Integrated application of biochar and chemical fertilizers improves growth and yield of wheat (Triticum aestivum) by altering soil biological heath

Integrated application of biochar and chemical fertilizers improves growth and yield of wheat (Triticum aestivum) by altering soil biological heath | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Integrated application of biochar and chemical fertilizers improves growth and yield of wheat (Triticum aestivum) by altering soil biological heath Muhammad Abdullah Aziz, Khalid Saifullah Khan, Rabia Khalid, Muhammad Shabaan, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2910777/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 17 Feb, 2024 Read the published version in Plant and Soil → Version 1 posted 6 You are reading this latest preprint version Abstract Aim Integration of chemical and organic fertilizers not only improves soil biological health and plant growth but also reduces costs of agricultural production, and hence, is an economically feasible approach to sustain plant growth in developing countries. Methods We performed a field experiment to evaluate the impact of poultry litter derived biochar (BC; 10 and 20 t ha -1 ) with and without chemical fertilizers (CF) on soil nutrient availability, microbial abundance, and soil enzymatic activity. Results Combined application of BC and CF significantly increased soil nutritional status and organic carbon (SOC), and these improvements were more prominent at higher BC level (20 tons ha -1 ), which increased soil microbial biomass carbon, nitrogen, and phosphorus by 27, 58, and 61%, respectively. Furthermore, BC20+CF treatment improved soil microbial abundances such as actinomycetes (24%), bacteria (70%), AMF (49%) and saprophytic fungi (38%). In terms of wheat growth and yield attributes, BC20+CF application enhanced spike length (55%), leaf area index (30%), tillers (12%), along with biological yield (26%) and grain yield (36%). BC20+CF application was also proved to be positively correlated with different soil enzymatic activities i.e., urease (24%), dehydrogenase (44%), and β-glucosidase (93%). Principal component analysis (PCA) analysis proved that co-application of BC @ 20 t ha -1 along with CF was the most efficient treatment in terms of improving soil nutrient status and microbial activities. Conclusions Hence, combined application of BC and CF could be an efficient tool to improve soil health, plant growth and reduce cost of agricultural production. Biochar Soil microbial abundance Soil enzymatic activity Soil fertility Crop yield Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1 Introduction Application of chemical fertilizer is a general practice adopted globally for ensuring increased crop yields and fulfilling the demands of human population (Liu et al. 2017 ). However, their continuous and repeated applications not only adversely affect soil quality but also lead to various environmental constraints such as soil acidification and groundwater pollution (Li et al. 2022 ; van der Bom et al. 2018 ). Furthermore, their excessive applications lead to diminishing returns and other associated economic risks such as increased costs of production followed by lower profit margins (Amjadian et al. 2021b ; Humbert et al. 2016 ). Substitution of chemical fertilizers with different organic amendments (OA) has proved to be a significant approach to improve crop yields, while sustaining soil health and environmental quality (He et al. 2022 ). Organic amendments are vital aspects of organic farming practices, that exert promising impacts on soil health and indigenous microbial communities, which in turn, perform nutrient cycling in soil and improve its fertility status (Chaudhari et al. 2021 ). They are rich source of essential plant nutrients and therefore, can reduce the farmer’s dependence on chemical fertilizers (Singh 2021 ). However, under tropical conditions where temperature is high and rainfall is lower, organic amendments get decomposed very quickly (Doan et al. 2021 ), which also exhibit their significant contribution towards global warming owing to abrupt mineralization (Xia et al. 2020 ). Application of OA can also ameliorate soil degradation and help in rebuilding a healthy and nutritious soil profile (Yang et al. 2021 ). Numerous researchers have reported regarding the beneficial role of OA in improving soil organic carbon, soil structure as well as fertility (Chen et al. 2021 ). For instance, application of livestock manure along with chemical fertilizers has reportedly resulted in enhanced soil C sequestration, soil fertility status and ultimately, the plant productivity (Deng et al. 2020 ). Palmer et al., ( 2017 ) reported that decomposition of soil organic matter (SOM) releases nitrogen (N), which can enhance crop yields under N limited conditions, as OA ensure slow and sustained release of essential nutrients to soil (Palmer et al. 2017 ), and thereby, their prolonged availability for plant uptake is enabled (Yang et al. 2021 ). In another study, Zhai et al. ( 2022 ) suggested that partial substitution of chemical fertilizers by organic fertilizers can improve the growth and physiological attributes of maize and can also improve soil quality. Among various amendments, biochar (BC) application as an organic amendment, has gained much attention, as it encompasses several positive implications on soil fertility, nutrient cycling, moisture retention, structure and carbon sequestration (Alkharabsheh et al. 2021 ; Elkhlifi et al. 2023 ; Semida et al. 2019 ). It is prepared by pyrolysis (burning of biomasses under oxygen deficient conditions) of different plant materials (Aziz et al. 2023 ) such as crop residue, food wastes, farmyard manure, and has recalcitrant properties, and can persist in soil for hundreds to thousands of years (Brassard et al. 2019 ; Budai et al. 2016 ). BC is comparatively more resistant to microbial degradation than other organic materials, and contains roughly 60% of stable organic carbon (Ameloot et al. 2013b ). Carbon atoms in BC molecules are firmly linked to each other, and therefore, restrict microorganisms to get access to carbon making it relatively more resistant to breakdown than other organic amendment (Lopez-Capel et al. 2016 ). Research on biochar has got significant attention in recent times due to its potential positive impacts towards environment sustainability, carbon sequestration, greenhouse gas emission and land degradation. Peng et al. 2021 stated that application of biochar in combination with N fertilizer positively affected wheat yield and soil nutrients by restraining soil nitrate leaching. Similarly, Sadaf et al. 2017 reported that co-application of biochar and chemical fertilizers led to significant increment in soil nutrient content and their uptake by wheat as compared to alone application of chemical fertilizer or biochar. Biochar also possesses the ability to reduce post application nutrient loss from soil, and thereby, can increase nutrient use efficiency (NUE) due to more uptake by plants. For instance, Win et al. ( 2019 ) reported that application of biochar with or without biofertilizers can improve the NUE particularly N, leading to its improved plant uptake leading to increased plant growth. Hence, incorporation of BC along with CF could be a suitable option to improve plant growth and enhance NUE by plant as availability and uptake of nutrients applied through CF may be increased under their combined application, and chances of nutrient leaching can also be minimized (Sadaf et al. 2017 ). Nutrient cycling in soil is affected by soil microbial processes, which reflect soil health under changing soil conditions. Among them, most important soil biological health indicators are microbial biomass carbon (MBC) and soil enzymes such as phosphatase, dehydrogenase and β-glycosidase (Saha et al. 2019 ). Biochar application has also been proved to be positively correlated with soil biological activities. It has been observed that improvement in wheat quality brought about by high-carbon fertilizers were linked to an increase in different bacterial genera such as Sphingomonas and Haliangium , which supported plant growth and development (Chen et al. 2021 ). Soil microbial activities in terms of soil enzymes exert a significant impact on various soil processes such as OM decomposition, mineralization, and nutrient cycling (Meetei et al. 2020 ; Sharma 2022 ). Charged surfaces present on biochar adsorb various enzyme substrates, increase enzyme reactions, and ultimately leads to better soil enzyme activity (Gasco et al. 2016 ). Alkaline phosphatase, which predominates in alkaline soils, plays a crucial role in the cycling of P by hydrolyzing ester phosphate bonds (Shabaan et al. 2022 ). Biochar also has positive priming effects on soil dehydrogenase activity (Azeem et al. 2019 ). β-glycosidase activity is a foremost indicator to observe the changes occur in soil (Stege et al. 2010 ). This enzyme is involved in hydrolysis of maltose and cellobiose, and acts as a catalyst (Matsuzawa and Yaoi 2017 ). Further, β-glycosidase also acts as an energy sources for the microorganisms during cellobiose degradation (Zang et al. 2018 ). Studies related to the impact of biochar on soil microorganisms are still poorly understood because of variation in feedstock type, pyrolysis temperature and abundance of carbon content in the biochar. Moreover, earlier studies comprising BC mainly focused on its positive role in improving soil fertility and crop growth and scanty data exists in literature regarding its beneficial role in soil biological health. Wheat is major agricultural crop of Pakistan, and acts as a staple food for almost every household, making it the most strategic commodity of the country, whose availability and affordability is the government’s main priority (Rasheed et al. 2021 ). Pakistan is the 8th largest wheat producer worldwide (Ahmad et al. 2021 ), where it is grown on an area of 8.9 million hectares (mha) with an annual production of 2.9 tons per ha (Pakistan 2022 ). It contributes 1.8% in the country’s gross domestic product (GDP), and 7.8% in the value addition in agriculture. Moreover, wheat performs an important role in providing various macromolecules such as carbohydrates, proteins and other essential minerals (Amjadian et al. 2021a ). One of the major reasons behind poor and sub-optimal yield of wheat in developing nations such as Pakistan resides with poor plant nutrition (Iqbal et al. 2015 ; Iqbal et al. 2021 ). Wang et al. ( 2022 ) checked the impact of integrated applications of chemical and organic fertilizers on community and diversity of prokaryotic microorganisms as well as on nutrient absorption by wheat and resultant increment in its growth. They observed that soil supplementation with OA enhanced the SOM contents, diversity of rhizosphere and endophytic microbes, and at the same time, augmented the relative abundance and activities of different microbes in endo- as well as rhizosphere. Thus, it is of great importance to examine the effects of biochar on microbial activities due to their sensitivity to environmental changes. We hypothesized that different doses of biochar with or without NPK fertilizer might stimulate the activity of soil bacteria with subsequent increased soil nutrient content and crop yield. Therefore, objective of this research was to assess the role of individual and combined application of BC and CF in altering the soil nutrient profile, bacterial activity and wheat growth in alkaline soil conditions. 2 Materials and Methods 2.1 Experimental site description Field experiment was performed at Research farm of Pir Mehr Ali Shah Arid Agriculture University, Koont Chakwal (latitude: 33° 38' 49.56" and longitude: 73° 4' 49.08") which is known as Pothwar plateau of northern Punjab. The area has a semi-arid subtropical climate with a pattern of bimodal rainfall occurring in late summer and winter seasons. Usually, around 60–70% of annual rainfall is received during the monsoon season, which runs from June 15th to September 15th. The remaining rainfall occurs during the winter months in the form of gentle showers. Experimental site is classified as Rawal series, Udic Haplustoll Alfisols. Generally, the soil was sandy clay loam (56% sand, 22.8% silt, 21.2% clay). Basic soil properties are given in Table 1 . Table 1 Basic characteristics of experimental site soil and biochar Properties Unit Soil Biochar pH 8.21 8.93 EC dS m − 1 0.32 1.73 Ash % --- 12.51 Total organic C % --- 42.58 NO 3 -N mg kg − 1 3.27 6.83 NH 4 -N mg kg − 1 2.37 7.37 Olsen P mg kg − 1 1.19 4.04 Extractable K mg kg − 1 62.59 92.60 Specific surface area m 2 g − 1 --- 2.88 Moisture content % 9.41 4.51 CEC cmol kg − 1 3.69 16.29 Bulk density gcm − 3 1.46 --- 2.2 Production of Biochar (BC) Biochar was prepared by using poultry litter as a feedstock material, which was collected from Poultry Research Institute Rawalpindi, Pakistan. Production took place at 400℃ for 3 h with 10℃ min − 1 heating rate in a muffle furnace. After that, BC was crushed and sieved from 2 mm sieve in order to get homogenized material prior to field application, and was applied on weight basis. 2.3 Experiment layout and treatment application Before seedbed preparation, field was ploughed twice and harrowed, and weeds and stubbles of previous crop were manually removed. Experimental plots were arranged by following randomized complete block design (RCBD), and each plot carried equal size (4m × 6m). All four treatments including control were applied in triplicates. Treatments of current study were control (C), BC @ 10 t ha − 1 (BC10), BC @ 20 t ha − 1 (BC20), BC 10 t ha − 1 + CF (BC10 + CF) and BC 20 t ha − 1 + CF (BC20 + CF). Furthermore, recommended doses of NPK (120:60:50 kg ha − 1 ) were applied as urea, DAP and sulfate of potash (SOP). 2.4 Soil Sampling Soil samples were collected from replicated plots in May 2022 using a soil auger followed by their pulverization and filtration through 2mm mesh. To evaluate microbial and enzymatic activities, soil samples were kept at 4℃, while for other nutrient analysis and physicochemical analyses, samples were air-dried at room temperature. 2.5 Analysis of soil and biochar physiochemical properties The pH of soil samples was measured by glass electrode method in H 2 O with 1:2.5 ratio of soil and water (Thomas 1996 ), and electrical conductivity (EC) was determined by using an EC meter (Page AL 1982). Soil nitrate nitrogen (NO 3 -N) determination was done by extraction method, where soil samples were extracted with 0.5 M K 2 SO 4 , after extracted the soil samples, the NO 3 -N was measured by using a spectrophotometer at the wavelength of 410 nm (Anderson and Ingram 1993a), for NH 4 -N soil samples were extracted by 2M KCl solution and measured using a spectrophotometer (Shimazadu UV-1280) (Keeney et al. 1983 ). Method of Olsen comprising NaHCO 3 (0.5 M) was used to determine P content from soil samples spectrophotometrically at spectrophotometer (Shimazadu UV-1280) (Olsen 1982). Soil K was determined by extraction of soil samples with 1 N ammonium acetate solution using the flame photometer (Jenway PFP7 flame photometer) (Helmke and Sparks 1996 ). For measuring soil organic carbon (SOC), K 2 Cr 2 O 7 titration method was followed (Nelson et al. 1983). Soil dissolved organic carbon (DOC) was determined by extraction of soil samples with 0.5M K 2 SO 4 (Song et al. 2018b ). Ash content of biochar was calculated by the following question $$Ash \left(\%\right)= \frac{D}{B}\times 100$$ Similarly, organic C content of biochar was calculated by using the following equation $$Organic Carbon \left(\%\right)= \frac{100-Ash}{1.724}$$ Cation exchange capacity (CEC) of BC was measured by mixing 1 N sodium acetate trihydrate (33 mL) with air-dried BC samples, and solution was shaken for five minutes (5000 rpm). After decanting, supernatant was discharged four times and 33 mL ethanol was added into the solution before being centrifuged. Then ammonium acetate solution was used for removing sodium and measured in flame photometer (Chapman 1965 ). The CEC was calculated using the following equation: $$CEC \left(meq 100{g}^{-1}\right)=\frac{meq}{1}Na \left(from calibration curve\right)\times \frac{A}{wt}\times \frac{100}{1000}$$ 2.6 Microbial biomass carbon (MBC) Soil MBC was estimated using fumigation-extraction method. Briefly, 30 mL of alcohol-free chloroform was placed in a beaker and was used as a fumigant, which was placed in a desiccator along with additional beakers filled with soil (10g). After the fumigation period, soil samples were extracted using 50 mL of 0.5 M K 2 SO 4 , and extract was filtered using Whatman 40. Same procedure was repeated for control. To measure the extracted samples, 4 mL of extracts were taken and mixed with 1 mL of potassium dichromate (0.0667 M). Mixture was then heated to 150°C for 30 minutes, and 5 mL of concentrated H 2 SO 4 was added. After adding 3–4 drops of phenanthroline monohydrate indicator, contents were titrated against ferrous ammonium sulfate solution. Following equation was used to calculate C mic (Brookes et al. 1985 ; Wu et al. 1990 ). $${C}_{mic}=\left(Extratcted {C}_{fumigated}-Extracted {C}_{unfumigated}\right)\times 2.64$$ 2.7 Microbial biomass nitrogen (MBN) To estimate MBN, gasification-extraction procedure was performed where extractant was digested in mixture of 4.4 mL of 30% hydrogen peroxide (H 2 O 2 ; 350 mL) and concentrated H 2 SO 4 (20 mL) at 360°C for 2 hours. After digestion, solution was placed in a steam distillation chamber where 25 mL alkaline mixture of NaOH and sodium thiosulfate (500 g and 25 g/L, respectively) were added. Further, boric acid (5 mL) was added to receiving end prior to distillation. Finally, samples were titrated to gray endpoint with hydrochloric acid (HCl; 0.1 N). For precision, amount of HCl was indexed (Brookes et al. 1985 ; Wu et al. 1990 ). $$TN \left(\%\right)=T \times 0.1 \times 0.001*) \times (S/A)/W \times 100/1$$ Where; * refers to the conversion factor (mg to g), T = Corrected titer (mL), S = final digestion solution volume (mL), A = aliquot (sample) volume (mL), and W = sample weight (g). $${N}_{mic}=Extracted {N}_{fumigated}- {Extracted N}_{unfumigated} \times 1.46$$ 2.8 Microbial biomass phosphorus (MBP) To estimate MBP, 2.5 g soil was oven-dried in three repetitions. One replica was incubated for 24 hours at 25°C in alcohol-free chloroform, while other two were aerobically incubated under same conditions. Samples were then shaken continuously for 30 minutes before being extracted with 0.5 M NaHCO 3 . Clear extract of 10 mL was then transferred to a 50 mL volumetric flask, to which 8 mL of reagent (ascorbic acid solution and ammonium molybdate solution) was added. After color development, samples were run on a spectrophotometer (Shimazadu UV-1280) at 882 nm. The KH 2 PO 4 was used for making five different levels of concentrations (0, 0.5, 1, 2, and 4 mg kg − 1 ) as for P standards. Following equation was used for analyzing MBP (Brookes et al. 1982 ; Joergensen et al. 1995 ). $${P}_{mic}= {(P}_{fumigated}- {P}_{unfumigated}) \times 2.5$$ 2.9 Soil enzymatic activities Urease activity (UA) analysis was done by using potassium chloride solution (50 mL). After filtration of soil extract, ammonium content was calculated at 690 nm optical density, and urease activity was determined (Kandeler and Gerber 1988 ). For the estimation of soil dehydrogenase activity (SDA), 5 g moist soil and 5 mL TTC solution were mixed in a test tube, and sealed in tube followed by its incubation for 24 h at 30°C. Later, 40 mL of acetone was added to extract, and incubated for 2 hours under darkness. Finally, soil solution was filtered, and optical density (OD) was measured at 546 nm. Triphenyl formazan (TPF) was developed at 0–40 g TPF mL − 1 for a calibration curve in Tris buffer (pH -7.6) and acetone (Alef 1995 ). Similarly, to estimate soil phosphatase activity (SPA), 10 g of moist soil was mixed with 1.5 mL of toluene and transferred to a 100 mL volumetric flask. Then, 10 mL of disodium phenyl phosphate and 20 mL of buffer were added in it, and incubated at 37°C for three hours. After incubation, extract was made up to 100 mL by adding distilled water and filtered. Then, 8 mL of filtrate was mixed with 5 mL of buffer and diluted up to 25 mL volume. 2,6 dibromo quinone chloramide was added, and final volume was made up to 100 mL. Mixture was then incubated at room temperature for about 30 minutes before reading at 600 nm. For standard calibration, 0-200 µg phenol was used, and phosphatase activity was expressed as phenol (µg g − 1 dwt 2 h − 1 ) (Alef 1995 ). Besides these, protease activity was measured by keeping samples for incubation at 50℃ with sodium caseinate by determining the absorbance at 700 nm of tyrosine content using Folin–Cicalteau reagent (Ladd and Butler 1972 ). Soil β-glucosidase enzyme activity was measured by colorimetric estimation of the p-nitrophenol method (Tabatabai 1982 ). In conditioning cabinet, 1 g of soil was incubated with a buffered solution of p -nitrophenyl-D-glucopyranoside (pH 6.0) and toluene at 37°C for one hour. After incubation, soil suspension was mixed with 1 mL of 0.M CaCl 2 and 4 mL of tris (hydroxymethyl) aminomethane buffer. Yellow filtrate obtained was used to estimate the quantity of p -nitrophenol produced by the hydrolysis of p -nitrophenyl-D-glucopyranoside at 37°C for 1 hour through calorimetric determination. 2.10 Plant Analysis Following the crop harvest, plant growth attributes i.e., root-shoot length, plants fresh-dry weight, and 100 grain weight were measured. An electrical balance was used to calculate the fresh weight of the roots and shoots. To calculate plants dry weight, their samples were oven dried at 65°C for 72 hours. These dried samples were subsequently pulverized, and kept in plastic bags for further chemical analysis. Plant samples were subjected to digestion in a block digester for 2 hours at 360℃. A digestion mixture consisting of Se powder, Li 2 SO 4 , and H 2 SO 4 was added to plant in 100 mL digestion tube. For plant samples, contents of N and P were determined using colorimetric method (Anderson and Ingram 1993b ). Before finding the plant nitrogen on Kjeldahl Nitrogen Analyzer (BKN- 983), the digested samples were mixed with 55 mL of NaOH and 50 mL of DI water (Kjeldahl 1883 ) then run on Kjeldahl apparatus. The P was quantified by mixing 5 mL of digest and 5mL of color reagent (ammonium heptamolybdate-ammonium vanadate) in nitric acid. Later, samples were run on spectrophotometer at 410 nm wavelength to check their absorbance (Shapter 1940 ). Total K contents of plants were determined by wet digestion of plant samples using a mixture of nitric and perchloric acid in a block digester. Then, samples were digested at 230°C for 3 hours and K concentration was determined using a flame photometer (Ryan et al. 2001 ). 2.11 Soil microbial abundance Soil phospholipid fatty acid analysis (PLFA) extraction method used in this study was explained by (Zhang et al. 2014a). Nonadecanoic acid methyl ester (19:0, Sigma) was employed as internal standard. Total microbial biomass was determined by the presence of 33 different PLFAs ranging from C14 to C20 in the samples. Gram-positive bacteria (G+) were indicated by the total of i14:0, a15:0, i15:0, a16:0, a17:0, and i17:0 (Yu et al. 2016 ), while gram-negative bacteria (G-) were represented by sum of 16:12OH, 16:1ω7c, 16:1ω9c, cy17:0, 17:1ω8c, 18:1ω7c, and cy19 (Yu et al. 2016 ). The sum of 18:2ω6,9c and 18:1w9c was used to indicate saprotrophic fungi (SF) (Zhang et al. 2014a), and 16:1w5c was used to indicate arbuscular mycorrhizal fungi (AMF) (Liu et al. 2015 ). The sum of 10Me16:0, 10Me17:0, and 10Me18:0 was employed to indicate actinomycetes, while 20:4ω6,9,12,15c was used to indicate protozoa (Zhang et al. 2014a). 2.12 Statistical Analysis Effects of all treatments (biochar and their interaction co-application with chemical fertilizers) on different physicochemical properties, microbial biomasses, and enzymatic activities in soil were studied by two-way analysis of variance (ANOVA) by using statistical software Statistix 8.1. Treatment means were compared for significant differences by using Tukey’s Honestly Significant Difference (HSD) test at p < 0.05 level (Steel et al. 1997 ). Principal component test (PCA) was carried out to determine the correlation of the environmental variables with all soil microbial, enzymatic and nutrients parameters. 3 Results 3.1 Soil pH and EC Biochar based treatments had variable effects on soil pH and EC. However, no significant impact on soil pH was pragmatic, and only in combined treatment (BC20 + CF), a slight improvement in pH was experienced as compared to sole BC treatments whereas, as compared to control treatment, maximum increment in soil pH was 14% (BC20 + CF). Besides, soil EC showed significant variations according to treatments application, and was significantly ( P < 0.05) increased by 96% under combined application (BC20 + CF), followed by sole application of BC @ 20 t ha − 1 (67%), BC10 + CF (64%) and BC @ 10t ha − 1 by (45%) (Fig. 1 ). 3.2 Soil nutrients content Addition of BC significantly improved soil nutrients status. Among them, soil NO 3 -N was significantly increased by 76% under BC20 + CF treatment however, in sole BC20 treatment this increment was approximately 23% higher than unamended soil. Whilst results of BC10 + CF and BC10 were marginally lower than other treatments except control (Fig. 1 ). Similarly, soil NH 4 -N was also increased under combined application of BC and chemical fertilizers i.e., BC10 + CF and BC20 + CF by 105 and 126% respectively as compared to control. Moreover, there was no statistically significant difference between BC20 and BC10 + CF treatments. Addition of BC led to greater soil P content, where maximum P contents were observed in combined application of BC and CF i.e., BC20 + CF and BC10 + CF exhibiting respective P contents (162 and 141%) as compared to unamended treatment. Sole application of BC10 and BC20 showed 42 and 17% lower P results as compared to their co-application with chemical fertilizers respectively. Soil K, SOC and DOC content had maximum increment of 44, 33 and 130% respectively, under BC20 + CF treatment while sole application of BC20 increased these nutrients status by 25, 28 and 89% respectively. Furthermore, results revealed that BC20 + CF enhanced the soil C/N ratio by 55% whereas BC10 + CF improved it by only 32% (Fig. 1 ). PCA analysis of soil nutrients with treatments also proved that integrated application of BC with CF significantly increased soil nutrient status (Fig. 4 ). 3.3 Microbial biomass and enzyme activity Maximum MBC (484 µg/g) was obtained in BC20 + CF treatment which was 27% higher than unamended control. Similarly, in sole BC20 treatment, this increment was only 21% followed by BC10 + CF and BC10 (17 and 14%) respectively. Likewise, maximum values of MBN and MBP were also obvious in BC20 + CF treated plots, where they were increased by 62 and 71% respectively (Fig. 2 ). However, in sole BC treatment, this increment was marginally lower than combined application (Fig. 4 ). Furthermore, combined application of BC and CF was the most efficient in improving soil enzyme activities i.e., urease activity was maximum under BC20 + CF treatment (24%) followed by BC20 (16%) than control. Moreover, BC10 + CF and BC10 enhanced UA by 14 and 12% respectively as compared to control. Similarly, soil dehydrogenase activity was also maximum under BC20 + CF (44%) treatment than control followed by BC10 + CF (31%), while BC20 and BC10 increased dehydrogenase activities by 26 and 12% respectively. In addition, phosphatase, protease and β-glucosidase activities were increased by 29, 107 and 93% respectively under BC20 + CF followed by BC20 sole application i.e., 23, 69 and 86% respectively (Fig. 2 ). 3.4 Soil microbial community Combine application of BC with CF significantly improved soil microbial biomass for instance PLFA had marginally better results under BC10 + CF (17%) and BC20 + CF (24%) as compare to control. Similarly, AMF and actinomycetes also has the highest value under BC20 + CF application i.e 49% and 24% respectively. F/B also increased under co-application BC10 + CF and BC20 + CF by 20% and 24% respectively (Fig. 3 ). PCA was conducted with 33 PLFA present in soil subjected to all five treatments and described the effect of each treatment on soil microbial community. Both PC1 and PC2 successfully explained about 93.75% and 4.53% of the total sample variation respectively (Fig. 5 ). Along with the PC1, it was observed that BC20 + CF significantly increased soil actinomycetes, SPF, gram+, gram-, total PLFA and F/B while BC10 + CF enhanced the soil fungi and AMF as compared to control. While BC20 increased only soil bacterial content i.e G + and G- in PC2 as compared to control. Overall, C had negative correlation with both PC1 and PC2. Generally, soil microbial abundance was maximum under BC20 + CF followed by BC10 + CF and BC20 however, BC10 did not produce any significant impact on it. 3.5 Plant growth and yield attributes Plant agronomic parameters significantly varied according to biochar application rate and method. Overall, combined application of BC with CF showed maximum improvements in all plant growth attributes i.e., plant shoot and root length were 22 and 37% higher than control under BC20 + CF respectively followed by BC10 + CF (15 and 25% respectively). Spike length, leaf area index and number of tillers’ maximum increment was also under BC20 + CF treatment i.e., 55, 30 and 12% higher than control respectively. BC10 + CF enhanced these parameters by 35, 21 and 8% respectively while, BC20 sole improved them only 26, 21 and 10% respectively. Similarly, plant total N, P, K was also maximum under BC20 + CF amendments i.e 10%, 58% and 80% higher than sole BC20 application (Table 2 ). Table 2 Plant physical and yield attributes under the effect of each treatment. Values denote the mean with 3 replications. Small letters with values indicate significant differences among treatments at the 5% probability level, where significance was tested by using the multiple comparison Tukey’s HSD test. (±) Standard error of the mean (n = 3). Treatments C BC10 BC20 BC10 + CF BC20 + CF Shoot Length (cm) 281.67 ± 14.73c 298.82 ± 11.27c 304.72 ± 8.18bc 325.61 ± 13.12ab 342.31 ± 9.54a Root length (cm) 8.43 ± 2.42b 10.05 ± 1.43ab 10.75 ± 1.56ab 10.47 ± 0.73ab 11.43 ± 0.88a Spike length (cm) 9.03 ± 0.27d 10.11 ± 0.46cd 11.41 ± 1.61bc 12.21 ± 0.86ab 14 ± 0.19a Leaf area index 2.91 ± 0.13c 3.33 ± 0.42b 3.56 ± 0.14ab 4.55 ± 0.17ab 3.81 ± 0.15a Tillers (m 2 ) 378.21 ± 16.11c 392 ± 12.39bc 416.67 ± 17.21ab 410.7 ± 14.38ab 426.61 ± 15.28a 1000 seed weight (g) 35.03 ± 1.81c 36.15 ± 1.42c 38.02 ± 0.31b 38.62 ± 0.91b 40.95 ± 1.11a Bio yield (t ha − 1 ) 8.8 ± 0.51d 9.03 ± 0.10cd 9.93 ± 0.17bc 9.72 ± 0.96b 11.13 ± 0.29a Grain yield (t ha − 1 ) 3.14 ± 0.17d 3.36 ± 0.11c 3.93 ± 0.21b 3.96 ± 0.09b 4.33 ± 0.13a Total N (mg g − 1 ) 1.89 ± 0.10c 1.47 ± 0.16bc 3.17 ± 0.11b 4.21 ± 0.48ab 5.28 ± 0.31a Total P (mg g − 1 ) 1.53 ± 0.13d 1.83 ± 0.09c 3.10 ± 0.14b 4.00 ± 0.71ab 4.91 ± 0.25a Total K (mg g − 1 ) 1.52 ± 0.02c 3.70 ± 0.36bc 5.39 ± 1.14bc 7.14 ± 1.19b 9.72 ± 1.13a Crop yield attributes significantly improved under higher BC application rate and its combination with NPK fertilizers such as BC20 + CF increased crop biological and grain yield by 26 and 36% respectively followed by BC10 + CF where these increments were 10 and 27% higher than control. Furthermore, 1000 grain weight and harvest index also significantly improved up to 17 and 14% respectively under BC20 + CF treatment (Table 2 ). 4 Discussion Biochar application at different rates and their co-application with CF improved soil nutrient status as compare to control (Murtaza et al. 2021 ). We observed that application of biochar improved the nutrient status of soil, and at the same time, enhanced C, N and P availability for microbial utilization as reported by previous studies (Bu et al. 2019 ; Naeem et al. 2019 ; Zhu et al. 2017a ). However, no significant change was observed in soil pH under all the applied BC rates, as previously stated by (Rashid et al. 2020 ) and (Zhu et al. 2017b ), who reported that alkaline soils have large buffering capacity, which resist towards any change in pH brought by BC application. (Griffin et al. 2017 ) found a little increment in soil pH after BC application, but significant enhancement in soil macro nutrients (K, NO 3 -N, Olsen P and NH 4 -N), as observed under current study, which indicate the direct role of BC on soil nutrient stock. Few other studies have indicated that the increase in soil nutrient levels is due to BC's labile C, N, P, and K contents which are continuously released in soil, and are readily available for plant uptake. In simple words, rise in soil nutrients can be attributed to the direct effects of BC on the soil (Aziz et al. 2023 ; Qayyum et al. 2017 ; Song et al. 2018a ). Moreover, BC contains larger surface area, negative surface charge and higher number of functional groups which play a key role against nutrients leaching (Yuan et al. 2016 ). Additionally, presence of oxonium functional groups on BC surface make increase its anion exchange capacity (AEC) which help to retain NO 3 -N and phosphate (Banik et al. 2018 ; Sorrenti et al. 2016 ), while the negative surface charge of BC can retain cations like NH 4 + and K + by sorption (Choudhary et al. 2021 ). It was observed that biochar-based treatments had a key role in improving soil enzyme activities as observed by (Song et al. 2019 ) and (Oladele 2019 ), who examined that presence of organic C, MBC and MBN pools of BC provided organic substrates for enzymes which lead towards higher enzymatic activity. Although, it has also been reported that BC addition lead to a reduction in soil enzyme activities because of synthetic C mineralization (El-Naggar et al. 2015 ). Moreover, few studies revealed that BC can adsorb a range of organic and inorganic molecules, this adsorption causes the blockage of reaction sites which decreases soil enzyme activities (He et al. 2021a ; Lehmann et al. 2011 ). Contrastingly, our study did not support this observation as results revealed that BC addition significantly increased the activity of all enzymes, as supported by the results of (Bailey et al. 2011 ) and (Ameloot et al. 2013a ), who revealed that BC has few volatile compounds which increase soil enzyme activity such as urease, protease, dehydrogenase phosphatase and β-glucosidase under alkaline conditions. Furthermore, BC can also increase enzymatic activity by absorbing soil toxic substances (Salam et al. 2019 ; Song et al. 2022 ) however, there was no sign of toxicity observed in the soil of current study. BC alters the soil biochemical properties (Elzobair et al. 2016 ) and nutrient availability (Lu et al. 2015 ), which lead to higher enzyme activities and therefore, significant correlation between BC concentration and soil enzymatic activity may directly support this suggestion. Interaction between soil enzymes and organic matter in soil determines the biogeochemical stability, which is manifested by the relative proportions of enzymes involved in CNP cycles. The balance between the ratio of microbial biomass and elemental composition of organic matter is essential for maintaining this stability (Waring et al. 2014 ; Wei et al. 2020 ). Inclusively we can suggest that, soil enzyme activity was improved with the increment in BC application rates although sometimes, it is based on the soil nutrients availability and climatic conditions (Waring et al. 2014 ). Generally, it has been reported that BC has the potential to improve soil biological properties, which leads to better soil microbial abundance because of BC's special structure and properties (Lucheta et al. 2016 ; Ren et al. 2022 ; Zhang et al. 2021 ). In current study, a significant increment was observed in soil total PLFA, bacterial and fungal in BC treated soils but this activity increased as the BC application rate increased with CFs. Our findings are closely related to the results of (Chen et al. 2017 ), where they reported that application of 3–9% of BC significantly improved the soil total PLFA content as compare to control. Moreover, the labile C and N pools of BC serve as a suitable habitat (i.e., energy and nutrient sources) for better growth and development of soil microbes (Zhao et al. 2016 ). Previous literature has proved that soil supplementation with biochar and animal manure marginally improves microbial community which lead towards better soil structure and C content (Bowles et al. 2014 ; Cheng et al. 2017 ). Bacterial and fungal biomass was increased with the BC application rate in current study but these results are distinct to the meta-analysis of (Xu et al. 2021 ), as they observed reduction in soil bacterial and fungal community which was proved by previous 107 research studies. Similarly, (Chen et al. 2013 ) also found some negative impacts of wheat straw derived BC on soil bacterial and fungal activity. This variation indicates that BC effects are based on its feedstock type, pyrolysis temperature, soil type and application rate (Xu et al. 2021 ). Further, it has also been revealed that soil microbial biomass is based on BC feedstock type as each feedstock have its own physicochemical characteristics and nutritious values (Luo et al. 2017 ). Additionally, soil pH is also influenced by the BC type, and this increase lead towards better soil microbial abundance. It has also been observed that porous structure of BC can be substantially affected by pyrolysis temperature (He et al. 2021b ), as the temperature rises, BC surface area and pore volume is increased, which provide desired habitat to microorganisms whilst lower BC pore volume and surface area favors the growth of fungal hypha (Muhammad et al. 2016 ). Conclusively, elevated BC pyrolysis temperature and pore spaces could be less conducive for fungal better growth and abundance. Bacterial fungal ratio indicate soil C sequestration capability, as the fungal content upsurges soil, carbon storage potential also increases which plays a pivotal role in environment safety (Malik et al. 2016 ). In our study, soil that received BC20 and BC20 + CF treatments had maximum fungal bacterial ratio, which reflects that application of BC alone or with chemical fertilizers could improve soil C sequestration potential and ecosystem stability. Moreover, in our findings, soil MBC, MBN and MBP were also increased i.e., particularly, at higher dose of BC application. Study of (Bhaduri et al. 2016 ) as well as (Biederman and Harpole 2013 ) support our results, where they observed that application of BC significantly increased soil MBC and MBP respectively. Similarly study of (Zhang et al. 2014b ) also revealed that application of corncob BC significantly increased soil MBC and MBN. Generally, MBC carries approximately 3–7% of the total soil organic carbon while MBN and MBP consist of 1–5% of total soil nitrogen and phosphorus respectively (Oladele et al. 2019 ). These shifts in soil microbial biomasses indicates the soil carbon mineralization, growth and mortality ratio of microbes (Fang et al. 2020 ). Our study revealed that poultry manure derived BC had significant positive effects on wheat crop growth and yield parameters. According to our results, crop growth was significantly improved as the BC application increased with NPK fertilizers. Our results are supported by (Khan et al. 2021 ) and (Aziz et al. 2023 ), as they applied different feedstock based biochars on alkaline soils, and observed a significant increment on crop growth, nutrients uptake and grain yield. Furthermore, it has also been examined that plant response to BC is based on feedstock type, pyrolysis temperature and soil type as each feedstock has its own properties (Choudhary et al. 2021 ). In our previous experiment, we applied three different types of BCs i.e., poultry litter biochar, Acacia modesta wood biochar and Dalbergia sissoo wood biochar, where we discovered that poultry litter biochar significantly influenced wheat growth along with their yield (Aziz et al. 2023 ). Being an organic amendment, BC reduces soil compaction, nutrients loss and increases water holding capacity along with nutrient use efficiency which leads toward improvements in crop agronomic and yield parameters (Khan et al. 2021 ). Generally, BC affect soil nutrients availability by two ways: directly, by providing nutrients (work as nutrient and reduces the nutrient release and leaching) indirectly, by increasing nutrient and water holding capacities of soil as well as by causing variations in soil pH, CEC and soil microbial abundance (Azeem et al. 2021 ). Moreover, BC has large surface area and high porosity, which provide suitable habitat to microbes for survival, which is very critical to nutrient solubilization, for instance, P solubilization leads to higher P uptake, which plays a significant impact on root growth, and ultimately increase nutrients uptake (Azeem et al. 2021 ; Wahid et al. 2020 ). Consequently, better nutrient uptake lead towards higher root-shoot growth and crop yield (Guo et al. 2019 ). Conclusion In our study, we examined the interactive effect of different application rates of poultry litter biochar with chemical fertilizers on soil biochemical characteristics and wheat yield. Soil microbial biomass and enzymatic activities were significantly affected by the sole and co-application of BC20 with chemical fertilizers. Our results also showed that BC20 + CF amendment led to significant improvement in soil nutrient content (i.e. NO 3 -N, NH 4 -H, Olsen P, K, SOC, DOC, C/N). Furthermore, higher rate of BC application along with chemical fertilizers significantly increased soil enzymatic activities (URE, DEH, PHO, PRO and β-GLU) and soil microbial community such as MBC, MBN, MBP, PLFA, AMF, fungi, actinomycetes, bacteria and protozoa. Similarly, a higher fungal: bacteria ratio was also observed under BC20 + CF which indicated higher C sequestration and ecosystem stability. Consequently, growth and yield of wheat crop was considerably enhanced by the application of BC20 + CF. Overall, our study confirmed that co-application of BC with CF is an effective approach to improve soil biochemical characteristics and crop yield. Further studies are needed to evaluate the effect of different feedstock-based biochar and fertilizers on soil bacterial and enzymatic activity for better crop growth and yield. Declarations Acknowledgement The authors are thankful to the Institute of soil and environmental sciences, Pir Mehr Ali Shah Arid Agriculture University Rawalpindi, Pakistan for providing facility to perform the experiment. Author contributions Conceptualization: Muhammad Abdullah Aziz, Khalid Saifullah Khan, Mosaed A. Majrashi; Methodology: Muhammad Abdullah Aziz, Rabia Khalid; Investigation: Khalid Saifullah Khan; Formal Analysis: Muhammad Abdullah Aziz; Data Analysis and visualization: Abdulaziz G. Alghamdi, Zafer Alasmary; Original draft preparation: Muhammad Abdullah Aziz, Muhammad Shabaan; Review and editing: all. Conflict of interest The authors declare no competing interests. References Ahmad TI, Khan RE, Soharwardi MA, Shafiq MN, Gillani S (2021) Socioeconomics and agronomy of wheat yield in cotton-wheat cropping system in Punjab, Pakistan: A quality-quantity assessment. International Journal of Agricultural Extension 9: 69-78. 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Win KT, Okazaki K, Ookawa T, Yokoyama T, Ohwaki Y (2019) Influence of rice-husk biochar and Bacillus pumilus strain TUAT-1 on yield, biomass production, and nutrient uptake in two forage rice genotypes. PLoS One 14: e0220236. Wu J, Joergensen R, Pommerening B, Chaussod R, Brookes P (1990) Measurement of soil microbial biomass C by fumigation-extraction-an automated procedure. Soil biology biochemistry 22: 1167-1169. doi: 10.1016/0038-0717(90)90046-3. Xia F, Mei K, Xu Y, Zhang C, Dahlgren RA, Zhang M (2020) Response of N2O emission to manure application in field trials of agricultural soils across the globe. Science of The Total Environment 733: 139390. doi: https://doi.org/10.1016/j.scitotenv.2020.139390. Xu W, Whitman WB, Gundale MJ, Chien CC, Chiu CY (2021) Functional response of the soil microbial community to biochar applications. GCB Bioenergy 13: 269-281. doi: https://doi.org/10.1111/gcbb.12773. Yang X, Zhang C, Ma X, Liu Q, An J, Xu S, Xie X, Geng J (2021) Combining organic fertilizer with controlled-release urea to reduce nitrogen leaching and promote wheat yields. Frontiers in Plant Science 12: 3087. Yu H, Gao Q, Shao Z, Ying A, Sun Y, Liu J, Mao W, Zhang B (2016) Decreasing nitrogen fertilizer input had little effect on microbial communities in three types of soils. PLoS One 11: e0151622. doi: https://doi.org/10.1371/journal.pone.0151622. Yuan H, Lu T, Wang Y, Chen Y, Lei T (2016) Sewage sludge biochar: Nutrient composition and its effect on the leaching of soil nutrients. Geoderma 267: 17-23. doi: https://doi.org/10.1016/j.geoderma.2015.12.020. Zang X, Liu M, Fan Y, Xu J, Xu X, Li H (2018) The structural and functional contributions of β-glucosidase-producing microbial communities to cellulose degradation in composting. Biotechnology for biofuels 11: 1-13. doi: https://doi.org/10.1186/s13068-018-1045-8. Zhai L, Wang Z, Zhai Y, Zhang L, Zheng M, Yao H, Lv L, Shen H, Zhang J, Yao Y (2022) Partial substitution of chemical fertilizer by organic fertilizer benefits grain yield, water use efficiency, and economic return of summer maize. Soil Tillage Research 217: 105287. Zhang, Li Y, Ren T, Tian Z, Wang G, He X, Tian C (2014a) Short-term effect of tillage and crop rotation on microbial community structure and enzyme activities of a clay loam soil. Biology fertility of soils 50: 1077-1085. Zhang M, Zhang L, Riaz M, Xia H, Jiang C (2021) Biochar amendment improved fruit quality and soil properties and microbial communities at different depths in citrus production. Journal of Cleaner Production 292: 126062. doi: https://doi.org/10.1016/j.jclepro.2021.126062. Zhang Q-z, Dijkstra FA, Liu X-r, Wang Y-d, Huang J, Lu N (2014b) Effects of biochar on soil microbial biomass after four years of consecutive application in the north China plain. PloS one 9: e102062. doi: https://doi.org/10.1371/journal.pone.0102062. Zhao M, Sun B, Wu L, Gao Q, Wang F, Wen C, Wang M, Liang Y, Hale L, Zhou J (2016) Zonal soil type determines soil microbial responses to maize cropping and fertilization. MSystems 1: e00075-00016. doi: https://doi.org/10.1128/mSystems.00075-16. Zhu L-x, Xiao Q, Cheng H-y, Shi B-j, Shen Y-f, Li S-q (2017a) Seasonal dynamics of soil microbial activity after biochar addition in a dryland maize field in North-Western China. Ecological engineering 104: 141-149. doi: https://doi.org/10.1016/j.ecoleng.2017.04.026. Zhu L-x, Xiao Q, Shen Y-f, Li S-q (2017b) Microbial functional diversity responses to 2 years since biochar application in silt-loam soils on the Loess Plateau. Ecotoxicology and Environmental Safety 144: 578-584. doi: https://doi.org/10.1016/j.ecoenv.2017.06.075. Cite Share Download PDF Status: Published Journal Publication published 17 Feb, 2024 Read the published version in Plant and Soil → Version 1 posted Editorial decision: Major revisions 25 Jul, 2023 Reviewers agreed at journal 16 Jun, 2023 Reviewers invited by journal 03 Jun, 2023 Editor invited by journal 14 May, 2023 Editor assigned by journal 14 May, 2023 First submitted to journal 12 May, 2023 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2910777","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":206552066,"identity":"7a7458f0-b510-4da4-90af-c80c46bd55c5","order_by":0,"name":"Muhammad Abdullah Aziz","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABDklEQVRIiWNgGAWjYFACHgYJhgILBgMg8wBDxT85kNiBBwS1GEhAtZw5YAzWkkCsFgbGtgOJDSAGPi267WcP3uYxkJA3Zz/+8OCPM3fS54cdfgi0xU5OtwG7FrMzecnWQC2GO3sSEg5IVDzL3Xg7zQCoJdnY7AAOLQdyzKSBWhg3HEg4cMDgDHPuxtkJIC0HErfh0nL+DViL/YbzDxsOJLYxpxvOTv+AX8sNiC2JG24kMxw42HY4QV46h4AtN94lW84xkEjecOMZw8GGM2mGG6RzCg4kGODxy/ncgzfeVNjYbjif/vjjjwobefnZ6Zs/fKiwk8OlBRMYgFUaEKscBOQbSFE9CkbBKBgFIwEAAO8ka5Bmag/2AAAAAElFTkSuQmCC","orcid":"","institution":"Pir Mehr Ali Shah Arid Agriculture University: PMAS-Arid Agriculture University Rawalpindi","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Muhammad","middleName":"Abdullah","lastName":"Aziz","suffix":""},{"id":206552067,"identity":"65d54b5d-99f7-40bb-aa33-046d9cb78e16","order_by":1,"name":"Khalid Saifullah Khan","email":"","orcid":"","institution":"","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Khalid","middleName":"Saifullah","lastName":"Khan","suffix":""},{"id":206552068,"identity":"e46b5d8d-df4c-49c8-9a0a-32a32d4bde33","order_by":2,"name":"Rabia Khalid","email":"","orcid":"","institution":"","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Rabia","middleName":"","lastName":"Khalid","suffix":""},{"id":206552069,"identity":"0874fe29-386a-488c-9807-35f28455f028","order_by":3,"name":"Muhammad Shabaan","email":"","orcid":"","institution":"","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Muhammad","middleName":"","lastName":"Shabaan","suffix":""},{"id":206552070,"identity":"a5ddddf1-e3ed-40e5-a2cf-4d78f7f2498c","order_by":4,"name":"Abdulaziz G. Alghamdi","email":"","orcid":"","institution":"","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Abdulaziz","middleName":"G.","lastName":"Alghamdi","suffix":""},{"id":206552071,"identity":"2f26bb12-6234-4b05-bb17-8818fd40c372","order_by":5,"name":"Zafer Alasmary","email":"","orcid":"","institution":"","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Zafer","middleName":"","lastName":"Alasmary","suffix":""},{"id":206552072,"identity":"9379f590-116a-45df-87b2-c388e7c5683a","order_by":6,"name":"Mosaed A. Majrashi","email":"","orcid":"","institution":"","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Mosaed","middleName":"A.","lastName":"Majrashi","suffix":""}],"badges":[],"createdAt":"2023-05-09 08:03:09","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2910777/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2910777/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s11104-024-06556-3","type":"published","date":"2024-02-17T15:01:37+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":38102340,"identity":"5fbfbf8b-65fe-4a47-a03f-0e9014d0430a","added_by":"auto","created_at":"2023-06-06 14:38:25","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":216733,"visible":true,"origin":"","legend":"\u003cp\u003eMacronutrient concentrations in soil under each treatment. Values represent the mean with three replications. The small letters on bars indicate significant differences among treatments at 5% probability levels, where significance was tested by using the multiple comparison Tukey’s HSD test. Error bars indicate the standard error of the mean (n = 3)\u003c/p\u003e","description":"","filename":"1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2910777/v1/0f3ca8e56cee405eb52321cc.jpg"},{"id":38102343,"identity":"05e16cde-377c-4e99-8298-4b29807ee442","added_by":"auto","created_at":"2023-06-06 14:38:25","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":199748,"visible":true,"origin":"","legend":"\u003cp\u003eSoil MBC, MBN, MBP and enzymatic activity under each treatment. Values represent the mean with three replications. The small letters on bars indicate significant differences among treatments at 5% probability levels, where significance was tested by using the multiple comparison Tukey’s HSD test. Error bars indicate the standard error of the mean (n = 3).\u003c/p\u003e","description":"","filename":"2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2910777/v1/6e254abff4cd3f03038cc900.jpg"},{"id":38102345,"identity":"ba1b83fd-ae20-475d-805a-60b9722f94d6","added_by":"auto","created_at":"2023-06-06 14:38:25","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":199748,"visible":true,"origin":"","legend":"\u003cp\u003eSoil microbial biomass under each treatment. Values represent the mean with three replications. The small letters on bars indicate significant differences among treatments at 5% probability levels, where significance was tested by using the multiple comparison Tukey’s HSD test. Error bars indicate the standard error of the mean (n = 3).\u003c/p\u003e","description":"","filename":"3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2910777/v1/7543030eb0c3995d66ccbc7b.jpg"},{"id":38102342,"identity":"58a57974-1fe7-4b41-8f2d-175fae6052cb","added_by":"auto","created_at":"2023-06-06 14:38:25","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":53541,"visible":true,"origin":"","legend":"\u003cp\u003ePrincipal Component Analysis (PCA) biplots of the soil microbial biomass C, N, P and nutrients influenced by the treatments.\u003c/p\u003e\n\u003cp\u003eNO\u003csub\u003e3\u003c/sub\u003e-N = Nitrate Nitrogen; NH\u003csub\u003e4\u003c/sub\u003e-N = Ammonium Nitrogen; Olsen P = Olsen Phosphorus; K = Potassium; MBC = Microbial biomass carbon; MBN = Microbial biomass nitrogen; MBP = Microbial biomass phosphorus; EC = Electrical conductivity; SOC = Soil organic carbon; DOC = Dissolved organic carbon.\u003c/p\u003e","description":"","filename":"4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2910777/v1/9c905b5f81acffab780199c7.jpg"},{"id":38104097,"identity":"a0dd4a58-9ea4-4f0d-b1c1-df29e0e7c64a","added_by":"auto","created_at":"2023-06-06 14:46:25","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":56495,"visible":true,"origin":"","legend":"\u003cp\u003ePrincipal Component Analysis (PCA) biplots of the soil microbial biomass and enzymatic activity influenced by the treatments.\u003c/p\u003e\n\u003cp\u003ePHO = Phosphatase; β-GLU = β-Glucosidase; URE = Urease; PRO = Protease; DEH = Dehydrogenase; F/B = Fungal bacterial ratio; PLFA = Phospholipid fatty acid; Gram+ = Gram positive bacteria; Gram- = Gram negative bacteria; SF = Saprophytic Fungi; AMF = Arbuscular mycorrhizal fungi.\u003c/p\u003e","description":"","filename":"5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-2910777/v1/277ad72736442f94d744048b.jpg"},{"id":51323169,"identity":"43f9461a-eadc-44ac-9e7e-092802f4284a","added_by":"auto","created_at":"2024-02-19 15:15:54","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1067031,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2910777/v1/790f5142-c14c-438e-90e4-ff600cea7d25.pdf"}],"financialInterests":"","formattedTitle":"Integrated application of biochar and chemical fertilizers improves growth and yield of wheat (Triticum aestivum) by altering soil biological heath","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003eApplication of chemical fertilizer is a general practice adopted globally for ensuring increased crop yields and fulfilling the demands of human population (Liu et al. \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). However, their continuous and repeated applications not only adversely affect soil quality but also lead to various environmental constraints such as soil acidification and groundwater pollution (Li et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; van der Bom et al. \u003cspan citationid=\"CR97\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Furthermore, their excessive applications lead to diminishing returns and other associated economic risks such as increased costs of production followed by lower profit margins (Amjadian et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2021b\u003c/span\u003e; Humbert et al. \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Substitution of chemical fertilizers with different organic amendments (OA) has proved to be a significant approach to improve crop yields, while sustaining soil health and environmental quality (He et al. \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Organic amendments are vital aspects of organic farming practices, that exert promising impacts on soil health and indigenous microbial communities, which in turn, perform nutrient cycling in soil and improve its fertility status (Chaudhari et al. \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). They are rich source of essential plant nutrients and therefore, can reduce the farmer\u0026rsquo;s dependence on chemical fertilizers (Singh \u003cspan citationid=\"CR87\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). However, under tropical conditions where temperature is high and rainfall is lower, organic amendments get decomposed very quickly (Doan et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), which also exhibit their significant contribution towards global warming owing to abrupt mineralization (Xia et al. \u003cspan citationid=\"CR104\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Application of OA can also ameliorate soil degradation and help in rebuilding a healthy and nutritious soil profile (Yang et al. \u003cspan citationid=\"CR106\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Numerous researchers have reported regarding the beneficial role of OA in improving soil organic carbon, soil structure as well as fertility (Chen et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). For instance, application of livestock manure along with chemical fertilizers has reportedly resulted in enhanced soil C sequestration, soil fertility status and ultimately, the plant productivity (Deng et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Palmer et al., (\u003cspan citationid=\"CR73\" class=\"CitationRef\"\u003e2017\u003c/span\u003e) reported that decomposition of soil organic matter (SOM) releases nitrogen (N), which can enhance crop yields under N limited conditions, as OA ensure slow and sustained release of essential nutrients to soil (Palmer et al. \u003cspan citationid=\"CR73\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), and thereby, their prolonged availability for plant uptake is enabled (Yang et al. \u003cspan citationid=\"CR106\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In another study, Zhai et al. (\u003cspan citationid=\"CR110\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) suggested that partial substitution of chemical fertilizers by organic fertilizers can improve the growth and physiological attributes of maize and can also improve soil quality.\u003c/p\u003e \u003cp\u003eAmong various amendments, biochar (BC) application as an organic amendment, has gained much attention, as it encompasses several positive implications on soil fertility, nutrient cycling, moisture retention, structure and carbon sequestration (Alkharabsheh et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Elkhlifi et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Semida et al. \u003cspan citationid=\"CR83\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). It is prepared by pyrolysis (burning of biomasses under oxygen deficient conditions) of different plant materials (Aziz et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e) such as crop residue, food wastes, farmyard manure, and has recalcitrant properties, and can persist in soil for hundreds to thousands of years (Brassard et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Budai et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). BC is comparatively more resistant to microbial degradation than other organic materials, and contains roughly 60% of stable organic carbon (Ameloot et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2013b\u003c/span\u003e). Carbon atoms in BC molecules are firmly linked to each other, and therefore, restrict microorganisms to get access to carbon making it relatively more resistant to breakdown than other organic amendment (Lopez-Capel et al. \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Research on biochar has got significant attention in recent times due to its potential positive impacts towards environment sustainability, carbon sequestration, greenhouse gas emission and land degradation. Peng et al. \u003cspan citationid=\"CR74\" class=\"CitationRef\"\u003e2021\u003c/span\u003e stated that application of biochar in combination with N fertilizer positively affected wheat yield and soil nutrients by restraining soil nitrate leaching. Similarly, Sadaf et al. \u003cspan citationid=\"CR80\" class=\"CitationRef\"\u003e2017\u003c/span\u003e reported that co-application of biochar and chemical fertilizers led to significant increment in soil nutrient content and their uptake by wheat as compared to alone application of chemical fertilizer or biochar. Biochar also possesses the ability to reduce post application nutrient loss from soil, and thereby, can increase nutrient use efficiency (NUE) due to more uptake by plants. For instance, Win et al. (\u003cspan citationid=\"CR102\" class=\"CitationRef\"\u003e2019\u003c/span\u003e) reported that application of biochar with or without biofertilizers can improve the NUE particularly N, leading to its improved plant uptake leading to increased plant growth. Hence, incorporation of BC along with CF could be a suitable option to improve plant growth and enhance NUE by plant as availability and uptake of nutrients applied through CF may be increased under their combined application, and chances of nutrient leaching can also be minimized (Sadaf et al. \u003cspan citationid=\"CR80\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eNutrient cycling in soil is affected by soil microbial processes, which reflect soil health under changing soil conditions. Among them, most important soil biological health indicators are microbial biomass carbon (MBC) and soil enzymes such as phosphatase, dehydrogenase and β-glycosidase (Saha et al. \u003cspan citationid=\"CR81\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Biochar application has also been proved to be positively correlated with soil biological activities. It has been observed that improvement in wheat quality brought about by high-carbon fertilizers were linked to an increase in different bacterial genera such as \u003cem\u003eSphingomonas\u003c/em\u003e and \u003cem\u003eHaliangium\u003c/em\u003e, which supported plant growth and development (Chen et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Soil microbial activities in terms of soil enzymes exert a significant impact on various soil processes such as OM decomposition, mineralization, and nutrient cycling (Meetei et al. \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Sharma \u003cspan citationid=\"CR86\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Charged surfaces present on biochar adsorb various enzyme substrates, increase enzyme reactions, and ultimately leads to better soil enzyme activity (Gasco et al. \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Alkaline phosphatase, which predominates in alkaline soils, plays a crucial role in the cycling of P by hydrolyzing ester phosphate bonds (Shabaan et al. \u003cspan citationid=\"CR84\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Biochar also has positive priming effects on soil dehydrogenase activity (Azeem et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). β-glycosidase activity is a foremost indicator to observe the changes occur in soil (Stege et al. \u003cspan citationid=\"CR94\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). This enzyme is involved in hydrolysis of maltose and cellobiose, and acts as a catalyst (Matsuzawa and Yaoi \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Further, β-glycosidase also acts as an energy sources for the microorganisms during cellobiose degradation (Zang et al. \u003cspan citationid=\"CR109\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Studies related to the impact of biochar on soil microorganisms are still poorly understood because of variation in feedstock type, pyrolysis temperature and abundance of carbon content in the biochar. Moreover, earlier studies comprising BC mainly focused on its positive role in improving soil fertility and crop growth and scanty data exists in literature regarding its beneficial role in soil biological health.\u003c/p\u003e \u003cp\u003eWheat is major agricultural crop of Pakistan, and acts as a staple food for almost every household, making it the most strategic commodity of the country, whose availability and affordability is the government\u0026rsquo;s main priority (Rasheed et al. \u003cspan citationid=\"CR76\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Pakistan is the 8th largest wheat producer worldwide (Ahmad et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), where it is grown on an area of 8.9\u0026nbsp;million hectares (mha) with an annual production of 2.9 tons per ha (Pakistan \u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). It contributes 1.8% in the country\u0026rsquo;s gross domestic product (GDP), and 7.8% in the value addition in agriculture. Moreover, wheat performs an important role in providing various macromolecules such as carbohydrates, proteins and other essential minerals (Amjadian et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2021a\u003c/span\u003e). One of the major reasons behind poor and sub-optimal yield of wheat in developing nations such as Pakistan resides with poor plant nutrition (Iqbal et al. \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Iqbal et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Wang et al. (\u003cspan citationid=\"CR99\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) checked the impact of integrated applications of chemical and organic fertilizers on community and diversity of prokaryotic microorganisms as well as on nutrient absorption by wheat and resultant increment in its growth. They observed that soil supplementation with OA enhanced the SOM contents, diversity of rhizosphere and endophytic microbes, and at the same time, augmented the relative abundance and activities of different microbes in endo- as well as rhizosphere. Thus, it is of great importance to examine the effects of biochar on microbial activities due to their sensitivity to environmental changes. We hypothesized that different doses of biochar with or without NPK fertilizer might stimulate the activity of soil bacteria with subsequent increased soil nutrient content and crop yield. Therefore, objective of this research was to assess the role of individual and combined application of BC and CF in altering the soil nutrient profile, bacterial activity and wheat growth in alkaline soil conditions.\u003c/p\u003e"},{"header":"2 Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Experimental site description\u003c/h2\u003e \u003cp\u003eField experiment was performed at Research farm of Pir Mehr Ali Shah Arid Agriculture University, Koont Chakwal (latitude: 33\u0026deg; 38' 49.56\" and longitude: 73\u0026deg; 4' 49.08\") which is known as Pothwar plateau of northern Punjab. The area has a semi-arid subtropical climate with a pattern of bimodal rainfall occurring in late summer and winter seasons. Usually, around 60\u0026ndash;70% of annual rainfall is received during the monsoon season, which runs from June 15th to September 15th. The remaining rainfall occurs during the winter months in the form of gentle showers. Experimental site is classified as Rawal series, Udic Haplustoll Alfisols. Generally, the soil was sandy clay loam (56% sand, 22.8% silt, 21.2% clay). Basic soil properties are given in Table \u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eBasic characteristics of experimental site soil and biochar\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eProperties\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnit\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSoil\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eBiochar\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003epH\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8.93\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eEC\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003edS m\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.32\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.73\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eAsh\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e---\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e12.51\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTotal organic C\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e---\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e42.58\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eNO\u003c/b\u003e\u003csub\u003e\u003cb\u003e3\u003c/b\u003e\u003c/sub\u003e\u003cb\u003e-N\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.27\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e6.83\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eNH\u003c/b\u003e\u003csub\u003e\u003cb\u003e4\u003c/b\u003e\u003c/sub\u003e\u003cb\u003e-N\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.37\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e7.37\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eOlsen P\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4.04\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eExtractable K\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e62.59\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e92.60\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eSpecific surface area\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003em\u003csup\u003e2\u003c/sup\u003e g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e---\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2.88\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eMoisture content\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9.41\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4.51\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eCEC\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ecmol kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.69\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e16.29\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eBulk density\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egcm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.46\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e---\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Production of Biochar (BC)\u003c/h2\u003e \u003cp\u003eBiochar was prepared by using poultry litter as a feedstock material, which was collected from Poultry Research Institute Rawalpindi, Pakistan. Production took place at 400℃ for 3 h with 10℃ min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e heating rate in a muffle furnace. After that, BC was crushed and sieved from 2 mm sieve in order to get homogenized material prior to field application, and was applied on weight basis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Experiment layout and treatment application\u003c/h2\u003e \u003cp\u003eBefore seedbed preparation, field was ploughed twice and harrowed, and weeds and stubbles of previous crop were manually removed. Experimental plots were arranged by following randomized complete block design (RCBD), and each plot carried equal size (4m \u0026times; 6m). All four treatments including control were applied in triplicates. Treatments of current study were control (C), BC @ 10 t ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (BC10), BC @ 20 t ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (BC20), BC 10 t ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e + CF (BC10\u0026thinsp;+\u0026thinsp;CF) and BC 20 t ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e + CF (BC20\u0026thinsp;+\u0026thinsp;CF). Furthermore, recommended doses of NPK (120:60:50 kg ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) were applied as urea, DAP and sulfate of potash (SOP).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Soil Sampling\u003c/h2\u003e \u003cp\u003eSoil samples were collected from replicated plots in May 2022 using a soil auger followed by their pulverization and filtration through 2mm mesh. To evaluate microbial and enzymatic activities, soil samples were kept at 4℃, while for other nutrient analysis and physicochemical analyses, samples were air-dried at room temperature.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.5 Analysis of soil and biochar physiochemical properties\u003c/h2\u003e \u003cp\u003eThe pH of soil samples was measured by glass electrode method in H\u003csub\u003e2\u003c/sub\u003eO with 1:2.5 ratio of soil and water (Thomas \u003cspan citationid=\"CR96\" class=\"CitationRef\"\u003e1996\u003c/span\u003e), and electrical conductivity (EC) was determined by using an EC meter (Page AL 1982). Soil nitrate nitrogen (NO\u003csub\u003e3\u003c/sub\u003e-N) determination was done by extraction method, where soil samples were extracted with 0.5 M K\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, after extracted the soil samples, the NO\u003csub\u003e3\u003c/sub\u003e-N was measured by using a spectrophotometer at the wavelength of 410 nm (Anderson and Ingram 1993a), for NH\u003csub\u003e4\u003c/sub\u003e-N soil samples were extracted by 2M KCl solution and measured using a spectrophotometer (Shimazadu UV-1280) (Keeney et al. \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e1983\u003c/span\u003e). Method of Olsen comprising NaHCO\u003csub\u003e3\u003c/sub\u003e (0.5 M) was used to determine P content from soil samples spectrophotometrically at spectrophotometer (Shimazadu UV-1280) (Olsen 1982). Soil K was determined by extraction of soil samples with 1 N ammonium acetate solution using the flame photometer (Jenway PFP7 flame photometer) (Helmke and Sparks \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e1996\u003c/span\u003e). For measuring soil organic carbon (SOC), K\u003csub\u003e2\u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e titration method was followed (Nelson et al. 1983). Soil dissolved organic carbon (DOC) was determined by extraction of soil samples with 0.5M K\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e (Song et al. \u003cspan citationid=\"CR90\" class=\"CitationRef\"\u003e2018b\u003c/span\u003e). Ash content of biochar was calculated by the following question\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$Ash \\left(\\%\\right)= \\frac{D}{B}\\times 100$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eSimilarly, organic C content of biochar was calculated by using the following equation\u003cdiv id=\"Equb\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equb\" name=\"EquationSource\"\u003e\n$$Organic Carbon \\left(\\%\\right)= \\frac{100-Ash}{1.724}$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eCation exchange capacity (CEC) of BC was measured by mixing 1 N sodium acetate trihydrate (33 mL) with air-dried BC samples, and solution was shaken for five minutes (5000 rpm). After decanting, supernatant was discharged four times and 33 mL ethanol was added into the solution before being centrifuged. Then ammonium acetate solution was used for removing sodium and measured in flame photometer (Chapman \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1965\u003c/span\u003e). The CEC was calculated using the following equation:\u003cdiv id=\"Equc\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equc\" name=\"EquationSource\"\u003e\n$$CEC \\left(meq 100{g}^{-1}\\right)=\\frac{meq}{1}Na \\left(from calibration curve\\right)\\times \\frac{A}{wt}\\times \\frac{100}{1000}$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.6 Microbial biomass carbon (MBC)\u003c/h2\u003e \u003cp\u003eSoil MBC was estimated using fumigation-extraction method. Briefly, 30 mL of alcohol-free chloroform was placed in a beaker and was used as a fumigant, which was placed in a desiccator along with additional beakers filled with soil (10g). After the fumigation period, soil samples were extracted using 50 mL of 0.5 M K\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and extract was filtered using Whatman 40. Same procedure was repeated for control. To measure the extracted samples, 4 mL of extracts were taken and mixed with 1 mL of potassium dichromate (0.0667 M). Mixture was then heated to 150\u0026deg;C for 30 minutes, and 5 mL of concentrated H\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e was added. After adding 3\u0026ndash;4 drops of phenanthroline monohydrate indicator, contents were titrated against ferrous ammonium sulfate solution. Following equation was used to calculate C\u003csub\u003emic\u003c/sub\u003e (Brookes et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e1985\u003c/span\u003e; Wu et al. \u003cspan citationid=\"CR103\" class=\"CitationRef\"\u003e1990\u003c/span\u003e).\u003cdiv id=\"Equd\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equd\" name=\"EquationSource\"\u003e\n$${C}_{mic}=\\left(Extratcted {C}_{fumigated}-Extracted {C}_{unfumigated}\\right)\\times 2.64$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e2.7 Microbial biomass nitrogen (MBN)\u003c/h2\u003e \u003cp\u003eTo estimate MBN, gasification-extraction procedure was performed where extractant was digested in mixture of 4.4 mL of 30% hydrogen peroxide (H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e; 350 mL) and concentrated H\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e (20 mL) at 360\u0026deg;C for 2 hours. After digestion, solution was placed in a steam distillation chamber where 25 mL alkaline mixture of NaOH and sodium thiosulfate (500 g and 25 g/L, respectively) were added. Further, boric acid (5 mL) was added to receiving end prior to distillation. Finally, samples were titrated to gray endpoint with hydrochloric acid (HCl; 0.1 N). For precision, amount of HCl was indexed (Brookes et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e1985\u003c/span\u003e; Wu et al. \u003cspan citationid=\"CR103\" class=\"CitationRef\"\u003e1990\u003c/span\u003e).\u003cdiv id=\"Eque\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Eque\" name=\"EquationSource\"\u003e\n$$TN \\left(\\%\\right)=T \\times 0.1 \\times 0.001*) \\times (S/A)/W \\times 100/1$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eWhere; * refers to the conversion factor (mg to g), T\u0026thinsp;=\u0026thinsp;Corrected titer (mL), S\u0026thinsp;=\u0026thinsp;final digestion solution volume (mL), A\u0026thinsp;=\u0026thinsp;aliquot (sample) volume (mL), and W\u0026thinsp;=\u0026thinsp;sample weight (g).\u003cdiv id=\"Equf\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equf\" name=\"EquationSource\"\u003e\n$${N}_{mic}=Extracted {N}_{fumigated}- {Extracted N}_{unfumigated} \\times 1.46$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e2.8 Microbial biomass phosphorus (MBP)\u003c/h2\u003e \u003cp\u003eTo estimate MBP, 2.5 g soil was oven-dried in three repetitions. One replica was incubated for 24 hours at 25\u0026deg;C in alcohol-free chloroform, while other two were aerobically incubated under same conditions. Samples were then shaken continuously for 30 minutes before being extracted with 0.5 M NaHCO\u003csub\u003e3\u003c/sub\u003e. Clear extract of 10 mL was then transferred to a 50 mL volumetric flask, to which 8 mL of reagent (ascorbic acid solution and ammonium molybdate solution) was added.\u003c/p\u003e \u003cp\u003eAfter color development, samples were run on a spectrophotometer (Shimazadu UV-1280) at 882 nm. The KH\u003csub\u003e2\u003c/sub\u003ePO\u003csub\u003e4\u003c/sub\u003e was used for making five different levels of concentrations (0, 0.5, 1, 2, and 4 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) as for P standards. Following equation was used for analyzing MBP (Brookes et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e1982\u003c/span\u003e; Joergensen et al. \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e1995\u003c/span\u003e).\u003cdiv id=\"Equg\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equg\" name=\"EquationSource\"\u003e\n$${P}_{mic}= {(P}_{fumigated}- {P}_{unfumigated}) \\times 2.5$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e2.9 Soil enzymatic activities\u003c/h2\u003e \u003cp\u003eUrease activity (UA) analysis was done by using potassium chloride solution (50 mL). After filtration of soil extract, ammonium content was calculated at 690 nm optical density, and urease activity was determined (Kandeler and Gerber \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e1988\u003c/span\u003e). For the estimation of soil dehydrogenase activity (SDA), 5 g moist soil and 5 mL TTC solution were mixed in a test tube, and sealed in tube followed by its incubation for 24 h at 30\u0026deg;C. Later, 40 mL of acetone was added to extract, and incubated for 2 hours under darkness. Finally, soil solution was filtered, and optical density (OD) was measured at 546 nm. Triphenyl formazan (TPF) was developed at 0\u0026ndash;40 g TPF mL\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e for a calibration curve in Tris buffer (pH -7.6) and acetone (Alef \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e1995\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eSimilarly, to estimate soil phosphatase activity (SPA), 10 g of moist soil was mixed with 1.5 mL of toluene and transferred to a 100 mL volumetric flask. Then, 10 mL of disodium phenyl phosphate and 20 mL of buffer were added in it, and incubated at 37\u0026deg;C for three hours. After incubation, extract was made up to 100 mL by adding distilled water and filtered. Then, 8 mL of filtrate was mixed with 5 mL of buffer and diluted up to 25 mL volume. 2,6 dibromo quinone chloramide was added, and final volume was made up to 100 mL. Mixture was then incubated at room temperature for about 30 minutes before reading at 600 nm. For standard calibration, 0-200 \u0026micro;g phenol was used, and phosphatase activity was expressed as phenol (\u0026micro;g g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e dwt 2 h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) (Alef \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e1995\u003c/span\u003e). Besides these, protease activity was measured by keeping samples for incubation at 50℃ with sodium caseinate by determining the absorbance at 700 nm of tyrosine content using Folin\u0026ndash;Cicalteau reagent (Ladd and Butler \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e1972\u003c/span\u003e). Soil β-glucosidase enzyme activity was measured by colorimetric estimation of the p-nitrophenol method (Tabatabai \u003cspan citationid=\"CR95\" class=\"CitationRef\"\u003e1982\u003c/span\u003e). In conditioning cabinet, 1 g of soil was incubated with a buffered solution of \u003cem\u003ep\u003c/em\u003e-nitrophenyl-D-glucopyranoside (pH 6.0) and toluene at 37\u0026deg;C for one hour. After incubation, soil suspension was mixed with 1 mL of 0.M CaCl\u003csub\u003e2\u003c/sub\u003e and 4 mL of tris (hydroxymethyl) aminomethane buffer. Yellow filtrate obtained was used to estimate the quantity of \u003cem\u003ep\u003c/em\u003e-nitrophenol produced by the hydrolysis of \u003cem\u003ep\u003c/em\u003e-nitrophenyl-D-glucopyranoside at 37\u0026deg;C for 1 hour through calorimetric determination.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e2.10 Plant Analysis\u003c/h2\u003e \u003cp\u003eFollowing the crop harvest, plant growth attributes i.e., root-shoot length, plants fresh-dry weight, and 100 grain weight were measured. An electrical balance was used to calculate the fresh weight of the roots and shoots. To calculate plants dry weight, their samples were oven dried at 65\u0026deg;C for 72 hours. These dried samples were subsequently pulverized, and kept in plastic bags for further chemical analysis. Plant samples were subjected to digestion in a block digester for 2 hours at 360℃. A digestion mixture consisting of Se powder, Li\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and H\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e was added to plant in 100 mL digestion tube. For plant samples, contents of N and P were determined using colorimetric method (Anderson and Ingram \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1993b\u003c/span\u003e). Before finding the plant nitrogen on Kjeldahl Nitrogen Analyzer (BKN- 983), the digested samples were mixed with 55 mL of NaOH and 50 mL of DI water (Kjeldahl \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e1883\u003c/span\u003e) then run on Kjeldahl apparatus. The P was quantified by mixing 5 mL of digest and 5mL of color reagent (ammonium heptamolybdate-ammonium vanadate) in nitric acid. Later, samples were run on spectrophotometer at 410 nm wavelength to check their absorbance (Shapter \u003cspan citationid=\"CR85\" class=\"CitationRef\"\u003e1940\u003c/span\u003e). Total K contents of plants were determined by wet digestion of plant samples using a mixture of nitric and perchloric acid in a block digester. Then, samples were digested at 230\u0026deg;C for 3 hours and K concentration was determined using a flame photometer (Ryan et al. \u003cspan citationid=\"CR79\" class=\"CitationRef\"\u003e2001\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e2.11 Soil microbial abundance\u003c/h2\u003e \u003cp\u003eSoil phospholipid fatty acid analysis (PLFA) extraction method used in this study was explained by (Zhang et al. 2014a). Nonadecanoic acid methyl ester (19:0, Sigma) was employed as internal standard. Total microbial biomass was determined by the presence of 33 different PLFAs ranging from C14 to C20 in the samples. Gram-positive bacteria (G+) were indicated by the total of i14:0, a15:0, i15:0, a16:0, a17:0, and i17:0 (Yu et al. \u003cspan citationid=\"CR107\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), while gram-negative bacteria (G-) were represented by sum of 16:12OH, 16:1ω7c, 16:1ω9c, cy17:0, 17:1ω8c, 18:1ω7c, and cy19 (Yu et al. \u003cspan citationid=\"CR107\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). The sum of 18:2ω6,9c and 18:1w9c was used to indicate saprotrophic fungi (SF) (Zhang et al. 2014a), and 16:1w5c was used to indicate arbuscular mycorrhizal fungi (AMF) (Liu et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). The sum of 10Me16:0, 10Me17:0, and 10Me18:0 was employed to indicate actinomycetes, while 20:4ω6,9,12,15c was used to indicate protozoa (Zhang et al. 2014a).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e2.12 Statistical Analysis\u003c/h2\u003e \u003cp\u003eEffects of all treatments (biochar and their interaction co-application with chemical fertilizers) on different physicochemical properties, microbial biomasses, and enzymatic activities in soil were studied by two-way analysis of variance (ANOVA) by using statistical software Statistix 8.1. Treatment means were compared for significant differences by using Tukey\u0026rsquo;s Honestly Significant Difference (HSD) test at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 level (Steel et al. \u003cspan citationid=\"CR93\" class=\"CitationRef\"\u003e1997\u003c/span\u003e). Principal component test (PCA) was carried out to determine the correlation of the environmental variables with all soil microbial, enzymatic and nutrients parameters.\u003c/p\u003e \u003c/div\u003e"},{"header":"3 Results","content":"\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Soil pH and EC\u003c/h2\u003e \u003cp\u003eBiochar based treatments had variable effects on soil pH and EC. However, no significant impact on soil pH was pragmatic, and only in combined treatment (BC20\u0026thinsp;+\u0026thinsp;CF), a slight improvement in pH was experienced as compared to sole BC treatments whereas, as compared to control treatment, maximum increment in soil pH was 14% (BC20\u0026thinsp;+\u0026thinsp;CF). Besides, soil EC showed significant variations according to treatments application, and was significantly (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) increased by 96% under combined application (BC20\u0026thinsp;+\u0026thinsp;CF), followed by sole application of BC @ 20 t ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (67%), BC10\u0026thinsp;+\u0026thinsp;CF (64%) and BC @ 10t ha\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e by (45%) (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e3.2 Soil nutrients content\u003c/h2\u003e \u003cp\u003eAddition of BC significantly improved soil nutrients status. Among them, soil NO\u003csub\u003e3\u003c/sub\u003e-N was significantly increased by 76% under BC20\u0026thinsp;+\u0026thinsp;CF treatment however, in sole BC20 treatment this increment was approximately 23% higher than unamended soil. Whilst results of BC10\u0026thinsp;+\u0026thinsp;CF and BC10 were marginally lower than other treatments except control (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Similarly, soil NH\u003csub\u003e4\u003c/sub\u003e-N was also increased under combined application of BC and chemical fertilizers i.e., BC10\u0026thinsp;+\u0026thinsp;CF and BC20\u0026thinsp;+\u0026thinsp;CF by 105 and 126% respectively as compared to control. Moreover, there was no statistically significant difference between BC20 and BC10\u0026thinsp;+\u0026thinsp;CF treatments. Addition of BC led to greater soil P content, where maximum P contents were observed in combined application of BC and CF i.e., BC20\u0026thinsp;+\u0026thinsp;CF and BC10\u0026thinsp;+\u0026thinsp;CF exhibiting respective P contents (162 and 141%) as compared to unamended treatment. Sole application of BC10 and BC20 showed 42 and 17% lower P results as compared to their co-application with chemical fertilizers respectively. Soil K, SOC and DOC content had maximum increment of 44, 33 and 130% respectively, under BC20\u0026thinsp;+\u0026thinsp;CF treatment while sole application of BC20 increased these nutrients status by 25, 28 and 89% respectively. Furthermore, results revealed that BC20\u0026thinsp;+\u0026thinsp;CF enhanced the soil C/N ratio by 55% whereas BC10\u0026thinsp;+\u0026thinsp;CF improved it by only 32% (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). PCA analysis of soil nutrients with treatments also proved that integrated application of BC with CF significantly increased soil nutrient status (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Microbial biomass and enzyme activity\u003c/h2\u003e \u003cp\u003eMaximum MBC (484 \u0026micro;g/g) was obtained in BC20\u0026thinsp;+\u0026thinsp;CF treatment which was 27% higher than unamended control. Similarly, in sole BC20 treatment, this increment was only 21% followed by BC10\u0026thinsp;+\u0026thinsp;CF and BC10 (17 and 14%) respectively. Likewise, maximum values of MBN and MBP were also obvious in BC20\u0026thinsp;+\u0026thinsp;CF treated plots, where they were increased by 62 and 71% respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e2\u003c/span\u003e). However, in sole BC treatment, this increment was marginally lower than combined application (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e4\u003c/span\u003e). Furthermore, combined application of BC and CF was the most efficient in improving soil enzyme activities i.e., urease activity was maximum under BC20\u0026thinsp;+\u0026thinsp;CF treatment (24%) followed by BC20 (16%) than control. Moreover, BC10\u0026thinsp;+\u0026thinsp;CF and BC10 enhanced UA by 14 and 12% respectively as compared to control. Similarly, soil dehydrogenase activity was also maximum under BC20\u0026thinsp;+\u0026thinsp;CF (44%) treatment than control followed by BC10\u0026thinsp;+\u0026thinsp;CF (31%), while BC20 and BC10 increased dehydrogenase activities by 26 and 12% respectively. In addition, phosphatase, protease and β-glucosidase activities were increased by 29, 107 and 93% respectively under BC20\u0026thinsp;+\u0026thinsp;CF followed by BC20 sole application i.e., 23, 69 and 86% respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003e3.4 Soil microbial community\u003c/h2\u003e \u003cp\u003eCombine application of BC with CF significantly improved soil microbial biomass for instance PLFA had marginally better results under BC10\u0026thinsp;+\u0026thinsp;CF (17%) and BC20\u0026thinsp;+\u0026thinsp;CF (24%) as compare to control. Similarly, AMF and actinomycetes also has the highest value under BC20\u0026thinsp;+\u0026thinsp;CF application i.e 49% and 24% respectively. F/B also increased under co-application BC10\u0026thinsp;+\u0026thinsp;CF and BC20\u0026thinsp;+\u0026thinsp;CF by 20% and 24% respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e3\u003c/span\u003e). PCA was conducted with 33 PLFA present in soil subjected to all five treatments and described the effect of each treatment on soil microbial community. Both PC1 and PC2 successfully explained about 93.75% and 4.53% of the total sample variation respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). Along with the PC1, it was observed that BC20\u0026thinsp;+\u0026thinsp;CF significantly increased soil actinomycetes, SPF, gram+, gram-, total PLFA and F/B while BC10\u0026thinsp;+\u0026thinsp;CF enhanced the soil fungi and AMF as compared to control. While BC20 increased only soil bacterial content i.e G\u0026thinsp;+\u0026thinsp;and G- in PC2 as compared to control. Overall, C had negative correlation with both PC1 and PC2. Generally, soil microbial abundance was maximum under BC20\u0026thinsp;+\u0026thinsp;CF followed by BC10\u0026thinsp;+\u0026thinsp;CF and BC20 however, BC10 did not produce any significant impact on it.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003e3.5 Plant growth and yield attributes\u003c/h2\u003e \u003cp\u003e Plant agronomic parameters significantly varied according to biochar application rate and method. Overall, combined application of BC with CF showed maximum improvements in all plant growth attributes i.e., plant shoot and root length were 22 and 37% higher than control under BC20\u0026thinsp;+\u0026thinsp;CF respectively followed by BC10\u0026thinsp;+\u0026thinsp;CF (15 and 25% respectively). Spike length, leaf area index and number of tillers\u0026rsquo; maximum increment was also under BC20\u0026thinsp;+\u0026thinsp;CF treatment i.e., 55, 30 and 12% higher than control respectively. BC10\u0026thinsp;+\u0026thinsp;CF enhanced these parameters by 35, 21 and 8% respectively while, BC20 sole improved them only 26, 21 and 10% respectively. Similarly, plant total N, P, K was also maximum under BC20\u0026thinsp;+\u0026thinsp;CF amendments i.e 10%, 58% and 80% higher than sole BC20 application (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePlant physical and yield attributes under the effect of each treatment. Values denote the mean with 3 replications. Small letters with values indicate significant differences among treatments at the 5% probability level, where significance was tested by using the multiple comparison Tukey\u0026rsquo;s HSD test. (\u0026plusmn;) Standard error of the mean (n\u0026thinsp;=\u0026thinsp;3).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colspan=\"5\" nameend=\"c6\" namest=\"c2\"\u003e \u003cp\u003eTreatments\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003eC\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003eBC10\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003eBC20\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003eBC10\u0026thinsp;+\u0026thinsp;CF\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003eBC20\u0026thinsp;+\u0026thinsp;CF\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eShoot Length (cm)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e281.67\u0026thinsp;\u0026plusmn;\u0026thinsp;14.73c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e298.82\u0026thinsp;\u0026plusmn;\u0026thinsp;11.27c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e304.72\u0026thinsp;\u0026plusmn;\u0026thinsp;8.18bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e325.61\u0026thinsp;\u0026plusmn;\u0026thinsp;13.12ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e342.31\u0026thinsp;\u0026plusmn;\u0026thinsp;9.54a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eRoot length (cm)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.43\u0026thinsp;\u0026plusmn;\u0026thinsp;2.42b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10.05\u0026thinsp;\u0026plusmn;\u0026thinsp;1.43ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10.75\u0026thinsp;\u0026plusmn;\u0026thinsp;1.56ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e10.47\u0026thinsp;\u0026plusmn;\u0026thinsp;0.73ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e11.43\u0026thinsp;\u0026plusmn;\u0026thinsp;0.88a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eSpike length (cm)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9.03\u0026thinsp;\u0026plusmn;\u0026thinsp;0.27d\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10.11\u0026thinsp;\u0026plusmn;\u0026thinsp;0.46cd\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e11.41\u0026thinsp;\u0026plusmn;\u0026thinsp;1.61bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e12.21\u0026thinsp;\u0026plusmn;\u0026thinsp;0.86ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e14\u0026thinsp;\u0026plusmn;\u0026thinsp;0.19a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eLeaf area index\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.91\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.42b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e3.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4.55\u0026thinsp;\u0026plusmn;\u0026thinsp;0.17ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e3.81\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTillers (m\u003c/b\u003e\u003csup\u003e\u003cb\u003e2\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e378.21\u0026thinsp;\u0026plusmn;\u0026thinsp;16.11c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e392\u0026thinsp;\u0026plusmn;\u0026thinsp;12.39bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e416.67\u0026thinsp;\u0026plusmn;\u0026thinsp;17.21ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e410.7\u0026thinsp;\u0026plusmn;\u0026thinsp;14.38ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e426.61\u0026thinsp;\u0026plusmn;\u0026thinsp;15.28a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e1000 seed weight (g)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e35.03\u0026thinsp;\u0026plusmn;\u0026thinsp;1.81c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e36.15\u0026thinsp;\u0026plusmn;\u0026thinsp;1.42c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e38.02\u0026thinsp;\u0026plusmn;\u0026thinsp;0.31b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e38.62\u0026thinsp;\u0026plusmn;\u0026thinsp;0.91b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e40.95\u0026thinsp;\u0026plusmn;\u0026thinsp;1.11a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eBio yield (t ha\u003c/b\u003e\u003csup\u003e\u003cb\u003e\u0026minus;\u0026thinsp;1\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.51d\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9.03\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10cd\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e9.93\u0026thinsp;\u0026plusmn;\u0026thinsp;0.17bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e9.72\u0026thinsp;\u0026plusmn;\u0026thinsp;0.96b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e11.13\u0026thinsp;\u0026plusmn;\u0026thinsp;0.29a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eGrain yield (t ha\u003c/b\u003e\u003csup\u003e\u003cb\u003e\u0026minus;\u0026thinsp;1\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3.14\u0026thinsp;\u0026plusmn;\u0026thinsp;0.17d\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.36\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e3.93\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e3.96\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTotal N (mg g\u003c/b\u003e\u003csup\u003e\u003cb\u003e\u0026minus;\u0026thinsp;1\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1.89\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.47\u0026thinsp;\u0026plusmn;\u0026thinsp;0.16bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e3.17\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4.21\u0026thinsp;\u0026plusmn;\u0026thinsp;0.48ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5.28\u0026thinsp;\u0026plusmn;\u0026thinsp;0.31a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTotal P (mg g\u003c/b\u003e\u003csup\u003e\u003cb\u003e\u0026minus;\u0026thinsp;1\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1.53\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13d\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.83\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e3.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.71ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.91\u0026thinsp;\u0026plusmn;\u0026thinsp;0.25a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTotal K (mg g\u003c/b\u003e\u003csup\u003e\u003cb\u003e\u0026minus;\u0026thinsp;1\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1.52\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.70\u0026thinsp;\u0026plusmn;\u0026thinsp;0.36bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e5.39\u0026thinsp;\u0026plusmn;\u0026thinsp;1.14bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e7.14\u0026thinsp;\u0026plusmn;\u0026thinsp;1.19b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e9.72\u0026thinsp;\u0026plusmn;\u0026thinsp;1.13a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eCrop yield attributes significantly improved under higher BC application rate and its combination with NPK fertilizers such as BC20\u0026thinsp;+\u0026thinsp;CF increased crop biological and grain yield by 26 and 36% respectively followed by BC10\u0026thinsp;+\u0026thinsp;CF where these increments were 10 and 27% higher than control. Furthermore, 1000 grain weight and harvest index also significantly improved up to 17 and 14% respectively under BC20\u0026thinsp;+\u0026thinsp;CF treatment (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e"},{"header":"4 Discussion","content":"\u003cp\u003eBiochar application at different rates and their co-application with CF improved soil nutrient status as compare to control (Murtaza et al. \u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). We observed that application of biochar improved the nutrient status of soil, and at the same time, enhanced C, N and P availability for microbial utilization as reported by previous studies (Bu et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Naeem et al. \u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Zhu et al. \u003cspan citationid=\"CR115\" class=\"CitationRef\"\u003e2017a\u003c/span\u003e). However, no significant change was observed in soil pH under all the applied BC rates, as previously stated by (Rashid et al. \u003cspan citationid=\"CR77\" class=\"CitationRef\"\u003e2020\u003c/span\u003e) and (Zhu et al. \u003cspan citationid=\"CR116\" class=\"CitationRef\"\u003e2017b\u003c/span\u003e), who reported that alkaline soils have large buffering capacity, which resist towards any change in pH brought by BC application. (Griffin et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2017\u003c/span\u003e) found a little increment in soil pH after BC application, but significant enhancement in soil macro nutrients (K, NO\u003csub\u003e3\u003c/sub\u003e-N, Olsen P and NH\u003csub\u003e4\u003c/sub\u003e-N), as observed under current study, which indicate the direct role of BC on soil nutrient stock. Few other studies have indicated that the increase in soil nutrient levels is due to BC's labile C, N, P, and K contents which are continuously released in soil, and are readily available for plant uptake. In simple words, rise in soil nutrients can be attributed to the direct effects of BC on the soil (Aziz et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Qayyum et al. \u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Song et al. \u003cspan citationid=\"CR88\" class=\"CitationRef\"\u003e2018a\u003c/span\u003e). Moreover, BC contains larger surface area, negative surface charge and higher number of functional groups which play a key role against nutrients leaching (Yuan et al. \u003cspan citationid=\"CR108\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Additionally, presence of oxonium functional groups on BC surface make increase its anion exchange capacity (AEC) which help to retain NO\u003csub\u003e3\u003c/sub\u003e-N and phosphate (Banik et al. \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Sorrenti et al. \u003cspan citationid=\"CR92\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), while the negative surface charge of BC can retain cations like NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e and K\u003csup\u003e+\u003c/sup\u003e by sorption (Choudhary et al. \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIt was observed that biochar-based treatments had a key role in improving soil enzyme activities as observed by (Song et al. \u003cspan citationid=\"CR89\" class=\"CitationRef\"\u003e2019\u003c/span\u003e) and (Oladele \u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e2019\u003c/span\u003e), who examined that presence of organic C, MBC and MBN pools of BC provided organic substrates for enzymes which lead towards higher enzymatic activity. Although, it has also been reported that BC addition lead to a reduction in soil enzyme activities because of synthetic C mineralization (El-Naggar et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Moreover, few studies revealed that BC can adsorb a range of organic and inorganic molecules, this adsorption causes the blockage of reaction sites which decreases soil enzyme activities (He et al. \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2021a\u003c/span\u003e; Lehmann et al. \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Contrastingly, our study did not support this observation as results revealed that BC addition significantly increased the activity of all enzymes, as supported by the results of (Bailey et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2011\u003c/span\u003e) and (Ameloot et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2013a\u003c/span\u003e), who revealed that BC has few volatile compounds which increase soil enzyme activity such as urease, protease, dehydrogenase phosphatase and β-glucosidase under alkaline conditions. Furthermore, BC can also increase enzymatic activity by absorbing soil toxic substances (Salam et al. \u003cspan citationid=\"CR82\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Song et al. \u003cspan citationid=\"CR91\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) however, there was no sign of toxicity observed in the soil of current study. BC alters the soil biochemical properties (Elzobair et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2016\u003c/span\u003e) and nutrient availability (Lu et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), which lead to higher enzyme activities and therefore, significant correlation between BC concentration and soil enzymatic activity may directly support this suggestion. Interaction between soil enzymes and organic matter in soil determines the biogeochemical stability, which is manifested by the relative proportions of enzymes involved in CNP cycles. The balance between the ratio of microbial biomass and elemental composition of organic matter is essential for maintaining this stability (Waring et al. \u003cspan citationid=\"CR100\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Wei et al. \u003cspan citationid=\"CR101\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Inclusively we can suggest that, soil enzyme activity was improved with the increment in BC application rates although sometimes, it is based on the soil nutrients availability and climatic conditions (Waring et al. \u003cspan citationid=\"CR100\" class=\"CitationRef\"\u003e2014\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eGenerally, it has been reported that BC has the potential to improve soil biological properties, which leads to better soil microbial abundance because of BC's special structure and properties (Lucheta et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Ren et al. \u003cspan citationid=\"CR78\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Zhang et al. \u003cspan citationid=\"CR112\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In current study, a significant increment was observed in soil total PLFA, bacterial and fungal in BC treated soils but this activity increased as the BC application rate increased with CFs. Our findings are closely related to the results of (Chen et al. \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), where they reported that application of 3–9% of BC significantly improved the soil total PLFA content as compare to control. Moreover, the labile C and N pools of BC serve as a suitable habitat (i.e., energy and nutrient sources) for better growth and development of soil microbes (Zhao et al. \u003cspan citationid=\"CR114\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Previous literature has proved that soil supplementation with biochar and animal manure marginally improves microbial community which lead towards better soil structure and C content (Bowles et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Cheng et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Bacterial and fungal biomass was increased with the BC application rate in current study but these results are distinct to the meta-analysis of (Xu et al. \u003cspan citationid=\"CR105\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), as they observed reduction in soil bacterial and fungal community which was proved by previous 107 research studies. Similarly, (Chen et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2013\u003c/span\u003e) also found some negative impacts of wheat straw derived BC on soil bacterial and fungal activity. This variation indicates that BC effects are based on its feedstock type, pyrolysis temperature, soil type and application rate (Xu et al. \u003cspan citationid=\"CR105\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Further, it has also been revealed that soil microbial biomass is based on BC feedstock type as each feedstock have its own physicochemical characteristics and nutritious values (Luo et al. \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Additionally, soil pH is also influenced by the BC type, and this increase lead towards better soil microbial abundance. It has also been observed that porous structure of BC can be substantially affected by pyrolysis temperature (He et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2021b\u003c/span\u003e), as the temperature rises, BC surface area and pore volume is increased, which provide desired habitat to microorganisms whilst lower BC pore volume and surface area favors the growth of fungal hypha (Muhammad et al. \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Conclusively, elevated BC pyrolysis temperature and pore spaces could be less conducive for fungal better growth and abundance.\u003c/p\u003e \u003cp\u003eBacterial fungal ratio indicate soil C sequestration capability, as the fungal content upsurges soil, carbon storage potential also increases which plays a pivotal role in environment safety (Malik et al. \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). In our study, soil that received BC20 and BC20 + CF treatments had maximum fungal bacterial ratio, which reflects that application of BC alone or with chemical fertilizers could improve soil C sequestration potential and ecosystem stability. Moreover, in our findings, soil MBC, MBN and MBP were also increased i.e., particularly, at higher dose of BC application. Study of (Bhaduri et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2016\u003c/span\u003e) as well as (Biederman and Harpole \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2013\u003c/span\u003e) support our results, where they observed that application of BC significantly increased soil MBC and MBP respectively. Similarly study of (Zhang et al. \u003cspan citationid=\"CR113\" class=\"CitationRef\"\u003e2014b\u003c/span\u003e) also revealed that application of corncob BC significantly increased soil MBC and MBN. Generally, MBC carries approximately 3–7% of the total soil organic carbon while MBN and MBP consist of 1–5% of total soil nitrogen and phosphorus respectively (Oladele et al. \u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). These shifts in soil microbial biomasses indicates the soil carbon mineralization, growth and mortality ratio of microbes (Fang et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOur study revealed that poultry manure derived BC had significant positive effects on wheat crop growth and yield parameters. According to our results, crop growth was significantly improved as the BC application increased with NPK fertilizers. Our results are supported by (Khan et al. \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) and (Aziz et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), as they applied different feedstock based biochars on alkaline soils, and observed a significant increment on crop growth, nutrients uptake and grain yield. Furthermore, it has also been examined that plant response to BC is based on feedstock type, pyrolysis temperature and soil type as each feedstock has its own properties (Choudhary et al. \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In our previous experiment, we applied three different types of BCs i.e., poultry litter biochar, \u003cem\u003eAcacia modesta\u003c/em\u003e wood biochar and \u003cem\u003eDalbergia sissoo\u003c/em\u003e wood biochar, where we discovered that poultry litter biochar significantly influenced wheat growth along with their yield (Aziz et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Being an organic amendment, BC reduces soil compaction, nutrients loss and increases water holding capacity along with nutrient use efficiency which leads toward improvements in crop agronomic and yield parameters (Khan et al. \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Generally, BC affect soil nutrients availability by two ways: directly, by providing nutrients (work as nutrient and reduces the nutrient release and leaching) indirectly, by increasing nutrient and water holding capacities of soil as well as by causing variations in soil pH, CEC and soil microbial abundance (Azeem et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Moreover, BC has large surface area and high porosity, which provide suitable habitat to microbes for survival, which is very critical to nutrient solubilization, for instance, P solubilization leads to higher P uptake, which plays a significant impact on root growth, and ultimately increase nutrients uptake (Azeem et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Wahid et al. \u003cspan citationid=\"CR98\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Consequently, better nutrient uptake lead towards higher root-shoot growth and crop yield (Guo et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e "},{"header":"Conclusion","content":"\u003cp\u003eIn our study, we examined the interactive effect of different application rates of poultry litter biochar with chemical fertilizers on soil biochemical characteristics and wheat yield. Soil microbial biomass and enzymatic activities were significantly affected by the sole and co-application of BC20 with chemical fertilizers. Our results also showed that BC20 + CF amendment led to significant improvement in soil nutrient content (i.e. NO\u003csub\u003e3\u003c/sub\u003e-N, NH\u003csub\u003e4\u003c/sub\u003e-H, Olsen P, K, SOC, DOC, C/N). Furthermore, higher rate of BC application along with chemical fertilizers significantly increased soil enzymatic activities (URE, DEH, PHO, PRO and β-GLU) and soil microbial community such as MBC, MBN, MBP, PLFA, AMF, fungi, actinomycetes, bacteria and protozoa. Similarly, a higher fungal: bacteria ratio was also observed under BC20 + CF which indicated higher C sequestration and ecosystem stability. Consequently, growth and yield of wheat crop was considerably enhanced by the application of BC20 + CF. Overall, our study confirmed that co-application of BC with CF is an effective approach to improve soil biochemical characteristics and crop yield. Further studies are needed to evaluate the effect of different feedstock-based biochar and fertilizers on soil bacterial and enzymatic activity for better crop growth and yield.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors are thankful to the Institute of soil and environmental sciences, Pir Mehr Ali Shah Arid Agriculture University Rawalpindi, Pakistan for providing facility to perform the experiment.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConceptualization: Muhammad Abdullah Aziz, Khalid Saifullah Khan, Mosaed A. Majrashi; Methodology: Muhammad Abdullah Aziz, Rabia Khalid; Investigation: Khalid Saifullah Khan; Formal Analysis: Muhammad Abdullah Aziz; Data Analysis and visualization: Abdulaziz G. Alghamdi, Zafer Alasmary; Original draft preparation: Muhammad Abdullah Aziz, Muhammad Shabaan; Review and editing: all.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAhmad TI, Khan RE, Soharwardi MA, Shafiq MN, Gillani S (2021) Socioeconomics and agronomy of wheat yield in cotton-wheat cropping system in Punjab, Pakistan: A quality-quantity assessment. 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Ecotoxicology and Environmental Safety 144: 578-584. doi: https://doi.org/10.1016/j.ecoenv.2017.06.075.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":true,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"plant-and-soil","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plso","sideBox":"Learn more about [Plant and Soil](https://www.springer.com/journal/11104)","snPcode":"11104","submissionUrl":"https://submission.nature.com/new-submission/11104/3","title":"Plant and Soil","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Biochar, Soil microbial abundance, Soil enzymatic activity, Soil fertility, Crop yield","lastPublishedDoi":"10.21203/rs.3.rs-2910777/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2910777/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eAim\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIntegration of chemical and organic fertilizers not only improves soil biological health and plant growth but also reduces costs of agricultural production, and hence, is an economically feasible approach to sustain plant growth in developing countries.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe performed a field experiment to evaluate the impact of poultry litter derived biochar (BC; 10 and 20 t ha\u003csup\u003e-1\u003c/sup\u003e) with and without chemical fertilizers (CF) on soil nutrient availability, microbial abundance, and soil enzymatic activity.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCombined application of BC and CF significantly increased soil nutritional status and organic carbon (SOC), and these improvements were more prominent at higher BC level (20 tons ha\u003csup\u003e-1\u003c/sup\u003e), which increased soil microbial biomass carbon, nitrogen, and phosphorus by 27, 58, and 61%, respectively. Furthermore, BC20+CF treatment improved soil microbial abundances such as actinomycetes (24%), bacteria (70%), AMF (49%) and saprophytic fungi (38%). In terms of wheat growth and yield attributes, BC20+CF application enhanced spike length (55%), leaf area index (30%), tillers (12%), along with biological yield (26%) and grain yield (36%). BC20+CF application was also proved to be positively correlated with different soil enzymatic activities i.e., urease (24%), dehydrogenase (44%), and β-glucosidase (93%). Principal component analysis (PCA) analysis proved that co-application of BC @ 20 t ha\u003csup\u003e-1\u003c/sup\u003e along with CF was the most efficient treatment in terms of improving soil nutrient status and microbial activities.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eHence, combined application of BC and CF could be an efficient tool to improve soil health, plant growth and reduce cost of agricultural production.\u003c/p\u003e","manuscriptTitle":"Integrated application of biochar and chemical fertilizers improves growth and yield of wheat (Triticum aestivum) by altering soil biological heath","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-06-06 14:38:20","doi":"10.21203/rs.3.rs-2910777/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revisions","date":"2023-07-25T05:10:59+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"","date":"2023-06-16T07:15:32+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2023-06-03T15:07:50+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"Plant and Soil","date":"2023-05-14T22:15:31+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2023-05-14T13:47:43+00:00","index":"","fulltext":""},{"type":"submitted","content":"Plant and Soil","date":"2023-05-12T08:58:13+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"plant-and-soil","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plso","sideBox":"Learn more about [Plant and Soil](https://www.springer.com/journal/11104)","snPcode":"11104","submissionUrl":"https://submission.nature.com/new-submission/11104/3","title":"Plant and Soil","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"53ac6291-52a4-4451-a4a3-3641dbf306ec","owner":[],"postedDate":"June 6th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-02-19T15:11:27+00:00","versionOfRecord":{"articleIdentity":"rs-2910777","link":"https://doi.org/10.1007/s11104-024-06556-3","journal":{"identity":"plant-and-soil","isVorOnly":false,"title":"Plant and Soil"},"publishedOn":"2024-02-17 15:01:37","publishedOnDateReadable":"February 17th, 2024"},"versionCreatedAt":"2023-06-06 14:38:20","video":"","vorDoi":"10.1007/s11104-024-06556-3","vorDoiUrl":"https://doi.org/10.1007/s11104-024-06556-3","workflowStages":[]},"version":"v1","identity":"rs-2910777","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2910777","identity":"rs-2910777","version":["v1"]},"buildId":"0shC4O-rRljfh4Nq9OyMh","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}