Stem rot of Anoectochilus formosanus (Af) caused by Fusarium oxysporum (Fo) is a major limiting factor to jewel orchid production in Taiwan. Fo causes discoloration in vascular tissues. However, some newly collected Fo isolates from Af stem rot do not cause vascular discoloration, suggesting changes may have occurred in the pathogen. Among recent Fo isolates from Af there are two colony types, the cottony alba (CA) and the sporodochial (S). In order to confirm that both colony types cause Af stem rot, 200 isolates were obtained from diseased stems in Nantou County and characterized by colony type and whether or not the infected plants had vascular discoloration. Isolates of both the CA and S types caused stem rot of Af; some isolates in each colony type caused vascular discoloration whilst others did not. Pathogenicity tests with 22 isolates resulted in stem rot disease severity ratings on Af of 3·1–4·0 and 2·1–4·0 with CA and S type colonies, respectively. The same isolates failed to cause disease on Cattleya, Dendrobium or Phalaenopsis plants. Phylogenetic analysis of partial intergenic spacer sequences showed that these isolates were distinguishable from other formae speciales of Fo and could be separated into two groups correlated with the CA or S type colonies with high bootstrap. Based on pathogenic, morphological and molecular characterizations, the Fo that causes stem rot of Af is proposed to be a new forma specialis, F. oxysporum f. sp. anoectochili, with different pathotypes.