Table 1.

Post-immunization hemagglutination inhibition (HI) antibody titers and lung viral titers 3 days post-challenge with a homologous H5N1 reassortant virus in vaccinated aged mice.

Group	Immunostimulation or Pre-immunization	Immunization	HI titers (Geometric mean)	Lung virus titer (Log10 EID50/ml ±SD)
Vector Control	HAd-ΔE1E3	HAd-ΔE1E3	≤10	6.50±0.10
Without immunostimulation	PBS	HAd-HA-NP	≤10	4.60±0.50
With Immunostimulation	HAd-Mbd2	HAd-HA-NP	50	3.10± 0.30

Aged mice (18 month-old) were inoculated intramuscularly (i.m.) either with PBS or with 1 × 10⁸ plaque-forming units (pfu) of HAd-Mbd2 [adenovirus vector expressing murine ß-defensin 2 (Mbd2)]. Seven days later, animals were immunized with 1 × 10⁸ pfu of HAd-HA-NP [adenovirus vector expressing hemagglutinin (HA) and nucleoprotein (NP) of a H5N1 influenza virus]. Animals that were similarly inoculated with 1 × 10⁸ pfu of HAdΔE1E3 on days 0 and 7 served as negative controls. Serum samples were obtained from all animals four weeks after the last inoculation and analyzed for A/Vietnam/1203/2004–specific hemagglutination inhibition (HI) titers by HI assay using 1% horse red blood cells. The titers are shown as geometric mean values. Four weeks after the last immunization, mice from each group were challenged with 100 MID50 (50% mouse infectious dose) of reverse genetics-derived A/Puerto Rico/8/1934 (H1N1) [PR8] containing HA and NA gene fragments of A/Vietnam/1203/04 (H5N1) [VNH5N1-PR8/CDC-RG]. Three days post-challenge, mice were euthanized, and the lungs were collected. The lung viral titers were determined to evaluate the protective efficacy of the vaccine. The detection limit of the lung viral titer was ≤1.5 Log10EID50 /ml. EID, egg infectious dose.