The effects of nitrogen and biofertilizers (mycorrhiza and azotobacter) on chlorophyll content, yield and grain filling components of wheat‏

Introduction
Nitrogen (N) is an essential element in crop plants and it’s ‎application is one of the important  factors for improving growth, chlorophyll content, rate and grain filling period as well as ‎grain yield of wheat. Although nitrogen is the major macronutrient determining grain filling components of wheat, but the release of large rates of nitrogen fertilizer into the environment can cause a serious environmental problem such as groundwater pollution. In such a situation, several strategies have been proposed to decrease the effects of pollution caused by large rates of nitrogen fertilizer. In this regard, application of biofertilizers such as Azotobacter and mycorrhiza can decrease the need for chemical fertilizers and adverse environmental effects, and increase crop yield. The aim of this study was to evaluate the effects of nitrogen and biofertilizers (mycorrhiza and Azotobacter) on chlorophyll content, yield and grain filling components of wheat.
Materials and methods
The factorial experiment was carried out in randomized complete block design with three replications at the research field of Faculty of Agriculture and Natural Resources, University of Mohaghegh Ardabili, Ardabil, Iran, in 2022-2023. The experimental factors were included the application of biofertilizers in eight levels (no application as control, and application of Azotobacter, mycorrhiza Glomus moseae, mycorrhiza Glomus intraradices, Glomus moseae and Azotobacter, Glomus intraradices and Azotobacter, Glomus moseae and Glomus intraradices, Glomus moseae and Glomus intraradices along with Azotobacter), and nitrogen fertilizer rates at three levels (no application as control, and application of 100 and 200 kg urea.ha^-1). The wheat cultivar ‘Chamran’ was used in this experiment with the optimal density of 400 seeds.m^-2. Mycorrhiza fungi (Glomus mosseae and Glomus intraradices) were obtained from Turan Zist Fanavar Institute, Shahrood, Iran. The soils with mycorrhiza fungi were treated based on the manufacturer’s protocol (20 g.m^-2). Azotobacter were isolated from the rhizosphere of wheat by Research Institute of Soil and Water, Tehran, Iran. The strain density of microorganism used as Azotobacter in this experiment were 1×10⁸ bacteria per milliliter (10⁸ cfu/ml⁻¹). To study the root volume, a number of plastic bags were put in each plot before planting. The roots were removed at maturity stage and after washing, root volume was measured by graduated cylinder. Two piecewise linear model was used to determine the grain filling parameters. To determinate grain filling components (such as grain filling period, grain filling rate, effective grain filling period), the first sampling was done 10 days after heading, and other samplings were done in 4-days intervals to determine the accumulation of grain weight. For determination of biological yield and grain yield, 0.2 m² was harvested in each plot.
Research findings
The results showed that application of Azotobacter and mycorrhiza (Glomus moseae and Glomus intraradices) with 200 kg urea.ha^-1 increased chlorophyll-a (48.52%), chlorophyll-b (39.47%), total chlorophyll (45.67%), carotenoid (58.94%), maximum grain weight (41.84%), grain filling rate (7.97%), grain filling period (18.44%), effective grain filling period (31.33%) and root volume (37.43%) compared to no application of biofertilizers and nitrogen. Also, application of biofertilizers (Glomus mosseae and Glomus intraradices with azotobacter) along with 200 kg urea.ha^-1 increased plant height, spike length, 1000 grain weight and grain yield of wheat by 51.5%, 42.85%, 47.19% and 36.68%, respectively, compared to no application of nitrogen and biofertilizers.
Conclusion
According to the results of this study, it seems that the application of biofertilizers (Azotobacter and Mycorrhiza) and nitrogen fertilizer can increase grain yield of wheat by improving the grain filling components and some physiological traits.