Highly Clonal Structure and Abundance of One Haplotype Characterise the Diplodia sapinea Populations in Europe and Western Asia

Abstract

Diplodia sapinea is a cosmopolitan endophyte and opportunistic pathogen having occurred on several conifer species in Europe for at least 200 years. In Europe, disease outbreaks have increased on several Pinus spp. in the last few decades. In this study, the genetic structure of the European and western Asian D. sapinea population were investigated using 13 microsatellite markers. In total, 425 isolates from 15 countries were analysed. A high clonal fraction and low genetic distance between most subpopulations was found. One single haplotype dominates the European population, being represented by 45.3% of all isolates and found in nearly all investigated countries. Three genetically distinct subpopulations were found: Central/North European, Italian and Georgian. The recently detected subpopulations of D. sapinea in northern Europe (Estonia) share several haplotypes with the German subpopulation. The northern European subpopulations (Latvia, Estonia and Finland) show relatively high genetic diversity compared to those in central Europe suggesting either that the fungus has existed in the North in an asymptomatic/endophytic mode for a long time or that it has spread recently by multiple introductions. Considerable genetic diversity was found even among isolates of a single tree as 16 isolates from a single tree resulted in lower clonal fraction index than most subpopulations in Europe, which might reflect cryptic sexual proliferation. According to currently published allelic patterns, D. sapinea most likely originates from North America or from some unsampled population in Asia or central America. In order to enable the detection of endophytic or latent infections of planting stock by D. sapinea, new species-specific PCR primers (DiSapi-F and Diplo-R) were designed. During the search for Diplodia isolates across the world for species specific primer development, we identified D. africana in California, USA, and in the Canary Islands, which are the first records of this species in North America and in Spain.

First report of Diplodia tip blight on Scots pine in Finland

“…The D. sapinea ITS sequences are deposited in GenBank with accession numbers OP103742-OP103748 (Table 1). The species-specific primers DiSapi-F (3´-CCCTTATATATCAAACTAT-GCTTTGT-5´) (Adamson et al 2021) and Diplo-R (3´-TTACATAGAGGATTGCCTTCG-5´) (Adamson et al 2021) confirmed the species identification. PCR protocol was 0.2 U/µl DNA polymerase (DreamTaq DNA Polymerase, Thermo Scientific™), 1X PCR Buffer, 200 µM dNTP, 0.5 µM primer 1, 0.5 µM primer 2, 10 µl of crude DNA (diluted to 1:50 from the original extracted concentration), and finally the reaction was adjusted to 25 µl with MQ H 2 O. PCR conditions were 95 °C for 5 min, 35 cycles of 95 °C for 30 s, 55 °C for 30 s, 72 °C for 1 min, and 72 °C for 10 min (Adamson et al 2021).…”

Host and abiotic constraints on the distribution of the pine fungal pathogen Sphaeropsis sapinea (= Diplodia sapinea)

“…countries where P. sylvestris is native, suggesting that the fungus is long established in this region (Adamson et al, 2021). On the other hand, the higher intraspecific diversity in populations of North America suggests that S. sapinea may originate from North America, or in Asia or central America (Adamson et al, 2021). However, S. sapinea was not found here to be endophytic in P. rigida, the host species of North American origin.…”

“…(B) Host of records of S. sapinea based on all GBIF records (GBIF all; n = 351), on combined data from GBIF and fungarium specimens at Botanical Garden and Botanical Museum (BGBM) of Berlin before 1910 with a recorded host (GBIF + BGBM < 1910; n = 33), and the subset of specimens for which the identity of S. sapinea was confirmed by PCR species-specific assay (PCR BGBM < 1910; n = 9). countries where P. sylvestris is native, suggesting that the fungus is long established in this region (Adamson et al, 2021). On the other hand, the higher intraspecific diversity in populations of North America suggests that S. sapinea may originate from North America, or in Asia or central America (Adamson et al, 2021).…”

“…influencing the fungus-host dynamics under climate change scenarios when potentially new hosts are encountered. High intraspecific diversity has been revealed within S. sapinea (Adamson et al, 2021) with different virulences between strains (Bußkamp et al, 2021). Conducting similar growth experiments assessing temperature niche parameters and maximum growth rate coupled with niche modeling with isolates of different haplotypes would help predict the future distribution of S. sapinea strains and their potential risk.…”

Diversity, migration routes, and worldwide population genetic structure of Lecanosticta acicola, the causal agent of brown spot needle blight

“…Due to the significant damage it causes, L. acicola is listed as a quarantine pathogen in numerous countries (EPPO, 2022 ) and extra measures for containment have been applied in the European Union, where the pathogen is classified as a regulated non‐quarantine pest (European Commission, 2019 ). Overall, climate extremes, global trade, and failure to implement proper quarantine measures have been commonly considered as essential factors exacerbating the spread of invasive plant pathogens, including L. acicola (Adamson et al, 2018b , 2021 ; Drenkhan et al, 2014b , 2020 ; Fisher et al, 2012 ; Ghelardini et al, 2017 ; Hanso & Drenkhan, 2009 ; Jürisoo et al, 2021 ). Furthermore, climate change, especially warmer winters, has been thought to be one of the main reasons for northwards spread of several forest pathogens (Hanso & Drenkhan, 2013 ).…”