Molecular evidence that Verticillium ablo-atrum Grp 2 isolates are distinct from V. albo-atrum Grp 1 and V. tricorpus

Abstract

Verticillium species-specific primers were used in a polymerase chain reaction to differentiate between Verticillium albo-atrum groups and V. tricorpus. Amplification with species-specific primers identified 21 isolates from the 64 V. albo-atrum isolates tested as Grp2. Genome analysis using RAPDs and restriction fragment length polymorphism (RFLP) of the intergenic (IGS) region of the rDNA showed that V. albo-atrum Grp2 isolate were genetically distinct from either V. albo-atrum Grp1 or V. tricorpus, demonstrating a significant differentiation between these species. The sizes of the amplified IGS fragment were different, with Grp1 isolates having a smaller fragment (≈ 2.1 kb) than either Grp2 (≈ 2.3 kb) or V. tricorpus (≈ 2.7 kb). Based on RAPD analysis, the average similarity coefficients between Grp1 and Grp2 were 35% and 34% between Grp2 and V. tricorpus. Multiple correspondence analysis separated the isolates into three major groups corresponding to Grp1, Grp2 and V. tricorpus. Surprisingly, isolates collected from Pisum sativa were distinct from other Grp1 V. albo-atrum isolates. The observed low levels of genetic similarity, the differences in sizes of IGS fragments, IGS-RFLP profiles and the RAPD patterns point to the possibility of Grp2 isolates comprising a different species of Verticillium than those occupied by either V. albo-atrum Grp1 isolates or V. tricorpus.