Analysis site

The study urban area includes c. dos km dos away from kinds-steeped lifeless calcareous grassland from inside the west Estonia (°N, °E). The fresh grasslands set-up over the years due to enough time-label comprehensive government, notably grazing by the residential pet, event hay and you may reducing shrubs and you can woods to own electricity. , 1999 ). This new grasslands generally had a max juniper (Juniperus communis) protection out of 29%, but are now much more overgrown which have Scots oak (Pinus sylvestris; around 80% cover).

Sampling construction

Testing are presented into the from the websites in which consistently handled (mown) unlock grassland stands (GR) were enclosed by really stands away from varying levels of over growing: transformation grasslands (TR) and you may young pine woods (FO). The three successional stages (GR, TR, FO) setting a-temporal succession of regeneration sequence level c. 60 year, and are also characterised by the a gradual over growing out of unlock grasslands by the shrubs and you will trees, that’s typical getting abandoned calcareous grasslands regarding the data part (Neuenkamp et al., 2016 ; Supporting Recommendations Fig. S1). Into the transformation grasslands government stopped c. 30–۳۵ yr in advance of testing and you will grasslands were partly overgrown of the pine and you may juniper; younger pine woods have been previous grasslands in which government stopped c. 60 year in advance of sampling and you may which were entirely overgrown because of the pine and you will juniper. Web site are defined as a location where the about three successional values was receive inside fifty m of each and every most other (GR–TR–FO). Altogether, 25 internet was basically sampled. A randomly discovered step 1 ? step 1 m patch are created in each successional phase (GR, TR, FO) of each webpages, causing a maximum of 75 plots (twenty five internet ? around three successional stages). The second services were discussed for every patch: the field level bush people, Are yeast organizations (in-plant origins, and rhizosphere crushed) and you may ecological parameters.

Research range

For every 1 ? step 1 yards plot, all hledání profilu the adult hub vascular plant types about crushed-layer area was in fact registered and their abundance are estimated while the payment shelter (Peet & Roberts, 2013 ). On top of that, percentage shelter quotes from shrubs and you may woods within a good ten ? ten meters city as much as for each and every spot was registered. Hereafter, the ground-layer bush society is known as ‘plant community’ as well as analyses is presented with this particular society until expressed if not.

An excellent ten ? ten ? 10 cm surface center are gathered throughout the middle of each and every plot and you can plant root was indeed split regarding the surface, dehydrated to own twenty four h from the 50°C and you will stored lifeless for additional unit analysis regarding sources-colonising Are fungus. Have always been yeast assortment in-plant origins ended up being analysed out-of mixed resources trials. Origins within the per attempt have been surface having fun with h2o nitrogen and you will thoroughly blended because the explained of the Garcia de- Leon et al. ( 2016 ). A great subsample out-of 70 mg of each and every blended sources test is useful unit analyses.

Bush groups was indeed revealed at a more impressive spatial scale weighed against In the morning fungal communities, as this scale would be to as much as echo the plant root neighborhood seized by the a great ten ? 10 ? 10 cm soil key. Small-scale bush range during the grasslands is actually highest below ground than just a lot more than soil (Hiiesalu ainsi que al., 2014 ), partially as the roomy supply system and you will much time rhizomes of a lot grassland vegetation connect the newest root within a localised crushed core that have significantly more than-surface organs growing numerous ).

To characterise AM fungal diversity in the soil, 10 g of soil was collected from each soil core, dried with silica gel and stored airtight at room temperature. A second subsample of 10 g of soil was collected from each soil core for measuring the concentration of an AM fungal neutral lipid fatty acid fungal abundance in the soil. This second soil sample was freeze-dried for 15 h at ?80°C and stored airtight at room temperature. The remainder of each soil core was air-dried for analysis of soil chemical properties (pH_KCl, N_tot, P, K, Mg, Ca, C_org) following International Organization for Standardization (ISO) protocols (see Methods S1).