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Inoculation of Ceratonia siliqua L. with native arbuscular mycorrhizal fungi mixture improves seedling establishment under greenhouse conditions
Abstract
The potential benefits of inoculation with arbuscular mycorrhizal (AM) fungi were investigated on carob
tree Ceratonia siliqua, a Mediterranean legume in Morocco. The parameters under study were the effect
of an inoculation on growth, mineral nutrition and roots mycorrhizal colonization of the plant under
nursery conditions. C. siliqua growth was measured after six months of culture in plastic bags arranged
in a randomised complete block under greenhouse conditions. Fungal inoculation consisted of a
mixture of native AM fungi propagated on Zea mays roots. Results show that the fungal symbionts were
effective to improve the growth of C. siliqua, confirming the requirement of mycorrhizal symbiosis for
the successful establishment of C. siliqua in a degraded soil. The approach used with indigenous AM
fungi complex isolated under C. siliqua appeared to be effective in promoting growth and nutrition of C.
siliqua. After 6 months of culturing in nursery conditions, height, shoot and root biomass, total
biomass, phosphorus and nitrogen foliar contents of the plants inoculated with native AM fungi were
significantly higher than in the control. Glomus spores were extracted from the soil under C. siliqua and
were observed on permanent slides under a microscope connected to a computer with digital image
analysis software. Seven spore morphotypes were detected under C. silqua in the Ourika Valley,
Morocco. Five Glomus species were classified as Glomus aggregatum, Glomus intraradices and
Glomus constrictum, whereas, two other Glomus species were not identified. The analysis of this spore
community revealed the presence of two other species belonging to Gigaspora genera. The use of a
mixture of native AM fungi as fungal inoculum improves clearly growth, nutrition and roots colonization
of C. siliqua seedling.
Key words: Arbuscular mycorrhizal fungi, diversity, growth, soil microbial activity, Ceratonia siliqua
... Here, saplings of five Mediterranean tree species, EM: Pinus halepensis and Quercus calliprinos (Torres & Honrubia, 1994;Trocha et al., 2012), and AM: Cupressus sempervirens (Zarik et al., 2016), Ceratonia siliqua (Essahibi et al., 2018;Lahcen et al., 2012) and ...
... Cupressus (Zarik et al., 2016) and Ceratonia (Essahibi et al., 2018;Lahcen et al., 2012). Most cases of transferred carbon occurred within the same guild, suggesting that the mycorrhizal fungi were active only in their traditional hosts. ...
Mycorrhizal fungi can colonize multiple trees of a single or multiple taxa, facilitating bidirectional exchange of carbon between trees. Mycorrhiza‐induced carbon transfer was shown in the forest, but it is unknown whether carbon is shared symmetrically among tree species, and if not, which tree species are better donors and which are better recipients. Here we test this question by investigating carbon transfer dynamics among five Mediterranean tree species in a microcosm system, including both ectomycorrhizal (EM) and arbuscular (AM) plants. Trees were planted together in 'community boxes' using natural soil from a mixed forest plot that serves as habitat for all five tree species and their native mycorrhizal fungi. In each box, only the trees of a single species were pulse‐labeled with 13CO2. We found that carbon transfer was asymmetric, with oak being a better donor, and pistacia and cypress better recipients. Shared mycorrhizal species may have facilitated carbon transfer, but their diversity did not affect the amount, nor timing, of the transfer. Overall, our findings in a microcosm system expose rich, but hidden, belowground interactions in a diverse population of trees and mycorrhizal fungi. The asymmetric carbon exchange among co‐habiting tree species could potentially contribute to forest resilience in an uncertain future.
... Benefits of mycorrhizal symbiosis were demonstrated on carob growth and drought stress tolerance (Essahibi et al., 2017;Manaut et al., 2015;Ouahmane et al., 2012), but different effects depending on Glomeromycota taxa was observed (Essahibi et al., 2017), highlighting the need to better characterize mycorrhizal partners of carob trees. The current results extended the range of Glomeromycota taxa previously described as associated with carob trees using older methodologies , with notably the detection of Diversispora spp., expanding the potentialities of carob habitats as reservoir of mycorrhizal resources to improve carob afforestation strategies . ...
... The current study focused on the belowground microbiota associated with several plant species representative of Mediterranean thermophilous carob woodlands in Morocco. The plant species investigated had been mainly assessed for their mycorrhizal community,(Alguacil et al., 2011;Azcón-Aguilar et al., 2003;Ferrol et al., 2004;Manaut et al., 2015;Ouahmane et al., 2012;Torrecillas et al., 2014;Turrini et al., 2010) notably because of the role of mycorrhiza in ecosystem functioning(Banerjee et al., 2018;Gianinazzi et al., 2010;Qin et al., 2019;Rillig, 2004b). The Glomeromycota in particular are well known for their benefits (1) on plant productivity, (2) on tolerance to drought stress and resistance to pathogens, as well as(3)in the process of plant succession(Smith and Read, 2009). ...
Le caroubier (Ceratonia siliqua L.), est une constituante majeure élément majeur de l’écosystème méditerranéen, jouant un rôle culturel et industriel important dans le pourtour méditerranéen. Cependant, ces dernières décennies connaissent un déclin du taux de réussite des plantations de caroubiers, notamment en condition d’irrigation limitante. La conservation de cette espèce est donc fortement dépendante de notre capacité à améliorer sa résistance face à l’aggravation des périodes de sécheresse. L’utilisation des champignons mycorhiziens arbusculaires, a été proposée comme une stratégie prometteuse pour améliorer la tolérance du caroubier face à la sécheresse. Cependant les mycorhizes ne représentent qu’une partie du microbiote bénéfique pour la croissance des plantes, et les cortèges microbiens natifs, par exemple associés aux plantes pionnières de l’habitat du caroubier, pourraient représenter des sources microbiennes bénéfiques pour la survie du caroubier face au changement climatique. Les objectifs de ce travail sont (i) la caractérisation du microbiote du sol dans différents habitats associés au caroubier au Maroc, et la détermination de leur spécificité en lien avec les composantes végétales, (ii) d’évaluer leur impact sur la croissance et résistance du caroubier en utilisant ces sols comme bio-inoculants. Les hypothèses sont (i) que le microbiote du sol est structuré au sein d’un habitat par la diversité floristique associée et que (ii) les spécificités du microbiote du sol et des racines sont des déterminants majeurs de la croissance du caroubier et de son niveau de résistance à la sécheresse. Les résultats ont montré une prédominance des actinobactéries, protéobactéries et ascomycètes dans deux habitats majeurs du caroubier, marquée par des divergences fonctionnelles entre le Nord et le Sud. L’utilisation comme bio-inoculant de microbiotes du sol associés à différentes composantes végétales des habitats à caroubier a montré des performances similaires entre le microbiote natif du caroubier et ceux associés à l’orge et au genêt blanc. Par contre, le microbiote natif du caroubier a conféré une faible résistance face des conditions de sécheresse, à l’inverse de celui provenant du pistachier lentisque. Les performances de croissance en condition non limitante en eau pourraient être liés à l’action conjointe de mycorhizes et de rhizobia, alors que la résistance à la sécheresse semble être conférée principalement par l’action de champignons endophytes de type DSE (dark septate endophytes), et potentiellement de mycorhizes. Le pistachier lentisque apparaît comme une cible prometteuse pour améliorer la résistance des caroubiers à la sécheresse en développant l'ingénierie de microbiotes naturels (transplantation de microbiotes du sol ou gestion des microbiotes du sol par couvert végétal).
... On the one hand, the effect of soil disturbance on spore viability can explain the negative correlation found between MPN and spore density. The same result was confirmed by Diop et al. [53], who demonstrated that disturbed ecosystem soils contain many non-viable spores that are not detected by the MPN method [54]. This discovery was also made in the soil of Retama monosperma following the degradation of Algeria's coastal ecosystem due to the overexploitation of coastal sands [55]. ...
The harm that invasive species cause to the environment has received a lot of attention. It is therefore appropriate that the current research was undertaken to evaluate the effects of invasion by Nicotiana glauca Graham on soil fertility by looking at (i) its contribution to the mycorrhizal potential of the soil, (ii) its impact on soil richness and diversity in terms of the arbuscular mycorrhizal fungi (AMF) community (iii), and its ability to modify the physicochemical characteristics in the invaded soil, specifically cleaning up heavy metal. The current study was conducted at Al Houz plain (Marrakesh region, Morocco), in heavily infested sites by N. glauca. The spores of AMF were isolated using the wet sieving process; the isolated spores were sorted for morphological features using a binocular microscope. The plant roots were thinned and colored before microscopic observation. The most probable number method was used to assess mycorrhizal soil infectivity. Heavy metal contamination in soils was characterized using an X-ray fluorescence spectrometer, and the pollution load index (PLI) was utilized to assess and compare the level of heavy metal contamination at each station. The ability of N. glauca to reproduce was evaluated in order to support one of its invasive characteristics. The estimate indicated that each plant might produce more than three million seeds. This significant number guarantees the plant a great capacity for reproduction and invasion. The extra-significant mycorrhizal potential, which can take the form of spores, mycelium, or vesicles that can regenerate mycorrhizae, was discovered by conducting soil analysis in the rhizospheric soils of N. glauca. This research demonstrated the strong mycotrophic capability of N. glauca and the large mycorrhizal potential of soils. Between 4.85 and 305.5 mycorrhizal propagules were considered to be the most probable number (MPN) per 100 g of dry soil. Based on color, shape and size, AMF were classified into five morphotypes corresponding to five genera. The isolated taxa of AMF with the most diverse spores were Glomus, Rhizophagus, Paraglomus, Scutellospora, and Sclerocystis. The Glomus genus was found to have spores in significant quantity. Furthermore, N. glauca demonstrated a potential involvement in the phytoremediation of damaged soils, with a high pollution load index demonstrating a particularly high accumulation of heavy metals. N. glauca is a highly mycotrophic plant that can boost soil mycorrhizal propagule stock. N. glauca has also been demonstrated to be a phytoremediation plant capable of cleansing contaminated soils. As a result, N. glauca could be considered as a prospective candidate for application in phytoremediation of polluted soils.
Atlantic forest natural soil flora was tested on two leguminous pioneer species Acacia gummifera and Retama monosperma to be used in rehabilitation programs of the coastal sand dunes forest of the Essaouira region. The rhizospheric soil of two endemic plant species: R. monosperma and Juniperus phoenicea was sampled and
split into two categories, one from native rhizospheric soil, the other of the sterilized rhizospheric soil. Investigation was focused on mycorrhizal formations, but other forms of beneficial symbiosis such as rhizobia and viable soil
microflora were also studied. Growth and nutrition variables assessed included lengths of roots and shoots, number of branches, ratio of root to shoot dry mass and water, nitrogen and phosphorus levels. Results showed important mycorrhizal associations in roots of both plants, presence of nodules and abundance of viable soil microflora. J. phoenicea had a 100% frequency of mycorrhizal formation
and an intensity of 80% compared to a frequency of 80% and intensity of 54% for R. monosperma. Nodules had the same density of CFU regardless of the origin. Abundance of viable microflora in rhisospheric soil of R. monosperma, J. phoenicea and the control differed significantly. Among the studied variables for A. gummifera and R. monosperma, nitrogen and phosphorus uptakes significantly wit the use of the nonsterilized rhizospheric soil. Both species doubled
their phosphorus uptake when colonized by mycorrhizal species, R. monosperma doubled its nitrogen uptake and A. gummifera increased it by seven times compared with the control. No significant difference was noted for the other variables
The carob tree has great potential for restoring vegetation to degraded areas around the Mediterranean and other dry areas of the world, and for improving the productivity of lands which would be marginal for other crops. There is scope for utilising an increase in the bean harvest, by expanding the volume of existing products/uses, and by developing new products such as protein rich foods and fodders, sugar and syrup, industrial gums and alcohol. The paper includes a discussion of botanical characteristics of the carob tree and of the Mediterranean ecosystems in which it is found.
The carob tree (Ceratonia siliqua L.) is an important economic resource for Morocco's rural populations. This species is used in reforestation actions and its cultivation in modern orchards is being undertaken to valorize marginal lands and substitute for drought sensitive species. However, little data is available on its intra-specific variability and its adaptability. Morphological characters of pods and seeds from 13 ecoregions of private-domesticated carob were used to assess phenotypic variation of this species. These stands extend from south-west to north-east and cover a wide range of Morocco's ecoregions. Pods length, width, thickness, seeds number, pulp weight, seeds yield and seeds length, width, thickness and weight were measured for 390 trees (30 trees per ecoregion). Statistically significant differences were found between ecoregions for all characters which were examined, what indicates a high phenotypic diversity. Principal component analysis (PCA) and hierarchical cluster of all ecoregions lead to identify two major and opposite groups (the northern ecoregions; and the central and south-western ecoregions). Ecoregions of the north of Morocco exhibited the largest and the thickest pods with the highest pulp weight while other ecoregions have relatively short pods but largest proportion of seed yield. Similarly, the northern ecoregions are characterized by the heaviest seeds. A correlation matrix between morphological characters, geographic parameters and precipitation exhibits a positive and a negative correlation of pods thickness and pulp weight with the latitude and the altitude, respectively. Seed yield and weight are negatively and positively correlated to pod width, pod thickness and pulp weight, respectively. In addition, seed weight is positively correlated with the latitude. The geographic pattern of the carob tree and its variability are discussed in this paper.
A new experimental procedure was developed to produce samples of leek roots containing early stages in colonization by vesicular–arbuscular mycorrhizal (VAM) fungi. This procedure involved transplanting leek seedlings into a pot culture containing an established symbiosis. Abundant colonization of seedling roots occurred in 1 week. Samples harvested at 2-day intervals after exposure to the inoculum were used to determine the time course of colonization. Contact of the root by external hyphae occurred after approximately 1 day, followed by penetration of hyphae into the root at about 2 days, arbuscule formation within 3–4 days, and vesicle formation after 4–5 days. The average distance from growing root apices at which each of the above events first occurred and the growth rate of intercellular hyphae were also estimated. Stages in root colonization were then documented using the chlorazol black E staining procedure. This transplant procedure should allow the production of abundant material at early stages in VAM development for chemical and morphological investigations as well as provide an efficient inoculation procedure.
There is now ample evidence to support the common assertion that most plants in natural ecosystems have mycorrhizal associations. Information about the worldwide distribution of plants with different types of mycorrhizal associations is used to establish correlations with the major climatic factors (water, temperature) which regulate the distribution of plants, as well as more localized edaphic conditions. Ecological implications of mycorrhizal associations in natural ecosystems and the role of soil or environmental factors, mycorrhizal fungus characteristics or host plant properties alone or in combination are considered. Factors which can influence the occurrence and effectiveness of mycorrhizal associations include (i) root properties (ii) edaphic or climatic factors (iii) soil organisms, (iv) soil disturbance and (v) host-fungus compatibility. More complex ecological topics (involving, the environment. plants and mycorrhizal fungi) that are discussed include (i) mycorrhizal phenology, (ii) factors responsible for varying degrees of mycorrhizal dependency in host plants, (iii) the role of mycorrhizal hyphae in soil, (iv) nutrient competition involving mycorrhizal and non-mycorrhizal plants and (v) mycorrhizal interactions involving pollution and other stresses, the rhizosphere, soil properties and allelopathy. The population ecology of mycorrhizal fungi and the influence of their associations on plant population ecology are also considered.
Aims:The study aimed to determine whether inoculation with native arbuscular mycorrhizal (AM) fungi could improve survival and growth of seedlings in degraded soils of Morocco. Methods and Results:Soil samples were collected from the rhizosphere of Cupressus atlantica trees in the N’Fis valley (Haut Atlas, Morocco). AM spores were extracted from the soil, identified and this mixture of native AM fungi was propagated on maize for 12 weeks on a sterilized soil to enrich the fungal inoculum. Then C. atlantica seedlings were inoculated with and without (control) mycorrhizal maize roots, cultured in glasshouse conditions and further,
transplanted into the field. The experiment was a randomized block design with one factor and three replication blocks. The results showed that a high AM fungal diversity was associated with C. atlantica; native AM fungi inoculation was very effective on the growth of C. atlantica seedlings in glasshouse conditions and this plant growth stimulation was maintained for 1 year after outplanting.
Conclusions:Inoculation of C. atlantica with AM fungi increased growth and survival in greenhouse and field.
Significance and Impact of the Study:The data indicate that use of native species of AM fungi may accelerate reforestation of degraded soils. Further studies have to be performed to determine the persistence of these mycorrhizae for a longer period of plantation and to measure the effects of this microbial inoculation
on soil biofunctioning.
The mycorrhizal status of several representative shrub species (Lavandula spp. and Thymus satureioides) in Moroccan semiarid ecosystems, was evaluated as well as their contribution to the mycorrhizal potential of the soil. Furthermore, the rhizosphere soils collected under these target species were tested for their influence on the growth of Cupressus atlantica, a tree species whose natural stands has declined in this area. Soil samples were collected from the rhizosphere of L. stoechas, L. dentata and of C. atlantica existing in the experimental area. Control samples were randomly collected from bare soil sites, away from plant influence.
All the target species formed AM symbiosis and the extent of AM fungal colonization was not significantly different between plant species. No significant difference was detected between the total number of AM fungal spores of the bare soil and those recorded in the root zones of target species and C. atlantica. Three genera of AM fungi (Scutellospora, Glomus and Acaulospora) were present in the rhizospheres of the plant species and in the bare soil.
The number of mycorrhizal propagules in soil originating from around the four target plant species was significantly higher than the one in the bare soil (Figure 1). The most probable number (MPN) of mycorrhizal propagules per 100 g of dry soil ranged from 7.82 (bare soil) to 179.7 (L. dentata and C. atlantica) and 244.5 (L. stoechas and T. satureioides). As the total number of spores was not different for the soil of different origins, the increase of the mycorrhizal soil infectivity (MSI) mainly resulted from larger AM mycelial networks that constituted the main source of AM fungal inoculum. In addition, this MSI enhancement was linked with changes in the functioning of soil microbial communities. In a glasshouse experiment, the growth of C. atlantica seedlings was significantly higher in the C. atlantica and in the shrub species soils than in the bare soil. Although the AM inoculum potential is not sufficient to ensure the development of forest trees in Mediterranean ecosystems, the use of plant nurses such as T. satureioides or Lavandula spp. could be of great interest to restore a self-sustaining vegetation cover to act against desertification.
The possibility of utilising chopped and deseeded carob pods (kibbles) as a source of polyphenolic antioxidants was examined by performing extractions with various solvent systems, in order to evaluate and optimize the conditions for the recovery of polyphenols. Maximum quantities of polyphenolic components were found in 80 % acetone extracts, as evaluated by measuring total polyphenol and total flavanol content. By contrast, ethyl ace-tate was inefficient in extracting polyphenols. The assessment of the antioxidant potency of carob pod extracts employing two characteristic in vitro models showed that carobs con-tain polyphenols with appreciable antiradical and reducing properties. The values obtained were compared to the data on red wines and pure polyphenolic antioxidants.
A survey of arbuscular mycorrhizal (AM) fungi was conducted in seven Tetraclinis woodlands. Microscopic analysis of the mycorrhizal status of T. articulata (Vahl) Masters roots revealed that all samples formed only AM, and no ectomycorrhizal fungi were detected. The mycorrhizal colonisation level was generally high (more than 80%), thus reflecting the mycotrophic nature of T. articulata. A "Paris-type" mycorrhizal structure was noted in all studied samples. The number of AM fungal spores detected in field-collected soils was relatively high. All recovered spores belonged to the Glomineae order, represented by Glomaceae and Acaulosporaceae families. Two groups were dominant: the first one included small (90 μm), hyaline, white to dark-yellow spores, and the second involved large (295 μm), light orange to dark orange-brown spores. The morphological characters indicated that the spore populations consisted of 3-6 morphotypes. The Glomus genus was represented by five species, i.e. Glomus aggregatum, Glomus constrictum, Glomus sp. 1, Glomus sp. 2, and Glomus sp. 3, while the Acaulospora genus was represented by only one unidentified species.
Disturbance of natural plant communities is the first visible indication of a desertification process, but damage to physical,
chemical, and biological soil properties is known to occur simultaneously. Such soil degradation limits reestablishment of
the natural plant cover. In particular, desertification causes disturbance of plant-microbe symbioses which are a critical
ecological factor in helping further plant growth in degraded ecosystems. Here we demonstrate, in two long-term experiments
in a desertified Mediterranean ecosystem, that inoculation with indigenous arbuscular mycorrhizal fungi and with rhizobial
nitrogen-fixing bacteria not only enhanced the establishment of key plant species but also increased soil fertility and quality.
The dual symbiosis increased the soil nitrogen (N) content, organic matter, and hydrostable soil aggregates and enhanced N
transfer from N-fixing to nonfixing species associated within the natural succession. We conclude that the introduction of
target indigenous species of plants associated with a managed community of microbial symbionts is a successful biotechnological
tool to aid the recovery of desertified ecosystems.
A survey of the natural mycorrhizal potential has been carried out in a representative area of a desertified semiarid ecosystem in the southeast of Spain. Many indigenous plants from the field site were mycorrhizal, including the dominant Anthyllis cytisoides, which had high levels of colonization by arbuscular mycorrhizal fungi (AMF). Low numbers of AMF spores were present in the soil, although a range of species, including Scutellospora calospora, Glomus coronatum, Glomus constrictum, and several Acaulospora species, was represented. Soil infectivities, as determined by a soil dilution method, were similar for most plants tested but were significantly lower for Anthyllis cytisoides. Nevertheless, when a less disruptive method to determine soil infectivity was used, the importance of the mycelial network in maintaining the infectivity of soil under perennial shrubs, such as Anthyllis cytisoides, was highlighted. Seasonal variations in the mycorrhizal infectivity showed that it was higher towards the end of the summer period than in midwinter. In screening trials in a greenhouse, the indigenous AMF did not significantly improve the growth of plants compared with that of noninoculated controls. Augmentation of the soil with an inoculum of Glomus intraradices resulted in improved growth of Anthyllis cytisoides in both sterile and nonsterile conditions, in contrast to results obtained following inoculation with Glomus mosseae or another Glomus sp. Our findings suggest that the indigenous inoculum levels of AMF are inadequate to support an extensive revegetation program in the absence of an additional mycorrhizal inoculum.
1 Removal experiments are widely used to study competition in natural ecosystems but suffer from a number of methodological drawbacks. We describe two glasshouse bioassays using field soil which were designed to test for effects of removal experiment methodology on plant growth. 2 Soil was sieved to remove plant roots which were then added back to some samples. The presence of decaying roots in the soil did not significantly alter the performance of Holcus lanatus plants over the time scale of the experiment (three months). 3 Over the same period of time, however, sieving the soil significantly reduced the mycorrhizal infection of Holcus lanatus roots and subsequent shoot growth. In the same experiment, a nonmycorrhizal species (Cerastium fontanum) was unaffected by soil disturbance. 4 These findings suggest that the disturbance resulting from total plant removal will have adverse effects on the performance of plants remaining in the sward. Since the presence of roots seems to have little effect, above-ground clipping and leaving roots to decay in the soil is likely to be a better approach to removal experiments.
The general objective of this study was to measure the mycorrhizal dependencies of Cupressus and Lavandula species, to assess the contribution of Lavandula species to the mycorrhizal soil infectivity (MSI) and to determine the main effects of the association between a Lavandula species (L. multifida) and Cupressus arizonica on the soil microbial activities, on the growth and the mycorrhizal status of each plant species in experimental conditions. Three species of Lavandula (L. stoechas L., L. dentata L. and L. multifida L.) and three species of Cupressus (C. atlantica, C. sempervirens and C. arizonica) were tested. Plant species were inoculated with an arbuscular mycorrhizal fungus, Glomus intraradices, to determine their mycorrhizal dependencies in controlled conditions. The results showed that L. multifida and L. dentata were representatives of highly mycorrhizal dependent plant species as well C. arizonica and C. sempervirens. The contribution of L. multifida to the MSI was determined by the most probable number (MPN) method. In six different soils, L. multifida enhanced the mycorrhizal soil infectivity, even more if the soil was P deficient. When C. arizonica and L. multifida were grown together, the growth and mycorrhizal colonization of each species were higher than those recorded when each plant species grew alone. The impact of this dual cultivation on the growth and mycorrhizal colonization of C. arizonica was comparable to those recorded with C. arizonica seedlings previously inoculated with G. intraradices. In contrast, the microbial activities of the soil collected from each treatment were different. These results emphasize the role of “resource islands” and “nurse plants” of Lavender plants in the regeneration processes of tree species such as Cupressus spp. In addition, they confirm the role of AM fungi as a major factor contributing to the growth and co-existence of each of the plant species (L. multifida and C. arizonica).
summaryEffects of inoculation of Hedysarum boreale ssp. boreale Nutt. (Fabaceae) with mycorrhizal fungi (topsoil inoculum) and Rhizobium (pure inoculum) were studied in the field in southwestern Wyoming, USA. After 3 years, plants receiving both inocula had greater total aboveground biomass and leaflet biomass, more leaves, higher aboveground nitrogen and phosphorus contents and greater survival than plants which received single or no inoculum. Soil 20–40 cm deep (where roots were concentrated) beneath Hedysarum individuals from all treatments had a lower concentration of total nitrogen than did soil between plants. Hedysarum plants, inoculated or not, apparently extracted sufficient nitrogen from the soil, as they had similar leaflet nitrogen concentrations. Hedysarum decreased the size of the pool of soil nitrogen even with Rhizobium inoculation on disturbed land in a semi-arid environment during the first 3 years of growth.
The infectivity of vesicular–arbuscular (VA) mycorrhizal fungi in three soils from different vegetation types was compared before and after disturbance. The relative quantities of infective propagules in the disturbed soils were estimated by an infectivity test after the soils were diluted. Spare numbers and mycorrhizal roots were also quantified in each soil.
The mycorrhizal colonization of clover roots in the infectivity st was not decreased after soil from an annual pasture had been disturbed. In contrast, in both a forest soil and a heathland soil, the percentage root length colonized of test plants was almost halved if the soils had been disturhtd. In the pasture soil up to 10 times or 25 times more propaguies survived disturbance than in the forest soil or the heathland soil respectively. The large number of propaguies may allow the VA mycorrhizal fungi in the pasture soil to maintain maximum infectivity after soil disturbance.
Extra-matrical spores of Endogone were found to be common in cultivated soils in Scotland. Six types of spores are described which are believed to represent distinct species. In preliminary inoculation experiments 4 of the 6 formed endotrophic mycorrhiza: three of these formed arbuscules and vesicles, and one formed only arbuscules. In field soil extra-matrical spores are much more abundant than spores formed in sporocarps.
Plants in nutrient-deficient soils often benefit when colonized by vesicular-arbuscular mycorrhizal fungi (VAMF). Plants vary, however, in responsiveness or mycorrhizal dependency (MD). The objective of this work was to evaluate MD within corn and soybean cultivars. Three improved and three unimproved corn (Zea mays L.) and three improved and two unimproved soybean [Glycine soja Siebold & Zucc. and Glycine max (L.) Merr.] cultivars were evaluated for growth response, nutrient uptake (N, P, K, Ca, Mg, and Zn), and root phosphatase activity. Greenhouse studies were conducted with VAMF (Gigaspora margarita or Glomus intradices) and without inoculation in a low-P soil. The VAMF colonization of roots with inoculation ranged from 62 to 87% for soybean and 49 to 68% for corn. Soybean had a higher MD than corn, but considerable variation occurred within soybean cultivars. Relative growth of the two unimproved soybean cultivars was significantly greater with VAMF colonization than without (Soja, > 1900%; Mandarin, > 400%). Among improved cultivars, relative growth was less enhanced with colonization (BSR 201, Richland, and Swift cultivars averaged 200% greater growth with VAMF than without VAMF colonization). An unimproved corn cultivar, Reid Yellow Dent, was unresponsive to mycorrhizal colonization, whereas another unimproved cultivar, Argentine Pop, increased growth 400% with colonization. Total uptakes of N, P, K, Ca, Mg, and Zn were significantly greater (P< 0.001) in mycorrhizal plants, but the concentrations (mg g −1 ) of N, Mg, and Ca were lower in mycorrhizal plants than in nonmycorrhizal plants. These results suggest that considerable variability exists in MD of corn and soybean cultivars when grown in P-deficient soils, and the variability extends across both improved and unimproved cultivars
The continuous production of ethanol from carob pod extract by immobilized Saccharomyces cerevisiae in a packed-bed reactor has been investigated. At a substrate concentration of 150 g dm−3, maximum ethanol productivity of 16 g dm−3 h−1 was obtained at D = 0·4 h−1 with 62·3% of theoretical yield and 83·6% sugars′ utilization. At a dilution rate of 0·1 h−1, optimal ethanol productivity was achieved in the pH range 3·5–5·5, temperature range 30–35·C and initial sugar concentration of 200 g dm−3. Maximum ethanol productivity of 24·5 g dm−3 h−1 was obtained at D = 0·5 h−1 with 58·8% of theoretical yield and 85% sugars′ utilization when non-sterilized carob pod extract containing 200 g dm−3 total sugars was used as feed material. The bioreactor system was operated at a constant dilution rate of 0·5 h−1 for 30 days without loss of the original immobilized yeast activity. In this case, the average ethanol productivity, ethanol yield (% of theoretical) and sugars′ utilization were 25 g dm−3 h−1, 58·8% and 85·5%, respectively.
Reconstitution of the potential of soil mycorrhizal inoculum is a key step in revegetation programs for semiarid environments. We tested the effectiveness of inoculation with native arbuscular mycorrhizal (AM) fungi or with an allochthonous AM fungus, Glomus claroideum, with respect to the growth of four shrub species, the release of mycorrhizal propagules in soil, within and outside the canopy, and the improvement of soil structural stability. Two years after outplanting, the mixture of native endophytes was more effective than, for Olea europaea subsp. sylvestris, Retama sphaerocarpa and Rhamnus lycioides, or equally as effective as, for Pistacia lentiscus, the non-native AM fungus Glomus claroideum, with respect to increasing shoot biomass and foliar NPK contents. The increases in glomalin concentration and structural stability produced by inoculation treatments in the rhizosphere soil of the all shrub species, except R. lycioides, ranged from about 55 to 173% and 13 to 21%, respectively. The mixture of native AM fungi produced the highest levels of mycorrhizal propagules in soil from the center of the canopy of P. lentiscus and R. lycioides, while plants of O. europaea and R. sphaerocarpa inoculated with G. claroideum had more mycorrhizal propagules than did those inoculated with the mixture of native fungi. The number of mycorrhizal propagules in soil outside the canopy of the four shrub species was 5–35 times higher in inoculation treatments than in soil of the non-inoculated plants.
Diurnal time course of the water relations of Ceratonia siliqua, Quercus coccifera, Pistacia terebinthus and Olea oleaster was studied on healthy and degraded sites for getting a suitable parameter describing the water stress impact on vegetation, and selecting the species suitable for reforestation. Out of the species used C. siliqua, showed high maximum stomatal conductance (g L), with high relative water content (RWC) under field conditions, and relatively constant minimum (midday) water potential (C min) and pre-dawn water potential (C pd 's). O. oleaster plants appeared to be unable to prevent dehydration inspite of consistent decrease of stomatal conductance (g L), when subjected to increasing water stress. Stomatal closure in fact, was not sufficient to prevent water loss and relative water content (RWC) dropped to about 70%. P. terebinthus species is water spender. Under water shortage condition inspite of complete closure of stomata (stomatal conductance dropped to 0.08 s cm À1), relative water content showed variable values. Q. coccifera typically showed high stomatal conductance activity and also high relative water content (average 83%) on healthy site, but on the degraded site it reduced stomatal conductance, thus maintaining high RWCs (over 80%) and preventing leaf water potential (C L) to drop to critical values.
The genetic diversity of 15 carob (Ceratonia siliqua L.) cultivars located in an experimental field from Algarve (Portugal) was evaluated over 7 years using 12 fruit and seed phenotypic characters, in order to characterize carob cultivars. The values of morphological traits obtained by cultivar were compared with those from other countries of the Mediterranean basin. Statistically significant differences were found between cultivars for all characters which were examined, what indicates a high genetic diversity. The relationship among these characters was analyzed by principal component analysis (PCA) resulting in the separation of these cultivars classed in four groups (clusters I–IV) and in four ungrouped cultivars. A three dimension of the model was found to be significant and explained 74.5% of the total variation, in which the first component accounting for 34.6% of the total variation is dominated by fruit characters, while the second component is dominated by seed characters. Cultivars plotted on the left-lower quadrant on the space determined by principal components 1 and 2 are characterized by fruits with high seed yield more appropriated for industrial rentability. The correlation analyses established by cultivar provided a specific understanding about the way how fruit and seed characteristics correlate within each cultivar. This approach can be useful for the development of a breeding programme, aiming to increase the seed yield, seed thickness, individual and total seed weight by fruit, characteristics that are determinant to improve the industrial exploitation of carob. # 2007 Published by Elsevier B.V.
Rates of photosynthesis and leaf conductance of the leaves of carob trees (Ceratonia siliqua L.) growing in natural conditions were measured during the course of the seasons to define the effects of the main climatic factors limiting growth in the region: temperature during the winter and water in the summer. The highest photosynthetic rates were measured in spring and autumn and could reach 25 μmol m−2 s−1 with optimal temperature and available water. Due to lower temperatures (4 to 6°C in the night) these values were frequently around 15 μmol m−2 s−1 during winter, but the strongest depression was due to prolonged drought in summer. However a reduction in photosynthesis rate down to 5 μmol m−2 s−1 occurred only after depletion of all the available water in the soil layer up to a depth of 50 cm. In the end of the summer, leaf conductance and water potential were in the order of 20 mmol m−2 s−1 and −3 MPa respectively. Compared to other trees that make up the Mediterranean sclerophyll forest, the photosynthetic activity of carob is high, and the tree tolerates a considerable depletion of soil water.
Phosphatase activity of arbuscular mycorrhizal (AM) fungi has attracted attention in three fairly distinct domains: intracellular
enzymes with defined metabolic functions that have been studied in intraradical hyphae, histochemical staining of alkaline
phosphatase as an indicator of fungal activity measured both intra- and extraradically, and extracellular activity related
to mineralization of organic P (Po) compounds that may enhance mycorrhizal utilization of an important nutrient pool in soil. This review focuses on the latter
subjects with emphasis on extraradical mycelium (ERM), while it draws on selected data from the vast material available concerning
phosphatases of other organisms. We conclude that histochemical staining of alkaline phosphatase is a sensitive and suitable
method for monitoring the effect of adverse conditions encountered by ERM both as a symbiotically functional entity in soil,
and in vitro without modifying interference of soil or other solid substrates. Furthermore, the quantitative importance of extracellular
enzymes for P nutrition of AM plants is estimated to be insignificant. This is concluded from the low quantitative contribution
extracellular hyphae of AM fungi give to the total phosphatase activity in soil, and from estimations of which processes that
may be rate limiting in organic P mineralization. Maximum values for the former is in the order of a few percent. As for the
latter, solubilization of Po seems to be far more important than Po hydrolysis for utilization of Po by AM fungi and plants, as both endogenous soil phosphatase activity and phosphatases of other soil organisms are ubiquitous
and abundant. Our discussion of mycorrhizal phosphatases supports the view that extracellular phosphatases of roots and micro-organisms
are to a large extent released incidentally into soil, and that the source has limited benefit from its activity.
An evaluation of the mycorrhizal status of desertification-threatened ecosystems has been recommended as a first step in rehabilitation/restoration approaches based on revegetation strategies using arbuscular mycorrhizal (AM) technology. Representative desertified semiarid areas were selected from southeast Spain where the vegetation is dominated by grasses, with Stipa tenacissima usually present, and with some patches of the shrubs Pistacia lentiscus, Rhamnus lycioides, Olea europaea subsp. sylvestris and Retama sphaerocarpa. The objective of this study was to evaluate the mycorrhizal potential in these soils, the contribution of the different species established to the mycorrhizal potential of the soils and to assess the main mycorrhizal propagules involved. There were more AM fungal propagules in the rhizospheres of all the shrub species studied compared with adjacent fallow soils, suggesting that AM propagules can be considered as a functional component of the resource islands developing around plant roots. R. sphaerocarpa and O. europaea had a higher capacity to enhance the development of mycorrhizal propagules in their rhizospheres than R. lycioides and P. lentiscus. Correlation analyses showed that the number of spores of the most representative AM fungal species, i.e. Glomus constrictum, and the total length of extraradical AM mycelium are the propagule sources which were best correlated with the mycorrhizal potential in terms of the number of “infective” AM propagules in the rhizosphere of the target plant species. The contribution of AM symbiosis to the potentiality of S. tenacissima as nurse plant was site dependent. Diversity of AM fungi present in the test area is rather low, indicating the high degree of degradation of the ecosystem. At most, only four AM fungal spore morphoecotypes were consistently detected in the rhizosphere of the target plant species.
Two techniques commonly used in soil restoration: (1) inoculation with selected microbial strains and (2) soil amendment with urban waste, were evaluated for afforestation of a degraded gypsiferous soil with Pinus halepensis Miller. One year after planting, survival of pines was not affected by any of these techniques, reaching values over 97% in all cases. Inoculation with two PGPR strains previously proven to enhance Pinus spp. growth, or with an ectomycorrhizal fungus separately did not affect pine growth or the nutrient content in needles. However, significant increments of pine growth due to soil amendment were detected. Most of the nutrients analysed in needles were found in higher amounts in plants grown in non-amended soil, except nitrogen that was higher in plants grown in the amended soil. The rhizospheric microbial activity was highly increased by soil amendment whereas inoculation with the different microorganisms had no effect on this parameter. The amendment of soil with urban waste drastically reduced the percentages of ectomycorrhizas on the pine roots and affected the frequency of the described ectomycorrhizal morphotypes compared with plants grown in the non-amended soil. Our results suggest that before undertaking a strategy for soil restoration, like plant inoculations with selected microorganisms or soil amendments, the autochthonous beneficial bacterial and fungal communities in the soil should be taken into account, as well as studies for adjusting the doses of amendments, to assure the establishment and development of plants and to keep the ecological equilibrium of soil.
Extracts from pods and leaves of carob (Ceratonia siliqua L.) were tested for their ability to inhibit cell proliferation of mouse hepatocellular carcinoma cell line (T1). The two extracts showed a marked alteration of T1 cell proliferation in a dose-related fashion reaching the maximal effect at 1 mg/ml. Moreover, we demonstrated that leaf and pod extracts were able to induce apoptosis in T1 cell lines after 24-h treatment mediating a direct activation of the caspase 3 pathway. HPLC analysis revealed the presence of gallic acid, (-) epigallocatechin-3-gallate and (-) epicatechin-3-gallate in pod and leaf extracts, compounds well known to exert antiproliferative effects. Their concentration reached 6.28 mg/g in carob leaves and 1.36 mg/g in carob pods extract. The discovery that carob pod and leaf extracts contained antiproliferative agents could be of practical importance in the development of functional foods and/or chemopreventive drugs.
Revegetation strategies, either for reclamation or for rehabilitation, are being used to recover desertified ecosystems. Woody legumes are recognized as species that are useful for revegetation of water-deficient, low-nutrient environments because of their ability to form symbiotic associations with rhizobial bacteria and mycorrhizal fungi, which improve nutrient acquisition and help plants to become established and cope with stress situations. A range of woody legumes used in revegetation programs, particularly in Mediterranean regions, were assayed. These legumes included both exotic and native species and were used in a test of a desertified semiarid ecosystem in southeast Spain. Screening for the appropriate plant species-microsymbiont combinations was performed previously, and a simple procedure to produce plantlets with optimized mycorrhizal and nodulated status was developed. The results of a 4-year trial showed that (i) only the native shrub legumes were able to become established under the local environmental conditions (hence, a reclamation strategy is recommended) and (ii) biotechnological manipulation of the seedlings to be used for revegetation (by inoculation with selected rhizobia and mycorrhizal fungi) improved outplanting performance, plant survival, and biomass development.
Hydrogels are hydrophilic macromolecular networks that are capable of retaining a large amount of water. A precise description of these systems is actually quite complex and the practical use of hydrogels for drug delivery and biomedical applications is often not supported by a well-defined knowledge of the overall structure of the polymeric network. In this paper, we report the characterization of two different systems: a chemical network based on Guar Gum (GG) and a physical gel prepared with Xanthan (Xanth) and Locust Bean Gum (LBG). The dynamo-mechanical properties of the gels were analysed: the cohesiveness and the adhesion of the networks were strongly dependent on time, temperature, and composition. The kinetics of the chemical crosslinking was followed by means of rheological measurements, i.e. recording the mechanical spectra of the gelling system, and the power law exponent at the gel point was evaluated. Furthermore, the networks, loaded with model drugs with different steric hindrance, were used as matrices for tablets and the rate of release of such model drugs was studied. The diffusion of the guest molecules was deeply dependent on their dimensions; in the case of Xanth-LBG tablets the release profiles were almost independent from the different cohesion properties of the starting hydrogel composition.