IGM-SPECIFIC SERODIAGNOSIS OF ACUTE HUMAN CYTOMEGALOVIRUS-INFECTION USING RECOMBINANT AUTOLOGOUS FUSION PROTEINS

Portions of three human cytomegalovirus (HCMV) polypeptides, which wer e shown previously to be highly reactive with patient sera, were expre ssed in Escherichia coli as autologous fusion proteins. Purified recom binant polypeptides were used as antigens in enzyme linked immunosorbe nt assay (ELISA) and compared against assays which use natural viral a ntigen from cell culture for their ability to improve IgM-specific ser ology of acute HCMV-infection. A fusion protein (CM2) which contained two copies of the C-terminal portion of pUL44 (p52, aa 297-433) and on e copy of a highly reactive fragment of the major DNA-binding protein pUL57 (aa 545-601) proved to be superior in sensitivity and specificit y compared to assays which use culture derived antigen, A construct ex pressing one copy of the fragments from pUL44 and pUL57 in fusion with the 54 amino terminal residues of pUL32 (pp150, aa 994-1048) did not lead to an improved sensitivity compared to CM2. Adversely, this polyp eptide reacted with a number of sera from asymptomatic blood donors in fected latently with HCMV indicating low specificity of this antigen f or the detection of acute infection. Concordant results were obtained with an antigen that combined only the C-terminal portions of pUL44 an d pUL32 (CM3). ELISA experiments with sequential sera from renal trans plant recipients demonstrated that detection of IgM-antibodies using C M2 as antigen correlated closely with acute infection, whereas high le vels of IgM-antibodies against CM1 and CM3 persisted for a month follo wing acute HCMV-infection. These results indicate that the application of a single autologous fusion protein like CM2 as antigen for recombi nant ELISAs can improve significantly IgM-serodiagnosis of acute HCMV infection.