Single nucleotide polymorphism-based multiplex PCR for identification and genotyping of the oomycete Pythium insidiosum from humans, animals and the environment

Rujirawat, Thidarat; Sridapan, Thanawat; Lohnoo, Tassanee; Yingyong, Wanta; Kumsang, Yothin; Sae‐Chew, Pattarana; Tonpitak, Walaiporn; Krajaejun, Theerapong

doi:10.1016/j.meegid.2017.08.004

Cited by 41 publications

(82 citation statements)

References 31 publications

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“…Based on the multiplex PCR targeting the three SNPs identified in the rDNA region, all the thirty-six South and Central American isolates of P. insidiosum and the five standard strains from Thailand, India and Japan were grouped in their respective clades, as suggested by RUJIRAWAT et al (2017) (Table 1, Figure 1). We observed that the American clinical isolates, grouped in clade I, generated amplicons of approximately 490 and 660bp when using the primers ITS1/R1 and ITS1/R2, respectively.…”

Section: Resultsmentioning

confidence: 99%

“…The multiplex PCR targeting the three SNPs identified in the rDNA region was developed by RUJIRAWAT et al (2017) and has many advantages, such as 100% of sensitivity, and specificity, rapid and cost-effective identification, and genotyping of P. insidiosum. As these authors evaluated only one Brazilian isolate of P. insidiosum in their study, we proposed to evaluate an expressive number of P. insidiosum clinical isolates from South America using this technique.…”

Section: Resultsmentioning

confidence: 99%

“…(5′-TTACCCGAAGGCGTCAAAGA-3′) (RUJIRAWAT et al, 2017). Amplifications were performed according to RUJIRAWAT et al (2017) with modifications.…”

Section: Methodsmentioning

confidence: 99%

“…(5′-TTACCCGAAGGCGTCAAAGA-3′) (RUJIRAWAT et al, 2017). Amplifications were performed according to RUJIRAWAT et al (2017) with modifications. Briefly, in a final volume of 25μL, the PCR reaction contained 1μM of the forward primer ITS1, 0.5μM each of the reverse primers (R1, R2, and R3), 1.5 units of Taq DNA polymerase (Invitrogen), 200μM of each deoxynucleotide, 1.5mM MgCl 2 , 1x enzyme buffer, and 100ng of DNA sample.…”

Section: Methodsmentioning

confidence: 99%

“…Simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers have been used for studies of diversity and relationship in different microorganisms, including P. insidiosum (SUPABANDHU et al, 2007;RUJIRAWAT et al, 2017). The purpose of our study was genotyping American clinical isolates of P. insidiosum using a fast, simple, and low-cost tool based on SNP multiplex PCR.…”

Section: Introductionmentioning

confidence: 99%

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

et al. 2019

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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.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

“…(5′-TTACCCGAAGGCGTCAAAGA-3′) (RUJIRAWAT et al, 2017). Amplifications were performed according to RUJIRAWAT et al (2017) with modifications.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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

et al. 2019

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Lagenidium , Paralagenidium , Pythium , Rhinosporidium , and Uncultivated Paracoccidioides Species (Formerly Lacazia loboi )

Vilela,

Mendoza

2023

ClinMicroNow

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The identification of Lagenidum and Paralagenidium species is only possible using molecular methodologies, such as PCR and sequencing. Based on taxonomic and other morphological characteristics, these four anomalous species, Lagenidium , Pythium , Rhinosporidium , and the uncultivated fungus previously known as Lacazia loboi , were not well understood. Vilela recently reported that the uncultivated fungus developing skin granulomas in dolphin and humans, long believed to be Lacazia loboi , comprised two different Paracoccidioides species: P. ceti and P. loboi . Because few DNA sequences from dolphins with the disease were used in phylogenetic analysis, the separation of the dolphin DNA from the cultivated Paracoccidioides species remained a puzzle. Deep skin scrapings and tissue biopsies from human and dolphins suspected of skin disease due to uncultivated Paracoccidioides species should be submitted to the laboratory.

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Pythiosis

Bosco¹,

Chechi²,

Paz³

et al. 2019

Recent Trends in Human and Animal Mycology

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Single nucleotide polymorphism-based multiplex PCR for identification and genotyping of the oomycete Pythium insidiosum from humans, animals and the environment

Cited by 41 publications

References 31 publications

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

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

Lagenidium , Paralagenidium , Pythium , Rhinosporidium , and Uncultivated Paracoccidioides Species (Formerly Lacazia loboi )

Pythiosis

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