Methicillin-resistant Staphylococcus saprophyticus in Sweden carries various types of staphylococcal cassette chromosome mec (SCCmec)

1176

5. 6. 7. 8.

9.

10.

11.

12.

13.

14.

15.

16.

Clinical Microbiology and Infection, Volume 15 Number 12, December 2009

Leishmania chagasi that detects specific antibody in African and American visceral leishmaniasis. Proc Natl Acad Sci USA 1993; 90: 775–779. Ancelle T. Statistique e´pide´miologie, 2nd edn. Maloin: Paris, France, 2002. Petrovic Z. Epidemiology of kala-azar in Serbia. Belgrade: Institute for Medical Research, 1980 (Serbian). Milovanovic M, Popovic D. Addition to research of kala-azar epidemic in NR Serbia. Voice Hyg Inst 1960; 9: 23–27 (Serbian). Ivovic V, Ivovic M, Miscevic Z. Sandflies (Diptera: Psychodidae) in the Bar area of Montenegro (Yugoslavia). Ann Trop Med Parasitol 2003; 97: 193–197. Colovic MD, Jankovic GM, Colovic NR, Cemerikic-Martinovic V. Kala-azar and myelodysplastic syndrome in the same patient. Haema 2002; 5: 246–248. Suvajdzic N, Pavlovic M, Misic S, Cemerikic V, Atkinson HDA, Colovic M. Secondary myelofibrosis in visceral leishmaniasis—case report. Haematologia (Budap) 2001; 31: 167–171. Bern C, Jha SN, Joshi AB, Thakur GD, Bista MB. Use of the recombinant K39 dipstick test and the direct agglutination test in a setting endemic for visceral leishmaniasis in Nepal. Am J Trop Med Hyg 2000; 63: 153–157. Da Silva MRB, Stewart J, Costa CHN. Sensitivity of bone marrow aspirates in the diagnosis of visceral leishmaniasis. Am J Trop Med Hyg 2002; 72: 811–814. Sundar S, Reed SG, Singh VP, Kumar PCK, Murray HW. Rapid accurate field diagnosis of Indian visceral leishmaniasis. Lancet 1998; 351: 563–565. Zijlstra E, Nur Y, Desjeux P, Khalil E, El-Hassan A, Groen J. Diagnosing visceral leishmaniasis with the recombinant K39 strip test: experience from the Sudan. Trop Med Int Health 2001; 6: 108–113. Carvalho SFG, Lemos EM, Corey R, Dietze R. Performance of recombinant K39 antigen in the diagnosis of Brazilian visceral leishmaniasis. Am J Trop Med Hyg 2003; 68: 321–324. Islam MZ, Itoh M, Shamsuzzaman SM et al. Diagnosis of visceral leishmaniasis by enzyme-linked immunosorbent assay using urine samples. Clin Diagn Lab Immunol 2002; 9: 789–794.

Methicillin-resistant Staphylococcus saprophyticus in Sweden carries various types of staphylococcal cassette chromosome mec (SCCmec) B. So¨derquist1,2 and C. Berglund1 Departments of 1) Clinical Microbiology and 2) Infectious Diseases, O¨rebro University Hospital, O¨rebro, Sweden

Abstract Staphylococcus saprophyticus is a common cause of uncomplicated urinary tract infections and is usually susceptible to the antimicrobial agents used for their treatment. However, S. saprophyticus resistant to b-lactam antibiotics and carrying mecA has been reported. Eight Swedish isolates of mecA-positive S. saprophyticus with diverse origin carrying at least three different types of staphylococcal described here.

cassette

chromosome

mec

(SCCmec)

are

CMI

Keywords: mecA gene, staphylococcal cassette chromosome mec, staphylococcus saprophyticus, urinary tract infection Original Submission: 30 June 2008; Revised Submission: 3 September 2008; Accepted: 12 September 2008 Editor: J-L. Mainardi Article published online: 18 May 2009 Clin Microbiol Infect 2009; 15: 1176–1178 10.1111/j.1469-0691.2009.02771.x

Corresponding author and reprint requests: B. So¨derquist, ¨ rebro Department of Clinical Microbiology and Infectious Diseases, O ¨ rebro, Sweden University Hospital, SE-70185 O E-mail: [email protected]

Staphylococcus saprophyticus, a coagulase-negative staphylococcus, is a common cause of uncomplicated urinary tract infections (UTI) in young and sexually active women. Routine laboratory identification of S. saprophyticus is mainly made on the basis of resistance to novobiocin, absence of haemolysis, and negative tests for coagulase and/or DNase. Staphylococcus saprophyticus is usually susceptible to the antimicrobial agents used for treatment of UTI. However, during the late summer in 2007, three isolates of S. saprophyticus were noted at the Department of ¨ rebro University Hospital, Sweden, Clinical Microbiology, O that were resistant to ampicillin and cephadroxil as determined by disc diffusion test (AB Biodisk, Solna, Sweden) according to the Swedish Reference Group for Antibiotics (SRGA, http://www.srga.org). Detection of mecA was performed by using a multiplex PCR adapted for the LightCycler System with SYBR Green I as described previously [1], and the three isolates were found to carry the mecA gene. A retrospective survey of S. saprophyticus isolated from the period 2000–2008 revealed in 15 urine samples containing S. saprophyticus regarded as resistant to cephadroxil. Five of these isolates had been stored at )70C and were subsequently further analysed. In addition, these five isolates were mecA-positive. Altogether, eight isolates were characterized by pulsed-field gel electrophoresis (PFGE) of chromosomal SmaI digests as described [1] and determined to the species level by ID32Staph (bioMe´rieux, Marcy l’Etoile, France). Determination of the staphylococcal cassette chromosome mec (SCCmec) type I, II, III, IV and V was performed as described [2] except for the class B mec complex, which was identified according to Okuma et al. [3].

ª2009 The Authors Journal Compilation ª2009 European Society of Clinical Microbiology and Infectious Diseases, CMI, 15, 1170–1191

Research Notes

CMI

The patients were all women aged 6–41 years (median 21 years), and the urine samples were collected at different ¨ rebro County outpatient clinics from various parts of O (ca. 280 000 inhabitants). MecA-positive isolates of S. saprophyticus were first reported from Japan in 1991 [4] and have thereafter been reported from Canada [5], the USA [6] and Brazil [7]. Although these are predominantly reports of sporadic cases, the prevalence of mecA among S. saprophyticus was in a recent study found to be 7.9% [8]. A large international survey [9] of the antimicrobial susceptibility of pathogens causing uncomplicated UTI from mainly European countries revealed that only 1.7% and 0.9% of S. saprophyticus isolates were resistant to ampicillin and cephadroxil, respectively. Nevertheless, eight ¨ rebro County were S. saprophyticus isolates obtained from O found to be resistant to ampicillin and cephadroxil, but susceptible to nitrofurantoin and trimethoprim. The mecA gene was detected by PCR, and PFGE showed several pulsotypes as shown in Fig. 1. In a recently published study [10] of the molecular epidemiology of S. saprophyticus isolated from UTIs in Sweden, no information was given about antimicrobial susceptibility. However, PFGE analysis of S. saprophyticus causing UTI showed a rather heterogeneous pattern, although some particular clones could be identified [10]. No epidemiological connection was identified among our patients since the urine samples were collected at various outpatient clinics in the county. Characterization of the SCCmec could assign five of the isolates as type III SCCmec, and those were indistinguishable or closely related according to the determination by PFGE, whereas three isolates remained non-typeable using our sets of primers. However, these isolates appear to represent at least two different SCCmec types, as shown in Table 1. Pulsotyping indicated that the non-typeable isolates 04T648 and 06T1173, which carried different mec complexes, were possibly related to each other, but not to the third isolate, 07T887(Fig. 1). Among S. epidermidis, type III and IV SCCmec seems to be the most prevalent SCCmec types [11–13]. Nevertheless, non-typeable isolates that probably carry novel 70

80

90

100

FIG. 1. Pulsed-field gel electrophoresis (PFGE) patterns of eight mecA-positive Staphylococcus saprophyticus isolated from urine samples in Sweden.

1177

TABLE 1. Results of SCCmec typing of eight mecA-positive Staphylococcus saprophyticus isolated from urine samples in Sweden ccr type

mec class

Isolate

1

2

3

C

IS1272

A

B

C

SCCmeca

02T589 04T648 04T741 06T599 06T1173 07T887 07T926 07T1006

n n n n n n n n

n n n n n n n n

Pos n Pos Pos n n Pos Pos

n n n n n n n n

Pos Pos Pos Pos Pos Pos Pos Pos

Pos n Pos Pos Pos Pos Pos Pos

n Pos n n n n n n

n n n n n n n n

III NT III III NT NT III III

a

NT, not typeable.

SCCmec types are commonly found. Of the non-typeable S. saprophyticus isolates in the present report, two carried the class A mec gene complex, but were negative for all the investigated ccr types. However, all eight isolates carried the insertion sequence IS1272 somewhere in the chromosome, yet only one isolate had the insertion in close proximity to mecA, indicating a class B mec complex. We believe that SCCmec in methicillin-resistant S. saprophyticus should be described with caution and should not be directly compared with the SCCmec described in methicillin-resistant Staphylococcus aureus (MRSA), since only the essential components of the elements were detected, and since only a handful of isolates have been investigated. However, the finding of similar mec complexes in methicillin-resistant S. saprophyticus, as in MRSA, may indicate a common origin. In contrast, the presence of a novel ccr gene complex, or perhaps the absence thereof, clearly indicates differences among various staphylococcal species and motivates further studies in order to clarify the complexity and obscurity of SCCmec in staphylococci, as shown by Hanssen et al. [14]. It is also interesting to note that, whereas the majority of S. epidermidis isolates carry mecA, it is much less prevalent in S. saprophyticus. This may indicate that the recipient chromosome differs among various staphylococcal species and that the SCCmec types are different. Most recently, characterization of the SCCmec of eight mecA-positive S. saprophyticus isolates from Japan has been reported [15]. These isolates were all non-typeable according to the current SCCmec classification, due to the presence of a class A mec complex and the absence of an amplification product for hitherto known ccr genes. Further analysis of a prototype S. saprophyticus isolate showed a novel composition of the class A mec gene complex in combination with the ccrA1/ccrB3 gene complex. In summary, this is the first report of characterization of methicillin-resistant S. saprophyticus from Europe, and these isolates have diverse origins and carry various types of SCCmec.

ª2009 The Authors Journal Compilation ª2009 European Society of Clinical Microbiology and Infectious Diseases, CMI, 15, 1170–1191

1178

Clinical Microbiology and Infection, Volume 15 Number 12, December 2009

Acknowledgements We thank B. Dragsten for performing the PFGE.

Transparency declaration B. So¨derquist has served as a consultant to Pfizer AB, Sweden.

CMI

13. Mombach Pinheiro Machado AB, Reiter KC, Paiva RM, Barth AL. Distribution of staphylococcal cassette chromosome mec (SCCmec) types I, II, III and IV in coagulase-negative staphylococci from patients attending a tertiary hospital in southern Brazil. J Med Microbiol 2007; 56: 1328–1333. 14. Hanssen AM, Kjeldsen G, Sollid JU. Local variants of staphylococcal cassette chromosome mec in sporadic methicillin-resistant Staphylococcus aureus and methicillin-resistant coagulase-negative staphylococci: evidence of horizontal gene transfer? Antimicrob Agents Chemother 2004; 48: 285–296. 15. Higashide M, Kuroda M, Omura CT et al. Methicillin-resistant Staphylococcus saprophyticus carrying staphylococcal cassette chromosome mec (SCCmec) have emerged in urogenital tract infections. Antimicrob Agents Chemother 2008; 52: 2061–2068.

References 1. Berglund C, Mo¨lling P, Sjo¨berg L, So¨derquist B. Predominance of staphylococcal cassette chromosome mec (SCCmec) type IV among methicillin-resistant Staphylococcus aureus (MRSA) in a Swedish county and presence of unknown SCCmec types with Panton–Valentine leukocidin genes. Clin Microbiol Infect 2005; 11: 447–456. 2. Berglund C, So¨derquist B. The origin of a methicillin-resistant Staphylococcus aureus (MRSA) at a neonatal ward in Sweden— possible horizontal transfer of a staphylococcal cassette chromosome mec between methicillin-resistant Staphylococcus haemolyticus and Staphylococcus aureus. Clin Microbiol Infect 2008; 14: 1048– 1056. 3. Okuma K, Iwakawa K, Turnidge JD et al. Dissemination of new methicillin-resistant Staphylococcus aureus clones in the community. J Clin Microbiol 2002; 40: 4289–4294. 4. Murakami K, Minamide W, Wada K, Nakamura E, Teraoka H, Watanabe S. Identification of methicillin-resistant strains of staphylococci by polymerase chain reaction. J Clin Microbiol 1991; 29: 2240– 2244. 5. Hussain Z, Stoakes L, Massey V et al. Correlation of oxacillin MIC with mecA gene carriage in coagulase-negative staphylococci. J Clin Microbiol 2000; 38: 752–754. 6. Swenson JM, Tenover FC. Cefoxitin disk study group. Results of disk diffusion testing with cefoxitin correlate with presence of mecA in Staphylococcus spp. J Clin Microbiol 2005; 43: 3818–3823. 7. Souza Antunes AL, Secchi C, Reiter KC, Rodrigues Perez LR, Peixoto de Freitas AL, Alves d’Azevedo P. Evaluation of oxacillin and cefoxitin disks for detection of resistance in coagulase negative staphylococci. Mem Inst Oswaldo Cruz 2007; 102: 719–723. 8. Higashide M, Kuroda M, Ohkawa S, Ohta T. Evaluation of a cefoxitin disk diffusion test for the detection of mecA-positive methicillin-resistant Staphylococcus saprophyticus. Int J Antimicrob Agents 2006; 27: 500– 504. 9. Kahlmeter G. An international survey of the antimicrobial susceptibility of pathogens from uncomplicated urinary tract infections: the ECO.SENS project. J Antimicrob Chemother 2003; 51: 69–76. 10. Widerstro¨m M, Wistro¨m J, Ferry S, Karlsson C, Monsen T. Molecular epidemiology of Staphylococcus saprophyticus isolated from women with uncomplicated community-acquired urinary tract infection. J Clin Microbiol 2007; 45: 1561–1564. 11. Wisplinghoff H, Rosato AE, Enright MC, Noto M, Craig W, Archer GL. Related clones containing SCCmec type IV predominate among clinically significant Staphylococcus epidermidis isolates. Antimicrob Agents Chemother 2003; 47: 3574–3579. 12. Miragaia M, Couto I, de Lencastre H. Genetic diversity among methicillin-resistant Staphylococcus epidermidis (MRSE). Microb Drug Resist 2005; 11: 83–93.

Molecular characterization of resistance to rifampicin in clinical isolates of Neisseria meningitidis A. Skoczynska1,2, C. Ruckly1, E. Hong1 and M-K. Taha1 1) Neisseria Unit, Institute Pasteur, Paris, France and 2) National Reference Centre for Bacterial Meningitis, National Medicines Institute, Warsaw, Poland

Abstract Among 3904 meningococcal isolates collected between October 2002 and June 2007 by the French Meningococcal Reference Centre, eight (0.20%) were resistant to rifampicin (Rif-R; MIC >1 mg/L) and 27 (0.69%) were intermediate-resistant to rifampicin (Rif-I; MICs between 0.38 mg/L and 1 mg/L) according to the E-test method. The MICs determined by agar dilution were lower, eliminating the E-test intermediate category. All Rif-R isolates had mutations in the rpoB gene, resulting in substitutions at or near amino acid position 552, which were absent in non-resistant isolates. These data suggest that a rifampicin clinical breakpoint of 1.0 mg/L should be adopted for Neisseria meningitidis.

Keywords: Meningococcus, mtrR, resistance mechanisms, rifampicin, rpoB Original Submission: 17 March 2008; Revised Submission: 27 August 2008; Accepted: 2 September 2008 Editor: A. Sundsfjord Article published online: 18 May 2009 Clin Microbiol Infect 2009; 15: 1178–1181 10.1111/j.1469-0691.2009.02783.x

ª2009 The Authors Journal Compilation ª2009 European Society of Clinical Microbiology and Infectious Diseases, CMI, 15, 1170–1191