Akbar Karimi; Habib Khodaverdiloo; MirHasan Rasouli Sadaghiani
Abstract
Introduction: Recently, due to enhancement of industrialization, urbanization and disposal of wastes, fertilizers and pesticides the concentration of heavy metals (HMs)in agricultural soil has increased. Heavy metals are serious threat for environment due to their hazardous effects. Lead (Pb) is one ...
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Introduction: Recently, due to enhancement of industrialization, urbanization and disposal of wastes, fertilizers and pesticides the concentration of heavy metals (HMs)in agricultural soil has increased. Heavy metals are serious threat for environment due to their hazardous effects. Lead (Pb) is one of the toxic heavy metal that threats the health of plants, living organisms and human. Excessive Pb concentrations in agricultural soils result in decreasing the soil fertility and health which affects the plant growth and leads to decrease in plant growth. Plants simultaneously exposed to Pb suffer morphological, biochemical and physiological injury. Pb adversely affect plant absorption of essential elements, chlorophyll biosynthesis and shoot and root growth. Arbuscular mycorrhizal fungi (AMF) and plant growth-promoting rhizobacteria (PGPR) are known to enhance nutrient uptake and improvement of plant growth and tolerance in heavy metal contaminated soils through different mechanisms including producing low molecular weight organic acids, siderophore, antibiotics and hormones. The objective of this study was to evaluate the effect of AMF and PGPR on yield, leaf relative water content (RWC), some biochemical properties and uptake of Pb, Fe and Zn by Hyoscyamus (Hyoscyamus niger L.) under soil Pb contamination.
Materials and Methods: This study was carried out in greenhouse condition as a factorial experiment based on a randomized complete block design with two factors including Pb concentration (in four levels) and microbial treatment (in three levels including arbuscular mycorrhizal fungi, plant growth-promoting rhizobacteria and control) and in three replications. Consequently, a soil was selected and spiked uniformly with concentrations of Pb (0, 250, 500 and 1000 mg Pb kg-1 soil). The contaminated soil was then sterilized and inoculated with the selected species of arbuscular mycorrhizal fungi (a mixture of Glomus species including G. intraradices, G. mosseae and G. fasciculatum) or plant growth-promoting rhizobacteria (a mixture of Pseudomonas species includeing P. putida, P. fluorescens, and P. aeruginosa). Seeds of Hyoscyamus niger L. plant were grown in pots containing the Pb spiked soil. At the end of growth period shoot length, dry weights of root and shoot, Fe, Zn and Pb concentration in shoot, and some biochemical and physiological properties of plant including relative water content (RWC) chlorophyll a, b and total chlorophyll, carotenoids, proline and soluble sugars, were measured.
Results and Discussion: Results indicated that with increasing soil Pb concentration, dry weights of root and shoot, shoot length, photosynthetic pigments contents (chlorophyll a, chlorophyll b, total chlorophyll and carotenoids), shoot Fe and Zn concentration decreased, while proline and soluble sugars contents and the shoot Pb concentration increased. With increasing of soil Pb concentration, relative water content decreased, however, this reduction in concentration of 1000 mg Pb kg-1 soil was not significant (P > 0.05) in compared with concentration of 1000 mg Pb kg-1 soil. Amounts of all measured properties in AMF and PGPR treatments were higher than that control treatment. The highest values of shoot weight and root weight, were observed in plants that inoculated with AMF. The lowest shoot weight was recorded in non-inoculated plants that were grown under 1000 mg Pb kg-1 soil concentration. In this study Arbuscular mycorrhizal fungi and plant growth-promoting rhizobacteria inoculation led to a significant increase (P≤0.05) in shoot length (12.9 -71.1%), shoot dry weight (11.5 – 81%), root dry weight (18.4 – 60.6%), chlorophyll (8.5 – 36.5%) and carotenoid (11.5 – 40.0%) pigments, proline (55 – 115.7%), soluble sugars (17.6 – 72.2%) and shoot Fe (9.5 – 57.2%) and Zn (25.0 – 165.5%) concentration in shoot at different levels of soil Pb. The highest and lowest amounts of shoot Fe, Zn and Pb concentration observed in AMF and control treatments respectively. Plant growth promoting rhizobacteria were more effective than arbuscular mycorrhizal fungi in shoot Fe, Zn and Pb concentration, while the mean of shoot length and shoot and root dry weight was higher in plants that inoculated with AMF compared to ones inoculated with PGPR. In general, there were not significant (P ≤ 0.05) differences in amounts of chlorophyll (chlorophyll a, b and chlorophyll a+b) and carotenoids pigments, proline and soluble sugars between AMF and PGPR treatments.
Conclusion: It could be concluded that microbial inoculation (mixture of AMF and PGPR species) with improvement of plant biochemical properties results in improved Hyoscyamus niger L. yield and increased tolerance to Pb toxicity. Thus, the use of microbial inoculation (mixture of AMF and PGPR species) inoculation might be suggested for enhancement of plant tolerance in Pb contaminated soils.
Morteza Pouzesh Shirazi; Hassan Haghightnia; Rahim Khademi
Abstract
Introduction: Tomato is one of the most important vegetables that are used by human. High price of tomato due to its out of season production in Bushehr province has made a great tendency for farmers of this area to cultivate it in fall and winter in nearly 12000 hectares in the province. Mean annual ...
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Introduction: Tomato is one of the most important vegetables that are used by human. High price of tomato due to its out of season production in Bushehr province has made a great tendency for farmers of this area to cultivate it in fall and winter in nearly 12000 hectares in the province. Mean annual precipitation is about 250 mm per year. Maximum and minimum annual temperatures are 51 and -1 Celsius degrees. Mean annual evaporation is 3000 mm. Recent droughts and going down of water table because of over usage of underground water are main problems of this region that have enforced farmers to decrease irrigation water utilization and try to increase water use efficiency by using new technologies. Therefore, any new technical methods which can increase water use efficiency will help crop production quality and quantity and result in yield stability in the region. Many researchers have proved that mycorrhizal fungi can improve agricultural systems sustainability because of their ability to increase water absorption due to their long and expanded hyphae. Also they are known to increase nutrient uptake from soils even poor in minerals. Therefore, a greenhouse project was designed to determine the effect of Arbuscular mycorrhizal fungi on nutrients uptake and water use efficiency of tomato under drought stress in Bushehr province in southern Iran.
Materials and Methods: The experiment was carried out on completely randomized block design in factorial with three replications during 2016-2017 in the greenhouse of Bushehr agricultural and natural resources research organization located in Southern Iran. Factors were as follows: 1) four Arbuscularmycorrhizal fungi (Glomus mosseae ،Glomus intraradices، Glomus vsersiform and Glomus caledonium) and a control (without inoculation), 2) irrigation at three levels based on soil moisture depletion at I1=25, I2=50 and I3=70 percent of available water to show stress (from non-stress to low and high stress levels). Soil used for this experiment was dried in sunshine for one month to be evacuated from any native fungi. Soils of the region are calcareous ones with low salinity limitation (EC= 3.70 dS/m) and poor nutrition elements and slightly alkali (pH=7.9). Inoculation of roots by fungi was done by pouring one spoon of fungi near tomato roots while cultivating the transported seedlings in the pot. This would help the roots to be exposed to fungi directly while spreading through growing. Irrigation water used were 22.3, 20.1 and 17.7 liter for each pot filled with 10 kg soil during the season. The experiment lasted 100 days and tomato properties were measured such as yield, colonization percent, nutrition elements concentration and also, water use efficiency. Dried leaves were milled and put in furnace at 550 Celsius degrees. Roots were soaked in water for five minutes to be detached from soil and cleaned toughly. Detached roots were maintained in small glass jars filled with alcohol and were kept in refrigerator at 5 Celsius degrees. Roots were then colored by Try pan blue method and root colonization percent was measured by Grid line intersect method.
Results and Discussion: Results showed that interaction effect of mycorrhizal fungi and irrigation levels was significant at 1% level on tomato properties (except for N concentration in leaves). Mean comparison with Duncan`s test proved that increase in drought stress caused decrease in all tomato properties but fungi inoculation could mitigate water shortage in comparison to non-inoculated tomatoes Higher drought stress decreased colonization significantly. Drought stress caused significant deficiency in nutrition elements such as N, P and Fe, however some elements such as K, Zn and Mn were increased in medium drought stress. Inoculation with all mycorrhizal fungi improved nutrition elements concentration in tomato leaves. Water use efficiency was increased 67.9, 49.6 and 52.1 percent from non to medium and high stress respectively in tomatoes treated with Glomus intraradices.
Conclusion: Using of mycorrhizal fungi specially Glomus intraradices and Glomus caledonium increased tomato water use efficiency and improved growth properties due to the increment of water resistance ability in plant. This phenomenon is caused by higher nutrition elements uptake by roots and optimization of water relationship in tomato in the presence of mycorrhizal fungi. According to results, it is suggested tomato roots inoculation with mycorrhizal fungi especially in regions with high water stress potential.
vahideh Shaabani Zenoozagh; Nasser Aliasgharzad; Jaffar Majidi; Roghaieh Hajiboland; Behzad Baradaran; Leili Aghebati-Maleki
Abstract
Introduction: Glomalin is a specific glycoprotein produced by the fungi belonging to phylum Glomeromycota and plays a key role in soil carbon and nitrogen storage. This also has a significant role in the stable aggregates formation and establishment of microbial communities in soil. Assimilated plant ...
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Introduction: Glomalin is a specific glycoprotein produced by the fungi belonging to phylum Glomeromycota and plays a key role in soil carbon and nitrogen storage. This also has a significant role in the stable aggregates formation and establishment of microbial communities in soil. Assimilated plant C which is allocated to the mycorrhizal fungus, appears as a recalcitrant glycoprotein (glomalin) in cell walls of hyphae and spores. Considering global warming due to increasing greenhouse gases, this phenomenon cab be important in carbon sequestration and reducing CO2 in atmosphere. Chemical fertilizers can affect symbiotic relations of these fungi, which in turn affect glomalin production.
Materials and Methods: In a factorial completely randomized design with three replication, clover plants (Trifolium repense L.) were included with Rhizophagus irregularis and/or Rhizobium leguminosarum bv. Trifolii. Four levels of nitrogen (0, 2, 6 and 10 mM as nitrate) in Newman & Romheld nutrient solution were applied to the pots containing 1.5 kg sterile sand. The pots were daily irrigated with nutrient solution containing the above-mentioned levels of nitrogen. Clover plants were excised after 12 weeks of growth. Fine roots were cleaned with %10 KOH and then stained using lactoglycerol trypan blue. Root colonization percentage was determined by grid line intersections method (GLM) described by Norrif et al (1992). For glomalin extraction, hyphal or root samples were autoclaved at 121 ⁰C with 50 mM sodium citrate buffer for 60 min in three cycles. Sand glomalin (SG) and root glomalin (RG) were measured by Bradford method after extraction. Nitrogen concentration in shoot and root was measured according to the standard method.
Results and Discussion: By increasing nitrogen level, the SG significantly decreased (p < 0.01), and at 2 mM, a 63.5 % decrease in SG was observed with relative to the nitrogen-free control. In the rhizobial treated pots, SG production increased by fungal inoculation (p < 0.01). The interaction between bacteria and AM was also significant in production of SG. At the presence of rhizobium bacteria, glomalin production by AM fungi increased significantly. The changes of glomalin content were not impacted by the presence of bacteria in the uninoculated pots with fungi. The highest amount of SG was recorded in the co-inoculated plants with nitrogen-free level. The amount of RG enhanced by increasing nitrogen concentration in nutrient solution. At 10 mM, RG increased by 12.90 %, 11.91 % and 1.44 % compared to the levels of 0, 2 and 6 mM, respectively. As the nitrogen level increased, the percentage of root colonization increased with respect to the control. Nitrogen concentration in shoot and root was enhanced by N increment to 10mM.
Conclusion: Carbon sequestration via glomali synthase by AM fungi is an important pathway for capturing CO2 from atmosphere. Field management measures help AM development of glomalin production. Based on our results, co-inoculated plants with AM and rhizobuim seem to positively affect the production of this glycoprotein. On the other hand, SG decreased significantly by increasing nitrogen concentrations in the nutrient solution. RG, however, increased significantly as a result of increased nitrogen in both fungal inoculations. The highest amount of RG was recorded in the co-inoculated plants with 10mM level. Glomalin synthesis by the fungi is positively affected by the soil nitrogen availability. Nitrogen is the main constituent of this glycoprotein. Plant photosynthates are translocated to the fungal organs via roots and mainly utilized for glomalin synthesis in hyphal and spore cell walls. During this process, nitrogen plays an important role as a constituent of the glycoprotein. The Bradford method was used for glomalin determination in this study. The method is not specific for glomalin and can also measure other glomalin related proteins and glycoproteins. Other proteins increased by N fertilization can hence be measured based on Bradford method. Once plant assimilates are translocated to the fungi, they may be transformed to the nitrogenous compounds if sufficient nitrogen sources are available. Accordingly, a considerable amount of fixed carbon is assimilated in fungal organs and soil particles. It can be concluded that carbon sequestration by arbuscular mycorrhizal symbiosis in terrestrial ecosystems can be improved by N fertilization at optimum level. In addition, the presence of rhizobium bacteria can meet the nitrogen requirement of plants through biological stabilization of nitrogen.
Elham Malekzadeh; Jafar Majidi; Nasser Aliasgharzad; Jalal Abdolalizadeh
Abstract
Introduction: Glomalin is known as a specific fungal glycoprotein belonging to the order Glomerales in phylum Glomeromycota and has been introduced as a heat shock protein. We hypothesised that increasing the level of Pb would lead to increase in glomalin production. Glomalin is usually determined by ...
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Introduction: Glomalin is known as a specific fungal glycoprotein belonging to the order Glomerales in phylum Glomeromycota and has been introduced as a heat shock protein. We hypothesised that increasing the level of Pb would lead to increase in glomalin production. Glomalin is usually determined by two methods, the Bradford protein dye-binding assay and the enzyme-linked immunosorbent assay (ELISA). Since many laboratories are not equipped to carry out the ELISA assay, many studies have measured glomalin-related soil protein using the Bradford colorimetric total protein assay. While, the ELISA method specifically measures glomalin by using monoclonal antibody MAb32B11.
Materials and Methods: The pot experiment was conducted in the sterile free-glomalin sand with Trifolium repens L. mycorrhized by Rhizophagus irregularis fungus and treated with the Pb levels of 0, 150, 300 and 450 µM. Thus, in vitro experiment was performed in two-compartments plates containing of the transformed carrot roots (Daucus carota L.) mycorrhized with the same fungus in root compartment and hyphal compartment treated with the Pb levels of 0, 0.01, 0.1 and 1 mM as Pb(NO3)2. For glomalin extraction, hyphal or root samples were autoclaved at 121 ⁰C with 50 mM sodium citrate buffer for 60 min in three cycles. Protein concentrations in the extracted samples were determined using a modified Bradford protein assay. Also, glomalin content in the samples were determined by indirect ELISA using monoclonal antibody MAb32B11. The percentages of the total root length were colonised by mycorrhizal fungi in pot culture and both hyphal and spore densities in the metal-containing hyphal compartment were determined.
Results and Discussion: In the in vitro culture the percentage of total hyphae and spore frequency decreased, while Bradford reactive total hyphal protein (BRHP) and Immunoreactive hyphal protein (IRHP) in hyphal compartment increased as the concentrations of Pb increased. Also, there was positive and significant correlation between immunoreactive hyphal protein (IRHP) and Bradford reactive total hyphal protein (BRHP) in hyphal compartment of in vitro culture (r= 0.941**). In the pot culture, the percentage of the total mycorrhized root length in all the treatments increased compared to the unleaded control as the concentrations of Pb raised. In general, Bradford reactive total protein and Immunoreactive protein in both the hyphal and root compartments of pot culture increased with increasing the Pb levels. Also, there were positive and significant correlations between immunoreactive hyphal protein (IRHP) with Bradford reactive total hyphal protein (BRHP) (r= 0.845 **) and immunoreactive root protein (IRRP) with Bradford reactive total root protein (BRRP) (r= 0.706 **) in pot experiment. Some previously researches had reported correlation between ELISA with Bradford contents, whether, Bradford and ELISA values were nearly the same, this means that the extraction process mostly separates glomalin. The results of non-mycorrhizal roots indicated that a small proportion of root protein is cross-reactive with the MAb32B11 antibody. There are some evidences that MAb32B11 is slightly cross-reactive with plant compounds, non-AMF species, and non-target proteins present in large concentration, such as BSA. Additionally, we found the increasing of BRRP contents of AMF-colonized root compared to the non-mycorhizal roots. This may be as a result of uptake and storage of arginine within AMF intraradical hyphae. Considering IRHP to BRHP ratio indicates that immunoreactivity percentage enhances by rising Pb levels. Immunoreactivity indicates a molecular configuration similar to the configuration of glomalin on hyphae. Our findings are in agreement with previous observations confirming that the toxicity-induced stress by metals may be enhancing glomalin production by AMF. The sequence of the glomalin gene revealed that is likely a 60-KDa heat-shock protein (Hsp) homolog. Glomalin relation with the heat shock proteins clarifies how stress is imposed by heavy metals may rapidly increase glomalin production by AMF and its concentrations in polluted soils.
Conclusion: The high contents of glomalin along with the increasing of Pb concentrations may be explained by the overexpression of this protein. This suggests that under Pb-induced stress and the toxic effect of Pb, the fungus exerts a protective mechanism against toxicant. Therefore, glomalin as a heat shock protein can involve in the reduction of possible cytosolic damages and the transfiguration of proteins under Pb toxicity. We can conclude that glomalin may reduce toxic elements availability via their stabilization and decrease their toxicity risk to other microorganisms and plants in heavy metal polluted sites.
F. Aghababaei; F. Raiesi; alireza hosseinpuor
Abstract
Soil biota such as earthworms and arbuscular mycorrhizal fungi (AMF) play an important role in the stability of ecosystem, and the bioavailability of soil elements, in particular heavy metals, in soils. To examine the effects of these organisms, a 3×2×3 factorial experiment arranged as randomized complete ...
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Soil biota such as earthworms and arbuscular mycorrhizal fungi (AMF) play an important role in the stability of ecosystem, and the bioavailability of soil elements, in particular heavy metals, in soils. To examine the effects of these organisms, a 3×2×3 factorial experiment arranged as randomized complete design was set up to study the individual and combined influence of earthworms (Lumbricus rubellus L.) and AMF (Glomus mosseae and Glomus intraradices) on soil organic matter (OM), dissolve organic carbon (DOC), soil respiration, microbial biomass carbon (MBC), soil enzyme activity and glomalin production in a calcareous soil contaminated with 0, 10, 20 mg of Cd kg-1 soil cropped with sunflower (Helianthus annuus L.) with three replications. Both earthworms and mycorrhizal fungi were able to survive in all the treatments with added Cd. Results showed that Cd pollution decreased all the measured microbial activities and properties in soil. Earthworm treatment increased DOC by 4-10% at all Cd levels. The amount of soil MBC in mycorrhizal treatments was greater (1.9-2.4 times) than that in non-mycorrhizal treatment, and AMF inoculation increased MBC/TOC ratio from 23% to 53% in Cd-polluted soils. Earthworm and AMF enhanced soil enzyme activity/MBC ratio, 10-18 and 40-54% for soil alkaline phosphatase and 4-9 and 40-55% for soil urease, respectively. The glomalin production increased at 20 mg kg-1 and was about 15% greater in G. mosseae than in G. intraradices species. Although soil respiration was decreased substantially with Cd pollution, inoculation of either earthworms or AMF enhanced soil respiration when compared with the corresponding controls.
samaneh ahmadi gheshlaghi
Abstract
Glomalin is a glycoprotein produced by arbuscular mycorrhizal (AM) fungi, and is a major component of soil organic matter, which plays an important role in soil aggregation and carbon sequestration. Glomalin is produced only by the AM fungi. On the other hand, stressful environments such as salinity ...
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Glomalin is a glycoprotein produced by arbuscular mycorrhizal (AM) fungi, and is a major component of soil organic matter, which plays an important role in soil aggregation and carbon sequestration. Glomalin is produced only by the AM fungi. On the other hand, stressful environments such as salinity can affect the AM fungi .The purpose of this study was to investigate the effect of NaCl salinity on glomalin production by Glomerales in symbiosis with corn plant. A factorial experiment was conducted in completely-randomized design (CRD) with four replications in a greenhouse. Factors were NaCl salinity with three levels (S0: 1.34, S1:4 and S2: 8 dS/m) and mycorrhizal fungi with four levels (non mycorrhizal, Glomus versiforme, G. intraradices, G. etunicatum). The results showed that the interaction of salinity and mycorrhizal fungi on plant dry weight, leaf proline, root colonization percentage, EEG and TG was significant at p
