A molecular analysis of species boundaries and relationships in Pythium

Restriction fragment length polymorphism (RFLP) analysis of the ribosomal DNA (rDNA) intergenic spacer (IGS) was used to resolve relationships among over 360 isolates representing 90 Pythium species. Multiple isolates of the same species usually exhibited minimal intraspecific variation in their restriction fragment profiles, while a high degree of intraspecific variation was usually observed when comparing fragment patterns between different species. In some cases though, different isolates of the same species had very different fragment profiles. This may be indicative of genetic variation within a species, or of the existence of cryptic speciation. However, some groups of morphologically similar species shared similar restriction fragment profiles. For such groups, RFLP analysis revealed that isolates representing different species often formed monophyletic clusters. The clustering of these species was usually supported by a high degree of morphological similarity among the species. This suggested that certain species may in fact be conspecific.; Pythium insidiosum, the only species in the genus capable of infecting animals, is the etiological agent of pythiosis, a disease characterized by cutaneous and subcutaneous lesions and some vascular diseases. RFLP analysis of the IGS for 28 isolates of P. insidiosum from a variety of animal hosts and geographic origins revealed the existence of three genetic clusters which exhibited a high degree of geographic isolation. These results were strengthened by a sequence analysis using rDNA internal transcribed spacer (ITS).; Accurate diagnosis of pythiosis and identification of its causal agent are often inconsistent with current diagnostic methods. The presence of other pathogenic organisms that produce symptoms similar to pythiosis in their hosts complicates matters further. A species-specific DNA probe was constructed using a 530 by HinfI fragment from the IGS of P. insidiosum. When the probe was incubated with dot blots of genomic DNA from 104 Pythium species, it hybridized only to the DNA of P. insidiosum and P. destruens, two species which are assumed to be conspecific. The probe also hybridized to DNA from all the isolates of P. insidiosum tested, regardless of their geographic origin or animal host. When tested against genomic DNA from other pathogenic organisms (Aspergillus fumigatus, Basidiobolus ranarum, Conidiobolus coronatus, Lagenidium giganteum, Paracoccidioides brasiliensis , and Prototheca wickerhamii), no cross-hybridization was detected. (Abstract shortened by UMI.)...