The inoculation response of single arbuscular mycorrhiza fungi (AMF) or rhizobia (Rh) in relation to nitrogen (N) acquisition of plants is well established, while the combined effect of both AMF and Rh is poorly known with regard to N assimilation for changes in amino acids of white clover (Trifolium repens). A pot study was carried out to evaluate the effect of single versus dual inoculation of AMF (Rhizophagus intraradices) and rhizobium (Rhizobium trifolii) on plant growth, leaf and root N contents, root amino acid contents, and root N-related enzyme activities in white clover. One hundred days after inoculations, Rh inoculation significantly stimulated the root colonization by R. intraradices. A single inoculation of AMF or Rh improved the plant growth (biomass production, root projected area, and root volume), root N acquisition, and dual inoculation of AMF and Rh further expanded some of these positive effects on root projected area and root N contents than single inoculation. All the inoculations notably increased activities of root asparagines synthase, nitrate reductase, and glutamate synthase, whilst dual inoculation displayed a much stronger effect in asparagine synthase activity than single inoculation. Single Rh treatment increased root glutamate and proline content, single AMF inoculation induced an increase in glutamate, aspartate, arginine, and ornithine content, while dual inoculation stimulated the accumulation of aspartate and proline. These results suggested the cooperation between the AMF and Rh inoculations, which magnified the positive effect on partly N metabolites and N-assimilation relevant enzymes of white clover.
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Akhtar MS, Siddiqui ZA (2008) Biocontrol of a root-rot disease complex of chickpea by Glomus intraradices, Rhizobium sp. and Pseudomonas straita. Crop Prot 27:410–417
Armengaud P, Thiery L, Buhot N, March GG, Savouré A (2004) Transcriptional regulation of proline biosynthesis in Medicago truncatula reveals developmental and environmental specific features. Physiol Plant 120:442–450
Bago B, Vierheilig H, Piché Y, Aguilar CA (1996) Nitrate depletion and pH changes induced by the extraradical mycelium of the arbuscular mycorrhizal fungus Glomus intraradices grown in monoxenic culture. New Phytol 133:273–280
Blilou J, Ocampo J, Garcia-Garrido J (1999) Resistance of pea roots to endomycorrhiza fungus or Rhizobium correlates with enhanced levels of endogenous salicylic acid. J Exp Bot 50:1663–1668
Bulgarelli RG, Marcos FCC, Ribeiro RV, Andrade SAL (2017) Mycorrhizae enhance nitrogen fixation and photosynthesis in phosphorus-starved soybean (Glycine max, L. Merrill). Environ Exp Bot 140:26–33
Clark RB, Zeto SK (2000) Mineral acquisition by arbuscular mycorrhizl plants. J Plant Nutr 23:867–902
Cruz C, Egsgaard H, Trujillo C, Ambus P, Requena N, Martins-Loução MA, Jakobsen I (2007) Enzymatic evidence for the key role of arginine in nitrogen translocation by arbuscular mycorrhizal fungi. Plant Physiol 144:782–792
Dunn MF (2015) Key roles of microsymbiont amino acid metabolism in rhizobia-legume interactions. Crit Rev Microbiol 41:411–451
Foyer CH, Valadier MH, Migge A, Becker TW (1998) Drought-induced effects on nitrate reductase activity and mRNA and on the coordination of nitrogen and carbon metabolism in maize leaves. Plant Physiol 117:283–292
Franzini VI, Azcon R, Mendes FL, Aroca R (2010) Interactions between Glomus species and Rhizobium strains affect the nutritional physiology of drought-stressed legume hosts. J Plant Physiol 167:614–619
Guo Q, Love J, Roche J, Song JC, Turnbull MH, Jameson PE (2017) A RootNav analysis of morphological changes in Brassica napus L. roots in response to different nitrogen forms. Plant Growth Regul 83:83–92
Hack CM, Porta M, Schäufele R, Grimoldi AA (2019) Arbuscular mycorrhiza mediated effects on growth, mineral nutrition and biological nitrogen fixation of Melilotus alba Med. in a subtropical grassland soil. Appl Soil Ecol 134:38–44
Hajong S, Kumaria S, Tandon P (2013) Comparative study of key phosphorus and nitrogen metabolizing enzymes in mycorrhizal and non-mycorrhizal plants of Dendrobium chrysanthum Wall. ex Lindl. Acta Physiol Plant 35:2311–2322
He JD, Chi GG, Zou YN, Shu B, Wu QS, Srivastava AK, Kuča K (2020) Contribution of glomalin-related soil proteins to soil organic carbon in trifoliate orange. Appl Soil Ecol 154:103592
Husted S, Mattsson M, Mollers C, Wallbraun M, Schjoerring JK (2002) Photorespiratory NH4+ production in leaves of wild-type and glutamine synthetase 2 antisense oilseed rape. Plant Physiol 130:989–998
Jin HR, Pfeffer PE, Douds DD, Piotrowski E, Lammers PJ, Shachar-Hill Y (2005) The uptake, metabolism, transport and transfer of nitrogen in an arbuscular mycorrhizal symbiosis. New Phytol 168:301–310
Joy KW, Ireland RJ, Lea PJ (1983) Asparagine synthesis in pea leaves, and the occurrence of an asparagine synthetase inhibitor. Plant Physiol 73:165–168
Lakshman HC, Patil GB (2004) Dual inoculation of VA-mycorrhiza Rhizobium beneficial to Acacia nilotica wild. Ecol Environ Conserv 10:461–464
Larimer AL, Clay K, Bever JD (2014) Synergism and context dependency of interactions between arbuscular mycorrhizal fungi and rhizobia with a prairie legume. Ecology 95:1045–1054
Lichtenthaler HK, Wellburn RR (1983) Determinations of total carotenoids and chlorophylls a and b of leaf extracts in different solvents. Biochem Soc Trans 11:591–592
Liyanaarachchi GV, Mahanama KR, Somasiri HP, Punyasiri PAN (2018) Development and validation of a method for direct, underivatized analysis of free amino acids in rice using liquid chromatography-tandem mass spectrometry. J Chromatogr A 1568:131–139
Liu CY, Zhang F, Zhang DJ, Zou YN, Shu B, Wu QS (2020) Transcriptome analysis reveals improved root hair growth in trifoliate orange seedlings by arbuscular mycorrhizal fungi. Plant Growth Regul 92:195–203
Lüscher A, Mueller-Harvey I, Soussana JF, Rees RM, Peyraud JL (2014) Potential of legume-based grassland–livestock systems in Europe: a review. Grass Forage Sci 69:206–228
Majumdar R, Barchi B, Turlapati SA, Gagne M, Minocha R, Long S, Minocha SC (2016) Glutamate, ornithine, arginine, proline, and polyamine metabolic interactions: the pathway is regulated at the post-transcriptional level. Front Plant Sci 7:1–17
Martin F, Cliquet JB, Stewart G (2001) Nitrogen acquisition and assimilation in mycorrhizal symbioses. In: Lea PJ, Morot-Gaudry JF (eds) Plant nitrogen. Springer, Berlin, pp 147–166
Meng LL, He JD, Zou YN, Wu QS, Kuča K (2020) Mycorrhiza-released glomalin-related soil protein fractions contribute to soil total nitrogen in trifoliate orange. Plant Soil Environ 66:183–189
Ndoye F, Kane A, Diedhiou AG, Bakhoum N, Fall D, Sadio O, Sy MO, Noba K, Diouf D (2015) Effects of dual inoculation with arbuscular mycorrhizal fungi and rhizobia on Acacia senegal (L.) Willd. seedling growth and soil enzyme activities in senegal. Int J Biosci 6:36–48
Neumann U, Kosier B, Jahnke J, Priefer UB, Al-Halbouni D (2011) Soil factors exhibit greater influence than bacterial inoculation on alfalfa growth and nitrogen fixation. FEMS Microbiol Ecol 77:590–599
Phillips JM, Hayman DS (1970) Improved procedures for clearing roots and staining parasitic and vesicular-arbuscular mycorrhizal fungi for rapid assessment of infection. Trans Br Mycol Soc 55:158–161
Ren CG, Kong CC, Wang SX, Xie ZH (2019) Enhanced phytoremediation of uranium-contaminated soils by arbuscular mycorrhiza and rhizobium. Chemosphere 217:773–779
Saxena AK, Rathi SK, Tilak KVBR (1997) Differential effect of various endomycorrhizal fungi on nodulating ability of green gram by Bradyrhizobium sp. (Vigna) strain S24. Biol Fert Soils 24:175–178
Singh RP, Srivastava HS (1986) Increase in glutamate synthase (NADH) activity in maize seedlings in response to nitrate and ammonium nitrogen. Physiol Plant 66:413–416
Tanaka Y, Yano K (2005) Nitrogen delivery to maize via mycorrhizal hyphae depends on the form of N supplied. Plant Cell Environ 28:1247–1254
Tuo XQ, Li H, Zou YN (2017) Alleviation of drought stress in white clover after inoculation with arbuscular mycorrhizal fungi. Not Bot Horti Agrobo 45:220–224
Wagemaker MJM, Eastwood DC, Drift CVD, Jetten MSM, Burton K, Griensven LJV, Camp HJOD (2007) Argininosuccinate synthetase and argininosuccinate lyase: two ornithine cycle enzymes from Agaricus bisporus. Mycol Res 111:493–502
Wild B, Alves RJE, Barta J, Capek P, Gentsch N, Guggenberger G, Hugelius G, Knoltsch A, Kuhry P, Lashchinskiy N, Mikutta R, Palmtag J, Prommer J, Schnecker J, Shibistova O, Takriti M, Urich T, Richte A (2018) Amino acid production exceeds plant nitrogen demand in Siberian tundra. Environ Res Lett 13:034002
Wu QS, He JD, Srivastava AK, Zou YN, Kuča K (2019) Mycorrhizas enhance drought tolerance of citrus by altering root fatty acid compositions and their saturation levels. Tree Physiol 39:1149–1158
Xavier LJC, Germida JJ (2003) Selective interactions between arbuscular mycorrhizal fungi and Rhizobium leguminosarum bv. viceae enhance pea yield and nutrition. Biol Fertil Soils 37:262–267
Yaseen T (2016) Effect of rhizobium, VAM and rock phosphate inoculation on growth and productivity and isolation of VAMF spores of Pisum sativum. Pesqui Agropecu Bras 5:564–572
Yang L, Zou YN, Tian ZH, Wu QS, Kuča K (2021) Effects of beneficial endophytic fungal inoculants on plant growth and nutrient absorption of trifoliate orange seedlings. Sci Hortic 277:109815
Yoneyama T, Suzuki A (2019) Exploration of nitrate-to-glutamate assimilation in non-photosynthetic roots of higher plants by studies of 15N-tracing, enzymes involved, reductant supply, and nitrate signaling: a review and synthesis. Plant Physiol Bioch 136:245–254
Zhang F, Wang P, Zou YN, Wu QS, Kuča K (2019) Effects of mycorrhizal fungi on root-hair growth and hormone levels of taproot and lateral roots in trifoliate orange under drought stress. Arch Agron Soil Sci 65:1316–1330
Zhang F, Zou YN, Wu QS, Kuča K (2020) Arbuscular mycorrhizas modulate root polyamine metabolism to enhance drought tolerance of trifoliate orange. Environ Exp Bot 171:103962
Zhao XY, Zhou S, Wang G, Xing G, Shi W, Xu RK, Zhu ZL (2012) Nitrogen balance in a highly fertilized rice-wheat double-cropping system in Southern China. Soil Sci Soc Am J 76:1068–1078
Zou YN, Wu QS, Kuča K (2020) Unravelling the role of arbuscular mycorrhizal fungi in mitigating the oxidative burst of plants under drought stress. Plant Biol. https://doi.org/10.1111/plb.13161
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Xie, MM., Chen, SM., Zou, YN. et al. Effects of Rhizophagus intraradices and Rhizobium trifolii on growth and N assimilation of white clover. Plant Growth Regul 93, 311–318 (2021). https://doi.org/10.1007/s10725-020-00689-y
- Amino acid
- Asparagine synthase