Microscopic detection of Leptospira bacterial organisms in urine sediment from a young dog with leptospirosis and a review of the pathobiology and diagnosis of canine leptospirosis.

Samples collected from an 11-month-old Dachshund-mix dog with a history of acute azotemia, fever, and enlarged and irregular kidneys were received at the Colorado State Veterinary Diagnostic Laboratory (CSU VDL). The submitting veterinarians were concerned about lymphoma versus acute nephritis/pyelonephritis. The CSU clinical pathology laboratory received urine for urinalysis and kidney aspirates for cytologic evaluation. Urine had also been submitted for aerobic culture and Leptospirosis PCR, and serum was submitted for Lepto-5 microscopic agglutination testing (MAT). Upon examination of a wet mount of the urine sediment, technical staff noted "vibrating" clumps of granular-appearing material throughout the slide, which prompted the preparation of a stained sediment slide for pathologist review. Very small, faintly staining organisms were observed, and an attempt was made to picture-match these with published reports of Leptospira in dog urine, but none could be found. In addition, some references claimed that Leptospira organisms are not seen in urine with light microscopy. The suspicion that these organisms were Leptospira sp. was supported by the MAT results and later confirmed by PCR. The organisms subsequently exhibited strong positive immunolabeling for the Leptospira antigen. This case report provides a searchable record of Leptospira organisms visualized by routine light microscopy in dog urine during natural infection and a review of canine leptospirosis pathobiology and diagnosis.

Analysis of Canine Peritoneal Fluid Analysis.

Canine peritoneal fluid analysis results were retrospectively reviewed to assess the appropriateness of different classification schemes. Cutoffs of 3000 cells/µL and 2.5 g/dL protein are recommended. Analyzing the total nucleated cell count and total protein concentration is only the first step in peritoneal fluid analysis; microscopic examination, clinical presentation, and other laboratory data are all important in determining the final classification of peritoneal fluid analysis, keeping in mind that the most important aspect of fluid analysis is not what something is called, but whether it helps achieve a diagnosis. Discussion of effusion mechanisms, study observations, and recommended diagnostic steps after fluid analysis are included.

A Quick Reference on Chloride.

Chloride is an essential element, playing important roles in digestion, muscular activity, regulation of body fluids, and acid-base balance. As the most abundant anion in extracellular fluid, chloride plays a major role in maintaining electroneutrality. Chloride is intrinsically linked to sodium in maintaining osmolality and fluid balance and has an inverse relationship with bicarbonate in maintaining acid-base balance. It is likely because of these close ties that chloride does not get the individual attention it deserves; we can use these facts to simplify and interpret changes in serum chloride concentrations.

Diagnosis of Disorders of Iron Metabolism in Dogs and Cats.

Iron is an essential element and is used by every cell in the body. This article summarizes iron metabolism and disorders associated with iron metabolism in dogs and cats. The diagnostic tests currently in use for assessing iron status are discussed.

Hematology of camelids.

Interpretation of camelid hematology results is similar to that of other mammals. Obtaining accurate results and using appropriate reference intervals can be a bit problematic, particularly when evaluating the erythron. Camelid erythrocytes vary from other mammals in that they are small, flat, and elliptical. This variation makes data obtained from samples collected from these species prone to error when using some automated instruments. Normal and abnormal findings in camelid blood are reviewed as well as how to ensure accurate results.

A comparison of peritoneal fluid values in mares following bilateral laparoscopic ovariectomy using a vessel sealing and dividing device versus placement of two ligating loops.

The objective of this study was to determine the effect of bilateral laparoscopic ovariectomy on peritoneal fluid values in mares and compare how this effect was modified by the method of ovarian vessel hemostasis used. Ten mares undergoing standing bilateral laparoscopic ovariectomy were used in a randomized clinical study. During surgery, blood vessels within the mesovarium were either: (1) sealed and transected with a vessel sealing and dividing device (VSDD), or (2) ligated using two loops placed proximal to each ovary and then the mesovarium transected using laparoscopic scissors. The ovaries were removed through the ipsilateral body wall. Abdominocentesis was performed before surgery and 24 h and 72 h after surgery. Markers of peritoneal inflammation, as measured by total nucleated cell count, total protein (TP) and red blood cell count via abdominocentesis, were consistently increased for all groups compared to pre-operative values. The mean (range) of TP for the VSDD group was 4.14 (3.9-4.5) g/dL, and that for the ligating loop group was 3.18 (2.7-3.5) g/dL. Use of the VSDD resulted in significantly greater TP concentrations in the abdominal fluid at 24 h and 72 h post-operatively when compared to a ligating loop (P <0.001 and 0.04, respectively).

Comparison of equine endometrial cytology samples collected with uterine swab, uterine brush, and low-volume lavage from healthy mares.

BACKGROUND: There have been no studies evaluating and comparing the quality of equine endometrial cytology samples obtained via the 3 most common sampling techniques from healthy mares. OBJECTIVES: The objective was to compare the quality and contents of equine endometrial samples obtained by 3 different sampling techniques: double-guarded uterine swab, double-guarded uterine brush, and low-volume lavage (LVL), all collected from clinically healthy mares. METHODS: Samples were collected from 24 healthy mares in early estrus. In 19 mares, samples were obtained in a sequential manner, first with the swab, then with the brush, followed by LVL. Cytologic evaluation included estimates of quality, cellularity, and presence of inflammatory cells. The clinical pathologist performing the evaluations was blinded to the collection technique. The Friedman test with Dunn's multiple comparisons was used to compare rankings of quality, cellularity, and the presence or absence of inflammatory cells. Observed cytologic differences were described. RESULTS: All techniques provided diagnostic samples, but swabs yielded the lowest quality sample. In our hands, the uterine brush provided the highest quality sample. Low-volume lavage samples contained higher numbers of neutrophils, although, in general, < 1 neutrophil/400× field is expected for all endometrial sampling techniques in healthy mares. CONCLUSIONS: All sampling techniques can be adequate methods for endometrial cytology, but the brush technique consistently provided the best sample. Sample contamination or poor slide quality can adversely affect interpretation. The most accurate criteria for determining what constitutes mild endometritis in mares have yet to be established.

Characterization of the canine urinary proteome.

BACKGROUND: Urine is an attractive biofluid for biomarker discovery as it is easy and minimally invasive to obtain. While numerous studies have focused on the characterization of human urine, much less research has focused on canine urine. OBJECTIVES: The objectives of this study were to characterize the universal canine urinary proteome (both soluble and exosomal), to determine the overlap between the canine proteome and a representative human urinary proteome study, to generate a resource for future canine studies, and to determine the suitability of the dog as a large animal model for human diseases. METHODS: The soluble and exosomal fractions of normal canine urine were characterized using liquid chromatography tandem mass spectrometry (LC-MS/MS). Biological Networks Gene Ontology (BiNGO) software was utilized to assign the canine urinary proteome to respective Gene Ontology categories, such as Cellular Component, Molecular Function, and Biological Process. RESULTS: Over 500 proteins were confidently identified in normal canine urine. Gene Ontology analysis revealed that exosomal proteins were largely derived from an intracellular location, while soluble proteins included both extracellular and membrane proteins. Exosome proteins were assigned to metabolic processes and localization, while soluble proteins were primarily annotated to specific localization processes. Several proteins identified in normal canine urine have previously been identified in human urine where these proteins are related to various extrarenal and renal diseases. CONCLUSIONS: The results of this study illustrate the potential of the dog as an animal model for human disease states and provide the framework for future studies of canine renal diseases.

Disease distribution in canine patients with acanthocytosis: 123 cases.

BACKGROUND: An acanthocyte is an abnormally shaped erythrocyte. In veterinary medicine, acanthocytes have historically been associated with canine hemangiosarcoma. In human medicine, acanthocytes are rarely observed with neoplastic disease and are more commonly associated with a variety of hereditary and acquired diseases. OBJECTIVES: The purpose of the study was to determine what disease processes are associated with the presence of acanthocytes in the peripheral blood of dogs. METHODS: Medical records for dogs presented to the Veterinary Teaching Hospital of Colorado State University during January 2004 through June 2008 with acanthocytes documented in their CBCs were retrospectively reviewed. RESULTS: A total of 123 dogs were included, 66 of which were diagnosed with neoplastic disease, most commonly hemangiosarcoma (n = 12), osteosarcoma (n = 11), and lymphoma (n = 11). The remaining 57 dogs had nonneoplastic disease, most commonly observed were gastrointestinal (n = 13), musculoskeletal (n = 8), renal (n = 8), and immune-mediated diseases (n = 7). No statistically significant difference was detected between percent acanthocytes present in dogs with neoplastic and nonneoplastic diseases. CONCLUSION: Acanthocytosis was observed with a variety of neoplastic and nonneoplastic diseases. While clearly commonly associated, the presence of acanthocytes in a blood smear should not be considered pathognomonic for hemangiosarcoma in dogs.

Diagnosis of disorders of iron metabolism in dogs and cats.

Iron is an essential element and is used by every cell in the body. This article summarizes iron metabolism and disorders associated with iron metabolism in dogs and cats. The diagnostic tests currently in use for assessing iron status are discussed.

Comparison of peritoneal fluid values after laparoscopic cryptorchidectomy using a vessel-sealing device (Ligasure™) versus a ligating loop and removal of the descended testis.

OBJECTIVE: To determine the effect of unilateral laparoscopic cryptorchidectomy and removal of the descended testis on peritoneal fluid values, and to compare effect between 2 methods for cryptorchid testis vessel hemostasis. STUDY DESIGN: Randomized clinical study. ANIMALS: Stallions (n = 10) with unilateral abdominal cryptorchid testis. METHODS: During standing laparoscopic cryptorchidectomy, blood vessels within the mesorchium of the cryptorchid testis were either sealed and transected with the LigaSure Atlas™ or 2 ligating loops were placed proximal to the testis and the tissue transected with laparoscopic scissors. The testis was removed through the body wall. After laparoscopic cryptorchidectomy, stallions were anesthetized and the descended testis was removed using a closed technique leaving the scrotal incision open. Abdominocenteses were performed before surgery, and 24, and 72 hours after surgery. RESULTS: Values for peritoneal total nucleated cell count (TNCC), total protein concentration (TP), and red blood cell count (RBCC) were all elevated at 24 and 72 hours when compared with baseline. Median TNCC for LigaSure™ (59,780 cells/µL) was nearly twice that of the ligating loop (32,880 cells/µL) at 24 hours postoperatively. There was no statistically significant difference in TNCC, TP, or RBCC between groups. CONCLUSIONS: TNCC, TP, and RBCC increase appreciably from baseline 24 hours after laparoscopic cryptorchidectomy and closed castration but are markedly reduced by 72 hours.

Using corrected serum chloride and predicted bicarbonate concentrations to interpret acid-base status in dogs.

BACKGROUND: Changes in water balance and the presence of unmeasured anions perturb the inverse relationship between serum chloride (Cl) and bicarbonate (HCO(3) ) concentrations in people, affecting accurate interpretation of acid-base status. OBJECTIVES: The aim of this study was to demonstrate that corrected serum Cl and predicted HCO(3) concentrations, based on serum sodium (Na) concentration and anion gap (AG), would be inversely correlated and could be used to better characterize causes of acid-base disorders in dogs. METHODS: In this retrospective study, electrolyte data from dogs with at least one abnormality in serum Na, Cl, or HCO(3) concentrations were analyzed. Profiles were classified before and after calculations using 2 methods, a modified Feldman and an institutional method, to correct Cl concentration and predict HCO(3) concentrations based on Na concentration and AG. Dogs were classified as low (L), normal (N), or high (H) based on Cl (first letter) and HCO(3) (second letter) concentrations, as follows: LL, LN, LH, NL, NN, NH, HL, HN, or HH. RESULTS: For profiles from 261 dogs, reclassifying corrected Cl and predicted HCO(3) concentrations resulted in a shift from the initial classification into a different one in 73% of dogs; in most cases, the shift was to LH, NN, or HL categories. Albumin concentration was a significant factor in acid-base balance. CONCLUSIONS: When interpreting acid-base status based on results of a standard biochemical panel, erroneous conclusions can be drawn if concentrations of Na, unmeasured anions, and albumin are not taken into account. The inverse relationship between serum Cl and HCO(3) concentrations may be used to identify frequent acid-base disorders as well as to unmask abnormalities obscured by irregularities in water balance or AG.

Detection of alkaline phosphatase in canine cells previously stained with Wright-Giemsa and its utility in differentiating osteosarcoma from other mesenchymal tumors.

BACKGROUND: Osteosarcoma (OSA) is a common primary bone tumor in dogs. Demonstration of alkaline phosphatase (ALP) reactivity by tumor cells on unstained slides is useful in differentiating osteosarcoma from other types of sarcoma. However, unstained slides are not always available. OBJECTIVES: The objectives of this study were to evaluate the diagnostic utility of detecting ALP expression in differentiating osteosarcoma from other sarcomas in dogs using cytologic material previously stained with Wright-Giemsa stain and to assess the sensitivity and specificity of ALP expression for diagnosing osteosarcoma using a specific protocol. METHODS: Archived aspirates of histologically confirmed sarcomas in dogs that had been previously stained with Wright-Giemsa stain were treated with 5-bromo, 4-chloro, 3-indolyl phosphate/nitroblue tetrazolium (BCIP/NBT) as a substrate for ALP. Cells were evaluated for expression of ALP after incubation with BCIP/NBT for 1 hour. Sensitivity and specificity of ALP expression for diagnosis of OSA were calculated. RESULTS: In samples from 83 dogs, cells from 15/17 OSAs and from 4/66 tumors other than OSA (amelanotic melanoma, gastrointestinal stromal tumor, collision tumor, and anaplastic sarcoma) expressed ALP. Sensitivity and specificity of ALP expression detected using BCIP/NBT substrate applied to cells previously stained with Wright-Giemsa stain for OSA were 88 and 94%, respectively. CONCLUSIONS: ALP expression detected using BCIP/NBT substrate applied to previously stained cells is useful in differentiating canine OSA from other mesenchymal neoplasms.

Influence of endurance exercise on serum concentrations of iron and acute phase proteins in racing sled dogs.

OBJECTIVE: To determine effects of long-distance racing exercise on iron status in endurance racing sled dogs, with or without anemia. DESIGN: Prospective cohort study. ANIMALS: 114 dogs that participated in the 2007 Iditarod Trail Sled Dog Race (59 and 55 dogs that did or did not complete the race, respectively). PROCEDURES: Stored serum samples obtained from 85 endurance-racing sled dogs that were expected to participate in the race were used to establish study reference intervals and prerace group values for iron-related variables. Blood samples collected from 114 study dogs before (ie, baseline) and after participation in the race were used to determine PCV and serum total protein concentrations before and after racing and assess iron-related variables after racing. RESULTS: Mean values for PCV and serum total protein concentration were decreased after racing, compared with baseline values in the same dogs. Mean serum iron concentration was low, and mean serum ceruloplasmin and C-reactive protein (CRP) concentrations were high in dogs after racing, compared with prerace group values. Mean serum ferritin concentration was high in dogs that did not complete the race, compared with the prerace group value and that of dogs that finished the race; 4 of 113 (3.5%) study dogs had low ferritin concentrations (< 73 ng/mL) after racing, suggestive of possible iron deficiency. CONCLUSIONS AND CLINICAL RELEVANCE: Decreased PCV and serum total protein concentrations were consistently detected, whereas iron deficiency appeared to be uncommon, in study dogs after race participation. High serum concentrations of ceruloplasmin and CRP after racing suggested that changes indicative of iron deficiency may be masked by inflammation. Alternatively, changes in serum iron and CRP concentrations may reflect a physiologic response.

Aryl hydrocarbon receptor targets pathways extrinsic to bone marrow cells to enhance neutrophil recruitment during influenza virus infection.

There is growing evidence that neutrophils influence host resistance during influenza virus infection; however, factors that regulate neutrophil migration to the lung during viral infection are unclear. Activation of the aryl hydrocarbon receptor (AhR) by the pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin) results in an increased number of neutrophils in the lung after influenza virus infection. The mechanism of AhR-mediated neutrophilia does not involve elevated levels of soluble neutrophil chemoattractants, upregulated adhesion molecules on pulmonary neutrophils, delayed neutrophil apoptosis, or increased vascular damage. In this study, we determined whether AhR activation increases neutrophil numbers systemically or only in the infected lung, and whether AhR-regulated events within the hematopoietic system underlie the dioxin-induced increase in pulmonary neutrophils observed during influenza virus infection. We report here that AhR activation does not increase neutrophil numbers systemically or increase neutrophil production in hematopoietic tissue, suggesting that the elevated number of neutrophils is restricted to the site of antigen challenge. The generation of CD45.2AhR-/--->CD45.1AhR+/+ bone marrow chimeric mice demonstrates that even when hematopoietic cells lack the AhR, TCDD treatment still results in twice as many pulmonary neutrophils compared with control-treated, infected CD45.2AhR-/--->CD45.1AhR+/+ chimeric mice. This finding reveals that AhR-mediated events extrinsic to bone marrow-derived cells affect the directional migration of neutrophils to the infected lung. These results suggest that the lung contains important and heretofore overlooked targets of AhR regulation, unveiling a novel mechanism for controlling neutrophil recruitment to the infected lung.

Cytology in food animal practice.

Diagnostic cytology can greatly aid the clinician in determining a more refined diagnosis, prognosis, and treatment plan to serve the client and patient better. Sample collection is not difficult and can be done in the field as well as in a hospital setting. The collection and sample handling procedures described in this article can help the clinician to obtain diagnostically valuable samples. In many cases, preliminary cytologic evaluation can be performed by the general practitioner. Additional diagnostic evaluation and interpretation are readily available from trained pathologists at diagnostic laboratories.