Application of PCR-RFLP technique to species identification and phylogenetic analysis of Streptomyces associated with potato scab in Brazil based on partial atpD gene sequences

Abstract

In the present study the atpD gene was evaluated as an alternative molecular marker for differentiation of Streptomyces species causing potato scab and to establish the phylogenetic relationships within this group. PCR-RFLP experiments using HaeIII and CfoI restriction enzymes allowed differentiation of most Streptomyces species, except the genomospecies S. reticuliscabiei and S. turgidiscabies which showed identical profiles between them. Phylogenetic relationships within Streptomyces spp. were also examined by comparing partial atpD gene sequences (858 nt) and the strains were separated in three main groups: Group I, supported by bootstrap values of 79%, comprised of S. scabiei, S. europaeiscabiei, S. stelliscabiei, S. reticuliscabiei, S. turgidiscabies and S. acidiscabies; Group II, supported by bootstrap values of 100%, comprised of S. sampsonii; and Group III supported by bootstrap values of 100%, comprised of S. caviscabies, S. setonii, S. luridiscabiei and S. puniciscabiei. The atpD gene sequence similarity observed among the species in this study ranged from 89.2% to 100%. Twenty-nine Streptomyces isolates obtained from potato growing areas in Brazil were analyzed by PCR-RFLP of atpD gene technique and identified as S. scabiei (15 strains), S. caviscabies/ S. setonii (12) and S. sampsonii (2). Using this tool S. europaeiscabiei strains were also detected from import material from the Netherlands. The presence of genes associated with the S. turgidiscabies pathogenicity island (PAI) was also determined and potato tuber slice assays were performed to confirm the pathogenicity of the strains. The data obtained herein indicated that the PCR-RFLP of atpD gene technique can be used for diagnosis and identification of phytopathogenic Streptomyces strains, representing a rapid and inexpensive tool. Moreover, this gene can be useful for phylogenetic analysis of Streptomyces spp. associated with potato scab.

Biological control of potato common scab by Bacillus amyloliquefaciens Ba01

“…S . scabies strains isolated from scabby potato tubers with nutrient agar or SCNA medium were identified by sequencing 16S rDNA with primers fD1 and rP2 and by PCR-RFLP with amplification of the partial atpD gene with primers atpDPF and atpDPR [ 27 ] (Table A in S1 File ). The sequence results were compared in the Basic Local Alignment Search Tool ( http://blast.ncbi.nlm.nih.gov/Blast.cgi ).…”

Streptomyces hilarionis sp. nov. and Streptomyces hayashii sp. nov., two new strains associated with potato scab in Brazil

“…The results showed that the PCR-RFLP of the atpD gene is an alternative tool in differentiating phytopathogenic species of Streptomyces , corroborating Corrêa et al . [7] [5]. Moreover, the in silico analysis used in this study allowed to differentiate genomospecies like S.…”

“…The PCR-RFLP of the atpD gene was used as an alternative approach for differentiation of phytopathogenic Streptomyces at species level according to Corrêa et al [7]. The PCR products (5 µl) were purified using the GFX PCR DNA and Gel Band Purification kit (GE Healthcare) under conditions specified by the manufacturer, and the sequencing reactions were performed using ABI Prism BigDye terminator cycle- sequencing kit (Applied Biosystems) with the same amplification conditions for MLSA.…”

“…In Brazil, potato scab has been associated with Streptomyces scabiei , Streptomyces caviscabiei and Streptomyces setonii species [7], and more recently Streptomyces brasiliscabiei [8]. In this study, strains IBSBF 2807 T and IBSBF 2953 T , associated with potato scab lesions in Brazil, were characterized using a polyphasic approach involving morphological, biochemical, physiological, pathogenic, and molecular assays in order to shed light into their correct taxonomic position.…”

Genotypic and phenotypic characterization of Streptomyces species associated with potato crops in the central part of Colombia

“…Studies from Latin American countries have focused on the identification and characterization of Streptomyces species causing scab disease, and little is known about the nonpathogenic Streptomyces communities present in potato crops (Corrêa et al, 2015; Lapaz et al, 2017). In Colombia, scab disease causes one of the main phytosanitary problems on potato crops (Instituto Colombiano Agropecuario [ICA], 2011).…”