Protective immunity induced by oral immunization with a rotavirus DNA vaccine encapsulated in microparticles

Abstract

DNA vaccines are usually given by intramuscular injection or by gene gun delivery of DNA-coated particles into the epidermis. Induction of mucosal immunity by targeting DNA vaccines to mucosal surfaces may offer advantages, and an oral vaccine could be effective for controlling infections of the gut mucosa. In a murine model, we obtained protective immune responses after oral immunization with a rotavirus VP6 DNA vaccine encapsulated in poly(lactide-coglycolide) (PLG) microparticles. One dose of vaccine given to BALB/c mice elicited both rotavirus-specific serum antibodies and intestinal immunoglobulin A (IgA). After challenge at 12 weeks postimmunization with homologous rotavirus, fecal rotavirus antigen was significantly reduced compared with controls. Earlier and higher fecal rotavirus-specific IgA responses were noted during the peak period of viral shedding, suggesting that protection was due to specific mucosal immune responses. The results that we obtained with PLG-encapsulated rotavirus VP6 DNA are the first to demonstrate protection against an infectious agent elicited after oral administration of a DNA vaccine.

FIG. 1

Diagram of the pCMVIA vector and the virus cDNA insert. SV 40 Ori, simian virus 40 origin of replication; CMV Pro, CMV immediate-early promoter, intron A (largest CMV intron); BGH, bovine growth hormone gene (provides polyadenylation [pA] signals).

FIG. 2

Rotavirus-specific serum ELISA antibodies mice from BALB/c mice that had been orally inoculated (by gavage) with PLG-encapsulated VP6 DNA vaccine (n = 13) or with PLG-encapsulated control plasmid DNA (n = 10). Serum was collected at the times indicated and tested by an ELISA for total antibody (IgG, IgM, and IgA) every 2 weeks for 12 weeks. Results are expressed as geometric mean titers ± standard error.

FIG. 3

Protection against EDIM rotavirus challenge in BALB/c mice. Mice were challenged with 100 ID₅₀ of virus per mouse 12 weeks after receiving PLG-encapsulated VP6 DNA vaccine (n = 13) or PLG-encapsulated control plasmid DNA (n = 10) by oral gavage. Virus shedding in feces, determined by an ELISA for detecting rotavirus antigen, is given as A₄₉₂ ± standard deviation. A positive test is one in which the A₄₉₂ is ≥0.1. There were significant differences (P < 0.0002) in viral shedding between the mice receiving the plasmid encoding VP6 and the plasmid control on the days indicated by an asterisk.

FIG. 4

ELISA for rotavirus-specific IgA in stool suspensions from mice that had been orally inoculated (by gavage) with PLG-encapsulated VP6 DNA vaccine (n = 13) or with PLG-encapsulated control plasmid DNA (n = 10). The stools were diluted 1:80 (wt/vol) in PBS. Results are expressed as A₄₉₂ ± standard deviation. Values of >0.1 are considered positive for IgA. There were significant differences (P < 0.004) in fecal IgA values between the mice receiving the plasmid encoding VP6 and the plasmid control on the days indicated by an asterisk.

FIG. 5

ELISA for rotavirus-specific IgA in stool suspensions from mice that had been orally inoculated (by gavage) with PLG-encapsulated VP6 DNA vaccine (n = 13) or with PLG-encapsulated control plasmid DNA (n = 10) and challenged with EDIM rotavirus. The stools were diluted 1:80 (wt/vol) in PBS. Results are expressed as A₄₉₂ ± standard deviation. An A₄₉₂ of >0.1 is considered positive for IgA. There were significant differences (P < 0.01) in fecal IgA values between the mice receiving the plasmid encoding VP6 and the plasmid control on the days indicated by an asterisk.