Root-knot nematodes (RKN), particularly , pose a significant threat to agriculture, with current management heavily reliant on agrochemicals due to a scarcity of resistant crop varieties. This study explores orchid mycorrhizae, specifically , as a sustainable biocontrol method to mitigate nematode infestation in tomato plants. The research aimed to compare different application methods and dosages of mycelial suspensions to reduce nematode reproduction and enhance plant health. Two application methods, root immersion for 24 hours and soil drenching, were tested with mycelial suspension concentrations ranging from 5 to 25 g.L. Results showed that a 15 g.L suspension significantly reduced nematode reproduction factor (RF) by 53.4% and nematode density (DENS) by 49.4% while increasing root fresh weight (RFW) by 53.8% in drenching. In subsequent experiments, soil drenching with 15 g.L mycelial suspension again reduced RF by 32.41% and DENS by 28.52%, with increases in shoot length (SL) by 26.31%, RFW by 20.42%, and shoot fresh weight (SFW) by 22.20%. Enzymatic analysis revealed that plants treated with and inoculated with nematodes (Wc+Me) showed a substantial decline in RF (71.13%) and DENS (76.96%). Additionally, there was a marked increase in peroxidase (POX) and catalase (CAT) activity, with Wc plants displaying a 180% increase in POX and a 112.5% increase in CAT at root colonization onset. By day 21, Wc+Me plants exhibited further enzyme activity increases, with POX up by 128% and CAT by 67.6%. This study emphasizes the potential of in enhancing plant resistance and reducing nematode impact, presenting a promising alternative to chemical control.