Carlos Montero

Field studies and cost-effectiveness analysis of vaccination with Gavac against the cattle tick boophilus microplus

The control of tick infestations and the transmission of tick-borne diseases remains a challenge for the cattle industry in tropical and subtropical areas of the world. Traditional control methods have been only partially successful and the parasites continue to result in significant losses for the cattle industry. Recently, vaccines containing the recombinant Boophilus microplus gut antigen Bm86 have been developed. These vaccines have been shown to control tick infestations in the field. However, extensive field studies investigating the efficacy and cost-effectiveness of vaccination have not been reported and are needed to appraise the effect of this new approach for tick control. Here is reported the results of the application of Gavac in a field trial including more than 260,000 animals in Cuba. In this study the correlation between the antibody response to vaccination and the effect on ticks fertility is determined. Physiological status of the animals was found to affect the primary response to vaccination but not the antibody titers after revaccination. A cost-effectiveness analysis showed a 60% reduction in the number acaricide treatments, together with the control of tick infestations and transmission of babesiosis, which resulted in savings of

Control of ticks resistant to immunization with Bm86 in cattle vaccinated with the recombinant antigen Bm95 isolated from the cattle tick, Boophilus microplus.

23.4 animal-1 year-1. These results clearly demonstrate the advantage of vaccination and support the application of Gavac for tick control.

VACCINATION AGAINST TICKS (BOOPHILUS SPP.) : THE EXPERIENCE WITH THE BM86-BASED VACCINE GAVAC

The recombinant Bm86-containing vaccine Gavac(TM) against the cattle tick Boophilus microplus has proved its efficacy in a number of experiments, especially when combined with acaricides in an integrated manner. However, tick isolates such as the Argentinean strain A, show low susceptibility to this vaccine. In this paper we report on the isolation of the Bm95 gene from the B. microplus strain A, which was cloned and expressed in the yeast Pichia pastoris producing a glycosylated and particulated recombinant protein. This new antigen was effective against different tick strains in a pen trial, including the B. microplus strain A, resistant to vaccination with Bm86. A Bm95-based vaccine was used to protect cattle against tick infestations under production conditions, lowering the number of ticks on vaccinated animals and, therefore, reducing the frequency of acaricide treatments. The Bm95 antigen from strain A was able to protect against infestations with Bm86-sensitive and Bm86-resistant tick strains, thus suggesting that Bm95 could be a more universal antigen to protect cattle against infestations by B. microplus strains from different geographical areas.

Integrated control of acaricide-resistant Boophilus microplus populations on grazing cattle in Mexico using vaccination with Gavac and amidine treatments.

The control of tick infestations and the transmission of tick-borne diseases remain a challenge for the cattle industry in tropical and subtropical areas of the world. Traditional control methods have been only partially successful and the parasites continue to result in significant losses for the cattle industry. Recently, vaccines containing the recombinant B. microplus gut antigen Bm86 have been developed. Our vaccine formulation (Gavac, Heber Biotec S.A., Havana, Cuba) has been registered and is commercially available in Cuba, Colombia, Dominican Republic, Brazil and Mexico. In controlled pen trials, Gavac has been effective for the control of artificial infestations of B. annulatus, B. decoloratus and chemical-sensitive and resistant B. microplus strains from Australia, Africa, America and Iran. In controlled field trials in Cuba, Brazil, Argentina and Mexico, Gavac has shown a 55-100% efficacy in the control of B. microplus infestations in grazing cattle 12-36 weeks after the first vaccination. Field trials under production conditions have been conducted in Cuba, Colombia, Brazil and Mexico in pure and cross-bred cattle herds. The application of Gavac has increased the time between acaricide treatments by an average of 32 /-21 days (P = 0.0005) resulting in important savings for the cattle industry. In Cuba, a cost-effectiveness analysis was conducted in more than 260000 animals. The cost-effectiveness analysis showed a 60% reduction in the number of acaricide treatments, together with the control of tick infestations and transmission of babesiosis, which resulted in savings of 23.4 dollars animal(-1) year (-1). These results clearly demonstrate the advantage of vaccination and support the application of Gavac for the control of Boophilus spp. infestations.

Effect of particulation on the immunogenic and protective properties of the recombinant Bm86 antigen expressed in Pichia pastoris

Throughout most of the twentieth century, tick infestations on cattle have been controlled with chemical acaricides, typically administered by dipping or spraying. This approach can cause environmental and residue problems and has created a high incidence of acaricide resistance within tick populations in the field. Recently we developed a vaccine against Boophilus microplus employing a recombinant Bm86 antigen preparation (Gavac)™, (Heber Biotec S.A., Havana, Cuba) which has been shown to induce a protective response in vaccinated animals. Here we show for the first time under field conditions a near 100% control of B. microplus populations resistant to pyrethroids and organophosphates, by an integrated system employing vaccination with Gavac™ and amidine treatments. This method effectively controls tick infestations while reducing the number of chemical acaricide treatments and consequently the rise of B. microplus populations resistant to chemical acaricides.

The evaluation of yeast derivatives as adjuvants for the immune response to the Bm86 antigen in cattle

The recombinant Bm86 tick antigen expressed in Pichia pastoris is obtained in a highly particulated form, as a distinguish feature of this expression system. This particulated protein, the active principle of the recombinant vaccine Gavac against the cattle tick, have shown high immunogenic and protective properties, probably associated with its own characteristics. To evaluate the effects of particulation on the properties of Bm86, three groups of calves were immunized with particulated or non-particulated recombinant Bm86 and the anti-Bm86 antibody response determined. Animals were challenged with a controlled tick infestation and the protective capacities of both proteins assessed. Humoral immune response and protection in cattle vaccinated with the particulated antigen were higher. These experiments suggested that particulation of the Bm86 expressed in P. pastoris is an important feature for the protective properties of the antigen in vaccine preparations.

The bovine IFN-ω1 is biologically active and secreted at high levels in the yeast Pichia pastoris

BackgroundThe Gavac™ vaccine against the cattle tick Boophilus microplus has proven its efficacy in a large number of controlled and field experiments. However, this vaccine could be further improved by searching for new alternative adjuvants that would induce a stronger long-lasting immune response. We conducted several experiments to assay the adjuvant effect of fractions of the recombinant yeast Pichia pastoris in mouse and cattle models. In previous experiments, the combination of the yeast membrane with saponin was the most effective formulation in stimulating the humoral immune response in mice, eliciting a response higher than Montanide 888. The response was predominantly of the IgG1 isotype. Here, we evaluated the response in cattle and compared the results with that obtained in mice.ResultsBm86 on the membrane of P. pastoris plus saponin produced high antibody titers in cattle, though the protection level against tick infestations was lower when compared to Gavac™, probably due to a decrease in the IgG1/IgG2 ratio. The predictive value of the mouse model was studied through correlation analysis between the isotype levels in mice and the efficacy of formulations in cattle. Good correlation was established between the level of antibodies in mice and cattle, and between the amount of anti -Bm86 IgG1 in mice and the degree of protection in cattle.ConclusionMouse model have the potential to predict immunogenicity and efficacy of formulations in cattle. These results also support the use of the yeast expression system for recombinant vaccine formulations, enabling the prediction of more cost - effective formulations.

Reproductive and safety assessment of vaccination with gavac against the cattle tick (Boophilus microplus)

Abstract The gene coding for bIFN- ω 1 was isolated from bovine genomic DNA by polymerase chain reaction (PCR). Recombinant bIFN- ω 1 was expressed in the yeast Pichia pastoris and high levels of the recombinant protein (0.4 mg ml −1 ) were secreted to the culture media. The obtained bIFN- ω 1 showed a cross-species antiviral activity on four mammalian cell lines of calf, pig, hamster and human origin, but this activity was absent on Madin–Darby canine kidney (MDCK) cells. A delivery carrier was developed to permit a better release of bIFN- ω 1. When compared with a control group, an increase in 6 days in the corpus luteum lifespan was obtained in cyclic ewes following three interferon (IFN) intrauterine administrations on days 9, 10 and 11 post-estrus. In summary, these results demonstrated for the first time that biologically active recombinant bIFN- ω 1 was highly secreted by P. pastoris showing antiviral activity in different cell lines and an antiluteolytic effect in cyclic ewes, with no detrimental effects on the animals.

Single dose adenovirus vectored vaccine induces a potent and long-lasting immune response against rabbit hemorrhagic disease virus after parenteral or mucosal administration

Recent developments in cattle tick control have incorporated the use of recombinant Bm86 vaccines against this ectoparasite. The vaccine developed by our group (Gavac) contains an antigen expressed in Pichia pastoris, and has been successfully employed for the control of tick infestations and transmission of tick-borne diseases. Here, we examined the safety and effect of the Gavac vaccine on reproductive parameters in cattle. Toxicity tests in mice and guinea pigs demonstrated the safety of Gavac. To study the adverse effects of vaccination on reproduction, a field trial involving 9,500 animals in Cuba was conducted. The cattle at 3 farms were vaccinated while those on a fourth farm were left unvaccinated and served as the control. Following vaccination, the control of tick infestation and the transmission of babesiosis were used to demonstrate the efficacy of the vaccine. No adverse effects were observed in any of the reproductive parameters studied when comparing the data before and after vaccination with Gavac and between the vaccinated farms and the control farm. These results demonstrate that under the conditions of our study vaccination with Gavac is safe for use on cattle.

Two initial vaccinations with the Bm86-based Gavacplus vaccine against Rhipicephalus (Boophilus) microplus induce similar reproductive suppression to three initial vaccinations under production conditions.

Rabbit hemorrhagic disease virus (RHDV) is the etiological agent of a lethal and contagious disease of rabbits that remains as a serious problem worldwide. As this virus does not replicate in cell culture systems, the capsid protein gene has been expressed in heterologous hosts or inserted in replication-competent viruses in order to obtain non-conventional RHDV vaccines. However, due to technological or safety issues, current RHDV vaccines are still prepared from organs of infected rabbits. In this work, two human type 5 derived replication-defective adenoviruses encoding the rabbit hemorrhagic disease virus VP60 capsid protein were constructed. The recombinant protein was expressed as a multimer in mouse and rabbit cell lines at levels that ranged from approximately 120 to 160 mg/L of culture. Mice intravenously or subcutaneously inoculated with a single 10(8) gene transfer units (GTU) dose of the AdVP60 vector (designed for VP60 intracellular expression) seroconverted at days 7 and 14 post-immunization, respectively. This vector generated a stronger response than that obtained with a second vector (AdVP60sec) designed for VP60 secretion. Rabbits were then immunized by parenteral or mucosal routes with a single 10(9)GTU dose of the AdVP60 and the antibody response was evaluated using a competition ELISA specific for RHDV or RHDVa. Protective hemagglutination inhibition (HI) titers were also promptly detected and IgG antibodies corresponding with inhibition percentages over 85% persisted up to one year in all rabbits, independently of the immunization route employed. These levels were similar to those elicited with inactivated RHDV or with VP60 obtained from yeast or insect cells. IgA specific antibodies were only found in saliva of rabbits immunized by intranasal instillation. The feasibility of VP60 production and vaccination of rabbits with replication-defective adenoviral vectors was demonstrated.