The short shelf life of Chinese steamed bread (CSB) at room temperature due to moldy contamination restricts its popularity. This study aimed to evaluate two selected lactic acid bacteria strains, namely Lactiplantibacillus plantarum LWQ17 (LWQ17) and Pediococcus pentosaceus LWQ1 (LWQ1), on their antifungal activities and potential applications in CSB making. The results showed that both strains, especially LWQ17, displayed remarkable inhibition against Aspergillus niger and Penicillium polonicum, the representative species found in CSB spoilage. LWQ17 and LWQ1 destroyed the cell membranes of the fungi and caused a loss of intracellular substances. When applied in CSB making, both strains could delay the germination of mold on the surface of CSB in a season-dependent way. LWQ17 delayed the germination of mold by 4 d in Spring and Winter, while 1 d in Summer and Autumn. Further study revealed that their antifungal activities were greatly influenced by temperatures and humidity. Both strains could completely inhibit the germination of mold during two weeks of storage under 37 °C but with different humidity levels. Moreover, our results demonstrated that acidic substances were crucial in inhibiting the fungi, while proteinic substances and hydrogen peroxide played an auxiliary role. Analysis of their acids by targeted metabolomics showed that LWQ17 produced more kinds of acids, and the amount of phenyllactic acid produced by LWQ17 was 60 times higher than that produced by LWQ1, which may explain its better antifungal efficacy. This study provides a reference for employing lactic acid bacteria in CSB preservation under different conditions.