Genotyping of South American clinical isolates of Pythium insidiosum based on single nucleotide polymorphism-based multiplex PCR

Abstract

We aimed to genotype the South American clinical isolates of Pythium insidiosum using the single nucleotide polymorphisms (SNP) of the ribosomal DNA sequences (rDNA). Previously, an SNP-based multiplex-PCR was able to distinguish three different clades of P. insidiosum isolates. Thus, we used this assay to evaluate South American clinical isolates of P. insidiosum (n=32), standard strains from Costa Rica (n=4), Thailand (n=3), Japan (n=1), and India (n=1), a standard strain of Pythium aphanidermatum, and Brazilian environmental isolates of Pythium torulosum, Pythium rhizo-oryzae and Pythium pachycaule voucher (n=3). It was possible to allocate each American P. insidiosum isolate to clade I, the isolates of India, Japan, and Thailand to clade II, and the Thai isolate to clade III. P. aphanidermatum, P.torulosum, P.rhizo-oryzae and P.pachycaule voucher isolates were not amplified. For the first time, a P. insidiosum isolate from Uruguay, South America, was included in molecular analyzes. By SNP-based multiplex-PCR, it was possible to perform the identification and genotyping of the South American isolates of P. insidiosum, demonstrating similar genetic characteristics of these isolates.

Nucleic Acid-Based Detection of Pythium insidiosum: A Systematic Review

“…The m-PCR assay exhibited 100% detection sensitivity and specificity based on the performance assessment against 53 P. insidiosum isolates and 22 control fungi. The SNP-based m-PCR assay has been used to identify and genotype P. insidiosum isolated from affected animals (primarily horses and dogs) in South America [ 26 ] and Egypt [ 25 ], as well as in the environment in Thailand [ 27 ]. Recently, Kulandai et al [ 35 ] described a duplex-PCR (d-PCR) assay for simultaneously detecting the 18S rRNA (using the primers Pin1 and Pin2) and ITS (using the primers ITSpy1 and ITSpy2 [ 36 ]) sequences of P. insidiosum in corneal specimens from patients with suspected ocular pythiosis.…”

Mefenoxam and pyraclostrobin: toxicity and in vitro inhibitory activity against Pythium insidiosum

“…equine and 1 canine) and 3 standard strains (CBS 119455, CBS 57585 e CBS 77784). All isolates were molecularly identified according to Weiblen et al (2019) (Table 2).…”

Avaliação Da Suscetibilidade in Vitro E Toxicidade in Vivo Do Fitoantifúngico Mefenoxam Frente a Isolados De Pythium Insidiosum

“…Para a realização dos testes de suscetibilidade in vitro, utilizou-se um total de 20 isolados de P. insidiosum, sendo 17 isolados clínicos brasileiros (16 de origem equina e 1 de origem canina) e três cepas padrão (CBS 119455, CBS 57585 e CBS 77784), todos os isolados estão identificados molecularmente, de acordo com Weiblen et al (2019). O mefenoxam (Metalaxyl ® , pureza 99,5%, Sigma-Aldrich Co., St Louis, MO, EUA) foi obtido comercialmente.…”