Population genetic analysis of invasive disease agent Lecanosticta acicola

Abstract

Magistritöö Metsamajanduse õppekavalL. acicola on Euroopas karantiinne patogeen, mis põhjustab paljudel männiliikidel varajast okaste varisemist. Ligi kümme aastat peale esmaleidu Eestist on see invasiivne patogeen levinud hajusalt kogu riigis, nakatades peamiselt mägimändi (Pinus mugo), mis on populaarne liik haljastuses. Seni on teadmata patogeeni geneetiline mitmekesisus Eestis, mis võimaldaks hinnata selle populatsiooni elujõulisust, patogeensust ning levikuteid. Magistritöö eesmärkideks on (1) dokumenteerida patogeeni geneetiline mitmekesisus Eestis, (2) hinnata sugulise paljunemise toimumise võimalikkust, (3) uurida patogeeni levikut Eestis ja analüüsida võimalikku antropogeenset mõju sellele ja (4) analüüsida patogeeni populatsioonigeneetilisi erinevusi kahel peremeestaimel: P. mugo ja P. sylvestris. Perioodil juuli 2015 kuni jaanuar 2017 koguti sümptomaatilisi okkaproove, millest patogeen isoleeriti puhaskultuuri molekulaarseteks uuringuteks. Lisaks kasutati isolaate varasematest L. acicola leidudest Eestis. Isolaadid jaotati populatsioonidesse nende geograafilise asukoha järgi. Kokku kasutati populatsioonianalüüsiks 104 isolaati, mida analüüsiti kasutades paarumistüüpide- ja mikrosatelliitide markereid. Andmeanalüüsiks kasutati programme GenAlex, GeneMapper ja Structure. Eesti L. acicola populatsiooni iseloomustab vähene geneetiline mitmekesisus ja suur kloonide osakaal. 104-lt isolaadilt tuvastati 51 erinevat haplotüüpi ning 11 analüüsitud lookuses 43 erinevat alleeli. Uurimuse tulemused viitavad patogeeni mitmekordsele sissetoomisele Eestisse ning inimese mõjule patogeeni levitamisel riigis. Esmakordselt Eestis ja PõhjaEuroopas tuvastati L. acicola harilikult männilt. Uue peremeestaimena maailma jaoks registreeriti Pinus x rhaetica. Eestis on levinum paarumistüüp MAT1-1 ning domineerib patogeeni suguta paljunemine. Kuid kuna populatsioonides esineb samaaegselt mõlemat paarumistüüpi, siis suguline paljunemine on võimalik. Edasise seire, populatsioonide analüüsi ja nakatamiskatsetega on oluline välja selgitada, kas patogeenist kujuneb tõsine oht harilikule männile Põhja-Euroopas ning jälgida patogeeni geneetilise mitmekesisuse muutuseid Eestis, mis viitaksid uute tüvede sissetoomisele või sugulisele paljunemisele.In Europe, L. acicola is a quarantine listed pathogen, which causes serious needle loss in many pine (Pinus) species. Approximately ten years after the first detection of the pathogen in Estonia, it has spread across the country, infecting and affecting mainly mountain pine (Pinus mugo), which is a popular species for greeneries and urban space. So far, there has been no data on the pathogens genetic diversity in Estonia, which would allow for the evaluation of its populations viability, pathogenicity and pathways. The aims of the thesis is (1) to document the pathogens genetic diversity in Estonia, (2) to evaluate the contingency of sexual reproduction in the population, (3) to investigate the spread of the pathogen in Estonia and to analyse the possible anthropogenic activity on it and (4) to analyse the genetic differences of the pathogen in two host species P. mugo and P. sylvestris. During the period of July 2015 to January 2017, samples of symptomatic needles were collected, from which the pathogen was isolated for molecular study. In addition, some earlier isolates of L. acicola in Estonia were used. Isolates were divided into populations according to their geographic locations in Estonia. All together, 104 isolates were used for the population analysis, which were analysed using mating type and microsatellite markers. Data analysis were done using programs GenAlex, GeneMapper and Structure. Estonian L. acicola population is characterised by low genetic variance and high number of clones. From the 104 isolates, 51 different haplotypes and 43 alleles on 11 loci were identified. Results indicate to multiple introduction events in Estonia and it supports human activity in the spread of the pathogen in the country. The first record of the L. acicola in Estonia and northern Europe, was identified on Scots pine (Pinus sylvestris). Pinus x rhaetica was registered as a new host for the pathogen in the world. In Estonia mating type idiomorph MAT1-1 is more common and it indicates that asexual reproduction dominates. However in populations, both mating types occur simultaneously, which means that sexual reproduction is possible. During future monitoring, population genetic analyses and inoculation tests, it is important to determine if the pathogen will develope into a serious threat to the Scots pine in northern Europe and to monitor changes in the pathogens genetic diversity, which would indicate to the introduction of new strain or sexual recombination