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Antifungal resistance has been associated with biofilm formation in many microorganisms, but not yet in Malassezia pachydermatis. This saprophytic yeast can cause otitis and dermatitis in dogs and has emerged as an important human pathogen, responsible for systemic infections in neonates in intensive care units. This study aims to evaluate the in vitro antifungal susceptibility of M. pachydermatis strains, in both their planktonic and sessile forms, to fluconazole, miconazole, ketoconazole, itraconazole, posaconazole, terbinafine and voriconazole using the XTT assay and Clinical and Laboratory Standards Institute (CLSI) microdilution method. The minimum inhibitory concentration (MIC) values recorded for each drug were significantly higher for sessile cells relative to planktonic cells to the extent that ≥ 90% of M. pachydermatis strains in their sessile form were classified as resistant to all antifungal agents tested. Data suggest that M. pachydermatis biofilm formation is associated with antifungal resistance, paving the way towards investigating drug resistance mechanisms in Malassezia spp.

The few studies attempting to specifically characterize dermatophytes from hair samples of dogs and cats using PCR-based methodology relied on sequence-based analysis of selected genetic markers. The aim of the present investigation was to establish and evaluate a PCR-based approach employing genetic markers of nuclear DNA for the specific detection of dermatophytes on such specimens. Using 183 hair samples, we directly compared the test results of our one-step and nested-PCR assays with those based on conventional microscopy and in vitro culture techniques (using the latter as the reference method). The one step-PCR was highly accurate (AUC > 90) for the testing of samples from dogs, but only moderately accurate (AUC = 78.6) for cats. A nested-PCR was accurate (AUC = 93.6) for samples from cats, and achieved higher specificity (94.1 and 94.4%) and sensitivity (100 and 94.9%) for samples from dogs and cats, respectively. In addition, the nested-PCR allowed the differentiation of Microsporum canis from Trichophyton interdigitale (zoophilic) and geophilic dermatophytes (i.e., Microsporum gypseum or Trichophyton terrestre), which was not possible using the one step-assay. The PCRs evaluated here provide practical tools for diagnostic applications to support clinicians in initiating prompt and targeted chemotherapy of dermatophytoses.

Dermatitis caused by Malassezia spp., one of most common skin disease in dogs, requires prolonged therapy and/or high doses of antifungal agents. In the present study, the antifungal susceptibility of M. pachydermatis to ketoconazole (KTZ), fluconazole (FLZ), itraconazole (ITZ), posaconazole (POS) and voriconazole (VOR) was evaluated in vitro using both CLSI reference broth microdilution (CLSI BMD) and E-test. A total of 62 M. pachydermatis strains from dogs with and without skin lesions were tested. M. pachydermatis strains were susceptible to ITZ, KTZ and POS using both test methods, with the highest MIC found in tests of FLZ. Essential agreement between the two methods ranged from 87.1% (VOR) to 91.9% (ITZ), and categorical agreement from 74.2% (FLZ) to 96.8% (ITZ). Minor error discrepancies were observed between the two methods, with major discrepancies observed for KTZ. A higher MIC(50) value for FLZ was noted with M. pachydermatis genotype B. The MICs(50) of M. pachydermatis genotype B for KTZ, VOR and POS were higher in isolates from dogs with skin lesions than those in isolates from animals without skin lesions. The results suggest a link between genotypes of M. pachydermatis and in vitro drug susceptibility. The categorical agreement for both E-test and CLSI BMD methods found in this investigation confirms the E-test as a reliable diagnostic method for routine use in clinical mycology laboratories.

Three lipid-dependent Malassezia isolates (here named 114A, 114B and 114C) recovered from a dog with skin lesions were phenotypically and genotypically characterized. All presented ovoid cells and buds formed on a narrow base. Most of the results from physiological tests were consistent with those of Malassezia furfur. The phylogenetic analysis of ITS-1 and LSU nucleotide sequences was concordant in placing all three clinical Malassezia isolates close to M. furfur. However, the phylogenetic data on the chs-2 sequence revealed that clinical isolate 114A is distinct from M. furfur and was closely affiliated to the sequence of M. pachydermatis with high nodal support. In particular, lipid-dependent isolates 114A displayed chs-2 sequences similar (100%) to that of the non-lipid dependent species Malassezia pachydermatis. The presence of the genetic and physiological polymorphisms detected in these three isolates of M. furfur could have resulted from a process of adaptation of this anthropophilic species to a new host.

While dermatophytoses of several animal species have been extensively investigated, information on their occurrence and epidemiology in rabbits is limited. We carried out a study from October 2006 to February 2007 of 23 rabbit farms in Apulia and Basilicata regions (southern Italy) in order to investigate the occurrence and risk factors associated with dermatophytoses in breeding rabbits. Dermatophytes were isolated from 86.9% (20/23) of the farms and from 51.8% (420/810) of the animals sampled. Trichophyton mentagrophytes (47.9%) and Microsporum canis (3.2%) were isolated from diseased (71.7%) and healthy (48.4%) animals as well from the surrounding environment (7.5%). The occurrence of lesions, the age of rabbits, and farm management (e.g., temperature, humidity and methods and frequency of disinfection practices) were identified as the most significant risk factors (P < 0.05) for the occurrence of dermatophytes. Animals in fattening and finishing stages were the most frequently infected (i.e., 58.2 and 61.6% respectively). Dermatophyte prevalence was significantly (P < 0.05) higher in areas with higher temperature (>20°C) and relative humidity ranging from 62-65%. The results of the present investigation suggest that zoonotic dermathophytes are present in rabbit farms and highlight the importance of correct management procedures for the control of the infections.