The Blue Mountains Conservation Society has concerns over the proposed Blackheath-Little Hartley tunnel, particularly about impacts on the World Heritage Area.
President, Madi Mclean, said there were potential dangers both during construction and when the tunnel is in operation.
"There is a significant risk that the tunnel's construction could negatively impact aquifers supplying groundwater to groundwater-dependent ecosystems, and their dependent species, along both sides of the proposed tunnel corridor, including within the World Heritage Area," she said.
"We are particularly concerned by the threat of reduced groundwater to the Commonwealth-listed peat swamps that align the corridor and their populations of endangered giant dragonfly and Blue Mountains water skink, both groundwater dependent species.
"Both species have been recorded in the swamps between Medlow Bath and Mt Victoria, including in the Greaves Creek, Govetts Leap, Popes Glen, Porters Pass, Centennial Glen, Hat Hill Creek, Victoria Creek, upper Grose River and Kerosene Vale catchments."
She was also concerned that tunnelling could have a negative impact on aquifers that supply water to moist cliff-face communities. This includes an endangered heath, Epacris hamiltonii, which only occurs in the Mountains and is restricted to moist rock overhangs along Greaves, Katoomba, and upper Govetts creeks.
The society said a detailed hydrogeological study must be carried out along the tunnel corridor to look at any aquifers which might be affected during tunnel construction and their relationship with the swamps and moist cliff-face ecological communities.
"Despite this EIS being released for comment, we are continuing to call on the federal minister for a full EIS across the entire Great Western Highway project upgrade to ensure that matters of national environmental significance are addressed," Ms Mclean said.
"The piecemeal approach currently being taken is an injustice to our unique environment and won't provide a true understanding of the overall impact and long-term consequences of the upgrade."
The EIS can be seen here. Submissions are open until Wednesday, March 1.