Detection of amitraz resistance in Rhipicephalus (Boophilus) microplus from SBS Nagar, Punjab, India.
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ABSTRACT:
SUBMITTER: Singh NK
PROVIDER: S-EPMC3934582 | biostudies-other | 2014
REPOSITORIES: biostudies-other
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Project description:The cattle tick Rhipicephalus microplus is one of the most important ectoparasites with great sanitary and economic impact for cattle rearing worldwide. Ivermectin is commonly used to control tick populations, but its use over the last 30 years has led to the development of resistant populations of R. microplus, and a concomitant loss of efficacy. In this context, we aimed to determine the metabolic mechanisms that contribute to ivermectin resistance in a resistant strain of this species. We performed lethal time bioassays with inhibitors of detoxifying enzymes and xenobiotic transporters (four detoxification pathways) using two strains of ticks: a susceptible strain, Mozo, and a resistant strain, Juarez. We used four inhibitors to test the involvement of different families of proteins responsible for detoxification of ivermectin, namely cytochrome P450, esterases, glutathione-S-transferase, and ATP Binding Cassette Transporters. We calculated the synergistic factor for each inhibitor and strain. To different degrees, all tested inhibitors altered the mortality rates in the strain Juarez, indicating that multiple mechanisms are responsible for the resistant phenotype. Detoxification mechanisms mediated by ABC transporters were observed to be the most important. Esterases, glutathione-S-transferases, and cytochrome-oxidases played less important roles in detoxification.
| S-EPMC6097998 | biostudies-literature
Project description:Susceptibility to synthetic pyrethroids (SP s) and the role of two major resistance mechanisms were evaluated in Mexican Rhipicephalus microplus tick populations. Larval packet test (LPT), knock-down (kdr) PCR allele-specific assay (PASA) and esterase activity assays were conducted in tick populations for cypermethrin, flumethrin and deltamethrin. Esterase activity did not have a significant correlation with SP s resistance. However a significant correlation (p < 0.01) was found between the presence of the sodium channel mutation, and resistance to SP s as measured by PASA and LPT respectively. Just over half the populations (16/28) were cross-resistant to flumethrin, deltamethrin and cypermethrine, 21.4% of the samples (6/28) were susceptible to all of the three pyrethroids 10.7 of the samples (3/28) were resistant to flumethrin, 3.4 of the samples (1/28) were resistant to deltamethrin only and 7.1% (2/28) were resistant to flumethrin and deltamethrin. The presence of the kdr mutation correlates with resistance to the SP s as a class. Target site insensitivity is the major mechanism of resistance to SP s in Mexican R. microplus field strains, involving the presence of a sodium channel mutation, however, esterase-based, other mutations or combination of mechanisms can also occur.
| S-EPMC2729983 | biostudies-literature
Project description:A R. microplus microarray was used to study differential gene expression in acaricide exposed larvae from an amitraz-resistant strain. The acaricide treatments were: organophosphate (OP), pyrethroid, ivermectin, and amitraz. The microarrays contained over 13,000 probes corresponding to each member of R. microplus gene index ESTs previously described (http://compbio.dfci.harvard.edu/tgi/cgi-bin/tgi/gimain.pl?gudb=b_microplus). Serial analysis of gene expression (SAGE) data from the OP treated R. microplus was used to verify the OP microarray data. The expression profiles of selected transcripts were verified by real time PCR. Among the significantly differentially expressed genes, were a tick legumain, involved in blood digestion, gluthathione S-transferase (GST), a detoxification enzyme involved in pesticide resistance, acyltransferase, several putative salivary sulfotransferases, and a glutamate receptor. **Note: contact person: Felix D. Guerrero email: felix.guerrero@ars.usda.gov Keywords: Rhipicephalus (Boophilus) microplus, acaricide resistance genes, organophosphates OP, microarrays, detoxification enzymes.
2008-12-31 | GSE10171 | GEO
Project description: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.
| S-EPMC4815859 | biostudies-literature
Project description:BACKGROUND: The southern cattle tick, Rhipicephalus (Boophilus) microplus, is an economically important parasite of cattle and can transmit several pathogenic microorganisms to its cattle host during the feeding process. Understanding the biology and genomics of R. microplus is critical to developing novel methods for controlling these ticks. RESULTS: We present a global comparative genomic analysis of a gene index of R. microplus comprised of 13,643 unique transcripts assembled from 42,512 expressed sequence tags (ESTs), a significant fraction of the complement of R. microplus genes. The source material for these ESTs consisted of polyA RNA from various tissues, lifestages, and strains of R. microplus, including larvae exposed to heat, cold, host odor, and acaricide. Functional annotation using RPS-Blast analysis identified conserved protein domains in the conceptually translated gene index and assigned GO terms to those database transcripts which had informative BlastX hits. Blast Score Ratio and SimiTri analysis compared the conceptual transcriptome of the R. microplus database to other eukaryotic proteomes and EST databases, including those from 3 ticks. The most abundant protein domains in BmiGI were also analyzed by SimiTri methodology. CONCLUSION: These results indicate that a large fraction of BmiGI entries have no homologs in other sequenced genomes. Analysis with the PartiGene annotation pipeline showed 64% of the members of BmiGI could not be assigned GO annotation, thus minimal information is available about a significant fraction of the tick genome. This highlights the important insights in tick biology which are likely to result from a tick genome sequencing project. Global comparative analysis identified some tick genes with unexpected phylogenetic relationships which detailed analysis attributed to gene losses in some members of the animal kingdom. Some tick genes were identified which had close orthologues to mammalian genes. Members of this group would likely be poor choices as targets for development of novel tick control technology.
| S-EPMC2100071 | biostudies-literature
Project description:The southern cattle fever tick, Rhipicephalus (Boophilus) microplus, is the most economically important ectoparasite of cattle worldwide. A limitation for sustainable control and eradication is the emergence of acaricide resistance among tick populations. Molecular diagnostic tools offer the opportunity to detect resistance rapidly, which can be complemented with confirmatory bioassays with larvae and adult ticks that are more resource and time consuming to generate. Synthetic pyrethroid resistance is one of the most prevalent and well-studied forms of resistance in arthropods, being linked with target site alterations in the sodium ion channel gene. Here, we report research on a novel molecular method to detect mutations in the para-sodium channel gene of R. microplus associated with acaricide resistance that is based on quantitative PCR high-resolution melt (HRM) analysis. Genomic DNA fragments of domains II and III of the para-sodium channel gene were amplified by real-time PCR in the presence of EVA®Green dye to test resistant and susceptible reference ticks from the U.S., Brazil, and Mexico. Larval packet tests with discriminating doses and a modified lethal time analysis were performed to confirm resistance to permethrin, cypermethrin, deltamethrin, and flumethrin in laboratory strains. Tick specimens collected from cattle that were inspected at the United States Port-of-Entry at the Texas-Mexico border were also genotyped. Previously described mutations associated with pyrethroid resistance (T170C, C190A, G184C, and T2134A) were successfully detected by qPCR-HRM in different genotypes and confirmed by sequencing. A novel non-synonymous SNP located at domain III (C2136A) and the G215T mutation in domain II, previously described only in Asian R. microplus and R. australis, were also detected with the HRM and confirmed by sequencing. This technique could be adapted for high-throughput screening, detection, and discovery of allele-specific mutations in cattle tick outbreak populations to inform eradication strategies in the USA. This knowledge could also be applied to integrated control programs in other parts of the world where R. microplus is endemic and where similar SNPs have been identified associated with pyrethroid resistance. This study highlights the existence of several mutations in the para-sodium channel gene in different combinations in field populations of R. microplus from Mexico.
| S-EPMC6423475 | biostudies-literature
Project description:Ticks are blood-feeding arthropods that can affect human and animal health both directly by blood-feeding and indirectly by transmitting pathogens. The cattle tick Rhipicephalus (Boophilus) microplus is one of the most economically important ectoparasites of bovines worldwide and it is responsible for the transmission of the protozoan Babesia bovis, the etiological agent of bovine babesiosis. Aquaporins (AQPs) are water channel proteins implicated in physiological mechanisms of osmoregulation. Members of the AQP family are critical for blood-feeding arthropods considering the extreme osmoregulatory changes that occur during their feeding. We investigated the pattern of expression of a newly identified AQP2 gene of R. microplus (RmAQP2) in different tick tissues and stages. We also examined in vivo the biological implications of silencing expression of RmAQP2 silencing during tick feeding on either uninfected or B. bovis-infected cattle.In silico gene analyses were performed by multiple alignments of amino acid sequences and topology prediction. Levels of RmAQP2 transcripts in different tick tissues and stages were analyzed by reverse transcriptase quantitative PCR. Patterns of expression of RmAQP2 protein were investigated by immunoblots. Gene silencing was performed by RNA interference and in vivo functional analyses carried out by feeding ticks on either uninfected or B. bovis-infected cattle.RmAQP2 transcripts were found in unfed larvae, engorged nymphs, and salivary glands and guts of partially engorged females; however, of all tick tissues and stages examined, RmAQP2 protein was found only in salivary glands of partially engorged females. RmAQP2 silencing significantly reduced tick fitness and completely abrogated protein expression. The effect of RmAQP2 silencing on fitness was more pronounced in females fed on a B. bovis-infected calf than in ticks fed on an uninfected calf and none of their larval progeny survived.Collectively, considering the gene expression and tick fitness data, we conclude that RmAQP2 is critical for tick blood feeding and may be a suitable candidate target for the development of novel strategies to control R. microplus and tick-borne parasites.
| S-EPMC4667534 | biostudies-literature
Project description:This study intends to characterize the sialotranscriptome profile of Rhipicephalus (Boophilus) microplus in response to Theileria equi and identify genes of interest with differential genomic expression, indicating relevant targets in the tick-protozoan interactions. The experimental design consisted of RNA sequencing from uninfected and T. equi-infected R. microplus salivary glands (SGs) to obtain transcriptomic profiles for characterization and comparison. A total of 288,952 transcripts were obtained from both tick profiles, 3456 transcripts (p < 0.05) differentially expressed in response to T. equi infection. The uninfected SGs' registered 231,179 transcripts, of which 155,359 were annotated. The most transcribed sequences were female-specific histamine binding protein and lipocalins. Regarding the T. equi-infected SGs, from the 238,964 assembled transcripts, 163,564 were annotated. The most transcribed sequences were histone demethylase JARID1 and Y-box-binding protein. Five transcripts (cystatin, arginase, nuclear factor κB kinase inhibitor subunit β (IκB), IκB delta, lysosomal-trafficking regulator, and reeler protein) presented the gene ontology (GO) category "response to protozoan" and were exclusively displayed in the T. equi-infected profile. The transcriptome of T. equi was also analyzed, registering 4728 hits. The study's genetic and molecular information would be of great value for future studies and biotechnological applications envisaging disease control.
| S-EPMC7913801 | biostudies-literature
Project description:BACKGROUND:Rhipicephalus (Boophilus) microplus (Rmi) a major cattle ectoparasite and tick borne disease vector, impacts on animal welfare and industry productivity. In arthropod research there is an absence of a complete Chelicerate genome, which includes ticks, mites, spiders, scorpions and crustaceans. Model arthropod genomes such as Drosophila and Anopheles are too taxonomically distant for a reference in tick genomic sequence analysis. This study focuses on the de-novo assembly of two R. microplus BAC sequences from the understudied R microplus genome. Based on available R. microplus sequenced resources and comparative analysis, tick genomic structure and functional predictions identify complex gene structures and genomic targets expressed during tick-cattle interaction. RESULTS:In our BAC analyses we have assembled, using the correct positioning of BAC end sequences and transcript sequences, two challenging genomic regions. Cot DNA fractions compared to the BAC sequences confirmed a highly repetitive BAC sequence BM-012-E08 and a low repetitive BAC sequence BM-005-G14 which was gene rich and contained short interspersed elements (SINEs). Based directly on the BAC and Cot data comparisons, the genome wide frequency of the SINE Ruka element was estimated. Using a conservative approach to the assembly of the highly repetitive BM-012-E08, the sequence was de-convoluted into three repeat units, each unit containing an 18S, 5.8S and 28S ribosomal RNA (rRNA) encoding gene sequence (rDNA), related internal transcribed spacer and complex intergenic region.In the low repetitive BM-005-G14, a novel gene complex was found between to 2 genes on the same strand. Nested in the second intron of a large 9 Kb papilin gene was a helicase gene. This helicase overlapped in two exonic regions with the papilin. Both these genes were shown expressed in different tick life stage important in ectoparasite interaction with the host. Tick specific sequence differences were also determined for the papilin gene and the protein binding sites of the 18S subunit in a comparison to Bos taurus. CONCLUSION:In the absence of a sequenced reference genome we have assembled two complex BAC sequences, characterised novel gene structure that was confirmed by gene expression and sequencing analyses. This is the first report to provide evidence for 2 eukaryotic genes with exon regions that overlap on the same strand, the first to describe Rhipicephalinae papilin, and the first to report the complete ribosomal DNA repeated unit sequence structure for ticks. The Cot data estimation of genome wide sequence frequency means this research will underpin future efforts for genome sequencing and assembly of the R. microplus genome.
| S-EPMC3160391 | biostudies-literature
Project description:BACKGROUND:The cattle tick, Rhipicephalus (Boophilus) microplus, economically impact cattle industry in tropical and subtropical regions of the world. The morphological and genetic differences among R. microplus strains have been documented in the literature, suggesting that biogeographical and ecological separation may have resulted in boophilid ticks from America/Africa and those from Australia being different species. To test the hypothesis of the presence of different boophilid species, herein we performed a series of experiments to characterize the reproductive performance of crosses between R. microplus from Australia, Africa and America and the genetic diversity of strains from Australia, Asia, Africa and America. RESULTS:The results showed that the crosses between Australian and Argentinean or Mozambican strains of boophilid ticks are infertile while crosses between Argentinean and Mozambican strains are fertile. These results showed that tick strains from Africa (Mozambique) and America (Argentina) are the same species, while ticks from Australia may actually represent a separate species. The genetic analysis of mitochondrial 12S and 16S rDNA and microsatellite loci were not conclusive when taken separately, but provided evidence that Australian tick strains were genetically different from Asian, African and American strains. CONCLUSION:The results reported herein support the hypothesis that at least two different species share the name R. microplus. These species could be redefined as R. microplus (Canestrini, 1887) (for American and African strains) and probably the old R. australis Fuller, 1899 (for Australian strains), which needs to be redescribed. However, experiments with a larger number of tick strains from different geographic locations are needed to corroborate these results.
| S-EPMC2656471 | biostudies-literature