Calonectria leaf blight caused by Calonectria species is one of the most important diseases associated with Eucalyptus plantations in Asia and South America. This study aimed to clarify the distribution patterns of Calonectria species residing in different species complexes associated with diseased trees and soils during leaf blight outbreak season in Eucalyptus plantations in southern China. In this study, 482 Calonectria isolates obtained from diseased Eucalyptus trees and soils under these trees in eight sampling sites in three provinces were identified by DNA sequence analyses of tef1, tub2, cmdA, and his3 gene regions. Six species residing in three species complexes were identified: Calonectria pseudoreteaudii and C. acaciicola in the Calonectria reteaudii species complex; C. hongkongensis, C. aconidialis, and C. chinensis in C. kyotensis species complex; and C. auriculiformis in C. cylindrospora species complex. The habitats of Calonectria in different species complexes differed, C. reteaudii species complex inhabits in both diseased trees and soils, C. kyotensis species complex only in soils. The Calonectria leaf blight in the sampled regions was caused by species in the C. reteaudii species complex but not by the species in the C. kyotensis species complex. These findings suggest that the species in the C. reteaudii species complex should receive more attention in disease management, as they are the primary cause of the disease in the sampled regions.