Location: Livestock Arthropod Pests Research2019 Annual Report
Objective 1: Sequencing and annotation of the genome of the horn fly and cattle fever tick. Subobjective 1A: Assembly and annotation of the cattle tick genome sequence. Subobjective 1B: Sequencing, assembly, and annotation of the genome sequence of the horn fly. Objective 2: Investigate molecular-based control and surveillance technologies. Subobjective 2A: Identify candidate antigens for anti-biting fly and anti-tick vaccines and formulate as vaccines for animal trials. Subobjective 2B: Identification of gene-based mechanisms of pesticide resistance and develop gene-based surveillance assays to monitor gene flow. Objective 3: Increase sequence information and genetic annotation of livestock pests, focusing on biological aspects likely to be affected by climate change.
Utilize advanced bioinformatic assembly and annotation protocols to attain a draft annotated R. microplus genome sequence of sufficient quality for publication in international peer-reviewed journals. The assembled and annotated sequence will be made available for the scientific community at CattleTickBase (http://ccg.murdoch.edu.au/cattletickbase) and GenBank. Sequence the horn fly genomic DNA with similar protocols and caveats utilized to sequence the tick genome. The horn fly genomic DNA to be sequenced will be obtained from a laboratory colony maintained since 1961 at our laboratory, reared in cages and feeding upon cotton pads saturated with citrated bovine blood. The assembled sequence will be available for the scientific community by submission of the data to GenBank. Identify candidate vaccine antigens through reverse vaccinology from datasets obtained from prior project or as part of objective 1 of this project. Utilize real-time PCR to study metabolic-based pesticide resistance and quantify gene expression of specific horn fly cytochrome P450s in populations of horn flies with known metabolic resistance-based mechanisms. Use a transcriptomic approach to sequence nuclear and mitochondrial genes from tick collections in Indian and Philippine collections to use for phylogenetic comparisons to the Texas outbreak R. microplus Deutsch population that was used in prior R. microplus transcriptome studies.
Accomplishments contributed to efforts by the ARS Veterinary Pest Genomics Center and included obtaining assembled and annotated genome and transcriptome sequences of the southern cattle fever tick, Rhipicephalus microplus, an important biological vector of cattle diseases, and the horn fly, Haematobia irritans, an economically important blood-feeding fly pest of cattle. The R. microplus genome contained over twice the amount of DNA as the human genome. Analysis of the horn fly genome identified and described genes involved in pesticide resistance and metabolism, sex determination, and blood feeding. Sequences of those livestock pests enabled research on novel control and surveillance technologies such as vaccines and molecular assays to track pesticide resistance. Several ARS patents covered sequences coding for R. microplus proteins that could be used as antigens in vaccine formulations. Some of these ARS-patented R. microplus antigens were tested for efficacy through agreements with industry partners. Likewise, the horn fly genome was mined applying reverse vaccinology to identify candidate antigens that could be produced in recombinant form for efficacy tests. The genome of the cattle fever tick, R. annulatus, which is the other fever tick species and vector of the microbes causing bovine babesiosis that threatens U.S. animal agriculture, was also sequenced. This achievement provided the unprecedented opportunity to conduct comparative genomic analysis that help understand the evolutionary history of R. microplus and R. annulatus. This information allows the identification of vulnerabilities in fever ticks that can be the target of sustainable eradication interventions. The Asian longhorned tick, Haemaphysalis longicornis, is a known serious pest of livestock and vector of several pathogens of livestock and humans where it is established. This invasive tick was detected in the U.S. in 2017. Sequencing of the H. longicornis genome was done using specimens from New Zealand. Genomic approaches refined during this project will be applied to innovate tactics against H. longicornis including vaccines. This impactful research project advanced the science of invasive tick genomics and delivered algorithms to mine genome data for the development of the next generation of tick management tools.
1. Comparative genomics of cattle fever ticks threatening the U.S. livestock industry. The cattle fever tick (CFT), R. annulatus, is an obligate ectoparasite of livestock and one of the fever tick species that threaten U.S. animal agriculture because of its vector ability to transmit the microbes causing bovine babesiosis, which is a malaria-like deadly disease of cattle. Collaborative efforts by ARS scientists in Kerrville, Texas, with the Veterinary Pest Genomics Center unraveled the CFT genome. Sequencing this genome provided access to all the information needed to build and maintain the CFT. This was done to translate genomic information and innovate technologies the Cattle Fever Tick Eradication Program can use to keep the U.S. free of CFT in a sustainable manner. A way to do this is through comparative genomics using sequences from the southern CFT discovered before and applied to research and develop anti-fever tick vaccines.
2. Functional genomics through RNA interference discovers paths to innovate cattle fever tick control technologies. Resistance to conventional acaricides offers the opportunity for discovery research that can be translated into safer technologies to keep the U.S. free of cattle fever ticks (CFT). Silencing genes through RNA interference (RNAi) helps validate targets for novel and safer acaricides, and it can serve as a platform to innovate tick control technologies. Silencing the gene coding for the leucokinin-like peptide receptor of the southern cattle fever tick by ARS scientists in Kerrville, Texas, in collaboration with academic partners resulted in decreased egg production and egg hatching, and increased time to lay eggs and for eggs to hatch. Disrupting the function of the leucokinin-like peptide receptor presents a way to control fever tick populations. Other experiments showed that advanced RNAi technology could be developed to control cattle fever ticks. Modified antisense oligonucleotides were shown to be biochemically stable and amenable to use in a drug delivery system that could be used to test for efficacy against fever tick infestation in cattle.
3. Advancements in our understanding of the molecular biology of cattle fever tick acetylcholinesterases. It was hypothesized that the presence of an active acetylcholinesterase (AChE) in the saliva of the southern cattle fever tick might be involved in the immunoregulation of the host response to tissue damage during blood feeding and thereby promote the transmission of disease-causing agents, or pathogens, like those that cause bovine babesiosis. ARS scientists in Kerrville, Texas, obtained further evidence consistent with this hypothetical paradigm by demonstrating that multiple arthropods and biological vectors of disease including several tick species, mosquitoes, and sand flies contain AChE in their saliva, whereas non-biological vectors biting arthropods such as horn flies and stable flies that also feed on blood lack salivary AChE. Functional studies are underway to test the effect of recombinant tick AChE activity on activation and subsequent developmental responses of various immune cell subsets. Science-based knowledge from confirmatory evidence that salivary AChE plays a role at the tick-host interface during blood feeding could be used to innovate tick control technologies that also block the transmission of fever tick-borne pathogens.
Domingues, L.N., Guerrero, F., Cameron, C., Farmer, A., Bendele, K.G., Foil, L.D. 2018. The assembled transcriptome of the adult horn fly, Haematobia irritans. Data in Brief. 19:1933-1940.
Brito, L., Barbieri, F., Rocha, R., Santos, A., Silva, R., Ribeiro, M., Guerrero, F., Foil, L., Oliveira, M. 2018. Pyrethroid and organophosphate pesticide resistance in field populations of horn fly in Brazil. Medical and Veterinary Entomology. 33:121-130. https://doi.org/10.1111/mve.12330.
Gundersen, D.E., Adrianos, S.L., Allen, M.L., Becnel, J.J., Chen, Y., Choi, M.Y., Estep, A., Evans, J.D., Garczynski, S.F., Geib, S.M., Ghosh, S.B., Handler, A.M., Hasegawa, D.K., Heerman, M.C., Hull, J.J., Hunter, W.B., Kaur, N., Li, J., Li, W., Ling, K., Nayduch, D., Oppert, B.S., Perera, O.P., Perkin, L.C., Sanscrainte, N.D., Sim, S.B., Sparks, M., Temeyer, K.B., Vander Meer, R.K., Wintermantel, W.M., James, R.R., Hackett, K.J., Coates, B.S. 2017. Arthropod genomics research in the United States Department of Agriculture-Agricultural Research Service: Applications of RNA interference and CRISPR gene editing technologies in pest control. Trends in Entomology. 13:109-137.
Brock, C.M., Temeyer, K.B., Tidwell, J.P., Yang, Y., Blandon, M.A., Carreon-Camacho, D., Longnecker, M.T., Almazan, C., Perez De Leon, A.A., Pietrantonio, P.V. 2019. The leucokinin-like peptide receptor from the cattle fever tick, Rhipicephalus microplus, is localized in the midgut periphery and receptor silencing with validated dsRNAs causes a reproductive fitness cost. International Journal for Parasitology. 49(3-4):287-299. https://doi.org/10.1016/j.ijpara.2018.11.006.