Antigen Localization Influences the Magnitude and Kinetics of Endogenous Adaptive Immune Response to Recombinant Salmonella Vaccines

ABSTRACT The use of recombinant attenuated Salmonella vaccine (RASV) strains is a promising strategy for presenting heterologous antigens to the mammalian immune system to induce both cellular and humoral immune responses. However, studies on RASV development differ on where heterologous antigens are expressed and localized within the bacterium, and it is unclear how antigen localization modulates the immune response. Previously, we exploited the plasmid-encoded toxin (Pet) autotransporter system for accumulation of heterologous antigens in cell culture supernatant. In the present study, this Pet system was used to express early secretory antigen 6 (ESAT-6), an immunodominant and diagnostic antigen from Mycobacterium tuberculosis, in Salmonella enterica serovar Typhimurium strain SL3261. Three strains were generated, whereby ESAT-6 was expressed as a cytoplasmic (SL3261/cyto), surface-bound (SL3261/surf), or secreted (SL3261/sec) antigen. Using these RASVs, the relationship between antigen localization and immunogenicity in infected C57BL/6 mice was systematically examined. Using purified antigen and specific tetramers, we showed that mice infected with the SL3261/surf or SL3261/sec strain generated large numbers of Th1 CD4+ ESAT-6+ splenic T cells compared to those of mice infected with SL3261/cyto. While all mice showed ESAT-6-specific antibody responses when infected with SL3261/surf or SL3261/sec, peak total serum IgG antibody titers were reached more rapidly in mice that received SL3261/sec. Thus, how antigen is localized after production within bacteria has a more marked effect on the antibody response than on the CD4+ T cell response, which might influence the chosen strategy to localize recombinant antigen in RASVs.

Zebra-borne neurotropic equid herpesvirus 1 meningoencephalitis in a Thomson’s gazelle (Eudorcas thomsonii)

We describe the histopathologic, immunohistochemical, and molecular features of a case of meningoencephalitis in a Thomson’s gazelle ( Eudorcas thomsonii) naturally infected with zebra-borne equid herpesvirus 1 (EHV-1) and the implications for the molecular detection of zebra-borne EHV-1. A 4-y-old female Thomson’s gazelle was submitted for postmortem examination; no gross abnormalities were noted except for meningeal congestion. Microscopic evaluation demonstrated multifocal nonsuppurative meningoencephalitis with intranuclear eosinophilic and amphophilic inclusion bodies and EHV-9 antigen in neurons. PCR demonstrated the presence of a herpesvirus with a nucleotide sequence 99–100% identical to the corresponding sequences of zebra-borne EHV-1 and of EHV-9 strains. To determine whether EHV-1 or EHV-9 was involved, a PCR with a specific primer set for EHV-9 ORF59/60 was used. The sequence was identical to that of 3 recognized zebra-borne EHV-1 strains and 91% similar to that of EHV-9. This isolate was designated as strain LM2014. The partial glycoprotein G ( gG) gene sequence of LM2014 was also identical to the sequence of 2 zebra-borne EHV-1 strains (T-529 isolated from an onager, 94-137 from a Thomson’s gazelle). The histologic lesions of encephalitis and antigen localization in this gazelle indicate prominent viral neurotropism, and lesions were very similar to those seen in EHV-1– and EHV-9–infected non-equid species. Histologic lesions caused by EHV-9 and zebra-borne EHV-1 are therefore indistinguishable.

FREQUENT PRESENCE OF PORCINE TESCHOVIRUS ANTIGENS IN VISCERAL AND LYMPHOID ORGANS OF NONSUPPURATIVE ENCEPHALITIC PIGS IN THE ENDEMIC FIELD SITUATION

This study characterized how porcine teschovirus (PTV) contributed to causing nonsuppurative encephalitis in the endemic field situation which was mostly subacute to chronic. Tissues from 57 encephalitic pigs, in the years 2005–2008, were randomly selected for detection of PTV antigens by immunohistochemistry. In only 3/57 (5.3%) pigs PTV signal was detected in brain cells of noninflammatory sites. On the other hand, PTV signals were present at significant rates in visceral and lymphoid organs. Spleen (33.3%), lung (24.6%) and kidney (14%) were among the higher, while those of lymphoid organs were 17.5%–26.3%, and intestines were 21.1%–24.6%. Nursery pigs not only comprised a bulk of the samples (n = 26), but also had a higher signal detection rate of 53.8% (by head) and average numbers of 2.65 positive organs per animal. Cellular tropism and antigen localization of PTVs are closely related to viremia, fecal and urinal sheddings, reinfection and recirculation of PTV within the herd.