<p>Trichomoniasis infected with Trichomonas vaginalis (T. vaginalis), can cause mild symptoms like itching and burning but can also lead to more serious adverse outcomes. While typically treated with metronidazole, this medication can face resistance from T. vaginalis and some individuals may experience side effects. Hence, the research on effective therapeutic methods is essential to improve traditional therapy for trichomoniasis. Alloferon, known as an anti-viral, anti-inflammatory, and anti-tumoral agent, has recently shown promise in regulating anti-oxidant proteins. This suggests potential as therapeutic agent for parasitic infection, as oxidative stress is associated with increased drug susceptibility in organisms. Therefore, we investigated the effect of alloferon in T. vaginalis and its potential enhancing effect in combination therapy with metronidazole. T. vaginalis treated with alloferon reduced the activity of its energy-producing organelles (hydrogenosomes) and changed structure of hydrogenosome. In addition, alloferon induced cell cycle arrest in the S phase of T. vaginalis, thereby leading to decreased proliferation. While metronidazole alone at a sublethal concentration (~ 50% inhibition) was ineffective, combining it with alloferon, significantly suppressed motility and proliferation in T. vaginalis. Therefore, our results indicated that alloferon could be an adjunct agent in combination therapy with metronidazole for trichomoniasis.</p>