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Evaluation and comparison of the potential of two ferritins as anti-tick vaccines against Haemaphysalis longicornis.

Galay RL, Miyata T, Umemiya-Shirafuji R, Maeda H, Kusakisako K, Tsuji N, Mochizuki M, Fujisaki K, Tanaka T - Parasit Vectors (2014)

Bottom Line: To demonstrate the effects of antibodies, oxidative stress was detected in the eggs and larvae.Reduced oviposition and hatching rate were observed in both rHlFER-immunized groups. rHlFER2 showed a higher vaccine efficacy.Collectively, these results showed that HlFER2 has a good potential as an anti-tick vaccine antigen that may affect multiple tick species.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathological and Preventive Veterinary Science, The United Graduate School of Veterinary Science, Yamaguchi University, Yoshida, Yamaguchi, 753-8515, Japan. rlgalay.dvm@gmail.com.

ABSTRACT

Background: Tick control is an essential aspect of controlling the spread of tick-borne diseases affecting humans and animals, but it presently faces several challenges. Development of an anti-tick vaccine is aimed at designing cost-effective and environmentally friendly protection against ticks and tick-borne diseases as an alternative to the use of chemical acaricides. A single vaccine from the tick midgut protein Bm86 is currently available for field applications, but its efficacy is limited to only some tick species. Identification of candidate vaccine antigens that can affect multiple tick species is highly desirable. The hard tick Haemaphysalis longicornis has two kinds of the iron-binding protein ferritin (HlFER), an intracellular HlFER1 and a secretory HlFER2, and RNA interference experiments showed that these are physiologically important in blood feeding and reproduction and in protection against oxidative stress. Here we investigated the potential of targeting HlFERs for tick control by immunizing the host with recombinant HlFERs (rHlFER1 and rHlFER2).

Methods: Rabbits were immunized with rHlFERs three times subcutaneously at two-week intervals. Antisera were collected before the first immunization and a week after each immunization to confirm the antigen-specific serum antibody titer by serum ELISA. Two weeks after the final immunization, the rabbits were challenged with tick infestation. After dropping, tick feeding and reproduction parameters were evaluated to determine vaccine efficacy. To demonstrate the effects of antibodies, oxidative stress was detected in the eggs and larvae.

Results: The antibody titer of rHlFER-immunized rabbits greatly increased after the second immunization. Antibodies exhibited cross-reactivity with rHlFERs and reacted with tick native HlFERs in Western blot analysis. Significantly lower bodyweight was observed in the ticks infested from the rHlFER2-immunized rabbit compared to those from the control rabbit. Reduced oviposition and hatching rate were observed in both rHlFER-immunized groups. rHlFER2 showed a higher vaccine efficacy. The antibodies against rHlFERs were detected in the eggs, and higher levels of oxidative stress biomarkers in the eggs and larvae, of ticks from rHlFER vaccinated rabbits.

Conclusion: Collectively, these results showed that HlFER2 has a good potential as an anti-tick vaccine antigen that may affect multiple tick species.

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Related in: MedlinePlus

Purification of recombinant HlFERs (rHlFERs). His-tagged rHlFERs (rHlFER1 and rHlFER2) were expressed in E. coli and then purified through Ni-affinity chromatography and dialysis against PBS. After refolding, 2 μg per protein was subjected to SDS-PAGE, and then the gel was stained with Coomassie brilliant blue. M, low molecular weight marker.
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Fig1: Purification of recombinant HlFERs (rHlFERs). His-tagged rHlFERs (rHlFER1 and rHlFER2) were expressed in E. coli and then purified through Ni-affinity chromatography and dialysis against PBS. After refolding, 2 μg per protein was subjected to SDS-PAGE, and then the gel was stained with Coomassie brilliant blue. M, low molecular weight marker.

Mentions: rHlFERs were expressed and purified from E. coli cells. SDS-PAGE showed that the purified rHlFER1 and rHlFER2 have a similar molecular mass of around 26 kDa (Figure 1) as described previously [17]. An antigen-specific ELISA was conducted to monitor the antibody titer of individual rabbits, and then the average antibody titer for each group from the two trials was calculated (Figure 2). Whereas no changes were observed in the antibody titers of the control group, the antibody titers of the groups immunized against rHlFER1 (Figure 2A) and rHlFER2 (Figure 2B) significantly increased after the second immunization. Furthermore, the antibodies also exhibited cross-reactivity to each antigen. rHlFER1-immunized rabbits showed an abrupt increase in antibody titer against rHlFER1 after the second immunization, which further increased after the third immunization, while the titer for rHlFER2 only significantly increased after the third immunization. For rHlFER2-immunized rabbits, the antibody titer against rHlFER2 abruptly increased after the second immunization but did not significantly increase further after the third immunization, while the level and the trend of the antibody titer against rHlFER1 were similar to those of the rHlFER1-immunized rabbits. Cross-reactivity of the antibodies was also observed on ELISA using rHlPrx2 as antigen but it was lower than that observed in HlFERs (Additional file 1: Figure S1).Figure 1


Evaluation and comparison of the potential of two ferritins as anti-tick vaccines against Haemaphysalis longicornis.

Galay RL, Miyata T, Umemiya-Shirafuji R, Maeda H, Kusakisako K, Tsuji N, Mochizuki M, Fujisaki K, Tanaka T - Parasit Vectors (2014)

Purification of recombinant HlFERs (rHlFERs). His-tagged rHlFERs (rHlFER1 and rHlFER2) were expressed in E. coli and then purified through Ni-affinity chromatography and dialysis against PBS. After refolding, 2 μg per protein was subjected to SDS-PAGE, and then the gel was stained with Coomassie brilliant blue. M, low molecular weight marker.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4197249&req=5

Fig1: Purification of recombinant HlFERs (rHlFERs). His-tagged rHlFERs (rHlFER1 and rHlFER2) were expressed in E. coli and then purified through Ni-affinity chromatography and dialysis against PBS. After refolding, 2 μg per protein was subjected to SDS-PAGE, and then the gel was stained with Coomassie brilliant blue. M, low molecular weight marker.
Mentions: rHlFERs were expressed and purified from E. coli cells. SDS-PAGE showed that the purified rHlFER1 and rHlFER2 have a similar molecular mass of around 26 kDa (Figure 1) as described previously [17]. An antigen-specific ELISA was conducted to monitor the antibody titer of individual rabbits, and then the average antibody titer for each group from the two trials was calculated (Figure 2). Whereas no changes were observed in the antibody titers of the control group, the antibody titers of the groups immunized against rHlFER1 (Figure 2A) and rHlFER2 (Figure 2B) significantly increased after the second immunization. Furthermore, the antibodies also exhibited cross-reactivity to each antigen. rHlFER1-immunized rabbits showed an abrupt increase in antibody titer against rHlFER1 after the second immunization, which further increased after the third immunization, while the titer for rHlFER2 only significantly increased after the third immunization. For rHlFER2-immunized rabbits, the antibody titer against rHlFER2 abruptly increased after the second immunization but did not significantly increase further after the third immunization, while the level and the trend of the antibody titer against rHlFER1 were similar to those of the rHlFER1-immunized rabbits. Cross-reactivity of the antibodies was also observed on ELISA using rHlPrx2 as antigen but it was lower than that observed in HlFERs (Additional file 1: Figure S1).Figure 1

Bottom Line: To demonstrate the effects of antibodies, oxidative stress was detected in the eggs and larvae.Reduced oviposition and hatching rate were observed in both rHlFER-immunized groups. rHlFER2 showed a higher vaccine efficacy.Collectively, these results showed that HlFER2 has a good potential as an anti-tick vaccine antigen that may affect multiple tick species.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathological and Preventive Veterinary Science, The United Graduate School of Veterinary Science, Yamaguchi University, Yoshida, Yamaguchi, 753-8515, Japan. rlgalay.dvm@gmail.com.

ABSTRACT

Background: Tick control is an essential aspect of controlling the spread of tick-borne diseases affecting humans and animals, but it presently faces several challenges. Development of an anti-tick vaccine is aimed at designing cost-effective and environmentally friendly protection against ticks and tick-borne diseases as an alternative to the use of chemical acaricides. A single vaccine from the tick midgut protein Bm86 is currently available for field applications, but its efficacy is limited to only some tick species. Identification of candidate vaccine antigens that can affect multiple tick species is highly desirable. The hard tick Haemaphysalis longicornis has two kinds of the iron-binding protein ferritin (HlFER), an intracellular HlFER1 and a secretory HlFER2, and RNA interference experiments showed that these are physiologically important in blood feeding and reproduction and in protection against oxidative stress. Here we investigated the potential of targeting HlFERs for tick control by immunizing the host with recombinant HlFERs (rHlFER1 and rHlFER2).

Methods: Rabbits were immunized with rHlFERs three times subcutaneously at two-week intervals. Antisera were collected before the first immunization and a week after each immunization to confirm the antigen-specific serum antibody titer by serum ELISA. Two weeks after the final immunization, the rabbits were challenged with tick infestation. After dropping, tick feeding and reproduction parameters were evaluated to determine vaccine efficacy. To demonstrate the effects of antibodies, oxidative stress was detected in the eggs and larvae.

Results: The antibody titer of rHlFER-immunized rabbits greatly increased after the second immunization. Antibodies exhibited cross-reactivity with rHlFERs and reacted with tick native HlFERs in Western blot analysis. Significantly lower bodyweight was observed in the ticks infested from the rHlFER2-immunized rabbit compared to those from the control rabbit. Reduced oviposition and hatching rate were observed in both rHlFER-immunized groups. rHlFER2 showed a higher vaccine efficacy. The antibodies against rHlFERs were detected in the eggs, and higher levels of oxidative stress biomarkers in the eggs and larvae, of ticks from rHlFER vaccinated rabbits.

Conclusion: Collectively, these results showed that HlFER2 has a good potential as an anti-tick vaccine antigen that may affect multiple tick species.

Show MeSH
Related in: MedlinePlus