Melina Garcia Guizzo

Effect of GSK-3 activity, enzymatic inhibition and gene silencing by RNAi on tick oviposition and egg hatching.

Glycogen synthase kinase-3 (GSK-3) is classically described as a key enzyme involved in glycogen metabolism in mammals. It has been shown to be highly conserved among several organisms, mainly in the catalytic domain region. This enzyme has already been described in Rhipicephalus (Boophilus) microplus and the ovaries of females appeared to be the major site of GSK-3 transcription. The treatment with GSK-3 specific inhibitor (alsterpaullone, bromo-indirubin-oxime 6 and indirubin-3-oxime) caused a reduction in oviposition and egg hatching in completely engorged female ticks. The effect was more pronounced in partially engorged females when alsterpaullone was administrated by artificial capillary feeding. Moreover, GSK-3 gene silencing by RNAi in partially engorged females reduced significantly both oviposition and hatching. The study of tick embryogenesis and proteins that participate in this process has been suggested as an important means for the development of novel strategies for parasite control. GSK-3 is an essential protein involved in embryonic processes and for this reason it has already been suggested as a possible antigen candidate for tick control.

Multi-antigenic vaccine against the cattle tick Rhipicephalus (Boophilus) microplus: a field evaluation.

The tick Rhipicephalus (Boophilus) microplus is a blood-sucking ectoparasite of cattle that severely impairs livestock production. Studies on tick immunological control address mostly single-antigen vaccines. However, from the commercial standpoint, so far no single-antigen vaccine has afforded appropriate protection against all R. microplus populations. In this context, multi-antigen cocktails have emerged as a way to enhance vaccine efficacy. In this work, a multi-antigenic vaccine against R. microplus was analyzed under field conditions in naturally infested cattle. The vaccine was composed by three tick recombinant proteins from two tick species that in previous single-vaccination reports provided partial protection of confined cattle against R. microplus infestations: vitellin-degrading cysteine endopeptidase (VTDCE) and boophilus yolk pro-cathepsin (BYC) from R. microplus, and glutathione S-transferase from Haemaphysalis longicornis (GST-Hl). Increased antibody levels against three proteins were recorded after immunizations, with a distinct humoral immune response dynamics for each protein. Compared to the control group, a statistically significant lower number of semi-engorged female ticks were observed in vaccinated cattle after two inoculations. This reduction persisted for 3 months, ranging from 35.3 to 61.6%. Furthermore, cattle body weight gain was significantly higher in vaccinated animals when compared to control cattle. Compared to the single-antigen vaccines composed by VTDCE, BYC or GST-Hl, this three-antigen vaccine afforded higher protection levels against R. microplus infestations.

Immunoprotective potential of a Rhipicephalus (Boophilus) microplus metalloprotease

Ticks have serious impacts on animal and human health, causing significant economic losses in cattle breeding. Besides damage due to the hematophagous behavior, they transmit several pathogens. Low cost and environmental safety have made vaccines a promising alternative control method against tick infestation. Metalloproteases (MPs) have been shown to be essential for diverse biological functions in hematophagous organisms, inhibiting blood clotting, degrading extracellular matrix proteins, and inhibiting host tissue repair via anti-angiogenic activity. In this study, we analyzed the immunoprotective potential of a recombinant MP against Rhipicephalus (Boophilus) microplus infestation. First, a cDNA encoding R. microplus amino acids sequence with highly conserved regions of the metzincin (reprolysin) group of MP was identified (BrRm-MP4). After expression and purification, recombinant BrRm-MP4 was used as a vaccinal antigen against R. microplus infestation in cattle (Bos taurus taurus). All vaccinated bovines developed immune response to the antigen, resulting in increased antibody level throughout the immunization protocol. Immunization with rBrRm-MP4 reduced tick feeding success, decreasing the number of engorged females and their reproduction potential, representing a 60% overall protection. These results show that rBrRm-MP4 provides protection against tick infestation, placing it is a potential candidate for an anti-tick vaccine.