Genomic and culture-based analysis of Cyclaneusma minus in New Zealand provides evidence for multiple morphotypes

Abstract

Cyclaneusma needle cast, caused by Cyclaneusma minus, affects Pinus species world wide. Previous studies suggested the presence of two distinct morphotypes in New Zealand, ‘verum’ and ‘simile’. Traditional mycological analyses revealed a third morphotype with clear differences in colony morphology and cardinal growth rates at varying temperatures. Genome sequencing of eight C. minus isolates provided further evidence of the existence of a third morphotype, named as ‘novus’ in this study. To further analyse these morphotypes, we predicted candidate effector proteins for all eight isolates, and also characterized a cell-death eliciting effector family, Ecp32, which is present in other pine phytopathogens. In concordance with their distinct classification into three different morphotypes, the number of Ecp32 family members differed, with patterns of pseudogenization in the ‘simile’ morphotype, and some members being found exclusively either in the ‘simile’ or ‘verum’ morphotypes. We also showed that the Ecp32 family proteins trigger cell death in non-host Nicotiana species, and, as previously demonstrated in other plant pathogens, the Ecp32 family proteins in C. minus adopt a β-trefoil fold. These analyses provide further evidence that the three morphotypes might be distinct species that need formal descriptions. Understanding the geographical range of different Cyclaneusma species and variations in virulence and pathogenicity will provide a better understanding of pine needle diseases and enable the development of more durable methods to control this disease.