ACVIM consensus statement on the diagnosis of immune thrombocytopenia in dogs and cats.

Immune thrombocytopenia (ITP) is the most common acquired primary hemostatic disorder in dogs. Immune thrombocytopenia less commonly affects cats but is an important cause of mortality and treatment-associated morbidity in both species. Immune thrombocytopenia remains a diagnosis of exclusion for which diagnostic guidelines are lacking. Primary, or non-associative, ITP refers to autoimmune platelet destruction. Secondary, or associative, ITP arises in response to an underlying disease trigger. However, evidence for which comorbidities serve as ITP triggers has not been systematically evaluated. To identify key diagnostic steps for ITP and important comorbidities associated with secondary ITP, we developed 12 Population Evaluation/Exposure Comparison Outcome (PECO) format questions. These questions were addressed by evidence evaluators utilizing a literature pool of 287 articles identified by the panelists using a structured search strategy. Evidence evaluators, using panel-designed templates and data extraction tools, summarized evidence and created guideline recommendations that then were integrated by diagnosis and comorbidity domain chairs. The revised PECO responses underwent a Delphi survey process to reach consensus on final guidelines. A combination of panel expertise and PECO responses were employed to develop algorithms for diagnosis of ITP in dogs and cats, which also underwent 4 iterations of Delphi review. Comorbidity evidence evaluators employed an integrated measure of evidence (IME) tool to determine evidence quality for each comorbidity; IME values combined with evidence summaries for each comorbidity were integrated to develop ITP screening recommendations, which also were subjected to Delphi review. Commentary was solicited from multiple relevant professional organizations before finalizing the consensus. The final consensus statement provides clinical guidelines for the diagnosis of, and underlying disease screening for, ITP in dogs and cats. The systematic consensus process identified numerous knowledge gaps that should guide future studies. This statement is a companion manuscript to the ACVIM Consensus Statement on the Treatment of Immune Thrombocytopenia.

ACVIM consensus statement on the treatment of immune thrombocytopenia in dogs and cats.

Management of immune thrombocytopenia (ITP) in dogs and cats is evolving, but there are no evidence-based guidelines to assist clinicians with treatment decisions. Likewise, the overall goals for treatment of ITP have not been established. Immunosuppressive doses of glucocorticoids are the first line treatment, but optimal treatment regimens beyond glucocorticoids remain uncertain. Additional options include secondary immunosuppressive drugs such as azathioprine, modified cyclosporine, and mycophenolate mofetil, usually selected based on clinician preference. Vincristine, human IV immunoglobulin (hIVIg), and transfusion of platelet or red blood cell-containing products are often used in more severe cases. Splenectomy and thrombopoietin receptor agonists are usually reserved for refractory cases, but when and in which patient these modalities should be employed is under debate. To develop evidence-based guidelines for individualized treatment of ITP patients, we asked 20 Population Intervention Comparison Outcome (PICO) format questions. These were addressed by 17 evidence evaluators using a literature pool of 288 articles identified by a structured search strategy. Evidence evaluators, using panel-designed templates and data extraction tools, summarized evidence and created guideline recommendations. These were integrated by treatment domain chairs and then refined by iterative Delphi survey review to reach consensus on the final guidelines. In addition, 19 non-PICO questions covering scenarios in which evidence was lacking or of low quality were answered by expert opinion using iterative Delphi surveys with panelist integration and refinement. Commentary was solicited from multiple relevant professional organizations before finalizing the consensus. The rigorous consensus process identified few comparative treatment studies, highlighting many areas of ITP treatment requiring additional studies. This statement is a companion manuscript to the ACVIM Consensus Statement on the Diagnosis of Immune Thrombocytopenia in Dogs and Cats.

How Changing Tick-Borne Disease Prevalence in Dogs Affects Diagnostic Testing.

Ixodes scapularis (the deer tick), Amblyomma americanum (the lone star tick) and Rhipicephalus sanguineus (the brown dog tick) are ticks that commonly parasitize dogs in the United States. In the first part of this article, we will examine their changing epidemiology to illustrate how being aware of their distribution and adapting diagnostic testing to include a broad range of pathogens may improve our ability to identify and help infected patients, especially those with suspected idiopathic immune-mediated disease. We will then discuss how to optimize testing for these pathogens using available panels.

Emerging Spotted Fever Rickettsioses in the United States.

Spotted fever rickettsioses are important causes of emerging infectious disease in the United States and elsewhere. Rocky Mountain Spotted Fever, caused by R. rickettsii causes a febrile, acute illness in dogs. Because it circulates in peripheral blood in low copy number and because of the acute nature of the disease, dogs may test PCR and seronegative at the time of presentation. Therefore, therapy with doxycycline must be initiated and continued based on the clinician's index of suspicion. Combining PCR with serologic testing, repeat testing of the same pre-antimicrobial blood sample, and testing convalescent samples for seroconversion facilitates diagnosis. The prognosis can be excellent if appropriate antimicrobial therapy is begun in a timely fashion. It is well established that dogs are sentinels for infection in people in households and communities. Whether R. rickettsii causes illness in cats is not well established. The role of other spotted fever group rickettsia in causing illness in dogs and cats is being elucidated. Veterinarians should keep in mind that novel and well characterized species of SFG Rickettsia are important causes of emerging infectious disease. Veterinarians can play an important role in detecting, defining, and preventing illness in their canine patients and their human companions.

Vector-borne disease and its relationship to hematologic abnormalities and microalbuminuria in retired racing and show-bred greyhounds.

BACKGROUND: Reference intervals for platelets and white blood cell (WBCs) counts are lower in greyhounds than other breeds. Proteinuria is common. Vector-borne diseases (VBD) cause thrombocytopenia, leukopenia, and proteinuria. Racing greyhounds are commonly exposed to vectors that carry multiple organisms capable of chronically infecting clinically healthy dogs. HYPOTHESIS/OBJECTIVES: Vector-borne disease prevalence is higher in retired racing greyhounds than in show-bred greyhounds. Occult infection contributes to breed-related laboratory abnormalities. ANIMALS: Thirty National Greyhound Association (NGA) retired racing and 28 American Kennel Club (AKC) show-bred greyhounds. METHODS: Peripheral blood was tested for Anaplasma, Babesia, Bartonella, Ehrlichia, hemotropic Mycoplasma, and Rickettsia species using PCR. Antibodies to Anaplasma, Babesia, Bartonella, Ehrlichia, and Rickettsia species and Borrelia burgdorferi were detected using immunofluorescence and ELISA assays. Complete blood counts, semiquantitative platelet estimates, and microalbuminuria concentration were determined. RESULTS: Seven of 30 NGA and 1/28 AKC greyhounds tested positive for ≥1 VBD (P = .05). More positive tests were documented in NGA (10/630) than in AKC dogs (1/588; P = .02). Exposure to Bartonella species (3/30), Babesia vogeli (2/30), Ehrlichia canis (1/30), and infection with Mycoplasma hemocanis (3/30) occurred in NGA dogs. Platelet counts or estimates were >170 000/µL. White blood cell counts <4000/µL (4/28 AKC; 5/30 NGA, P > .99; 1/8 VBD positive; 8/51 VBD negative, P = .99) and microalbuminuria (10/21 AKC; 5/26 NGA, P = .06; 1/8 VBD positive; 14/25 VBD negative, P = .41) were not associated with VBD. CONCLUSIONS AND CLINICAL IMPORTANCE: The prevalence of thrombocytopenia and B. vogeli exposure was lower than previously documented. Larger studies investigating the health impact of multiple VBD organisms are warranted.

Effects of storage and leukocyte reduction on the concentration and procoagulant activity of extracellular vesicles in canine packed red cells.

OBJECTIVES: To characterize the size and procoagulant activity of extracellular vesicles (EV) that accumulate in canine packed red blood cells (pRBCs) over time and the effect of leukocyte reduction on these characteristics. DESIGN: Prospective cohort study. SETTING: Private small animal specialty referral hospital and university research laboratories. ANIMALS: Ten healthy blood donor dogs. INTERVENTIONS: Five pRBCs units were obtained according to standard protocols, and 5 were leukocyte-reduced prior to processing. Platelet-free supernatant from the pRBC units was collected on days 0, 10, 20, 32, and 42. MEASUREMENTS AND MAIN RESULTS: Nanoparticle tracking analysis was performed to determine the size and concentration of EVs. Thrombin generation associated with phosphatidylserine-positive EVs was determined using a capture assay. Factor Xa generation associated with phosphatidylserine-positive EVs and tissue factor-positive EVs was measured in a subset of EVs isolated by centrifugation of the supernatant at 20,000 × g. R package nparLD and the Mann-Whitney U-test were used to determine the effect of duration of storage and the effect of leukocyte reduction, respectively. Small (mean < 125 nm) procoagulant EVs accumulated over time, with significant increases occurring on or after day 20 in both non-leukocyte reduced and leukocyte-reduced units. The procoagulant activity of the EVs was due to phosphatidylserine, not tissue factor. Increases in EV concentration and procoagulant activity occurred earlier in non-leukocyte reduced units. Extracellular vesicle accumulation and procoagulant activity were not decreased at any individual time point by leukocyte reduction. CONCLUSIONS: Further studies characterizing and determining the clinical relevance of small procoagulant EVs in pRBCs are warranted.

Prospective evaluation of the influence of sampling method on bacterial culture results and antimicrobial selection in 52 dogs with infected wounds.

OBJECTIVE: To determine the influence of sampling methods on culture results and selection of antimicrobials for treating infected wounds in dogs. DESIGN: Prospective study from January to July 2016. SETTING: Three private multispecialty referral centers. ANIMALS: Fifty-two client-owned dogs with infected wounds. INTERVENTIONS: Each wound was sampled for culture by 3 methods: swabbing prior to preparation (dirty swabs), swabbing after debridement and sterile lavage (clean swabs), and biopsy. Bacterial species and phenotypes were compared. Three clinicians unaware of patient, wound, and sampling information selected antimicrobial drugs based on culture and sensitivity reports. Antimicrobials were divided into class I, II, or III based on established guidelines. The number, highest class of antimicrobial chosen, and inter-investigator agreement were examined. MEASUREMENTS AND MAIN RESULTS: Identical populations of bacteria were isolated for all 3 sampling techniques in only 31% of wounds. Significantly fewer bacterial species were isolated from biopsy samples (1.87 bacterial species per wound ± 1.14) than from clean swab samples (2.29 ± 1.18; P = 0.009) but not dirty swab samples (2.29 ± 1.29; P = 0.06). The recovery frequency for gram-positive bacteria was lower for biopsy compared to either swabbing technique (P = 0.001 for both comparisons). No difference was observed between clean and dirty swabbing techniques for any parameter examined. Sampling technique did not affect the proportion of wounds with anaerobic, gram-negative, or multi-drug resistant bacteria. The number (P = 0.28) and highest class of antimicrobial (P = 0.9) selected per wound did not differ between the 3 sampling techniques (P = 0.28). Clinician agreement was 83-90% depending on sampling technique. CONCLUSION: Although there were some differences in bacteria isolated from biopsy samples compared to swab samples from infected wounds, technique did not influence the number and highest class of antimicrobial selected by clinicians. Wound debridement prior to sampling by swabbing did not alter the number or type of bacteria isolated, nor the number or the highest class of antimicrobial selected by clinicians.

Optimal Vector-borne Disease Screening in Dogs Using Both Serology-based and Polymerase Chain Reaction-based Diagnostic Panels.

Vector-borne disease and idiopathic immune-mediated disease present similarly. Diagnostic panels that include multiple organisms help detect infection and identify coinfections. Comprehensive diagnostic panels that combine polymerase chain reaction (PCR) and serology should be used in initial screening to maximize sensitivity and identify infection. Repeat testing using PCR is warranted in dogs at high risk of infection with organisms that circulate in blood in low numbers or intermittently. Convalescent serologic testing can help diagnose acute infection. This article discusses the pathophysiology and epidemiology of the organisms, panel selection, and how to recognize when more aggressive testing for an organism is warranted.

ACVIM consensus statement on the diagnosis of immune-mediated hemolytic anemia in dogs and cats.

Immune-mediated hemolytic anemia (IMHA) is an important cause of morbidity and mortality in dogs. IMHA also occurs in cats, although less commonly. IMHA is considered secondary when it can be attributed to an underlying disease, and as primary (idiopathic) if no cause is found. Eliminating diseases that cause IMHA may attenuate or stop immune-mediated erythrocyte destruction, and adverse consequences of long-term immunosuppressive treatment can be avoided. Infections, cancer, drugs, vaccines, and inflammatory processes may be underlying causes of IMHA. Evidence for these comorbidities has not been systematically evaluated, rendering evidence-based decisions difficult. We identified and extracted data from studies published in the veterinary literature and developed a novel tool for evaluation of evidence quality, using it to assess study design, diagnostic criteria for IMHA, comorbidities, and causality. Succinct evidence summary statements were written, along with screening recommendations. Statements were refined by conducting 3 iterations of Delphi review with panel and task force members. Commentary was solicited from several professional bodies to maximize clinical applicability before the recommendations were submitted. The resulting document is intended to provide clinical guidelines for diagnosis of, and underlying disease screening for, IMHA in dogs and cats. These should be implemented with consideration of animal, owner, and geographical factors.

Post-chemotherapy perforation in cats with discrete intermediate- or large-cell gastrointestinal lymphoma.

Objectives Gastrointestinal (GI) perforation is a well described complication of GI lymphoma in people, commonly occurring within days of initiation of chemotherapy. There are no studies documenting the prevalence of GI perforation in cats with intermediate- or large-cell GI lymphoma or whether it is associated with induction of chemotherapy. The objectives of this study were to document the prevalence and timing of post-chemotherapy perforation in cats with discrete GI masses caused by intermediate- or large-cell lymphoma. Methods Cats with a diagnosis of intermediate- or large-cell lymphoma based on cytologic or histopathologic examination of a mass lesion of the GI tract and treated with chemotherapy were identified by searching the patient record database of three large specialty referral hospitals. Cats undergoing surgical resection of a GI mass prior to chemotherapy were excluded from the study. A clinical diagnosis of GI perforation was made using ultrasound findings and analysis of abdominal fluid. Results Twenty-three cats with intermediate- (n = 3) or large-cell (n = 20) lymphoma were included in the study. GI perforation was confirmed in 4/23 cats (17%), and occurred at 23, 56, 59 and 87 days after induction. There was no association between tumor size, the presence of hypoproteinemia or suppurative inflammation within the mass at the time of diagnosis and subsequent perforation. Post-hoc analysis revealed that the magnitude of weight loss within 15-28 days of diagnosis was greater in cats with perforation. Conclusions and relevance In this pilot study, we found that post-chemotherapy GI perforation in cats with intermediate- or large-cell GI lymphoma occurs. Acute perforation after induction of chemotherapy was not documented. Larger prospective studies are needed to determine risk factors associated with perforation and whether surgical excision would reduce the risk of subsequent GI perforation in these patients.

Seasonality of immune-mediated hemolytic anemia in dogs from southern California.

OBJECTIVE: To determine if there is seasonal variation in the incidence of canine idiopathic immune-mediated hemolytic anemia (IMHA) at 2 specialty hospitals in southern California. DESIGN: Retrospective study. To determine seasonality, cases of idiopathic IMHA presenting between June 2006 and June 2012 were identified by searching medical record databases of 2 large specialty hospitals. The presence of anemia with autoagglutination, spherocytosis or a positive Coomb's test and no identifiable underlying cause were required for inclusion. Dogs with a history of recent travel (within 1 year) were excluded. One hundred and twenty-six dogs, 57 from a practice in San Diego, and 69 from a practice in Los Angeles, met the criteria. RESULTS: The pattern of seasonality differed with practice location (P = 0.02). More cases of idiopathic IMHA presented during spring and summer (n = 39) than winter and fall (n = 18) for the San Diego practice (P = 0.008) but not the Los Angeles practice (n = 32 and 37, respectively, P = 0.63). CONCLUSIONS: The incidence of idiopathic IMHA in dogs presenting to specialty hospitals located in 2 different climates of southern California was different, suggesting environmental triggers may be involved. Larger, prospective studies are needed to determine whether environmental parameters or undetected infectious disease account for some cases of idiopathic IMHA in dogs.

Distributive veterinary clinical education: a model of clinical-site selection.

The distributive model at the Western University of Health Sciences College of Veterinary Medicine (WesternU-CVM) utilizes third-party clinical sites rather than a traditional on-campus teaching hospital during years 3 and 4 of the curriculum. All veterinary schools are required by the American Veterinary Medical Association's accreditation standards to ensure that students are exposed to a diverse case load of sufficient number with active participation in the diagnostic work-up and treatment of patients. With one centralized teaching hospital, monitoring this aspect of the student experience is relatively straightforward. The distributive model of clinical veterinary education poses several challenges not encountered in a teaching hospital due to the number of clinical sites involved in delivering the curriculum. This article describes a clinical-site and preceptor selection process and the guidelines currently used to evaluate whether clinical sites and preceptors are suitable for initial inclusion in the program at WesternU-CVM. Outcomes data regarding the number and variety of student case exposures, student involvement in case management, and student evaluations of clinical experience are presented. These data suggest that the recruitment and selection process described here results in diverse and ample case-load exposure opportunities in a distributive model of veterinary education.

Prothrombotic mechanisms and anticoagulant therapy in dogs with immune-mediated hemolytic anemia.

OBJECTIVE: To review the pathophysiology of thrombosis in hemolytic disease, and the efficacy of thromboprophylaxis in dogs with immune-mediated hemolytic anemia (IMHA). DATA SOURCES: Computerized searches of Pubmed, INDEX VETERINARIUS, and the journal database of the Veterinary Information Network, and a manual search of bibliographies of published manuscripts. HUMAN DATA SYNTHESIS: Experimental data suggest that hemolysis leads to the induction of the potent procoagulant tissue factor on monocytes and endothelial cells and subsequent activation of coagulation. In addition, damaged red cells, activated platelets, and small cell-derived membrane vesicles called microparticles may contribute to coagulation by providing membrane surfaces containing exposed anionic phospholipids that serve as docking sites for prothrombinase (factor Va-factor Xa) and tenase (factor VIIIa-factor IXa) complexes of the coagulation cascade. Some microparticles also contain tissue factor, further fueling coagulation. Thromboprophylaxis for hemolytic disease in people primarily targets the coagulation cascade rather than platelets, as most thromboemboli are of venous rather than arterial origin. The use of unfractionated heparin is closely monitored to ensure therapeutic levels are reached. VETERINARY DATA SYNTHESIS: Thromboembolic disease is a major factor affecting survival in dogs with IMHA. It is likely that hemolysis contributes to the prothrombotic state. Thrombosis occurs in both veins and arteries, with pulmonary thromboembolism (a venous thrombus) occurring very commonly. Evidence suggests that tissue factor mediates the development of the prothrombotic state. Heparin, and the anti-platelet agents aspirin, and clopidogrel have been used for thromboprophylaxis in dogs with IMHA. However, a lack of validated therapeutic endpoints and controlled studies make it difficult to determine if survival is affected or if 1 drug is more effective than another. CONCLUSIONS: Prospective clinical trials comparing individually adjusted heparin or other anti-coagulant drugs to anti-platelet drugs are needed to make evidence-based recommendations for thromboprophylaxis in dogs with IMHA.

Re-emergence of Babesia conradae and effective treatment of infected dogs with atovaquone and azithromycin.

Babesia conradae (B. conradae) causes hemolytic anemia in dogs. This organism has not been reported clinically since it was originally described in southern California in 1991. To date, no anti-protozoal therapies have been associated with clearance of B. conradae. This report describes the use of atovaquone and azithromycin for the treatment of dogs naturally infected with B. conradae and report the re-emergence of B. conradae in southern California. Twelve dogs naturally infected with B. conradae were identified by practicing veterinarians and public health officials in southern California. Treatments consisted of a 10 day course of atovaquone (13.3mg/kg PO q 8h) and azithromycin (10-12.5mg/kg PO q 24h). Four dogs were treated in a randomized blinded placebo-controlled fashion, four additional cases were treated in a non-random, non-blinded fashion and one dog received no treatment. All dogs were tested for B. conradae DNA by polymerase chain reaction (PCR) initially and then once or 3 times post treatment (60-210 days). B. conradae infected dogs that received treatment did not have any detectable Babesia DNA by PCR after treatment. In contrast, dogs receiving placebo had detectable Babesia DNA by PCR throughout the study period. Combination therapy with atovaquone and azithromycin appears to be effective for acute and chronic babesiosis caused by B. conradae.

Clinical presentation of 26 anaplasma phagocytophilum-seropositive dogs residing in an endemic area.

Anaplasma (A.) phagocytophilum, the etiological agent of canine granulocytic anaplasmosis, is capable of inciting moderate to severe clinical disease in a variety of mammals and is endemic in the upper midwest. The purpose of this study was fourfold: to describe the range of clinical signs in dogs seropositive to A. phagocytophilum; to examine the prevalence of immune-mediated hemolytic anemia (IMHA) in this population; to evaluate whether specific clinical signs were associated with coexposure to Borrelia (B.) burgdorferi in actively infected dogs; and to determine whether clinical response to doxycycline was complete in treated dogs. Medical records of dogs seropositive for A. phagocytophilum were reviewed retrospectively. Peripheral blood smears were also reviewed retrospectively for granulocytic Anaplasma morulae. Lethargy (81%), inappetence (58%), and lameness (50%) were the most common clinical signs, followed by fever (46%). Thrombocytopenia was the most common laboratory abnormality, and IMHA was diagnosed in three dogs. Dogs that were thrombocytopenic and had antibodies to both A. phagocytophilum and B. burgdorferi had a median platelet count of 51,000/µL (range 20,000 to 171,000/µL), which was significantly lower than the count in dogs with antibodies only to A. phagocytophilum (P=0.04). Some dogs had an apparent relapse of clinical signs after an appropriate course of doxycycline. Testing for A. phagocytophilum by polymerase chain reaction, serum antibody assays, and/or blood smear evaluation should be considered in dogs with IMHA, cough, or epistaxis and that reside in A. phagocytophilum-endemic areas. If moderate to severe thrombocytopenia is present, testing for concurrent B. burgdorferi infection may be warranted.

Lack of evidence for perinatal transmission of canine granulocytic anaplasmosis from a bitch to her offspring.

Granulocytic anaplasmosis is an emerging infectious disease affecting dogs and humans in the United States and other regions of the world. Relatively few cases have been described in pregnant women, and perinatal transmission appears to occur infrequently in humans. Infection in pregnant dogs has not been reported. Diagnosis of infection during pregnancy poses therapeutic challenges, because doxycycline, the treatment of choice, is teratogenic. Also, infection during pregnancy may result in more severe disease. When infection is diagnosed after parturition, knowledge of the risk of perinatal transmission to offspring is important, because prophylactic therapy in neonates is also not without risk. In this report, we describe relatively severe clinical manifestations of Anaplasma phagocytophilum infection in a postpartum bitch and a lack of perinatal transmission to her puppies.

Insulin activation of the phosphatidylinositol 3-kinase/protein kinase B (Akt) pathway reduces lipopolysaccharide-induced inflammation in mice.

Insulin is used to control pro-inflammatory hyperglycemia in critically ill patients. However, recent studies suggest that insulin-induced hypoglycemia may negate its beneficial effects in these patients. It is noteworthy that recent evidence indicates that insulin has anti-inflammatory effects that are independent of controlling hyperglycemia. To date, the mechanism by which insulin directly reduces inflammation has not been elucidated. It is well established that insulin activates phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling in many cell types. We and others have shown that this pathway negatively regulates LPS-induced signaling and pro-inflammatory cytokine production in monocytic cells. We hypothesized that insulin inhibits inflammation during endotoxemia by activation of the PI3K/Akt pathway. We used a nonhyperglycemic mouse model of endotoxemia to determine the effect of continuous administration of a low dose of human insulin on inflammation and survival. It is noteworthy that insulin treatment induced phosphorylation of Akt in muscle and adipose tissues but did not exacerbate lipopolysaccharide (LPS)-induced hypoglycemia. Insulin decreased plasma levels of interleukin-6, tumor necrosis factor-alpha, monocyte chemotactic protein 1 (MCP1)/JE, and keratinocyte chemoattractant, and decreased mortality. The PI3K inhibitor wortmannin abolished the insulin-mediated activation of Akt and the reduction of chemokine and interleukin-6 levels. We conclude that insulin reduces LPS-induced inflammation in mice in a PI3K/Akt-dependent manner without affecting blood glucose levels.

Molecular characterization of Rickettsia rickettsii infecting dogs and people in North Carolina.

Rocky Mountain spotted fever (RMST) is an important cause of morbidity and mortality in people and dogs in the United States. Disease manifestations are strikingly similar in both species, and illness in dogs can precede illness in people. R. rickettsii has been identified as a Select Agent by the CDC as a Category C priority pathogen by the National Institute of Allergic and Infectious Diseases because it is amenable to use as a bioterror agent. The clinical and temporal relationship of naturally occurring diseases in dogs and people suggests that dogs could serve as sentinels for natural infection and bioterrorist attacks using this organism. Recognizing genetic modifications in naturally occurring disease agents in order to distinguish them from intentionally released agents are priorities put forth by the NIAID. To determine whether the rickettsiae naturally infecting dogs is the same as those that infect persons in a given geographical region, we characterized rickettsial isolates obtained from three dogs and two persons diagnosed with RMSF in North Carolina. Portions of three genes (ompA, rrs, and gltA) amplified by PCR were cloned and sequenced or directly sequenced. Reactions were run in duplicate in forward and reverse directions. Gene sequences were aligned with known sequences deposited in GenBank and with each other. Sequences of the 5' region of the ompA gene were 100% homologous with a tick strain (Bitterroot) of R. rickettsii for all five isolates. Sequences of the rrs gene were 99.8 99.9% homologous with a tick strain (Sawtooth) of R. rickettsii. rrs gene sequences from one dog and the two persons was identical. Sequences of one dog isolate differed from these by one base pair. Sequences from another dog isolate differed by two base pairs. Sequences of the gltA gene are pending. This confirms on a molecular level that R. rickettsii causing naturally occurring RMSF in dogs in North Carolina is highly homologous to R. rickettsii that causes the disease in people in the same region. Sequence data will be deposited in GenBank, thereby providing genetic information regarding naturally occurring R. rickettsii.

Morphometric assessment of the proximal portion of the tibia in dogs with and without cranial cruciate ligament rupture.

Based on the clinical observation that dogs with a steep tibial plateau slope had variable tibial morphology, we hypothesized that these dogs could be further characterized using measurements developed by examining computer generated models of specific proximal tibial malformations. A 3D tibial model was created from a normal canine tibia. The model was manipulated to reproduce two specific proximal tibial anomalies representing deformities originating from the tibial plateau or the proximal tibial shaft. Data from these models were used to create specific measurements that would characterize the shape of these anomalies. These measurements included the diaphyseal tibial axis (DTA)/proximal tibial axis (PTA) angle, which defined the orientation of the proximal portion of the shaft in relation to the tibial mid-shaft. These measurements were then made on radiographs of dogs with and without cranial cruciate ligament (CCL) rupture. Models with tibial plateau and proximal shaft deformities had a steep tibial plateau slope (TPS). Models with proximal shaft deformity had a markedly increased DTA/PTA angle. The model with a 10 degree proximal shaft deformity had a DTA/PTA angle of 11.23 degrees. Six dogs (9.0%) had a DTA/PTA angle larger than 11.23 degrees (range, 11.4-13.9 degrees). Dogs in this group had ruptured CCL and a steep TPS. Dogs with CCL rupture had higher TPS (mean, 31.8 +/- 4.1 degrees) and DTA/PTA angle (mean, 6.0 +/- 3.3 degrees) than dogs without CCL rupture (means, 23.6 +/- 3.4 degrees and 4.1 +/- 2.2 degrees, respectively). Dogs with proximal shaft deformity represented a distinct group, which could not be identified using the magnitude of the TPS alone. Characterizing more precisely the shape of the proximal portion of the tibia in dogs contributes to our understanding of the pathogenesis of steep TPS and may facilitate the optimization of the surgical management of dogs with CCL rupture.

Febrile illness associated with Rickettsia conorii infection in dogs from Sicily.

We report serologic and molecular evidence of acute, febrile illness associated with Rickettsia conorii in 3 male Yorkshire terriers from Sicily (Italy).