Efficacy of a Replikin Peptide Vaccine Against Low-Pathogenicity Avian Influenza H5 Virus

Abstract

In this study, the sequence of the H5 and PB1 genes of the low-pathogenic avian influenza virus (LPAI) A/Black Duck/NC/674-964/06 isolate were determined for replikin peptides and used to design and chemically synthesize a vaccine. The vaccine was used to immunize specific-pathogen-free (SPF) leghorn chickens held in Horsfall isolation units, by the upper respiratory route, at 1, 7, and 14 days of age. The birds were challenged at 28 days of age with 1 x 10(6) 50% embryo infective dose of the LPAI Black Duck/NC/674-964/06 H5N1 virus per bird. Oropharyngeal and cloacal swabs were collected at 2, 4, and 7 days postinoculation (PI) for virus detection by real-time RT-PCR. Serum was collected at 7, 14, and 21 days PI and examined for antibodies against avian influenza virus by the enzyme-linked immunosorbent assay and hemagglutination inhibition (HI) tests. Tissue samples for histopathology were collected from three birds per group at 3 days PI. The experimental design consisted of a negative control group (not vaccinated and not challenged) and a vaccinated group, a vaccinated and challenged group, and a positive control group (challenged only). None of the nonchallenged birds, the vaccinated birds, or the vaccinated and challenged birds showed overt clinical signs of disease during the study. A slight depression was observed in the nonvaccinated challenged birds on day 2 postchallenge. Although the numbers of birds per group are small, no shedding of the challenge virus was detected in the vaccinated and challenged birds, whereas oropharyngeal and cloacal shedding was detected in the nonvaccinated and challenged birds. HI antibodies were detected in the vaccinated and nonchallenged group as well as in the vaccinated and challenged group, but rising antibody titers, indicating infection with the LPAI challenge virus, were not detected. Rising HI titers were observed in the nonvaccinated and challenged group. In addition, no antibodies were detected in the nonchallenged birds. Noteworthy microscopic lesions were not observed in the vaccinated and challenged birds, whereas nonvaccinated-challenged birds had microscopic lesions consistent with infection with LPAI viruses. Taken together, these data indicate that a replikin peptide vaccine, specifically made against the H5N1 Black Duck/NC/674-964/06 isolate, and administered three times to the upper respiratory tract, was capable of protecting chickens from infection and from shedding of the homologous virus, which is extremely important because reduced virus shedding and transmission decreases the potential for H5 LPAI viruses to become HPAI viruses. The study is also important because it shows that the vaccine can be effectively mass-delivered to the upper respiratory tract.

Genomic Replikin Counts of Infectious Salmon Anemia Virus (ISAV) in Canada Exceed the Counts in Lethal Outbreaks in Norway, Chile, and Scotland. Real-Time Tracking of the Evolution of the ISAV Genome and the Resultant Replikins Solid Phase ISAV Vaccine Make ISAV Pandemic Prevention Possible.

“…The Replikin Count has been used to predict the relative lethality of shrimp Taura Syndrome Virus (TSV) isolates, the accuracy of the prediction confirmed in the linear relationship observed in the laboratory. Replikins TSV Synthetic Vaccines have been shown in the laboratory to block TSV infection in shrimp (12). Regional temporal cyclic expansion preceded by increased concentration of Replikins observed in other viruses (9) also may be occurring for ISAV, as seen in the Figure. The present data suggest that the marked decline in salmon yields in recent years in Canadian and U.S. waters (10,19), at least in part, may be related to ISAV.…”

“…The ISAV vaccine is similar in its basis to the oral Replikins TSV Vaccine, and to the TransFlu TM Vaccine which has been demonstrated to block H5N1 virus and its excretion by chickens (12). For the first time it may be possible to counter virus reservoir formation where mutation can occur to more lethal strains (12). The ReplikinsISAV TM Vaccine will be applied in the same way to counter reservoir formation in Canada and elsewhere.…”

Bogoch Replikins Pandemic Prevention: Increase of Strain-Specific Influenza Genomic Replikin Counts, Having Predicted Outbreaks and their Location Seven Times Consecutively, Up to Two Years in Advance, Provides Time for Prevention of Pandemics

“…Real-time Replikin analysis of the evolution of the virus genome identified both mutations and structural reorganization of the hemagglutinin and p B1 genes over several years before each outbreak. This information, together with the specific Replikin sequences so obtained, permitted solid-phase synthesis of Replikin vaccines in seven days, which blocked H5N1 in chickens (13). The information also now provides up to two years of time to thoroughly test and distribute vaccines to high risk individuals in the countries identified; thus for the first time, a quantitative genomic Replikins method to both predict initial outbreaks and to prevent the development of a pandemic.…”

“…The time taken for these changes to occur, encourages the consideration that prevention of a pandemic is possible for the first time. In contrast, the exact Replikin sequences revealed make it possible to produce in as few as 7 days an effective vaccine for the most recent mutations (13).…”

Prediction of specific virus outbreaks made from the increased concentration

“…However, with simultaneous increases in the Replikin Counts of each strain, outbreaks of H1N1 and H5N1 are now in progress in Mexico and Egypt repectively (2,41,44), and in preparation to meet this threatened combination, the authors have prepared the first completely synthetic TransFlu TM Replikins Vaccine against these two and the other common influenza strains. TransFlu TM was successful in blocking H5N1 (23) in the first of its continuing independent trials in the U.S. and elsewhere.…”