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Background/Objectives: Amblyomma sculptum is among the most dangerous ticks in South America, as it is the species most associated with humans and is the main vector of Rickettsia rickettsii. In the face of the problems related to tick control based on chemical acaricides, vaccines emerge as a promising method. In previous works, three salivary recombinant proteins (rAs8.9kDa, rAsKunitz, and rAsBasicTail) and one protein based on intestinal immunogenic regions (rAsChimera) were described with 59 to 92% vaccine efficacy against A. sculptum females. Here, we evaluate novel vaccine formulations containing binary or multiple combinations of the antigens rAs8.9kDa, rAsKunitz, rAsBasicTail, and rAsChimera against the three instars of the tick. Methods: A control group of mice was immunized with adjuvant alone (aluminum hydroxide gel) and compared to five groups immunized with formulations containing two, three, or four of the antigens. Results: The formulations were safe, with no significant alterations to host behavior and hematological or biochemical parameters. Immunizations induced a significant increase in the CD19+ B lymphocyte percentage in all groups, but no difference was seen for CD8+ and CD4+ T lymphocytes or CD14+ monocytes. The best protection was observed for the formulations containing two antigens, which reached above 98% efficacy, while the groups containing three or four antigens presented 92.7 and 94.4% efficacy, respectively. Conclusions: All antigen combinations were promising as vaccine formulations against A. sculptum. The formulation containing rAs8.9kDa and rAsChimera showed the best efficacy and should be focused on in further experiments. [ABSTRACT FROM AUTHOR]

The cattle tick, Rhipicephalus microplus, is the most significant ectoparasite in the cattle industry. The application of acaricides constitutes the main control method. However, inadequate treatments have serious drawbacks, including the appearance of multi-resistant ticks. Tick vaccines offer a safe and economically sustainable alternative for controlling R. microplus. Nevertheless, the efficacy of existing vaccines has been limited by polymorphisms in target antigens among strains from different geographical regions. In this study, we characterized a putative Metalloprotease from the ADAMTSL family. We analyzed three regions to evaluate their transcriptional profiling in different R. microplus tick tissues, using two constitutive genes (β-tubulin and Elfa-1) as references. The expression levels showed that ADAMTSL-R1 was upregulated 39.37-fold (p ≤ 0.05) in salivary glands. The ADAMTSL-R2 showed the highest expression, rising 7.69-fold (p ≤ 0.05) in ovaries and up to 59.39-fold (p ≤ 0.05) in egg mass. Furthermore, this region showed the highest level of conservation among Rhipicephalus isolates. The ADAMTSL-R3 was upregulated only in the egg mass. The results of this study provide a basis for future research focused on elucidating the role of these protein variants in tick biology, including their feeding mechanisms and potential implications in pathogen transmission. Understanding these factors may aid in developing an effective tick vaccine. [ABSTRACT FROM AUTHOR]

The cattle tick, Rhipicephalus microplus, is the most significant ectoparasite in the cattle industry. The application of acaricides constitutes the main control method. However, inadequate treatments have serious drawbacks, including the appearance of multi-resistant ticks. Tick vaccines offer a safe and economically sustainable alternative for controlling R. microplus. Nevertheless, the efficacy of existing vaccines has been limited by polymorphisms in target antigens among strains from different geographical regions. In this study, we characterized a putative Metalloprotease from the ADAMTSL family. We analyzed three regions to evaluate their transcriptional profiling in different R. microplus tick tissues, using two constitutive genes (β-tubulin and Elfa-1) as references. The expression levels showed that ADAMTSL-R1 was upregulated 39.37-fold (p ≤ 0.05) in salivary glands. The ADAMTSL-R2 showed the highest expression, rising 7.69-fold (p ≤ 0.05) in ovaries and up to 59.39-fold (p ≤ 0.05) in egg mass. Furthermore, this region showed the highest level of conservation among Rhipicephalus isolates. The ADAMTSL-R3 was upregulated only in the egg mass. The results of this study provide a basis for future research focused on elucidating the role of these protein variants in tick biology, including their feeding mechanisms and potential implications in pathogen transmission. Understanding these factors may aid in developing an effective tick vaccine.

Background/Objectives: Amblyomma sculptum is among the most dangerous ticks in South America, as it is the species most associated with humans and is the main vector of Rickettsia rickettsii. In the face of the problems related to tick control based on chemical acaricides, vaccines emerge as a promising method. In previous works, three salivary recombinant proteins (rAs8.9kDa, rAsKunitz, and rAsBasicTail) and one protein based on intestinal immunogenic regions (rAsChimera) were described with 59 to 92% vaccine efficacy against A. sculptum females. Here, we evaluate novel vaccine formulations containing binary or multiple combinations of the antigens rAs8.9kDa, rAsKunitz, rAsBasicTail, and rAsChimera against the three instars of the tick. Methods: A control group of mice was immunized with adjuvant alone (aluminum hydroxide gel) and compared to five groups immunized with formulations containing two, three, or four of the antigens. Results: The formulations were safe, with no significant alterations to host behavior and hematological or biochemical parameters. Immunizations induced a significant increase in the CD19+ B lymphocyte percentage in all groups, but no difference was seen for CD8+ and CD4+ T lymphocytes or CD14+ monocytes. The best protection was observed for the formulations containing two antigens, which reached above 98% efficacy, while the groups containing three or four antigens presented 92.7 and 94.4% efficacy, respectively. Conclusions: All antigen combinations were promising as vaccine formulations against A. sculptum. The formulation containing rAs8.9kDa and rAsChimera showed the best efficacy and should be focused on in further experiments.

Abstract Background Tick control is mostly hampered by the rise of acaricide-resistant tick populations. Significant efforts have focused on developing alternative control methods, including cross-species protective and/or cocktail-based anti-tick vaccines, to achieve protection against various tick species. Methods In this study, full-length open reading frames encoding subolesin (SUB) from Rhipicephalus microplus and ferritin 2 (FER2) from Hyalomma anatolicum as well as the partial 60S acidic ribosomal protein (P0) from R. microplus were cloned, expressed in Escherichia coli and used as vaccine antigens against Rhipicephalus sanguineus sensu lato (R. sanguineus s.l.) infestation in rabbits. Results In silico analyses revealed that the SUB, P0 and FER2 proteins were antigenic and displayed limited similarity to the host's homologous proteins. The proteins shared identities of 97.5%, 100% and 89.5% with their SUB, P0 and FER2 R. sanguineus s.l. orthologous sequences, respectively. Antibodies against each recombinant protein cross-recognized the native proteins in the different tissues and developmental stages of R. sanguineus s.l. Overall efficacy of the SUB, FER2 and cocktail (SUB+FER2+P0) vaccines against R. sanguineus s.l. infestation was 86.3%, 95.9% and 90.9%, respectively. Conclusions Both mono-antigen and the cocktail anti-tick vaccines affected the biological parameters of R. sanguineus s.l. infestation in the rabbit model, which could be extrapolated to its infested host under natural conditions. These findings support the possibility of using mono-antigenic and cocktail-based vaccines for large-scale anti-tick vaccine development against multiple tick species. Graphical Abstract

This comprehensive review explores the field of anti-tick vaccines, addressing their significance in combating tick-borne diseases of public health concern. The main objectives are to provide a brief epidemiology of diseases affecting humans and a thorough understanding of tick biology, traditional tick control methods, the development and mechanisms of anti-tick vaccines, their efficacy in field applications, associated challenges, and future prospects. Tick-borne diseases (TBDs) pose a significant and escalating threat to global health and the livestock industries due to the widespread distribution of ticks and the multitude of pathogens they transmit. Traditional tick control methods, such as acaricides and repellents, have limitations, including environmental concerns and the emergence of tick resistance. Anti-tick vaccines offer a promising alternative by targeting specific tick proteins crucial for feeding and pathogen transmission. Developing vaccines with antigens based on these essential proteins is likely to disrupt these processes. Indeed, anti-tick vaccines have shown efficacy in laboratory and field trials successfully implemented in livestock, reducing the prevalence of TBDs. However, some challenges still remain, including vaccine efficacy on different hosts, polymorphisms in ticks of the same species, and the economic considerations of adopting large-scale vaccine strategies. Emerging technologies and approaches hold promise for improving anti-tick vaccine development and expanding their impact on public health and agriculture. [ABSTRACT FROM AUTHOR]

Background: Tick control is mostly hampered by the rise of acaricide-resistant tick populations. Significant efforts have focused on developing alternative control methods, including cross-species protective and/or cocktail-based anti-tick vaccines, to achieve protection against various tick species. Methods: In this study, full-length open reading frames encoding subolesin (SUB) from Rhipicephalus microplus and ferritin 2 (FER2) from Hyalomma anatolicum as well as the partial 60S acidic ribosomal protein (P0) from R. microplus were cloned, expressed in Escherichia coli and used as vaccine antigens against Rhipicephalus sanguineus sensu lato (R. sanguineus s.l.) infestation in rabbits. Results: In silico analyses revealed that the SUB, P0 and FER2 proteins were antigenic and displayed limited similarity to the host's homologous proteins. The proteins shared identities of 97.5%, 100% and 89.5% with their SUB, P0 and FER2 R. sanguineus s.l. orthologous sequences, respectively. Antibodies against each recombinant protein cross-recognized the native proteins in the different tissues and developmental stages of R. sanguineus s.l. Overall efficacy of the SUB, FER2 and cocktail (SUB+FER2+P0) vaccines against R. sanguineus s.l. infestation was 86.3%, 95.9% and 90.9%, respectively. Conclusions: Both mono-antigen and the cocktail anti-tick vaccines affected the biological parameters of R. sanguineus s.l. infestation in the rabbit model, which could be extrapolated to its infested host under natural conditions. These findings support the possibility of using mono-antigenic and cocktail-based vaccines for large-scale anti-tick vaccine development against multiple tick species. [ABSTRACT FROM AUTHOR]

The control of ticks through vaccination offers a sustainable alternative to the use of chemicals that cause contamination and the selection of resistant tick strains. However, only a limited number of anti-tick vaccines have reached commercial realization. In this sense, an antigen effective against different tick species is a desirable target for developing such vaccines. A peptide derived from the tick P0 protein (pP0) conjugated to a carrier protein has been demonstrated to be effective against the Rhipicephalus microplus, Rhipicephalus sanguineus, and Amblyomma mixtum tick species. The aim of this work was to assess the efficacy of this peptide when conjugated to the Bm86 protein against Dermacentor nitens and Ixodes ricinus ticks. An RNAi experiment using P0 dsRNA from I. ricinus showed a dramatic reduction in the feeding of injected female ticks on guinea pigs. In the follow-up vaccination experiments, rabbits were immunized with the pP0-Bm86 conjugate and challenged simultaneously with larvae, nymphs, and the adults of I. ricinus ticks. In the same way, horses were immunized with the pP0-Bm86 conjugate and challenged with D. nitens larva. The pP0-Bm86 conjugate showed efficacies of 63% and 55% against I. ricinus and D. nitens ticks, respectively. These results, combined with previous reports of efficacy for this conjugate, show the promising potential for its development as a broad-spectrum anti-tick vaccine. [ABSTRACT FROM AUTHOR]

Rhipicephalus microplus economically impacts cattle production in tropical and subtropical countries. Application of acaricides constitutes the major control method; however, inadequate use has increased resistant tick populations, resulting in environmental and cattle product contamination. Anti-tick vaccines based on the Bm86 antigen are an environmentally friendly, safe, and economically sustainable alternative for controlling R. microplus infestations. Nevertheless, variable efficacy has been experienced against different geographic tick strains. Herein, we evaluated the efficacy of a conserved polypeptide Bm86 derived from a Mexican R. microplus strain previously characterized. Twelve cows were assigned to three experimental groups and immunized with three doses of the polypeptide Bm86 (pBm86), adjuvant/saline alone, and Bm86 antigen (control +), respectively. Specific IgG antibody levels were measured by ELISA and confirmed by Western blot. In addition, the reproductive performance of naturally infested R. microplus was also determined. The more affected parameter was the adult female tick number, with a reduction of 44% by the pBm86 compared to the controls (p < 0.05), showing a vaccine efficacy of 58%. Anti-pBm86 IgG antibodies were immunogenic and capable of recognizing the native Bm86 protein in the eggs, larvae, and guts of R. microplus. The negative correlation between antibody levels and the reduction of naturally tick-infested cattle suggested that the effect of the polypeptide Bm86 was attributed to the antibody response in immunized cattle. In conclusion, the polypeptide Bm86 showed a specific immune response in cattle and conferred protection against R. microplus in a Mexican tropical region. These findings support further experiments with this antigen to demonstrate its effectiveness as a regional vaccine. [ABSTRACT FROM AUTHOR]

Although vaccines are one of the environmentally friendly means to prevent the spread of ticks, there is currently no commercial vaccine effective against Haemaphysalis longicornis ticks. In this study, we identified, characterized, localized, and evaluated the expression patterns, and tested the immunogenic potential of a homologue of Rhipicephalus microplus ATAQ in H. longicornis (HlATAQ). HlATAQ was identified as a 654 amino acid-long protein present throughout the midgut and in Malpighian tubule cells and containing six full and one partial EGF-like domains. HlATAQ was genetically distant (homology < 50%) from previously reported ATAQ proteins and was expressed throughout tick life stages. Its expression steadily increased (p < 0.001) during feeding, reached a peak, and then decreased slightly with engorgement. Silencing of HlATAQ did not result in a phenotype that was significantly different from the control ticks. However, H. longicornis female ticks fed on a rabbit immunized with recombinant HlATAQ showed significantly longer blood-feeding periods, higher body weight at engorgement, higher egg mass, and longer pre-oviposition and egg hatching periods than control ticks. These findings indicate that the ATAQ protein plays a role in the blood-feeding-related physiological processes in the midgut and Malpighian tubules and antibodies directed against it may affect these tissues and disrupt tick engorgement and oviposition. [ABSTRACT FROM AUTHOR]

This comprehensive review explores the field of anti-tick vaccines, addressing their significance in combating tick-borne diseases of public health concern. The main objectives are to provide a brief epidemiology of diseases affecting humans and a thorough understanding of tick biology, traditional tick control methods, the development and mechanisms of anti-tick vaccines, their efficacy in field applications, associated challenges, and future prospects. Tick-borne diseases (TBDs) pose a significant and escalating threat to global health and the livestock industries due to the widespread distribution of ticks and the multitude of pathogens they transmit. Traditional tick control methods, such as acaricides and repellents, have limitations, including environmental concerns and the emergence of tick resistance. Anti-tick vaccines offer a promising alternative by targeting specific tick proteins crucial for feeding and pathogen transmission. Developing vaccines with antigens based on these essential proteins is likely to disrupt these processes. Indeed, anti-tick vaccines have shown efficacy in laboratory and field trials successfully implemented in livestock, reducing the prevalence of TBDs. However, some challenges still remain, including vaccine efficacy on different hosts, polymorphisms in ticks of the same species, and the economic considerations of adopting large-scale vaccine strategies. Emerging technologies and approaches hold promise for improving anti-tick vaccine development and expanding their impact on public health and agriculture.

Abstract Background There have been ongoing efforts to identify anti-tick vaccine targets to protect cattle from infestation with cattle fever ticks Rhipicephalus (Boophilus) microplus. Two commercial vaccines based on the tick gut protein Bm86 have had variable effectiveness, which has led to poor acceptance, and numerous studies have attempted to identify vaccine antigens that will provide more consistently effective protection. Transcriptomic analysis of R. microplus led to identification of three aquaporin genes annotated to code for transmembrane proteins involved in the transport of water across cell membranes. Previous work showed that vaccination with full-length recombinant aquaporin 1 (RmAQP1) reduced tick burdens on cattle. Targeted silencing of aquaporin 2 (RmAQP2) expression suggested it might also be a good anti-tick vaccination target. Methods Three synthetic peptides from the predicted extracellular domains of RmAQP2 were used to vaccinate cattle. Peptides were conjugated to keyhole limpet hemocyanin (KLH) as an antigenic carrier molecule. We monitored the antibody response with ELISA and challenged vaccinated cattle with R. microplus larvae. Results There was a 25% reduction overall in the numbers of ticks feeding to repletion on the vaccinated cattle. Immune sera from vaccinated cattle recognized native tick proteins on a western blot and reacted to the three individual synthetic peptides in an ELISA. The vaccinated calf with the highest total IgG titer was not the most effective at controlling ticks; ratios of IgG isotypes 1 and 2 differed greatly among the three vaccinated cattle; the calf with the highest IgG1/IgG2 ratio had the fewest ticks. Ticks on vaccinated cattle had significantly greater replete weights compared to ticks on controls, mirroring results seen with RNA silencing of RmAQP2. However, protein data could not confirm that vaccination had any impact on the ability of the tick to concentrate its blood meal by removing water. Conclusions A reduced number of ticks feed successfully on cattle vaccinated to produce antibodies against the extracellular domains of RmAQP2. However, our predicted mechanism, that antibody binding blocks the ability of RmAQP2 to move water out of the blood meal, could not be confirmed. Further study will be required to define the mechanism of action and to determine whether these vaccine targets will be useful components of an anti-tick vaccine cocktail. Graphical Abstract

The control of ticks through vaccination offers a sustainable alternative to the use of chemicals that cause contamination and the selection of resistant tick strains. However, only a limited number of anti-tick vaccines have reached commercial realization. In this sense, an antigen effective against different tick species is a desirable target for developing such vaccines. A peptide derived from the tick P0 protein (pP0) conjugated to a carrier protein has been demonstrated to be effective against the Rhipicephalus microplus, Rhipicephalus sanguineus, and Amblyomma mixtum tick species. The aim of this work was to assess the efficacy of this peptide when conjugated to the Bm86 protein against Dermacentor nitens and Ixodes ricinus ticks. An RNAi experiment using P0 dsRNA from I. ricinus showed a dramatic reduction in the feeding of injected female ticks on guinea pigs. In the follow-up vaccination experiments, rabbits were immunized with the pP0-Bm86 conjugate and challenged simultaneously with larvae, nymphs, and the adults of I. ricinus ticks. In the same way, horses were immunized with the pP0-Bm86 conjugate and challenged with D. nitens larva. The pP0-Bm86 conjugate showed efficacies of 63% and 55% against I. ricinus and D. nitens ticks, respectively. These results, combined with previous reports of efficacy for this conjugate, show the promising potential for its development as a broad-spectrum anti-tick vaccine.

Ixodes ricinus and Ixodes scapularis are the main vectors for the causative agents of Lyme borreliosis and a wide range of other pathogens. Repeated tick-bites are known to lead to tick rejection; a phenomenon designated as tick immunity. Tick immunity is mainly directed against tick salivary gland proteins (TSGPs) and has been shown to partially protect against experimental Lyme borreliosis. TSGPs recognized by antibodies from tick immune animals could therefore be interesting candidates for an anti-tick vaccine, which might also block pathogen transmission. To identify conserved Ixodes TSGPs that could serve as a universal anti-tick vaccine in both Europe and the US, a Yeast Surface Display containing salivary gland genes of nymphal I. ricinus expressed at 24, 48 and 72 h into tick feeding was probed with either sera from rabbits repeatedly exposed for 24 h to I. ricinus nymphal ticks and/or sera from rabbits immune to I. scapularis. Thus, we identified thirteen TSGP vaccine candidates, of which ten were secreted. For vaccination studies in rabbits, we selected six secreted TSGPs, five full length and one conserved peptide. None of these proteins hampered tick feeding. In contrast, vaccination of guinea pigs with four non-secreted TSGPs – two from the current and two from a previous human immunoscreening - did significantly reduce tick attachment and feeding. Therefore, non-secreted TSGPs appear to be involved in the development of tick immunity and are interesting candidates for an anti-tick vaccine. [ABSTRACT FROM AUTHOR]

African swine fever (ASF), a highly contagious, deadly infectious disease, has caused huge economic losses to animal husbandry with a 100% mortality rate of the most acute and acute infection, which is listed as a legally reported animal disease by the World Organization for Animal Health (OIE). African swine fever virus (ASFV) is the causative agent of ASF, which is the only member of the Asfarviridae family. Ornithodoros soft ticks play an important role in ASFV transmission by active biological or mechanical transmission or by passive transport or ingestion, particularly in Africa, Europe, and the United States. First, this review summarized recent reports on (1) tick species capable of transmitting ASFV, (2) the importance of ticks in the transmission and epidemiological cycle of ASFV, and (3) the ASFV strains of tick transmission, to provide a detailed description of tick-borne ASFV. Second, the dynamics of tick infection with ASFV and the tick-induced immune suppression were further elaborated to explain how ticks spread ASFV. Third, the development of the anti-tick vaccine was summarized, and the prospect of the anti-tick vaccine was recapitulated. Then, the marked attenuated vaccine, ASFV-G-ΔI177L, was compared with those of the anti-tick vaccine to represent potential therapeutic or strategies to combat ASF.

Rhipicephalus microplus economically impacts cattle production in tropical and subtropical countries. Application of acaricides constitutes the major control method; however, inadequate use has increased resistant tick populations, resulting in environmental and cattle product contamination. Anti-tick vaccines based on the Bm86 antigen are an environmentally friendly, safe, and economically sustainable alternative for controlling R. microplus infestations. Nevertheless, variable efficacy has been experienced against different geographic tick strains. Herein, we evaluated the efficacy of a conserved polypeptide Bm86 derived from a Mexican R. microplus strain previously characterized. Twelve cows were assigned to three experimental groups and immunized with three doses of the polypeptide Bm86 (pBm86), adjuvant/saline alone, and Bm86 antigen (control +), respectively. Specific IgG antibody levels were measured by ELISA and confirmed by Western blot. In addition, the reproductive performance of naturally infested R. microplus was also determined. The more affected parameter was the adult female tick number, with a reduction of 44% by the pBm86 compared to the controls (p < 0.05), showing a vaccine efficacy of 58%. Anti-pBm86 IgG antibodies were immunogenic and capable of recognizing the native Bm86 protein in the eggs, larvae, and guts of R. microplus. The negative correlation between antibody levels and the reduction of naturally tick-infested cattle suggested that the effect of the polypeptide Bm86 was attributed to the antibody response in immunized cattle. In conclusion, the polypeptide Bm86 showed a specific immune response in cattle and conferred protection against R. microplus in a Mexican tropical region. These findings support further experiments with this antigen to demonstrate its effectiveness as a regional vaccine.

Tick saliva has been extensively studied in the context of tick-host interactions because it is involved in host homeostasis modulation and microbial pathogen transmission to the host. Accumulated knowledge about the tick saliva composition at the molecular level has revealed that serine protease inhibitors play a key role in the tick-host interaction. Serpins are one highly expressed group of protease inhibitors in tick salivary glands, their expression can be induced during tick blood-feeding, and they have many biological functions at the tick-host interface. Indeed, tick serpins have an important role in inhibiting host hemostatic processes and in the modulation of the innate and adaptive immune responses of their vertebrate hosts. Tick serpins have also been studied as potential candidates for therapeutic use and vaccine development. In this review, we critically summarize the current state of knowledge about the biological role of tick serpins in shaping tick-host interactions with emphasis on the mechanisms by which they modulate host immunity. Their potential use in drug and vaccine development is also discussed. [ABSTRACT FROM AUTHOR]

Tick saliva has been extensively studied in the context of tick-host interactions because it is involved in host homeostasis modulation and microbial pathogen transmission to the host. Accumulated knowledge about the tick saliva composition at the molecular level has revealed that serine protease inhibitors play a key role in the tick-host interaction. Serpins are one highly expressed group of protease inhibitors in tick salivary glands, their expression can be induced during tick blood-feeding, and they have many biological functions at the tick-host interface. Indeed, tick serpins have an important role in inhibiting host hemostatic processes and in the modulation of the innate and adaptive immune responses of their vertebrate hosts. Tick serpins have also been studied as potential candidates for therapeutic use and vaccine development. In this review, we critically summarize the current state of knowledge about the biological role of tick serpins in shaping tick-host interactions with emphasis on the mechanisms by which they modulate host immunity. Their potential use in drug and vaccine development is also discussed.

Background: There have been ongoing efforts to identify anti-tick vaccine targets to protect cattle from infestation with cattle fever ticks Rhipicephalus (Boophilus) microplus. Two commercial vaccines based on the tick gut protein Bm86 have had variable effectiveness, which has led to poor acceptance, and numerous studies have attempted to identify vaccine antigens that will provide more consistently effective protection. Transcriptomic analysis of R. microplus led to identification of three aquaporin genes annotated to code for transmembrane proteins involved in the transport of water across cell membranes. Previous work showed that vaccination with full-length recombinant aquaporin 1 (RmAQP1) reduced tick burdens on cattle. Targeted silencing of aquaporin 2 (RmAQP2) expression suggested it might also be a good anti-tick vaccination target. Methods: Three synthetic peptides from the predicted extracellular domains of RmAQP2 were used to vaccinate cattle. Peptides were conjugated to keyhole limpet hemocyanin (KLH) as an antigenic carrier molecule. We monitored the antibody response with ELISA and challenged vaccinated cattle with R. microplus larvae. Results: There was a 25% reduction overall in the numbers of ticks feeding to repletion on the vaccinated cattle. Immune sera from vaccinated cattle recognized native tick proteins on a western blot and reacted to the three individual synthetic peptides in an ELISA. The vaccinated calf with the highest total IgG titer was not the most effective at controlling ticks; ratios of IgG isotypes 1 and 2 differed greatly among the three vaccinated cattle; the calf with the highest IgG1/IgG2 ratio had the fewest ticks. Ticks on vaccinated cattle had significantly greater replete weights compared to ticks on controls, mirroring results seen with RNA silencing of RmAQP2. However, protein data could not confirm that vaccination had any impact on the ability of the tick to concentrate its blood meal by removing water. Conclusions: A reduced number of ticks feed successfully on cattle vaccinated to produce antibodies against the extracellular domains of RmAQP2. However, our predicted mechanism, that antibody binding blocks the ability of RmAQP2 to move water out of the blood meal, could not be confirmed. Further study will be required to define the mechanism of action and to determine whether these vaccine targets will be useful components of an anti-tick vaccine cocktail. [ABSTRACT FROM AUTHOR]

Although vaccines are one of the environmentally friendly means to prevent the spread of ticks, there is currently no commercial vaccine effective against Haemaphysalis longicornis ticks. In this study, we identified, characterized, localized, and evaluated the expression patterns, and tested the immunogenic potential of a homologue of Rhipicephalus microplus ATAQ in H. longicornis (HlATAQ). HlATAQ was identified as a 654 amino acid-long protein present throughout the midgut and in Malpighian tubule cells and containing six full and one partial EGF-like domains. HlATAQ was genetically distant (homology < 50%) from previously reported ATAQ proteins and was expressed throughout tick life stages. Its expression steadily increased (p < 0.001) during feeding, reached a peak, and then decreased slightly with engorgement. Silencing of HlATAQ did not result in a phenotype that was significantly different from the control ticks. However, H. longicornis female ticks fed on a rabbit immunized with recombinant HlATAQ showed significantly longer blood-feeding periods, higher body weight at engorgement, higher egg mass, and longer pre-oviposition and egg hatching periods than control ticks. These findings indicate that the ATAQ protein plays a role in the blood-feeding-related physiological processes in the midgut and Malpighian tubules and antibodies directed against it may affect these tissues and disrupt tick engorgement and oviposition.

• Amblyomma sculptum is one the most important tick species in south America. • Few works have focused on the search for anti- A. sculptum vaccines. • Concealed gut proteins may be promissing vaccine antigens against A. sculptum. • A chimeric protein was constructed with promising efficacy against A. sculptum. • Vaccine induced a reduction in female fertility and nymph mortality. Amblyomma sculptum is widely distributed in Brazil and is the main vector of Rickettsia rickettsii , the causative agent of the Brazilian spotted fever (BSF). Tick gut proteins play an essential role in blood feeding, digestion, and protection of gut epithelium. Therefore, many of these were investigated as potential vaccine targets for tick-control strategies. The present study aimed to select transcripts corresponding to putative immunogenic proteins in the A. sculptum gut epithelial membrane, produce recombinant proteins and evaluate them as antigens against A. sculptum infestations. Three gut proteins − AsMucin, AsAPP, and AsLAMP − and a chimeric protein (rAsChimera) based on 22 peptides containing putative B cell epitopes from seven different gut proteins were evaluated as anti- A. sculptum antigens. Mice immunizations revealed that all recombinant targets elicited humoral response with significantly increased IgG levels compared to controls. For rAsChimera, IgG levels remained significantly higher than controls up to 75 days after the end of the immunization. Challenge trials revealed that vaccination with the chimeric protein was the most effective against A. sculptum , inducing 100 % nymph mortality and reaching 80.8 % efficacy against females. The other three proteins did not induce relevant protection, as AsAPP had only 26.6 % efficacy, whereas AsMucin and AsLAMP induced no protection. These data indicate that targeting gut protein immunogenic regions may be an effective strategy for a vaccine formulation against A. sculptum. [ABSTRACT FROM AUTHOR]

A peptide from the P0 acidic ribosomal protein (pP0) of ticks conjugated to keyhole limpet hemocyanin from Megathura crenulata has shown to be effective against different tick species when used in host vaccination. Turning this peptide into a commercial anti-tick vaccine will depend on finding the appropriate, technically and economically feasible way to present it to the host immune system. Two conjugates (p64K-Cys1pP0 and p64K-βAla1pP0) were synthesized using the p64K carrier protein from Neisseria meningitidis produced in Escherichia coli, the same cross-linking reagent, and two analogues of pP0. The SDS-PAGE analysis of p64K-Cys1pP0 showed a heterogeneous conjugate compared to p64K-βAla1pP0 that was detected as a protein band at 91kDa. The pP0/p64K ratio determined by MALDI-MS for p64K-Cys1pP0 ranged from 1 to 8, being 3-5 the predominant ratio, while in the case of p64K-βAla1pP0 this ratio was 5-7. Cys1pP0 was partially linked to 35 out of 39 Lys residues and the N-terminal end, while βAla1pP0 was mostly linked to the six free cysteine residues, to the N-terminal end, and, in a lesser extent, to Lys residues. The assignment of the conjugation sites and side reactions were based on the identification of type 2 peptides. Rabbit immunizations showed the best anti-pP0 titers and the highest efficacy against Rhipicephalus sanguineus ticks when the p64K-Cys1pP0 was used as vaccine antigen. The presence of high molecular mass aggregates observed in the SDS-PAGE analysis of p64K-Cys1pP0 could be responsible for a better immune response against pP0 and consequently for its better efficacy as an anti-tick vaccine. [ABSTRACT FROM AUTHOR]

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Acquired tick resistance is a phenomenon wherein the host elicits an immune response against tick salivary components upon repeated tick infestations. The immune responses, potentially directed against critical salivary components, thwart tick feeding, and the animal becomes resistant to subsequent tick infestations. The development of tick resistance is frequently observed when ticks feed on non‐natural hosts, but not on natural hosts. The molecular mechanisms that lead to the development of tick resistance are not fully understood, and both host and tick factors are invoked in this phenomenon. Advances in molecular tools to address the host and the tick are beginning to reveal new insights into this phenomenon and to uncover a deeper understanding of the fundamental biology of tick‐host interactions. This review will focus on the expanding understanding of acquired tick resistance and highlight the impact of this understanding on anti‐tick vaccine development efforts. [ABSTRACT FROM AUTHOR]

Ticks are ubiquitous blood-sucking ectoparasites capable of transmitting a wide range of pathogens such as bacteria, viruses, protozoa, and fungi to animals and humans. Although the use of chemicals (acaricides) is the predominant method of tick-control, there are increasing incidents of acaricide tick resistance. Furthermore, there are concerns over accumulation of acaricide residues in meat, milk and in the environment. Therefore, alternative methods of tick-control have been proposed, of which anti-tick cattle vaccination is regarded as sustainable and user-friendly. Over the years, tremendous progress has been made in identifying and evaluating novel candidate tick vaccines, yet none of them have reached the global market. Until now, Bm86-based vaccines (Gavac™ in Cuba and TickGARDPLUS™ Australia-ceased in 2010) are still the only globally commercialized anti-tick vaccines. In contrast to Bm86, often, the novel candidate anti-tick vaccines show a lower protection efficacy. Why is this so? In response, herein, the potential bottlenecks to formulating efficacious anti-tick vaccines are examined. Aside from Bm86, the effectiveness of other anti-tick vaccines is rarely assessed. So, how can the researchers assess anti-tick vaccine effectiveness before field application? The approaches that are currently used to determine anti-tick vaccine efficacy are re-examined in this review. In addition, a model is proposed to aid in assessing anti-tick vaccine effectiveness. Finally, based on the principles for the development of general veterinary vaccines, a pipeline is proposed to guide in the development of anti-tick vaccines.

Abstract Background Ticks cause massive damage to livestock and vaccines are one sustainable substitute for the acaricides currently heavily used to control infestations. To guide antigen discovery for a vaccine that targets the gamut of parasitic strategies mediated by tick saliva and enables immunological memory, we exploited a transcriptome constructed from salivary glands from all stages of Rhipicephalus microplus ticks feeding on genetically tick-resistant and susceptible bovines. Results Different levels of host anti-tick immunity affected gene expression in tick salivary glands; we thus selected four proteins encoded by genes weakly expressed in ticks attempting to feed on resistant hosts or otherwise abundantly expressed in ticks fed on susceptible hosts; these sialoproteins mediate four functions of parasitism deployed by male ticks and that do not induce antibodies in naturally infected, susceptible bovines. We then evaluated in tick-susceptible heifers an alum-adjuvanted vaccine formulated with recombinant proteins. Parasite performance (i.e. weight and numbers of females finishing their parasitic cycle) and titres of antigen-specific antibodies were significantly reduced or increased, respectively, in vaccinated versus control heifers, conferring an efficacy of 73.2%; two of the antigens were strong immunogens, rich in predicted T-cell epitopes and challenge infestations boosted antibody responses against them. Conclusion Mining sialotranscriptomes guided by the immunity of tick-resistant hosts selected important targets and infestations boosted immune memory against salivary antigens.

The medical and veterinary public health importance of ticks and tick-borne pathogens is increasing due to the expansion of the geographic ranges of both ticks and pathogens, increasing tick populations, growing incidence of tick-borne diseases, emerging tick transmitted pathogens, and continued challenges of achieving effective and sustained tick control. The past decades show an increasing interest in the immune-mediated control of tick infestations and pathogen transmission through the use of vaccines. Bovine tick resistance induced by repeated infestations was reported over a century ago. This review addresses the phenomena and immunological underpinning of resistance to tick infestation by livestock and laboratory animals; the scope of tick countermeasures to host immune defenses; and the impact of genomics, functional genomics, and proteomics on dissecting complex tick–host–pathogen interactions. From early studies utilizing tick tissue extracts to salivary gland derived molecules and components of physiologically important pathways in tick gut and other tissues, an increased understanding of these relationships, over time, impacted the evolution of anti-tick vaccine antigen selection. Novel antigens continue to emerge, including increased interest in the tick microbiome. Anti-tick and transmission blocking vaccines targeting pathogen reservoirs have the potential to disrupt enzootic cycles and reduce human, companion, domestic animal, and wildlife exposure to infected ticks.

Ticks and the diseases they transmit are of huge veterinary, medical and economic importance worldwide. Control of ticks attacking livestock and companion animals is achieved primarily by application of chemical or plant-based acaricides. However, ticks can rapidly develop resistance to any new product brought onto the market, necessitating an ongoing search for novel active compounds and alternative approaches to tick control. Many aspects of tick and tick-borne pathogen research have been facilitated by the application of continuous cell lines derived from some of the most economically important tick species. These include cell lines derived from acaricide-susceptible and resistant ticks, cell sub-lines with in vitro -generated acaricide resistance, and genetically modified tick cells. Although not a replacement for the whole organism, tick cell lines enable studies at the cellular and molecular level and provide a more accessible, more ethical and less expensive in vitro alternative to in vivo tick feeding experiments. Here we review the role played by tick cell lines in studies on acaricide resistance, mode-of-action of acaricides, identification of potential novel control targets through better understanding of tick metabolism, and anti-tick vaccine development, that may lead to new approaches to control ticks and tick-borne diseases. [ABSTRACT FROM AUTHOR]

• Protection capacity of Subolesin against multi-tick infestations. • DNA constructs and chimeric polypeptides formulations as tools to evaluate anti-tick vaccine candidates. • High vaccination efficacies were obtained following immunization protocols involved vaccine different preparations. Vaccination against ticks is an environmentally friendly alternative control method compared to chemical acaricide applications. Subolesin is a conserved protein in ticks, which can provide protection against some tick species. In this study, we evaluated the capacity of cocktail vaccination with Subolesin and ribosomal acidic protein 0 (P0) peptide against adults of Haemaphysalis longicornis. Priming with DNA vaccine expressing subolesin, followed by boosters of a single antigen (rRhSub) or a chimeric polypeptide (rRhSub/P0), provided cross protection. This treatment resulted in significant mortality, reduced blood ingestion and reduced reproduction in H. longicornis adults. Vaccination efficacies of 79.3% and 86.6% are reported in groups supplemented with rRhSub and rRhSub/P0, respectively. Conserved antigens, such as subolesin , formulated as DNA vaccine and enhanced with chimeric polypeptides, could be used as an anti-tick vaccine application, especially for control of infestation involving several tick species. [ABSTRACT FROM AUTHOR]

Bovine babesiosis, caused by Babesia bovis and B. bigemina, is a major tick-borne disease of cattle with global economic impact. The disease can be prevented using integrated control measures including attenuated Babesia vaccines, babesicidal drugs, and tick control approaches. Vaccination of cattle with the Rhipicephalus microplus Bm86-based recombinant vaccine reduces the fitness of R. microplus and R. annulatus, but several booster inoculations are required to maintain protection. Herein, we generated a stable transfected strain of B. bovis expressing an enhanced GFP (eGFP) and a chimeric version of Bm86 (B. bovis/Bm86/eGFP). The eGFP was expressed in the parasite cytoplasm, whereas Bm86 was displayed on the surface of merozoites. Three splenectomized calves experimentally infected with B. bovis/Bm86/eGFP showed mild signs of acute disease and developed long-lasting antibody responses to B. bovis and native Bm86. No evidence of sequestration of parasites in the cerebral capillaries was found upon postmortem analysis, confirming attenuation of the strain. This is the first report of transfected B. bovis expressing the tick antigen Bm86 on the merozoite surface that elicits an antibody response to native Bm86. These results represent a proof of concept for a novel live, attenuated, tagged dual-vaccine approach to attempt simultaneous control of babesiosis and tick infestation.

A synthetic 20 amino acid peptide of the ribosomal protein P0 from ticks, when conjugated to keyhole limpet hemocyanin from Megathura crenulata and used as an immunogen against Rhipicephalus microplus and Rhipicephalus sanguineus s.l. species, has shown efficacies of around 90%. There is also experimental evidence of a high efficacy of this conjugate against Amblyomma mixtum and Ixodes ricinus species, which suggest that this antigen could be a good broad-spectrum anti-tick vaccine candidate. In this study, the P0 peptide (pP0) was chemically conjugated to Bm86 as a carrier protein. SDS-PAGE analysis of this conjugate demonstrated that it is highly heterogeneous in size, carrying from 1 to 18 molecules of pP0 per molecule of Bm86. Forty-nine out of the 54 lysine residues and the N-terminal end of Bm86 were found partially linked to pP0 by using LC-MS/MS analysis and the combination of four different softwares. Several post-translational modifications of Bm86 protein were also identified by mass spectrometry. High immunogenicity and efficacy were achieved when dogs and cattle were vaccinated with the pP0–Bm86 conjugate and challenged with R. sanguineus s.l. and R. microplus, respectively. These results encourage the development of this antigen with promising possibilities as an anti-tick vaccine.

Anti-tick vaccines have the potential to be an environmentally friendly and cost-effective option for tick control. In vaccine development, the identification of efficacious antigens forms the major bottleneck. In this study, the efficacy of immunization with recombinant ferritin 2 and native tick protein extracts (TPEs) against Ixodes ricinus infestations in calves was assessed in two immunization experiments. In the first experiment, each calf (n = 3) was immunized twice with recombinant ferritin 2 from I. ricinus (IrFER2), TPE consisting of soluble proteins from the internal organs of partially fed I. ricinus females, or adjuvant, respectively. In the second experiment, each calf (n = 4) was immunized with protein extracts from the midgut (ME) of partially fed females, the salivary glands (SGE) of partially fed females, a combination of ME and SGE, or adjuvant, respectively. Two weeks after the booster immunization, calves were challenged with 100 females and 200 nymphs. Blood was collected from the calves before the first and after the second immunization and fed to I. ricinus females and nymphs using an in vitro artificial tick feeding system. The two calves vaccinated with whole TPE and midgut extract (ME) showed hyperemia on tick bite sites 2 days post tick infestation and exudative blisters were observed in the ME-vaccinated animal, signs that were suggestive of a delayed type hypersensitivity (DTH) reaction. Significantly fewer ticks successfully fed on the three animals vaccinated with TPE, SGE, or ME. Adults fed on the TPE and ME vaccinated animals weighed significantly less. Tick feeding on the IrFER2 vaccinated calf was not impaired. The in vitro feeding of serum or fresh whole blood collected from the vaccinated animals did not significantly affect tick feeding success. Immunization with native I. ricinus TPEs thus conferred a strong immune response in calves and significantly reduced the feeding success of both nymphs and adults. In vitro feeding of serum or blood collected from vaccinated animals to ticks did not affect tick feeding, indicating that antibodies alone were not responsible for the observed vaccine immunity.

After Rhipicephalus microplus, the most important tick species affecting livestock industry in Cuba belong to the Amblyomma genus. There are few reports of effective vaccine antigens for these species. Recently, vaccination and challenge trials using a peptide from the P0 acidic ribosomal protein of R. microplus ticks (pP0) as antigen have shown an efficacy around 90% against tick species from the Rhipicephalus genus. Given the high degree of sequence conservation among tick species, pP0 could be an antigen of versatile use in anti‐tick vaccine formulations. In this paper, seven rabbits were immunized with a chemical conjugate of pP0 to keyhole limpet haemocyanin. Rabbits were challenged with an average of 1,900 Amblyomma mixtum larvae from a Cuban tick strain. The average number of recovered fed larvae and the viability of larvae in the moulting process were significantly lower in vaccinated animals compared with the control group. The overall vaccine efficacy of the P0 peptide antigen is 54% according to the calculated parameters. [ABSTRACT FROM AUTHOR]

Crimean‐Congo haemorrhagic fever (CCHF) is a tick borne viral disease reported from different parts of the world. The distribution of the CCHF cases are linked with the distribution of the principal vector, Hyalomma anatolicum in the ecosystem. Presently, vector control is mainly dependent on repeated application of acaricides, results in partial efficacy and generated acaricide resistant tick strains. Amongst the different components of integrated management programme, immunization of hosts is considered as one of the sustainable component. To restrict CCHF virus spreading, use of anti‐Hyalomma vaccines appears as a viable solution. Accordingly, present study was under taken to characterize and evaluate vaccine potential of two conserved molecules, ferritin2 (FER2) and tropomyosin (TPM). Silencing of the genes conferred a cumulative reduction (rejection + unable to engorge) of 61.3% in FER2 and 70.2% in TPM respectively. Furthermore, 44.2% and 72.7% reduction in engorgement weight, 63.6% and 94.9% reduction in egg masses in FER2 and TPM silenced ticks in comparison to LUC‐control group was recorded. The recombinant protein, rHaFER2 was characterized as 35 kDa protein with pI of 5.84 and possesses iron binding domains. While rHaTPM is a 51kDa protein with pI of 4.94 having calcium binding domains. Immunization of cross‐bred calves by rHaFER2 conferred 51.7% and 51.2% protection against larvae and adults of H. anatolicum challenge infestations. While rHaTPM conferred 63.7% and 66.4% protection against larvae and adults infestations, respectively. The results were comparable with the data generated by RNAi and it clearly showed the possibility for the development of anti‐hyalomma vaccine to manage CCHF virus and Theileria annulata infection in human and animals. [ABSTRACT FROM AUTHOR]

Anti-tick vaccines have the potential to be an environmentally friendly and cost-effective option for tick control. In vaccine development, the identification of efficacious antigens forms the major bottleneck. In this study, the efficacy of immunization with recombinant ferritin 2 and native tick protein extracts (TPEs) against Ixodes ricinus infestations in calves was assessed in two immunization experiments. In the first experiment, each calf (n = 3) was immunized twice with recombinant ferritin 2 from I. ricinus (IrFER2), TPE consisting of soluble proteins from the internal organs of partially fed I. ricinus females, or adjuvant, respectively. In the second experiment, each calf (n = 4) was immunized with protein extracts from the midgut (ME) of partially fed females, the salivary glands (SGE) of partially fed females, a combination of ME and SGE, or adjuvant, respectively. Two weeks after the booster immunization, calves were challenged with 100 females and 200 nymphs. Blood was collected from the calves before the first and after the second immunization and fed to I. ricinus females and nymphs using an in vitro artificial tick feeding system. The two calves vaccinated with whole TPE and midgut extract (ME) showed hyperemia on tick bite sites 2 days post tick infestation and exudative blisters were observed in the ME-vaccinated animal, signs that were suggestive of a delayed type hypersensitivity (DTH) reaction. Significantly fewer ticks successfully fed on the three animals vaccinated with TPE, SGE, or ME. Adults fed on the TPE and ME vaccinated animals weighed significantly less. Tick feeding on the IrFER2 vaccinated calf was not impaired. The in vitro feeding of serum or fresh whole blood collected from the vaccinated animals did not significantly affect tick feeding success. Immunization with native I. ricinus TPEs thus conferred a strong immune response in calves and significantly reduced the feeding success of both nymphs and adults. In vitro feeding of serum or blood collected from vaccinated animals to ticks did not affect tick feeding, indicating that antibodies alone were not responsible for the observed vaccine immunity. [ABSTRACT FROM AUTHOR]

Abstract INCOGARR is a thematic network recently approved to be financially supported by the Ibero-American Program of Science and Technology for Development (CYTED). The objectives of this Network are the design and evaluation of an efficient and feasible anti-tick vaccine candidate from the technical and economical points of view and also sharing experiences in the immunological control of ticks as part of an Integrated Control Program. The Network consists of seven laboratories and one company from six countries. The first meeting of the Network took place with the representation of each laboratory involved. In the meeting, general and specific objectives and activities of the Network were discussed and it was a very nice example of international collaboration to address an unsolved worldwide topic on tick control in which laboratories with different competencies and expertise join their efforts in a common goal. [ABSTRACT FROM AUTHOR]

Due to the continuous threat of ticks and tick-borne diseases to human and animal health worldwide, and the drawbacks of chemical acaricide application, many researchers are exploring vaccination as an alternative tick control method. Earlier studies have shown that host antibodies can circulate in the ticks, but it has not been confirmed whether these antibodies can be passed on to the eggs. We previously reported that ticks infesting rabbits immunized with a recombinant secretory ferritin of Haemaphysalis longicornis (HlFER2) had reduced egg production and hatching. Here we attempted to detect the presence of antibodies against HlFER2 in the ovary and eggs of female ticks through immunofluorescent visualization. Purified anti-HlFER2 antibodies or rabbit IgG for control was directly injected to engorged female H. longicornis . Ovaries and eggs after oviposition were collected and prepared for an indirect immunofluorescent antibody test. Positive fluorescence was detected in ovaries one day post-injection of anti-HlFER2 antibodies. Through silencing of Hlfer2 gene, we also determined whether the injected antibodies can specifically bind to native HlFER2. Immunofluorescence was observed in the oocytes of dsLuciferase control ticks injected with anti-HlFER2 antibodies, but not in the oocytes of Hlfer2 -silenced ticks also injected with anti-HlFER2 antibodies. Our current findings suggest that host antibodies can be passed on to the oocytes, which is significant in formulating a vaccine that can disrupt tick reproduction. [ABSTRACT FROM AUTHOR]

Haemaphysalis longicornis can transmit high varieties of tick-borne pathogens (TBPs), and a primary strategy for preventing the transmission of those TBPs is to control ticks. Hemalin, a thrombin inhibitor of the Kunitz-type family and a crucial component in H. longicornis feeding process has been isolated from parthenogentic ticks. This study aimed to evaluate the validity of a recombinant Hemalin (rHlHemalin) vaccination as an anti-tick vaccine against H. longicornis in rabbits to find a new candidate for an effective tick control. In this study, mouse splenocytes were isolated and used to investigate immune responses after rHlHemalin stimulation. The rabbits were vaccinated with the rHlHemalin protein. After tick challenges, body weight at engorgement, egg mass, and the reproductive cycle of H. longicornis were evaluated. To confirm the vaccination, the passive immunization tests of α-rHlHemalin sera were performed. The results showed that the rHlHemalin protein could stimulate cytokine production in mouse splenocytes. Vaccination assay revealed that the periods from tick infestations to egg-hatch in the vaccination group were significantly longer than those in the phosphate buffer saline (PBS) group (P = 0.0003). In addition, the tick body weight at engorgement (P = 0.0019) and egg mass at 10 days after oviposition (P = 0.0232) were higher than those in the PBS group. These findings were consistent with the current passive immunization results and suggest rHlHemalin vaccination extended the reproductive cycle in H. longicornis but did not decrease the body weight at engorgement or weight of egg mass. Therefore, it is debatable whether Hemalin vaccination is highly-effective anti-tick vaccine or not. However, due to the importance of thrombin inhibitors in tick blood feeding and blood digestion, additional inhibitor-based vaccines should be developed aiming to find an effective and environmentally friendly biological strategy to combat ticks. • The recombinant Hemalin protein of Haemaphysalis longicornis could stimulate cytokine production in mouse splenocytes. • rHlHemalin vaccination extended the reproductive cycle of H. longicornis. • The passive immunization of α-rHlHemalin sera modulated the reproductive cycle of H. longicornis. [ABSTRACT FROM AUTHOR]

Ticks are the most important vectors of pathogens affecting both domestic and wild animals worldwide. Hard tick feeding is a slow process—taking up to several days—and necessitates extended control over the host response. The success of the feeding process depends upon injection of tick saliva, which not only controls host hemostasis and wound healing, but also subverts the host immune response to avoid tick rejection that creates a favorable niche for the survival and propagation of diverse tick-borne pathogens. Here, we report on the molecular and biochemical features and functions of an Ixodes ricinus serine protease inhibitor (IrSPI). We characterize IrSPI as a Kunitz elastase inhibitor that is overexpressed in several tick organs—especially salivary glands—during blood-feeding. We also demonstrated that when IrSPI is injected into the host through saliva, it had no impact on tissue factor pathway-induced coagulation, fibrinolysis, endothelial cell angiogenesis or apoptosis, but the protein exhibits immunomodulatory activity. In particular, IrSPI represses proliferation of CD4+ T lymphocytes and proinflammatory cytokine secretion from both splenocytes and macrophages. Our study contributes valuable knowledge to tick-host interactions and provides insights that could be further exploited to design anti-tick vaccines targeting this immunomodulator implicated in I. ricinus feeding.

Background: Ticks cause massive damage to livestock and vaccines are one sustainable substitute for the acaricides currently heavily used to control infestations. To guide antigen discovery for a vaccine that targets the gamut of parasitic strategies mediated by tick saliva and enables immunological memory, we exploited a transcriptome constructed from salivary glands from all stages of Rhipicephalus microplus ticks feeding on genetically tick-resistant and susceptible bovines. Results: Different levels of host anti-tick immunity affected gene expression in tick salivary glands; we thus selected four proteins encoded by genes weakly expressed in ticks attempting to feed on resistant hosts or otherwise abundantly expressed in ticks fed on susceptible hosts; these sialoproteins mediate four functions of parasitism deployed by male ticks and that do not induce antibodies in naturally infected, susceptible bovines. We then evaluated in tick-susceptible heifers an alum-adjuvanted vaccine formulated with recombinant proteins. Parasite performance (i.e. weight and numbers of females finishing their parasitic cycle) and titres of antigen-specific antibodies were significantly reduced or increased, respectively, in vaccinated versus control heifers, conferring an efficacy of 73.2%; two of the antigens were strong immunogens, rich in predicted T-cell epitopes and challenge infestations boosted antibody responses against them. Conclusion: Mining sialotranscriptomes guided by the immunity of tick-resistant hosts selected important targets and infestations boosted immune memory against salivary antigens. [ABSTRACT FROM AUTHOR]

The southern cattle fever tick, Rhipicephalus (Boophilus) microplus, is no doubt the most economically important ectoparasite of cattle globally. The inappropriate use of chemical acaricides has driven the evolution of resistance in populations of R. (B.) microplus. Anti-tick vaccines represent a technology that can be combined with acaricides in integrated control programs to mitigate the impact of R. (B.) microplus. The recombinant form of Bm86 antigen from the Campo Grande (rBm86-CG) strain of R. (B.) microplus was produced using the Pichiapastoris expression system to test its ability to immunoprotect cattle against tick infestation. Secretion of rBm86-CG by P. pastoris through the bioprocess reported here simplified purification of the antigen. A specific humoral immune response was detected by ELISA in vaccinated cattle. Immunoblot results revealed that polyclonal antibodies from vaccinated cattle recognized a protein in larval extracts with a molecular weight corresponding to Bm86. The rBm86-CG antigen showed 31% efficacy against the Campo Grande strain of R. (B.) microplus infesting vaccinated cattle. The rBm86-CG is an antigen that could be used in a polyvalent vaccine as part of an integrated program for the control of R. (B.) microplus in the region that includes Mato Grosso do Sul.O carrapato Rhipicephalus (Boophilus) microplus é, sem dúvidas, o ectoparasito economicamente mais importante para o gado a nível mundial. A utilização inadequada de acaricidas tem impulsionado a evolução da resistência em populações de R. (B.) microplus. Vacinas contra o carrapato representam uma tecnologia que pode ser combinada com acaricidas em programas de controle integrado para diminuir o impacto de R. (B.) microplus. A forma recombinante da Bm86 da cepa Campo Grande (rBm86-CG) de R. (B.) microplus foi produzido utilizando o sistema de expressão em Pichia pastoris para testar sua capacidade de imunoproteção ao gado contra a infestação de carrapatos. A secreção de rBm86-CG em P. pastoris pelo bioprocesso, simplificou a purificação do antígeno. A resposta imune humoral específica foi detectada por ELISA em soros de bovinos vacinados. Resultados de "imunoblot" revelaram que anticorpos policlonais de bovinos vacinados reconheceram uma proteína em extratos de larvas com um peso molecular correspondente à Bm86. O antígeno rBm86-CG mostrou eficácia de 31% contra a amostra CG de R. (B.) microplus utilizada para infestar os bovinos vacinados. Pelos resultados obtidos, concluímos que a rBm86-CG é um antígeno que pode ser usado em uma vacina polivalente, como parte de um programa integrado para o controle de R. (B.) microplus no estado do Mato Grosso do Sul, Brasil.

Immunological studies against ixodid ticks, Hyalomma anatolicum anatolicum (Linnaeus, 1758) were conducted on six-month old domestic goat breed Capra hircus lehri, randomly selected from different farms of districts of Balochistan, Pakistan. Goats were allocated into four groups of twenty animals each for immunization experiments. Each group was used to assess the effect of immunization against ticks Hyalomma a. anatolicum by using extracts of attachment cones made up of cement proteins taken from mouthparts of tick whole instars as source of antigen. Effects of different antigen doses emulsified with different adjuvants were analyzed in four groups, each cohort comprising twenty goats. First group was immunized with alum adjuvant, second with Freund's complete adjuvant (FCA), third with Incomplete Freund's adjuvant (IFA) and fourth with Montanide ISA-50 (Incomplete Seppic) adjuvant. Goats administered with different doses (5-500 µg) of tick cement antigen emulsified with Montanide adjuvant showed effective immune response through reduction in tick engorgement (decreased weight) and increased percentage tick rejection which proved that Montanide adjuvant is the most potent adjuvant. Results also showed that lower doses of tick cement antigen (5 µg of antigen/ml of adjuvant/animal) emulsified with Montanide ISA-50 adjuvant induced a better stimulatory effect on percentage tick rejection. [ABSTRACT FROM AUTHOR]

To identify suitable targets for development of cross-protective tick vaccine, in silico analysis was attempted and male tick derived molecule, voraxin-α was targeted. The voraxin- α homologue of Rhipicephalus ( Boophilus) microplus was cloned, sequenced and analyzed employing standard methods. The deduced amino acids sequence analysis of the 419 bp cloned voraxin-α gene of R. ( B.) microplus indicated very high (94.6 %) similarity with voraxin- α of the R. appendiculatus and moderate to low identity with Amblyomma hebraeum, Dermacentor silvarum and Haemaphysalis longicornis. The results suggest that recombinant voraxin-α might be a good candidate as cross-protective anti-tick vaccine. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Highlights: [•] Subolesin orthologs of two major ticks were cloned and sequenced. [•] Calves were immunized with recombinant Subolesin. [•] Moderate protective vaccine efficacy (40%) was obtained against R. (B.) microplus. [ABSTRACT FROM AUTHOR]

Abstract: Prior exposure of vertebrate hosts to tick salivary proteins can induce specific immunity to tick infestation, as well as affording protection against tick-transmitted Borrelia burgdorferi infection in the mammalian host. Vaccination using an adenovirus expression system to deliver 4 tick salivary proteins (Ad-Salps) derived from Ixodes scapularis, Salp15, Salp25A, Salp25D, and Isac, was explored. Results indicate that vaccination with tick salivary proteins in an adenoviral vector can be used to modulate a Th1 response in the host and partially control spirochete load in immunized mice after infected tick challenge. [Copyright &y& Elsevier]

Abstract: Vaccines are among the alternative tick control methods expected to replace at least in part the volumes of chemical acaricides currently used worldwide. However, a vaccination approach depends on a host immune response against proteins that are essential to tick physiology. The cystatin family is a protein class recently investigated to compose an effective antigen in a tick vaccine. In this study, a cDNA from Rhipicephalus appendiculatus with high sequence similarity to cystatins type 2 was identified by random sequencing analysis and called R. appendiculatus cystatin 1 (Ra-cyst-1). DNA sequence analysis showed that the cloned Ra-cyst-1 has a 423-bp open reading frame and codified to a 140-amino acid polypeptide. The putative mature protein consists of 115 amino acid residues with a deduced molecular weight of 12.8kDa. The highly conserved G (P-I), QxVxG (P-II), and PW (P-III) type 2 cystatins motifs are present in Ra-cyst-1 cDNA. RT-PCR analysis showed that the Ra-cyst-1 gene is expressed in nymph, male, and female midgut following blood feeding, but not in the salivary glands of fed females. In addition, Western blot revealed that recombinant Ra-cyst-1 was not recognized by sera derived from rabbits infested with ticks, suggesting that this cystatin is not secreted into the host during infestation. We hypothesize that Ra-cyst-1 may play a role in the tick feeding process and could be a concealed antigen candidate in further anti-tick vaccination trials. [Copyright &y& Elsevier]