Factors associated with Giardia infection in dogs in southern Ontario, Canada.

Information concerning risk factors associated with Giardia infection in dogs in southern Ontario, Canada, is currently lacking. This study therefore aimed to identify risk factors for Giardia infection in dogs that visit off-leash dog parks in southern Ontario. From May-November 2018, fecal samples were collected from 466 dogs in 12 off-leash dog parks in the Niagara and Hamilton regions of Ontario. A survey that asked questions pertaining to travel history (i.e., area of residence, locations and regions visited in the previous 6 months), basic medical history (i.e., spay/neuter status, veterinary visits, use of deworming medication), consumption of a raw diet, and the physical (i.e., age, sex, breed) and behavioral characteristics (i.e., off-leash activities, hunting activities) of each dog sampled was administered to the respective owner. All fecal samples were examined with the Giardia plate ELISA (IDEXX Laboratories) for parasite antigen. Multivariable logistic regression analyses were conducted on the survey data to investigate putative risk factors for Giardia infection. Overall, 11.8% (95% CI: 9.2-15.1%) of samples tested positive for Giardia antigen. Results from the multivariable logistic regression analyses identified an interaction between dog age and spay/neuter status that was significantly associated with Giardia infection. The odds of infection were greater in intact as compared to neutered adult dogs (OR: 3.6, 95% CI: 1.7-7.9, p = 0.001), and in neutered juvenile dogs as compared to neutered adults (OR: 5.2, 95% CI: 2.2-12.2, p < 0.001). The results provide veterinarians with evidence-based information for identifying dogs at greatest risk of Giardia infection in southern Ontario.

Evaluation of a commercial coproantigen immunoassay for the detection of Toxocara cati and Ancylostoma tubaeforme in cats and Uncinaria stenocephala in dogs.

Coproantigen immunoassays (IDEXX Fecal Dx® antigen tests) were evaluated for their ability to identify Toxocara cati and Ancylostoma tubaeforme infections in cats and Uncinaria stenocephala infection in dogs. Five cats were experimentally infected with 500 embryonated eggs of T. cati, eight cats with 500 third-stage larvae (L3) of A. tubaeforme and seven dogs with 500 L3 of U. stenocephala. In addition to the three coproantigen tests, the course of infection was monitored by a combined sedimentation-flotation method with ZnSO4 as flotation medium (specific gravity: 1.28-1.30) and a modified McMaster method in case of copromicroscopically positive samples. Eggs of T. cati were first observed between 28 and 54 days post infection (dpi) in four of the five infected cats. In these four cats, positive roundworm coproantigen signals were obtained between 16 and 44 dpi. Positive coproantigen signal always preceded egg observations, but the interval varied between 6 and 30 days. Hookworm-specific positive coproantigen signals were detected in seven of the eight A. tubaeforme infected cats between 10 and 52 dpi, while consecutive egg excretion was observed in three cats between day 26 and 54 pi. Of these three, coproantigen signal preceded egg observation by 12 to 24 days. Four cats had positive coproantigen results in the absence of egg excretion, and one cat never achieved a positive result for egg or coproantigen. In six of seven U. stenocephala infected dogs, infection was confirmed by copromicroscopy between 16 and 24 dpi as well as for hookworm coproantigen between 10 and 14 dpi. Coproantigen signal was detected prior to egg observation by 2 to 14 days. No cross-reactions between the roundworm, hookworm und whipworm tests occurred in study animals. The results of this study demonstrate the ability of the coproantigen tests to detect the common roundworm and hookworm infections in cats and U. stenocephala infections in dogs as well as the ability to detect the prepatent stage of infection.

Prevalence of intestinal parasites in dogs in southern Ontario, Canada, based on fecal samples tested using sucrose double centrifugation and Fecal Dx® tests.

In southern Ontario, Canada, there is a lack of information concerning the prevalence of intestinal parasites in dogs. As such, this study aimed to characterize the prevalence of intestinal parasites in dogs visiting off-leash parks in the region using sucrose double centrifugation and Fecal Dx® tests. Additionally, data obtained via the sucrose double centrifugation method were used to evaluate the performance of the Fecal Dx® tests. Fecal samples were collected from 466 dogs aged ≥6 months from May to November 2018 (mean age = 3.7 years). Overall, eleven intestinal parasites were identified using sucrose double centrifugation. Roundworm eggs (Toxocara canis and Baylisascaris procyonis), hookworm eggs (Ancylostoma caninum and Uncinaria stenocephala), and whipworm eggs (Trichuris vulpis) were identified in 1.07% (95% confidence interval [CI] 0.38-2.56%), 5.79% (95% CI 3.85-8.31%), and 5.15% (95% CI 3.33-7.57) of samples, respectively. Using the Fecal Dx® tests, 1.07% (95% CI 0.38-2.56%), 4.29% (95% CI 2.64-6.55%), and 2.15% (95% CI 1.03-3.91) of the samples tested positive for roundworm, hookworm, and whipworm antigen, respectively. To assess the level of agreement between the Fecal Dx® tests and sucrose double centrifugation, three methods were used. Cohen's kappa indicated a fair-to-moderate level of agreement between Fecal Dx® tests and sucrose double centrifugation. In contrast, the prevalence-adjusted bias-adjusted kappa and Gwet's first-order agreement coefficient indicated almost perfect agreement between these tests, ranging from 0.87 to 0.99 among the parasites examined. This study provides valuable information on the prevalence of intestinal parasites in mature dogs in southern Ontario that will help guide parasite control recommendations for dogs in this region.

Experimental comparison of Baylisascaris procyonis definitive host competence between domestic dogs and raccoons (Procyon lotor).

Domestic dogs can function as either paratenic or definitive hosts for the zoonotic raccoon roundworm Baylisascaris procyonis. However, factors leading to development of patent infections in dogs are under-studied. Here we compared infection dynamics of B. procyonis in dogs vs the natural raccoon host. Dogs and raccoons were inoculated 5000 or 500 B. procyonis eggs (n = 3 per dose) or were fed B. procyonis-infected laboratory mice (n = 3 per dose; mice inoculated with 1000 or 250 eggs). Fecal samples were analysed via flotation and a commercial coproantigen ELISA designed for detection of Toxocara spp. Two of 12 dogs (both received low dose larvae) developed patent infections; all 12 raccoons became infected with 10 developing patent infections. Compared with dogs, prepatent periods were shorter in raccoons and maximum egg outputs were much greater. Baylisascaris procyonis coproantigens were detectable via ELISA in all raccoons and the patently infected dogs. Finally, dogs spontaneously lost infections while all patently infected raccoons shed eggs until conclusion of the study. Our results demonstrate that dogs are clearly suboptimal hosts showing limited parasite establishment and fecundity vs raccoons. Despite the low competence, patently infected dogs still pose a risk for human exposure, emphasizing the importance of control measures.

Antigen detection: Insights into Toxocara and other ascarid infections in dogs and cats.

Detection of ascarid excreted or secreted (E/S) molecules is an alternative approach to the identification of infection by egg flotation. E/S molecules serve as direct markers for the ascarid nematode commonly found in cats and dogs (Toxocara spp., Toxascaris leonina and Baylisascaris procyonis). The nematode derived E/S material mixes with the intestinal contents of the host animal and is available for detection as a coproantigen in the host's faeces. Antigen capture immunological techniques allow sensitive coproantigen detection. Different patterns of antigen to egg agreement are demonstrated in an experimental Toxocara canis infection throughout the prepatent, patent, and post-treatment phases. Examination of faecal samples from a large field population of dogs and cats tested for both egg shedding and antigen indicates that more infections were identified by antigen. Host age influences the agreement of antigen and Toxocara egg results. Older dogs and cats were less likely to have a patent infection (egg positive and antigen positive) result pattern. An egg observation in the absence of antigen detection may indicate a spurious egg. The impact of spurious eggs was further examined by comparisons of cohorts of dogs separated by presence or absence of a pseudoparasite observation or by egg semi-quantification bin. Lastly, the antigen to egg agreement was calculated for other ascarid species.

Coproantigen Detection Augments Diagnosis of Common Nematode Infections in Dogs.

Microscopic methods which employ active or passive flotation have been used to detect parasite diagnostic stages in the feces of companion animals for many years. More recently, coproantigen ELISAs for the detection of excretory/secretory products from intestinal nematodes have been introduced. These assays can identify the presence of parasites when eggs are not recovered by flotation (e.g. prepatent infection or intermittent egg shedding). The study was designed to assess the added benefit of these coproantigen tests in canine fecal diagnostics. The work was performed at 3 separate sites where canine fecal samples were each independently evaluated by both centrifugal flotation with an expert examiner (CFE) and passive flotation with a less experienced examiner. All samples were also tested using coproantigen ELISA to detect ascarid, hookworm, or whipworm antigen (IDEXX Laboratories, Inc, Westbrook, Maine). A total of 1202 samples were collected; 626 were from shelter dogs and 576 were from pet dogs. CFE recovered ascarid eggs in 58 samples, hookworm eggs in 229 samples, and whipworm eggs in 95 samples. Of the positive samples identified by CFE, the PFE and ELISA identified 40 and 51 ascarid samples, 188 and 203 hookworm samples, and 65 and 67 whipworm positive samples, respectively. The coproantigen ELISA identified 8 ascarid, 82 hookworm, and 22 whipworm positive samples that were not detected by CFE. The combined results of passive flotation and the coproantigen ELISA improved the percent agreement with centrifugal flotation, suggesting that greater sensitivity of detection may be achieved through the use of complementary diagnostic methods. However, errors of misidentification and poor recovery apparently introduced by less experienced examiners using an inferior flotation method remained. A diagnostic approach that combines coproantigen assays with centrifugal flotation and examination by an expert allows detection of more ascarid, hookworm, and whipworm infections.

Parasite survey on a captive wolf population using classical techniques and ELISA coproantigen detection, USA.

As laws change around the United States, wildlife that were once kept as companion animals are now often confiscated by local authorities. They are then euthanized unless a home is found for them at a sanctuary. Wolf sanctuaries are, therefore, becoming increasingly important for their conservation and management. However, little data is available on best practices for the health management of captive wolves, including data on parasitic diseases. Our objective was to assess the prevalence of parasites of captive wolves combining classical coprological techniques and immunoassays based on the detection of coproantigen of selected canid parasites. Fecal samples of 39 animals were collected upon observation of individual animals defecating. All samples were processed using the Fecal Dx® tests, a suite of coproantigen ELISAs for detection of ascarid, hookworm, whipworm, and Giardia (IDEXX Laboratories Inc.). Out of the 39 samples, 38 were processed using the double-centrifugation sugar flotation (DCSF) and 34 using a modification of the Baermann technique. Twenty-eight samples (71.8%) were positive for hookworm, and none positive for the other parasites tested using coproantigen ELISA. Ancylostoma sp. (26, 68.4%), Eucoleus boehmi (13, 34.2%), and Trichuris sp. (2; 5.3%), and Sarcocystis sp. (13, 34.2%) were detected using DCSF. No metastrongyloid lungworm larvae were found. The Cohen's kappa index (0.97) showed excellent agreement between the hookworm coproantigen ELISA and the DCSF using feces preserved in ethanol for a short period of time. This study provides a baseline on the parasites of captive wolves, and shows that recent innovative diagnostics in veterinary parasitology, developed and optimized for dogs, may be used for assessing the health of wolves.

Diagnosis of feline whipworm infection using a coproantigen ELISA and the prevalence in feral cats in southern Florida.

Trichuris felis, the whipworm of cats, is a relatively rare parasite, although more common in tropical and sub-tropical regions such as the Caribbean and South America. In southern Florida, T. felis is known to occur, but estimating prevalence can be challenging using fecal egg counts due to low intensity and single sex infections. A microplate enzyme-linked immunosorbent assays (ELISAs) for parasite-specific antigen in feces could increase the ability to detect these infections. In this study, the IDEXX Fecal Dx™ antigen ELISA for whipworm (designed for detection of Trichuris vulpis) was evaluated for detection of T. felis using 35 feral cats from St. Kitts, euthanized for non-study related reasons. Twenty-nine of the cats were positive for T. felis with worm counts ranging from 1 to 66 per cat (mean 9.6) and egg counts ranging from 0 to >500 (mean 109.8). The ELISA detected 26 of 29 positive cats while flotation (centrifugation with Sheather's sugar solution) detected 24 of the 29 positive cats. To estimate prevalence in southern Florida, feces from 65 feral cats from the greater Miami area were tested using the ELISA and fecal flotation (centrifugation with zinc sulfate). Twenty-five cats (38%) were identified as positive with the ELISA compared to 17 using fecal flotation. This prevalence was surprising and further investigated by reviewing results of feline samples from Florida submitted to IDEXX Reference Laboratories between 2010 and 2017 and analyzed using fecal flotation. While prevalence was below 1%, there was an apparent trend in increasing prevalence. The results of this study confirm that the IDEXX Fecal Dx™ antigen test for whipworm ELISA, while developed for T. vulpis, can detect T. felis infections. It also suggests that prevalence might be higher than previously known in Florida and might be increasing. Further studies are required to determine the distribution of this higher prevalence and if the distribution and prevalence of T. felis are changing.

Enzyme-linked immunosorbent assays for coproantigen detection of Ancylostoma caninum and Toxocara canis in dogs and Toxocara cati in cats.

We report the development and field validation of 2 ELISAs for the detection of Ancylostoma caninum or Toxocara canis coproantigens in the feces of dogs with experimental and natural infections, and evidence of cross-reactivity with respective feline counterparts. A. caninum-specific coproantigens were detected in feces of experimentally infected dogs starting at 9 d post-infection (dpi), whereas eggs were not seen until 23 dpi. T. canis-specific coproantigens were detected in 3 of 5 experimentally infected dogs by 31 dpi, and 4 of the 5 animals by 38 dpi. T. canis eggs were seen in feces of 4 of the 5 animals by 38 dpi. One dog had delayed coproantigen detection and low egg output. Additionally, 817 canine and 183 feline fecal samples from naturally infected animals tested by flotation were subjected to coproantigen ELISA testing. Of these 1,000 canine and feline samples, 13 and 23 samples, respectively, were positive for "hookworm" or "roundworm" eggs; 19 and 26 samples were ELISA positive, respectively. The T. canis ELISA detected T. cati coproantigen in cat fecal samples. Discrepant ELISA and flotation results were obtained for 16 hookworm- and 13 roundworm-positive samples. Re-examination of the egg-positive, ELISA-negative samples indicated several instances of possible misidentification or coprophagy, whereas detection of antigen in samples without egg observations is likely a reflection of true infection status with egg shedding below detection levels. There is good indication, based on accumulated field data, that these antigen tests also detect other hookworm and ascarid species.

Enzyme-linked immunosorbent assay for coproantigen detection of Trichuris vulpis in dogs.

Infections with Trichuris vulpis, the canine whipworm, may be challenging to diagnose even though characteristic bipolar eggs are shed by mature worms and may be recovered from feces. Decreased detection sensitivities because of using flotation solutions with specific gravities <1.3 and a lengthy prepatent period can lessen the diagnostician's ability to detect infection. Coproantigen detection in feces is becoming an accepted form of diagnosing parasitic infections and can circumvent some of the factors that affect egg recovery. The development of an enzyme-linked immunosorbent assay (ELISA) for the detection of whipworm-specific coproantigens in the feces of dogs with experimental and natural T. vulpis infections is reported herein. Whipworm-specific coproantigens were evidenced in feces from experimentally infected dogs using the newly developed ELISA starting as early as day 23 postinfection, while eggs were not detected in feces until day 69. In addition, 1,156 field fecal samples were tested using fecal flotation methods and the newly developed whipworm ELISA. Of these, 27 samples were found by flotation to be whipworm egg positive, while 35 had detectable antigen on the ELISA. Discrepant results were obtained in 12 samples; 2 egg-positive samples tested ELISA negative, and 10 ELISA-positive samples did not contain detectable egg levels. Using the fecal ELISA for the detection of whipworms in dogs should allow for earlier detection of infection, aid the identification of cases in the face of low egg shedding, and increase detection sensitivity as most commercial laboratories are using flotation solutions not optimal for T. vulpis egg detection.

Pharmacological and pharmacokinetic characterization of the cannabinoid receptor 2 agonist, GW405833, utilizing rodent models of acute and chronic pain, anxiety, ataxia and catalepsy.

To date, two cannabinoid receptors have been identified, CB1 and CB2. Activation of these receptors with non-selective cannabinoid receptor agonists reduces pain sensitivity in animals and humans. However, activation of CB1 receptors is also associated with central side effects, including ataxia and catalepsy. More recently, a role for selective CB2 agonists in pain modification has been demonstrated. GW405833, a selective CB2 agonist, was recently reported to partially reverse the inflammation and hyperalgesia in a rat model of acute inflammation. In the current report, we extend the characterization and therapeutic potential of this compound. For the first time, we show that GW405833 selectively binds both rat and human CB2 receptors with high affinity, where it acts as a partial agonist (approximately 50% reduction of forskolin-mediated cAMP production compared to the full cannabinoid agonist, CP55,940). We also report for the first time that intraperitoneal administration of GW405833 (0.3-100 mg/kg) to rats shows linear, dose-dependent increases in plasma levels and substantial penetration into the central nervous system. In addition, GW405833 (up to 30 mg/kg) elicits potent and efficacious antihyperalgesic effects in rodent models of neuropathic, incisional and chronic inflammatory pain, the first description of this compound in these models. In contrast, analgesia, sedation and catalepsy were not observed in this dose range, but were apparent at 100 mg/kg. Additionally, GW405833 was not antihyperalgesic against chronic inflammatory pain in CB2 knockout mice. These data support the tenet that selective CB2 receptor agonists have the potential to treat pain without eliciting the centrally-mediated side effects associated with non-selective cannabinoid agonists, and highlight the utility of GW405833 for the investigation of CB2 physiology.