Recombinant vaccine against hepatitis E: duration of protective immunity in rhesus macaques

Hepatitis E virus (HEV) is an enterically transmitted virus that causes acute hepatitis. A candidate vaccine containing recombinant HEV capsid protein (56kDa) expressed in insect cells was shown previously to be highly immunogenic when administered in two doses to rhesus monkeys and to protect them from hepatitis E when challenged with a large intravenous dose of homologous or heterologous HEV. In the present study, the effect of a third dose of the same vaccine lot was evaluated and more importantly, the duration of protection following two doses was determined. Rhesus monkeys vaccinated twice with the 56kDa capsid protein were challenged with homologous virus 6 or 12 months after the last vaccination: 3 of 4 monkeys challenged 6 or 12 months later, respectively were protected against viral hepatitis. Similarly, all four of the rhesus monkeys given a third dose of vaccine 1 month prior to challenge were protected against hepatitis. In contrast, all four monkeys given placebo developed hepatitis following challenge. In summary, two doses of HEV vaccine partially protected rhesus monkeys from hepatitis E following intravenous challenge 6 or 12 months after vaccination.     

Rabies virus pathogenesis in relationship to intervention with inactivated and attenuated rabies vaccines

Despite progress in vaccine development in the past century the mechanisms behind immune responses elicited by rabies biologics or via natural infection remain largely unknown. In this study, we compared protection elicited by standard, early, or delayed prophylaxis with a reduced number of vaccine doses using inactivated and live-attenuated vaccines. Two-month-old Syrian hamsters, 4-week-old ICR mice or adult rhesus macaques were inoculated with canine rabies virus variants. Thereafter, prophylaxis was initiated 6 h, 1, 2, 3, 4, 5, 6 or 7 days post-exposure (p.e.). One or several doses of inactivated (HDCV), or reverse genetically attenuated (live), or gamma-irradiated (inactivated)-ERAG333 vaccines were administered intramuscularly. The dynamics of virus spread were measured over time in the rodent models. Rabies virus reached the spinal cord at day 4 and brain at day 6 p.e. All hamsters succumbed in groups in which live ERAG333 was delayed until days 5 and 6 p.e. However, 78%, 44%, 56% and 22% of hamsters survived when one dose of live ERAG333 was administered 6 h, 1, 2, 3, and 4 days p.e., respectively. Similarly, 67% survived when inactivated ERAG333 was administered at 24 h p.e. All hamsters succumbed when standard prophylaxis (the Essen regimen) was delayed until days 3–6, but 67% and 33% of hamsters survived when PEP began 1 or 2 days p.e., respectively. Macaques were protected by one dose of attenuated ERAG333 at 24 h p.e. The highly attenuated (live) and inactivated ERAG333 vaccines elicited potent protective immune responses, even when prophylaxis initiation was delayed. When 2–5 doses of commercial vaccine and HRIG were administered according to the Essen scheme, 89–100% of the animals survived. Reduced vaccine schedules provided efficacious intervention, regardless of the total number of vaccine doses administered.     

Respiratory tract immunization of non-human primates with a Newcastle disease virus-vectored vaccine candidate against Ebola virus elicits a neutralizing antibody response

We previously developed a respiratory tract vaccine candidate against Ebola virus (EBOV) based on human parainfluenza virus type 3 (HPIV3), a respiratory paramyxovirus, expressing the EBOV GP envelope protein (HPIV3/GP) from an added gene. Two doses of this vaccine candidate delivered by the intranasal and intratracheal route protected monkeys against intraperitoneal challenge with EBOV; however, concerns exist that the vaccine candidate may have reduced immunogenicity in the adult human population due to pre-existing immunity against HPIV3. Here we developed a new vaccine candidate (NDV/GP) based on Newcastle disease virus (NDV), an avian paramyxovirus that is antigenically distinct from human viral pathogens and is highly attenuated in monkeys. Following one intranasal and intratracheal inoculation of Rhesus monkeys with NDV/GP, titers of EBOV-specific antibodies in respiratory tract secretions and serum samples determined by ELISA, as well as serum EBOV-neutralizing antibodies, were undetectable or low compared to those induced by HPIV3/GP. A second immunization resulted in a substantial boost in serum IgG ELISA titers, yet the titers remained lower than those induced by a second dose of HPIV3/GP. In contrast, the ELISA IgA titers in respiratory tract secretions and, more importantly, the serum EBOV-neutralizing antibody titers were equal to those induced after the second dose of HPIV3/GP. These data suggest that NDV/GP can be effective for immunization against EBOV alone, or in combination with either HPIV3/GP or another vaccine platform in a heterologous prime-boost regimen.     

A dengue virus serotype-1 DNA vaccine induces virus neutralizing antibodies and provides protection from viral challenge in Aotus monkeys

A DNA vaccine that expresses the premembrane/membrane (prM) and envelope (E) genes of dengue virus serotype-1 was tested for immunogenicity and protection against dengue-1 virus challenge in Aotus nancymae monkeys. The vaccine, in 1 mg doses, was administered intradermally (i.d.) to three monkeys and intramuscularly (i.m.) to three others. For controls, a 1 mg dose of vector DNA was administered i.d. to two monkeys and i.m. to one. All animals were primed and then boosted at one and five months post priming. Sera were collected monthly and analyzed for dengue-1 antibodies by enzyme linked immunosorbent assay (ELISA) and plaque reduction neutralization test (PRNT). Dengue-1 antibodies were detectable in the sera from i.d. and i.m. vaccine inoculated animals one month after the first boost and peaked one month after the second boost. The antibody levels from sera of animals that received the vaccine via the i.d. route were twice those from sera of animals that received the vaccine via the i.m. route. Six months after the second boost all inoculated and two naive monkeys were challenged with 1.25x10(4) plaque forming units (PFU) of dengue-1 virus. Two vaccine immunized animals were protected from viremia while the others showed a reduction in viremia. The mean days of viremia were 1 and 1.3 for the animals that were immunized with the vaccine via the i.d. or i.m. route, respectively vs 4 and 2 mean days of viremia in the animals inoculated with control DNA. Naive animals were viremic for an average of 4 days. All of the three control monkeys that received control DNA inoculum by either the i.d. or i.m. route had an intermittent viremia pattern with one or more negative days interspersed between the positive days. This pattern was not observed in any of the vaccine recipients or the na?ve control monkeys. These results demonstrate that DNA immunization is a promising approach for the development of dengue vaccines and that A. nancymae monkeys are suitable for dengue vaccine trials.     

The effect of mucoadhesive excipient on the nasal retention time of and the antibody responses induced by an intranasal influenza vaccine

IntroductionRecently, we reported that intranasal vaccination of humans with whole inactivated influenza vaccine in the absence of mucosal adjuvant induced neutralizing antibody responses in the serum and nasal mucus. The mucoadhesive excipient carboxy-vinyl polymer (CVP) increases the viscosity and therefore mucoadhesiveness of intranasal medicaments and is an authorized excipient in Japan. In the present study, we analyzed the effect of adding CVP on intranasal whole inactivated influenza vaccine antigen dynamics and antibody responses.MethodsMice and nonhuman primates (NHPs) were intranasally administered the [¹⁸F]-radiolabeled vaccine and subjected to positron emission tomography analysis for 6 h. Dendritic cells were stimulated in vitro with the vaccine mixed with or without a mucosal adjuvant (Ampligen) and/or CVP, after which the tumor necrosis factor (TNF)-α and interferon (IFN)-β levels in the supernatants were measured. Cynomolgus monkeys were immunized intranasally with the vaccine mixed with Ampligen and/or CVP and their vaccine-specific serum IgG and IgA titers were measured on days 0 and 33.ResultsThe vaccine was retained significantly longer in the nasal cavity of both mice and NHPs when it was delivered with CVP rather than PBS. Accumulation of the radiolabeled vaccine in the central nervous system was not detected in either model regardless of whether CVP was used. CVP only very weakly increased the TNF-α production of vaccine-stimulated dendritic cells. IFN-β production was not observed regardless of the presence or absence of CVP. CVP increased the vaccine-specific IgA antibody responses of the intranasally vaccinated cynomolgus macaques.ConclusionCVP increased intranasal retention of whole inactivated influenza vaccine, did not promote antigen redirection to the central nervous system, and improved mucosal antibody responses. The mechanism probably relates to its mucoadhesive properties rather than its ability to directly stimulate the immune system. Intranasal vaccines with CVP may be a promising candidate vaccine formulation for humans.     

Effective protection of monkeys against death from street virus by post-exposure administration of tissue-culture rabies vaccine

Three series of experiments on rabies vaccines were carried out on rhesus monkeys using suckling-mouse-brain vaccine, rabbit-brain vaccine, duck-embryo vaccine, and purified, concentrated tissue-culture vaccine. The latter was prepared in a human diploid cell strain and inactivated with β-propiolactone, and consisted of tissue-culture fluid concentrated 200-fold with a final infectivity titre of 10^9.8 plaque-forming units per ml before inactivation. In the first two series of experiments, several vaccines were tested for relative immunogenicity on a pre-exposure basis. In the third series, a successful model was developed in which a single inoculation of the tissue-culture vaccine administered after exposure to rabies virus, with or without accompanying standard doses of antirabies serum, was evaluated as a method of prevention. A single dose of the tissue-culture vaccine protected 7 out of 8 monkeys from death by street virus. Homologous or heterologous antirabies serum alone gave poor results. The results indicate great promise for prophylaxis in man with one dose, or perhaps a few doses, of highly concentrated, purified tissue-culture vaccine.     

Immunogenicity of hepatitis B vaccine among hemodialysis patients: effect of revaccination of non-responders and duration of protection

Background

Hepatitis B vaccination is recommended for patients on hemodialysis, however, seroprotection after a primary vaccine series is suboptimum. Limited data are available on the effect of revaccination of non-responders and on persistence of immunity in this population.

Methods

Hepatitis B vaccine (40 μg/dose) was given to 77 susceptible patients on hemodialysis (0, 1, and 6 month schedule). Levels of hepatitis B surface antibody (anti-HBs) were tested ≥28 days after the third dose was administered, and non-responders revaccinated with an additional 3-dose series. Vaccine responders (anti-HBs ≥10 mIU/mL) were re-tested every 6 months and booster doses given as needed. Kaplan-Meier survival curve was used to estimate the probability of maintaining protective antibody level. Cox-proportional hazards models were used to assess the association between time to loss of protective antibody levels and certain explanatory variables.

Results

Overall primary vaccine-induced response was 79.2% (95% CI 68.2%, 87.3%), including 49/77 (63.6%; 95% CI 51.8%, 74.7%) patients who received the initial primary hepatitis B vaccine series and 12/21 (57.1%; 95% CI 34.4%, 77.4%) non-responders who were revaccinated with an additional series. Among weak responders (anti-HBs level 10.0-99.9 mIU/mL), protective antibody levels persisted in 44% for 12 months post-vaccination; whereas among strong responders (anti-HBs level ≥100 mIU/mL), protective antibody levels persisted in 92% for 12 months, and 68% for 24 months post-vaccination. A weak post-vaccination response increased the risk of losing protective antibody levels (adjusted hazard ratio, 9.7; 95% confidence interval, 3.5-28.5; p < 0.0001).

Conclusion

Revaccinating patients undergoing hemodialysis who do not respond to a primary vaccine series substantially increases the pool of protected patients. The threshold for defining hepatitis B vaccine-induced immunity should be revisited in this patient population to maximize the duration of protection.     

Intramuscular immunization with a vesicular stomatitis virus recombinant expressing the influenza hemagglutinin provides post-exposure protection against lethal influenza challenge

Vaccines currently licensed for the prevention of seasonal influenza induce antibodies against the influenza hemagglutinin (HA) and neuraminidase (NA) contained in the vaccine preparation but require at least 2 weeks after immunization for the development of protective immunity. These vaccines do not induce protective responses quickly enough to blunt the effects of infection when administered after exposure. We have developed a novel vaccine based on recombinant vesicular stomatitis virus which expresses the influenza hemagglutinin (rVSV HA) and protects mice from lethal influenza challenge when the vaccine is administered intramuscularly at least 24 h after delivery of the influenza challenge virus. To our knowledge this is the first vaccine that effectively protects animals from lethal influenza challenge when delivered by a systemic route after influenza exposure has occurred. The induction of HA-specific immune responses by the vaccine is necessary for full protection from challenge, because animals immunized with an empty rVSV vector were not protected equally. Our results are consistent with a model in which vaccination induces an immediate antiviral cytokine response, followed by development of humoral and cellular immune responses which act to reduce pulmonary viral loads and accelerate recovery. Consistent with this model, mice vaccinated with the specific vaccine rVSV HA had high levels of IFN-α in the serum by 24 h after challenge/vaccination, developed serum neutralizing Ab to influenza 2 days prior to control animals, and had detectable anti-HA CD8 T cells present in the peripheral blood 3 days prior to control mice.     

Protective effect and the therapeutic index of indralin in juvenile rhesus monkeys

The radioprotective effect of indralin in rhesus monkeys was examined over 60 d following gamma irradiation. Male and female rhesus macaques (Macaca mulatta) 2–3-years-old and weighing 2.1–3.5 kg were used. Animals were exposed to total-body gamma irradiation from ⁶⁰Co at a dose of 6.8 Gy (lethal dose, 100% lethality over 30 days). Indralin (40–120 mg kg^–1) was administered intramuscularly 5 min prior to radiation exposure. Indralin taken at a dose of 120 mg kg^–1 protected five out of six monkeys (compared with the radiation control group, in which all 10 animals died). The average effective dose of indralin in the monkeys exposed to gamma irradiation for 30 min was equal to 77.3 (63.3–94.3) mg kg^–1, and the maximum tolerated dose of indralin administered to monkeys was 800 mg kg^–1. Indralin reduced radiation-induced injuries in macaques, thus resulting in a less severe course of acute radiation syndrome. Delayed and less pronounced manifestation of the haemorrhagic syndrome of the disease, and milder forms of both leukopenia and anaemia were also noted. The therapeutic index for indralin, expressed as the ratio of the maximum tolerated dose to the average effective dose_, was equal to 10. Therefore, indralin has a significant radioprotective effect against radiation and has a high therapeutic index in rhesus monkeys.     

Protective effects of high-potency FMDV O1 Manisa monovalent vaccine in cattle challenged with FMDV O/SKR/2010 at 7 or 4 days post vaccination

Serotype O foot-and-mouth disease (FMD) virus belonging to the SEA topotype continues to be a significant problem in the Eastern Asia region, with outbreaks in Japan and South Korea resulting in the culling of over 3.5 million cattle and pigs in recent years. High-potency O1 Manisa vaccine was previously shown to provide protection in cattle 21 days post vaccination (dpv) following challenge with a representative virus, O/SKR/2010. This study tested the ability of the O1 Manisa vaccine to protect cattle from infection and disease with the O/SKR/2010 virus within just 4 or 7 days post vaccination. The vaccine protected 50% of cattle from clinical disease when administered 7 days prior to challenge, but was not protective with just 4 days between vaccination and challenge. Viraemia was significantly reduced in animals challenged 7 dpv but not 4 dpv, compared to unvaccinated controls, however, there were no effects on the level of virus detected in nasal and oral secretions regardless of vaccination time. The level of neutralising antibodies detected in cattle challenged 7 dpv correlated with protection from clinical disease. All animals seroconverted to FMDV non-structural proteins, suggesting no sterile protection. An equal number of animals became persistently infected in both vaccine groups. The results indicated that high-potency O1 Manisa vaccine administered just 7 days prior to challenge should provide partial protection of cattle if an outbreak of O/SKR/2010, or related viruses, occurs, and would be useful to limit spread of FMDV when used in conjunction with other control measures.     

Challenge studies in Rhesus monkeys immunized with candidate hepatitis E vaccines: DNA,DNA-prime-protein-boost and DNA-protein encapsulated in liposomes

Complete ORF2 gene (1983 bp) of hepatitis E virus (HEV) and the 450 bp region within ORF2 containing neutralizing epitope (NE) cloned in pVAX1 and corresponding proteins expressed in baculovirus and prokaryotic systems respectively were evaluated as vaccine candidates. Two doses of liposome encapsulated DNA plus corresponding protein with both ORF2 and NE regions (Lipo-ORF2-DP and Lipo-NE-DP) showed 100% seroconversion and comparable anti-HEV titres in Swiss albino mice. These vaccine candidates were further evaluated as DNA, DNA-prime-protein-boost (DPPB) and liposome formulations in Rhesus monkeys. Monkeys receiving ORF2/NE DNA seroconverted after fourth dose while those immunized employing ORF2-DPPB format seroconverted at 7 weeks post third dose. In view of the delayed weak antibody response, these monkeys were not challenged. Though Lipo-ORF2-DP was immunogenic, 2 of the 4 monkeys developed HEV infection following homologous virus challenge of 100 Monkey Infectious Dose₅₀. Both monkeys immunized with Lipo-NE-DP and 1 of the 2 monkeys immunized with NE-DPPB showed complete protection, the second monkey being protected from hepatitis with limited viral replication. Irrespective of the type of immunogen, all challenged monkeys were protected from hepatitis. The results document Lipo-NE-DP to be a promising vaccine candidate needing further evaluation.     

Immunization of rhesus monkeys with a mucosal prime, parenteral boost strategy protects against infection with Helicobacter pylori.

Rhesus monkeys were immunized with recombinant Helicobacter pylori urease vaccine given solely by the parenteral route or preceded by a priming dose given by the oral route. Two groups of monkeys received parenteral urease with either a synthetic glycolipid adjuvant (Bay) or aluminum hydroxide (alum) as adjuvants. A third group of monkeys received a priming dose of oral urease given with the mucosal adjuvant LT (Escherichia coli heat labile enterotoxin), followed by parenterally administered booster doses of urease adsorbed to alum. Monkeys receiving placebo served as controls. The monkeys received a total of 4 doses of vaccine with the first 3 doses given every 3 weeks and the last booster dose administered 14 weeks later. The monkeys were challenged orally with H. pylori one week after the last vaccine dose and euthanized 10 weeks after challenge, at which time, their stomachs were collected for determination of bacterial colonization and histopathology. Monkeys primed with the oral vaccine and boosted with the parenteral vaccine showed a statistically significant reduction in bacterial colonization when compared to sham-immunized control animals (P = 0.05; Wilcoxon rank sums test). Monkeys receiving parenteral only regimes of urease plus Bay or alum showed no difference in bacterial colonization compared with sham-immunized controls (P = 1.00 and P = 0.33, respectively). The mucosal prime-parenteral boost regime did not cause gastropathy. There was no difference in any of the 3 treatment groups with respect to gastric epithelial changes compared to control animals. There was also no difference in the type and extent of gastric inflammatory cell infiltrates between animals vaccinated by the mucosal prime-parenteral boost strategy and sham immunized controls. However, monkeys receiving the two parenteral-only regimens had slightly elevated gastritis scores.     

Enhanced pulmonary pathology associated with the use of formalin-inactivated respiratory syncytial virus vaccine in cotton rats is not a unique viral phenomenon

The specificity of viral antigens in the formalin-inactivated, alum-precipitated respiratory syncytial virus (FI-RSV) vaccine in augmenting the pulmonary inflammatory response was evaluated. Cotton rats were immunized with a FI-RSV vaccine derived from Vero cells, a monkey cell line, or HEp-2 cells, a human cell line. The FI-RSV/Vero and the FI-RSV/HEp-2 vaccines were prepared similarly to the original Lot-100 FI-RSV vaccine that was associated with enhanced disease in the mid-1960s field trials. Each vaccine was administered intramuscularly at various doses and intervals. At I, 4 or 7 weeks after the last vaccine dose, cotton rats were challenged with 10⁶ plaque-forming units of live RSV grown in HEp-2 cells. For controls, FI-parainfluenza, FI-HEp-2 and alum vaccines, and live RSV primary infection were used. For measuring virus replication and histopathology, lungs were harvested at 4 and 8 days postchallenge. A dose-response relationship to vaccine dose was observed for ELISA, neutralizing and antifusion antibodies. All animals given three doses or two of the higher doses of FI-RSV/Vero vaccine developed significant neutralizing antibody, were protected against pulmonary virus replication and had similar low levels of histopathology compared with live RSV and controls. Two immunizations of the lowest dose of FI-RSV/Vero vaccine did not induce neutralizing antibody, did not provide protection of the lung against RSV and did not enhance the pulmonary cellular response. However, FI-RSV/HEp-2 vaccine was associated with significant enhanced pulmonary histopathology despite inducing high titres of neutralizing antibody and protecting the lungs against RSV infection. FI-HEp-2 vaccine was also associated with significantly enhanced pulmonary histopathology. Under the conditions tested, non-viral antigens played a major role in the augmentation of the pulmonary cellular response in cotton rats that were given FI-RSV/HEp-2 vaccine.     

Similarity of Bisphenol A Pharmacokinetics in Rhesus Monkeys and Mice: Relevance for Human Exposure

Objective

Daily adult human exposure to bisphenol A (BPA) has been estimated at < 1 μg/kg, with virtually complete first-pass conjugation in the liver in primates but not in mice. We measured unconjugated and conjugated BPA levels in serum from adult female rhesus monkeys and adult female mice after oral administration of BPA and compared findings in mice and monkeys with prior published data in women.

Methods

Eleven adult female rhesus macaques were fed 400 μg/kg deuterated BPA (dBPA) daily for 7 days. Levels of serum dBPA were analyzed by isotope-dilution liquid chromatography–mass spectrometry (0.2 ng/mL limit of quantitation) over 24 hr on day 1 and on day 7. The same dose of BPA was fed to adult female CD-1 mice; other female mice were administered ³H-BPA at doses ranging from 2 to 100,000 μg/kg.

Results

In monkeys, the maximum unconjugated serum dBPA concentration of 4 ng/mL was reached 1 hr after feeding and declined to low levels by 24 hr, with no significant bioaccumulation after seven daily doses. Mice and monkeys cleared unconjugated serum BPA at virtually identical rates. We observed a linear (proportional) relationship between administered dose and serum BPA in mice.

Conclusions

BPA pharmacokinetics in women, female monkeys, and mice is very similar. By comparison with approximately 2 ng/mL unconjugated serum BPA reported in multiple human studies, the average 24-hr unconjugated serum BPA concentration of 0.5 ng/mL in both monkeys and mice after a 400 μg/kg oral dose suggests that total daily human exposure is via multiple routes and is much higher than previously assumed.     

Enteric administration of a live attenuated measles vaccine does not induce protective immunity in a macaque model

To test the option of oral vaccination with a live attenuated measles vaccine (LAV), we have evaluated the potential of an orally administered enteric-coated tablet containing a candidate LAV (strain Leningrad-16, MV-L16). To this end three groups of two cynomolgus macaques each were vaccinated via different routes with 10(3.8) TCID(50) MV-L16 vaccine: intramuscularly (i.m.), intraintestinally (i.i.) upon laparotomy and via enteric-coated tablets. Upon vaccination, MV-L16 could only be isolated from one of the i.m.-vaccinated monkeys and not from any of the other five. Both the i.m.-infected monkeys and one of the i.i.-infected monkeys developed a MV-specific serum antibody response. Also, MV-specific CD8(+) IFN gamma-producing T cells could be demonstrated in all three monkeys that had seroconverted. Upon challenge with wild-type MV 1 year after vaccination, only these three monkeys proved to be protected. These data do not support the viability of the concept of oral vaccination with LAVs.     

Immunization of rhesus monkeys with a mucosal prime, parenteral boost strategy protects against infection with Helicobacter pylori

Rhesus monkeys were immunized with recombinant Helicobacter pylori urease vaccine given solely by the parenteral route or preceded by a priming dose given by the oral route. Two groups of monkeys received parenteral urease with either a synthetic glycolipid adjuvant (Bay) or aluminum hydroxide (alum) as adjuvants. A third group of monkeys received a priming dose of oral urease given with the mucosal adjuvant LT (Escherichia coli heat labile enterotoxin), followed by parenterally administered booster doses of urease adsorbed to alum. Monkeys receiving placebo served as controls. The monkeys received a total of 4 doses of vaccine with the first 3 doses given every 3 weeks and the last booster dose administered 14 weeks later. The monkeys were challenged orally with H. pylori one week after the last vaccine dose and euthanized 10 weeks after challenge, at which time, their stomachs were collected for determination of bacterial colonization and histopathology. Monkeys primed with the oral vaccine and boosted with the parenteral vaccine showed a statistically significant reduction in bacterial colonization when compared to sham-immunized control animals (P=0.05; Wilcoxon rank sums test). Monkeys receiving parenteral only regimes of urease plus Bay or alum showed no difference in bacterial colonization compared with sham-immunized controls (P=1.00 and P=0.33, respectively). The mucosal prime–parenteral boost regime did not cause gastropathy. There was no difference in any of the 3 treatment groups with respect to gastric epithelial changes compared to control animals. There was also no difference in the type and extent of gastric inflammatory cell infiltrates between animals vaccinated by the mucosal prime–parenteral boost strategy and sham immunized controls. However, monkeys receiving the two parenteral-only regimens had slightly elevated gastritis scores.     

Immunization with recombinant Helicobacter pylori urease decreases colonization levels following experimental infection of rhesus monkeys

Rhesus monkeys, naturally colonized with H. pylori as indicated by culture and histology were immunized with either 40 mg recombinant H. pylori urease administered orally together with 25 microg Escherichia coli heat-labile enterotoxin (LT) or immunized with LT alone. An initial 6 doses were administered over an 8 week period. All five vaccinated monkeys had a greater than two-fold rise in urease-specific serum IgG and IgA level and urease-specific salivary IgA was induced in 3 of 5 vaccinated animals after 6 or 7 doses of vaccine. Vaccination had no measurable therapeutic effect on H. pylori colonization. H. pylori was eradicated from these monkeys with a course of antimicrobials plus omeprazole, a 7th vaccine dose was given (10 months after the 6th dose) and they were rechallenged with H. pylori. Necropsy was performed 23 weeks after rechallenge and H. pylori colonization was determined by histological examination of 12 individual gastric sites. A significant reduction in colonization (p < or = 0.0001; Friedman's analysis of variance) was found in the vaccinated animals. Histopathologic examination of necropsy tissues also revealed a trend towards reduced gastritis and epithelial alterations in the vaccinated group compared to animals receiving LT alone. This study provides the first evidence for effective vaccination of nonhuman primates against H. pylori, and preliminary evidence that a reduction in bacterial density attributable to immunization may lessen gastric inflammation.     

Induction of protective immunity in swine by immunization with live attenuated recombinant pseudorabies virus expressing the capsid precursor encoding regions of foot-and-mouth disease virus

Foot-and-mouth disease (FMD) causes morbidity to livestock and serious economic consequences to its associated industry and therefore it is necessary to develop a safe and efficient vaccine to prevent or control this disease. A recombinant live attenuated virus vaccine, designated PRV-P1, was generated by insertion of an expression cassette containing CMV promoter, FMDV P1 gene and SV 40 poly-A into the gG gene region of a live attenuated pseudorabies virus vaccine strain (TK⁻/gG⁻/LacZ⁺). To determine the induction of protective immunity, 16 FMDV and PRV seronegative white swine were randomly divided into four groups and immunized intramuscularly. The parental virus (TK⁻/gG⁻/LacZ⁺) was injected into three pigs, the recombinant virus PRV-P1 into five pigs and commercial FMD-inactivated vaccine into five pigs, with PBS (negative control) into three pigs. All animals were immunized again 4 weeks later to boost the immune response and challenged with virulent type O FMDV O/ES/2001 strain 4 weeks after the second immunization. Results showed PRV-P1 vaccinated pigs induced high-level neutralizing antibody response to both FMDV and PRV, and strong CTL response against FMD antigen activation. Three of five pigs were completely protected against challenge with FMDV, one pig minimally protected and the other one had increased protection but not complete. However, one pig vaccinated with commercial FMD vaccine developed constant pyrexia. Average levels of antibodies against non-structural 3ABC proteins were significantly lower and efficacy on inhibition of FMDV replication was much increased in swine vaccinated with PRV-P1 than those immunized with commercial FMD vaccine after FMDV challenge. Our results showed that the recombinant PRV-P1 can induce not only humoral and cell-mediated immune responses but also partial protection against FMDV challenge, making it a good candidate for future development of the FMD vaccine.     

Lack of viraemia and limited antibody response of dengue virus immune rhesus monkeys after vaccination with DEN-2/S-1 vaccine

A dengue-2 live virus vaccine was tested in monkeys immune to heterologous dengue serotypes to determine if, as with wild DEN-2 virus, antibody-enhanced viraemia and seroconversion would occur. A low dose of 900 plaque-forming units (PFU) of the DEN-2/S-1 vaccine virus was inoculated subcutaneously into rhesus monkeys six months after they had received wild DEN-1, DEN-2 or DEN-3 viruses, and into non-immune monkeys. As previously reported for non-immune monkeys, there was little, if any, detectable vaccine viraemia in any of the groups of monkeys. There was no difference in seroconversion between the dengue heterologously immune (3/6) and non-immune (1/3) monkeys. These data indicated that (i) the vaccine virus may differ from the parent virus in the ability to complex with heterologous antibody and, thus, in the ability to infect Fc receptor bearing cells in monkeys; (ii) 10(3) PFU of vaccine virus is approximately the 50% infectious dose in monkeys as measured by seroconversion.