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Anthocyanidin reductase (ANR) is an important regulator of flavonoid metabolism, and proanthocyanidins, the secondary metabolites of flavonoids, play an important role in the response of plants to pathogenic stress. Therefore, in this study, the expression analysis of the ANR gene family of <i>Gossypium barbadense</i> after inoculation with <i>Fusarium oxysporum</i> f. sp. <i>vasinfectum</i> (FOV) was performed at different time points. It was found that <i>Gb_ANR-47</i> showed significant differences in the disease-resistant cultivar 06-146 and the susceptible cultivar Xinhai 14, as well as in the highest root expression. It was found that the expression of <i>Gb_ANR-47</i> in the resistant cultivar was significantly higher than that in the susceptible cultivar by MeJA and SA, and different amounts of methyl jasmonate (MeJA) and salicylic acid (SA) response elements were found in the promoter region of <i>Gb_ANR-47</i>. After silencing <i>GbANR-47</i> in 06-146 material by VIGS technology, its resistance to FOV decreased significantly. The disease severity index (DSI) was significantly increased, and the anthocyanin content was significantly decreased in silenced plants, compared to controls. Our findings suggest that <i>GbANR-47</i> is a positive regulator of FOV resistance in <i>Gossypium barbadense</i>. The research results provide an important theoretical basis for in-depth analysis of the molecular mechanism of <i>GbANR-47</i> and improving the anti-FOV of <i>Gossypium barbadense</i>.