Paecilomyces variotii is a commonly occurring species in air and food, but it is also associated with many types of human infections and is among the emerging causative agents of opportunistic mycoses in immunocompromised hosts. Paecilomyces can cause hyalohyphomycosis, and two species, Paecilomyces lilacinus and P. variotii, are the most frequently encountered organisms. In the present study, a set of 34 clinical isolates morphologically identified as P. variotii or P. lilacinus were formally identified by sequencing intergenic transcribed spacer regions 1 and 2 (including 5.8S rDNA) and a part of the -tubulin gene. Three isolates were identified as P. lilacinus, and five of the presumptive P. variotii isolates did not belong to the genus Paecilomyces but were identified as Talaromyces eburneus (anamorph, Geosmithia argillacea) or Hamigera avellanea (anamorph, Merimbla ingelheimense). Applying the most recent taxonomy, we found that the clinical P. variotii isolates could be identified as P. variotii sensu stricto (14 strains), P. formosus (11 strains), and P. dactylethromorphus (1 strain). These data indicate that P. formosus occurs in clinical samples as commonly as P. variotii. Susceptibility tests showed that the antifungal susceptibility profiles of P. variotii, P. formosus, and P. dactylethromorphus are similar and that all strains tested were susceptible to amphotericin B in vitro. P. lilanicus, T. eburneus, and H. avellanea had different susceptibility profiles; and flucytosine and voriconazole were the least active of the antifungal drugs tested against these species. Our results indicate that correct species identification is important to help guide appropriate antifungal therapy.Paecilomyces variotii is a commonly occurring species that has previously been isolated from various substrates, including (pasteurized) foods, soil, indoor air, and wood (23,36,41,42,43). However, it is also associated with many types of human infections and is listed among the emerging causative agents of opportunistic mycoses in immunocompromised hosts. Paecilomyces can cause hyalohyphomycosis (1), and two species, Paecilomyces lilacinus and P. variotii, are the most frequently encountered (20, 52). Both species are morphologically similar but can be differentiated on the basis of conidial color and growth rates (41). However, small-subunit ribosomal gene sequences showed that the two species are unrelated: P. variotii belongs to the order Eurotiales, while P. lilacinus is a member of the order Hypocreales (27). Although they are uncommon, Paecilomyces infections are associated with almost any organ or system of the human body (40). Most cases concern immunocompromised patients and are cutaneous or catheter related. However, dissemination, for example, that involving the central nervous system, has been observed in a number of cases (15,24). Ocular infections associated with prolonged contact lens use or ocular surgery have also been reported (40), as has peritonitis in patients with continuous ambulatory peritoneal dialy...