Contents lists available at ScienceDirect Applied Soil Ecology journal homepage: www.elsevier.com/locate/apsoil Contribution of glomalin-related soil proteins to soil organic carbon in trifoliate orange Jia-Dong He a , Ge-Ge Chi a , Ying-Ning Zou a , Bo Shu a , Qiang-Sheng Wu a,b, ⁎ , A.K. Srivastava c , Kamil Kuča b a College of Horticulture and Gardening, Yangtze University, Jingzhou, Hubei 434025, China b Department of Chemistry, Faculty of Science, University of Hradec Kralove, Hradec Kralove 50003, Czech Republic c ICAR-Central Citrus Research Institute, Nagpur 440033, Maharashtra, India ARTICLE INFO Keywords: Carbon pool Citrus Extraradical hyphae Glomalin Rhizosphere ABSTRACT Arbuscular mycorrhizal fungi (AMF) produce glomalin-related soil protein (GRSP) that influences organic carbon (C) storage in soil; however, how much purified GRSP fractions contribute to soil organic carbon (SOC) is yet not known. The present study evaluated the contribution of GRSP towards changes in SOC in trifoliate orange grown in a rootbox divided into a roots + hyphae chamber (roots colonized by AMF hyphae and AMF extra- radical hyphae) and a hyphae chamber (only the presence of AMF extraradical hyphae, without roots). Three AMF species (Diversispora epigaea, Paraglomus occultum and Rhizoglomus intraradices) were inoculated into the roots + hyphae chamber. Following four months of plant growth, P. occultum showed higher AMF hyphal growth in roots, soils and nylon mesh than D. epigaea or R. intraradices. Mycorrhizal inoculation improved the plant growth performance and increased easily extractable GRSP (EE-GRSP) and difficultly extractable GRSP (DE-GRSP) concentrations in both chambers, regardless of AMF species. The C content observed in total GRSP of the soil after purification was 2.71 ± 0.49 mg g -1 , while purified EE-GRSP and DE-GRSP showed the C content of 1.01 ± 0.19 mg g -1 and 1.70 ± 010.34 mg g -1 , respectively. The C contribution by purified EE-GRSP and DE-GRSP accounted for 8.67 ± 0.95% and 14.59 ± 2.21%, respectively, of total SOC, with a total C con- tribution of purified GRSPs accounting for 23.26 ± 2.67% of total SOC. A significantly higher C content of GRSP and the C contribution of GRSP to SOC were observed in DE-GRSP than in EE-GRSP, as well as the soil of the roots + hyphae chamber than the soil of the hyphae chamber. The proportionate distribution of water-stable aggregate in 2–4 and 1–2 mm sizes and their stability were higher under AMF hyphae than under non-AMF hyphae. This study thus provided a database evidence of increased contribution of GRSP towards build-up of SOC in response to mycorrhizal symbiosis. 1. Introduction Arbuscular mycorrhizal fungi (AMF) are one of the most pre- dominant soil microbial communities and can establish symbiotic as- sociations, namely, arbuscular mycorrhizas (AMs), with up to 80% of terrestrial plants (Keymer and Gutjahr, 2018). AMF are known to fa- cilitate several important functions of host plants including improved plant performance, changed rhizosphere microbial diversity, and en- hanced ability of plants to tolerate abiotic stress (Park et al., 2016; Jiang et al., 2017; Wu et al., 2019; Zhang et al., 2020). On the other hand, hyphae and spores of AMF secrete a kind of glycoprotein (glo- malin), which is deposited into the soil named as glomalin-related soil protein (GRSP) (Wright and Upadhyaya, 1996; Rillig, 2004; Schindler et al., 2007). The GRSPs are broadly divided into three types: easily extractable GRSP (EE-GRSP), difficultly extractable GRSP (DE-GRSP), and total GRSP (T-GRSP) as the sum of EE-GRSP and DE-GRSP (Wu et al., 2014). The carbon (C) stored in GRSP is considered highly re- calcitrant in nature, lasting for a minimum of 12–22 years (Zou et al., 2016), and plays an active role in the development of soil structure (Chi and Wu, 2017). AMF are reported to consume 4–20% of the photosynthates syn- thesized by host plants. In return, C from mycorrhizas is deposited into the soil via extraradical hyphae as sink for C storage, where mycorrhizal contribution towards organic C accumulated into soil ecosystem in- volved approximately 54–900 kg hm -2 (Zhu and Miller, 2003). Col- lectively, AMF hyphae and AMF-released GRSP are important https://doi.org/10.1016/j.apsoil.2020.103592 Received 12 November 2019; Received in revised form 10 March 2020; Accepted 15 March 2020 ⁎ Corresponding author at: College of Horticulture and Gardening, Yangtze University, Jingzhou, Hubei, 434025, China. E-mail address: wuqiangsheng@yangtzeu.edu.cn (Q.-S. Wu). Applied Soil Ecology 154 (2020) 103592 0929-1393/ © 2020 Elsevier B.V. All rights reserved. T