Abstract
BackgroundTurfgrass species are agriculturally and economically important perennial crops. Turfgrass species are highly susceptible to a wide range of fungal pathogens. Dollar spot and brown patch, two important diseases caused by fungal pathogens Sclerotinia homoecarpa and Rhizoctonia solani, respectively, are among the most severe turfgrass diseases. Currently, turf fungal disease control mainly relies on fungicide treatments, which raises many concerns for human health and the environment. Antimicrobial peptides found in various organisms play an important role in innate immune response.Methodology/Principal FindingsThe antimicrobial peptide - Penaeidin4-1 (Pen4-1) from the shrimp, Litopenaeus setiferus has been reported to possess in vitro antifungal and antibacterial activities against various economically important fungal and bacterial pathogens. In this study, we have studied the feasibility of using this novel peptide for engineering enhanced disease resistance into creeping bentgrass plants (Agrostis stolonifera L., cv. Penn A-4). Two DNA constructs were prepared containing either the coding sequence of a single peptide, Pen4-1 or the DNA sequence coding for the transit signal peptide of the secreted tobacco AP24 protein translationally fused to the Pen4-1 coding sequence. A maize ubiquitin promoter was used in both constructs to drive gene expression. Transgenic turfgrass plants containing different DNA constructs were generated by Agrobacterium-mediated transformation and analyzed for transgene insertion and expression. In replicated in vitro and in vivo experiments under controlled environments, transgenic plants exhibited significantly enhanced resistance to dollar spot and brown patch, the two major fungal diseases in turfgrass. The targeting of Pen4-1 to endoplasmic reticulum by the transit peptide of AP24 protein did not significantly impact disease resistance in transgenic plants.Conclusion/SignificanceOur results demonstrate the effectiveness of Pen4-1 in a perennial species against fungal pathogens and suggest a potential strategy for engineering broad-spectrum fungal disease resistance in crop species.
Highlights
Turfgrasses, agriculturally and economically important crop species, are used worldwide for lawns of buildings, roadsides, athletic and recreational fields providing numerous benefits including reducing soil erosion, trapping dust and pollutants, moderating temperature, safer playing grounds and beautifying the environment [1,2]
To generate transgenic plants expressing Pen4-1 and study the role Pen4-1 plays in plant disease resistance, two chimeric DNA constructs were prepared containing either the coding sequence of a single peptide Pen4-1 (Figure 1a) or the DNA sequence coding for the transit signal peptide of the secreted tobacco AP24 protein translationally fused to Pen4-1 coding sequence (Figure 1b)
Using Agrobacterium-mediated transformation of embryogenic callus derived from mature seeds and phosphinothricin selection, we separately introduced the two chimeric gene constructs (Figure 1a, 1b) into a creeping bentgrass (A. stolonifera L.) cultivar, Penn A-4, producing a total of 25 independent T0 transgenic lines transformed with the construct, p35S-bar/Ubi-Pen4-1, and 5 with the construct, p35S-bar/Ubi-AP24::Pen4-1
Summary
Turfgrasses, agriculturally and economically important crop species, are used worldwide for lawns of buildings, roadsides, athletic and recreational fields providing numerous benefits including reducing soil erosion, trapping dust and pollutants, moderating temperature, safer playing grounds and beautifying the environment [1,2]. Dollar spot and brown patch, two important diseases caused by fungal pathogens Sclerotinia homoecarpa and Rhizoctonia solani respectively, are among the most severe and frequently occurring diseases on turfgrass lawns in the summer [3,4]. Fungicides are commonly applied to control fungal diseases This raises concerns about the potential emergence of new pathogen strains as a result of intensive use of chemicals [5,6,7]. Large scale agricultural use of DMIs since 1970s has led to the emergence of resistant genotypes of several phytopathogenic fungi impacting different crop and fruit species including turfgrass [6,8,9,10]. Dollar spot and brown patch, two important diseases caused by fungal pathogens Sclerotinia homoecarpa and Rhizoctonia solani, respectively, are among the most severe turfgrass diseases. Antimicrobial peptides found in various organisms play an important role in innate immune response
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