Folia Microbiologica

, Volume 64, Issue 2, pp 161–170 | Cite as

A highly efficient electrophoretic method for discrimination between two Neoscytalidium species using a specific fungal internal transcribed spacer (ITS) fragment

  • Mohammed Baqur S. Al-ShuhaibEmail author
  • Hawraa N. Al-Kaaby
  • Sabah L. Alwan
Original Article


Neoscytalidium (or N.) dimidiatum and N. novaehollandiae are two aggressive plant pathogenic species that affect several agricultural crops. Early detection and identification of these fungi are of critical importance to bring about the effective minimization to the threat they pose to the infected plants. Herein, two species of Neoscytalidium were rapidly discriminated by utilizing the rRNA internal transcribed (ITS4-5.8S-ITS5) PCR primers. A total of 100 isolates of Neoscytalidium species, which were isolated from Iraqi canker-infected fig trees, were included in this study. Two discrete electrophoretic PCR bands were observed in Neoscytalidium isolates—A-variants were about 546 bp, while B-variants were about 993 bp in length. The comprehensive phylogenetic analysis of both DNA variants revealed that A-variants resided between N. novaehollandiae and N. hyalinum, while B-variants were closely related to N. dimidiatum. Furthermore, the highly specific re-constructed tree of both electrophoretic variants demonstrated that B-variants share a high similarity with N. novaehollandiae. Additionally, the secondary structures for both variants were predicted computationally to reveal the structural patterns that each variant follows. In conclusion, a small rRNA locus comprising 22 nucleotides that differs in the two variants is potentially responsible for this species-specific classification. The main divergence in the amplified loci led to the classification of these fungal variants into two main species, namely N. dimidiatum and N. novaehollandiae, demonstrating that the amplification by ITS4–ITS5 rRNA fragment is a beneficial strategy that can be employed for the assessment of Neoscytalidium diversity in the natural ecosystems.

Supplementary material

12223_2018_641_MOESM1_ESM.pdf (149 kb)
Supplementary Fig. 1 The main morphological and microbiological characterization of Neoscytalidium species that isolated from cankered stems of fig trees in the middle Euphrates region of Iraq. A, Stem canker symptoms caused by Neoscytalidium species observed in the fields. B, Morphological characterization of Neoscytalidium species in the Petri dish. C, Conidiogenous cells in the branched and brown hyphae. D, Mature conidia. (PDF 148 kb)
12223_2018_641_MOESM2_ESM.docx (17 kb)
Supplementary Table 1 (DOCX 17 kb)


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Copyright information

© Institute of Microbiology, Academy of Sciences of the Czech Republic, v.v.i. 2018

Authors and Affiliations

  1. 1.Department of Animal Production, College of AgricultureAl-Qasim Green UniversityAl-QasimIraq
  2. 2.Department of Plant Protection, College of AgricultureUniversity of KufaKufaIraq

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