Simian Virus 40 (SV40) antibodies in elderly subjects

We read with interest the article of Wong et al.1 They described the prevalence and distribution among ethnic groups of polyomavirus SV40 antibodies in women in Houston, Texas. Their data were obtained using a plaque-reduction neutralization assay, which is a high specific method to detect SV40 antibodies in human serum samples. However, this approach has significant disadvantages, such as: (i) time consuming, (ii) intensive labour, (iii) data are obtained in weeks, (iv) high cost. SV40-neutralization assay, because of these limitations, cannot be employed for large sample size. SV40 was recognized in the 1960 as contaminant of both inactivated (Salk) and live (Sabin) anti-poliomyelitis vaccines. 2,3 After its isolation, SV40 was experimentally characterized as a transforming and oncogenic virus. 2,3 Detection of SV40 antibodies has been attempted using different serologic methods, but the high protein homology among the three main polyomaviruses, SV40, BK virus (BKV) and JC virus (JCV), hampered the results, which were always affected by some cross-reactivity. 4,5,6 Thus, it remains to be determined whether SV40 is also a human virus and the extend of its infection rate. The identification of SV40-seropositive healthy individuals and serum antibody reactivity to SV40 antigens are of paramount importance in revealing the prevalence of SV40 infection in humans. In particular, little information is available about SV40 infection in elderly subjects that are prove to develop tumors more often than individuals of younger age. We development a specific and sensitive indirect ELISA to detect SV40 antibodies in serum samples of normal subjects 6 and patients affected by malignant pleural mesothelioma. 7 In the present study, the same immunologic test which employs synthetic peptides as mimotopes from the SV40 capsid viral protein 1-3 (VPs 1-2-3) antigens was used to investigate and compare age-specific seroprevalence for SV40 in elderly healthy subjects. Serum samples (n=105) from subjects aged 66-91 ys old, diluted at 1/20, were analyzed by indirect ELISAs. Samples were tested for reactivity to SV40 epitopes of VP1 and VP2/3 capsid proteins, named VP1 B peptide and VP2/3 peptide, respectively. 6,7 Sera were considered SV40-positive when reacting with both VP1 B and VP2/3 C peptides. The overall prevalence by combining SV40-positive sera, both for VP1 B and VP2/3 C peptides, was 21.9% (23/105) (Figure 1). No positive results were obtained with human peptide used as a control, 6,7 which had an OD of less than 0.1 (0.088-0.098). This OD value is usually consistent with SV40-negative sera. 6,7 A prevalence selection corresponding to 15.5%, 16.6%, and 36.6%, was observed in subjects within the cohort aged 66-74 ys, 75-81 ys, and 82-91 ys old, respectively. However, the different prevalence of SV40 antibodies among these cohorts of individuals was not statistically significant (Chi-square test with Yates’ correction and Fischer’s exact test). The two indirect ELISA tests, with two distinct VP, B and C peptides, gave overlapping results, thus confirming the presence of antibodies against SV40 VPs in human sera from elderly healthy subjects. SV40-positive sera, diluted at 1/20, had a mean value of approximately 0.27-0.31 OD in the cohorts of subjects aged 66-74 ys and 75-81 ys, while a lower mean value of 0.19 OD was detected in sera from individuals aged 82-91 ys old. This difference is not statistically significant (P > 0.05, Anova and Newman-Keuls Comparison test). Figure 1. To verify the antibody titer, 12 sera from elderly individuals found to be SV40 positive for both VP1 B and VP2/3 C peptides were serially diluted from 1/20 to 1/160 and further investigated by indirect ELISA. The assay indicated that these sera carry antibodies against SV40 that remain positive at a 1/80 dilution. Figure 2. Our study indicates that serum samples from elderly healthy individuals react with specific synthetic peptides, mimicking SV40 capsid protein antigens, with an overall prevalence of 21.9%. Immunologic data were obtained by the indirect ELISA with viral peptides/mimotopes, which are specific for SV40 viral capsid proteins. No cross-reactivity occurred with the closely related human polyomaviruses BK and JC, 6,7 which are ubiquitous in human populations with a prevalence of serum antibodies of 60-90%. 8 Interestingly, our immunological data on SV40-prevalence in sera from elderly subjects are unique data for this cohort. Indeed, this is the first study reporting on a specific immunological assay for the detection in elderly subjects of antibodies against SV40. Our result suggests that SV40 infects the human population with a low prevalence and a low titer. A low percentage of adults positive for SV40 antibodies was reported in other studies where SV40 VLPs were used as antigens. 4,6 In these investigations, human sera were adsorbed with BKV and JCV VLPs, thus subtracting a substantial amount of antibodies cross-reacting with SV40 VP epitopes. It has also been proposed that a distinct prevalence of SV40 infection occurs in different ethnic groups, with distinct hygienic and environmental standards. 1 Our research data obtained by indirect ELISAs might be applied to future practice in the clinical laboratory analysis detecting SV40 antibodies and their titers in serum samples from individuals/patients infected by SV40. Immunologic data, obtained with these SV40 VP mimotopes, indicate that (i) specific SV40 antibodies can be detected in serum samples from elderly healthy subjects; (ii) in these healthy individuals, aged 66-91 ys, the SV40 infection rate increases in elderly with the age; (iii) the proportion of elderly subjects with high SV40 antibody levels decreases with age. We showed that SV40 antibodies are more prevalent in the cohort of 82-91 ys, and that the differences in seroprevalence among age cohorts are suggestive of differences in the route of the infection. Interestingly, the human population we studied is to old to have been infected by SV40 through the early anti-polio vaccines contaminated by this virus, thus suggesting that SV40 is spreading in human independently from those vaccines. Immunological data from this study and other investigations 1,3,7,8 indicate that SV40 is also a human virus. Alternatively, it may be that another, as yet undiscovered, polyomavirus infect humans. In this case, positive immunologic data could be due to a new polyomavirus closely related to SV40.