Effects And Mechanism Of Arbuscular Mycorrhizal Fungi On Iron Uptake In Citrus

Arbuscular mycorrhizal(AM) fungus is a kind of microbe widely present in the earth, which could form mutualistic symbioses with host plants.AM fungi play very important role in plant growth due to it's improvement of mineral elements and water.Citrus has no or,if any short root hairs in field and is fairly dependent on AM fungi that are most of Glomus species.Citrus is very susceptible to iron chlorosis that is the most frequent nutritional problem in this type of fruit due to high calcium or high pH in soil,which cause decreased yield and neglectly affected of fruit quality.This study deals with two citrus rootstoks(Poncirus trifoliata(L.) Raf.and Citrus reticulata Blanco) and three arbuscular mycorrhizal fungi of Glomus in order to evaluate the effect and mechanism of AM fungi on iron uptake in citrus,so as to provide basic theory for improvement of mineral elements.The main contents and results in the present study are as follows:(1)Study on effects of AM fungi on citrus and microenvironmentExperiment was carried out in pot culture and effects of three Glomus species(Glomus mosseae,G.versiforme and G.diaphanum) on mycorrhizal development,plant growth and soil microbes in trifoliate orange(Poncirus trifoliata(L.) Raf.) were studied. Different AM fungi showed the difference of inter-species in citrus seedlings.Inoculation with G.versiforme showed the greatest effects compared with G mosseae and G. diaphanum in trifoliate orange seedlings which had highest AM colonization,hyphal density,plant height,stem diameter,root volume and density of bacteria(×10⁶cfu/g), fungus(×10⁶cfu/g) and actonimycetes(×10⁵ cfu/g) in rhizospheric soil.It suggested that G. versiforme was the best in the three AM fungus in citrus.(2)Effects of AM fungus on iron uptake in citrus under different pH conditionsThe effect of the arbuscular mycorrhizal(AM) fungus(Glomus versiforme) on iron contents in two citrus rootstocks[trifoliate orange(Poncirus trifoliata L.Raf) and red tangerine(Citrus reticulata Blanco)]was studied in sand culture under different pH conditions.The experiment was carried out at four pH levels by applying nutrient solution at pH 5.0,6.0,7.0 or 8.0 to P.trifoliata and pH 5.2,6.2,7.2 or 8.2 to C.reticulata.Plant biomass,chlorophyll,leaf active iron and H⁺ excreted were used to analyze the effects of AM fungi on iron uptake.The results indicated that colonization by G.versiforme led to higher plant height,dry weights,chlorophyll content,active iron and total iron of leaves compared with non-mycorrhizal treatments and the maximum extent was 33.34%, 26.83%,32.72%,24.37%and 13.75%.In heigh pH treatments,amounts of H⁺ excreted from the two rootstock roots without AM fungi inoculum showed strong signal of iron deficient.However,H⁺ excreted less in AM treatments,suggesting that.AM fungi improved the iron nutrition in citrus.(3)Function of AM fungi on iron uptake in citrus under iron deficiency and heavy bicarbonate stressEffects of arbuscular mycorrhizal fungus(Glomus versiforme) on iron nutriton of trifoliate orange and red tangerine were investigated with a sand culture under iron deficiency and heavy bicarbonate stress.The experiment was carried out at four pH levels by applying nutrient solution at CK(pH 6.0),-Fe(pH 6.0),CaCO₃(pH 7.0) or CaCO₃ (pH 8.0) to trifoliate orange and CK(pH 6.2),-Fe(pH 6.2),CaCO₃(pH 7.2) or CaCO₃ (pH 8.2) to red tangerine.The plant biomass,chlorophyll,active iron,tissue structure and ratios of mineral elements in leaves,root ferric-chelate reductase(FCR) activities and total phenolics were measured.Results showed that the frontier between palisade and spongy tissue got vague and cells were arranged tightly.The colonization of G versiforme significantly increased active iron accumulation and the sythesis of chlorophyll,improved Fe/Mn and K/Ca,and enhanced activities of POD,CAT and root FCR and the contents of total phenolics in citrus roots,and the maximum extent was 66.67%,22.46%,54.04%, 50.17%,12.87%and 12.70%respectively;decreased P/Fe,50(10P+K)/Fe,and the depth downrange was 52.33%and 35.58%respectively.(4)Relation ship between AM fungi and each iron species in different rhizospherie soil of citrusSand culture was carried out in trifoliate orange and red tangerine inoculded with G versiforme.Seasonal variation of soil iron species such as Exch-Fe,MnOX-Fe,Carb-Fe, OM-Fe,AOFe-Fe,COFe-Fe,RES-Fe,available Fe and glomalin in different soil range (0-2 cm,2-4 cm,4-8 cm) apart from taproot by root confinement were studied in normal management.The results indicated that RES-Fe,OM-Fe and Exch-Fe made up of soil available Fe,and Carb-Fe and AOFe-Fe showed the negative effects.Colonization of G versiforme decreased RES-Fe,OM-Fe and Exch-Fe,and the depth downrange was 55.27%,42.15%and 59.10%respectively,improved available Fe uptake.Furthermore,Fe, Mn,Cu and Zn could be sequestered by soil glomalin,showing the buffer action of iron deficient.(5)FRO1(FCR gene) gene clone in citrus root and effects of AM fungi on FRO1 expressionTotal RNA was extracted from trifoliate orange and red tangerine roots and cDNA fragment encoding FRO1 gene was obtained by RT-PCR.Effects of AM fungi on FRO1 expression was studied via Real time PCR.Results showed that the partial cDNA length of FRO1 gene in trifoliate orange and red tangerine was 350 bp in size,and above 80% homology was found with Pisum sativum and Medicago truncatula.Inoculated with AM fungi enhanced the gene expression during different stage and higher in iron dificient treatment than normal iron conditions.