Differentiation of Basidiobolus spp. Isolates: RFLP of a Diagnostic PCR Amplicon Matches Sequence-Based Classification and Growth Temperature Preferences

Abstract

The genus Basidiobolus, known since 1886, is primarily associated with reptiles and amphibians. Although globally distributed, rare infections caused by members of this genus mainly occur in tropical and subtropical regions. Morphological and physiological characteristics were used in the past for the description of species. However, some of these characteristics vary depending on culture conditions. Therefore, most species names are regarded as synonyms of B. ranarum as the only pathogenic species. Yet, not all environmental isolates are necessarily pathogenic. This study aimed to analyze if environmental Basidiobolus isolates can be distinguished reliably based on morpho-physiological and molecular characteristics. Eleven isolates originally obtained from feces of south African reptiles and one type strain, Basidiobolus microsporus DSM 3120, were examined morpho-physiologically. Sequence analysis of the 18S and partial 28S rRNA gene and restriction analysis of a diagnostic amplicon (restriction fragment length polymorphism, RFLP) were performed for all 12 strains. Based on the results obtained, morphological features and the 18S rRNA sequence proved insufficient for the reliable differentiation of isolates. However, isolates were distinguishable by growth temperature profiles, which matched isolate clusters established by partial 28S rRNA gene sequence and restriction analysis of a Basidiobolus specific diagnostic PCR amplicon. Our results indicate that RFLP analysis can be used as a fast screening method to identify Basidiobolus isolates with similar physiological characteristics.