The melanose disease causing Diaporthe citri is important pathogen in citrus plants worldwide. Currently, there are few researches related to biological control of melanose disease. In this study, we investigated control of Diaporthe citri by Bacillus amyloliquefaciens Y1 isolated from the soil. Low-cost fertilizer based PB medium and large-scale fermenter have been developed for field application of biological control. We prove that Y1 can be successfully grown in large-scale fermenters using PB medium without contamination. In addition, Y1 bacterial culture filtrates (BCF) of large-scale cultivation in PB medium (LP-Y1), present better growth inhibition of D. citri than small-scale cultivation in TSB medium (ST-Y1). Microscopic observations also showed that pathogen mycelial morphology was significantly affected by BCF, presumably due to metabolites produced by Y1. Finally, in pot experiment, pre-treatment with fungicide and Y1 to leaves significantly reduced disease incidence compared to the control group. This study showed that Y1 incubated with fertilizer based medium inhibited the occurrence of D. citri and could be grown in large scale field conditions without contamination as well as cost optimization.