DIGITALNA ARHIVA ŠUMARSKOG LISTA
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MATERIALS AND METHODS
MATERIJALI I METODE
Sampling – Uzorkovanje
In order to properly evaluate individuals for oxidative stress screening tests, we first selected them according to phenophase [early (var. praecox) vs late (var. tardissima)] and physiological groups (vital or damaged). The main criterion for determining the affinity of trees to one of the physiological status groups (vital or damaged) was the degree of crown damage, which was assessed according to Dubravac et al. (2011): trees with defoliation above 25 % were considered significantly damaged trees, while trees with canopy damage up to 25 % were considered undamaged or vital trees. Presence of damage caused by insect pests and pathogens, such as damage to stems caused by xylophagous and phloemophagous insects (e.g., Scolytus spp., Agrilus spp., Cerambyx cerdo, Zeuzera pyrin) (Marković and Stojanović, 2011; Mihajlović et al., 2015), and necrotic lesions on stems caused by Diplodia seriata (Zlatković et al., 2018) were used as an additional criterion in tree selection, especially in discrimination between vital and damaged trees. Crown condition was assessed in the same year as sampling. Leaf samples from 42 individuals [21 individuals of Quercus robur var. praecox (11 vital + 10 damaged) and 21 individuals of Q. robur var. tardissima (11 vital + 10 damaged)] were collected from seed stand located in the Northern part of Serbia (N 45° 00‘12.66”; E 019° 08‘50.4”) during springtime in April and May 2013. The altitude of the examined stand was ranging from 81 to 83 m above sea level (a.s.l.). The geological substrate was defined as alluvial sand sediments with mostly gley soils ranging from riparian black soil and black meadow soil to brown forest soil. Hydrological conditions were characterized by the absence of flooding and soil moisture was strongly dependent on fluctuations of groundwater table levels. Climate in the sampling area is defined as temperate continental to modified continental, with semi-humid and warm summers (Kottek et al., 2006).
Plant material – Biljni material
Phenological varieties [early (var. praecox) vs late (var. tardissima)] and physiological group (vital or damaged) were the observed variables in tested individuals of pooled early oak with vital (PEOV), pooled early oak with damaged (PEOD), pooled late oak with vital (PLOV) and pooled late oak with damaged (PLOD) canopies. Each pooled group comprised 10 or 11 samples in total, measured in triplicates in order to achieve optimal statistical significance for each group (for more, see Statistics sub-section). Collected leaves were immediately put and labeled on aluminum foliage, preserved on dry ice, and transported to the Laboratory for Chemical and Biochemical Analysis at the Institute of Lowland Forestry and Environment (ILFE) for further processing.
Oxidative stress screening tests – Testovi praćenja oksidacijskog stresa
Two grams of oak leaves were homogenized in a cooled mortar with pestle with 10 ml of phosphate buffer (0.1 M KH2PO4, pH=7), centrifuged for 10 min at 11000 rpm and supernatant was used for further oxidative stress screening test (Table 1), as per authors instructions. All analyses were determined spectrophotometrically in microwell-plates by Multiplate reader MultiScan GO (Thermo Scientific, Germany).
Statistics – Statistika
Values of the biochemical parameters were expressed as mean value ± standard error of determinations made in