Poljski jasen (Fraxinus angustifolia Vahl), gospodarski i ekološki vrlo značajna vrsta nizinskih šuma Hrvatske, posljednjih nekoliko godina pokazuje sve izraženije simptome odumiranja. Iako je na nekoliko lokacija u Republici Hrvatskoj u krošnjama osutih stabala utvrđena patogena gljiva Hymenoscyphus fraxineus (T. Kowalski) Baral, Queloz & Hosoya, još uvijek nije potvrđena kao primarni uzročnik odumiranja poljskoga jasena u nizinskim šumama već se smatra samo jednim od više različitih biotičkih i abiotičkih čimbenika koji sudjeluju u navedenom procesu. Svrha ovog istraživanja je bila ispitati ulogu ostalih gljiva te gljivama sličnih organizama u okolnom tlu te na korijenovom sustavu i donjem dijelu debla stabala poljskoga jasena kako bi se uz prijašnja istraživanja koja su se uglavnom odnosila na krošnju, dobio sveobuhvatan uvid u ulogu navedenih skupina organizama u odumiranju. Istraživanje je provedeno u prirodnim sastojinama sa simptomima odumiranja na tri različite lokacije, pri čemu su uzimani uzorci okolnog tla, korijena te donjeg dijela debla na ukupno 90 stabala svrstanih prema osutosti krošnje u tri kategorije zdravstvenog stanja. Iz navedenih su uzoraka izolirani miceliji gljiva i njima sličnih organizama koji su potom identificirani molekularnim metodama. U istraživanju je dobiveno ukupno 1497 izolata gljiva i njima sličnih organizama koji su molekularnim II analizama svrstani u 80 taksona, te prema načinu života i ishrane kategorizirani kao saprotrofi, endofiti i paraziti. Utvrđena distribucija identificiranih taksona s obzirom na različite kategorije uzorka, kategorije zdravstvenog stanja stabala te geografske lokacije uzorkovanja je pokazala specijalizaciju samo za određeni tip biljnog tkiva i supstrata. Izračunata je raznolikost (bogatstvo) vrsta pomoću Shannonovog indeksa raznolikosti te indeksa ujednačenosti, čije se vrijednosti nisu statistički značajno mijenjale za zajednice na stablima različitog zdravstvenog stanja. Statističkim je analizama utvrđeno kako je većina identificiranih taksona (77) jednako distribuirana na stablima različitog zdravstvenog stanja, dok je manji broj onih (69) koji su jednako distribuirani na različitim geografskim lokacijama uzorkovanja. Od značajnijih biotičkih čimbenika koji sudjeluju u odumiranju poljskoga jasena su na korijenu i bazi debla stabala utvrđene vrste Hymenoscyphus fraxineus, Armillaria spp., Ganoderma adspersum, Fusarium solani i Diaporthe cotoneastri. U tlu odumirućih sastojina nije potvrđena prisutnost patogenih vrsta roda Phytophthora. S obzirom na statistički značajnu povezanost razvoja nekroza na deblu i korijenu te osutosti krošnje, navedeni se parametar pokazao dobrim pokazateljem zdravstvenog stanja stabala.
|Abstract (english)|| |
Narrow-leaved ash (Fraxinus angustifolia Vahl), ecologically and economically very important species in lowland forests of the Republic of Croatia, has been showing more prominent symptoms of dieback. According to the crown defoliation monitoring data as a part of the ICP Forests program, narrow-leaved ash is currently the most damaged forest tree species in the Republic of Croatia, with more than 70% of trees with significantly defoliated crown in 2016. Although has the pathogenic fungus Hymenoscyphus fraxineus (T. Kowalski) Baral, Queloz & Hosoya been determined in the crowns of diseased trees in several locations, it has still not been confirmed as the primary cause of narrow-leaved ash dieback in the lowland forests and it is rather considered to be one of the multiple factors affecting the process. Goal of this research was to investigate the role of other fungi and fungus-like organisms present in the adjacent soil, tree roots and stem collars in the decline of this tree species. IV Research has been conducted at three different locations in the natural stands affected by dieback during the growth period in 2016. Trees were categorized according to the crown defoliation in three different categories of health condition: 1) healthy - with crown defoliation < 25%; 2) with impaired health status - crown defoliation 26-60%; 3) declining - crown defoliation 61-99%. From total of 90 trees, 30 in each health category, samples of adjacent soil, root and stem collar were taken for the isolation of fungi and fungus-like organisms using conventional plating on artificial media method for the plant tissue and three different methods for the soil (soil dilution plate, soil plate and soil baiting method). Isolated mycelia were sorted into morphotypes according to their morphological characteristics. From each morphotype one to four representative isolates were chosen for the molecular identification of the species. In the second part of the research, 30 extra soil samples were taken from the vicinity of the part of already examined trees in order to detect the presence of pathogenic Phytophthora species using the soil baiting method and molecular methods for the species identification. All of the soil samples, 86% of root and 85% of stem samples gave fungal growth. In average, three or four different taxa were found per tree, depending on the type of the sample. Research has resulted with total of 1497 isolates sorted into 80 taxa, of which 59 were identified to the species level. Identified taxa were categorized according to their known ecology noted in the various literature into endophytes, saprotrophs and parasites. Most of the taxa were found in just one sample type, indicating their specialization for different substrates. In contrast, most of the taxa were identified in all three health categories and sampling locations. Species diversity in different sample types, shown by Shannon diversity and Evenness indices, did not significantly differ among different health categories in observed locations. For the most taxa, there was no statistically significant difference in their distribution in different health categories. Analysis of taxa distribution among different sampling locations revealed significant difference for 11 taxa, three on stems, six on roots and two in soil, which can indicate different habitat conditions or temporal phases of the dieback process in the observed localities. In order to evaluate does crown defoliation reflect the health status of tree roots and V stems, connection between health category and presence of necroses in plant tissue was tested and the results justified its use for this purposes. Results of the second part of the research did not confirm the presence of pathogenic Phytophthora species in soils of declining stands, excluding them so far as the contributing factor to the decline. Results of this research point out several conclusions. There is high abundance and diversity of taxa of fungi and fungus-like organisms in the soil, tree roots and tree stems in the declining narrow-leaved ash stands, where number of them are officially reported for the first time in the Republic of Croatia, and some of them could be completely new to science since they were not identified to the species level. In general, approximately equal numbers of isolates appointed to endophytes, saprotrophs and parasites are present in soil and trees. Communities of fungi and fungus-like organisms differ more among different sample types, since larger number of taxa seems to be specialized for specific substrate, whereas community compositions in different health categories and sampling localities are relatively similar. The fact that there was no significant difference in distribution of the most taxa in different health categories indicates that taxa present in healthy tissues tend to be primary colonizers of the tissue of impaired health due to some other factors. Although some of the taxa can be considered as opportunistic pathogens based on their equal frequency in tissues of different health categories and assertions found in literature, this assumptions have to be further investigated in order to confirm them. Among the identified taxa, ones with the most prominent role in the narrow-leaved ash decline in the roots and stems of the trees are considered to be Hymenoscyphus fraxineus, Armillaria spp., Ganoderma adspersum, Fusarium solani and Diaporthe cotoneastri.