Spring low-temperature stress (LTS) has become a major limiting factor for the development of high yield, quality and efficiency in wheat production. It not only affects the function of wheat leaves and the development of spikes but also impacts stem lodging resistance, and may experience elevated risk of stem lodging. This study conducted a field pot experiment to assess the effect of phosphorus fertilizer application mode on wheat stem lodging resistance under spring LTS. Two wheat varieties, Yannong19 (YN19, cold-tolerant variety) and Xinmai26 (XM26, cold-sensitive variety) used as the experiment material. Two phosphorus fertilizer application models including traditional phosphorus application (TPA) and optimized phosphorus application (OPA) were employed. Temperature treatment was conducted at 15 °C (CK) and −4 °C (LT) in a controlled phytotron. Our results showed that spring LTS decreased the stem wall thickness and internode fullness, and altered stem anatomical structure and chemical composition, resulting in a decrease in wheat stem mechanical strength and lodging resistant index. Compared with TPA, the OPA increased the stem wall thickness and internode fullness. The thickness of the stem mechanic tissue layer and parenchymatous tissue, and the area of the large vascular bundle and small vascular bundle were increased by the OPA, which alleviated the damage to stem cell walls caused by spring LTS. At the same time, the OPA also increased the contents of lignin, cellulose, and soluble sugar, improving the C/N ratio in wheat stem. Due to the improved stem morphological characteristics, anatomical structure, and chemical compositions, the wheat stem exhibited enhanced lodging resistance, which increased the lodging resistant index of the 2nd and 3rd internodes of YN19 and XM26 by 27.27%, 11.63% and 14.15%, 15.73% at the dough stage compared with TPA under spring LTS. Meanwhile, OPA could not only alleviate the yield loss caused by spring LTS, but also increase the grain yield without spring LTS. This study indicated that OPA enhances wheat stem lodging resistance under spring LTS, and would be meaningful and practical for improving wheat resistance to low-temperature stress.