# Ciprofloxacin-resistant Salmonella Kentucky in travelers

François-Xavier Weill, Sophie Bertrand, Françoise Guesnier, Sylvie

Baucheron-Monnier, Patrick A.D Grimont, Axel Cloeckaert

► To cite this version:

François-Xavier Weill, Sophie Bertrand, Françoise Guesnier, Sylvie Baucheron-Monnier, Patrick A.D Grimont, et al.. Ciprofloxacin-resistant Salmonella Kentucky in travelers. Emerging Infectious Diseases, 2006, 12 (10), pp.1611-1612. 10.3201/eid1210.060589. hal-02667389

HAL Id: hal-02667389

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Submitted on 31 May 2020

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cing of products. J Med Virol. 2001; 65:138-48.

9. Palacios G, Casas I, Cisterna D, Trallero G, Tenorio A, Freire C. Molecular epidemiology of echovirus 30: temporal circulation and substitution of single lineages. J Virol. 2002;76:4940-9.

10. Avellon A, Casas I, Trallero G, Perez C, Tenorio A, Palacios G. Molecular analysis of echovirus 13 isolates and aseptic meningitis, Spain. Emerg Infect Dis. 2003;9:934-41.

Address for correspondence: Ana Avellón, Spanish Enterovirus Reference Laboratory, National Center of Microbiology, Carlos III Institute of Health, Carretera de Majadahonda a Pozuelo, Km 2, Majadahonda 28220, Madrid, Spain; email: aavellon@isciii.es

Ciprofloxacin-
resistant
*Salmonella*
Kentucky in
Travelers

**To the Editor:** Ciprofloxacin is the treatment of choice of severe non-typhoidal *Salmonella* infections in adults. Resistance to ciprofloxacin has been found exceptionally in nontyphoidal *Salmonella enterica* isolates and only in serotypes Typhimurium, Choleraesuis, and Schwarzengrund (1-8). Such isolates have been collected from humans and animals in Europe, Asia, and North America.

We report the emergence of ciprofloxacin-resistant isolates of S. Kentucky since 2002 in French travelers returning from northeast and eastern Africa. From 2000 through 2005, 197 S. Kentucky isolates from humans (1 per patient) were serotyped, from 69,759 total S. enterica isolates serotyped at the French National Reference Centre for *Salmonella*. Antimicrobial drug sus-

ceptibility was determined for 186 isolates by the disk-diffusion method with 32 antimicrobial drugs, as previously described (9). Resistance to several drugs, amoxicillin (18%), gentamicin (16%), nalidixic acid (21%), sulfonamides (24%), and tetracycline (24%), has been observed from 2000 through 2005.

A total of 17 (9%) ciprofloxacin-
resistant S. Kentucky strains were iso-
lated. A resistant isolate that was unty-
pable by conventional serotyping
(rough) but that had a pulsed-field gel
electrophoresis (PFGE) profile assos-
ciated with serotype Kentucky, was
included in this study. Ciprofloxacin
MIC levels in these isolates, deter-
mined by standard agar doubling dilu-
tion as previously described (2), were
4–16 mg/L. The first ciprofloxacin-
resistant strain was isolated in
December 2002 from a French tourist
who had gastroenteritis during a Nile
cruise in Egypt. In 2004 and 2005, 17
ciprofloxacin-resistant isolates were
identified in unrelated adults who
lived in different cities of France at
different times of the year. The 16
patients we contacted acquired the
infection during or immediately after
travel to Egypt (10 patients), Kenya
and Tanzania (3), or Sudan (1). In 2
cases, gastroenteritis occurred 2
months after travel to Egypt. None of
the investigated cases were fatal or
life-threatening.

The 18 ciprofloxacin-resistant iso-
lates (17 serotype Kentucky and 1
rough) displayed various susceptibili-
ty patterns, from single resistance to
quinolones to multiple resistance (up
to 9 antimicrobial agents). To identify
mutations responsible for cipro-
floxacin resistance, the quinolone
resistance-determining regions
(QRDRs) of *gyrA*, *gyrB*, *parC*, and
*parE* were amplified by PCR and
sequenced as described previously
(3,9), except that different forward
primers for *gyrB* (5'-TTATCGACGC-
CGCGCGTGCGC-3') and *parE* (5'-
CGCGTAACTGCATCG-GGTTC-3')

were used. The 18 ciprofloxacin-
resistant isolates had different double
mutations in *gyrA* leading to amino
acid substitutions, Ser83Phe and
Asp87Gly (8 isolates), Ser83Phe and
Asp87Asn (5), and Ser83Phe and
Asp87Tyr (5), but had identical muta-
tions in *parC* (resulting in Ser80Ile).
An additional substitution was
observed in ParC, Thr57Ser. This sub-
stitution, however, did not appear to
be associated with quinolone resist-
ance because it was also identified in
nalidixic acid-susceptible isolates.
No isolates had substitutions in the
QRDRs of GyrB and ParE. All iso-
lates tested by PCR for the plasmid-
mediated quinolone resistance-con-
ferring gene *qnrA* (9) were negative.
In the presence of the efflux pump
inhibitor Phe-Arg-β-naphthylamide,
the MICs of ciprofloxacin were
reduced from 4-fold to 16-fold, which
suggests that an active efflux mecha-
nism was present (2). The involve-
ment of the AcrAB-TolC efflux sys-
tem was determined by measuring
AcrA expression with a method previ-
ously described (5). A moderate pro-
duction of AcrA (3- to 4-fold increase
when compared with the baseline pro-
duction of AcrA in reference strain
98K) was observed in all but 1
ciprofloxacin-resistant isolate. This
isolate overproduced (6-fold) AcrA,
which correlated with a higher
ciprofloxacin MIC (16 mg/L).

The 18 ciprofloxacin-resistant iso-
lates and 14 ciprofloxacin-susceptible
S. Kentucky isolates used for compar-
ison were genotyped by PFGE with
*Xba*I restriction and PulseNet's run-
ning conditions, as described previ-
ously (9). Each profile that differed by
≥1 clear band >50 kb was considered a
distinct profile. The 18 resistant iso-
lates displayed 9 profiles that differed
by 1 to 3 bands (Dice correlation coef-
ficient 55%) (Figure). Profile X1c was
predominant (7 [39%] of 18). The 6
pansusceptible isolates tested dis-
played 5 different patterns unrelated to
those of resistant isolates. Use of a
<figure><img src="image_2.png" /></figure>

Figure. Dendrogram generated by BioNumerics version 4.1 (Applied Maths, Sint-Martens-Latem, Belgium) showing the results of cluster analysis on the basis of *Xbal* pulsed-field gel electrophoresis (PFGE) of *Salmonella enterica* serotype Kentucky isolates. Similarity analysis was performed by using the Dice coefficient, and clustering was performed by the unweighted pair-group method with arithmetic means with an optimization parameter of 0.5% and a 0.5% band position tolerance. The different PFGE profiles, the phenotypes of resistance to quinolones (R-type), and corresponding number of isolates are indicated. CIP, ciprofloxacin; NAL, nalidixic acid.

second restriction enzyme, *SpeI*, for the resistant isolates of X1 cluster enhanced discrimination. No clear correlations between combined PFGE patterns, *gyrA* mutations, and probable country of infection were observed.

Since *S*. Kentucky is infrequently isolated from human, animal, or environmental sources in France, these isolates must have been acquired abroad. Unfortunately, no investigations have been thus far conducted to identify the source of the contamination in probable countries of infection. Poultry products may be of particular interest because poultry is the main animal reservoir of *S*. Kentucky. Another possible source in East Africa is pork because a recent report identified quinolone-resistant (ciprofloxacin MIC >0.125 mg/L) *S*. Kentucky isolates in slaughtered pigs in Ethiopia (10). After identifying the source of the contamination, appropriate control measures should be implemented in the affected countries to control the spread of these isolates.

## Acknowledgments

We thank all the corresponding laboratories of the French National Reference Center Salmonella network and C. Mouline for expert technical assistance.

François-Xavier Weill,*

Sophie Bertrand,†

Françoise Guesnier,*

Sylvie Baucheron,‡ Patrick A.D.

Grimont,* and Axel Cloeckaert‡

*Institut Pasteur, Paris, France; †Scientific Institute of Public Health, Brussels, Belgium; and ‡Institut National de la Recherche Agronomique, Nouzilly, France

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Address for correspondence: François-Xavier Weill, Centre National de Référence des *Salmonella*, Unité de Biodiversité des Bactéries Pathogènes Emergentes, Institut Pasteur; 28 Rue du Docteur Roux, 75724 Paris CEDEX 15, France; email: fxweill@pasteur.fr