Application of allelopathic bacteria (AB) for weed suppression may be helpful to solve various environmental challenges posed by conventional weed control techniques. In our earlier studies, around 400 strains of rhizobacteria of five weeds and wheat were isolated, screened for production of phytotoxic substances, and tested for phytotoxic activity on wild oat and little seed canary grass, and possible effects on wheat under laboratory conditions. We obtained 13 strains inhibitory to wild oat (Avena fatua L.) and 11 to little seed canary grass (Phalaris minor Retz.). Five of these (13 and 11) strains also suppressed wheat (Triticum aestivum L.) while others either stimulated or remained ineffective on wheat in separate bioassays. The success of any weed biocontrol technique, however, depends on its response under field conditions. Therefore, the present study was conducted to investigate biological weed control of the five most efficient strains of AB under natural conditions in pot and field trials. Wheat was artificially invaded with wild oat in the pot trial through seeding. Wheat of the field trial was artificially invaded with wild oat and little seed canary through seeding. The selected strains belonged to pseudomonads (Pseudomonas putida, P. fluorescence, P. aeruginosa, and P. alcaligenes) and their inocula were prepared using sterilized peat. The inoculated seeds of wild oat and wheat were sown together in a pot trial. The inoculated seeds of wild oat, little seed canary grass, and wheat were sown together in the field experiment. The field was selected based on chronic infestation of these weeds. However, weed invasion was ensured by adding seeds of weeds (inoculated with the respective strains of AB, according to treatment plan). A severe invasion of wild oat was observed in the pot trial, which reduced the grain yield of infested wheat up to 60.8%. The effectiveness of applied strains controlled 22.0–76.3% loss of grain yield of infested wheat. Weed invasion in the field trial reduced the grain yield of the crop up to 56.3% and effectiveness of the applied strains controlled 29.0–60.7% loss of grain yield of infested wheat. The study of other agronomic, physiological, and chemical parameters of the crop and weeds supported these findings. Harnessing the potential of these strains exhibited in our studies may be helpful to introduce an innovative, sustainable, and eco-friendly weed control technique for production of wheat.