Polymorphism of the bm86 Gene in South American Strains of the Cattle Tick Boophilus microplus

Sossai, Sidimar; Peconick, Ana Paula; Sales-Junior, Policarpo A.; Marcelino, F. C.; Vargas, Marlene; Neves, Elisangela S.; Patarroyo, J. H.

doi:10.1007/s10493-005-3262-7

Cited by 43 publications

(27 citation statements)

References 22 publications

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“…The number and location of isolates selected reflects the proportion of current outbreaks in their respective counties, as well as the number of female ticks submitted through the CFTEP that were available for oviposition. This number of isolates is also comparable to that used in previous descriptions of Bm86 geographic sequence variability [11]. The majority of documented infestations in South Texas are R. microplus , and this is reflected in the number of isolates sequenced: 24 R. microplus and 3 R. annulatus (Table 1).…”

Section: Resultssupporting

confidence: 81%

Bm86 midgut protein sequence variation in South Texas cattle fever ticks

et al. 2010

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BackgroundCattle fever ticks, Rhipicephalus (Boophilus) microplus and R. (B.) annulatus, vector bovine and equine babesiosis, and have significantly expanded beyond the permanent quarantine zone established in South Texas. Currently, there are no vaccines approved for use within the United States for controlling these vectors. Vaccines developed in Australia and Cuba based on the midgut antigen Bm86 have variable efficacy against cattle fever ticks. A possible explanation for this variation in vaccine efficacy is amino acid sequence divergence between the recombinant Bm86 vaccine component and native Bm86 expressed in ticks from different geographical regions of the world.ResultsThere was 91.8% amino acid sequence identity in Bm86 among R. microplus and R. annulatus sequenced from South Texas infestations. When South Texas isolates were compared to the Australian Yeerongpilly and Cuban Camcord vaccine strains, there was 89.8% and 90.0% identity, respectively. Most of the sequence divergence was focused in one region of the protein, amino acids 206-298. Hydrophilicity profiles revealed that two short regions of Bm86 (amino acids 206-210 and 560-570) appear to be more hydrophilic in South Texas isolates compared to vaccine strains. Only one amino acid difference was found between South Texas and vaccine strains within two previously described B-cell epitopes. A total of 4 amino acid differences were observed within three peptides previously shown to induce protective immune responses in cattle.ConclusionsSequence differences between South Texas isolates and Yeerongpilly and Camcord strains are spread throughout the entire Bm86 sequence, suggesting that geographic variation does exist. Differences within previously described B-cell epitopes between South Texas isolates and vaccine strains are minimal; however, short regions of hydrophilic amino acids found unique to South Texas isolates suggest that additional unique surface exposed peptides could be targeted.

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Section: Resultssupporting

confidence: 81%

Bm86 midgut protein sequence variation in South Texas cattle fever ticks

et al. 2010

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“…This sequence was not found in any of the five sequenced clones and may be less abundant, have been the result of alternative splicing or represent an allele which was lost in the R. microplus tick population since the source material was collected several generations earlier. The presence of Bm86 alleles within the same tick population has been previously reported [18]. Two alleles, Ra86-1 and Ra86-2, were also found to be transcribed in the midgut of R. appendiculatus females.…”

Section: Discussionsupporting

confidence: 70%

Selection of reference genes for quantitative RT-PCR studies in Rhipicephalus (Boophilus) microplus and Rhipicephalus appendiculatus ticks and determination of the expression profile of Bm86

et al. 2009

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BackgroundFor accurate and reliable gene expression analysis, normalization of gene expression data against reference genes is essential. In most studies on ticks where (semi-)quantitative RT-PCR is employed, normalization occurs with a single reference gene, usually β-actin, without validation of its presumed expression stability. The first goal of this study was to evaluate the expression stability of commonly used reference genes in Rhipicephalus appendiculatus and Rhipicephalus (Boophilus) microplus ticks. To demonstrate the usefulness of these results, an unresolved issue in tick vaccine development was examined. Commercial vaccines against R. microplus were developed based on the recombinant antigen Bm86, but despite a high degree of sequence homology, these vaccines are not effective against R. appendiculatus. In fact, Bm86-based vaccines give better protection against some tick species with lower Bm86 sequence homology. One possible explanation is the variation in Bm86 expression levels between R. microplus and R. appendiculatus. The most stable reference genes were therefore used for normalization of the Bm86 expression profile in all life stages of both species to examine whether antigen abundance plays a role in Bm86 vaccine susceptibility.ResultsThe transcription levels of nine potential reference genes: β-actin (ACTB), β-tubulin (BTUB), elongation factor 1α (ELF1A), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), glutathione S-transferase (GST), H3 histone family 3A (H3F3A), cyclophilin (PPIA), ribosomal protein L4 (RPL4) and TATA box binding protein (TBP) were measured in all life stages of R. microplus and R. appendiculatus. ELF1A was found to be the most stable expressed gene in both species following analysis by both geNorm and Normfinder software applications, GST showed the least stability. The expression profile of Bm86 in R. appendiculatus and R. microplus revealed a more continuous Bm86 antigen abundance in R. microplus throughout its one-host life cycle compared to the three-host tick R. appendiculatus where large variations were observed between different life stages.ConclusionBased on these results, ELF1A can be proposed as an initial reference gene for normalization of quantitative RT-PCR data in whole R. microplus and R. appendiculatus ticks. The observed differences in Bm86 expression profile between the two species alone can not adequately explain the lack of a Bm86 vaccination effect in R. appendiculatus.

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“…(B.) microplus, Sossai et al (2005) suggested that geographic isolation is not the sole determinant of genomic variation. It has been suggested that diversity greater than 3% in amino acid sequence between the recombinant antigen and native Bm86 protein in target tick populations results in vaccination inefficiencies .…”

Section: Introductionmentioning

confidence: 99%

Bovine immunoprotection against Rhipicephalus (Boophilus) microplus with recombinant Bm86-Campo Grande antigen

Cunha

León

Pinto

et al. 2012

Rev. Bras. Parasitol. Vet.

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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 Pichia pastoris 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.Keywords: Tick, Rhipicephalus (Boophilus) microplus, anti-tick vaccine, recombinant Bm86, Pichia pastoris. ResumoO 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.

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Polymorphism of the bm86 Gene in South American Strains of the Cattle Tick Boophilus microplus

Cited by 43 publications

References 22 publications

Bm86 midgut protein sequence variation in South Texas cattle fever ticks

Bm86 midgut protein sequence variation in South Texas cattle fever ticks

Selection of reference genes for quantitative RT-PCR studies in Rhipicephalus (Boophilus) microplus and Rhipicephalus appendiculatus ticks and determination of the expression profile of Bm86

Bovine immunoprotection against Rhipicephalus (Boophilus) microplus with recombinant Bm86-Campo Grande antigen

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