Leaf surface fertilization with liquid fertilizer produced from amino acids constitutes a potentially important source of nitrogen and is important for plant production. However, few reports have focused on the plant growth promotion by novel liquid fertilizers created by new amino acid resources, let alone the influence on leaf microbiota. In this study, the effects of liquid fertilizer, created by amino acids hydrolyzed from animal hairs with or without the PGPR strain Bacillus amyloliquefaciens SQR9, on crop yield and leaf microbiota were investigated. The results showed that leaves sprayed with amino acid liquid fertilizer (AA) and liquid biological fertilizer (AA9) persistently increased cowpea yields compared to the control amended with chemical fertilizer (CF). Fertilization with amino acid fertilizer showed no significant difference in microbial composition compared with the CF treatment; however, the introduction of functional microbes altered the microbial composition. Pearson correlation analysis, VPA analysis and SEM models all revealed that the amino acids liquid fertilizer application, but not the functional strain or the altered microbiota, performed as the direct driver attributing to yield enhancement. We conclude that leaf fertilization with a novel amino acid liquid fertilizer can greatly enhance the crop yield and that the addition of beneficial microbes may perform the role in further altering the composition of leaf microbiota.