Alleviating arsenic toxicity in Spinacia oleracea by modulating accumulation and speciation using fungal intergeneric...

Protoplast fusion has potential for developing improved hybrid strains from genetically diverse fungal groups possessing different arsenic detoxification traits to combat as toxicity in crops. Arsenic (As) contamination of agricultural soil poses a threat to the agroecosystem and food safety. Fungal capacity to mitigate As toxicity (via biosorption, methylation, volatilization, and oxidation) could be enhanced through genetic improvement using protoplast fusion. Unlike conventional microbial consortia, protoplast fusion can produce stable fusants and may overcome incompatibility among strains, though fusants should be tested for long-term stability. In this study, intergeneric protoplast fusion was carried out between As-methylating strain FNBR-FA-03 and arsenite-oxidizing strain FNBR-FA-06 to generate novel fusants with improved As detoxification efficiency. The fusants were screened based on As volatilization, arsenite oxidation, plant growth promotion, and their potential to reduce As accumulation in Spinacia oleracea L tissues. The best performing fusant strain, FU 26(4), exhibited a substantial increase in As volatilization (9.7-fold) and arsenite oxidation (3.6-fold) compared to the parent strain FNBR-FA-03 and FNBR-FA-06, respectively. The fusant FU 26(4) alleviated As stress to Spinacia oleracea L. grown in As-contaminated soil (20 mg kg^-1) in terms of a 73.4% and 62.7% decrease in leaf and root As contents, respectively. The fusants also resulted in methylation of inorganic As to relatively less toxic MMA(As⁺⁵) and DMA(As⁺⁵), detected in the plant petiole and leaf. Apart from this, the fusant strains exhibited plant growth-promoting traits, with potential to improve the overall plant growth and physiology. The results of this study demonstrate the potential of the protoplast fusion technique to develop novel fusants that combine parental traits for the sustainable bioremediation of As-contaminated soils.