Phylogenetic and morphological analyses of Pythium graminicola and related species

Abstract

Isolates of Pythium graminicola and related species were differentiated using restriction fragment length polymorphism (RFLP) analyses of the internal transcribed spacer (ITS) regions of rDNA and the cytochrome c oxidase subunit II (COX II) gene. These sequences were used in subsequent phylogenetic analyses. Finally, the phylogenetic placement of species was compared to that determined from morphological characteristics. The 62 isolates tested were divided into seven groups, A-G, based on RFLP analysis of the rDNA-ITS region. In the RFLP analysis of the COX II gene, isolates were divided into groups similar to those based on ITS-RFLP. Groups A and B were each separated into two additional subgroups. Grouping of isolates based on RFLP analyses agreed with the morphological differentiation. Groups A, B, D, E, F, and G were identified as P. graminicola, P. arrhenomanes, P. aphanidermatum, P. myriotylum, P. torulosum, and P. vanterpoolii, respectively. Group C was closely related to group B based on phylogenetic analysis of the rDNA-ITS region and the COX II gene and is similar to P. arrhenomanes. Each of the other species occupied their own individual clades. Although P. arrhenomanes is morphologically similar to P. graminicola, our phylogenetic analyses revealed that it was evolutionarily distant from P. graminicola and more closely related to P. vanterpoolii. Our analysis also revealed that P. torulosum with smaller oogonia is more closely related to P. myriotylum with large oogonia than to P. vanterpoolii, which forms smaller oogonia and is morphologically similar to P. torulosum. P. aphanidermatum with large oogonia and aplerotic oospores was not related to the morphologically similar species P. myriotylum. Results suggest that P. graminicola and related species are phylogenetically distinct, and molecular analyses, in addition to morphological analyses, are necessary for the accurate taxonomic placement of species in this complex.

Molecular Characterization of Pythium Spp. Isolated from Tomato Seedlings in the Syrian Coast

“…Moreover, cultural conditions can influence the variability of morphological characteristics and lead to misidentification of species (Kageyama et al, 2005). The result of failure could be in identification of some characters of Pythium.…”

Characterization of species associated with Pythium soft rot of ginger and evaluation of Pythium Oligandrum as a biocontrol

“…in other studies (Francis et al, 1994;Gómez-Alpízar et al, 2011;Harvey et al, 2000;Kageyama et al, 2005;Matsumoto et al, 2000;Rafin et al, 1995). However, with rapid progress in developing novel molecular tools, PCR-RFLP is considered to be a time consuming approach compared to isothermal amplification of DNA techniques such as LAMP (loop-mediated isothermal amplification) and SMAP 2 (smart amplification process version 2) (Mitani et al, 2007;Notomi et al, 2000).…”

“…intermedium (Allain-Boulé et al, 2004) or intraspecific variation within species like P. myriotylum (Kageyama et al, 2005;Perneel et al, 2006), P. arrhenomanes (Chen and Hoy, 1993) and P. mercurial sp. nov. .…”

“…Hence, Schroeder et al (2013) arrhenomanes and on this molecular evidence, Chen et al (1992) suggested the consolidation of these two species. However, Chen and Hoy (1993) were successful in applying two enzyme sets: Watanabe et al, 2007), the CoxI gene alone Martin and Kistler, 1990) or in conjunction with the CoxII gene (Kageyama et al, 2005). Fukuta et al (2013Fukuta et al ( , 2014 successfully developed a set of six primers, including two loop primers, to specifically detect P. aphanidermatum and P. myriotylum using LAMP.…”

“…(Schroeder et al, 2013), sequencing alternative genes has been deployed. These include nuclear encoded β-tubulin, SSU (small subunit) and LSU (large subunit) Kumar and Shukla, 2006;Lévesque and De Cock, 2004;Liu et al, 2012;Moralejo et al, 2008;Mu et al, 1999;Spies et al, 2011a;Tao et al, 2011), CoxI (cytochrome c oxidase subunit 1) (Kroon et al, 2004;Robideau et al, 2011), CoxII (Garzón et al, 2005b;Kageyama et al, 2005;Kammarnjesadakul et al, 2011;Martin, 2000;Nechwatal et al, 2005;Spies et al, 2011a;Villa et al, 2006) and NADH (dehydrogenase subunit 1) (Kroon et al, 2004;Moralejo et al, 2008).…”