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Antimicrobial Agents and Chemotherapy, October 2003, p. 3371-3372, Vol. 47, No. 10
0066-4804/03/$08.00+0     DOI: 10.1128/AAC.47.10.3371-3372.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

LETTER TO THE EDITOR

Comparison of In Vitro Antifungal Susceptibilities of Conidia and Hyphae of Dermatophytes with Thick-Wall Macroconidia


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LETTER
 
Although the invasive forms of filamentous fungi are generally hyphae, the reference methods (6) recommend the use of conidial suspensions, such as inocula. Some studies testing Aspergillus spp. have compared the MICs obtained with conidial suspensions with the MICs obtained with hyphal suspensions by using both forms of inocula, and the MICs were similar (1, 3, 7). This was probably because the thickness of the cell walls of the conidia and hyphae of the fungi tested were similar. However, in some fungi, such as several dermatophytes, the wall of the conidia is considerably thicker than the wall of the hyphae, and we do not know whether the antifungal susceptibilities of these two morphological forms would be different. We have compared how inocula of conidia and hyphae of five species of dermatophytes with thick-wall macroconidia affect the MICs of five antifungal agents.

A total of 14 strains of dermatophytes were tested three times on different days against five antifungal drugs, i.e., clotrimazole (CTZ), itraconazole (ITZ), ravuconazole (RVZ), terbinafine (TF), and voriconazole (VCZ), using both hyphal and conidial inocula (Table 1). Drug dilutions were prepared by the reference methods (6). The final concentrations were 16 to 0.01 µg/ml for all drugs. To obtain an inoculum of macroconidia, the fungus was grown on wheat flour-cow milk agar supplemented with honey at 28°C for 7 to 10 days. The inoculum was prepared by flooding the surface of the agar with distilled water containing 0.05% Tween 80 and scraping the sporulated colonies with the tip of a Pasteur pipette. The suspensions were filtered through sterile gauze to remove the microconidia. The macroconidia retained in the gauze were washed with distilled water and adjusted to a concentration of 5 x 106 macroconidia/ml with a hemacytometer. To obtain an inoculum of hyphae, the fungus was grown on Sabouraud agar at 37°C for 5 to 7 days. Hyphal suspensions were prepared as described previously. They were then vortexed while the pieces of mycelium were broken with a glass rod and filtered through sterile gauze to remove hyphal mats. Hyphal fragments were adjusted to 5 x 106 hyphal fragments/ml. Tests were performed using a broth microdilution method (6). Microdilution trays were incubated at 28 and 37°C (macroconidia and hyphae, respectively) for 4 days. MIC was defined as the lowest drug concentration showing 100% growth inhibition. The significance of the different MICs obtained with the two types of inocula for each antifungal was determined by the Wilcoxon test.


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  		TABLE 1. In vitro activities of five antifungal agents against 14 isolates of dermatophytes using two types of inoculaa

The MICs obtained with conidia were generally higher than those obtained with hyphae for all drugs tested (P < 0.05) (Table 1). The highest MICs (>=8 µg/ml) were reached with macroconidial inocula of the following species and drugs: Microsporum canis and TF; Microsporum cookei and RVZ; Microsporum gypseum and CTZ; Microsporum racemosum and TF, VCZ, and ITZ; and Trichophyton ajelloi and ITZ.

For all species and antifungal drugs, overall agreement (differences not greater than one dilution) between MICs obtained with both types of inocula was low (25.7%). The highest agreement was with M. canis (73.4%), and the lowest agreement was with M. cookei and T. ajelloi (0%).

It is difficult to compare these results with other studies because of the scarcity of data on how the inoculum form affects the MICs for dermatophytes. Nardoni et al. (5) have been the only researchers to have evaluated this. They tested TF against M. canis, and the MICs obtained with both inocula were similar. The results obtained with other fungal research groups have been contradictory. Nakai et al. (4) tested dimorphic fungi and observed that the MIC of micafungin was higher for the yeast-like form than for the mycelial form. With fluconazole, the results were just the opposite. Other studies (1, 2, 7) indicated that MICs were independent of the form of inoculum. Our results highlight the importance of the type of inoculum and the thickness of the cell walls in determining the in vitro antifungal susceptibility of dermatophytes. Further studies are needed to determine which type of inoculum is more predictive.


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REFERENCES
 
    1
  1. Espinel-Ingroff, A. 2001. Germinated and nongerminated conidial suspensions for testing of susceptibilities of Aspergillus spp. to amphotericin B, itraconazole, posaconazole, ravuconazole, and voriconazole. Antimicrob. Agents Chemother. 45:605-607.[Abstract/Free Full Text]
    2. 2
  2. González, G. M., R. Tijerina, D. A. Sutton, J. R. Graybill, and M. G. Rinaldi. 2002. In vitro activities of free and lipid formulations of amphotericin B and nystatin against clinical isolates of Coccidioides immitis at various saprobic stages. Antimicrob. Agents Chemother. 46:1583-1585.[Abstract/Free Full Text]
    3. 3
  3. Manavathu, E. K., J. Cutright, and P. H. Chandrasekar. 1999. Comparative study of susceptibilities of germinated and ungerminated conidia of Aspergillus fumigatus to various antifungal agents. J. Clin. Microbiol. 37:858-861.[Abstract/Free Full Text]
    4. 4
  4. Nakai, T., J. Uno, F. Ikeda, S. Tawara, K. Nishimura, and M. Miyaji. 2003. In vitro antifungal activity of micafungin (FK463) against dimorphic fungi: comparison of yeast-like and mycelial forms. Antimicrob. Agents Chemother. 47:1376-1381.[Abstract/Free Full Text]
    5. 5
  5. Nardoni, S., F. Millanta, and F. Mancianti. 2000. In vitro sensitivity of two different inocula of Microsporum canis versus terbinafine. J. Mycol. Med. 10:148-151.
    6. 6
  6. National Committee for Clinical Laboratory Standards. 2002. Reference method for broth dilution antifungal susceptibility testing of conidium-forming filamentous fungi. Proposed approved standard. NCCLS document M38-AP. National Committee for Clinical Laboratory Standards, Wayne, Pa.
    7. 7
  7. Pujol, I., J. Fernández-Ballart, and J. Guarro. 2001. Effect of inoculum form on in vitro antifungal susceptibilities of Aspergillus spp. J. Antimicrob. Chemother. 47:715-718.[Abstract/Free Full Text]
Belkys Fernández-Torres
Isabel Inza
Josep Guarro*
Unitat de Microbiología
Facultat de Medicina
Universitat Rovira i Virgili
Reus, Spain

* Phone: 34 977 759359
Fax: 34 977 759322
E-mail: umb{at}astor.urv.es

Antimicrobial Agents and Chemotherapy, October 2003, p. 3371-3372, Vol. 47, No. 10
0066-4804/03/$08.00+0     DOI: 10.1128/AAC.47.10.3371-3372.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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  • Santos, D. A., Barros, M. E. S., Hamdan, J. S. (2006). Establishing a Method of Inoculum Preparation for Susceptibility Testing of Trichophyton rubrum and Trichophyton mentagrophytes. J. Clin. Microbiol. 44: 98-101 [Abstract] [Full Text]  
  • Santos, D. A., Hamdan, J. S. (2005). Evaluation of Broth Microdilution Antifungal Susceptibility Testing Conditions for Trichophyton rubrum. J. Clin. Microbiol. 43: 1917-1920 [Abstract] [Full Text]  

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