Typing of Scedosporium apiospermum by multilocus enzyme electrophoresis and random amplification of polymorphic DNA

Abstract

The genetic diversity among epidemiologically unrelated strains of"the human pathogenic fungus Scedosporium apiospermum or its teleomorph, Pseudallescheria boydii, from different areas in Europe, was investigated by multilocus enzyme electrophoresis (MLEE) and random ampli®cation of polymorphic DNA (RAPD). Fourteen enzyme activities were analysed by starch gel electrophoresis, corresponding to 27 polymorphic loci and 43 iso-enzymes. Among the enzymes studied, propionate esterase, carboxyl esterase, superoxide dismutase, carbonate dehydratase and malate dehydrogenase were the most polymorphic, allowing the classi®cation of the strains into 6±11 groups each. Combination of the data obtained for the different enzyme activities studied allowed differentiation of the strains. Similarly, a high polymorphism was also revealed by each of the 20 RAPD primers tested, but no single primer was able to differentiate all the strains. The most ef®cient primers were GC70, UBC-701 and UBC-703, which revealed 17 distinct genotypes each, and combination of the results obtained with this threeprimer set allowed complete discrimination of the strains. The dendrograms obtained from MLEE or RAPD by the unweighted pair-group method using arithmetic average cluster analysis did not reveal any clustering according to the geographic origin of the strains or their pathogenicity.

Molecular Typing of AustralianScedosporiumIsolates Showing Genetic Variability and NumerousS. aurantiacum

“…For all 3 Scedosporium spp., there was no clustering of strains according to their geographic or body site of origin or by their ability to cause invasive disease, which is in agreement with previous fi ndings for S. apiospermum (20,30) and S. prolifi cans (16,17,21). Of note, no specifi c genotypes were associated with underlying medical conditions or risk factors.…”

“…This diversity is generally consistent with the high degree of polymorphism (15-20 genotypes) previously found (10,20,32). In contrast, genetic variation was lowest (28%) among the S. aurantiacum isolates (online Appendix Figure 1; Figure 3).…”

“…The moderate genetic diversity among S. prolificans confi rms previous fi ndings (19). Despite the observed polymorphisms, PcoA of PCR fi ngerprint profi les showed dense clustering for S. prolifi cans (Figure 3), which is consistent with the low to absent intraspecies variability in S. prolifi cans found by others (20,21,33). These apparently contradictory fi ndings emphasize the importance of choosing the optimum molecular typing tool with the most appropriate discriminatory power for the organism or species being studied.…”

“…Molecular typing techniques now provide the means to elucidate the epidemiology of Scedosporium infections and to investigate potential case clusters (16,18,19). Strains recovered from patients with cystic fi brosis have demonstrated a high degree of genetic variability (10,20), although a single genetic profi le predominated in 1 study (8). The degree of genetic variation within S. prolifi cans is more controversial.…”

Genotyping Study of Scedosporium apiospermum Isolates from Patients with Cystic Fibrosis

“…In addition, little information is available concerning the epidemiology of the colonization of the airways by this fungus. In order to improve our knowledge in this field, sequential and multiple isolates from different CF patients were typed by random amplification of polymorphic DNA (RAPD) with a three-primer set previously selected for its ability to discriminate among epidemiologically unrelated strains of S. apiospermum (26).…”