The design and construction of functional interlayers for lithium–sulfur (Li–S) batteries has attracted much attention and was demonstrated to be effective to alleviate the notorious “shuttle effect.” An often neglected issue is that the introduction of interlayer will reduce the overall energy density of the battery. In this work, we report a sulfur-infiltrated mesoporous silica/carbon nanotube (CNT) composite as an interlayer for Li–S batteries. The mesoporous silica with large surface area (842 m2 g−1) and pore volume (0.85 cm3 g−1) can not only ensure abundant exposed sites for polysulfide capture but also accommodate a large amount of sulfur inside the pore structure. CNT was composited with silica to enhance the electronic conductivity of the interlayer, which is beneficial for fast sulfur redox reaction kinetics and improved utilization of sulfur. Compared to the pristine and CNT-modified separator, the mesoporous silica/CNT composite-modified separator enables better cycling stability and rate performance. More importantly, it was demonstrated that separately incorporating sulfur into a cathode and interlayer enables better battery performance than locating all the sulfur in the cathode. At a total sulfur loading of 4 mg cm−2 (3 mg cm−2 sulfur on the cathode and 1 mg cm−2 on the mesoporous silica/CNT interlayer), a high initial discharge capacity of 1410 mAh g−1 and a retained capacity of 952 mAh g−1 after 100 cycles were exhibited. This work provides important guidance for future design of functional interlayers for practical Li–S batteries.