Manure substitution of chemical fertilizer affect soil microbial community diversity, structure and function in greenhouse vegetable production systems

Soil microbial community and enzyme activities together affect various ecosystem functions of soils. Fertilization, as important agricultural management practices, are known to modify soil microbial characteristics; however, inconsistent results have been reported. The aim of this research therefore was to make a comparative study of the effects of different fertilization patterns (No N inputs (No N), 100% chemical fertilizer-N (CN) inputs (4/4CN) and different substitution rates of CN by organic manure-N (MN) (3/4CN+1/4MN, 2/4CN+2/4MN and 1/4CN+3/4MN)) on soil physicochemical properties, enzyme activities and microbial attributes in a GVP of Tianjin, China. Manure substitution of chemical fertilizer, especially at higher substitution rate (2/4CN+2/4MN and 1/4CN+3/4MN), improved soil physicochemical properties (higher soil organic C (SOC) and nutrient contents; lower bulk densities), promoted microbial growth (higher total phospholipid fatty acids and microbial biomass C contents) and activity (higher soil hydrolase activities). Manure addition caused a remarkable increase of the fungi/bacteria ratio and a distinct shift in the fungal (bacterial) community to greater abundance of arbuscular mycorrhizal fungi (G+ bacteria) compared with saprotrophic fungi (G− bacteria). These changes drove shifts toward fungal-dominated soil microbial communities and then optimized microbial community structure. Also, manure application increased soil biodiversity (microbial community and enzyme function), indicated by increased Shannon–Wiener diversity. Redundancy analysis indicated that the most possible mechanism of the impacts of different fertilization patterns on soil microbial characteristics may be the mediation of SOC and nutrient (N) availability (especially SOC) in this GVP of China. In conclusion, manure substitution of chemical fertilizer, especially at higher substitution rate, was more efficient for improving soil quality and biological functions.

to greater abundance of arbuscular mycorrhizal fungi (G+ bacteria) compared with saprotrophic fungi 23 (G− bacteria). These changes drove shifts toward fungal-dominated soil microbial communities and 24 then optimized microbial community structure. Also, manure application increased soil biodiversity 25 (microbial community and enzyme function), indicated by increased Shannon-Wiener diversity.

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Redundancy analysis indicated that the most possible mechanism of the impacts of different 27 fertilization patterns on soil microbial characteristics may be the mediation of SOC and nutrient (N) 28 availability (especially SOC) in this GVP of China. In conclusion, manure substitution of chemical 29 fertilizer, especially at higher substitution rate, was more efficient for improving soil quality and 30 biological functions.

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Vegetable production is a vital part of agricultural sector in China. In 2015, the areas of vegetable 36 planting in China were 2.1 × 10 7 ha [1], which was around 9.5 and 210.0 times, respectively, of that in 37 Europe (2.2 × 10 6 ha [2]) and Canada (1.0 × 10 5 ha [3]). Over the last 30 years, greenhouse vegetable 38 productions (GVPs) in China have grown rapidly and become the main type of vegetable production 39 due to its higher economic benefits relative to vegetable productions in open-air fields [4]. As a high-40 intensity agricultural ecosystem, the GVPs has been characterized as large cropping index, high study exploring the changes of soil microbial properties under different fertilization regimes in GVPs.

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Thus, the impact of manure application on soil microbial properties and the underlying mechanisms 65 still need to be investigated in GVPs.

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The mean annual temperature is 11.6°C with mean annual precipitation of 586 mm. The frost-free period

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This study comprised five treatments as follows: (1)  plot was fitted with a water meter to ensure the amount of irrigation applied was accurately controlled.

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If the field capacity dropped below 60%, we irrigated the soil to replenishing water to 75% of field 114 capacity. The total irrigation amounts during the celery and tomato growing periods were 3334 and 3889 115 m 3 ha -1 year -1 .

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After the celery (the fifteenth-season) harvest in January 2017, ten soil cores (3 cm in diameter, 0-20 cm    189 where Pi is the ratio of each enzyme activity to the sum of all enzyme activities.

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The PC loadings for soil enzyme activities (Fig. 3b) and PC scores indicated that higher manure-  were significantly (P < 0.05) higher than those in chemical fertilization treatment (48.46-51.98 nmol 256 g −1 soil), and total PLFA contents increased as the manure application rate increased (Fig. 4a). No 257 significant differences were found in total PLFA contents between chemical fertilization treatments.  (Fig. 4a, 4b, 4c and 4d).

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The ratios of fungi to bacteria (F/B) and arbuscular mycorrhizal fungi to saprotrophic fungi (AMF/SF)

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respectively, compared to those in chemical fertilization treatments ( Fig. 4a and 4c). significantly higher by 26.33% than 4/4CN treatment (Fig. 4b). 269 The PCA of the 24 PLFAs data indicated that soil microbial community structure was markedly

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Soil microbial community and functional (enzyme) diversity in this study varied among different 279 fertilization treatments (

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Compared to the 4/4CN treatment, the (i17:0 + i15:0)/(a17:0 + a15:0) values slightly increased under    (Table 3), which was similar to previous studies [41]. These positive effects could be 311 attributed to manure containing amounts of carbon resources, and subsequently release of these 312 resources into soil [42]. Manure application could improve the nutrient retention ability of soil, which 313 could explain the results described above [43]. Additionally, we found manure addition decreased soil

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AK contents. This is in contrast to most studies that annual addition of manure increased soil AK

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Another possible explanation was that great demands for nutrients during microbial proliferation 319 reduced short-term nutrients (K) availability [48]. In other words, manure application in this study 320 promoted vegetable growth (Table S1) and soil microbial proliferation ( Fig. 1 and Fig. 4), which 321 consumed large amounts of nutrients (K) and therefore reduce soil nutrients (K) availability.   levels ("metrics") of soil microbial community, from a whole-community profile to specific group 358 abundance, microbial diversity, and physiological stress (Fig. 4, Fig. 6 and Table 5

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Manure application not only increased all and individual microbial groups biomass, but also changed 368 microbial community structure (Fig. 4). In line with the results of previous research on agricultural soil

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These changes indicated that some species (fungi, G+ bacteria and AMF) adapted better to manure application than other species (bacteria, G− bacteria and SF). On the basis of previous studies, the 372 utilization of microorganisms for exogenous organic materials usually has a community succession effect:  388 richness (SR), but also reduced soil microbial community evenness (Table 5). It has been suggested 389 that manure application could (1) improve soil structure and (2) increase carbon and nutrient resource 390 diversity, and subsequently improve soil microbial community diversity and richness [62,70].

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Interestingly, different microorganisms have been reported to be differently affected by fertilization,

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owing to their diverse structure and physiology (e.g., manure addition was more favorable to the growth of some species, such as fungi etc.) [71,72]. This could be the reason that soil microbial 394 community evenness was decreased in manure-amended soils.

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Higher values of these indices are associated with decreased bacterial growth rate and increased SOC  represents an important challenge in microbial ecology [49]. RDA (Fig. 7a)