Wildlife are hosts of ectoparasites, such as fleas and ticks that may transmit human and animal pathogens. Little is known about the ecology of many ectoparasite species native to southern Texas, or their role in pathogen maintenance and transmission. Much attention has been given to the role of nonnative nilgai antelope as cattle fever tick hosts and agents of dispersal, but little attention has been given to other ectoparasites that may utilize nilgai antelope as hosts. As southern Texas is a hot-spot for flea-borne (murine) typhus, it is important to examine flea species presence, abundance, and host use in this region. Fleas were opportunistically collected during wildlife depredation activities, from hunter-harvested animals, or during handling in the course of other research activities in several southern Texas counties. A total of 9 wildlife species were sampled, from which 3 flea species were identified. A total of 83 Pulex porcinus (Jordan and Rothschild) were collected from nilgai, coyotes, bobcats, javelina, feral swine, and a black-tailed jackrabbit. In total, 9 Euhoplopsyllus glacialis affinis (Baker) were collected from cottontail rabbits, and 1 Echidnophaga gallinacea (Westwood) was collected from a raccoon. To our knowledge, this represents the first report of fleas from nilgai antelope. Pulex porcinus, although often considered a specialist species, was collected from a wide range of hosts, including 2 (nilgai antelope and black-tailed jackrabbit) that represent new host records for this species. The role of P. porcinus as a pathogen vector is unknown, but its apparent abundance in this region warrants further investigation.

Rhipicephalus (Boophilus) microplus is globally one of the most economically important ectoparasites of cattle costing the agriculture industry billions of dollars annually. Resistance to chemical control measures has prompted the development of novel methods of control. Recent advancements in genetic control measures for human and other animal vectors have utilized sex determination research to manipulate sex ratios, which have shown promising results in mosquitoes, namely Aedes aegypti and Anopheles stephensi. Here, we use R. (B.) microplus as a model to provide foundational research to allow similar avenues of investigation in ticks using R. (B.) microplus as a model. Karyotypes for R. (B.) microplus show an XX:XO sex determining system with the largest chromosome being the sex chromosome. Using flow cytometric methods, the size of the sex chromosome was estimated at 526.91 Mb. All measures to identify the sex chromosome within the cattle tick genome assembly associated sex chromosomal characteristics to 2 chromosomes. This discrepancy between the assembly and karyotypes of the tick led to generating a new genome assembly with a single adult male specimen. The 2 chromosomes in question aligned with a single scaffold within the new genome that had a length of 513.29 Mb and was the first time the sex chromosome was identified in an Ixodid genome assembly.

Abstract Background The cattle fever tick, Rhipicephalus (Boophilus) microplus, is a vector of pathogens causative of babesiosis and anaplasmosis, both highly lethal bovine diseases that affect cattle worldwide. In Ecdysozoa, neuropeptides and their G-protein-coupled receptors play a critical integrative role in the regulation of all physiological processes. However, the physiological activity of many neuropeptides is still unknown in ticks. Periviscerokinins (CAP2b/PVKs) are neuropeptides associated with myotropic and diuretic activities in insects. These peptides have been identified only in a few tick species, such as Ixodes ricinus, Ixodes scapularis and R. microplus, and their cognate receptor only characterized for the last two. Methods Expression of the periviscerokinin receptor (Rhimi-CAP 2b R) was investigated throughout the developmental stages of R. microplus and silenced by RNA interference (RNAi) in the females. In a first experiment, three double-stranded (ds) RNAs, named ds680-805, ds956-1109 and ds1102-1200, respectively, were tested in vivo. All three caused phenotypic effects, but only the last one was chosen for subsequent experiments. Resulting RNAi phenotypic variables were compared to those of negative controls, both non-injected and dsRNA beta-lactamase-injected ticks, and to positive controls injected with beta-actin dsRNA. Rhimi-CAP 2b R silencing was verified by quantitative reverse-transcriptase PCR in whole females and dissected tissues. Results Rhimi-CAP 2b R transcript expression was detected throughout all developmental stages. Rhimi-CAP 2b R silencing was associated with increased female mortality, decreased weight of surviving females and of egg masses, a delayed egg incubation period and decreased egg hatching (P < 0.05). Conclusions CAP2b/PVKs appear to be associated with the regulation of female feeding, reproduction and survival. Since the Rhimi-CAP 2b R loss of function was detrimental to females, the discovery of antagonistic molecules of the CAP2b/PVK signaling system should cause similar effects. Our results point to this signaling system as a promising target for tick control. Graphical Abstract

Abstract Background Rhipicephalus microplus is the vector of deadly cattle pathogens, especially Babesia spp., for which a recombinant vaccine is not available. Therefore, disease control depends on tick vector control. However, R. microplus populations worldwide have developed resistance to available acaricides, prompting the search for novel acaricide targets. G protein-coupled receptors (GPCRs) are involved in the regulation of many physiological processes and have been suggested as druggable targets for the control of arthropod vectors. Arthropod-specific signaling systems of small neuropeptides are being investigated for this purpose. The pyrokinin receptor (PKR) is a GPCR previously characterized in ticks. Myotropic activity of pyrokinins in feeding-related tissues of Rhipicephalus sanguineus and Ixodes scapularis was recently reported. Methods The R. microplus pyrokinin receptor (Rhimi-PKR) was silenced through RNA interference (RNAi) in female ticks. To optimize RNAi, a dual-luciferase assay was applied to determine the silencing efficiency of two Rhimi-PKR double-stranded RNAs (dsRNA) prior to injecting dsRNA in ticks to be placed on cattle. Phenotypic variables of female ticks obtained at the endpoint of the RNAi experiment were compared to those of control female ticks (non-injected and beta-lactamase dsRNA-injected). Rhimi-PKR silencing was verified by quantitative reverse-transcriptase PCR in whole females and dissected tissues. Results The Rhimi-PKR transcript was expressed in all developmental stages. Rhimi-PKR silencing was confirmed in whole ticks 4 days after injection, and in the tick carcass, ovary and synganglion 6 days after injection. Rhimi-PKR silencing was associated with an increased mortality and decreased weight of both surviving females and egg masses (P < 0.05). Delays in repletion, pre-oviposition and incubation periods were observed (P < 0.05). Conclusions Rhimi-PKR silencing negatively affected female reproductive fitness. The PKR appears to be directly or indirectly associated with the regulation of female feeding and/or reproductive output in R. microplus. Antagonists of the pyrokinin signaling system could be explored for tick control. Graphical abstract

The southern cattle fever tick (SCFT) Rhipicephalus (Boophilus) microplus, is considered the most important ectoparasite of livestock in the world because of high financial losses associated with direct feeding and transmission of the hemoparasites Babesia bovis, B. bigemina, and Anaplasma marginale. Unfortunately, SCFT in many parts of the world have evolved resistance to all market-available pesticides thus driving development of new control technologies. Vaccination against ticks using the tick gut protein Bm86 has been shown to be effective against acaricide-resistant ticks. This technique has been successfully implemented in Puerto Rico for the control of acaricide-resistant R. microplus on dairy and beef cattle. Observations from Puerto Rico indicate a potentially positive interaction between anti-tick vaccination when used in conjunction with systemic acaricide treatment. In this project, controlled animal studies were completed directly comparing efficacy of anti-tick vaccination with and without systemic acaricide. Results show that the Bm86 anti-tick vaccine in combination with the macrocyclic lactone, Moxidectin, expressed a synergistic interaction, providing greater and longer efficacy than either treatment alone.

Ticks from the genus Rhipicephalus have enormous global economic impact as ectoparasites of cattle. Rhipicephalus microplus and Rhipicephalus annulatus are known to harbor infectious pathogens such as Babesia bovis, Babesia bigemina, and Anaplasma marginale. Having reference quality genomes of these ticks would advance research to identify druggable targets for chemical entities with acaricidal activity and refine anti-tick vaccine approaches. We sequenced and assembled the genomes of R. microplus and R. annulatus, using Pacific Biosciences and HiSeq 4000 technologies on very high molecular weight genomic DNA. We used 22 and 29 SMRT cells on the Pacific Biosciences Sequel for R. microplus and R. annulatus, respectively, and 3 lanes of the Illumina HiSeq 4000 platform for each tick. The PacBio sequence yields for R. microplus and R. annulatus were 21.0 and 27.9 million subreads, respectively, which were assembled with Canu v. 1.7. The final Canu assemblies consisted of 92,167 and 57,796 contigs with an average contig length of 39,249 and 69,055 bp for R. microplus and R. annulatus, respectively. Annotated genome quality was assessed by BUSCO analysis to provide quantitative measures for each assembled genome. Over 82% and 92% of the 1066 member BUSCO gene set was found in the assembled genomes of R. microplus and R. annulatus, respectively. For R. microplus, only 189 of the 1066 BUSCO genes were missing and only 140 were present in a fragmented condition. For R. annulatus, only 75 of the BUSCO genes were missing and only 109 were present in a fragmented condition. The raw sequencing reads and the assembled contigs/scaffolds are archived at the National Center for Biotechnology Information.

Ticks are pests and vectors of diseases that are of public health and veterinary importance. The cattle tick, Rhipicephalus microplus (Canestrini, 1888), is one of the most studied tick species because of its impact on livestock health and production in the tropical and subtropical parts of the world, costing the cattle industry billions annually. Control methods have evolved throughout the years but so has R. microplus. Reliance upon chemical control has created a consistent need to develop new technologies to overcome the pesticide resistance that occurs as the ticks adapt. In order to utilize the more advanced tools such as RNAi or Crispr/Cas9 systems, tick tissues need to be isolated and manipulated. Unfortunately, there are a limited number of dissection guides available providing a detailed view of tick internal anatomy. This manual includes photomicrographs to guide the dissection of R. microplus adults, male and female. Topography and anatomical differences between the internal organs of unfed and gravid adult females are described. We were able to locate the crucial tissues for cattle tick physiology and lay out spatial and temporal guidelines for their identification and dissection. Examples of how this information can be used at the nexus between organismal and molecular research to innovate tick control technologies is discussed.

Acetylcholinesterase (AChE) was previously reported to be present in saliva of the southern cattle tick, Rhipicephalus (Boophilus) microplus (Canestrini), with proposed potential functions to 1) reduce acetylcholine toxicity during rapid engorgement, 2) modulate host immune responses, and 3) to influence pathogen transmission and establishment in the host. Potential modulation of host immune responses might include participation in salivary-assisted transmission and establishment of pathogens in the host as has been reported for a number of arthropod vector-borne diseases. If the hypothesis that tick salivary AChE may alter host immune responses is correct, we reasoned that similar cholinesterase activities might be present in saliva of additional arthropod vectors. Here, we report the presence of AChE-like activity in the saliva of southern cattle ticks, Rhipicephalus (Boophilus) microplus; the lone star tick, Amblyomma americanum (Linnaeus); Asian tiger mosquitoes, Aedes albopictus (Skuse); sand flies, Phlebotomus papatasi (Scopoli); and biting midges, Culicoides sonorensis Wirth and Jones. Salivary AChE-like activity was not detected for horn flies Haematobia irritans (L.), stable flies Stomoxys calcitrans (L.), and house flies Musca domestica L. Salivary cholinesterase (ChE) activities of arthropod vectors of disease-causing agents exhibited various Michaelis-Menten KM values that were each lower than the KM value of bovine serum AChE. A lower KM value is indicative of higher affinity for substrate and is consistent with a hypothesized role in localized depletion of host tissue acetylcholine potentially modulating host immune responses at the arthropod bite site that may favor ectoparasite blood-feeding and alter host defensive responses against pathogen transmission and establishment.

Rhipicephalus annulatus field populations collected from small cattle farms in Beni-Suef province in Egypt were evaluated for deltamethrin resistance by toxicological in vitro bioassays (adult immersion test-AIT and larval packet test-LPT). Moreover, a quantitative PCR high resolution melting (PCR-HRM) technique was used to detect nucleotide substitutions in the voltage-gated sodium channel (Na-channel) gene. By the in vitro bioassays, the examined ticks were phenotypically categorized as deltamethrin susceptible (populations El-Wasta -A, and El-Hakamna -C) or resistant (populations El-Wasta -B, El-Hakamna -D, EL-Halabia -E, and Kom-abokhalad -F). Based on LPT findings, the phenotypic resistant populations were found to have a resistance ratio between 6.5 - 10.8. The PCR-HRM genotyping of the ticks showed variable melting curves among the populations in domain II of the Na-channel gene. Analysis of the curves showed the presence of wild type, mutant homozygous, and mutant heterozygous tick individuals. By sequencing the PCR amplified fragments, the C190A mutation was the only detected nucleotide polymorphism of domain II among the phenotypically resistant populations, which was present in 39.5 % (34/86) of the ticks tested. On the other hand, the phenotypically susceptible populations A and C did not show C190A mutant homozygous (RR) individuals. Meanwhile, in domain III all of the examined populations revealed melting curves like the wild type. Furthermore, the sequence analysis of these populations confirmed the absence of SNPs in domain III. The C190A single point mutation was detected for the first time in domain II of the Na-channel gene of deltamethrin-resistant R. annulatus in Egypt using PCR-HRM. Screening for efficacy of chemical compounds used by farmers to control ticks on cattle should be considered as part of animal health programs to manage the emerging resistance to acaricides in R. annulatus populations.

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.

The cattle fever tick, Rhipicephalus microplus (Canestrini) (Acari: Ixodidae), is a one-host tick that infests primarily cattle in tropical and sub-tropical regions of the world. This species transmits deadly cattle pathogens, especially Babesia spp., for which a recombinant vaccine is not available. Therefore, disease control depends on tick vector control. Although R. microplus was eradicated in the USA, tick populations in Mexico and South America have acquired resistance to many of the applied acaricides. Recent acaricide-resistant tick reintroductions detected in the U.S. underscore the need for novel tick control methods. The octopamine and tyramine/octopamine receptors, both G protein-coupled receptors (GPCR), are believed to be the main molecular targets of the acaricide amitraz. This provides the proof of principle that investigating tick GPCRs, especially those that are invertebrate-specific, may be a feasible strategy for discovering novel targets and subsequently new anti-tick compounds. The R. microplus leucokinin-like peptide receptor (LKR), also known as the myokinin- or kinin receptor, is such a GPCR. While the receptor was previously characterized in vitro, the function of the leucokinin signaling system in ticks remains unknown. In this work, the LKR was immunolocalized to the periphery of the female midgut and silenced through RNA interference (RNAi) in females. To optimize RNAi experiments, a dual-luciferase system was developed to determine the silencing efficiency of LKR-double stranded RNA (dsRNA) constructs prior to testing those in ticks placed on cattle. This assay identified two effective dsRNAs. Silencing of the LKR with these two validated dsRNA constructs was verified by quantitative real time PCR (qRT-PCR) of female tick dissected tissues. Silencing was significant in midguts and carcasses. Silencing caused decreases in weights of egg masses and in the percentages of eggs hatched per egg mass, as well as delays in time to oviposition and egg hatching. A role of the kinin receptor in tick reproduction is apparent.

The cattle tick, Rhipicephalus annulatus (Say) is a vector of bovine babesiosis and responsible for direct and indirect losses to cattle producing areas located in temperate and subtropical dry regions. Resistance against pyrethroids has been reported for this species in Asia and Africa, but never before in North America. An outbreak strain, Rio Lado, collected close to the border between Mexico and the United States, in Maverick County, Texas, showed low level of resistance to permethrin, a pyrethroid pesticide. We used genomic material from different strains of cattle ticks collected within the Permanent Quarantine Zone (Rio Lado, Vega and Klein Grass strains) to partially characterize the coding gene of the voltage-gated sodium channel (Na-channel), target-site of pyrethroid pesticides, and search for putative mutations associated with resistance using quantitative PCR high resolution melt (HRM) analysis. The two amplified fragments, corresponding to domains II and III of the Na-channel, were 100 % identical to its ortholog in Rhipicephalus microplus (Canestrini). No nucleotide polymorphisms in the Na-channel gene were observed in the pyrethroid-resistant Rio Lado strain, when compared to the susceptible strains Klein Grass and Vega. This study reports the first case of pyrethroid resistance in R. annulatus collected in the United States. Also, we provide new genomic data for this species of tick that allows for the development of a new method to screen for mutations associated with pyrethroid resistance.

Amitraz is an important product for the control of cattle ticks around the world. In comparison with other products for the control of ticks, it is quite affordable and it has a rapid knock-down effect. It binds with and activates adrenergic neuro-receptors of animals and it inhibits the action of monoamine oxidases (MAO). Resistance to amitraz has been documented in Rhipicephalus microplus, R. decoloratus and R. appendiculatus. Four mechanisms of resistance have been proposed, each of which is supported by evidence but none of which has been definitively confirmed as the cause of resistance in the field. The proposed mechanisms include genetic target site insensitivity in two G protein-coupled receptors, the beta-adrenergic octopamine receptor (BAOR) and the octopamine/tyramine receptor (OCT/Tyr), increased expression or activity of monoamine oxidases and increased expression or activity of the ATP binding cassette transporter.

The southern cattle fever tick, Rhipicephalus microplus Canestrini, is widely distributed in tropical and subtropical regions, causing huge economic losses to milk and meat production. Application of acaricides is the most widely used control strategy, but because of increasing evidence of resistance to acaricide and environmental concerns, novel strategies are needed. Use of entomopathogenic nematodes is a promising alternative for tick control. Six species of nematodes, Steinernema riobrave Cabanillas, Poinar, and Raulston (355 strain), S. carpocapsae (Weiser) (All strain), S. feltiae Filipjev (SN strain), Heterorhabditis bacteriophora Poinar (VS strain), H. indica Poinar, Karunakar, and David (HOM1 strain), and H. floridensis Nguyen, Gozel, Koppenhofer, and Adams (K22 strain) were tested for pathogenicity against R. microplus (Deutch strain). Engorged female ticks were exposed to 1,250, 2,500, and 5,000 infective juvenile nematodes for 24, 48, and 72 hours in Petri dishes, and tick mortality over time, pre-oviposition period, egg mass weight, reproductive index, percentage inhibition of oviposition, and hatching percentage were evaluated. Dose-dependent mortality was recorded in Heterorhabditis species S. riobrave and S. carpocapsae which increased with exposure time. Minimum LC50 (95% CL) values of 1999 (1947-2052) and 69 (63-75) infective juveniles per Petri dish were calculated for S. riobrave and H. floridensis, respectively, at 72 hours of exposure. Infective juvenile concentration and exposure time did not influence the pre-oviposition period. Differences in egg mass weight and reproductive index among the treatments and check group were observed with respect to concentration and exposure time. Reduction in hatchability was recorded only in ticks exposed to S. riobrave. Overall, H. floridensis was the most virulent species against engorged R. microplus females, adversely impacting most biological parameters. S. riobrave showed potential especially because it is native to South Texas and commercially produced. Future studies are warranted to evaluate the potential of this nematode species for biological control of the cattle fever tick under field conditions.

The brown dog tick, Rhipicephalus sanguineus (Latrielle) sensu lato, is an important ectoparasite of dogs and occasionally humans, capable of transmitting several pathogens, such as Rickettsia and Ehrlichia, which are of veterinary and medical importance. The brown dog tick is distributed worldwide and has an affinity for human habitations in much of its range. In some populations, lack of integrated pest management plans and overuse of pyrethroid pesticides and other sodium channel inhibitors has resulted in high levels of resistance to permethrin. Recently, a highly conserved region of the R. sanguineus sodium channel was sequenced, indicating that a single nucleotide polymorphism of thymine to cytosine on domain III segment VI of the sodium channel could confer resistance. A molecular assay targeting a point mutation in the sodium channel was developed and optimized to separate ticks expressing permethrin resistance from those from a susceptible colony. Thereafter, multiple field-collected phenotypically permethrin-resistant populations were evaluated using this molecular assay to determine genotype. As confirmed by DNA sequencing, a point mutation was present at a high rate in phenotypically resistant tick populations that was not present in the susceptible strain. These data suggest an additional permethrin resistance mechanism to metabolic resistance, which has been reported for this tick species, and confirm its association with phenotypic resistance. The results of this study further emphasize the need to preserve acaricide chemistry through rotation of active ingredients used to control ectoparasites.

The brown dog tick, Rhipicephalus sanguineus sensu lato (Latreille), is a cosmopolitan ectoparasite and vector of pathogens that kill humans and animals. Pyrethroids represent a class of synthetic acaricides that have been used intensely to try to control the brown dog tick and mitigate the risk of tick-borne disease transmission. However, acaricide resistance is an emerging problem in the management of the brown dog tick. Understanding the mechanism of resistance to acaricides, including pyrethroids, is important to adapt brown dog tick control strategies. The main objective of this study was to determine if target-site mutations associated with pyrethroid resistance in other pests could be associated with phenotypic resistance detected in a brown dog tick population from Florida. We amplified segment 6 of the domain III of the voltage-sensitive sodium channel protein, using cDNAs synthesized from pyrethroid-susceptible and pyrethroid-resistant tick strains. A single nucleotide point mutation (SNP) identified in a highly conserved region of domain III S6 in the resistant ticks resulted in an amino acid change from phenylalanine to leucine. This mutation is characteristic of resistance phenotypes in other tick species, and is the first report of this mutation in R. sanguineus. Molecular assays based on this knowledge could be developed to diagnose the risk for pyrethroid resistance, and to inform decisions on integrated brown dog tick management practices.

The giant reed, Arundo donax, is a perennial grass species that has become an invasive plant in many countries. Expansive stands of A. donax have significant negative impacts on available water resources and efforts are underway to identify biological control agents against this species. The giant reed grows under adverse environmental conditions, displaying insensitivity to drought stress, flooding, heavy metals, salinity and herbaceous competition, thus hampering control programs. To establish a foundational molecular dataset, we used an llumina Hi-Seq protocol to sequence the transcriptome of actively growing shoots from an invasive genotype collected along the Rio Grande River, bordering Texas and Mexico. We report the assembly of 27,491 high confidence transcripts (≥200 bp) with at least 70% coverage of known genes in other Poaceae species. Of these 13,080 (47.58%), 6165 (22.43%) and 8246 (30.0%) transcripts have sequence similarity to known, domain-containing and conserved hypothetical proteins, respectively. We also report 75,590 low confidence transcripts supported by both trans-ABBySS and Velvet-Oases de novo assembly pipelines. Within the low confidence subset of transcripts we identified partial hits to known (19,021; 25.16%), domain-containing (7093; 9.38%) and conserved hypothetical (16,647; 22.02%) proteins. Additionally 32,829 (43.43%) transcripts encode putative hypothetical proteins unique to A. donax. Functional annotation resulted in 5,550 and 6,070 transcripts with assigned Gene Ontology and KEGG pathway information, respectively. The most abundant KEGG pathways are spliceosome, ribosome, ubiquitin mediated proteolysis, plant–pathogen interaction, RNA degradation and oxidative phosphorylation metabolic pathway. Furthermore, we also found 12, 9, and 4 transcripts annotated as stress-related, heat stress, and water stress proteins, respectively. We envisage that these resources will promote and facilitate studies of the abiotic stress capabilities of this exotic plant species, which facilitates its invasive capacity.

Background: Tick parasitism is a major impediment for cattle production in many parts of the world. The southern cattle tick, Rhipicephalus (Boophilus) microplus, is an obligate hematophagous parasite of domestic and wild animals that serves as vector of infectious agents lethal to cattle. Tick saliva contains molecules evolved to modulate host innate and adaptive immune responses which facilitates blood feeding and pathogen transmission. Tick feeding promotes CD4 T cell polarization to a Th2 profile usually accompanied by down-regulation of Th1 cytokines through as yet undefined mechanisms. Co-stimulatory molecules on antigen presenting cells are central to development of T cell responses including Th1 and Th2 responses. Tick induced changes to antigen presenting cell signal transduction pathways are largely unknown. Here we document the ability of R. microplus salivary gland extracts (SGE) to effect differential CD86 expression.Results: We examined changes in co-stimulatory molecule expression in murine RAW 264.7 cells in response to R. microplus SGE exposure in the presence of the toll-like receptor 4 (TLR4) ligand, LPS. After 24 hrs, CD86, but not CD80, was preferentially up-regulated on mouse macrophage RAW 264.7 cells when treated with SGE and then LPS, but not SGE alone. CD80 and CD40 expression was increased with LPS, but the addition of SGE did not alter expression. Higher concentrations of SGE were less effective at increasing CD86 RNA expression. The addition of mitogen or extracellular kinase (MEK) inhibitor, PD98059, significantly reduced the ability for SGE to induce CD86 expression, indicating activation of MEK is necessary for SGE induced up-regulation.Conclusions: Molecules in SGE of R. microplus have a concentration-dependent effect on differential up-regulation of CD86 in a macrophage cell line activated by the TLR4 ligand, LPS. This CD86 up-regulation is at least partially dependent on the ERK1/2 pathway and may serve to promote Th2 polarization of the immune response.