A multi-omics approach for understanding blood digestion dynamics in Ixodes scapularis and identification of anti-tick vaccine targets.

Ixodes scapularis, the black-legged tick, is a major arthropod vector that transmits the causative agents of Lyme disease and several other pathogens of human significance. The tick midgut is the main tissue involved in blood acquisition and digestion and the first organ to have contact with pathogens ingested through the blood meal. Gene expression in the midgut before, during, and after a blood meal may vary in response to the physiological changes due to blood feeding. A systems biology approach based on RNA and protein sequencing was used to gain insight into the changes in tick midgut transcripts and proteins during blood ingestion (unfed and partially fed) and digestion (1-, 2-, 7-, and 14 days post detachment from the host) by the Ixodes scapularis female ticks. A total of 2,726 differentially expressed transcripts, and 449 proteins were identified across the time points. Genes involved in detoxification of xenobiotics, proteases, protease inhibitors, metabolism, and immunity were differentially expressed in response to blood feeding. Similarly, proteins corresponding to the same groups were also differentially expressed. Nine genes from major gene categories were chosen as potential vaccine candidates, and, using RNA interference, the effect of these gene knockdowns on tick biology was investigated. Knockdown of these genes had variable negative impacts on tick physiology, such as the inability to engorge fully and to produce eggs and increased mortality. These and additional gene targets provide opportunities to explore novel tick control strategies.

The highly improved genome of Ixodes scapularis with X and Y pseudochromosomes.

Ixodes scapularis, the black-legged tick, is the principal vector of the Lyme disease spirochete, Borrelia burgdorferi, and is responsible for most of the ∼470,000 estimated Lyme disease cases annually in the USA. Ixodes scapularis can transmit six additional pathogens of human health significance. Because of its medical importance, I. scapularis was the first tick genome to be sequenced and annotated. However, the first assembly, I. scapularis Wikel (IscaW), was highly fragmented because of the technical challenges posed by the long, repetitive genome sequences characteristic of arthropod genomes and the lack of long-read sequencing techniques. Although I. scapularis has emerged as a model for tick research because of the availability of new tools such as embryo injection and CRISPR-Cas9-mediated gene editing yet the lack of chromosome-scale scaffolds has slowed progress in tick biology and the development of tools for their control. Here we combine diverse technologies to produce the I. scapularis Gulia-Nuss (IscGN) genome assembly and gene set. We used DNA from eggs and male and female adult ticks and took advantage of Hi-C, PacBio HiFi sequencing, and Illumina short-read sequencing technologies to produce a chromosome-level assembly. In this work, we present the predicted pseudochromosomes consisting of 13 autosomes and the sex pseudochromosomes: X and Y, and a markedly improved genome annotation compared with the existing assemblies and annotations.

Cas9-mediated gene editing in the black-legged tick, Ixodes scapularis, by embryo injection and ReMOT Control.

Despite their capacity to acquire and pass on an array of debilitating pathogens, research on ticks has lagged behind other arthropod vectors, such as mosquitoes, largely because of challenges in applying available genetic and molecular tools. CRISPR-Cas9 is transforming non-model organism research; however, successful gene editing has not yet been reported in ticks. Technical challenges for injecting tick embryos to attempt gene editing have further slowed research progress. Currently, no embryo injection protocol exists for any chelicerate species, including ticks. Herein, we report a successful embryo injection protocol for the black-legged tick, Ixodes scapularis, and the use of this protocol for genome editing with CRISPR-Cas9. We also demonstrate that the ReMOT Control technique could be successfully used to generate genome mutations outside Insecta. Our results provide innovative tools to the tick research community that are essential for advancing our understanding of the molecular mechanisms governing pathogen transmission by tick vectors and the underlying biology of host-vector-pathogen interactions.

Ecology of Ixodes pacificus Ticks and Associated Pathogens in the Western United States.

Lyme disease is the most important vector-borne disease in the United States and is increasing in incidence and geographic range. In the Pacific west, the western black-legged tick, Ixodes pacificus Cooley and Kohls, 1943 is an important vector of the causative agent of Lyme disease, the spirochete, Borrelia burgdorferi. Ixodes pacificus life cycle is expected to be more than a year long, and all three stages (larva, nymph, and adult) overlap in spring. The optimal habitat consists of forest cover, cooler temperatures, and annual precipitation in the range of 200-500 mm. Therefore, the coastal areas of California, Oregon, and Washington are well suited for these ticks. Immature stages commonly parasitize Western fence lizards (Sceloporus occidentalis) and gray squirrels (Sciurus griseus), while adults often feed on deer mice (Peromyscus maniculatus) and black-tailed deer (Odocoileus h. columbianus). Ixodes pacificus carry several pathogens of human significance, such as Borrelia burgdorferi, Bartonella, and Rickettsiales. These pathogens are maintained in the environment by many hosts, including small mammals, birds, livestock, and domestic animals. Although a great deal of work has been carried out on Ixodes ticks and the pathogens they transmit, understanding I. pacificus ecology outside California still lags. Additionally, the dynamic vector-host-pathogen system means that new factors will continue to arise and shift the epidemiological patterns within specific areas. Here, we review the ecology of I. pacificus and the pathogens this tick is known to carry to identify gaps in our knowledge.