Toxoids

We used data from a study that determined the antibody responses to a number of vaccine and viral antigens in a group of 45 individuals over a period of between 5 and 26 years, with a mean range of about 15.2 years (see [12 (link)] for details). These samples were taken as part of a center-wide, comprehensive program to permit serologic testing of people working in close proximity to nonhuman primates. We focused on the antibody responses to measles, rubella, vaccinia, mumps, and VZV antigens that were elicited either by a live-attenuated vaccine or natural infection, as well as antibodies to tetanus and diphtheria antigens that were elicited by immunization with inactivated protein toxin (i.e., toxoid) vaccines.
Because we wanted to consider the decay of antibody in the absence of boosting, the antibody data were curated to remove spikes due to revaccination or infection as described in more detail previously [12 (link)]. This involved censoring to exclude the following: timepoints for 3 years after immunization, when there was a rapid change in antibody levels [12 (link),16 (link)], seronegative or unvaccinated individuals, and individuals with fewer than 4 contiguous data points. We then kept the time series with the largest number of contiguous data points for the response of each individual to each vaccine or virus antigen.
As previously described, antibody titers were measured using ELISA and calibrated when possible in terms of international units (IUs). This allowed us to rescale the antibody concentration by dividing it by the level at which protection is lost [8 (link),17 (link)–21 (link)]. In our plots and analysis, the magnitude of responses is shown as the log of the scaled titer. We did not have a level at which protection is lost for mumps and VZV, and for these, we scaled against the threshold of detection for the ELISA assay for that antigen. The level of antibody required for protection for different infections was taken from the literature and was assumed to be the same for all individuals—we did not consider variation in the protective threshold between different individuals due to lack of relevant data (see Discussion). Note that we did not consider the absolute magnitude of the antibody response (e.g., moles/L or mg/mL) because the ELISA assays used do not measure this quantity.
Estimating the variability in the magnitude of the responses of different individuals to a given vaccine required taking into account uncertainty in the time of vaccination or infection and the different ages covered by the time series for different individuals. Because we do not find a significant correlation of antibody titer with age (and gender), but do find a strong correlation with time (see analysis in [12 (link)]), we used time rather than age as the main factor governing antibody titer. Because we did not know the time of vaccination, we shifted the time axis so that time equal to 0 corresponded to the mean age of the time series for each individual, and we used the intercept as a summary measure of the average magnitude of the response of the individual. We emphasize that the magnitude is not the peak magnitude just after vaccination or infection but rather the magnitude at the mean timepoint for that time series, which is expected to be many years or decades after vaccination or infection.

In the controlled experimental pen studies, cross-bred gilts from a high-health farm were used. The gilts were 8–15 months of age at first vaccination and were seronegative or low seropositive to PCV2 and Mycoplasma hyopneumoniae (MHP). None of the gilts were PCV2 viremic at first vaccination. Gilts were housed in gestation pens and in farrowing crates. Cross fostering in the lactation study was allowed only before Day 0.
In the field study, sows and gilts from two commercial farms in the USA were enrolled. The herds were of high health status. No clinical signs of PCV2 or MHP were present in the herds at the time of vaccination. All parities were represented at both sites. Sows/gilts were housed in conventional gestation crates (Site A) or gestation pens (Site B) and in farrowing crates for both sites. The piglets were kept with their birth sow throughout the study when possible. Cross fostering was permitted for animal welfare reasons only.
Farm A was a 1300-sow farrow-to-wean closed/internal multiplication facility, while farm B was a 600-sow farrow-to-wean facility. Both sites vaccinated gilts/sows with a Rotavirus, Clostridium perfringens Type C and Escherichia coli bacterin toxoid two weeks prior to their anticipated farrowing dates. Average production variables are shown in Table 3.

The antigen adsorption capacity of alhydrogel and nanoalum was analysed via SDS-PAGE using formaldehyde-inactivated C. botulinum Type D toxoid (OBP, SA) as a model antigen. The BoNT D toxoid vaccine was formulated with (10% v/v) either alhydrogel or nanoalum and stirred for 24 h at RT. The formulation was sampled at 0 h and 24 h; the samples were centrifuged at 13,000× g, 10 min, 4 °C. The supernatant was analysed on a 7.5% non-reducing SDS-PAGE according to the method described by Laemmli, 1970 [16 (link)]. The inactivated BoNT D toxoid was included as a reference (initial sample), and the same result would be expected for BoNT C.