Ronald B. Davey

Identification of a Third Boophilus microplus (Acari: Ixodidae) cDNA Presumptively Encoding an Acetylcholinesterase

Abstract Oligodeoxynucleotide primers, based on amino acid sequences conserved in known acetylcholinesterases (AChEs), were used in reverse-transcription polymerase chain reaction (RT-PCR) with mRNA from Boophilus microplus (Canestrini) as the template. Primer walking and rapid amplification of cDNA ends (RACE) techniques were used to complete the cDNA sequence identified by RT-PCR. The complete B. microplus cDNA sequence contained an open reading frame encoding a 620 amino acid protein with a 20 amino acid signal peptide at the N-terminus targeting the protein for the secretion pathway. BLAST searches of GenBank using the presumptively encoded protein revealed highest sequence similarity to AChEs. The presumptively encoded protein was of similar size and structural properties to other identified AChEs, including the presence of the catalytic triad (Ser, Glu, His) and appropriate placement of internal cysteines to yield three internal disulfide bonds corresponding to those of known AChEs. Putative conserved domains identified the sequence as a member of the carboxylesterase family, pfam00135.8, of which AChE is a member. This cDNA therefore presumptively encodes a third transcribed AChE (AChE3) cDNA of B. microplus. Comparison of the three AChE cDNA sequences expressed in B. microplus demonstrated no discernible nucleotide sequence homology and relatively low amino acid sequence homology, strongly suggesting that they are not alleles of one another. The potential presence of multiple expressed AChEs in B. microplus suggests alternative mechanisms for development of resistance to pesticides that target AChE. The homology-based identification of a third expressed AChE in B. microplus is a surprising result and strongly implies the need for confirmation of gene identity for presumptive AChEs.

Introduced ticks and tick-borne diseases: the threat and approaches to eradication.

Exotic tick species and tick-borne diseases are serious threats to live-stock, companion animals, and wildlife in the United States. Recurring introductions of exotic tick species into the United States are a significant indicator of the degree of risk. Successful tick-eradication campaigns, such as the national program that eradicated Boophilus annulatus and B. microplus from the United States, the Cattle Fever Tick Eradication Program of the US Department of Agricultures Veterinary Services that protects against the re-entry and dissemination of Boophilus ticks from Mexico back into their former haunts in the southern states, and the eradication action that eliminated Rhipicephalus evertsi from a game park in Florida, are sources of useful information that aid in elucidating essential elements of successful eradication programs. Examples of failed eradication programs in places such as Puerto Rico and St. Croix also have heuristic value. Among the varieties of tick species and related infectious agents that threaten the United States, Boophilus ticks and bovine babesiosis, Amblyomma species (especially the tropical bont tick) and heartwater, and equine babesiosis, for which endemic vectors exist, are of special concern. Risk assessments to accumulate, evaluate, and synthesize information needed to appraise risks, consequences, and preparedness are necessary not just to inform federal, state, and local officials, as well as producers and stakeholders, but also to facilitate the creation of emergency response plans.

Distribution of Rhipicephalus (Boophilus) microplus and Rhipicephalus (Boophilus) annulatus (Acari: Ixodidae) Infestations Detected in the United States Along the Texas/Mexico Border

ABSTRACT Species identification and global positioning system coordinates of infestations of cattle fever ticks, Rhipicephalus (Boophilus) annulatus (Say) and Rhipicephalus (Boophilus) microplus (Canestrini), were determined for 790 specimens submitted to the National Veterinary Services Laboratory between 1 October 1999 and 30 September 2010. Cattle fever tick specimens obtained by personnel of the United States Department of Agriculture-Animal and Plant Health Inspection Service-Cattle Fever Tick Eradication Program from infested cattle and wildlife along the Texas/ Mexico border were submitted for identification, as required by the program. A geographic information system database was developed that incorporates location, collection, and infestation records. Submitted ticks came from 11 Texas counties and were comprised of 19.5% R. (B.) annulatus and 80.5% R. (B.) microplus. Maps produced from this study locate and define the parapatric boundary between R. (B.) annulatus and R. (B.) microplus.

Survival of larvae of Boophilus annulatus, Boophilus microplus, and Boophilus hybrids (acari: ixodidae) in different temperature and humidity regimes in the laboratory

The survival period for larvae of Boophilus annulatus (Say), Boophilus microplus (Canestrini) and hybridized Boophilus ticks was determined by exposure to various combinations of temperature (20, 25, 30 and 35 degrees C) and relative humidity (32, 63, 75, 84 and 97% RH) in the laboratory. Results indicated that within a given temperature and RH regime, there was no difference (P greater than 0.05) in larval survival among the three species tested, indicating that these ticks respond similarly over a wide range of temperature and RH combinations. Larval survival in all three species was longest (P less than 0.05) at 20 degrees C and either 84 or 97% RH. With each increase in temperature at the 84 and 97% RH treatment levels, there was a corresponding significant (P less than 0.05) decrease in larval survival. When the temperature reached 35 degrees C at all humidities or when the RH was 63% or less at all temperatures, the mean larval survival period was 43 days or less in all cases and little difference (P greater than 0.05) was observed among the treatment regimes included. Results suggest that at a RH of 75% and more, the temperature is the determining factor in larval survival, whereas at a RH of 63% and less the RH is the determining factor in larval survival, regardless of temperature.

Efficacy of amitraz (Taktic® 12.5% EC) as a dip for the control of Boophilus microplus (Canestrini) (Acari: Ixodidae) on cattle

Four groups of cattle infested with Boophilus microplus (Canestrini) were each dipped in a different concentration of amitraz diluted from a 12.5% EC formulation to determine the efficacy and performance of the product in an 11,400 l dipping vat. Except for the period when heifers were dipped, animals were restrained in stanchions placed individually inside 3.3 x 3.3 m2 stalls within an open-sided barn. The amitraz in the vat was stabilized with hydrated lime to maintain a pH of ca. 12. Analyses of vat samples showed that concentrations of amitraz in the vat were 7.6 to 13% lower than the targeted concentrations of 0.010, 0.015, 0.020, and 0.025% active ingredient (AI) for dilutions prepared according to instructions on the manufacturers label. The large quantity of hydrated lime added to the vat (10 kg/1000 l) interfered with the HPLC analysis of vat samples. Therapeutic efficacy of each of the four observed concentrations (0.0088, 0.0131, 0.0174, and 0.0231% AI) of amitraz was excellent (> 99% control). However, the rapid detachment of all ticks from an animal within a few hours after treatment with amitraz, that has been frequently observed, was not pronounced in the present study. Only 47% of the B. microplus detached in the first 4 h post-treatment, and 84% detached within the first 24 h. All of the treatments, except the lowest concentration, provided protection of cattle against re-infestation by B. microplus larvae for 14 days post-treatment. Possibly as a result of the formation of a compact layer of lime and amitraz on the bottom after the vat was undisturbed for six weeks, intense agitation was required to re-suspend the active ingredient.

Efficacy of sprays of amitraz against Boophilus ticks on cattle

Abstract Cattle infested with all parasitic life-stages of Boophilus microplus (Canestrini) and B. annulatus (Say) were sprayed with 0.0125 or 0.025% active ingredient (AI) Amitraz. The detachment pattern of engorged or partially engorged females indicated that both concentrations of Amitraz caused premature detachment, with at least 90% of all females detaching within 24 h post-treatment. A large portion of the females collected during the first 24 h post-treatment were not fully engorged. The 2 tick species were equally sensitive to both concentrations tested. At 0.0125%, control of Index of Reproduction (IR) of B. annulatus was 98.9% while control of B. microplus was 97.8%. At 0.025% control of IR was 99.2 and 98.0% for B. annulatus and B. microplus , respectively. Amitraz adversely affected size and oviposition of females and the hatchability of eggs laid by treated females. Based on known detachment intervals of females after infestation, it appeared that Amitraz-treated animals were protected against larval reinfestation of both species for ca. 7 days at 0.0125% and for ca. 10 days at 0.025%.

Efficacy of Spinosad Against Acaricide-Resistant and -Susceptible Rhipicephalus (Boophilus) microplus and Acaricide-Susceptible Amblyomma americanum and Dermacentor variabilis

ABSTRACT Various acaricide-resistant strains of Rhipicephalus (Boophilus) microplus, representative of the major resistance mechanisms found in Mexico and Brazil, were exposed to spinosad using the Food and Agriculture Organization of the United Nations larval packet test and adult immersion test (AIT). Larvae of all strains tested were found to be susceptible to spinosad. Conversely, spinosad did not show toxic activity toward engorged females used in the AIT. In vitro tests against larvae, nymphs, and adults of acaricide-susceptible Amblyomma americanum and Dermacentor variabilis revealed differences in activity that were dependent on tick species and parasitic life stage. Spinosad seemed to be a viable alternative to current acaricides available for tick control in the species tested. The larval packet test should be used for future monitoring of resistance, as the AIT did not provide useful information with this chemical. The potential benefit for the use of spinosad in integrated pest management or eradication programs is discussed.

Efficacy of freshly mixed coumaphos suspensions adjusted to various pH levels for treatment of cattle infested with Boophilus annulatus (Say) (Acari: Ixodidae)

Abstract Reductive dechlorination of coumaphos results in the formation of a highly toxic material, potasan, but lower pH (≤ 5.5) of the liquid containing coumaphos may eliminate this degradation process. However, the effects of lower pH on the rapid settling of coumaphos particles and on the efficacy of the pesticide are not known. Three suspensions were made from a freshly mixed stock solution of a flowable formulation of coumaphos at 0.125% active ingredient at pH values of 6.7, 5.5, and 4.5. Coumaphos particles were observed to remain well suspended in all pH treatments before, during, and for up to 1 h after applications were made on tick infested cattle, suggesting that there was no break in any of the coumaphos suspensions. Likewise, observations on the behavior of each of the animals (both treated and untreated) appeared to be similar and uniform across all groups, suggesting that the lowering of the pH would not cause any noticeable adverse reaction in the treated animals. Data obtained from the coumaphos suspensions at each of the three pH treatments (pH 6.7, 5.5, and 4.5) against Boophilus annulatus (Say) resulted in control of the index of reproduction (IR) that ranged from 94.4 to 96.5%, indicating that pH had little effect on the efficacy of the acaricide. Results also indicated that the pH of coumaphos suspensions had little effect on engorgement weights (range 270–280 mg), egg mass weights (range 88–100 mg), or percent hatch of eggs (range 57–71%) of females that survived the treatments. Therefore, we concluded that coumaphos suspensions with a pH as low as 4.5 would have no observable adverse effect on the rapid settling of coumaphos particles in suspension, cause no noticeable adverse reactions in treated animals, and remain highly efficacious against B. annulatus .

Efficacy and blood sera analysis of a long-acting formulation of moxidectin against Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) on treated cattle.

ABSTRACT The therapeutic and persistent efficacy of a single subcutaneous injection of a long-acting formulation of moxidectin at a concentration of 1 mg/kg body weight was determined against Rhipicephalus (Boophilus) microplus (Canestrini), along with the concentration-time blood sera profile in treated cattle. The therapeutic efficacy against ticks of all parasitic stages on cattle at the time of treatment was >99.9%, and the mean tick number, index of fecundity, engorgement weight, and egg mass weight of ticks recovered from treated animals were all significantly lower than ticks from untreated animals. The index of fecundity, engorgement weight of females, and egg mass weight of ticks recovered from treated animals infested at weekly (7-d) intervals between 14 and 63 d posttreatment were significantly lower than for ticks on untreated animals, whereas the number of ticks per animal recovered from treated cattle remained lower than that of untreated cattle for up to 49 d posttreatment. The percentage control remained >99% at weekly intervals between 14 and 49 d posttreatment, which is the minimum level of efficacy considered acceptable for use in the United States Cattle Fever Tick Eradication Program. The serum concentration of moxidectin in treated cattle increased to 25.6 ppb (parts per billion) within 1 d after treatment, and peaked at 47.3 ppb at 8 d posttreatment. Moxidectin sera levels remained above the estimated 100% threshold level for elimination of feeding ticks (5–8 ppb) for 44–53 d after treatment. The label claim of 50 d of prevention against reinfestation for the long-acting moxidectin formulation used in the study was supported by the efficacy and sera concentration data obtained. Based on these results, cattle could be treated at 63-d intervals with minimal risk of viable ticks detaching from treated animals. This treatment interval would be 4.5-fold longer than the presently required treatment interval of 14 d, thus leading to ≈75% reduction in gathering and handling costs of cattle incurred by producers.

Liquid Chromatographic Detection of Permethrin from Filter Paper Wipes of White-tailed Deer

White-tailed deer, Odocoileus virginianus (Zimmermann), are keystone hosts for the adult stage of lone star ticks, Amblyomma americanum (L.), (Patrick and Hair 1978, J. Parasitol. 64:1100 -1106; Bloemer et al. 1986. J. Econ. Entomol. 79: 679 683) and blacklegged ticks, Ixodes scapularis Say, (Barbour and Fish 1993, Science 260: 1610-1616), both of which transmit a variety of agents that cause disease in humans, including Lyme disease, human ehrlichioses, and human babesiosis in the eastern U.S. (Lockhart et al. 1996, J. Med. Entomol. 33: 153 158; Varela et al. 2004, J. Clin. Microbiol. 42:1163 1169). White-tailed deer are also marginal hosts of the cattle tick, Rhipicephalus (Boophilus) annulatus (Say), and the southern cattle tick, R. (B.) microplus (Canestrini), particularly in the absence of cattle (Cooksey et al. 1989, J. Med. Entomol. 26: 155 158). Increasing populations of white-tailed deer along the Texas-Mexico border are compromising efforts by federal agencies to prevent the reestablishment of these vectors of bovine babesiosis from Mexico, where these disease vectors are endemic (George 1990, J. Agric. Entomol. 7:119 -125, Pound et al. 2010, J. Econ. Entomol. 103:211 -218). In an effort to manage the disease risks posed by the increasing numbers of whitetailed deer and subsequent tick populations in the northeastern U.S. and reinfestations of cattle fever ticks (southern cattle ticks and cattle ticks) in South Texas, the U.S. Department of Agriculture (USDA) Agricultural Research Service (ARS) developed two passive treatment devices designed to apply permethrin acaricide onto the ears