Biological management of root rot disease of sour orange (Citrus aurantium L.) caused by Phytophthora nicotianae

Abstract

Root rot disease of sour orange (Citrus aurantium L.) caused by Phytophthora nicotianae was investigated using morphological, molecular, and biological control approaches. Seven Phytophthora spp. isolates were obtained from infected citrus roots collected from different locations in Babil Governorate, Iraq. Morphological characterization on PDA medium revealed fast-growing, white to cream-colored, cottony colonies with irregular margins. Pathogenicity tests indicated that P. nicotianae isolates Phy1 and Phy5 exhibited the highest disease severity, reaching 74.87% and 71.53%, respectively, after 30 days of inoculation. Molecular identification based on ITS1 and ITS4 regions confirmed the morphological diagnosis. The pathogenic fungus P. nicotianae was registered under accession number PV99958.1 and showed 85% sequence similarity with the Moscow isolate (AY773099.1). The biocontrol fungus Trichoderma harzianum was registered under accession number MT358869.1 and exhibited 98.44% similarity with the reference isolate (PQ409493.1). In vitro antagonistic assays demonstrated that Lysinibacillus sphaericus completely inhibited the growth of P. nicotianae (100% inhibition) at a concentration of 160 × 10⁵ CFU ml⁻¹. Brevibacillus paramycoides also showed strong antifungal activity, achieving 94.44% inhibition at 8 × 10⁸ CFU ml⁻¹, while T. harzianum reduced fungal growth by 91.11%. Greenhouse experiments revealed that biological control agents (L. sphaericus, B. paramycoides, and T. harzianum), in addition to animal organic wastes (cow, sheep, and poultry manure), significantly reduced disease severity and seedling mortality compared with the untreated control. These results indicate that integrated biological control strategies represent a promising and eco-friendly approach for managing root rot disease caused by P. nicotianae in sour orange