studied strains,
we PCR amplified
and sequenced each of the candidate genes and
thereby identified
restriction sites that differed usefully between
the parental strains.
The PCR product corresponding to each candidate
gene from each of the
12 Mtz
r transformants was then analyzed by restriction with
appropriate
enzymes (Table
1).
In each case, except for
rdxA (see below), the results
showed unambiguously that the Mtz
r transformants contained
alleles of the Mtz
s NCTC11638 recipient at each of these
loci. This is exemplified
by the
Sau3AI and
MseI
RFLP patterns of
sodB shown in Fig.
1.
Thus, the ability of NCTC11637 genomic
DNA to transform NCTC11638
to Mtz
r did not depend on
acquisition of special donor alleles of any
of these previously
suggested candidate genes.
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FIG. 1.
Ethidium bromide-stained agarose gel displaying the
restriction endonuclease digestion pattern of the sodB
amplicon. (A) Digestion with Sau3AI; (B) digestion with
MseI. Lane 1, H. pylori NCTC11638
(Mtzs); lane 2, H. pylori NCTC11637
(Mtzr); lanes 3 to 14, H. pylori NCTC11638
transformants (Mtzr). The positions of the molecular size
markers (1-kb ladder; Stratagene) are indicated on the left.
|
|
In contrast to these negative results, PCR amplification from NCTC11637
rdxA, the gene whose inactivation had been implicated
recently in the Mtz
r of fresh clinical isolates, yielded a
product of 650 bp, 200
bp smaller than expected from the published
sequences and as observed
with NCTC11638 (Fig.
2). Each of the 12 Mtz
r
transformants of NCTC11638 also yielded a product of only 650
bp with
the
rdxA-specific primers. This result indicated that
the
Mtz
r of NCTC11637 was due to mutation in
rdxA.
Independent confirmation
for the involvement of
rdxA in the
Mtz
r phenotype of strain NCTC11637 was obtained by
transformation
of wild-type NCTC11638 with the aberrant 650-bp PCR
product obtained
from NCTC11637. This resulted in high numbers of
Mtz
r transformants, while no such transformants were
observed in the
control transformation, where we used the 850-bp
rdxA PCR fragment
obtained from strain NCTC11638.
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FIG. 2.
Ethidium bromide-stained agarose gel of the
rdxA amplicon. Lane 1, H. pylori NCTC11638
(Mtzs); lane 2, H. pylori NCTC11637
(Mtzr); lanes 3 to 14, H. pylori NCTC11638
transformants (Mtzr). The positions of the molecular size
markers (1-kb ladder; Stratagene) are indicated on the left.
|
|
Sequence analysis of the rdxA genes.
The basis of
the unexpected shortness of the NCTC11637 rdxA locus was
determined by sequencing. In the donor strain and also in the
transformants analyzed, we found an insertion of a novel 260-bp segment
just before the start of HP0953 (the gene just downstream of
rdxA) and a deletion of a 462-bp segment including 177 bp
from the 3' end of rdxA (Fig.
3). A BLAST search showed that 41 and 33 bp at the left and right ends, respectively, of this 260-bp element are
closely matched to the corresponding left and right ends of the 2-kb
insertion sequence (IS) IS650, whereas its central 186 bp
were unrelated to sequences in full-length IS605
(2). Rather, this segment corresponded to the
mini-IS605 (Fig. 4) that was
found at the right end of the cag pathogenicity island in
one strain (NCTC11638, where it is referred to as is605, in
contrast to IS605) (8). The genome sequence of
the fully sequenced strain 26695 contains eight such
mini-IS650 elements (Fig. 4), but none is in its
cag pathogenicity island (28). Just after we
submitted the manuscript, the complete genomic DNA sequence of H. pylori J99 was published (4), and this sequence reveals
the presence of five partial IS605 elements (Fig. 4).
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FIG. 3.
Alignment of the DNA sequences and the predicted encoded
proteins of the rdxA amplicons. Mtzs, H. pylori NCTC11638; Mtzr, H. pylori NCTC11637
and the metronidazole-resistant transformants of NCTC11638. The
sequence of the donor NCTC11637 amplicon is identical to that of all
the metronidazole-resistant NCTC11638 transformants. The primers used
to amplify the rdxA region (and amino acids derived
therefrom) are indicated in boldface; the inverted repeats of the
IS605 (7) are in boldface and underlined. The
amino acids directly above the DNA sequence correspond to H. pylori NCTC11638; the ones directly below the DNA sequence
correspond to NCTC11637 and the transformants. Gaps in the DNA sequence
are marked with dashes, and identical nucleotides are indicated by
dots.
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FIG. 4.
Alignment of the mini-IS605 sequence present
in the rdxA gene from strain NCTC11637 (see Fig. 3); the
cag pathogenicity island (cag PAI) of strain
NCTC11638 (7), GenBank accession no. U60176 bp 22476 to
22735; one of the eight partial copies of IS605 present in
strain 26695 (27), GenBank accession no. HPAE000537 bp 2867 to 2606; and one of the five partial copies present in strain J99
(4), GenBank accession no. HPAE000537 bp 2633 to 2891. The
inverted repeats of IS605 (7) are indicated in
boldface and underlined. For definitions of the dots and dashes, please
see the legend to Fig. 3.
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|
| |
DISCUSSION |
The results of our study of the type strain NCTC11637 confirm the
conclusions of Goodwin et al. (12) that inactivation of rdxA is responsible for much or all of the high-level
metronidazole resistance so often seen with H. pylori
clinical isolates. There was no evidence that mutation in
sodB, katA, ferredoxin, flavodoxin, por, or recA genes made any
contribution to the high-level metronidazole resistance of NCTC11637.
Although the PCR products for the above loci did not always cover the
complete gene, it is generally assumed (as was confirmed by our
analysis of the rdxA gene sequences) that natural
transformation results in the exchange of a large fragment of DNA.
Hence, not finding any indications for gene exchange in the central
portion of these six genes in 12 independent Mtz-resistant
transformants provides strong evidence against the involvement of these
genes and the loci adjacent to them. However, our data do not exclude
the possibility that, for the low-level resistance that is observed
with some H. pylori strains (13), these genes may
still play a role. Only determination of enzyme activities can provide
the definitive proof for the relative contribution of each gene in the
Mtzr of an individual strain.
All Mtzr mutations in rdxA that Goodwin et al.
had observed were small point mutations (base substitutions), whereas
the loss of RdxA function in NCTC11637 was caused by a deletion of much of the rdxA gene, associated with insertion of
mini-IS605. This is a novel transposable element that does
not contain any significant open reading frame and that accordingly, we
conjecture, may depend on another, larger element such as full-length
(2 kb) IS605 for its movement. Associated with
mini-IS605 in this strain was a deletion that removed 462 bp
of rdxA. Several different IS elements that are not related
to IS605 or mini-IS605 had been shown in other
species to generate deletions adjacent to their sites of insertion
(14, 25, 29). In two cases at least (IS1 and
IS50) (25, 32), deletion is independent of
recA function, and we propose that an equivalent IS
element-mediated adjacent-deletion process may have operated here. In
one scenario, mini-IS605 was inserted between
rdxA and the adjacent HP0953 gene, leaving a strain that was
still metronidazole susceptible but highly prone to mutate to
Mtzr by adjacent deletion. An alternative possibility of
insertion and concomitant deletion has not been ruled out.
This is the first report of a transposon-based spontaneous mutation
associated with antibiotic resistance in H. pylori. However, some one-third of strains from Europe and the Americas carry
full-length IS605, a similar fraction carry the distantly
related IS606 element, and more than half of the strains
seem to carry mini-IS605 (whether full-length
IS605 is present or not) (19). Endogenous IS
elements are well known in other species as agents of spontaneous
mutation and genome rearrangements and evolution. We propose that they may function similarly in H. pylori, perhaps often
contributing to drug resistance in the face of mounting
antibiotic usage and other adaptive changes of considerable
evolutionary significance.
| |
ACKNOWLEDGMENTS |
This work was supported by the Netherlands Digestive Disease
Foundation, the Medical Research Council of Canada grant R-14292 and
Astra Canada, and the National Institutes of Health (DK48029, AI38166,
and HG00820).
We thank A. B. Brinkman for his technical support.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Department of
Medical Microbiology and Infection Control, Vrije Universiteit
Amsterdam, Van der Boechorststraat 7, 1081 BT Amsterdam, The
Netherlands. Phone: 31-20-4448319. Fax: 31-20-4448318. E-mail:
JG.Kusters.mm{at}med.vu.nl.
| |
REFERENCES |
| 1.
|
Akopyants, N.,
N. O. Bukanov,
T. U. Westblom, and D. E. Berg.
1992.
PCR-based RFLP analysis of DNA sequence diversity in the gastric pathogen Helicobacter pylori.
Nucleic Acids Res.
20:6221-6225[Abstract/Free Full Text].
|
| 2.
|
Akopyants, N. S.,
S. W. Clifton,
D. Kersulyte,
J. E. Crabtree,
B. E. Youree,
C. A. Reece,
N. O. Bukanov,
E. S. Drazek,
B. A. Roe, and D. E. Berg.
1998.
Analysis of the cag pathogenicity island of Helicobacter pylori.
Mol. Microbiol.
28:37-53[Medline].
|
| 3.
|
Akopyants, N. S.,
Q. Jiang,
D. E. Taylor, and D. E. Berg.
1997.
Corrected identity of isolates of Helicobacter pylori reference strain NCTC11637.
Helicobacter
2:48-52[Medline].
|
| 4.
|
Alm, R. A.,
L. S. Ling,
D. T. Moir,
B. L. King,
E. D. Brown,
P. C. Doig,
D. R. Smith,
B. Noonan,
B. C. Guild,
B. L. deJonge,
G. Carmel,
P. J. Tummino,
A. Caruso,
M. Uria-Nickelsen,
D. M. Mills,
C. Ives,
R. Gibson,
D. Merberg,
S. D. Mills,
Q. Jiang,
D. E. Taylor,
G. F. Vovis, and T. J. Trust.
1999.
Genomic-sequence comparison of two unrelated isolates of the human gastric pathogen Helicobacter pylori.
Nature
397:176-180[Medline].
|
| 5.
|
Ausubel, F. M.,
R. Brent,
R. E. Kingston,
D. D. Moore,
J. G. Seidman,
J. A. Smith, and K. S. Struhl.
1989.
Current protocols in molecular biology.
Greene Publishing and Wiley Interscience, New York, N.Y
|
| 6.
|
Buckley, M. J.,
H. X. Xia,
D. M. Hyde,
C. T. Keane, and C. A. Morain.
1997.
Metronidazole resistance reduces efficacy of triple therapy and leads to secondary clarithromycin resistance.
Dig. Dis. Sci.
42:2111-2115[Medline].
|
| 7.
|
Cederbrant, G.,
G. Kahlmeter, and A. Ljungh.
1992.
Proposed mechanism for metronidazole resistance in Helicobacter pylori.
J. Antimicrob. Chemother.
29:115-120[Abstract/Free Full Text].
|
| 8.
|
Censini, S.,
C. Lange,
Z. Xiang,
J. E. Crabtree,
P. Chiara,
M. Borodovsky,
R. Rappuoli, and A. Covacci.
1996.
Cag, a pathogenicity island of Helicobacter pylori, encodes type I-specific and disease associated virulence factors.
Proc. Natl. Acad. Sci. USA
93:14648-14653[Abstract/Free Full Text].
|
| 9.
|
Chang, K. C.,
S. W. Ho,
J. C. Yang, and J. T. Wang.
1997.
Isolation of a genetic locus associated with metronidazole resistance in Helicobacter pylori.
Biochem. Biophys. Res. Commun.
236:785-788[Medline].
|
| 10.
|
Debets-Ossenkopp, Y. J.,
A. B. Brinkman,
E. J. Kuipers,
C. M. J. E. Vandenbroucke-Grauls, and J. G. Kusters.
1998.
Explaining the bias in the 23S rRNA gene mutations associated with clarithromycin resistance in clinical isolates of Helicobacter pylori.
Antimicrob. Agents Chemother.
42:2749-2751[Abstract/Free Full Text].
|
| 11.
|
Edwards, D. I.
1993.
Nitroimidazole drugs: action and resistance mechanisms. I. Mechanisms of action.
J. Antimicrob. Chemother.
31:9-20[Free Full Text].
|
| 12.
|
Goodwin, A.,
D. Kersulyte,
G. Sisson,
S. J. O. Veldhuysen van Zanten,
D. E. Berg, and P. S. Hoffman.
1998.
Metronidazole resistance in Helicobacter pylori is due to null mutations in a gene (rdxA) that encodes an oxygen-insensitive NADPH nitroreductase.
Mol. Microbiol.
28:383-393[Medline].
|
| 13.
|
Graham, D. Y.
1998.
Antibiotic resistance in Helicobacter pylori: implications for therapy.
Gastroenterology
115:1272-1277[Medline].
|
| 14.
|
Habermann, P., and P. Starlinger.
1982.
Bidirectional deletions associated with IS4.
Mol. Gen. Genet.
185:216-222[Medline].
|
| 15.
|
Hoffman, P. S.,
A. Goodwin,
J. Johnsen,
K. Magee, and S. J. O. Veldhuyzen van Zanten.
1996.
Metabolic activities of metronidazole-sensitive and -resistant strains of Helicobacter pylori: repression of pyruvate oxidoreductase and expression of isocitrate lyase activity correlate with resistance.
J. Bacteriol.
178:4822-4829[Abstract/Free Full Text].
|
| 16.
|
Hughes, N. J.,
P. A. Chalk,
C. L. Clayton, and D. J. Kelly.
1995.
Identification of carboxylation enzymes and characterization of a novel four-subunit pyruvate:flavodoxin oxidoreductase from Helicobacter pylori.
J. Bacteriol.
177:3953-3959[Abstract/Free Full Text].
|
| 17.
|
IARC (EUROGAST Study Group).
1993.
An international association between Helicobacter pylori infection and gastric cancer.
Lancet
341:1359-1362[Medline].
|
| 18.
|
Kelleher, J. E., and E. A. Raleigh.
1991.
A novel activity in Escherichia coli K-12 that directs restriction of DNA modified at CG dinucleotide.
J. Bacteriol.
173:5220-5223[Abstract/Free Full Text].
|
| 19.
|
Kersulyte, D.,
N. S. Akopyants,
S. W. Clifton,
B. A. Roe, and D. E. Berg.
1998.
Novel sequence organization and insertion specificity of IS605 and IS606: chimaeric transposable elements of Helicobacter pylori.
Gene
223:175-186[Medline].
|
| 20.
|
Logan, R. P. H.,
P. A. Gummet,
J. J. Misiewicz,
Q. N. Karim,
M. M. Walker, and J. H. Baron.
1991.
One week eradication regimen for Helicobacter pylori.
Lancet
338:1249-1252[Medline].
|
| 21.
|
Mégraud, F.
1998.
Epidemiology and mechanism of antibiotic resistance in H. pylori.
Gastroenterology
115:1278-1282[Medline].
|
| 22.
|
Moore, R. A.,
B. Beckthold, and L. E. Bryan.
1995.
Metronidazole uptake in Helicobacter pylori.
Can. J. Microbiol.
41:746-749[Medline].
|
| 23.
|
NIH Consensus Conference.
1994.
H. pylori in peptic ulcer disease.
JAMA
272:65-69[Abstract/Free Full Text].
|
| 24.
|
Noach, L. A.,
W. L. Langenberg,
M. A. Bertola,
J. Dankert, and G. N. J. Tytgat.
1994.
Impact of metronidazole resistance on the eradication of Helicobacter pylori.
Scand. J. Infect. Dis.
26:321-327[Medline].
|
| 25.
|
Reif, H. J., and H. Saedler.
1975.
IS1 is involved in deletion formation in the gal region of E. coli K12.
Mol. Gen. Genet.
137:17-28[Medline].
|
| 26.
|
Smith, M. A., and D. I. Edwards.
1995.
Redox potential and oxygen concentration as factors in the susceptibility of Helicobacter pylori to nitroheterocyclic drugs.
J. Antimicrob. Chemother.
35:751-764[Abstract/Free Full Text].
|
| 27.
|
Thompson, S. A., and M. T. Blaser.
1995.
Isolation of the Helicobacter pylori recA gene and involvement of the recA region in resistance to low pH.
Infect. Immun.
63:2185-2193[Abstract].
|
| 28.
|
Tomb, J. F.,
O. White,
A. R. Kerlavage,
R. A. Clayton,
G. G. Sutton,
R. D. Fleischmann,
K. A. Ketchum,
H. P. Klenk,
S. Gill,
B. A. Dougherty,
K. Nelson,
J. Quackenbush,
L. Zhou,
E. F. Kirkness,
S. Peterson,
B. Loftus,
D. Richardson,
R. Dodson,
H. G. Khalak,
A. Glodek,
K. McKenney,
L. M. Fitzegerald,
N. Lee,
M. D. Adams, and J. C. Venter.
1997.
The complete genome sequence of the gastric pathogen Helicobacter pylori.
Nature
388:539-547[Medline].
|
| 29.
|
Tomcsanyi, T.,
C. M. Berg,
S. H. Phadnis, and D. E. Berg.
1990.
Intramolecular transposition by a synthetic IS50 (Tn5) derivative.
J. Bacteriol.
172:6348-6354[Abstract/Free Full Text].
|
| 30.
|
Walsh, J. H., and W. L. Peterson.
1995.
The treatment of Helicobacter pylori infection in the management of peptic ulcer disease.
N. Engl. J. Med.
333:984-991[Free Full Text].
|
| 31.
|
Wang, Y.,
K. P. Roos, and D. E. Taylor.
1993.
Transformation of Helicobacter pylori by chromosomal metronidazole resistance and by a plasmid with a selectable chloramphenicol resistance marker.
J. Gen. Microbiol.
139:2485-2493[Abstract/Free Full Text].
|
| 32.
|
York, D.,
K. Welch,
I. Y. Goryshin, and W. S. Reznikoff.
1998.
Simple and efficient generation in vitro of nested deletions and inversions: Tn5 intramolecular transposition.
Nucleic Acids Res.
26:1927-1933[Abstract/Free Full Text].
|
Antimicrobial Agents and Chemotherapy, November 1999, p. 2657-2662, Vol. 43, No. 11
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Copyright © 1999, American Society for Microbiology. All rights reserved.
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Abstract
Introduction
