JAC Advance Access published online on September 28, 2008
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dkn403
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Research letter |
Comparison of killing activity of caspofungin against Candida parapsilosis, Candida orthopsilosis and Candida metapsilosis
1 Faculty of Dentistry, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary 2 Department of Medical Microbiology, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary 3 Department of Ophthalmology, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary
* Corresponding author. Tel: +36-52-417-565; Fax: +36-52-417-565; E-mail: major{at}med.unideb.hu
Key Words: paradoxical growth , fungicidal , time–kill method , medium dependence
Candida orthopsilosis and Candida metapsilosis are Candida species recently separated from Candida parapsilosis sensu stricto, replacing the formerly existing designation of C. parapsilosis groups II and III.1 Retrospective analysis with DNA-based techniques of the formerly identified C. parapsilosis strain collections revealed that C. orthopsilosis and C. metapsilosis together account for fewer than 10% of all infections previously attributed to C. parapsilosis.2,3
Information about antifungal susceptibilities of C. orthopsilosis and C. metapsilosis is scant.2,3 However, based on newer data, echinocandin MICs for C. orthopsilosis and C. metapsilosis strains seem to be statistically significantly lower compared with C. parapsilosis.3
The aim of our study was to determine the killing activity of caspofungin against C. parapsilosis, C. orthopsilosis and C. metapsilosis using time–kill methodology.
We used six, three and four C. parapsilosis, C. orthopsilosis and C. metapsilosis isolates identified with DNA-based methods;1 C. parapsilosis ATCC 22019 strain was also included for reference. Two C. parapsilosis, three C. orthopsilosis and three C. metapsilosis isolates were from an Italian collection of clinical isolates; all others were isolated in the University of Debrecen, Hungary.
MICs were determined according to the CLSI (formerly the NCCLS) method as well as with its slightly modified version using antibiotic medium 3 (AM3) (Fluka), but otherwise the same experimental conditions.4,5 The caspofungin concentration range was 0.015–8 mg/L; MICs were read after 24 h using the partial inhibition criterion.4 Time–kill studies were performed as described previously using RPMI-1640 and AM3 in two simultaneous tests.5 Caspofungin concentrations ranged from 0.06 to 16 mg/L in both media. Aliquots were removed at 0, 4, 8, 12, 24 and 48 h and plated onto Sabouraud dextrose agar. Plates were incubated at 35°C for 48 h, and fungicidal activity was defined as a 99.9% reduction in viable cell count compared with the starting inoculum.5 As C. parapsilosis shows high (90%) frequency of paradoxical growth (PG) in RPMI-1640,6 PG was also evaluated after 24 and 48 h. PG was defined as killing activity (99.9% reduction of the starting inoculum) observed at least at two supra-MIC concentrations, but consistent lack of killing by higher concentrations.5 All tests were performed twice with all strains.
Table 1 shows caspofungin MICs read at 24 h and killing activity after 24 and 48 h in RPMI-1640 and AM3. PG was observed neither in RPMI-1640 nor in AM3 in MIC tests.
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In the killing studies using RPMI-1640, caspofungin showed a fungistatic effect in the case of C. parapsilosis and C. orthopsilosis after 24 h (Table 1). However, CP85 C. orthopsilosis was killed by 2 mg/L (16xMIC) caspofungin, but not by any other concentrations. After 48 h, decreases in viable cfu numbers at 1–16 mg/L caspofungin concentrations for C. parapsilosis and C. orthopsilosis varied between 1.07 and 2.95 log10 cfu/mL and between 1.18 and 2.97 log10 cfu/mL, respectively. After 48 h, caspofungin was fungicidal at concentrations of 1–8 mg/L (16–128xMIC) against all C. metapsilosis isolates apart from isolate number CP92.
In killing studies with AM3, a fungistatic effect was observed after 24 h; two of the seven C. parapsilosis isolates showed PG (Table 1). After 48 h, four of the seven C. parapsilosis isolates were killed at 
0.5 mg/L caspofungin concentrations (0.5–2xMIC); the remaining three isolates (isolates 9150, 896/1 and CP117) showed PG. All C. orthopsilosis isolates showed PG after 24 h (Table 1). However, after 48 h, all isolates were killed by caspofungin concentrations of 0.12–1 mg/L (2–16xMIC) or higher. All C. metapsilosis strains were killed at caspofungin concentrations of 0.06 mg/L (1xMIC) after 48 h, but not after 24 h (Table 1).
In our work, the three C. orthopsilosis isolates behaved similar to C. parapsilosis after 24 h; fungistatic or PG effects were observed regardless of the medium used. However, in AM3 after 48 h, caspofungin proved to be clearly fungicidal against C. orthopsilosis, but not against C. parapsilosis isolates. C. metapsilosis isolates, regardless of the medium used, were more susceptible to the killing activity of caspofungin.
This study is the first comparing caspofungin killing activity against the closely related species C. parapsilosis, C. orthopsilosis and C. metapsilosis. Killing curves, regardless of the medium used, showed a decreasing order of susceptibility to caspofungin: C. metapsilosis > C. orthopsilosis > C. parapsilosis.
Based on high echinocandin MICs for C. parapsilosis sensu stricto, in the case of isolates identified as C. parapsilosis sensu lato low MICs of echinocandins may be regarded as an indicator that an isolate is in fact C. orthopsilosis or C. metapsilosis; in the case of isolates with low echinocandin MICs, DNA-based identification1–3 of the isolates is desirable. Because C. orthopsilosis and C. metapsilosis seem to be relevant species among bloodstream isolates in some countries,2,3 this distinction may be particularly important in some epidemiological situations or in clinical situations when the use of echinocandins as therapy or prophylaxis is planned.
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Funding |
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The study did not receive financial support from third parties.
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TopFundingTransparency declarationsReferences |
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None to declare.
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Acknowledgements |
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We thank Ariana Tavanti for kindly providing the Italian isolates. Caspofungin pure powder was kindly provided by Merck Research Laboratories, Rahway, NJ, USA.
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References |
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TopFundingTransparency declarationsReferences |
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1 . Tavanti A, Davidson AD, Gow NAR, et al. Candida orthopsilosis and Candida metapsilosis spp. nov. to replace Candida parapsilosis groups II and III. J Clin Microbiol (2005) 43:284–92.
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Gomez-Lopez A, Alastruey-Izquierdo A, Rodriguez D, et al. Prevalence and susceptibility profile of Candida metapsilosis and Candida orthopsilosis: results from population-based surveillance of candidemia in Spain. Antimicrob Agents Chemother (2008) 52:1506–9.
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Lochart SR, Messer SA, Pfaller MA, et al. Geographic distribution and antifungal susceptibility of the newly described species Candida orthopsilosis and Candida metapsilosis, in comparison to the closely-related species Candida parapsilosis. J Clin Microbiol (2008) 46:2659–64.
4 . National Committee for Clinical Laboratory Standards. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts—Second Edition: Approved Standard M27-A2 (2002) Wayne, PA, USA: NCCLS.
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Varga I, Soczo G, Kardos G, et al. Time–kill studies investigating the killing activity of caspofungin against Candida dubliniensis: comparing RPMI-1640 and antibiotic medium 3. J Antimicrob Chemother (2008) 62:149–52.
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Chamilos G, Lewis RE, Albert N, et al. Paradoxical effect of echinocandins across Candida species in vitro: evidence for echinocandin-specific and Candida species-related differences. Antimicrob Agents Chemother (2007) 51:2257–9.
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