Skip directly to site content Skip directly to page options Skip directly to A-Z link Skip directly to A-Z link Skip directly to A-Z link
Volume 20, Number 4—April 2014

Salmonella Subtypes with Increased MICs for Azithromycin in Travelers Returned to the Netherlands

Robert-Jan HassingComments to Author , Wil H.F. Goessens, Wilfrid van Pelt, Dik J. Mevius, Bruno H. Stricker, Nicky Molhoek, Annelies Verbon, and Perry J.J. van Genderen
Author affiliations: Erasmus Medical Centre, Rotterdam, the Netherlands (R.-J. Hassing, W.H.F. Goessens, B.H. Stricker, A. Verbon); Inspectorate of Health Care, The Hague, the Netherlands (R.-J. Hassing, B.H. Stricker); National Institute for Public Health and the Environment (RIVM), Bilthoven, the Netherlands (W. van Pelt); Utrecht University, Utrecht, the Netherlands (D.J. Mevius); Central Veterinary Institute of Wageningen UR, Lelystad, the Netherlands (D.J. Mevius); Travel Clinic Havenziekenhuis, Rotterdam (N. Molhoek, P.J.J. van Genderen); Institute for Tropical Diseases, Havenziekenhuis, Rotterdam (P.J.J. van Genderen)

Cite This Article


Antimicrobial susceptibility was analyzed for 354 typhoidal Salmonella isolates collected during 1999–2012 in the Netherlands. In 16.1% of all isolates and in 23.8% of all isolates that showed increased MICs for ciprofloxacin, the MIC for azithromycin was increased. This resistance may complicate empirical treatment of enteric fever.

Enteric fever caused by Salmonella enterica serotypes Typhi and Paratyphi A, B, and C is mainly a disease of the developing world, and it is occasionally diagnosed as an imported disease in countries where the disease is not endemic (1). Its empirical treatment has been hampered by resistance to ampicillin, chloramphenicol, and trimethoprim and by decreased ciprofloxacin susceptibility (MIC for ciprofloxacin 0.125–1.0 µg/mL), and ciprofloxacin resistance (MIC for ciprofloxacin >1.0 µg/mL) (2,3). As a consequence, third-generation cephalosporins are used as first-line drugs for intravenous treatment, and azithromycin is frequently used for empirical treatment of uncomplicated enteric fever.

Although no clinical breakpoints are available to define azithromycin susceptibility or resistance, several clinical studies have demonstrated good efficacy of azithromycin for the treatment of uncomplicated enteric fever in clinical and in vitro studies (410). Regarding MIC breakpoints, the European Committee on Antimicrobial Susceptibility Testing states that isolates with an MIC <16 µg/mL for azithromycin should be considered as wild-type organisms that are responsive to treatment (3). The Clinical and Laboratory Standards Institute ( does not provide clinical breakpoints for macrolides for the group of Enterobacteriaceae. In previous studies of typhoidal Salmonella isolates, MICs for azithromycin ranged from 4 µg/mL to 64 µg/mL (813).

The first clinical case for which treatment of illness caused by typhoidal Salmonella spp. with azithromycin (MIC 256 µg/mL) failed was reported during 2010, evidenced by testing a S. enterica Paratyphi A isolate from Pakistan (14). Further, in a study of isolates from blood samples collected in India during 2005–2008, MICs >16 µg/mL for azithromycin were observed in 34.7% (35/101 isolates) of the typhoidal Salmonella isolates; clinical non-response was reported in 19 of 36 patients treated with azithromycin (13). Whether this problem of increasing MICs for azithromycin is limited to India or is emerging globally is not clear. The objective of our study was to investigate azithromycin susceptibility and trends in antibacterial drug resistance over time in isolates collected during 1999–2012 in the Netherlands.

The Study

Enteric fever is a notifiable disease in the Netherlands. During January 1999–December 2012, a total of 354 isolates were submitted by microbiology laboratories to the Salmonella National and Community Reference Laboratory ( 177 (50%) Salmonella Typhi isolates, 98 (27.7%) S. enterica Paratyphi A isolates, 78 (22.0%) S. enterica Paratyphi B isolates, and 1 (0.3%) S. enterica Paratyphi C isolate. There was no statistically significant difference in sex among patients whose tests showed S. enterica Typhi isolates (56.3% male, 43.7% female, p = 0.18) and S. enterica Paratyphi isolates (51.6% male, 48.4% female, p = 0.18). Patients ranged in age from 0 to 92 years. The median ages, 28.4 and 29.5 years, respectively (p = 0.60), did not differ between patients in whose samples S. enterica Typhi and Paratyphi were isolated. Trends in cumulative 1-year incidence were determined by linear regression analysis; data were weighted on the number of isolates collected each year. The cumulative 1-year incidence of enteric fever was relatively stable during 1999–2012, with an average of 25 isolates (4–39/year) for that period (p = 0.42).

Figure 1

Thumbnail of MICs of azithromycin of 354 Salmonella enterica serotypes Typhi and Paratyphi A, B, and C isolates from samples collected from ill returned travelers in the Netherlands, 1999–2012. For wild type isolates, MIC&lt;16 μg/mL.

Figure 1. . MICs of azithromycin of 354 Salmonella enterica serotypes Typhi and Paratyphi A, B, and C isolates from samples collected from ill returned travelers in the Netherlands, 1999–2012. For wild type...

Figure 2

Thumbnail of MICs of azithromycin in relation to ciprofloxacin susceptibility of 354 Salmonella enterica serotypes Typhi and Paratyphi isolates. Increased MICs for azithromycin (MIC&gt;16 μg/mL) in isolates with decreased ciprofloxacin susceptibility or ciprofloxacin resistance (MIC&gt;0.125 μg/mL) versus ciprofloxacin-susceptible isolates (MIC&lt;0.125 μg/mL) (p = 0.004).

Figure 2. . MICs of azithromycin in relation to ciprofloxacin susceptibility of 354 Salmonella enterica serotypes Typhi and Paratyphi isolates. Increased MICs for azithromycin (MIC>16 μg/mL) in isolates with decreased ciprofloxacin susceptibility or...

All MICs were determined by using the broth microdilution method with Mueller-Hinton II cation–adjusted broth (Difco, Franklin Lakes, NJ, USA). We applied European Committee for Antimicrobial Susceptibility Testing guidelines for category interpretation for different antibacterial drugs (3). Azithromycin MICs were 2–256 µg/mL among the 354 isolates and were increased in 57 (16.1%) of the isolates (Figure 1). The distribution of azithromycin MICs of S. enterica Typhi and Paratyphi A and B peaked at 8, 16, and 8 µg/mL, respectively (Figure 1). Trend analysis showed no increased MIC over time for all isolates (p = 0.21) or for S. enterica Typhi (p = 0.35) or Paratyphi (p = 0.70). One Paratyphi A isolate, from a sample acquired in Malaysia in 2007, required an MIC of 256 µg/mL. Decreased susceptibility to ciprofloxacin was observed in 116 (32.8%) and ciprofloxacin resistance in 6 (1.7%) of the 354 isolates. Cumulative 1-year incidence of isolates with decreased susceptibility or resistance to ciprofloxacin increased significantly from 0% (0/12 isolates) in 1999 to 64.3% (18/28 isolates) in 2012 (p<0.001) (Technical Appendix Figure). Among isolates with decreased susceptibility or resistance to ciprofloxacin, 23.8% (29/122 isolates) showed an increased MIC for azithromycin; MIC increased for 12.1% (28/232 isolates) of the ciprofloxacin-susceptible isolates (p = 0.004) (Figure 2). No significant increase in amoxicillin, trimethoprim, or chloramphenicol resistance was observed (amoxicillin, p = 0.97; trimethoprim, p = 0.95; and chloramphenicol, p = 0.99) (Technical Appendix Figure). For all isolates, the MICs for erythromycin ranged from 64 to ≥512 µg/mL. Resistance to third-generation cephalosporins was not observed in the isolates.

The origins of isolates for which the country of acquisition was known (205 of 354 strains) were distributed among geographic regions by using the United Nations geoscheme ( Besides imported cases from countries in which enteric fever is highly endemic, such as India, Indonesia, and Pakistan, rates of importation were high for travelers from Turkey and Morocco (Table).

Percentages of elevated MICs for azithromycin were highest for isolates acquired in regions that had concurrent high proportions of isolates with decreased susceptibility or resistance to ciprofloxacin (Table). In isolates acquired in countries from Southern Asia, increased MICs for ciprofloxacin and increased MICs for azithromycin were observed in 21.4% (18/84 isolates) of the isolates.


We found high percentages of elevated azithromycin MICs in typhoidal Salmonella isolates collected in the Netherlands during 1999–2012. MICs >16 µg/mL for azithromycin were found in 16.1% of all isolates and in 23.8% of isolates with elevated MICs for ciprofloxacin. This observation may be explained by increased use of azithromycin in countries from which samples yielded high rates of typhoidal Salmonella isolates with decreased susceptibility or resistance to ciprofloxacin. Moreover, our findings are aligned with an alarming report from India on increasing MICs for azithromycin (13). Our study shows higher MICs than anticipated based on another Western case series (11), implying that these potentially resistant strains are not confined to India.

Besides treatment with third-generation cephalosporins, empirical treatment options may be scarce for patients with potential azithromycin-resistant Salmonella serotypes. Reuse of antibacterial drugs, such as ampicillin, chloramphenicol, or trimethoprim may be a valuable treatment option upon proven susceptibility, but widespread use of these antibacterial drugs as first-line treatment will likely result in rapid reemergence of multidrug resistance and associated drug-related adverse effects. Further, increasing the dose of ciprofloxacin or using alternative fluoroquinolones has been suggested as an effective treatment in some cases (15). This option will not be feasible for empirical treatment because it applies only in a minority of cases and may be associated with drug toxicity. The danger of losing azithromycin to antimicrobial resistance could be detrimental in countries faced with endemic or epidemic enteric fever and complicated by poverty; therefore, azithromycin should be used with care. The results of this study also implicate the importance of developing more effective vaccines as control measures for enteric fever. Future research is needed to evaluate clinically relevant breakpoints of azithromycin by analyzing the treatment outcome of azithromycin in relation to their MICs.

In conclusion, typhoidal Salmonella isolates in ill returned travelers from the Netherlands already show a high percentage of increased MICs for azithromycin. Because the highest proportions of increased MICs for azithromycin are found in isolates with increased MICs for ciprofloxacin and in regions where decreased susceptibility or resistance to ciprofloxacin is already widely prevalent among S. enterica Typhi and Paratyphi isolates, this resistance may further limit future treatment options for enteric fever.

Dr Hassing is a PhD candidate and an infectious diseases specialist at the Erasmus Medical Centre, Rotterdam, the Netherlands, and a pharmacovigilance inspector at the Health Care Inspectorate, The Hague, the Netherlands. His research interests include antimicrobial resistance in Enterobacteriaceae.



We thank Marian ten Kate and Aart van der Meijden for performing susceptibility testing and Kees Veldman for sending the isolates.



  1. Parry  CM. Typhoid fever. Curr Infect Dis Rep. 2004;6:2733. DOIPubMedGoogle Scholar
  2. Booker  BM, Smith  PF, Forrest  A, Bullock  J, Kelchlin  P, Bhavnani  SM, Application of an in vitro infection model and simulation for reevaluation of fluoroquinolone breakpoints for Salmonella enterica serotype typhi. Antimicrob Agents Chemother. 2005;49:177581. DOIPubMedGoogle Scholar
  3. Leclercq  R, Cantón  R, Brown  DF, Giske  CG, Heisiq  P, MacGowan  AP, EUCAST expert rules in antimicrobial susceptibility testing. Clin Microbiol Infect. 2013;19:14160. DOIPubMedGoogle Scholar
  4. Gordillo  ME, Singh  KV, Murray  BE. In vitro activity of azithromycin against bacterial enteric pathogens. Antimicrob Agents Chemother. 1993;37:12035. DOIPubMedGoogle Scholar
  5. Metchock  B. In-vitro activity of azithromycin compared with other macrolides and oral antibiotics against Salmonella typhi. J Antimicrob Chemother. 1990;25 Suppl A:29–31.
  6. Butler  T, Sridhar  CB, Daga  MK, Pathak  K, Pandit  RB, Khakhria  R, Treatment of typhoid fever with azithromycin versus chloramphenicol in a randomized multicentre trial in India. J Antimicrob Chemother. 1999;44:24350. DOIPubMedGoogle Scholar
  7. Parry  CM, Ho  VA, Phuong  LT, Truong  NT, Bay  PVB, Lahn  MN, Randomized controlled comparison of ofloxacin, azithromycin, and an ofloxacin-azithromycin combination for treatment of multidrug-resistant and nalidixic acid-resistant typhoid fever. Antimicrob Agents Chemother. 2007;51:81925. DOIPubMedGoogle Scholar
  8. Girgis  NI, Butler  T, Frenck  RW, Sultan  Y, Brown  FM, Tribble  D, Azithromycin versus ciprofloxacin for treatment of uncomplicated typhoid fever in a randomized trial in Egypt that included patients with multidrug resistance. Antimicrob Agents Chemother. 1999;43:14414 .PubMedGoogle Scholar
  9. Chinh  NT, Parry  CM, Ly  NT, Ha  HD, Thong  MX, Diep  TS, A randomized controlled comparison of azithromycin and ofloxacin for treatment of multidrug-resistant or nalidixic acid-resistant enteric fever. Antimicrob Agents Chemother. 2000;44:18559. DOIPubMedGoogle Scholar
  10. Dolecek  C, Tran  TP, Nguyen  NR, Le  TP, Ha  V, Phung  QT, A multi-center randomised controlled trial of gatifloxacin versus azithromycin for the treatment of uncomplicated typhoid fever in children and adults in Vietnam. PLoS ONE. 2008;3:e2188. DOIPubMedGoogle Scholar
  11. Sjölund-Karlsson  M, Joyce  K, Blickenstaff  K, Ball  T, Haro  J, Medalla  FM, Antimicrobial susceptibility to azithromycin among Salmonella enterica isolates from the United States. Antimicrob Agents Chemother. 2011;55:39859. DOIPubMedGoogle Scholar
  12. Capoor  MR, Rawat  D, Nair  D, Hasan  AS, Deb  M, Aggarwal  P, In vitro activity of azithromycin, newer quinolones and cephalosporins in ciprofloxacin-resistant Salmonella causing enteric fever. J Med Microbiol. 2007;56:14904. DOIPubMedGoogle Scholar
  13. Rai  S, Jain  S, Prasad  KN, Ghoshal  U, Dhole  TN. Rationale of azithromycin prescribing practices for enteric fever in India. Indian J Med Microbiol. 2012;30:303. DOIPubMedGoogle Scholar
  14. Molloy  A, Nair  S, Cooke  FJ, Wain  J, Farrington  M, Lehner  PJ, First report of Salmonella enterica serotype paratyphi A azithromycin resistance leading to treatment failure. J Clin Microbiol. 2010;48:46557. DOIPubMedGoogle Scholar
  15. Hassing  RJ, Goessens  WH, Mevius  DJ, van Pelt  W, Mouton  JW, Verbon  A, Decreased ciprofloxacin susceptibility in Salmonella Typhi and Paratyphi infections in ill-returned travellers: the impact on clinical outcome and future treatment options. Eur J Clin Microbiol Infect Dis. 2013;32:1295301 . DOIPubMedGoogle Scholar




Cite This Article

DOI: 10.3201/eid2004.131536

Table of Contents – Volume 20, Number 4—April 2014

EID Search Options
presentation_01 Advanced Article Search – Search articles by author and/or keyword.
presentation_01 Articles by Country Search – Search articles by the topic country.
presentation_01 Article Type Search – Search articles by article type and issue.



Please use the form below to submit correspondence to the authors or contact them at the following address:

Robert-Jan Hassing, Erasmus Medical Centre, PO Box 2040, 3000 CA Rotterdam, the NetherlandsRobert-Jan Hassing, Erasmus Medical Centre, PO Box 2040, 3000 CA Rotterdam, the Netherlands

Send To

10000 character(s) remaining.


Page created: March 18, 2014
Page updated: March 18, 2014
Page reviewed: March 18, 2014
The conclusions, findings, and opinions expressed by authors contributing to this journal do not necessarily reflect the official position of the U.S. Department of Health and Human Services, the Public Health Service, the Centers for Disease Control and Prevention, or the authors' affiliated institutions. Use of trade names is for identification only and does not imply endorsement by any of the groups named above.