Location: Veterinary Pest Genetics Research Unit
2022 Annual Report
Objectives
Objective 1: Perform bioinformatic analysis of tick and fly genomes to find new targets of control and methods of surveillance.
Objective 2: Develop genetic methods to control ticks and dipteran pests of livestock and wildlife, to include new vaccines through reverse vaccinology, gene editing/silencing, gene drives, and other genetic approaches to mitigate acaricide resistance.
Approach
Genetic studies on arthropods of medical and veterinary importance helps identify and elucidate gene activities in target organisms. Genomic techniques provide information on population genetics and support identifying invasive insect source populations and migration pathways, the development of gene targets for disruption or RNAi gene suppression, identifying pest species that are difficult to identify morphologically, reverse vaccinology for the discovery of new antigen candidates, and more rapid discovery of natural enemies using next generation sequencing and metagenomics. Research on fly and tick metagenomes (the entire genomic complement found in the environment) and microbiomes (the universe of microbes living in association with each pest) generates information about pathogens and commensal and symbiotic microorganisms that can be used to solve agricultural problems associated with these organisms. Scientists have worked with U.S. university partners and international institutions to generate ‘omic resources such as transcriptomes, microbiomes, and genomes for a number of livestock and human pests, including biting midges, mosquitoes, and house flies. Mosquito mitochondrial genomes sequenced using nanopore technology support the development of databases for rapid identification of field samples. Sequencing of pooled RNA-Seq is valuable for gene expression analysis of pathogens that cause disease in humans, livestock, and other animals. Understanding of genetics and bionomics are critical in the developing sustainable integrated pest management (IPM) programs. Defining the problem and selecting appropriate control strategies requires obtaining background information on pest identification (systematics and taxonomy), distribution (spatial and temporal), and behavior (particularly behaviors that cause or have the potential to cause damage). Fundamental research on pest genetics generates information that can be used to identify weaknesses of the pest; findings can also be used to help develop models that assess entomological and/or epidemiological risk to host populations. Genetic studies of pests are rapidly generating a wealth of information that can be used to develop new and adaptive pest control measures using CRISPR CAS9 and Gene Drive technologies. In practice, each genetically based control method is applied individually and locally, so it is especially challenging to project if or how laboratory results will be replicated in integrated field studies. Developing new genetic and chemical pest control measures that result in commercial IPM products often requires coordinating stakeholder efforts, funding, and other resources.
Progress Report
Project #3094-22320-001-000D, Genetics of Veterinary Pests, is a new project beginning in fiscal year (FY) 2022 as the result of a realignment Program Direction and Resource Allocation Memo (PDRAM)(Nov. 2021). Objective 1 is newly created (no FY22 milestones) and Objective 2 was transferred from Project #3094-32000-042-000D, Integrated Pest Management of Cattle Fever Ticks.
In support of Objective 1, perform bioinformatic analysis of tick and fly genomes to find new targets of control and methods of surveillance, ARS scientists at Kerrville, Texas, utilized the newly acquired Pacbio and Illumina sequencers to complete the sequencing of two Dermacentor and two Amblyomma tick genomes. These genomes are currently being assembled into scaffolds, and the transcriptomes of these respective species are being sequenced to generate gene annotations. In addition, the unit has completed the sequencing of the stable fly, horn fly, and face fly. These genomes are similarly undergoing assembly and annotation. Direct applications of these genomes are currently being applied to population genetic datasets and gene expression studies to better understand the underlying biological processes involved in host-pest interactions.
In support of Subobjective 2A, evaluate the efficacy of novel acaricides and delivery systems against ticks on livestock, and through the targeted treatment of infested pastures and protected and sensitive habitats, research conducted by ARS scientists at Kerrville, Texas, in collaboration with scientists at Texas A&M University in College Station, Texas, demonstrated that silencing expression of the pyrokinin receptor gene in adult R. microplus ticks reduced tick survival and reproduction providing strong evidence that interfering with pyrokinin function in ticks has potential for the development of novel acaricides. Further, research conducted by ARS scientists at Kerrville, Texas, in collaboration with ARS scientists at Fort Pierce, Florida, has presented evidence that gene silencing in ticks may be adaptable as a novel method of tick control.
ARS researchers at Kerrville, Texas, selected specific recombinant DNA molecules encoding specific tick antigens as potential anti-tick vaccine antigen candidates in support of Subobjective 2B, identify candidate antigens for anti-tick vaccines and formulate as vaccines for animal trials. Computational methods and reverse vaccinology algorithms were used to initially screen over 75,000 transcripts and rank their anti-tick vaccine potential. Ten protein antigen candidates were selected for recombinant protein expression in P. pastoris. Two of these candidates were then successfully purified in milligram quantities sufficient to formulate vaccine doses that will be utilized to conduct cattle stall tests to evaluate vaccine efficacy against R. microplus.
In support of Subobjective 2C, compare the genomes of R. microplus, R. annulatus, and Haemaphysalis longicornis to identify sex determination genes for the development of genetic control methods, researchers at Edinburg, Texas, conducted a comparative analysis of sex determining genes utilizing published datasets from 17 species of hard ticks including R. microplus, R. annulatus, H. longicornis, D. andersoni, A. maculatum, R. sanguineus, and I. scapularis. Two Ornithodoros soft tick species and four species of mites were also included in the analysis. This analysis will be used to identify sex determination pathway gene orthologues for comparison to R. microplus.
Accomplishments
1. Metabolic enzyme transcripts sequenced from pesticide resistant horn flies. The horn fly is a blood-feeding parasite of cattle causing over $800 million in economic losses in the U.S. each year. ARS researchers in Kerrville, Texas, and Parlier, California, collaborated with researchers at Abilene Christian University, Louisiana State University, and the National Center for Genome Resources to develop a comprehensive dataset of metabolic enzyme transcripts from populations of pesticide resistant horn flies. Specific enzyme transcripts were identified that are responsible for pesticide detoxification. Identification of these enzyme transcripts may lead to tools to help combat pesticide resistance.
2. RNAi silencing of pyrokinin receptor negatively affects reproduction in female fever ticks. Cattle fever ticks have global economic impact as ectoparasites of cattle and are known to harbor pathogens that cause diseases in cattle like cattle fever and anaplasmosis. ARS researchers in Kerrville and Edinburg, Texas, in collaboration with Texas A&M University, demonstrated that silencing expression of the pyrokinin receptor gene in adult female fever ticks reduced tick survival and reproduction providing evidence that interfering with pyrokinin function in ticks could be exploited for the development of novel acaricides.
Review Publications
Bendele, K.G., Bodine, D.L., Xu, Q., Foil, L.D., Cameron, C., Perez De Leon, A.A., Farmer, A., Retzel, E., Moore, V., Lohmeyer, K.H., Guerrero, F. 2022. The adult horn fly transcriptome and the complement of transcripts encoding cytochrome P450s, glutathione S-transferases, and esterases. Veterinary Parasitology. https://doi.org/10.1016/j.vetpar.2022.109699.
Leal-Galvan, B., Harvey, C., Thomas, D.B., Saelao, P., Oliva Chavez, A. 2022. A method for the isolation of miRNAs from tick ex vivo salivary gland cultures and extracellular vesicles. The Journal of Visualized Experiments (JoVE). https://doi.org/10.3791/63618.
Wulff, J.P., Temeyer, K.B., Tidwell, J.P., Schlechte, K.G., Lohmeyer, K.H., Pietrantonio, P.V. 2022. Pyrokinin receptor silencing in females of the southern cattle tick Rhipicephalus (Boophilus) microplus is associated with a reproductive fitness cost. Parasites & Vectors. https://doi.org/10.1186/s13071-022-05349-w.
