Phenology of five tick species in the central Great Plains.

The states of Kansas and Oklahoma, in the central Great Plains, lie at the western periphery of the geographic distributions of several tick species. As the focus of most research on ticks and tick-borne diseases has been on Lyme disease which commonly occurs in areas to the north and east, the ticks of this region have seen little research attention. Here, we report on the phenology and activity patterns shown by tick species observed at 10 sites across the two states and explore factors associated with abundance of all and life specific individuals of the dominant species. Ticks were collected in 2020-2022 using dragging, flagging and carbon-dioxide trapping techniques, designed to detect questing ticks. The dominant species was A. americanum (24098, 97%) followed by Dermacentor variabilis (370, 2%), D. albipictus (271, 1%), Ixodes scapularis (91, <1%) and A. maculatum (38, <1%). Amblyomma americanum, A. maculatum and D. variabilis were active in Spring and Summer, while D. albipictus and I. scapularis were active in Fall and Winter. Factors associated with numbers of individuals of A. americanum included day of year, habitat, and latitude. Similar associations were observed when abundance was examined by life-stage. Overall, the picture is one of broadly distributed tick species that shows seasonal limitations in the timing of their questing activity.

Multiple species of canine Rhipicephalus complex detected in Canada.

Multiple species of brown dog ticks have been described in the United States and the Caribbean: Rhipicephalus sanguineus sensu stricto (s.s.), also referred to as temperate lineage; R. linnaei (=tropical lineage); and R. rutilus (=southeastern Europe lineage) However, Rhipicephalus spp. are rarely recovered from dogs in Canada. To identify canine Rhipicephalus spp. in Canada and determine the influence of travel history on infestation, ticks morphologically identified as brown dog ticks (n = 93) collected from dogs (n = 13) in British Columbia, Ontario, and Québec, Canada were submitted with information regarding each dog's geographic location and travel history. Nucleic acid was extracted from available individual ticks (n = 86) and PCR was used to amplify sequences of a 12S rRNA mitochondrial gene fragment. Sequences were compared to published reference sequences of known species and a phylogenetic tree constructed. Twenty-three ticks (26.7%) consistent with R. linnaei were identified on seven dogs, including dogs from British Columbia and Ontario, with a median infestation intensity of 2 ticks/dog (mean = 3.3 ticks/dog). Sixty-one ticks (70.9%) consistent with R. sanguineus s.s. were found on two dogs from Québec and Ontario (median = 30.5 ticks/dog; mean = 30.5 ticks/dog). One dog from Ontario was infested with R. rutilus (n = 2) (2.3%). Species could not be determined for ticks from three dogs from Ontario and Québec. Most infested dogs (10/13; 76.9%) had a recent (< 1 month) international travel history. These data confirm that multiple species of canine Rhipicephalus are occasionally found in Canada and suggest introduction following travel is likely responsible for these infestations. Further analysis will allow for greater understanding of the range and diversity of canine Rhipicephalus spp. in North America and may reveal risk factors for infestation.

Diagnosis of canine intestinal parasites: Improved detection of Dipylidium caninum infection through coproantigen testing.

Intestinal parasites, including cestodes like Dipylidium caninum, are common in dogs in the United States of America (USA), but fecal flotation consistently, and, at times, dramatically, fails to identify many of these infections. To determine the extent to which including coproantigen testing for D. caninum would improve the identification of dogs infected with this cestode, we evaluated fecal samples from 877 dogs (589 pet and 288 from municipal shelters) from six USA states using zinc sulfate (specific gravity 1.24) fecal flotation with centrifugation along with coproantigen detection for Giardia sp., hookworms, ascarids, and Trichuris vulpis. For D. caninum, PCR of perianal swabs was included. Intestinal parasite infections were identified, using centrifugal fecal flotation or coproantigen, in 265 dogs (13.2 % pet, 64.9 % shelter). Dipylidium caninum infection was detected in 5.6 % of dogs with the combination of coproantigen and centrifugal fecal flotation, and 7.3 % of dogs when perianal swab results were included; prevalence varied by diagnostic method, population, and geographic region. In pet dogs, D. caninum infection was identified by fecal flotation (0), coproantigen (2.2 %), or perianal swabs (1.2 %). The same methods revealed infection in 0.3 %, 12.5 %, and 11.1 % of shelter dogs, respectively. Frequent use of praziquantel in shelter dogs (116/288; 40.3 %) may have reduced prevalence. Positive and negative agreement of D. caninum coproantigen with perianal swab PCR in pet dogs was 85.7 % and 98.8 %, respectively. Multiple logistic regression analysis accounting for region, population, and age found D. caninum infection to be more common in shelter dogs relative to pet (adjusted OR 4.91 [2.48, 10.24]) and in the Southcentral and Southeast regions relative to North (adjusted OR 9.59 [1.92, 174.13] and 17.69 [3.67, 318.09] respectively). Coproantigen testing also enhanced the detection of other intestinal parasites over fecal flotation alone, including Giardia sp. (14.7 % vs 3.3 %), hookworms (13.8 % vs 8.4 %), ascarids (2.9 % vs 2.2 %), and T. vulpis (2.9 % vs 1.4 %). Together, these data indicate that the coproantigen assay employed increases detection of D. caninum infections several fold, supporting the use of this test in clinical practice, and add to a growing body of research documenting enhanced diagnosis through implementation of multiple laboratory-based methods.

Seasonality of Amblyomma americanum (Acari: Ixodidae) Activity and Prevalence of Infection with Tick-Borne Disease Agents in North Central Oklahoma.

Background: Amblyomma americanum is the most common tick infesting both animals and humans in the southern United States and transmits a variety of zoonotic agents. The rise in tick-borne diseases (TBD) globally imparts a need for more active surveillance of tick populations to accurately quantify prevalence and risk of tick-borne infectious organisms. To better understand TBD risk in north central Oklahoma, this study aimed to describe the current seasonal activity of A. americanum in this region and investigate the seasonality of tick-borne infectious agents. Materials and Methods: Tick collections were performed twice a month for a duration of 2 years at a field site in Payne County, Oklahoma. Total nucleic acid was extracted from a subset of adult A. americanum and tested for Rickettsia spp., Ehrlichia spp., and Borrelia spp. using established PCR protocols. Results: Peak activity times for each life stage were observed, with adults primarily active 1 month earlier than historical seasonal trends describe, and male A. americanum active earlier in the year than female A. americanum. Rickettsia spp., Ehrlichia chaffeensis, Ehrlichia ewingii, and Borrelia lonestari were found in 26.4%, 6.1%, 2.5%, and 1.1% of adult A. americanum, respectively. No seasonal trend in spotted fever group Rickettsia spp. (SFGR) was observed in peak activity months. Conclusions: This study found an apparently shifting phenology for A. americanum adults in Oklahoma. While these results did not show a trend in SFGR, further investigation is needed to better understand the potential seasonality of infection prevalence within A. americanum across the expanding range of this vector, especially considering the extended activity of males in winter months.

Fluralaner (Bravecto®) treatment kills Aedes aegypti after feeding on Dirofilaria immitis-infected dogs.

BACKGROUND: Transmission of canine heartworm (Dirofilaria immitis) from infected to naïve dogs is dependent on successful mosquito feeding and survival. METHODS: To determine whether treating heartworm-infected dogs with fluralaner (Bravecto®) limits the survival of infected mosquitoes, and potentially the transmission of D. immitis, we allowed female mosquitoes to feed on microfilaremic dogs and evaluated mosquito survival and infection with D. immitis. Eight dogs were experimentally infected with D. immitis. On day 0 (~ 11 months post-infection), four microfilaremic dogs were treated with fluralaner according to label directions while the other four were non-treated controls. Mosquitoes (Aedes aegypti Liverpool) were allowed to feed on each dog on days -7, 2, 30, 56, and 84. Fed mosquitoes were collected, and the number of live mosquitoes determined at 6 h, 24 h, 48 h, and 72 h post-feeding. Surviving mosquitoes held for 2 weeks were dissected to confirm third-stage D. immitis larvae; PCR (12S rRNA gene) was performed post-dissection to identify D. immitis in mosquitoes. RESULTS: Prior to treatment, 98.4%, 85.1%, 60.7%, and 40.3% of mosquitoes fed on microfilaremic dogs were alive at 6 h, 24 h, 48 h, and 72 h post-feeding, respectively. Similarly, mosquitoes fed on microfilaremic, non-treated dogs were alive 6 h post-feeding (98.5-100%) throughout the study. In contrast, mosquitoes fed on fluralaner-treated dogs 2 days after treatment were dead or severely moribund by 6 h post-feeding. At 30 and 56 days post-treatment, > 99% of mosquitoes fed on treated dogs were dead by 24 h. At 84 days post-treatment, 98.4% of mosquitoes fed on treated dogs were dead by 24 h. Before treatment, third-stage larvae of D. immitis were recovered from 15.5% of Ae. aegypti 2 weeks after feeding, and 72.4% were positive for D. immitis by PCR. Similarly, 17.7% of mosquitoes fed on non-treated dogs had D. immitis third-stage larvae 2 weeks after feeding, and 88.2% were positive by PCR. Five mosquitoes fed on fluralaner-treated dogs survived 2 weeks post-feeding, and 4/5 were from day 84. None had third-stage larvae at dissection, and all were PCR-negative. CONCLUSION: The data indicate that fluralaner treatment of dogs kills mosquitoes and thus would be expected to reduce transmission of heartworm in the surrounding community.

Phylogeography of the blacklegged tick (Ixodes scapularis) throughout the USA identifies candidate loci for differences in vectorial capacity.

The blacklegged tick (Ixodes scapularis (Journal of the Academy of Natural Sciences of Philadelphia, 1821, 2, 59)) is a vector of Borrelia burgdorferi sensu stricto (s.s.) (International Journal of Systematic Bacteriology, 1984, 34, 496), the causative bacterial agent of Lyme disease, part of a slow-moving epidemic of Lyme borreliosis spreading across the northern hemisphere. Well-known geographical differences in the vectorial capacity of these ticks are associated with genetic variation. Despite the need for detailed genetic information in this disease system, previous phylogeographical studies of these ticks have been restricted to relatively few populations or few genetic loci. Here we present the most comprehensive phylogeographical study of genome-wide markers in I. scapularis, conducted by using 3RAD (triple-enzyme restriction-site associated sequencing) and surveying 353 ticks from 33 counties throughout the species' range. We found limited genetic variation among populations from the Northeast and Upper Midwest, where Lyme disease is most common, and higher genetic variation among populations from the South. We identify five spatially associated genetic clusters of I. scapularis. In regions where Lyme disease is increasing in frequency, the I. scapularis populations genetically group with ticks from historically highly Lyme-endemic regions. Finally, we identify 10 variable DNA sites that contribute the most to population differentiation. These variable sites cluster on one of the chromosome-scale scaffolds for I. scapularis and are within identified genes. Our findings illuminate the need for additional research to identify loci causing variation in the vectorial capacity of I. scapularis and where additional tick sampling would be most valuable to further understand disease trends caused by pathogens transmitted by I. scapularis.

Geographic Distribution and Seasonality of Brown Dog Tick Lineages in the United States.

Two lineages of brown dog ticks (Rhipicephalus sanguineus sensu lato (s.l.), Latreille [Acari: Ixodidae]) have been described in North America: temperate and tropical. To characterize the distribution of these lineages across this region and evaluate seasonal activity, a 12S rRNA mitochondrial gene fragment was sequenced from R. sanguineus s.l. collected from hundreds of dogs and cats from different locations across 25 of the 50 states from 2018 to 2021. Infestations with temperate lineage predominated (78.5%) and were identified on pets from 20 states, with most (83.5%) from areas with annual mean daily average temperature <20°C. Tropical lineage submissions were less common (19.3%), submitted from 15 states, and most (80.0%) tropical lineage ticks were from areas with an annual mean daily average temperature >20°C. Although travel history was not obtained for all dogs, when tropical lineage infestations were found in colder regions, follow up conversations with veterinarians suggested some of these infestations may have resulted from recent travel of dogs. A limited number (2.2%) of dogs from Arizona and Texas were co-infested with both lineages. Both temperate and tropical lineage ticks were collected from pets in every month of the year. Temperate lineage infestations were primarily collected March through August while tropical lineage infestations were more often collected June through November. These data confirm at least two lineages of R. sanguineus s.l. are present in the United States, each predominating in distinct, overlapping geographies, and suggest that peak activity of each lineage occurs at different times of the year.

Diversity and geographic distribution of rickettsial agents identified in brown dog ticks from across the United States.

Rhipicephalus sanguineus sensu lato, or brown dog ticks, transmit a variety of pathogens of veterinary and public health importance globally. Pathogens vectored by brown dog ticks and identified in the United States include Babesia vogeli, Ehrlichia canis, and several spotted fever group Rickettsia spp. (SFGR). Due to the challenge of collecting canine blood samples nationwide to screen for exposure to these pathogens, we took an indirect approach and tested brown dog ticks for molecular evidence of infection. Brown dog ticks (616 adults and 65 nymphs) collected from dogs and cats across the nation were tested by separate PCR assays detecting Babesia spp., E. canis, and SFGR. While no Babesia sp. was found, we identified rickettsial agents in 3.5% (24/681; 95% CI 2.4-5.2%) of the ticks. Pathogens and related organisms detected in ticks included E. canis (n = 1), Rickettsia amblyommatis (n = 3), Rickettsia massiliae (n = 11), Rickettsia monacensis (n = 3), Rickettsia montanensis (n = 5), and an undefined Rickettsia species (n = 1). These data demonstrate a wider geographic distribution of R. massiliae than previously known, and to the authors' knowledge, reports R. monacensis in brown dog ticks for the first time. Due to the close association that brown dog ticks have with domestic dogs and humans, more research is needed to understand the full array of organisms, some of which are zoonotic, potentially transmitted by this widespread tick complex.

Survey on the Presence of Equine Tick-Borne Rickettsial Infections in Southcentral United States.

Although ticks are known vectors of pathogens across a range of hosts, there is limited research on emerging tick-borne diseases of horses in the United States. Tick surveys from other regions suggest Rickettsia spp. and Ehrlichia spp. may be clinically relevant in horses. To better understand the transmission risk of these tick-borne rickettsial disease agents to horses, ticks were collected from horses in Oklahoma. Ticks for the current study (306 Amblyomma americanum, 20 Dermacentor albipictus, 19 D. variabilis, and 7 A. maculatum) were evaluated for Rickettsia spp. and Ehrlichia spp. using polymerase chain reaction and sequence analysis. Serum samples from infested and noninfested horses were evaluated for antibodies to R. rickettsii using indirect fluorescence antibody testing and Ehrlichia spp. and Anaplasma spp. using a commercial enzyme-linked immunoassay. Of the horses with tick infestations, 71.4% hosted at least one tick with a rickettsial agent detected. Rickettsia spp. were identified in 25.9% (91/352) of the ticks tested with R. amblyommatis (80.2%; 73/91) most often detected. Ehrlichia spp. were identified in 2.8% (10/352) of the ticks tested with E. ewingii most often identified. Serologic screening revealed no horses with antibodies to R. rickettsii or Anaplasma spp., but 29.6% of the examined horses had circulating antibodies to Ehrlichia spp. Together, these results demonstrate the presence of Rickettsia spp. and Ehrlichia spp. in equine ticks and evidence of past or current infection with Ehrlichia spp. in Oklahoma horses which strongly suggests there is a need to explore the relationship between these agents and equine health.

Influence of tick sex and geographic region on the microbiome of Dermacentor variabilis collected from dogs and cats across the United States.

As tick-borne diseases continue to increase across North America, current research strives to understand how the tick microbiome may affect pathogen acquisition, maintenance, and transmission. Prior high throughput amplicon-based microbial diversity surveys of the widespread tick Dermacentor variabilis have suggested that life stage, sex, and geographic region may influence the composition of the tick microbiome. Here, adult D. variabilis ticks (n = 145) were collected from dogs and cats from 32 states with specimens originating from all four regions of the United States (West, Midwest, South, and Northeast), and the tick microbiome was examined via V4-16S rRNA gene amplification and Illumina sequencing. A total of 481,246 bacterial sequences were obtained (median 2924 per sample, range 399-11,990). Fifty genera represented the majority (>80%) of the sequences detected, with the genera Allofrancisella and Francisella being the most abundant. Further, 97%, 23%, and 5.5% of the ticks contained sequences belonging to Francisella spp., Rickettsia spp., and Coxiella spp., respectively. No Ehrlichia spp. or Anaplasma spp. were identified. Co-occurrence analysis, by way of correlation coefficients, between the top 50 most abundant genera demonstrated five strong positive and no strong negative correlation relationships. Geographic region had a consistent effect on species richness with ticks from the Northeast having a significantly greater level of richness. Alpha diversity patterns were dependent on tick sex, with males exhibiting higher levels of diversity, and geographical region, with higher level of diversity observed in ticks obtained from the Northeast, but not on tick host. Community structure, or beta diversity, of tick microbiome was impacted by tick sex and geographic location, with microbiomes of ticks from the western US exhibiting a distinct community structure when compared to those from the other three regions (Northeast, South, and Midwest). In total, LEfSe (Linear discriminant analysis Effect Size) identified 18 specific genera driving these observed patterns of diversity and community structure. Collectively, these findings highlight the differences in bacterial diversity of D. variabilis across the US and supports the interpretation that tick sex and geographic region affects microbiome composition across a broad sampling distribution.

Identification of Rickettsia spp. and Babesia conradae in Dermacentor spp. Collected from Dogs and Cats Across the United States.

In the United States, Dermacentor variabilis and Dermacentor andersoni are considered key vectors for Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever. Through regional surveillance, a wide diversity of Rickettsia spp. have been documented in D. variabilis, and Dermacentor spp. has been suggested as potential vectors for various other pathogens, including Babesia spp. and Ehrlichia canis. To better define the prevalence and diversity of pathogens in Dermacentor spp. across the United States, 848 ticks collected from dogs and cats in 44/50 states in 2018-2019 were tested by PCR for Rickettsia spp.-specific 17 kDa and ompA gene fragments; a subset of Dermacentor spp. was also tested with PCR, targeting fragments of the 18S and large subunit region rRNA genes of Babesia spp. and 16S rRNA genes of E. canis. Rickettsia spp. was identified in 12.5% (106/848) of ticks. Species detected include Rickettsia montanensis (n = 64 ticks), Rickettsia bellii (n = 15 ticks), Rickettsia rhipicephali (n = 13 ticks), Rickettsia peacockii (n = 8 ticks), Rickettsia amblyommatis (n = 3 ticks), Rickettsia cooleyi (n = 1 tick), and unclassified Rickettsia spp. (n = 2 ticks). Ticks with R. montanensis and R. bellii were submitted from every U.S. region; R. rhipicephali was predominantly detected in ticks from the southern half of the United States, and all R. peacockii-positive ticks were D. andersoni that originated from the Rocky Mountain states. Ehrlichia canis was not detected in any Dermacentor spp., and Babesia conradae was detected in two Dermacentor albipictus. Because most ticks had fed on dogs or cats before submission, these findings do not implicate a given Dermacentor sp. as a primary vector of these agents, but in regard to Rickettsia spp., the data do support other published work showing D. variabilis harbors a diversity of Rickettsia species with unknown implications for animal and human health.

Molecular detection of Cercopithifilaria bainae in brown dog ticks collected from dogs across the United States.

Cercopithifilaria bainae is a filarioid nematode of dogs shown to use Rhipicephalus sanguineus sensu lato (s.l.), the brown dog tick, as the vector. Previously in the United States, C. bainae infections have been reported in a dog from Florida, and in dogs and ticks in Oklahoma, but data are lacking from other areas of the country. Here, we tested brown dog ticks from across the United States for C. bainae DNA to assess the geographic distribution of where this novel parasite may be cycling in ticks and dogs. Archival brown dog ticks were available for testing through the national tick survey Show Us Your Ticks. Ticks were morphologically identified, dissected, and tested by PCR to detect filarioid mitochondrial DNA. A total of 1400 brown dog ticks were tested from 321 separate animals from 23 states, with 5.7 % (80/1400) of the ticks testing positive for C. bainae DNA. At least one positive tick was detected in submissions from 9 states in addition to Florida and Oklahoma. Cercopithifilaria bainae DNA was detected in larval, nymphal, and adult stages of brown dog ticks and only in ticks removed from dogs. Of all dogs with brown dog ticks collected from them, 17.6 % (55/312) were infested with at least one tick that harbored C. bainae DNA. Findings from this study demonstrate a wider geographic range of C. bainae than previously known, and that dogs are commonly infested with brown dog ticks with molecular evidence of C. bainae infection.

Equine attachment site preferences and seasonality of common North American ticks: Amblyomma americanum, Dermacentor albipictus, and Ixodes scapularis.

BACKGROUND: Ticks are common on horses, but recent publications characterizing equine tick infestations in North America are lacking. METHODS: To further understand attachment site preferences of common ticks of horses, and to document the seasonality of equine tick infestation in northeastern Oklahoma, horses from eight farms were evaluated twice a month over a 1-year period. Each horse was systematically inspected beginning at the head and moving caudally to the tail. Attachment sites of ticks were recorded and all ticks collected were identified to species and stage. RESULTS: Horses (26 males and 62 females) enrolled in the study ranged in age from 1 to 23 years (mean = 12, 95% CI 11-13). A total of 2731 ticks were collected; 84.1% (74/88) of the horses were infested (median = 3 ticks) at one or more examinations. Five tick species were identified, including Amblyomma americanum (78.2%; 2136/2731), Ixodes scapularis (18.2%; 497/2731), Dermacentor albipictus brown variant (2.6%; 71/2731), Dermacentor variabilis (0.7%; 20/2731), and Amblyomma maculatum (0.3%; 7/231). Most ticks were adults (83.6%; 2282/2731), but immature A. americanum (436/2136; 20.4%), D. albipictus (12/71; 16.9%), and A. maculatum (n = 1) were occasionally recovered. Amblyomma americanum were most often attached to the inguinal area, and I. scapularis and D. albipictus were most commonly found on the chest and axillary region (P < 0.0001). Ticks were found on horses in every month of the year. The largest number of ticks (638/2731; 23.4%) were collected in May (P < 0.0001). Amblyomma americanum, primarily immature, was the only tick recovered in September, I. scapularis and D. albipictus predominated October through February, and both A. americanum and I. scapularis were common in March. In the warmer months, April through August, A. americanum was the most common tick, followed by D. variabilis and A. maculatum. CONCLUSIONS: This research confirms that ticks common on horses in North America have attachment site preferences and that ticks infest horses in Oklahoma throughout the year, including during the winter. Additional research is warranted to fully understand the risk these infestations pose to equine health.

Optimizing heartworm diagnosis in dogs using multiple test combinations.

BACKGROUND: Various heartworm (HW) diagnostic testing modalities detect products of, or reactions to, different life cycle stages of Dirofilaria immitis. Microfilariae (Mf) can be directly visualized in blood, antigen (Ag) from immature and adult heartworms may be detected on commercial assays, and antibody (Ab) tests detect the host immune response to larval stages. Ag and Mf tests are commonly used in dogs, which frequently carry adult HW infections, but Ab tests have only been validated for use in cats. In some HW-infected dogs, Ag is blocked by immune complexing leading to false-negative results. Heat-treatment (HT) to disrupt these complexes can increase the sensitivity of HW Ag tests. The aim of this study was to compare different methods for diagnosing HW infection in dogs at high risk using individual and paired diagnostic tests, including an exploration of using Ab tests designed for cats to test canine samples. METHODS: One hundred stray adult (≥ 2-year-old) dogs in Florida shelters were tested using Mf, HW Ag, and HW Ab tests (feline HW Ab tests currently not commercially validated/approved for use in dogs); two versions of each test platform were used. RESULTS: Fourteen dogs tested positive using point-of-care (POC) Ag tests; an additional 2 dogs tested positive with microtiter well assay, and an additional 12 dogs tested positive using HT Ag testing. For individual tests, Ag test sensitivity/specificity compared to HT Ag was 50-57%/100%, and Ab tests were 46-64%/82-94%. Sensitivity estimates for individual tests were higher when comparing to non-HT Ag. Pairing POC Ag tests with Mf tests improved sensitivity without loss of specificity, while pairing POC Ag and Ab tests modestly increased sensitivity at the expense of specificity. CONCLUSIONS: Screening dogs for HW infection using both POC Ag and Mf detection, which is recommended by the American Heartworm Society, improved diagnostic performance in this study compared to single Ag test use, but may have missed more than one in four infected dogs. The need to improve access to highly accurate, rapid, and inexpensive large-scale HW testing for dogs in animal shelters remains largely unmet by current testing availability. The development of practical and validated protocols that incorporate heat or chemical treatment to disrupt Ag-Ab complexes in POC testing or decreasing the cost and time required for such testing in reference laboratories might provide solutions to this unmet need. Similar studies performed in countries where the prevalence of parasites such as D. repens or A. vasorum is different to the USA could potentially yield very different positive predictive values for both HT and non-HT Ag tests.

Dermacentor variabilis is the Predominant Dermacentor spp. (Acari: Ixodidae) Feeding on Dogs and Cats Throughout the United States.

Throughout North America, Dermacentor spp. ticks are often found feeding on animals and humans, and are known to transmit pathogens, including the Rocky Mountain spotted fever agent. To better define the identity and distribution of Dermacentor spp. removed from dogs and cats in the United States, ticks submitted from 1,457 dogs (n = 2,924 ticks) and 137 cats (n = 209 ticks) from veterinary practices in 44/50 states from February 2018-January 2020 were identified morphologically (n = 3,133); the identity of ticks from regions where Dermacentor andersoni (Stiles) have been reported, and a subset of ticks from other regions, were confirmed molecularly through amplification and sequencing of the ITS2 region and a 16S rRNA gene fragment. Of the ticks submitted, 99.3% (3,112/3,133) were Dermacentor variabilis (Say), 0.4% (12/3,133) were D. andersoni, and 0.3% (9/3,133) were Dermacentor albipictus (Packard). While translocation of pets prior to tick removal cannot be discounted, the majority (106/122; 87%) of Dermacentor spp. ticks removed from dogs and cats in six Rocky Mountain states (Montana, Idaho, Wyoming, Nevada, Utah, and Colorado) were D. variabilis, suggesting this species may be more widespread in the western United States than is currently recognized, or that D. andersoni, if still common in the region, preferentially feeds on hosts other than dogs and cats. Together, these data support the interpretation that D. variabilis is the predominant Dermacentor species found on pets throughout the United States, a finding that may reflect recent shifts in tick distribution.

Ixodes spp. from Dogs and Cats in the United States: Diversity, Seasonality, and Prevalence of Borrelia burgdorferi and Anaplasma phagocytophilum.

Ixodes spp. are commonly found on dogs and cats throughout the world. In the eastern United States, 16S rDNA sequence of Ixodes scapularis, the predominant species, reveals two clades-American and Southern. To confirm the species and clades of Ixodes spp. ticks submitted from pets, we examined ticks morphologically and evaluated 16S rDNA sequence from 500 ticks submitted from 253 dogs, 99 cats, 1 rabbit, and 1 ferret from 41 states. To estimate pathogen prevalence, flaB of Borrelia burgdorferi (Bb) sensu stricto and 16S rDNA of Anaplasma phagocytophilum (Ap) were amplified and sequenced. Most Ixodes spp. from the Northeast (n = 115/115; 100%) and the Midwest (n = 77/80; 96.3%) were I. scapularis, American clade. Borrelia spp. were identified in 34 of 192 (17.8%) and Ap in 5 of 192 (2.6%) I. scapularis. Two Ixodes cookei and one Ixodes texanus were identified from Ohio, Illinois, and Michigan. In contrast, 156 of 261 (59.8%) Ixodes spp. from the Southeast were I. scapularis, American clade; 86 of 261 (33.0%) were I. scapularis, Southern clade; 9 of 261 (3.4%) were Ixodes affinis; and 10 of 261 (3.8%) were I. cookei. Southern clade was significantly more common in Florida and less common in the upper South (p < 0.0001). One I. scapularis (1/242; 0.4%) from the Southeast (Kentucky) tested positive for Bb and 6 of 242 (2.5%) were positive for Ap. In the West, most (34/44; 77.3%) Ixodes spp. were Ixodes pacificus, with Ixodes angustus (n = 6) submitted from dogs in Alaska, Washington, and Oregon and Ixodes haerlei (n = 4) preliminarily identified from a dog in Montana. Pathogens were not detected in any ticks from the West. Although I. scapularis, American clade, predominated in the Northeast and Midwest, additional Ixodes spp. were found on dogs and cats in other regions and pathogens were less commonly detected. The role of less common Ixodes spp. as disease vectors, if any, warrants continued investigation.

Recent reports of winter tick, Dermacentor albipictus, from dogs and cats in North America.

Dermacentor albipictus, a common one-host tick of large animals in North America, is most often reported from moose (Alces alces) and is rarely implicated as a parasite of cats and dogs. From 2018 to 2020, 4 dogs and 4 cats from United States and 3 dogs from Canada were infested with D. albipictus. The specimens were collected and submitted to university diagnostic specialists by veterinary clinics in Indiana, Kentucky, Minnesota, Colorado, Wyoming, Utah, Alberta, and British Columbia between the months of October to February (United States) and April to June (Canada). Six adults and five nymphal D. albipictus were collected in the United States while three adults were collected from pets in Canada, and most often a single D. albipictus was present. Identification of specimens collected in the United States were confirmed by amplification and sequencing of 16S rRNA and ITS-2 gene fragments. Rickettsia spp. were not detected in any D. albipictus collected in the United States by 17 kDa-based PCR. As tick populations continue to increase and expand in North America, correct identification of ticks collected from pets is critical to accurately track the progression and spread of ticks and tick-borne diseases.

Heartworm prevalence in dogs versus cats: Multiple diagnostic modalities provide new insights.

The cornerstones of diagnosis of heartworm (HW) in dogs are the detection of circulating antigen from adult female Dirofilaria immitis or the visualization of microfilariae in whole blood. These tests are less sensitive in cats because of the feline immune response leading to low numbers of adult worms, but heartworm antibody tests are also licensed for use in cats. HW antibodies in cats are detectable when there has at least been larval development in the tissues, but positive antibody tests cannot distinguish between current and previous larval infections; thus, cats with positive antibody test results are considered currently or previously infected with D. immitis. The aim of the present study was to use multiple HW diagnostic modalities to maximize detection of infection in dogs and cats at high risk of infection and to compare infection prevalence between these two hosts. Blood samples collected from 100 stray dogs and 100 stray cats at Florida animal shelters were tested for HW antigen (before and after heat treatment) and microfilariae; cats were also tested for HW antibody. Dogs were significantly (P = 0.0001) more likely to be diagnosed with adult HW infection (28 %; 95 % CI: 20.1-37.6%) when compared with cats (4 %; 95 % CI: 1.6-10.2%) on the basis of positive antigen and microfilariae test results. Cats with current or previous adult, immature adult, or larval HW infections comprised 19 % (95 % CI: 12.4%-27.9%) of the feline population, which was not significantly different (P = 0.1) from the prevalence of adult D. immitis infection in dogs. Testing unprotected cats for heartworm antibodies demonstrated a similar, high risk of infection to the matched unprotected dog population in Florida, which supports the use of HW preventives in cats in areas where HW transmission occurs.

Haemaphysalis longicornis, the Asian longhorned tick, from a dog in Virginia, USA.

The Asian longhorned tick, Haemaphysalis longicornis, was only recently recognized in North America and has since been identified on a wide range of domestic and wild animal hosts in multiple states throughout the eastern United States. An H. longicornis nymph was submitted for identification from a dog in central Virginia, USA. Identification was made using standard keys and confirmed molecularly. No products are currently FDA label-approved as effective for H. longicornis in the USA; however, many acaricides commercially available in the United States are known to be effective against H. longicornis in other areas of the world where this tick is endemic. Veterinarians should be aware H. longicornis can commonly be found infesting dogs, cats, livestock, and wildlife, and should continue recommending year-round tick prevention for all pets and routine monitoring for tick-borne infections.