Screening of nitrogen-fixing bacteria based on a novel colorimetric method

Document Type : Original Article

Authors

1 Department of Microbiology, Faculty of Biological Sciences, Alzahra University, Tehran, Iran

2 Microbial Biotechnology Department, Agricultural Biotechnology Research Institute of Iran, Agricultural Research, Education and Extension Organization, 3135933151, Karaj, Iran

10.30466/asr.2025.55844.1870

Abstract

Sustainable agriculture development has led to a reduction in nitrogen fertilizer consumption and allocating 70% share of nitrogen biofertilizers in the global microbial market. The complexity and high cost of conventional methodologies is one of the main challenges for diazotrophic bacteria screening and then simple, precise and low cost techniques should be introduced. In this research, a novel colorimetric method developed based on the converting spectrophotometric data into RGB and HSV items through smartphone Color Grab application to estimate the biological nitrogen fixation. A total of 12 Bacillus subtilis isolates and Azotobacter chroococcum were screened for nitrogen fixing based on the color change of NFB medium containing bromothymolblue. The nitrogen concentration was quantified using a standard curve obtained via spectrophotometric data. Conversion of spectroscopic data into smartphone colorimetry items provides the fast, accurate and low-cost monitoring of nitrogen-fixing strains. According to the colorimetric results, S indice (R2 ≤ 0.97) in both open and closed systems were successfully used for standard curve drawing in the range of zero to 18 µg/ml to estimate different nitrogen concentrations. Based on the NFB medium color shift, all bacterial isolates except A15d and g33b strains were able to ammonia production.The nitrogen fixation capacity of bacterial isolates was in the range of 1.65 µg/ml to 13.32 µg/ml. Strains g19l and W exhibited the highest nitrogen concentration estimated as 13.32 and 12.49 µg/ml, respectively. The high concordance between the spectroscopic and colorimetric data substantiated the efficacy of the employed method for precise and streamlined screening of diazotrophic bacterial strains.

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History

  • Receive Date: 03 January 2025
  • Revise Date: 08 May 2025
  • Accept Date: 12 May 2025

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