2022 AAHA Canine Vaccination Guidelines.

These guidelines are an update and extension of previous AAHA peer-reviewed canine vaccination guidelines published in 2017. Vaccination is a cornerstone of canine preventive healthcare and one of the most cost-effective ways of maintaining a dog's health, longevity, and quality of life. Canine vaccination also serves a public health function by forming a barrier against several zoonotic diseases affecting dogs and humans. Canine vaccines are broadly categorized as containing core and noncore immunizing antigens, with administration recommendations based on assessment of individual patient risk factors. The guidelines include a comprehensive table listing canine core and noncore vaccines and a recommended vaccination and revaccination schedule for each vaccine. The guidelines explain the relevance of different vaccine formulations, including those containing modified-live virus, inactivated, and recombinant immunizing agents. Factors that potentially affect vaccine efficacy are addressed, including the patient's prevaccination immune status and vaccine duration of immunity. Because animal shelters are one of the most challenging environments for prevention and control of infectious diseases, the guidelines also provide recommendations for vaccination of dogs presented at or housed in animal shelters, including the appropriate response to an infectious disease outbreak in the shelter setting. The guidelines explain how practitioners can interpret a patient's serological status, including maternally derived antibody titers, as indicators of immune status and suitability for vaccination. Other topics covered include factors associated with postvaccination adverse events, vaccine storage and handling to preserve product efficacy, interpreting product labeling to ensure proper vaccine use, and using client education and healthcare team training to raise awareness of the importance of vaccinations.

Novel Penicillium species causing disseminated disease in a Labrador Retriever dog.

This report describes the phenotypic characteristics of a novel Penicillium species, Penicillium labradorum, isolated from a 3-year-old male, castrated, Labrador retriever with disseminated fungal disease. The dog's presenting clinical signs included lethargy, lymphadenopathy, tachypnea, moderate pitting edema, and nonweight bearing lameness associated with the right hind limb. Fine-needle aspirate biopsies from the sublumbar and prescapular lymph nodes were initially examined. The cytologic findings were consistent with pyogranulomatous inflammation with abundant extracellular and phagocytized fungal fragments and hyphae. Based on the morphology of the organisms and lack of endogenous pigment, hyalohyphomycosis was considered most likely, with Fusarium, Penicillium, and Paecilomyces species being considerations. Fungal isolates were obtained via culture of samples from the lymph nodes, and molecular identification testing originally identified an undescribed Penicillium species belonging to the Penicillium section Exilicaulis. BLAST searches and phylogenetic analyses performed approximately 1 year and 9 months after the isolation date revealed an isolate within the Penicillium parvum clade in the Penicillium section Exilicaulis but phylogenetically distant from the other species in the section, thus representing a new species, Penicillium labradorum. Antifungal susceptibility testing was also performed on the isolate and low minimum inhibitory concentrations were observed with terbinafine, voriconazole, and posaconazole, while in vitro resistance was observed with fluconazole. The dog had been previously treated with fluconazole, itraconazole, amphotericin B lipid complex, voriconazole, and terbinafine. Approximately 587 days after the initial diagnosis, the dog was euthanized due to worsening of clinical signs and concerns for quality of life.

Identification of Cytauxzoon felis antigens via protein microarray and assessment of expression library immunization against cytauxzoonosis.

BACKGROUND: Cytauxzoonosis is a disease of felids in North America caused by the tick-transmitted apicomplexan parasite Cytauxzoon felis. Cytauxzoonosis is particularly virulent for domestic cats, but no vaccine currently exists. The parasite cannot be cultivated in vitro, presenting a significant limitation for vaccine development. METHODS: Recent sequencing of the C. felis genome has identified over 4300 putative protein-encoding genes. From this pool we constructed a protein microarray containing 673 putative C. felis proteins. This microarray was probed with sera from C. felis-infected and naïve cats to identify differentially reactive antigens which were incorporated into two expression library vaccines, one polyvalent and one monovalent. We assessed the efficacy of these vaccines to prevent of infection and/or disease in a tick-challenge model. RESULTS: Probing of the protein microarray resulted in identification of 30 differentially reactive C. felis antigens that were incorporated into the two expression library vaccines. However, expression library immunization failed to prevent infection or disease in cats challenged with C. felis. CONCLUSIONS: Protein microarray facilitated high-throughput identification of novel antigens, substantially increasing the pool of characterized C. felis antigens. These antigens should be considered for development of C. felis vaccines, diagnostics, and therapeutics.

18 F-FDG-PET/CT as adjunctive diagnostic modalities in canine fever of unknown origin.

Fever of unknown origin (FUO) is a persistent or recurrent fever for which the underlying source has not been identified despite diagnostic investigation. In people, 18 F-fluoro-2-deoxyglucose positron emission tomography (18 F-FDG-PET) alone or in combination with computed tomography (CT) is often beneficial in detecting the source of fever when other diagnostics have failed. Veterinary reports describing use of these modalities in animals with fever of unknown origin are currently lacking. Aims of this retrospective case series were to describe 18 F-FDG-PET or 18 F-FDG-PET/CT findings in a group of dogs with fever of unknown origin. Dogs presenting to a single center between April 2012 and August 2015 were included. A total of four dogs met inclusion criteria and underwent either positron emission tomography (n = 2) or positron emission tomography/CT (n = 2) as a part of their diagnostic investigation. All subjects underwent extensive diagnostic testing prior to 18 F-FDG-PET/CT. Initial diagnostic evaluation failed to identify either a cause of fever or an anatomic location of disease in these four dogs. In each dog, positron emission tomography or positron emission tomography/CT was either able to localize or rule out the presence of focal lesion thereby allowing for directed sampling and/or informed disease treatment. Follow up 18 F-FDG-PET/CT scans performed in two patients showed improvement of observed abnormalities (n = 1) or detected recurrence of disease allowing for repeated treatment before clinical signs recurred (n = 1). Fever resolved after specific treatment in each dog. Findings from the current study supported the use of positron emission tomography or positron emission tomography/CT as adjunctive imaging modalities for diagnosis and gauging response to therapy in dogs with fever of unknown origin.

Rapid High-Resolution Melt Analysis of Cytauxzoon felis Cytochrome b To Aid in the Prognosis of Cytauxzoonosis.

Cytauxzoon felis is a virulent, tick-transmitted, protozoan parasite that infects felines. Cytauxzoonosis was previously thought to be uniformly fatal in domestic cats. Treatment combining atovaquone and azithromycin (A&A) has been associated with survival rates of over 60%. Atovaquone, a ubiquinone analogue, targets C. felis cytochrome b (cytb), of which 30 unique genotypes have been identified. The C. felis cytb genotype cytb1 is associated with increased survival rates in cats treated with A&A. The purpose of this study was to design a PCR panel that could distinguish C. felis cytb1 from other cytochrome b genotypes. Primer pairs were designed to span five different nucleotide positions at which single-nucleotide polymorphisms in the C. felis cytb gene had been identified. Through the use of high-resolution melt analysis, this panel was predicted to distinguish cytb1 from other cytb genotypes. Assays were validated using samples from 69 cats with cytauxzoonosis for which the C. felis cytb genotypes had been characterized previously. The PCR panel identified C. felis cytb1 with 100% sensitivity and 98.2% specificity. High-resolution melt analysis can rapidly provide prognostic information for clients considering A&A treatment in cats with cytauxzoonosis.

COMPARISON OF LUNG ATTENUATION AND HETEROGENEITY BETWEEN CATS WITH EXPERIMENTALLY INDUCED ALLERGIC ASTHMA, NATURALLY OCCURRING ASTHMA AND NORMAL CATS.

Airway remodeling is a prominent feature of feline allergic asthma but requires biopsy for characterization. Computed tomography (CT) has appeal as a minimally invasive diagnostic test. The purpose of this prospective case-control study was to compare indices of airway remodeling between cats with experimentally induced, spontaneous asthma and healthy unaffected cats using CT. We hypothesized that experimental and spontaneous feline asthma would have similar CT airway remodeling characteristics and that these would be significantly different in healthy cats. Experimentally induced asthmatic research cats (n = 5), spontaneously asthmatic pet cats (n = 6), and healthy research cats (n = 5) were scanned unrestrained using a 64-detector row CT scanner. Inspiratory breath-hold CT scans were also performed in experimentally induced asthmatic and healthy cats. Mean ± extent variation of lung attenuation for each cat was determined using an airway inspector software program and CT images were scored for lung heterogeneity by a board-certified veterinary radiologist who was unaware of cat group status. Groups were compared using one-way ANOVA (unrestrained scans) and the Student's t-test (anesthetized scans) with significance defined as P < 0.10. Experimentally asthmatic and spontaneously asthmatic cats had significantly (P = 0.028 and P = 0.073, respectively) increased lung attenuation compared to healthy cats. Heterogeneity scores were higher in experimentally induced asthmatic cat than in healthy cats. Objective quantification of lung heterogeneity and lung volume did not differ among the three groups (P = 0.311, P = 0.181, respectively). Findings supported our hypothesis. Inspiratory breath-hold anesthetized CT scans facilitated discrimination between asthmatic and healthy cats in comparison to unrestrained CT scans.

Pharmacogenomics of Cytauxzoon felis cytochrome b: implications for atovaquone and azithromycin therapy in domestic cats with cytauxzoonosis.

Cytauxzoon felis, an emerging virulent protozoan parasite that infects domestic cats, is treated with atovaquone and azithromycin (A&A). Atovaquone targets parasite cytochrome b. We characterized the C. felis cytochrome b gene (cytb) in cats with cytauxzoonosis and found a cytb genotype that was associated with survival in A&A-treated cats.

Oral administration of an extended-release formulation of nitrofurantoin results in high concentrations in the urine of dogs.

OBJECTIVE: Sporadic bacterial cystitis in both dogs and humans is often caused by Escherichia coli. In humans, nitrofurantoin is a first-line antimicrobial for the treatment of bacterial cystitis but in dogs a lack of available data may be part of the reason it is only recommended as a second-line treatment. The objective of this preliminary study was to determine the plasma pharmacokinetics and urine concentrations of nitrofurantoin monohydrate-macrocrystalline in dogs. ANIMALS: 8 healthy female hound dogs. PROCEDURES: From July 26 to July 28, 2021, dogs received a single oral dose of nitrofurantoin monohydrate-macrocrystalline 100 mg with food. Blood and urine were collected at predetermined times. Nitrofurantoin concentrations were assayed by UPLC-MS/MS and plasma data were analyzed using noncompartmental methods. RESULTS: Plasma concentrations were low for all dogs with a mean ± SD maximum concentration (Cmax) of 0.242 ± 0.098 µg/mL (range, 0.14 to 0.42 µg/mL) occurring between 2 and 24 hours. Urine concentrations were manyfold higher than for plasma. Cmax in urine was 134 ± 54 µg/mL (range, 49.1 to 218 µg/mL) occurring between 6 and 36 hours. As seen in other species, nitrofurantoin concentrated in urine with concentrations being 500 times higher than the concentration in plasma. CLINICAL RELEVANCE: Results suggested that nitrofurantoin monohydrate-macrocrystalline formulation of nitrofurantoin should be effective in treating bacterial cystitis caused by susceptible uropathogens.

Evaluation of serum 25-hydroxyvitamin D, C-reactive protein, and haptoglobin as biomarkers in dogs newly diagnosed with histoplasmosis.

BACKGROUND: Serum 25-hydroxyvitamin (OH)D, C-reactive protein (CRP), and haptoglobin are useful biomarkers in various infectious diseases and inflammatory disorders in dogs, but their utility in histoplasmosis is unknown. OBJECTIVE: Determine if serum 25(OH)D, CRP, and haptoglobin concentrations are different in dogs with histoplasmosis compared to healthy controls and whether serum globulin, albumin, CRP, or haptoglobin are associated with 25(OH)D concentration. ANIMALS: Twenty-two client-owned dogs (histoplasmosis, n = 12; controls, n = 10). METHODS: Prospective case-control study. Dogs with histoplasmosis were categorized as pulmonary, disseminated, or gastrointestinal (GI) tract. Serum 25(OH)D was measured using modified high-performance liquid chromatography (HPLC). Serum CRP and haptoglobin were measured with ELISA assays. RESULTS: Dogs with histoplasmosis were grouped as disseminated (n = 8) and GI tract (n = 4). No dogs had pulmonary tract involvement alone. Dogs with histoplasmosis (median, interquartile range [IQR]; 11.6 ng/mL, 16.8) had lower serum 25(OH)D concentrations than controls (35.7 ng/mL, 17.6; P < .001). Serum CRP and haptoglobin concentrations were higher in dogs with histoplasmosis (CRP: median, IQR; 63.5 mg/L, 37.1 and haptoglobin: 459.7 mg/dL, 419.6) than controls (CRP: 1.9 mg/L, 2; P < .001 and haptoglobin: 85.5 mg/dL, 106.7; P = .003). Serum 25(OH)D concentration was positively associated with fold change in serum albumin concentration (ρ = 0.77; P < .001), and negatively associated with fold change in serum globulin (ρ = -0.61; P = .003) and CRP concentrations (ρ = -0.56; P = .01). CONCLUSION AND CLINICAL IMPORTANCE: Assay of serum 25(OH)D, CRP, and haptoglobin could have clinical value in dogs with histoplasmosis.

Urinary cystatin B differentiates progressive versus stable IRIS Stage 1 chronic kidney disease in dogs.

BACKGROUND: Early identification of dogs with progressive vs stable chronic kidney disease (CKD) might afford opportunity for interventions that would slow progression. However, currently no surrogate biomarker reliably predicts CKD progression. HYPOTHESIS/OBJECTIVES: Urinary cystatin B (uCysB), a novel kidney injury biomarker, predicts progressive disease in International Renal Interest Society (IRIS) CKD Stage 1. ANIMALS: Seventy-two dogs, including 20 dogs from 4 university centers with IRIS CKD Stage 1, with IDEXX symmetric dimethylarginine (SDMA) concentration up to 17 µg/dL and no systemic comorbidities, and 52 clinically healthy staff-owned dogs from a fifth university center. METHODS: A multicenter prospective longitudinal study was conducted between 2016 and 2021 to assess uCysB concentration in IRIS CKD Stage 1 and control dogs. Dogs were followed to a maximum of 3 years (control) or 25 months (CKD). Stage 1 IRIS CKD was classified as stable or progressive using the slope of 1/SDMA, calculated from 3 timepoints during the initial 90-day period. Dogs with slope above or below -0.0007 week × dL/µg were classified as stable or progressive, respectively. Mixed effects modeling was used to assess the association between uCysB and progression rate. RESULTS: Estimates of first visit uCysB results predictive of active ongoing kidney injury based on the mixed effects models were 17 ng/mL for control, 24 ng/mL for stable CKD, and 212 ng/mL for progressive CKD (P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: Urinary cystatin B differentiated stable vs progressive IRIS CKD Stage 1. Identification of dogs with progressive CKD may provide an opportunity for clinicians to intervene early and slow progression rate.

Differentiation of stable kidney function versus progressive dysfunction in dogs.

BACKGROUND: Circulating creatinine and symmetric dimethylarginine (SDMA) are biomarkers of kidney function that have been used variously to define stable vs progressive chronic kidney disease (CKD). Slope monitoring of inverse biomarker values (creatinine-1 or SDMA-1 ) has shown promise, but quantitative criteria to distinguish stable vs progressive CKD using this approach are lacking. OBJECTIVE: Assessment of creatinine-1 and SDMA-1 slope cutoffs to distinguish stable vs progressive CKD. ANIMALS: One hundred ten clinically healthy university staff-owned dogs and 29 male colony dogs with progressive X-linked hereditary nephropathy (XLHN). METHODS: Retrospective analysis combining 2 prospective observational studies, 1 tracking kidney function biomarkers in healthy dogs (HDs) to a maximum of 3 years, and 1 tracking kidney function biomarkers in male colony dogs with progressive XLHN to a maximum of 1 year. The minimum slope of creatinine-1 or SDMA-1 as measured using the IDEXX SDMA test from HD was assigned as the slope cutoff for stable kidney function. RESULTS: The stable vs progressive slope cutoff was -0.0119 week × dL/mg for creatinine-1 and -0.0007 week × dL/µg for SDMA-1 . CONCLUSIONS AND CLINICAL IMPORTANCE: In the studied CKD population, progressive dysfunction can be distinguished from stable kidney function by using the slope of creatinine-1 or SDMA-1 . These criteria may serve to characterize CKD in other cohorts of dogs and to establish guidelines for degrees of progression rate in dogs with naturally occurring CKD.

Prevalence of antibiotic use for dogs and cats in United States veterinary teaching hospitals, August 2020.

BACKGROUND: Awareness of prescribing practices helps identify opportunities to improve antibiotic use (AU). OBJECTIVES: To estimate AU prevalence in dogs and cats in U.S. veterinary teaching hospitals (VTHs) and identify antibiotic drugs commonly prescribed, indications for use, and evidence of bacterial infection. ANIMALS: Medical record data were collected from dogs and cats examined at 14 VTHs. METHODS: Data were collected from VTH medical records of dogs and cats examined by primary care, urgent care, emergency and critical care, internal medicine, and surgery services on a single day during August 13-September 3, 2020. Data included signalment; clinical service; inpatient or outpatient status; clinical conditions; diagnostic tests; evidence of bacterial infection; intended reason for AU; name and route of antibiotics prescribed. RESULTS: Of 883 dogs and cats, 322 (36.5%) were prescribed at least 1 antibiotic. Among 285 antibiotics administered systemically intended for treatment of infection, 10.9% were prescribed without evidence of infection. The most common class of antibiotics presribed for systemic administration was potentiated penicillin for dogs (115/346, 33.3%) and cats (27/80, 33.8%). For dogs and cats, first-generation cephalosporins (93/346, 26.9% and 11/80, 13.8%, respectively) and fluoroquinolones (51/346, 14.7% and 19/80, 23.8%, respectively) was second or third most-prescribed. Common AU indications included skin, respiratory, and urinary conditions, and perioperative use. CONCLUSIONS AND CLINICAL IMPORTANCE: Collaborative data collection provides a sustainable methodology to generate national AU prevalence estimates and bring attention to areas requiring additional research and detailed data collection. These efforts can also identify practice improvement opportunities in settings where future veterinarians are trained.

The tyrosine kinase inhibitor masitinib blunts airway inflammation and improves associated lung mechanics in a feline model of chronic allergic asthma.

BACKGROUND: Blockade of tyrosine kinase signaling by masitinib, a c-kit/PDGF receptor tyrosine kinase inhibitor, can modulate allergic airway inflammation, but effects on lung mechanics have not been well characterized. We hypothesized masitinib would decrease airway eosinophilia and consequently improve pulmonary mechanics in a feline allergic asthma model. METHODS: Asthma was induced in 12 cats using Bermuda grass allergen (BGA). Cats received 50 mg/day oral masitinib or placebo. Bronchoalveolar lavage fluid (BALF) was analyzed for eosinophils, total protein (TP) and BGA-specific IgE. Ventilator-acquired mechanics after methacholine (MCh) challenge determined MCh concentration needed to increase baseline airway resistance by 200% (EC(200)R(aw)), positive end expiratory occlusion pressure (PEEP) and end inspiratory breath hold pressure (P(plat)). An inverse correlate of respiratory system compliance P(plat)-PEEP was also calculated. Data were analyzed using the Wilcoxon test, with one-tailed significance set at p < 0.1. RESULTS: After 4 weeks, percent eosinophils in BALF was lower in masitinib-treated cats (7 ± 9%) versus controls (30 ± 27%, p = 0.023). BALF TP significantly differed (p = 0.047) between groups, decreasing with masitinib and increasing with placebo. BALF BGA-specific IgE was unaffected by masitinib. Both groups showed an improvement in EC(200)R(aw) (masitinib, p = 0.015; control, p = 0.078) but no significant change in PEEP after 4 weeks. Masitinib-treated cats demonstrated decreased P(plat) (p = 0.033) and P(plat)-PEEP (p = 0.075) at week 4, suggesting an improvement in respiratory compliance. CONCLUSIONS: Masitinib reduced BALF eosinophilia and TP, indicating improved airway inflammation and edema, and improved P(plat) and P(plat)-PEEP, suggesting benefit to respiratory compliance influenced by airway inflammation/edema. Masitinib deserves further study in humans with chronic allergic asthma.

Successful treatment of acute respiratory distress syndrome in 2 dogs.

Acute respiratory distress syndrome (ARDS) was diagnosed in 2 dogs with acute dyspnea. Short-term positive pressure ventilation and intense critical and nursing care were provided. Both dogs improved and were discharged. Few reports describe successful recovery from ARDS. Due to advances in positive pressure ventilation and improvement in the supportive care of critically ill veterinary patients, the prognosis for ARDS may improve.

Cytauxzoonosis.

Cytauxzoon felis is a hematoprotozoan parasite with a complex life cycle involving a tick-vector and a mammalian host. The mammalian hosts are all felidae but in the bobcat reservoir host, the parasite typically causes only a brief, self-resolving illness followed by a prolonged subclinical infection. In domestic cats, however, infection often leads to an acute febrile illness characterized by severe morbidity and mortality. Diagnosis is based on microscopic identification of parasites or molecular testing. Treatment for ill cats is expensive, difficult, and often unsuccessful. Prevention is quite possible and depends on avoidance of feeding by vector ticks.

Fluid Therapy for Pediatric Patients.

Pediatric dogs and cats within their first 12 weeks of life have important electrolyte requirements and physiologic considerations that may impact fluid therapy. Fluid requirements are higher in pediatrics, while fluid losses are greater due to underdeveloped physiologic responses. Hydration and volume status are difficult to assess in young animals, and their small size makes intravenous (IV) access difficult to obtain. Young patients can quickly deteriorate from dehydration, poor husbandry, and infection and become critically ill, requiring prompt recognition, treatment, intensive care, and monitoring. Clinicians should be aware of all available routes of fluid administration including oral, subcutaneous (SC), intraperitoneal (IP), IV, and intraosseous (IO), and the limitations associated with each route.

Etiology and outcome of extreme neutrophilic leukocytosis: A multi-institutional retrospective study of 269 dogs.

BACKGROUND: The magnitude of diagnostic abnormalities can influence the perception of clinical outcome. Extreme neutrophilic leukocytosis (ENL) is an uncommon finding caused by markedly increased granulopoiesis. A lack of recent, large-scale studies limits our understanding of the importance, causation, and prognosis associated with ENL in dogs. HYPOTHESIS/OBJECTIVES: Describe disease categories (DC) identified in dogs with ENL and identify variables associated with survival. We hypothesized that factors including fever, segmented and band neutrophil counts, and DC would be negatively associated with survival. ANIMALS: Two-hundred sixty-nine dogs with ENL (segmented neutrophils ≥50 × 103  cells/µL) presented to the veterinary teaching hospitals at Auburn University (n = 164), the University of Missouri (n = 81), and Oklahoma State University (n = 24) between January 1, 2009 and December 31, 2019. METHODS: Retrospective study. Demographic data and outcome variables including temperature, CBC findings, DC, duration of hospitalization (DOH) and outcome were acquired from the medical record. Statistical analyses included chi-squared and Kruskal-Wallis tests, and Pearson product moment correlations with a P < .05 significance level. RESULTS: Mortality was 41%. Survival differed with DC (P = .002). Mortality was higher (P < .05) in dogs with neoplasia (56.2%) vs immune-mediated disease (20.5%) or tissue damage/necrosis (19%). Weight (P = .001, r = -0.14) and total neutrophil count (P = .04, r = -0.02) were weakly negatively associated with survival whereas DOH was weakly positively associated with survival (P = .03, r = 0.14). CONCLUSIONS AND CLINICAL IMPORTANCE: Mortality in dogs with ENL is high but differed according to DC. Only weak correlations between clinical or clinicopathologic variables and mortality were identified. Extreme neutrophilic leukocytosis should be interpreted in conjunction with the underlying disease process, and not broadly used to predict clinical outcome.

Evaluation of various sample sources for the cytologic diagnosis of Cytauxzoon felis.

BACKGROUND: Cytauxzoon felis is a life-threatening protozoan disease of cats. Identification of schizont-laden macrophages is a point-of-care diagnostic test for acute cytauxzoonosis. HYPOTHESIS/OBJECTIVES: The primary objective determined cytologic agreement between sample types to diagnose acute cytauxzoonosis. The secondary objective evaluated novices' ability to identify cytauxzoon organisms in blood films and tissue aspirates. ANIMALS: Thirty-eight cats with suspected acute cytauxzoonosis and 5 controls examined postmortem. METHODS: Cases were prospectively submitted and collected. Blood film, lymph node, and splenic aspirates were blindly reviewed for sample quality, presence of schizont-laden macrophages, and agreement between sample types. A subset of cases and controls were evaluated by 12 blinded novice observers to determine sensitivity and specificity for identifying organisms in various sample types. RESULTS: Acute cytauxzoonosis diagnosis was made on at least 1 sample type in 28/38 cats. Schizont-laden macrophages were seen on 33% (10/30) of blood films, 56% (19/34) lymph node aspirates, 77% (26/34) splenic aspirates. Schizont-laden macrophages were more likely seen on splenic than lymph node aspirates (McNemar's, P = .03) or blood film (McNemar's, P = <.001). Novice observers were more likely to agree with experts when identifying schizont-laden macrophages in splenic aspirates (sensitivity = 77.1%, specificity = 94.4%) versus lymph node aspirates (sensitivity = 52.8%, specificity = 96.4%) or blood films (sensitivity = 41.7%, specificity = 96.9%). CONCLUSIONS AND CLINICAL IMPORTANCE: Schizont-laden macrophages are most frequently identified in spleen, even by novice observers. If the diagnosis of acute cytauxzoonosis cannot be confirmed via blood film, then splenic, followed by peripheral lymph node aspirates can be considered.

Direct injection of Amblyomma americanum ticks with Cytauxzoon felis.

Cytauxzoon felis is a tick-borne hemoprotozoan parasite that causes life-threatening disease in domestic cats in the United States. Currently, the platforms for C. felis research are limited to natural or experimental infection of domestic cats. This study aims to develop an alternative model by infecting Amblyomma americanum ticks with C. felis via direct injection. Amblyomma americanum adults were injected with C. felis-infected feline erythrocytes through two routes: directly into the digestive tract through the anal pore (IA injection), or percutaneously into the tick hemocoel (IH injection). RNAscope® in situ hybridization (ISH) was used to visualize the parasites within the ticks at different time points after injection. Four months after injection, ticks were divided into 3 infestation groups based on injection methods and inoculum type and fed on 3 naïve cats to assess the ticks' ability to transmit C. felis. Prior to the transmission challenge, selected ticks from each infestation group were tested for C. felis RNA via reverse transcription-PCR (RT-PCR). In both IA- and IH-injected ticks, ISH signals were observed in ticks up to 3 weeks after injection. The number of hybridization signals notably decreased over time, and no signals were detected by 4 months after injection. Prior to the transmission challenge, 37-57% of the sampled ticks were positive for C. felis RNA via RT-PCR. While the majority of injected ticks successfully attached and fed to repletion on all 3 cats during the transmission challenge, none of the cats became infected with C. felis. These results suggest that injected C. felis remained alive in ticks but was unable to progress to infective sporozoites after injection. It is unclear why this infection technique had been successful for other closely related tick-borne hemoprotozoa and not for C. felis. This outcome may be associated with uncharacterized differences in the C. felis life cycle, the lack of the feeding or molting in our model or absence of gametocytes in the inoculum. Nonetheless, our study demonstrated the potential of using ticks as an alternative model to study C. felis. Future improvement of a tick model for C. felis should consider other tick species for the injection model or utilize infection methods that more closely emulate the natural infection process.

An Improved Point-of-Care ELISA for the Diagnosis of Anaplasmosis and Ehrlichiosis During the Acute Phase of Tick-Borne Infections in Dogs.

Veterinarians often test for serologic evidence of vector-borne infections in sick dogs presenting with clinical signs or to screen for subclinical chronic infections. Additional peptide targets for the detection of antibodies to Anaplasma phagocytophilum, Anaplasma platys, and Ehrlichia canis were added to an existing point-of-care (POC) ELISA test (SNAP 4Dx Plus Test, IDEXX Laboratories, Westbrook, ME). This second-generation, multi-analyte test detects Dirofilaria immitis antigen and antibodies to Anaplasma spp., Borrelia burgdorferi, and Ehrlichia spp. The second-generation test is expected to better meet the needs of practicing veterinarians and their patients. To assess this expectation, the second-generation POC test was evaluated with serum samples from experimentally infected dogs and a broader field population of dogs. Compared to the first-generation test, most dogs experimentally infected with A phagocytophilum (n = 7/8), A platys (n = 4/6), or E canis (n = 4/6) had detectable antibody responses 3-22 days earlier post-infection; these results demonstrated better alignment with polymerase chain reaction (PCR) amplification results and the onset of clinical signs. Using a convenience sample set of 510 sera from both academic and commercial veterinary diagnostic laboratories, the second-generation test had sensitivities greater than 90% for Anaplasma spp. (94.1%), B burgdorferi (95.5%), Ehrlichia spp. (93.4%) and D immitis (98.0%). Specificity ranged from 96.8% - 100% across the four assays. Results from this study demonstrate that the second-generation POC ELISA had an improved ability to detect serologic responses during the acute phase of A phagocytophilum, A platys, and E canis experimental infections. The results from the broader field samples support overall high sensitivity and specificity, consistent with the historical performance of the first-generation POC ELISA test.