The genomic origin of alien chromosomes carrying resistance to wheat streak mosaic virus (WSMV) and the wheat curl mite (WCM), Aceria tosichella Keifer, was evaluated in nine wheat-alien addition, substitution, or translocation lines, derived from hybrids of wheat with Thinopyrum ponticum (Podp.) Barkworth & D.R. Dewey (2n = 10x = 70), or with Th. intermedium (Host) Barkworth & D.R. Dewey (2n = 6x = 42). One of the four wheat × Th. ponticum lines was resistant to WSMV and WCM. The other three lines were resistant to WCM, but susceptible to WSMV. Five wheat × Th. intermedium lines were susceptible to WCM, but were highly resistant to WSMV. Genomic in situ hybridization (GISH) using different genomic DNA probes demonstrated that all WSMV- and WCM-resistant lines carried an alien chromosome(s) related to the Js genome in Th. ponticum and Th. intermedium. The Js alien chromosome(s) displayed a special GISH hybridization pattern with the S genomic DNA probe, in which S genome DNA hybridized strongly in the centromeric regions and occasionally in the terminal regions with no or a weak hybridization signal in the middle of the two arms of the chromosomes. The WCM-resistant wheat × Th. ponticum lines N/5.10.10 and 63-30-2-2-2-8-1 carried the same short arm of chromosome 6Js derived from the partial amphiploid Agrotana, while the WSMV-resistant wheat × Th. intermedium lines T-Ai, CI15092, CI17766, A29-13-3-1 and KS93WGRC27 all had the same alien chromosome or chromosome arm of 4Js from Th. intermedium. The Js genome present in Th. ponticum and Th. intermedium is an important source of WSMV and WCM resistance that can be transferred into wheat.Key words: wheat-Thinopyrum lines, WSMV- and WCM-resistance, genomic in situ hybridization, Js genome.