Plasmid-mediated Quinolone Resistance in Salmonella enterica, United Kingdom

To the Editor: Fluoroquinolones are broad-spectrum antimicrobial drugs used to treat many clinical infections. Salmonellosis is treated with fluoroquinolones only in elderly or immunocompromised patients, but these drugs are also used for treating patients with enteric fever, invasive disease, or long-term salmonellae carriage. High-level fluoroquinolone resistance is uncommon, but reduced susceptibility is increasing.

Since 1998, plasmid-mediated quinolone resistance encoded by qnr genes A, B, and S that confer low-level resistance to nalidixic acid and reduced susceptibility to ciprofloxacin has been identified in several enterobacterial species, including Salmonella. Their clinical importance is in facilitating resistance to potentially lethal levels of quinolone. Additionally, qnr genes are often associated with strains that produce extended-spectrum β-lactamases.

We recently reported identification of qnr genes in Salmonella in the United Kingdom (1). Most isolates were associated with the Far East. Two isolates of S. Virchow were part of an outbreak associated with imported cooked chicken from Thailand. During October 2006–April 2007, we monitored qnr genes in nontyphoidal salmonellae isolated in the United Kingdom that expressed reduced susceptibility to ciprofloxacin (MIC 0.125–1.0 μg/mL) with concomitant susceptibility to nalidixic acid (MIC <16 μg/mL). This resistance phenotype is a useful marker for the qnr gene as the sole quinolone resistance determinant (1).

Recent studies showed that isolates of Salmonella spp. and Escherichia coli with decreased susceptibility to ciprofloxacin (MICs >0.06 μg/mL and 0.5 μg/mL, respectively), but with susceptibility or intermediate resistance to nalidixic acid (MIC 8–16 μg/mL and 4–8 μg/mL, respectively), all had qnrA or qnrS genes but lacked mutations in the topoisomerase genes (2,3). Strains with ciprofloxacin MICs >1 μg/mL were also included to monitor involvement of qnr genes in development of high-level ciprofloxacin resistance. Breakpoint concentrations used are based on long-term studies within the Health Protection Agency Laboratory of Enteric Pathogens. Ciprofloxacin Etest (AB Biodisk, Solna, Sweden) results were interpreted according to manufacturer’s procedures. A total of 45 Salmonella spp. strains were tested. Screening for qnr genes by multiplex PCR identified 37 isolates with qnrS and 2 carrying qnrB variants (Table) (4). However, the qnrB primer pair in this multiplex did not fully match all qnrB gene variants. PCR and sequencing using primers FQ1 and FQ2 (5) and qnrS-F and qnrS-R (1), were used to identify specific qnrB and qnrS gene variants.

Table. Isolates of Salmonella enterica with plasmid-mediated qnr genes, United Kingdom, October 2006–April 2007.

Salmonella serotype	Phage type*	No. isolates	VNTR profile†	Ciprofloxacin MIC (μg/mL)‡	Additional resistance to antimicrobial drugs§	qnr identified
Corvallis	–	1	–	0.25	S, Su, T	qnrS1
Corvallis	–	2	–	0.38	S, Su, T	qnrS1
Corvallis	–	1	–	1.0	S, Su, T ,Cf	qnrS1
Corvallis	–	1	–	0.25	None	qnrS1
Corvallis	–	1	–	0.38	None	qnrS1
Schwarzengrund	–	1	–	0.25	T	qnrB5
Typhimurium	DT120	4	1–6-0–0-3	0.38	S, Su, T	qnrS1
Typhimurium	DT120	3	1–6-0–0-3	0.50	S, Su, T	qnrS1
Typhimurium	DT120	3	1–4-0–0-3	0.38	S, Su, T	qnrS1
Typhimurium	DT120	1	1–4-0–0-3	0.50	S, Su, T	qnrS1
Typhimurium	DT120	1	1–4-0–0-3	0.38	None	qnrS1
Typhimurium	DT120	1	1–5-0–0-3	0.38	S, Su, T	qnrS1
Typhimurium	DT193	1	1–6-0–0-3	0.50	S, Su, T	qnrS1
Typhimurium	DT193	1	1–4-0–0-3	0.38	C, S, Su, Sp, T, Tm	qnrS1
Typhimurium	DT193	1	1–4-0–0-3	0.38	S, Su, T	qnrS1
Typhimurium	DT193	2	1–5-0–0-3	0.38	S, Su, T	qnrS1
Typhimurium	DT193	1	1–4-0–0-3	0.50	A, Su	qnrS1
Typhimurium	49b	1	1–4-19–1-3	0.25	A, G, Ne, K, S, Su, Sp, T, Tm, Ak, Cx, Cr, Cf, Cn, Ct	qnrB2
Typhimurium	NC	1	1–4-0–0-3	0.25	S, Su, T	qnrS1
Typhimurium	UT	1	3–8-19–1-2	>32	A, C, G, S, Su, Sp, T, Tm, Fu, Nx	qnrS1
Virchow	43	5	–	1.0	A, Fu, Nx	qnrS1
Virchow	43	2	–	1.5	A, Fu, Nx	qnrS1
Virchow	25a	1	–	0.75	Tm	qnrS1
Virchow	11	1	–	1.0	A, Fu, Nx	qnrS1
Virchow	NC	1	–	1.5	A, C, G, Ne, K, S, Su, Sp, T, Tm, Fu, Nx, Cx, Cr, Cf, Cn, Ct	qnrS1

*DT, definitive type; NC, does not conform to a recognized pattern; UT, untypeable.
†VNTR, variable number tandem repeat. Loci of the VNTR profiles are presented in the following order: STTR9-STTR5-STTR6-STTR10pl-STTR3. The number 0 in the VNTR profile represents cases with no amplification of PCR product.
‡Determined by Etest.
§Antimicrobial drugs (breakpoint final concentrations): S, streptomycin (16 mg/L); Su, sulfonamide (64 mg/L); T, tetracycline (8 mg/L); Cf, cefuroxime (16 mg/L); C, chloramphenicol (8 mg/L); Sp, spectinomycin (64 mg/L); Tm, trimethoprim (2 mg/L); A, ampicillin (8 mg/L]; G, gentamicin (4 mg/L); Ne, neomycin (8 mg/L) K, kanamycin (8 mg/L); Ak, amikacin (4 mg/L); Cx, cefalexin (16 mg/L); Cr, cefradine (16 mg/L); Cn, ceftriaxone (1 mg/L); Ct, cefotaxime (1 mg/L); Fu, furazolidone (8 mg/L); Nx, nalidixic acid (16 mg/L).

The qnrS1-positive salmonellae belong to serotypes Typhimurium (21 isolates), Virchow (10), and Corvallis (6). Most S. Typhimurium isolates were either definitive phage type 120 or 193, and most S. Virchow isolates were phage type 43 (Table). Thirteen qnrS1-positive isolates were from patients who reported recent travel to Egypt, India, Malaysia, Morocco, Thailand, or an undisclosed destination.

Twelve isolates from patients who had not traveled abroad were assumed to be from UK-acquired infections. S. Virchow isolates had been associated with cooked chicken from Thailand (1), and qnrS1 has recently been described in S. Corvallis strains from humans in Denmark or isolated in Thailand from humans, chicken, pork, and beef (3). Comparison of pulsed-field gel electrophoresis patterns and resistance phenotypes of qnrS1-positive S. Corvallis strains identified common types, suggesting that some UK patients may have acquired S. Corvallis from chicken from Thailand.

Thirteen isolates showed resistance to ceftriaxone, cefotaxime, or ampicillin. Plasmids with qnr genes have been found to co-transfer TEM, SHV, and CTX-M genes (1,5,6). Co-transmission of fluoroquinolone and β-lactamase resistance is clinically important because co-selection of resistance by use of either drug may occur.

Twenty-one qnrS1-positive S. Typhimurium were subtyped by variable number tandem repeat (VNTR) analysis to determine whether the increase was caused by spread of >1 distinct strains (7). Twenty isolates produced 1 of 3 related profiles (loci of VNTR profiles are ordered STTR9-STTR5-STTR6-STTR10pl-STTR3): 1–4-0–0-3, 9 isolates; 1–5-0–0-3, 3 isolates; or 1–6-0–0-3, 8 isolates. Alleles 4 and 5, and 5 and 6 at locus STTR5 only differed by an extra 6-bp repeat, which suggests a clonal relationship between the qnrS1-positive S. Typhimurium in this study (Table) (8). S. Typhimurium isolates with the 1–6-0–0-3 profile have been isolated from tourists returning from Asia (7), which suggests that the UK qnrS1-positive S. Typhimurium isolates have originated in the Far East.

These findings show increased occurrence of qnr genes, particularly qnrS1, in nontyphoidal salmonellae in the United Kingdom. These data are in contrast to those of recent studies in the United States and France, which show low incidences of qnrS genes in larger strain collections (9,10). The qnr phenotype is in contrast to resistance mediated by mutations in the topoisomerase genes whereby 1 mutation confers low-level resistance to fluoroquinolones and full resistance to nalidixic acid. Our previous study demonstrated that qnrS1 was sufficient to cause decreased susceptibility to ciprofloxacin in the absence of mutations in gyrA (1). In this study, a qnr gene was sufficient to increase the ciprofloxacin MIC to 0.38–0.75 μg/mL. In addition, a qnr gene contributed to high-level ciprofloxacin resistance in 10 isolates, thereby potentially jeopardizing first-line treatment of vulnerable patient groups with ciprofloxacin.