Cryphonectria parasitica, the causal agent of chestnut blight, has been present in Slovenia since at least 1950. To improve understanding of its diversity, 254 isolates of the fungus from 11 Slovenian populations were sampled. Fifteen vegetative compatibility (vc) types were identified. The dominant vc type was EU-13, comprising 40AE1% of all isolates tested, followed by EU-1 (19AE7%), EU-2 (12AE2%) and EU-12 (9%). The vc type diversity in the most diverse population sampled in Slovenia was higher than in the populations found previously in northern Italy and Croatia. Both mating types and perithecia were observed in surveyed populations. Natural hypovirulence was found in six out of seven populations tested, with frequencies ranging from 72AE2% in the population sampled near the Croatian border to 11AE1% in the population sampled near the Austrian border. All identified hypoviral isolates (21) belonged to the Italian subtype of Cryphonectria hypovirus 1 and were closely related to the hypoviruses found in other European countries. Despite the high vc type diversity, incidence of hypovirulence was also high, indicating widespread natural biological control of the disease.

Candidatus (Ca.) Phytoplasma ulmi', the causal agent of elm yellows, was found widely distributed across elm populations in Croatia, infecting Ulmus laevis and Ulmus minor. Especially high prevalence of the infection, approximately 75%, was detected in U. laevis, but more than half of the trees were symptomless. 'Candidatus Phytoplasma solani' and 'Ca. Phytoplasma asteris' were also detected. The latter could possibly represent a new 16SrI subgroup, most closely related to 16SrI-B. Diversity of 'Ca. Phytoplasma ulmi' in Croatia was determined by sequencing of the 16S rRNA gene, ribosomal protein genes rpl22 and rps3, secY and secY-map genes, in 62 phytoplasma isolates. Phylogenetic analysis indicated that Croatian isolates share a common origin and are closely related to strains of 'Ca. Phytoplasma ulmi' from southeastern Europe. However, comparative sequence analysis revealed mutations at positions where variability has never been detected before, including positions within sequences unique to 'Ca. Phytoplasma ulmi' in the 16S rRNA gene, rpl22, rps3 and secY. New genotypes were identified based on the sequenced genes. This study points to a significantly higher genetic diversity than previously reported, and a necessity to revise the formal description of this phytoplasma species and to include newly discovered characteristics.

No abstract

The ascomycete fungus Cryphonectria parasitica, causal agent of chestnut blight, is probably one of the best known invasive fungal pathogens in forests of Europe and North America. Mycovirus that reduces virulence of C. parasitica can be used as a biocontrol agent of the chestnut blight. However, anastomosis-mediated virus transmission is limited by a vegetative (in)compatibility (vc) system involving at least six known diallelic vic genetic loci. This study looked at vegetative compatibility (vc) diversity in two populations of C. parasitica in Croatia. For that purpose, a PCR assay was validated and implemented using already known/published and newly designed primers for amplification of six known vic loci. The vc genotypes determined by PCR for 158 C. parasitica isolates investigated in this study were in complete agreement with the vc genotypes determined by pairwise co-culturing of the same isolates, revealing the specificity and accuracy of the PCR-based molecular vic genotyping assay. Twenty-six unique vc genotypes were found among 158 isolates, and 19 vc types per population, which makes Croatian C. parasitica populations among the most diverse in Europe regarding the number of vc types and genetic diversity. Low values of multilocus linkage disequilibrium suggest sexual reproduction as a major contributor to high C. parasitica genetic diversity in studied populations.

Some CHV1 strains belonging to the Italian subtype have similar hypovirulent effects on C. parasitica to those belonging to the French subtype. Furthermore, chestnut susceptibility and recovery could be influenced by the response of chestnut trees to particular hypovirulent C. parasitica isolates, and virus-fungus-chestnut interactions could have significant implications for the success of chestnut blight biocontrol. © 2016 Society of Chemical Industry.