bWe have determined the in vitro activity of several antibacterial and antifungal drugs against Pythium insidiosum using broth microdilution (BMD), disk diffusion, and Etest methods. The largest zones of inhibition (disk diffusion) and the lowest BMD and Etest MICs were observed for azithromycin, clarithromycin, linezolid, mupirocin, doxycycline, minocycline, and tigecycline. The in vitro activities observed suggest that antibacterials, which act by inhibiting protein synthesis, are promising candidate therapies for the treatment of pythiosis. P ythiosis is a life-threatening and chronic pyogranulomatous disease caused by the fungus-like pathogen Pythium insidiosum, the main oomycete species capable of infecting humans and other animals (1, 2). Although there have been a few reports of clinical cures of cases of pythiosis with antifungal therapy (3, 4), the data from the literature on the clinical management of pythiosis patients with treatment by antifungals indicate that such therapy has been largely ineffective (2,5,6). The genus Pythium is unable to synthesize ergosterol, the active target of most antifungals, which partly explains why this class of drugs has been ineffective (2).Studies on the in vitro susceptibility of the clinical isolates of P. insidiosum to antifungal drugs have shown divergent results, and there are no international protocols approved for evaluating the in vitro susceptibility of P. insidiosum. Interestingly, previous studies have shown that P. insidiosum is quite sensitive to antibacterials belonging to the classes of macrolides, tetracyclines, and glycylcyclines (7,8) and that its combination with antifungal drugs can result in synergistic interactions in vitro (9). However, the effects of antibacterial agents that inhibit protein synthesis on P. insidiosum have not been studied extensively. Thus, the objective of this study was to compare the in vitro susceptibilities of P. insidiosum to a number of antibacterial and antifungal drugs using the Clinical and Laboratory Standards Institute (CLSI) M38-A2 broth microdilution, CLSI M51-A disk diffusion, and Etest methods.We evaluated the susceptibility of 25 Brazilian P. insidiosum strains isolated from equine pythiosis lesions. All of the isolates were identified using a PCR-based assay according to Botton et al. (10). The reference strains included P. insidiosum CBS 101555 from an equine pythiosis case and P. insidiosum CBS 119452 and Pythium aphanidermatum CBS 128995 from human pythiosis cases.The broth microdilution (BMD) reference assay was carried out following the CLSI M38-A2 guidelines (11), as previously described (12, 13). The final concentrations of the antimicrobial agents tested in the wells ranged from 1,024 to 0.5 g/ml for mupirocin and tobramycin and from 256 to 0.125 g/ml for azithromycin, clarithromycin, clindamycin, chloramphenicol, doxycycline, erythromycin, florfenicol, fluconazole, fusidic acid, lincomycin, linezolid, minocycline, roxithromycin, terbinafine, tetracycline, tigecycline, and tilmicosin. The final conce...
eThe present study investigated the in vitro and the in vivo interactions among azithromycin, clarithromycin, minocycline, and tigecycline against Pythium insidiosum. In vitro antimicrobial activities were determined by the broth microdilution method in accordance with CLSI document M38-A2, and the antibiotic interactions were assayed using the checkerboard MIC format. In vivo efficacy was determined using a rabbit infection model. The geometric mean MICs of azithromycin, clarithromycin, minocycline, and tigecycline against P. insidiosum were, respectively, 1.91, 1.38, 0.91, and 0.79 g/ml. By checkerboard testing, all combinations resulted in in vitro synergistic interactions (>60%). Antagonism was not observed. The in vivo studies showed that azithromycin (20 mg/kg/day twice daily) alone or in combination with minocycline (10 mg/kg/day twice daily) significantly decreased the fungal burden. This study demonstrates that azithromycin possesses potent curative efficacy against subcutaneous pythiosis in the rabbit model.
We describe the in vitro activity of macrolides and tetracycline antibiotics against Pythium insidiosum. The MICs were determined according to CLSI procedures (visual MIC) and by a colorimetric method [3-(4,5-dimethyl-2-thiazyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT)]. The lowest geometric mean (GM) MIC (MICs in g/ml) (0.39 and 0.7 by visual reading and colorimetric method, respectively) and MIC ranges (0.125 to 2.0) were obtained for minocycline, while the highest MICs were shown for erythromycin (GM of 7.58 and 12.25 by visual reading and colorimetric method, respectively, and MIC ranged from 2 to 32). This significant in vitro activity makes these classes of antibiotics good candidates for experimental treatment of pythiosis.
Here, a microdilution technique based on the M27-A2 protocol (NCCLS, 2002) was employed to compare the susceptibilities of Candida albicans and Candida dubliniensis to essential oils extracted from plants used as spices. The chemical compositions of the essential oils were defined based on the analysis of retention indices obtained by gas chromatography-mass spectroscopy. Taken together, the results showed that the activity of the compounds against the two species was similar.
Candida dubliniensis is an emerging pathogen first described in 1995, which shares many phenotypic features with Candida albicans and therefore may be misidentified in microbial laboratories. Despite various phenotypic techniques described in the literature to differentiate the two species, the correct identification of C. dubliniensis remains problematic due to phenotypic similarities between these species. Thus, as the differences between both are best characterized at genetic levels, several molecular methods have been proposed to provide a specific and rapid identification of this species. Epidemiological studies have shown that C. dubliniensis is prevalent throughout the world and it is primarily associated with oral carriage and oropharyngeal infections in patients infected with human immunodeficiency virus (HIV). However, data acquired from its isolation from other healthy and immunocompromised patients are variable, and there is still no real consensus on the epidemiological relevance of this species. In this article, we review the various phenotypic methods used in the identification of C. dubliniensis and the epidemiological impact of this new species.
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