Climatic changes are supposed to negatively affect forest ecosystems in Central and Southern Europe. The severe impact of elevated air temperature and prolonged summer drought periods predicted by dramatic climate change scenarios, can lead not only to migrations of different tree species, but also to the forest decline, and even mass mortality of trees (Stojanović et al., 2013). According to projections of Stojanović et al. (2014), at the territory of Republic of Serbia pedunculate oak (Quercus robur L.) was marked as the most affected tree species to upcoming climate perturbations. The decline of pedunculate oak trees can cause great economical losses, since the forests of this species are considered as the most valuable in Serbia. Although oaks are known as slow growing species with a long- life span, they exhibit high sensitivity towards various abiotic (e.g. drought and heat stress) and biotic (e.g. powdery mildew has been described as is one the most threatening) stress factors, which often act synergistically in natural environments.
Drought and elevated temperatures (heat) have been described as two synchronized consequences of climate change, jointly initiating oxidative stress in plants (Wahid et al., 2007). Likewise, powdery mildew fungi (Ascomycota: Erysiphales: Erysiphaceae) represents an important group of obligate, biotrophic fungal pathogens of European oak trees, that drastically reduce leaf lifespan, functional leaf area, net photosynthesis and carbon assimilation, thereby affecting secondary metabolism, involved in plant defense (Hajji et al., 2009).
Oxidative stress represents imbalance between generation of reactive oxygen and nitrogen species (ROS and RNS) and antioxidant defense mechanisms (Štajner et al., 2013). Besides expressing highly detrimental effects in cell by causing process of lipid peroxidation, oxidation of proteins and DNA mutations, ROS play essential role as ubiquitous messengers of stress responses and as a signaling molecules in adaptive processes to both biotic and abiotic stresses. On one side, some of the ROS and RNS, such as NO and H2O2 are capable to initiate stomata closure thus limit transpiration and water loss and prevent the negative effects of drought (Agurla et al., 2018) while on the other hand these molecules play an important role in conveying and amplifying signals during biotic stress (e.g., beneficial and pathogenic microbes, fungi, insects, other herbivores).
During abiotic and biotic stress, to neutralize harmful effects of ROS and oxidative stress, plants activate whole plant defense machinery consisted of antioxidants and antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APx), guaiacol peroxidase (POD), glutathione reductase (GR), etc. Furthermore, drought and heat stress up-regulate expression and activity of antioxidant enzymes, as well as enzymes involved in biosynthesis of phenolic compounds, such as phenyl-alanine ammonium lyase (PAL) or enzymes in charge of biosynthesis of osmolytes such as proline (PRO) or glycine betain (GB) (Laxa et al., 2019; Ashraf and Foolad, 2005).
One of the most remarkable group of biomolecules regarding the improvement of plants’ resilience to abiotic and biotic stress factors is definitely the group of polyamines (PAs), that exert different effects that help plants to deal with stressful conditions due to their antioxidant, osmoprotective and antimicrobial activity against plant pathogens (Romero et al., 2018). Polyamines are low molecular-weight organic polycations displaying high biological activity. These universal multifunctional regulators of physiological processes manifest distinct anti-stress protective action and improve resistance to drought, heat, salinity, pathogens as well as heavy metals in plants that over-express PAs biosynthetic genes or accumulate some of specific PAs (Kebert et al., 2016; Wen et al., 2008). They can also regulate genome activity, cell division and expansion, plant growth and development embryogenesis, leaf senescence, abiotic and biotic stress responses and infection by pathogenic fungi and viruses (Alcázar et al., 2010). The common PAs in plants are spermidine (Spd), spermine (Spm) and their diamine precursor, putrescine (Put). In plants, two different routes of PAs biosynthesis are known: the ornithine decarboxylase (ODC, EC 220.127.116.11) pathway or the arginine decarboxylase (ADC, EC 18.104.22.168) pathway via several intermediates. Polyamine degradation in plants is catalyzed by the two oxidative enzymes: copper-containing diamine oxigenase (DAO, EC22.214.171.124) and flavoprotein-dependent polyamine oxigenase (PAO, EC 126.96.36.199.) (Moshou et al., 2008). Evidences point to an interplay between polyamines with ROS generation and NO signalling in ABA-mediated stress responses, therefore PAs participate in stomata closure (Yamasaki and Cohen, 2006). Due to polyamines ubiquitous nature, plants interaction with both beneficial (including mycorrhiza) and pathogenic microbes is followed by changes in PAs metabolism in the plant as well as in the microbes (Jiménez-Bremont et al., 2014). PAs act as antioxidants, but they also exhibit prooxidative properties since during their catabolism they generate ROS as a by-product (i.e. H2O2), which commonly happens during plant’s infection with pathogenic fungi, since H2O2 acts as a defense signaling molecule (Kim et al., 2013). Spermidine is especially known for activation of hypersensitive reaction (HR) related genes as well genes that are important for cell redox homeostasis, protein metabolism and plant defense. Sannazzaro et al. (2004) emphasized significant role of PAs in initial stages of plant infection with mycorrhiza fungi since different PAs were discovered in the spores of AMF. The stability of the entire forest ecosystem and the carbon balance under climate change depends on the trees’ intimate root-soil interaction with soil microbes, especially with mycorrhizal fungi.
Mycorrhizal fungi make mutualistic association with more than 90% of plant species and represent the key players in carbon dynamics and carbon fluxes among plants, soil and the atmosphere (Simard and Austin, 2010). Mycorrhizal fungi are also well documented to improve plants tolerance to unfavorable abiotic stress factors such as heat, drought, salinity or presence of heavy metals, as well as to boost plants immunity and increase resistance to pathogens and provide other ecosystem services (Kumar et al., 2017; Kivlin et al., 2013). Considering the fact that the small and profuse hyphae have 60 times more absorptive area than fine roots, during drought stress plants employ strategy to invest their photosynthate carbon (around 20%) in the development of hyphae rather than fine roots (Simard et al., 2002). Through complex network of mycelium mycorrhizal fungi facilitates communication between trees in the forest and have a pivotal role in linking aboveground and belowground components of biogeochemical cycles.
Interestingly, one mode of action of mechanism how mycorrhiza fungi alleviate drought and heat stress in plants is related to their ability to boost antioxidant defense systems (Marulanda et al. 2007; Ruíz-Sánchez et al., 2010). Modulation of antioxidant enzymes by mycorrhiza was reported in black locust Robinia pseudoacacia exposed to drought, where mycorrhizal plants had higher SOD, POD, CAT, APX, and GR activities and over-expressed transcript levels of Cu/Zn-SOD, APX and GR (He at al., 2017). Similarly, seedlings of drought-resistant Elaeagnus angustifolia colonized by Rhizophagus irregularis showed notably higher activities of superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX) in leaves of mycorrhizal seedlings during salinity treatment compared to those of the non-mycorrhizal seedlings (Chang et al., 2018). Nevertheless, the mechanism of how mycorrhizal fungi alleviate drought stress in different woody plant species is still not completely explained and further investigation on this topic are necessary. Therefore, new findings that are going to be obtained through the MYCOCLOMArt would opened new perspective and offered solutions for prominent problem of Serbian oak forests which are threatened by abiotic and biotic stress factors.
During mycorrhiza establishment it comes to modulation of plant defense responses and molecular reprogramming, which leads to an effective activation of the plant immune responses and expression of defense genes, very similarly to Induced Systemic Resistance (ISR) (Cameron et al., 2013). This kind of induced resistance is called mycorrhiza induced resistance (MIR) and it has been reported that MIR results in an active suppression of components of salyciclic acid (SA)-dependent defense pathways, causing systemic activation of jasmonate (JA)-dependent defense pathways (Pozo and Azcon-Aguilar, 2007; Pieterse et al., 2012). To escape infection by pathogens, plants have evolved a battery of defense mechanisms mediated by multiple pathogen-specific signal transduction pathways (van Kal, 2006). Beside defense mechanisms that are mediated and regulated by the plant hormones, plants during pathogenesis exhibit distinct differences of enzymes involved in polyamine and polyphenolic metabolism at both transcriptomic and metabolic level (Zhang et al., 2015). In respect to that, resistant genotypes of oat (Avena sativa L.) showed increased levels of polyamines, while exogenous application of polyamines onto leaf surface caused an increased resistance towards the powdery mildew in oat (Montilla-Bascón et al., 2014). Several fold increase of free polyamines, significantly increased polyamine oxidase (PAO) activity and a moderate increase in ornithine decarboxylase (ODC) activity were detected in leaves of barley infected by powdery mildew compared to the non-infected controls (Coghlan and Walters, 1990). Noteworthy, volatile organic compounds (VOCs) emitted by plants represent another key factor in plants’ defense and have pivotal role, not only to attract pollinators, but also to repel, deter or kill both pathogens and herbivores, and they could even serve as a markers of plant disease (Moore et al., 2014). Diverse groups of molecules belonging to VOCs demonstrate multitude of functions, where one of the main functions could be that VOCs are controlling multitrophic interactions, both belowground and aboveground and contribute to overall plants fitness (Bezemer et al., 2005). Several authors documented that during MIR, mycorrhizal fungi is capable to alter amounts and composition of volatile organic compounds blends emitted by plants, which significantly modulates susceptibility of host plants towards different pathogenic fungi including powdery mildew (Rapparini et al., 2008; Asensio et al., 2012; Thirkell et al., 2017). However, little is known about the mechanisms how VOC signals are connected with fungal infections, since majority of studies have been focused on plant–insect interactions (Schaub et al., 2010).