Phylogeny of Pythium insidiosum by the genes encoding the translation elongation factor (Tef-1α), α and β-tubulin and restriction pattern analysis by Pulse-Field Gel Electrophoresis (PFGE)

Author(s)

Ana Carolina do Prado

• Document type: Master's Dissertation
• Press: Botucatu. 2020-06-30.
• Institution: Universidade Estadual Paulista (Unesp). Instituto de Biociências. Botucatu
• Defense date: 2020-02-28
• Advisor: Sandra de Moraes Gimenes Bosco

Abstract

Pythium insidiosum is the etiological agent of pythiosis, a chronic granulomatous infection, prevalent in tropical and subtropical regions that affects mammals, especially horses, dogs and humans. This is a fungus-like microorganism that has a wide geographical distribution and is very prevalent in South America (equine and canine pythiosis) and Thailand (human pythiosis). Molecular studies have allowed early diagnosis and better understand of phylogenetic relationships, and the pathogen is currently divided into three clades (I, II and III or A, B and C). However, this information is still quite limited for this pathogen and new gene regions could help understanding the evolutionary history of this species. Therefore, this study aimed to establish the phylogenetic relationships among 52 american and asian P. insidiosum isolates by sequencing three new gene regions: the translation elongation factor (Tef-1α), α and β tubulin, in addition to the standardization of the Pulse Filed Gel Electrophoresis (PFGE) technique for this species. The Tef-1α region showed little polymorphic in relation to α and β tubulin, however it separated the strains here studied into two distinct clades, being composed only of strains previously classified as clade III (Asian), and grouped all the American strains next to the clade II strains. The β tubulin gene-based phylogeny separated the isolates into the three expected clades. Regarding α tubulin there was differentiation of isolates in clades I and II, not observing amplification of clade III. The clade II strains were subdivided into two monophyletic clades. We conclude that the results of the phylogeny of the genes here evaluated are consistent with the data previously studied for this species, corroborating the hypothesis of clonal expansion of Asian strains to the American continent. PFGE analyzes are in the phase of standardization of electrophoretic profiles and we believe it may complement the data on the evolutionary relationships of Pythium insidiosum. (AU)

• FAPESP's process: 18/24507-0 - Molecular characterization of Pythium insidiosum by the sequencing of genes coding for Tef-1± and ± and ² tubulin and analysis of the restriction pattern by pulse-field gel electrophoresis (PFGE)
• Grantee: Ana Carolina do Prado
• Support Opportunities: Scholarships in Brazil - Master