In the present study, we describe the isolation and the characterization of three different Hyposoter didymator virus (HdV) lepidopteran host-expressed genes, the products of which might interfere with the host physiology during parasitism. In this report, we study the expression of HdV genes in Sf9 cells infected with HdV since results indicate that the Sf9 model mimics to some extent the in vivo model and may be utilized to study expression of HdV genes in lepidopteran host cells. This system allowed us to isolate three HdV-specific cDNAs, termed M24, M27, and M40. cDNA nucleotide sequence analysis demonstrated significant regions of homology. The three cDNAs displayed repeated sequences arranged in tandem array that might have evolved through domain duplication. Similar to other previously described polydnavirus host-expressed genes, two intron positions have been found in the M24 leader region. The cDNAs corresponded to RNAs of 1.5, 1.6, and 2.3 kb that are also detected in parasitized Spodoptera littoralis larvae. They are encoded by different genes likely located on different HdV DNA molecules. Corresponding RNAs are detected early postinfection and remain detectable for at least 10 days postinfection. They encode secreted glycine- and proline-rich proteins. An antiserum raised against a baculovirus recombinant M24-encoded protein detected similar proteins in the culture medium of infected lepidopteran cells and in parasitized host hemolymph. We propose that the three cloned genes belong to an HdV gene family specifically expressed in parasitized lepidopteran hosts.