High‐mobility group box 1 as a surrogate prognostic marker in dogs with systemic inflammatory response syndrome

Objective – To evaluate various surrogate markers associated with the inflammatory and counter‐inflammatory responses with respect to mortality in dogs with systemic inflammatory response syndrome (SIRS). Design – Prospective observational study. Setting – Veterinary Teaching Hospital. Animals – Twenty‐eight dogs with naturally occurring diseases and SIRS from January 2007 to May 2009. Interventions – Upon admission to the veterinary hospital, history and baseline data from the physical examination, including parameters previously defined for meeting SIRS criteria, were documented. Heparinized blood samples were collected and plasma cytokines interleukin‐6 (IL‐6), IL‐10, and high‐mobility group box 1 (HMGB1) were measured by sandwich ELISA. Measurements and Main Results – In nonsurvivors, median plasma HMGB1 concentrations (0.718 μg/L, interquartile range [IQR]; 0.300–1.626 μg/L) and the ratio of HMGB1 to IL‐10 (2.236, IQR; 0.972–5.367) were significantly increased as compared with those found in survivors (0.300 μg/L, IQR; 0.300–0.312 μg/L for HMGB1; 1.017, IQR; 0.862–1.126 for the ratio of HMGB1 to IL‐10, P=0.007 and 0.024, respectively). Plasma IL‐6, IL‐10, and the ratio of IL‐6 to IL‐10 were not significantly different between groups. Among the parameters studied, HMGB1 and the ratio of HMGB1 to IL‐10 performed the best in discriminating outcome in dogs with SIRS according to receiver operator characteristic curve analysis. Conclusions – Increases in plasma HMGB1 concentration and the ratio of HMGB1 to IL‐10 may predict poorer outcomes in dogs with SIRS. The approach described may lead to reliable prognostic biomarkers and new therapeutic concepts in the study of SIRS in dogs.     

Thrombelastography in dogs admitted to an intensive care unit

Background: Underlying conditions in dogs admitted to an intensive care unit (ICU) can cause hemostatic dysfunction. Thrombelastography (TEG) may be useful in detecting hemostatic alterations as compared with standard coagulation tests. Objectives: The purpose of this study was to compare TEG results and those of standard coagulation tests in identifying hemostatic dysfunction in dogs admitted to an ICU and to investigate associations among the variables measured. Methods: Tissue factor‐activated TEG analysis, d ‐dimer and fibrinogen concentrations, antithrombin (AT) activity, prothrombin time (PT), activated partial thromboplastin time (aPTT), and platelet count were measured using standard techniques on 27 dogs admitted to ICU with a disease known to be associated with hemostatic dysfunction and in 31 clinically healthy control dogs. Results were compared between groups using nonparametric tests and κ analysis; principal component analysis (PCA) and Spearman rank correlation were used to measure associations among variables. Results: Fourteen of 27 ICU dogs had abnormal TEG tracings, which were used to classify the dogs as hypercoagulable (n=11), hypocoagulable (n=3), or normocoagulable (n=13). Hypercoagulable dogs had significantly increased d ‐dimer (P=.03) and fibrinogen (P=.01) concentrations compared with normocoagulable dogs. In ICU dogs, positive associations were identified between maximum amplitude (MA), α‐angle, fibrinogen concentration, and platelet count, and between PT, aPTT, and reaction time (R). Significant correlations were found between MA and fibrinogen (r_s=.76, P<.001) and between reaction time (R) and PT (r_s=.51, P=.003). Conclusions: TEG was useful in detecting hemostatic dysfunction in dogs in an ICU. Positive associations among variables may provide insight as to how overall coagulation status reflects alterations in clot strength and coagulation time. Dogs with TEG tracings indicative of hypercoagulability are likely in procoagulant states. Future studies of the incidence of thrombotic complications in dogs with hypercoagulable TEG tracings are warranted.     

The role of Acinetobacter baumannii as a nosocomial pathogen for dogs and cats in an intensive care unit

Acinetobacter baumannii is a nosocomial pathogen associated with high morbidity and mortality in humans. Whereas infections with strains of Acinetobacter species have been reported in various situations, the importance of A baumannii as a nosocomial pathogen in veterinary hospitals has not been studied so far. In this retrospective case series, we describe 17 dogs and 2 cats from which A baumannii had been isolated during a 21/2-year period. In 7 dogs, A baumannii induced systemic signs of illness, whereas 12 animals showed signs of local infection. In all animals with systemic infection, and in 2 with localized infection, A baumannii contributed to the death of the animal or contributed to euthanasia; the remaining 8 dogs and both cats recovered. Molecular typing of the isolates with restriction polymorphisms of ribosomal DNA provided evidence of nosocomial spread of this pathogen and for the presence of several strains of A baumannii in the hospital environment.     

Immunocytochemical study of Ki-67 as a prognostic marker in canine mammary neoplasia

Background: Canine mammary tumors are challenging for clinicians and pathologists because of complex histologic classification, low specificity of cytologic diagnosis, and unpredictable biological behavior. In histologic specimens, expression of tumor proliferation marker Ki‐67, a nuclear nonhistone protein, has been shown to have prognostic value for canine mammary tumors and to correlate with malignancy and low survival rates. Objective: The objective of this study was to measure the proliferation index of canine mammary tumors by immunochemical detection of Ki‐67 in cytologic specimens and to determine its relationship to clinical and pathologic variables and patient outcome. Methods: Spontaneous mammary tumors from 31 female dogs were surgically excised. Imprint specimens for cytologic evaluation were wet‐fixed in ethanol; histologic specimens were prepared routinely. Immunostaining was performed with the PH 177 monoclonal antibody against Ki‐67; proliferation index was graded from negative to +++. Dogs were followed for 18 months. Multivariate logistic regression analysis was used to determine correlations between immunocytochemical results, tumor and clinical variables, and patient outcome. Results: Ki‐67 proliferation indices in cytologic specimens were significantly lower for nonmalignant tumors than for malignant tumors. High index values of Ki‐67 were positively correlated with metastasis, death from neoplasia, low disease‐free survival rates, and low overall survival rate. With the exception of 4 specimens for which cellularity was insufficient, positive expression of Ki‐67 in cytologic specimens correlated with that of histologic specimens. Conclusions: The prognostic value of the Ki‐67 index in canine mammary tumors by using wet‐fixed cytology imprint specimens was similar to that observed previously for histologic specimens. Immunocytochemical detection of Ki‐67 could improve the accuracy and value of cytology by providing safe and rapid information about malignancy and patient outcome.     

Evaluation of cystatin C as an endogenous marker of glomerular filtration rate in dogs

Cystatin C is a cysteine protease inhibitor produced by all nucleated cells. It is freely filtered by the glomerulus and is unaffected by nonrenal factors such as inflammation and gender. Because of greater sensitivity and specificity, cystatin C has been proposed to replace creatinine as a marker of glomerular filtration rate (GFR) in humans. The aims of this study were to validate an automated assay in canine plasma and to evaluate the usefulness of cystatin C as a marker of GFR in dogs. Western blotting was used to demonstrate cross-reactivity of an anti-human cystatin C antibody. An immunoturbidimetric assay was used to detect cystatin C in 25 clinically healthy dogs and 25 dogs with renal failure. Mean cystatin C concentration in the healthy dogs and the dogs with renal failure was 1.08 +/- 0.16 mg/L and 4.37 +/- 1.79 mg/L respectively. Intra- and interassay variability was <5%. The assay was linear (r = .974) between 0.14 and 7.53 mg/L. Both cystatin C and creatinine concentrations were measured in banked, frozen serum from 20 remnant kidney model dogs and 10 volume-depleted dogs for which GFR measurements by exogenous creatinine clearance had been determined previously. In the remnant kidney model, cystatin C was better correlated with GFR than creatinine (r = .79 versus .54) but was less well correlated with GFR in volume-depleted dogs (r = .54 versus .95). GFR measurements were repeated in the remnant kidney model dogs 60 days after initial GFR measurements. At this time, cystatin C and creatinine concentrations correlated equally well with GFR (r = .891 versus .894, respectively). Cystatin C concentration is a reasonable alternative to creatinine for screening dogs with decreased GFR due to chronic renal failure.     

Diagnosis of disseminated intravascular coagulation in dogs admitted to an intensive care unit.

OBJECTIVE: To describe and evaluate hemostatic function in critically ill dogs with clinical signs of diseases that predispose to disseminated intravascular coagulation (DIC). DESIGN: Prospective case series. ANIMALS: 59 critically ill dogs (affected dogs) with clinical signs of diseases known to predispose to DIC and 52 clinically normal dogs (control dogs). PROCEDURE: Activated partial thromboplastin time (aPTT), prothrombin time (PT), thrombin clotting time (TCT), plasma fibrinogen concentration, serum concentration of fibrin and fibrinogen-related antigens (FRA), and plasma antithrombin III (AT III) activity were determined for all dogs. Results from affected dogs were compared with those of control dogs. In some affected dogs, postmortem tissue specimens were examined for evidence of microvascular thrombosis. A diagnosis of DIC was made by fulfilling at least 3 of the following criteria: 1) abnormal aPTT, PT, or TCT value, 2) low plasma fibrinogen concentration, 3) low plasma AT III activity, 4) high serum FRA concentration, or 5) low platelet count. To evaluate the severity of hemostatic dysfunction, 3 arbitrary categories (mild, moderate, and severe) were proposed. RESULTS: A diagnostic strategy based on moderate hemostatic dysfunction identified DIC in 16 of 59 (27.1%) affected dogs. The AT III activity was < 70% in 15 of 16 dogs with DIC. Microvascular thrombosis was observed in tissue specimens from 7 of 8 affected dogs. Serum FRA and plasma fibrinogen concentrations did not contribute in establishing a diagnosis of DIC. CONCLUSIONS AND CLINICAL RELEVANCE: A diagnosis of DIC can be made when hemostatic dysfunction is moderate in dogs with clinical signs of diseases associated with DIC.     

Surveillance of infections associated with intravenous catheters in dogs and cats in an intensive care unit

Positive catheter-tip culture rates and risk factors associated with bacterial colonization of intravenous (i.v.) catheters were assessed in dogs and cats. Aerobic and anaerobic bacterial cultures were performed on 151 catheters, and 24.5% were positive. Of the positive cultures, 46.0% grew Enterobacter spp. The type of catheter used, blood sampling through the catheter, the type of i.v. infusate administered, the duration the catheter was in place, the catheter location, complications with the catheter, and the final outcome of the animal were not associated with an increased risk of a positive bacterial culture from the catheter tip.     

Incidence of catheter-associated urinary tract infection among dogs in a small animal intensive care unit

OBJECTIVE: To determine incidence of and possible risk factors for catheter-associated urinary tract infection (UTI) among dogs hospitalized in an intensive care unit and compare results of bacterial culture of urine samples with results of bacterial culture of catheter tips. DESIGN: Prospective study. ANIMALS: 39 dogs. PROCEDURE: A standard protocol for aseptic catheter placement and maintenance was used. Urine samples were obtained daily and submitted for bacterial culture. When possible, the urinary catheter tip was collected aseptically at the time of catheter removal and submitted for bacterial culture. Bacteria that were obtained were identified and tested for antimicrobial susceptibility. RESULTS: 4 of the 39 (10.3%) dogs developed a UTI. The probability of remaining free from UTI after 1 day in the intensive care unit was 94.9%, and the probability of remaining free from UTI after 4 days was 63.3%. Bacteria isolates were generally common urinary tract pathogens and were susceptible to most antimicrobials. Specific risk factors for catheter-associated UTI, beyond a lack of antimicrobial administration, were not identified. Positive predictive value of bacterial culture of urinary catheter tips was only 25%. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that placement of an indwelling urinary catheter in dogs is associated with a low risk of catheter-associated UTI during the first 3 days after catheter placement, provided that adequate precautions are taken for aseptic catheter placement and maintenance. Results of bacterial culture of urinary catheter tips should not be used to predict whether dogs developed catheter-associated UTI.     

Comparison of fentanyl and hydromorphone constant rate infusions for pain management in dogs in an intensive care unit

Objective

To compare the efficacy and quality of analgesia provided by constant rate infusions (CRIs) of hydromorphone and fentanyl in dogs in the intensive care unit (ICU).

Prevalence and risk factors for Clostridium difficile colonization in dogs and cats hospitalized in an intensive care unit

Clostridium difficile is the most common cause of hospital- and antimicrobial-associated diarrhea in hospitalized humans however the role of C. difficile in diarrhea in dogs has not been defined. A prospective study of C. difficile colonization in dogs and cats was conducted in a veterinary teaching hospital intensive care unit (ICU). Rectal swabs were taken from patients upon admission to the ICU and every third day of hospitalization until discharge or death. C. difficile was isolated from 73/402 (18%) animals; 69% of isolates were toxigenic. Community-associated colonization was identified in 39/366 (11%) of animals that were sampled at the time of admission, while C. difficile was subsequently isolated from 27 of the remaining 327 (8.3%) animals that had a negative admission swab. C. difficile was isolated from seven other dogs during hospitalization, but the origin was unclear because the admission swab was not collected. Administration of antimicrobials prior to admission and administration of immunosuppressive drugs during hospitalization were risk factors for hospital-associated colonization (P=0.006, OR 4.05, 95% CI 1.4-10.8). Acquisition of C. difficile during hospitalization in the ICU was associated with the development of diarrhea (P=0.004). Two ribotypes, one toxigenic and one non-toxigenic, predominated.     

Development of antimicrobial drug resistance in rectal Escherichia coli isolates from dogs hospitalized in an intensive care unit

OBJECTIVE: To determine whether duration of hospitalization in the intensive care unit (ICU) of a veterinary teaching hospital was associated with prevalence of antimicrobial resistance among rectal Escherichia coli isolates from dogs, whether antimicrobial treatment was associated with prevalence of antimicrobial resistance, and whether there were associations among antimicrobial drugs to which isolates were resistant. DESIGN: Prospective observational study. ANIMALS: 116 dogs hospitalized in an ICU for >or= 3 days. PROCEDURES: Rectal swab specimens were obtained every 3 days and submitted for bacterial culture for E coli. Isolates were tested for susceptibility to 12 antimicrobial agents by means of disk diffusion. RESULTS: For each additional day that a dog was hospitalized in the ICU, the odds of being colonized with an E coli isolate resistant to 1 or more of the 12 antimicrobials tested increased by a factor of 1.5, independent of antimicrobial treatment. Dogs that were treated with enrofloxacin were 25.6 times as likely to be colonized by a quinolone-resistant E coli strain as were dogs that did not receive any antimicrobials. Significant correlations were found for resistance to agents in the extended-spectrum cephalosporin group and the quinolone group. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that the proportion of rectal E coli isolates obtained from dogs housed for >or= 3 days in a veterinary teaching hospital ICU that were resistant to antimicrobial agents increased as the duration of hospitalization in the ICU increased. Thus, ICU hospitalization time should be as short as possible to prevent development of antimicrobial resistance among rectal E coli isolates.     

Evaluation of point-of-care tests for diagnosis of disseminated intravascular coagulation in dogs admitted to an intensive care unit.

OBJECTIVE: To evaluate the accuracy of point-of-care tests for the diagnosis of disseminated intravascular coagulation (DIC) in dogs and assess the correlation and agreement of results between point-of-care and laboratory tests in the evaluation of hemostatic function. DESIGN: Prospective case series. ANIMALS: 59 critically ill dogs (affected dogs) with clinical signs of diseases known to predispose to DIC and 52 clinically normal dogs. PROCEDURES: Accuracy of the point-of-care tests (activated clotting time [ACT], estimated platelet count and number of schizocytes from a blood smear, plasma total solids [TS] concentration, and the protamine sulfate test) was evaluated, using receiver operating characteristic curves and likelihood ratios. A strategy, using likelihood ratios to calculate a posttest probability of DIC, was tested with 65% used as a threshold for initiation of treatment. Results of laboratory tests (coagulogram and plasma antithrombin III activity) were used as the standard for comparison in each dog. RESULTS: ACT and estimated platelet count provided the best accuracy for detection of DIC. The plasma TS concentration, schizocyte number, and protamine sulfate test had poor accuracy. The strategy using post-test probability of DIC identified 12 of 16 affected dogs that had DIC. Estimated platelet count was correlated and had acceptable clinical agreement with automated platelet count (r = 0.70). The plasma TS (r = 0.28) concentration and serum albumin (r = 0.63) concentration were not accurate predictors of plasma antithrombin III activity. The ACT did not correlate with activated partial thromboplastin time (r = 0.28). CONCLUSIONS AND CLINICAL RELEVANCE: Strategic use of likelihood ratios from point-of-care tests can assist clinicians in making treatment decisions for dogs suspected to have DIC when immediate laboratory support is unavailable.     

Post‐natal Changes in Testicular Concentrations of Interleukin‐1 Alpha and Beta and Interleukin‐6 during Sexual Maturation in Bulls

Based on observations in laboratory animals interleukins could be regulators of testicular development. The objects of this study were to see if interleukins (IL‐1 and IL‐6) are present in the developing bull testis and to establish the temporal patterns of concentrations of IL‐1 and IL‐6 in the bovine testis during development. Separate groups of six bull calves were castrated every 4 weeks from 5 to 33 weeks of age, and at 56 weeks of age. Mean testicular IL‐1 alpha concentrations decreased (p < 0.01) from 5 to 9 weeks of age and 13 to 21 weeks of age. Mean testicular IL‐1 beta concentrations decreased (p < 0.01) from 13 to 17 weeks of age and from 29 to 33 weeks of age. Mean IL‐1 bioactivity increased from 13 to 17 weeks of age, decreased to 21 weeks, increased to 25 weeks, decreased to 29 weeks and decreased from 33 to 56 weeks of age (p < 0.05). Mean testicular IL‐6 concentrations decreased (p < 0.05) from 9 to 13 weeks of age, increased (p < 0.05) to 21 weeks, decreased (p < 0.05) to 25 weeks, increased (p < 0.05) to 29 weeks and decreased (p < 0.01) to 56 weeks of age. In conclusion, testicular IL‐1 alpha, IL‐1 beta and IL‐6 were found in the bovine testis and concentrations were age dependent. Testicular IL‐1 alpha and IL‐1 beta concentrations were highest in the early post‐natal period; however, IL‐1 bioactivity and IL‐6 concentrations were greatest in the immediate pre‐pubertal period. These findings suggest a functional role for interleukins in testicular development in the bull.     

Cosegregation of a factor VIII microsatellite marker with mild hemophilia A in Golden Retriever dogs

Mild hemophilia A (factor VIII deficiency) was diagnosed in Golden Retrievers and pedigree studies were undertaken to test the cosegregation of an intragenic factor VIII marker with the disease phenotype. The study population consisted of 30 client-owned dogs (22 males and 8 females). Hemophilic males (n = 12) typically demonstrated prolonged bleeding after trauma or surgery rather than spontaneous hemorrhagic events. The affected males had a proportionate reduction in factor VIII coagulant activity (mean FVIII:C = 4%) and factor VIII protein concentration (mean FVIII:Ag = 3%). Twenty-five dogs (10 affected males, 8 clear males, 2 obligate carrier dams, and 5 suspect carrier daughters) were genotyped for a factor VIII microsatellite marker, with allele size assigned by an automated capillary electrophoresis system. Five distinct marker alleles were present in the study pedigree and a 300-base pair allele was found to segregate with the hemophilia A phenotype. The inheritance of the hemophilia-associated allele defined carrier status for 5 suspect daughters of obligate carrier dams. The limitations inherent to linkage analyses (i.e., lack of access to key family members and homozygosity at the marker locus) did not preclude carrier detection in this pedigree. We conclude that genotype analysis for the intragenic factor VIII marker can aid in control of canine hemophilia A through enhanced carrier detection.