The western Hunan–eastern Guizhou Zn-Pb metallogenic belt is one of the important Zn-Pb mineralization regions in China. The Dadongla deposit, located in the northeast of Guizhou Province, is one of the typical Zn-Pb deposits in the region and has estimated resources more than 12 million metric tons (Mt) with an average grade of 4.11 wt% Zn+Pb. Its orebodies are hosted in the lower Cambrian Aoxi Formation dolomite, occurring as bedded, para-bedded in shape, and in conformity with the wall rock. The ore mineral assemblage is simple, dominated by sphalerite with minor pyrite and galena, and the gangue minerals are composed of dolomite, calcite with minor bitumen and barite. In view of the lack of geological and geochemical researches, the genesis of Zn-Pb ore is still unclear. Laser ablation-inductively coupled plasma mass spectrometry (LA-ICPMS) spot and mapping analyses were used to obtain sphalerite trace element chemistry in the Dadongla Zn-Pb deposit in Guizhou, China, aiming to constrain its ore genesis. The results show that sphalerite is characterized by the enrichment of Cd, Fe, Ge and Hg, corresponding with that of typical MVT deposits. Four zones were identified in the sphalerite crystal from Dadongla from the center to margin according to the color bands, in which the zone in the center, representing the early ore-stage sphalerite, is characterized by enrichment of Cd relatively, while the zone forming at late ore-stage is enriched in Ge and Hg relatively. The finding was controlled by differential leached metals content in ore-forming fluid from its source rock. Notably, critical element Ge trends to be enriched at the late ore-stage and follows a substitution of 2Zn2+? Ge4++□ (vacancy). Moreover, the calculated ore-forming temperature ranges from 79.9°C to 177.6°C by the empirical formula, which is similar to that of typical Mississippi Valley-type (MVT) deposits. Compared with the features of trace elements in sphalerite from different types of deposits, together with the geology, the Dadongla deposit belongs to an MVT Zn-Pb deposit.