Abstract A new isolate of the human immunodeficiency virus (HIV) related to the HIV-2 strain was isolated from peripheral blood lymphocytes of an Ivory Coast patient with AIDS. This isolate referred to as HIV-2 EHO could be differentiated by its envelope precursor and external glycoprotein which are 20-kDa smaller than those of HIV-2 ROD isolate. Furthermore, the apparent molecular weight of the major core protein of HIV-2 EHO is 27 kDa instead of 26 kDa as in HIV-2 ROD isolate. In addition, the product of the vpx gene which is a characteristic feature of the HIV-2 strain, is 14 kDa in HIV-2 EHO compared with 16 kDa in HIV-2 ROD. In contrast to these, the envelope precursor of HIV-2 EHO forms a transient dimer during its maturation as is the case for HIV-2 ROD. In both cases the transmembrane proteins are 36 kDa and exist as homodimers of 80 kDa. Endoglycosidase H digestion experiments indicated that the 20-kDa difference between the two HIV-2 isolates is not due to a difference in the number of N-linked oligosaccharide chains per polypeptide, since deglycosylated envelope precursors of HIV-2 ROD and EHO have an apparent molecular weight of 80 and 60 kDa, respectively. Partial proteolysis of the envelope precursors from the two isolates with Staphylococcus aureus V8 protease gave a distinct pattern of polypeptides. These results suggest that the differences between the external envelope proteins of the two HIV-2 isolates are due to their amino acid composition. Accordingly, polyclonal antibodies raised against HIV-2 ROD envelope do not recognize the corresponding envelope proteins of HIV-2 EHO by immunoblotting and immunoprecipitation assays. These data illustrate that analysis of viral proteins could be useful for a rapid characterization of new viral isolates and show the heterogeneity of HIV-2 isolates in West Africa.