The zinc mines of Tennessee are known to contain high concentrations of certain trace elements (Cu, Ga, Ge, Fe, and Cd). Here, we investigate and compare sphalerites from two mining districts in the center (Central Tennessee Mines: CT) and east (East Tennessee Mines: ET) of Tennessee in order to (1) understand the substitution mechanisms that control the incorporation of trace elements in sphalerite, and (2) relate these mechanisms to the conditions of metal deposition. Electron microprobe and LA-ICP-MS analyses reveal complex chemical zoning in the sphalerites together with coupling of two main groups of trace elements: copper, germanium, and gallium; and iron and cadmium. The zoning is interpreted in terms of substitution mechanisms using different equations, including those previously described, such as 2Cu+ + Ge4+ ↔ 3Zn2+, Cu+ + Ga3+ ↔ 2Zn2+, Fe2+ + Cd2+ ↔ 2Zn2+, as well as new equations involving Ge2+ and vacancies: Ge2+ ↔ Zn2+, □ + Ge4+ ↔ 2Zn2+, and □ + 2Ga3+ ↔ 3Zn2+.
Electron microprobe analyses of euhedral crystals of sphalerite highlight a correlation between the presence of trace elements and preferential crystallographic growth directions: copper and germanium are preferentially incorporated along the  growth direction, while cadmium is preferentially incorporated along the  growth direction. Raman spectroscopy highlights for the first time the possible coexistence of two different crystallographic forms of ZnS in some samples: cubic sphalerite and a non-cubic form, possibly a wurtzite-type structure. This is supported by the LA-ICP-MS data, which demonstrates a relationship between the trace element content and crystallographic structure: the non-cubic form of ZnS is enriched in copper, germanium, and gallium, while iron and cadmium are the most abundant trace elements in the cubic sphalerite. Although most of the main substitution mechanisms are observed in sphalerites from the two mining districts, the sphalerites differ in terms of their germanium grade and valency. These differences can be interpreted both in terms of fluid composition and redox conditions, suggesting that two different episodes of mineralization occurred in Tennessee.