The pol and gag gene fragments of small ruminant lentivirus field isolates collected in the last decade in Italy were amplified, sequenced, and analyzed. Phylogenetic analysis revealed that the majority of ovine isolates form a distinct cluster more similar to caprine lentivirus prototypes than to the visna virus prototype. These findings confirm and extend those reported by Leroux et al. (Arch. Virol., 142:1125-1137, 1997). Moreover, we observed that a variable region of Gag, included in the fragment analyzed, corresponded to one of the three major capsid antigen epitopes, which suggests that the antibody response to this epitope may be type specific. To test this hypothesis, two recombinant peptides, derived from the Icelandic prototype K1514 and this novel genotype, were expressed and used in an enzyme-linked immunosorbent assay to screen a panel of ovine and caprine sera collected from different geographical locations in Italy. Several sera reacted in a type-specific manner, indicating that in a diagnostic setting the combination of at least these two type-specific peptides is necessary to cover a wide range of infections. Additionally, these results support the hypothesis of cross-species transmission based on the phylogenetic analysis described above. This has implications for the control and eradication of small ruminant lentivirus infections.To date, maedi-visna virus (MVV) and caprine arthritis encephalitis virus (CAEV) are considered to be two antigenically related and genetically distinct lentiviruses of the Retroviridae family (3). Since the cross-reactivity between MVV and CAEV involves the major structural proteins (7), a number of serological tests have been proposed, based on ovine strains, to detect specific antibodies in both species. Ovine lentiviruses are usually easier to grow in tissue culture, and well-adapted strains have been extensively used to develop native-based immunoassays (9, 19). Furthermore, since sequence information was first produced from ovine strains, recombinant antigens have been largely employed and characterized from ovine isolates (12,13,14,25,33). The development of a diagnostic test capable of detecting the widest range of infection is obvious from a practical point of view. The current concept of the universality of a single-strain-based immunoassay is based on the finding that the gag-encoded capsid antigen (CA) and the env-encoded transmembrane (TM) protein are conserved among small ruminant lentiviruses (7, 21), despite the variability of the env-encoded surface antigen (1, 31). However, further characterization of the immunodominant epitopes of the major CA has shown that these epitopes are at least in part quite variable, questioning the use of single-strain-based immunoassays for diagnostic purposes. In particular, the immunodominant region of the CA involved in the cross-reaction between ovine and caprine infection was recently identified (22). Partial mapping studies suggested that at least two consecutive linear epitopes, located in the N-terminal half of t...