Degradation of mycorrhizal fungal communities associated with cork oak and understory vegetation by the anthropogenic factors
Article Category: Short Communication
Pubblicato online: 22 set 2022
Pagine: 184 - 194
Ricevuto: 28 set 2021
Accettato: 04 mag 2022
DOI: https://doi.org/10.2478/ffp-2022-0018
Parole chiave
© 2022 Hana Ksentini et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
The cork oak (
They play a major role in the landscape formation and are of great ecological and socio-economic value (soil protection, cork production, logging, grazing space, mushroom production, aromatic and medicinal plants, ecotourism, etc.) for Mediterranean populations (Eriksson et al. 2017; Ksentini et al. 2020). However, declining regeneration of cork oak was observed, notably in Algerian cork oak populations (Bouhraoua 2008). The cork oak health status degradation is an ancient problem, going back to the end of the 19th century (Bouhraoua 2008). However, it was not until these years that cork oak mortality became a matter of concern for the future of these ecosystems. In fact, cork oak decline was defined as a gradual process involving several factors acting sequentially or simultaneously and leading directly or indirectly to a gradual tree weakness (Ben Jamaa 2011), followed by a decrease in its defensive ability and the establishment of favourable conditions for infection by biotic agents (De Sousa et al. 2008).Cork oak decline has multiple impacts at aboveground and belowground levels, strongly affecting resilience and productivity of cork oak forests. In addition to symptoms directly affecting the cork oak itself, cork oak decline was shown to negatively impact key elements of ecosystem functioning, notably, the soil microbiome(Lancellotti and Franceschini 2013). Indeed, deterioration of the condition of cork oak forests has been worsened by the mentioned anthropogenic activities and climatic disturbances, combined with a lack or scarcity of natural cork oak regeneration (Maghnia et al. 2019).
Among soil microbiota, mycorrhizal fungi are considered as key components of the sustainable soil–plant system (Dickie et al. 2004). They are generally considered to be ‘soil engineers’ that are particularly involved in the productivity and stability of ecosystems and agrosystems. Ectomycorrhizal (EcM) symbiosis is a mutualistic relationship in which the fungal hyphae surround and grow between the cortical cells of specialised fine plant roots. EcM fungi play an important role in the regeneration of forests by facilitating nutrient and water uptake by their host plants (Smith and Read 2008). However, in contrast to arbuscular mycorrhizal fungi (AMF), EcM symbionts are not uniformly distributed within ecosystems in terms of presence/absence, abundance or community composition (Boudiaf et al. 2013).
Several studies reported the strong impact of cork oak degradation on soil microbiome, with the main focus being on the about the role and diversity of soil EcM fungal communities establishing a symbiosis with cork oaks (Ksentini et al. 2017; Boudiaf et al. 2013; Lancellotti and Franceschini 2013). However,
The effects of some shrub species on cork oak recruitment and survival have also been observed. For example,
Therefore, the objectives of this study were to determine the impact of degradation by anthropogenic pressure on (i) the rate of mycorrhizal colonization of root-associated fungal community of
The study was carried out in the forest ‘Brabtia’ (36°52′09.27″ N, 8°20′16.58″ E) located in northeastern Algeria at an altitude of 30 m (Fig. 1).The climate is sub-humid with a hot and dry season between May and November, while the rest of the year, it is humid and rainy(Sarri et al. 2014). The average annual rainfall is 225 mm. The Brabtia forest is composed of various shrub species such as
Figure 1
Location of the cork oak forest ‘Brabtia’ and the study stations (Diaf et al. 2019). ND – non-degraded station, D – degraded station
Two stations within the Brabtia forest (Fig. 1), 17 km apart, have been chosen: one (ND) non-degraded and the other (D) degraded by various anthropogenic pressures (harvesting cork oak, cutting wood, grazing). The non-degraded station was chosen according to the following criteria: (i) less accessible to grazing livestock, (ii) lower frequency of fires and (iii) high density of cork oak trees and their understory shrub vegetation. The degraded station was chosen according to the following criteria: (i) decrease in the density of the cork oak and its understory shrubs vegetation to about 50 trees and (ii) higher repeated fires.
At each station, soil samples of cork oak (CO), Cistus (CI), lavender (L) and thyme (T) were collected from the foot of six trees (2 kg/tree) at a depth of 20 cm. After a composite soil sample was collected in every station, very fine roots of cork oak, CI, L and T more likely to be mycorrhizal and more easily observable under a microscope were collected at the same time as the soil. Soils were air-dried for further physicochemical analyses, assessment of the AM propagules in the soil, spore isolation and morphological identification.
Rhizosphere soil of cork oak (
EcM colonization assessment was made by counting the presence or absence of colonized root tips under a stereomicroscope according to the method of Brundrett et al. (1996). The percentage of root colonization was determined for each sample by examining three hundred 1cm root pieces and was calculated using the following formula:
The sampling of the same roots, performed as described above, was cleared and stained for assessment of mycorrhizal colonization using the technique of Phillips and Hayman (1970), and the percent of AM colonization was quantitatively estimated following Trouvelot and Kough (1986). Samples were processed through the following stages: soil washing, cutting into segments of 1.5 cm, hot cleaning with KOH 10% (15 min), immersed into a solution of HCl 20% (10 min) and then staining with 0.03% trypan blue solution at 90°C. Root observations were done made in five replicates for 30 root fragments of 1 cm length, which were then placed between the slide and cover in a drop of glycerol. The annotation was made using the software MYCOCALC (
The mycorrhizal potential of the rhizosphere of four tested species (cork oak, lavender, thyme, and cistus) collected from both stations (degraded and non-degraded) was measured by the most probable number (MPN) of AM propagule (Porter1979), which consists in making a series of successive soil dilutions and determining the limiting dilution at which no AMF propagules can be detected. For this, the collected soil was mixed with sterilised soil to make five serial dilutions (1/10) and five replicates of 50 g per pot. One pre-germinated clover (
Spores were extracted for the rhizosphere of the four tested species (cork oak, lavender, thyme, and cistus) collected in winter, 201 from both stations (degraded and non-degraded), using the wet sieving method (Gerdemann and Nicholson 1963). In a 1 L beaker, 100 g of each composite sample of the soil was immersed with 0.5 L of tap water and stirred for 1 min with a spatula. After 10–30 seconds of decanting, the supernatant was passed through two superposed sieves with decreasing meshes 250 and 50 μm, respectively). This operation was repeated twice. The contents retained by the sieves were divided in two tubes and centrifuged for 4 min at 2000 rpm. The supernatant was discarded, and a viscosity gradient was created by adding 20 mL of sucrose solution at 50% in each centrifuge tube (Brundrett et al. 1996). The mixture was rapidly stirred, and the tube provided was centrifuged again for 1 min at 3000 rpm. Unlike the first centrifugation, the supernatant was poured onto the sieve of 50 μm; the resulting substrate was rinsed with distilled water to remove sucrose. The spores were then recovered with little distilled water in an Erlenmeyer flask and counted, and their density (spore number per 50 g dry soil) was determined.
Apparently viable spores were observed in a stereomicroscope and separated into morphotypes according to their colour, size and shape. They were mounted on slides with polyvinyl lactoglycerol (PVLG) and Melzer's reagent for observation under a microscope and taxonomic identification through spore morphology. Spores were extracted and separated into morphotypes by their colour, shape and size. Characteristics of spores, such as the number of spore layers, ornamentation of the outer layers, type of hyphal attachments and the wall layer reactions to Melzer's reagent, were also recorded. The identification of species was made according to the description of the species provided by the International Culture Collection of Vesicular Arbuscular Mycorrhizal Fungi (INVAM) following the classification of Redecker et al. (2013).
Data are expressed as the mean ± standard error of the mean. Statistically significant differences between the groups were evaluated by one-way of variance (ANOVA) followed by post hoc multiple comparisons with Tukey's honestly significant difference (HSD) test using IBM Statistical Package for the Social Sciences (SPSS) statistical software, ver.25.0 (SPSS, Inc., Chicago, IL, USA). Differences with
Degradation by anthropogenic pressure had a significant negative influence on most of the tested physicochemical parameters of the soil of
Soil physicochemical properties of
| Station | pH | OM (%) | N % | P (mg/kg) | K total |
|---|---|---|---|---|---|
| ND | 5.95a | 3.50a | 0.31a | 6.80b | 21.17b |
| D | 5.88a | 2.00b | 0.27a | 4.60a | 11.27a |
Note: ND – non-degraded station; D – degraded station; OM – organic matter; N – nitrogen; P – phosphorus; K – potassium.
EcM root colonization was assessed in the roots of cork oak and cistus in the degraded and non-degraded stations (Fig. 2). A negative influence of anthropogenic pressure on the EcM root colonization of
Figure 2
Rate of EcM colonization of cork oak (
The estimation of mycorrhizal intensity revealed, on average, significantly higher rates (
Figure 3
Mycorrhizal intensity (M%) of the root system of
COND: cork oak in the non-degraded station; COD: cork oak in the degraded station; CIND:
The results obtained show that the number of AMF propagules in the four studied soils of both stations had been affected negatively by degradation due to anthropogenic pressure. The soil of cork oak roots had the lowest number of 133 propagules per kilogram of soil (Fig. 4). However, under the T and CI, the number of propagules (in the non-degraded and degraded stations) was higher (566 and 400, respectively). On the other hand, the number of propagules in the soil under L was 666 and 466 in both stations, respectively.
Figure 4
The number of AM propagules per kilogram of soil in the degraded and the non-degraded stations
Estimation of the number of arbuscular fungal spores in 50 g of soil taken from cork oak, cistus, lavender and thyme in the non-degraded station showed high values compared to the number in the soil taken from the degraded station. The soil of T was the richest in fungal frequency with the total number of AM spores being 125 (Fig. 5), followed by L with 89 spores, while the lowest numbers of spores were recorded in the soil of CI and the CO with 52 and 50 spores, respectively. On the other hand, the degraded station showed lower numbers with 30, 39, 52 and 57 in the soil of CO, CI, L and T, respectively. The morphological criteria of the AMF spores isolated from the soil under study allowed us to find five spore species in thyme and lavender and three spore species in cork oak and cistus. The various genera and species identified in all rhizospheres belonged to the genera
Figure 5
The total number of the fungal diversity of spores of the four rhizospheres in the degraded and the non-degraded stations
Frequency of morphotypes of spores in the four rhizospheres
| Fractions | Spores collected from | ||
|---|---|---|---|
| 250 μm |
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| 250 μm |
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| 250 μm |
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| 250 μm |
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Note: + present, ++ abundant, +++ highly abundant.
Mediterranean forest ecosystems are severely threatened by the impact of humans and climatic conditions in numerous ways (Gauquelin et al. 2018). For this purpose, before restoring a degraded ecosystem, it was essential to know the effect of this degradation on the mycorrhizal communities associated with
In our study, the soil in both stations was characterized by low chemical fertility due to low levels of major assimilable nutrients (nitrogen and phosphorus). This degradation of our soil may be due to the combined factors of repeated fires, overgrazing, and anthropogenic pressure. Requena et al. (2001) also showed that loss and degradation of plant cover affect the physicochemical and biological properties of soils. Indeed, the influence of these factors on soil degradation mainly leads to deterioration of the ecological characteristics of the environment, whose first sign is degradation of the physical, chemical and biological properties of the soil, which makes the soil poor in mycorrhizae and fungal diversity (Zhao and Zhao 2007).
The results of EcM root colonization of cork oak and the cistus showed high EcM colonization in the non-degraded station, while in the degraded station, these rates were low. This may be due to the greater competition among these plants for water and nutrients in the soil, especially in more degraded soils. Our findings are consistent with the results obtained by Albuquerque-Martins et al. (2019) and de Vega et al. (2010) who have found that under different growing conditions, shrubby Cistaceae species showed a greater degree of EcM colonization. According to Shi et al. (2017), specific EcM fungi are capable of improving the juvenile growth of some forest species and mitigating the effects of transplant stress. The interaction between trees and EcM fungi is dependent on many factors, namely, tree species, environmental conditions and belowground interactions, among others (Opik et al. 2006). Even seasonal variations have an important role in EcM fungal dynamics in the soil. According to Voriskova et al. (2014), seasonal changes have a significant impact on fungal activity, biomass content and composition, as well as on the relative abundance of different fungal groups in temperate oak forests.
The presence of anthropogenic pressure had also negatively affected the rate of AM colonization and the number of spores. AMF are obligate biotrophic symbionts that require host plants to live (Smith and Read 2010), and changing the dominant plant species in the ecosystem could change the established balance in favor of certain fungal species. However, reduction in the diversity of plant communities that occurs in the degraded site should decrease the density of the AMF community. We suggest that the change in plant composition and dominance of ecosystem structure may be the primary forces driving the change in AMF structure. Therefore, selective forces towards more competitive and adaptable fungal species are likely active, contributing to the change in AMF community structure.
In Mediterranean ecosystems, the host tree can interact with a hundred different EcM fungal species. The functional complementarity of symbionts is key to the flexibility and resilience of forests to changing natural conditions (Azul et al. 2010; Courty et al. 2010). The majority of studies treated ectomycorrhizas because of their economic importance, using mainly artificial inoculation of seedlings (Agueda et al. 2006).
The number of AM propagules found in the soil of cork oak and cistus was low in the degraded station compared with the non-degraded station, whereas under the rhizosphere of lavender and thyme, this number was higher. The low number of propagules could be due to the nature of the soil, which is strongly anthropogenic due to human pressure, and overgrazing, which causes degradation of native plant communities (population structure and species diversity), resulting in loss or decrease of mycorrhizal propagules in the soil (Asmelash et al. 2016). Soil degradation and vegetation cover perturbation lead to loss of nutrients, reduction of microbial diversity (Kennedy and Smith 1995) and, in particular, propagules of mycorrhizal fungi, which therefore the reduction of the potential of the inoculum for the formation of soil mycorrhizae (Azcon-Aguilar et al. 2003). Our results are also in agreement with the findings of Duponnois et al. (2001), who have shown that degradation of soil by anthropogenic activities decreased its diversity and the number of AM propagules.
Indeed, degraded areas still show low densities of native mycorrhizal propagules. AMF species also have very different life cycles, and the relative abundance of every species at each stage of the cycle (spores, hyphae, colonised roots) may change over time (Nehila et al. 2015).
The natural diversity of spores was low in all the rhizospheres studied, with the dominance of the genus
Asmelash et al. (2016) showed that the degraded lands harbour low levels of AMF abundance and diversity. It was also reported that human disturbance decreased AMF spore density, root colonization and nutrient availability (Birhane et al. 2017). Smith (1987) also showed that the variation in the number of spores of AMF depend on the season, formation processes, and germination. Glomus species are widely distributed in various habitats of the world, and they dominate in degraded soils (Uhlmann et al. 2006). According to Schenck and Perez (1990), this AM species is ubiquitous. However, we noticed that the genus
The degradation of forest sites by anthropogenic pressure has been found to have a negative impact on the physicochemical properties of soil in degraded soils compared to the non-degraded station. The changes in soil properties demonstrated a negative impact on the EM-root colonization levels of
The mycorrhizal potential of the soil under cork oak was the lowest in the degraded station compared to that assessed under lavender, cistus and thyme in both stations. The presence of fungal spores was also low in all rhizospheres studied, with the dominance of the genus
Thus, in the case of our degraded Mediterranean ecosystems, the mycorrhizal potential is very low to ensure the development of forest species the selection of native AM and EcM fungi and their multiplication in trap plants could be reintroduced in the degraded oak forests and thus improve the quality of seedlings of this last in order to use them in the ecological restoration of degraded ecosystems.