Feline Forensics: Revealing the Unique Decomposition of Cats.

Limited data about the differences between the decomposition of animals with fur exist within the forensic veterinary medicine discipline. Due to the sheer number of animals used as animal models and the number of animals that exist, determining stages of decomposition that are applicable for all species is difficult. Typically, assessing what stage of decomposition a decedent is in is subjective due to the visual assessment of decomposition changes. A method developed to rectify this issue was the total body score (TBS) method, which assigns a numerical value based on the degree of decomposition to the head, torso, and limbs. The purpose of this study was to examine the decomposition of cats, identify any unique decomposition characteristics, and determine the rate of decomposition using total body score and accumulated degree days (ADDs). Twelve domestic short-haired cats were placed in a grassy field in West Lafayette, IN. An assessment of TBS was documented for each cat and each experimental group. An exponential relationship between TBS and ADD was documented. Overall, there was not a significant difference in the rate of decomposition or total body scores between the groups (Z = -91.00, p = 0.0672).

Immunoassay testing for barbiturates using alternative matrices in postmortem tissues from cats and dogs.

The barbiturate drug pentobarbital is commonly used by veterinarians for the euthanasia of domestic animals. During the veterinary forensic autopsy, it is sometimes necessary to determine whether the animal was chemically euthanized with pentobarbital. The use of a human immunochromatographic test for barbiturate screening utilizing dog or cat urine has been previously validated; however, the use of alternative matrices for this purpose is yet to be explored when urine is not available. Postmortem heart, liver, spleen, skeletal muscle, blood and/or urine samples from 20 dogs and 26 cats with a reported chemical euthanasia status were processed using two different methods, bead homogenization and sonication, and screened for barbiturates using a human immunochromatographic test. There was 100% agreement of the immunochromatographic test results using the sonication method with the reported euthanasia status of both dogs and cats. Using the bead homogenization method, agreement with the reported euthanasia status was 93.3% and 96.7% for dogs and cats, respectively, due to invalid test results from four dog and two cat samples. A subset of liver samples (10 canine and 10 feline) was analyzed via gas chromatography-mass spectrometry, and there was 100% agreement between the immunochromatographic test results and gas chromatography-mass spectrometry results for both cats and dogs. Overall, our results support the use of a variety of alternative matrices for barbiturate screening in cats and dogs.

Postmortem Analysis of Vitreous Urea Nitrogen, Creatinine, and Magnesium of Renal and Post-Renal Disease in Cats.

Analysis of vitreous urea nitrogen (VUN) and creatinine (CREA) has been shown to be a useful indicator of azotemia in human postmortem examinations. Previous studies in cats, dogs, horses, and cattle have shown a good correlation between serum and postmortem ocular fluid for urea and creatinine. The aim of this study was to evaluate differences in VUN, CREA, and magnesium (MG) concentrations in vitreous humor collected postmortem from cats that presented with renal or post-renal disease and cats without renal or post-renal disease. Nine cats with renal or post-renal disease and twenty cats without renal or post-renal disease that underwent autopsy were used in this study. Collection of postmortem vitreous humor was performed, and vitreous samples were frozen at -80 °C for a minimum of 24 h prior to analysis using an in-clinic dry chemistry analyzer. Overall, there were statistically significant differences for VUN, CREA, and MG between cats with renal or post-renal disease and cats without renal or post-renal disease. Analysis of vitreous humor from cats suspected to have renal or post-renal disease can provide useful diagnostic information pertaining to renal function and issues in the urethra and urinary bladder. Future studies exploring postmortem vitreous chemistry in cats are warranted.

A non-inferiority study comparing the ultrasound-guided parasacral with a novel greater ischiatic notch plane approach in canine cadavers.

OBJECTIVE: To describe the gross and ultrasound anatomy of the parasacral region and an ultrasound-guided greater ischiatic notch (GIN) plane approach aimed at staining the lumbosacral trunk (LST) in canine cadavers. To evaluate if the ultrasound-guided GIN plane approach is non-inferior to the previously described ultrasound-guided parasacral approach at staining the LST. STUDY DESIGN: Prospective, randomized, non-inferiority experimental anatomic study. ANIMALS: A total of 17 (23.9 ± 5.2 kg) mesocephalic canine cadavers. METHODS: Anatomic and echographic landmarks, and the feasibility of performing a GIN plane technique were evaluated using two canine cadavers. The remaining 15 cadavers had each hemipelvis randomly assigned to be administered either parasacral or GIN plane injection of 0.15 mL kg-1 dye solution. The parasacral region was dissected after injections to assess the staining of LST, cranial gluteal nerve, pararectal fossa and pelvic cavity. The stained LST were removed and processed for histological evaluation of intraneural injections. A one-sided z-test for non-inferiority (non-inferiority margin -14%) was used to statistically evaluate the success of the GIN plane versus the parasacral approach. Data were considered statistically significant when p < 0.05. RESULTS: The GIN plane and parasacral approach stained the LST in 100% and 93.3% of the injections, respectively. The success rate difference between treatments was 6.7% [95% confidence interval, -0.6 to 19.0%; p < 0.001 for non-inferiority]. The GIN plane and parasacral injections stained the LST for 32.7 ± 16.8 mm and 43.1 ± 24.3 mm, respectively (p = 0.18). No evidence of intraneural injection was found. CONCLUSIONS AND CLINICAL RELEVANCE: The ultrasound-guided GIN plane technique resulted in nerve staining that was non-inferior to the parasacral technique and may be considered an alternative to the parasacral approach to block the LST in dogs.

Lumbosacral plexus block using a combination of ultrasound-guided lateral pre-iliac and parasacral approaches in cats.

OBJECTIVE: To describe an ultrasound-guided lateral pre-iliac (LPI) and parasacral (PS) approach in feline cadavers (phase I) and compare the perioperative analgesic use and complications in cats administered LPI and PS blocks (group PNB) or epidural anesthesia (group EPI) for pelvic limb surgery (phase II). STUDY DESIGN: Experimental uncontrolled, anatomic and retrospective cohort study. ANIMALS: A group of eight feline cadavers and 52 medical records. METHODS: Bilateral LPI and PS approaches with 0.1 mL kg-1 of dye to stain the femoral and obturator nerves and the lumbosacral trunk, respectively, were performed on each cadaver. Nerve staining effect was evaluated upon dissections (phase I). Perioperative analgesics use, and complication rates were retrospectively compared between groups PNB and EPI (phase II). Continuous data were compared using the Mann-Whitney U test and the prevalence of events with Fisher's exact test. Differences were considered significant when p < 0.05. RESULTS: Dissections revealed that the LPI approach stained 94% and 75% of the femoral and obturator nerves, respectively. The PS approach stained 100% of the lumbosacral trunks. Cats enrolled in group PNB (n = 23) were administered lower doses of intraoperative opioids than those in group EPI (n = 25) (p = 0.006). Intraoperative rescue analgesia was required in 60% and 17.4% of cats enrolled in groups EPI and PNB, respectively (p = 0.003). Group PNB required more intraoperative anticholinergics than group EPI (p = 0.02). There were no differences in postoperative pain scores, analgesic use and complication rates. CONCLUSIONS AND CLINICAL RELEVANCE: The ultrasound-guided LPI and PS approach stained the femoral/obturator nerves and the lumbosacral trunk, respectively, in feline cadavers. Furthermore, PNB was associated with lower intraoperative opioid use and similar postoperative pain and analgesic use compared with epidural anesthesia in a cohort of cats undergoing surgery of the pelvic limb.

Erector spinae plane block at the thoracolumbar spine: a canine cadaveric study.

OBJECTIVE: To investigate the injectate spread and nerve staining of ultrasound-guided erector spinae plane (ESP) injections at the thoracolumbar spine in canine cadavers. STUDY DESIGN: Prospective, randomized, descriptive, anatomic study. ANIMALS: A total of 15 canine cadavers. METHODS: The location of the medial and lateral branches of the dorsal branches of the spinal nerves (DBSN) from the tenth thoracic (T10) to the third lumbar vertebra (L3) were identified by dissection of three cadavers. ESP injections of dye (0.5 mL kg-1) were performed in seven cadavers using as landmarks the T12 transverse process (ESPTp) on one side and the lateral aspect of the T12 mammillary process (ESPMp) on the opposite side. Additionally, five cadavers were injected with dye (0.5 mL kg-1) bilaterally on the lateral aspect of the L2 mammillary process (ESPMp_L2). Nerve staining effect was analyzed after gross anatomic dissections. The number of stained nerves was analyzed using the Mann-Whitney U test. RESULTS: Gross anatomic dissections showed that the medial and lateral branches of the DBSN change their path in relation to the epaxial muscles caudal to T11. Approaches ESPTp and ESPMp at T12 stained 2 (0-2) and 3 (2-4) medial (p = 0.01) and 3 (3-4) and 2 (0-2) lateral (p = 0.03) branches, respectively. Injection ESPMp_L2 stained 3 (2-4) medial and 2 (0-3) lateral branches. Injections ESPMp and ESPMp_L2 produced a preferential cranial spread from the injection site. No ventral branches of the spinal nerves were stained with either technique. CONCLUSIONS AND CLINICAL RELEVANCE: These results suggest that the mammillary process should be used as anatomic landmark to perform ultrasound-guided ESP blocks in the thoracolumbar spine caudal to T11 when targeting the medial branches of the DBSN. Injections should be performed one spinal segment caudal to the level intended to desensitize.

Evaluation of a human urine barbiturate test to screen for pentobarbital euthanasia of dogs and cats.

Pentobarbital is used commonly to euthanize animals. Occasionally during a death investigation, it is necessary to determine whether a cat or dog was euthanized via pentobarbital overdose. Screening for the detection of barbiturates including pentobarbital can be performed using commercial immunochromatographic tests. We used a commercial immunochromatographic test for barbiturates in humans to screen for barbiturates in urine collected postmortem from 20 dogs and 20 cats to determine whether they had been euthanized with pentobarbital. Additionally, we analyzed the urine for pentobarbital using liquid chromatography-mass spectrometry as a confirmatory test. Screening and confirmation testing revealed 100% agreement between the tests and with the euthanasia status of each animal. Our results support the use of the immunochromatographic test for the screening of urine collected postmortem to assess for the presence of barbiturates, specifically pentobarbital, used for euthanasia.

Postmortem Vitreous Humor Analysis in Dogs, Cats and Horses.

Postmortem chemistry can be a useful ancillary technique that the forensic pathologist can use during a death investigation. In stark contrast, there is limited information available for use of postmortem vitreous humor analysis in animals. In order to use postmortem vitreous humor in veterinary forensic investigations, validation of a method to analyze vitreous humor is required. The goal of this study was to determine the precision, bias, TEobs and sigma (σ) of the Element DC chemistry analyzer; assess its precision using the vitreous humor collected postmortem from dogs, cats and horses and assess the stability of postmortem vitreous humor from all the three species. Analysis of quality control material (QCM) and pooled vitreous humor samples for the three species was used to test for sodium (Na), chloride (Cl), potassium (K), magnesium (Mg), creatinine (Crea) and blood/vitreous urea nitrogen. Analysis of QCM showed that the Element DC was both precise and accurate. When analyzing the pooled vitreous humors, most within-run coefficients of variance (CVs) were found to be <5% and the between-run CVs for five out of six analytes were found to be <5% for dogs, cats and horses. In all the three species, the capped samples of vitreous humor were stable out of refrigeration for up to 5 h. The results of this study show that the Element DC can successfully be used to analyze the postmortem vitreous humor from dogs, cats and horses.

Feasibility of using tissue autolysis to estimate the postmortem interval in horses.

Estimation of the postmortem interval (PMI) is a poorly studied field in veterinary pathology. The development of field-applicable methods is needed given that animal cruelty investigations are increasing continually. We evaluated various histologic criteria in equine brain, liver, and muscle tissue to aid the estimation of PMI in horses, which is central to forensic investigations of suspicious death. After death, autolysis proceeds predictably, depending on environmental conditions. Currently, no field-applied methods exist that accurately estimate the PMI using histology in animals or humans through quantification of autolysis. Brain, liver, and skeletal muscle from 12 freshly euthanized horses were held at 22°C and 8°C for 72 h. Tissues were sampled at T0h, T1h, T2h, T4h, T6h, T12h, T24h, T36h, T48h, T60h, and T72h. For each tissue, we quantified 5 to 7 criteria associated with autolysis, based on the percentage of microscopic field involved. Each criterion was modeled, with temperature and time as independent variables. Changes were most predictable in liver and muscle over the first 72 h postmortem. The criteria for autolysis that were present most extensively at both temperatures were hepatocyte individualization and the separation of bile duct epithelium from the basement membrane. The changes that were present next most extensively were disruption of myofiber continuity, hypereosinophilia, and loss of striation. Brain changes were highly variable. The high statistical correlation between the parameter "autolysis" and the variables "time/temperature", indicates that autolysis is progressive and predictable. Further investigation of these criteria is needed to establish histologic algorithms for PMI.

Sylvatic Canine Morbillivirus in Captive Panthera Highlights Viral Promiscuity and the Need for Better Prevention Strategies.

Canine Distemper Virus (CDV) is a multi-host morbillivirus that infects virtually all Carnivora and a few non-human primates. Here we describe a CDV outbreak in an exotic felid rescue center that led to the death of eight felids in the genus Panthera. Similar to domestic dogs and in contrast to previously described CDV cases in Panthera, severe pneumonia was the primary lesion and no viral antigens or CDV-like lesions were detected in the central nervous system. Four tigers succumbed to opportunistic infections. Viral hemagglutinin (H)-gene sequence was up to 99% similar to strains circulating contemporaneously in regional wildlife. CDV lesions in raccoons and skunk were primarily encephalitis. A few affected felids had at least one previous vaccination for CDV, while most felids at the center were vaccinated during the outbreak. Panthera sharing a fence or enclosure with infected conspecifics had significantly higher chances of getting sick or dying, suggesting tiger-tiger spread was more likely than recurrent spillover. Prior vaccination was incomplete and likely not protective. This outbreak highlights the need for further understanding of CDV epidemiology for species conservation and public health.

Microscopic examination of dog chews: correlation of histological findings to product labeling.

The purpose of this study was to use routine morphologic-based staining techniques to examine the histology of commercially labeled rawhide and rawhide-free dog chew products and compare the results to the product labeling. Ten dog chew products were examined by light microscopy using hematoxylin and eosin and Masson's trichrome stains. The products were labeled by the manufacturer as rawhide, beef hide, beef chew/rawhide free, and rawhide free. Four of the products were composed of two separate materials, a main chew roll and a second substance (filler) which was coated on or between the layers of the main chew roll. These materials were processed independently. Microscopically, a variety of tissues and materials were identified including collagen, skeletal muscle, fat, plant material, and starch. The products and their fillers were separated into four distinct groups based on microscopic appearance. The components identified in eight of the products appeared consistent with the product labeling. Two products labeled as rawhide free appeared similar to the dermis and this was inconsistent with product labeling. Masson's trichrome stain was not helpful in distinguishing tissue types in the tested products and this may have been due to the heat processing the products underwent during manufacturing. Bacteria and/or fungi were identified by microscopy in the H&E stained sections in four rawhide-free products.

MORTALITY INVESTIGATION OF MONITORED EASTERN BOX TURTLES (TERRAPENE CAROLINA CAROLINA) IN CENTRAL ILLINOIS, USA, FROM 2016-18.

Mortality events in eastern box turtles (Terrapene carolina carolina) threaten conservation efforts across the species range. These events are often under-diagnosed and, when observed, predictive health factors are unavailable prior to death. At Kickapoo State Park in central Illinois, USA, ranaviruses caused observed mortality events in amphibians and chelonians in 2014 and 2015. Following these outbreaks, eastern box turtles (n=36) were affixed with radio transmitters and temperature data loggers to obtain repeated location and temperature data from spring 2016-spring 2018. Bimonthly, samples of blood and oral and cloacal swabs were collected to investigate health parameters (hematology and cytokine transcription) and presence of multiple pathogens. Deaths of instrumented turtles occurred in 2016 (n=5), 2017 (n=15), and 2018 (n=2). The largest single die-off occurred in February 2017 (n=7). Seventeen turtles were necropsied and multiple pathologic processes were identified, most frequently decreased adipose stores (n=6). Two turtles had pathologic findings consistent with multisystemic inflammation. In addition, infectious pathogens were identified in turtles prior to death, but no single agent was associated with each mortality event. Ranavirus was not detected in any turtle. Hot spot analysis revealed spatial clustering at the center and edges of the study area for body temperature as well as for relative cytokine transcription of interleukin-1 beta, tumor necrosis factor alpha, and interleukin-10 associated with turtle death. Though no single causal factor could be identified, the information from this mortality event can direct future chelonian mortality investigations by providing baseline longitudinal data prior to death and in surviving turtles.

Understanding the Efficiency of Splenic Hemangiosarcoma Diagnosis Using Monte Carlo Simulations.

Hemangiosarcoma is a common neoplasm of the spleen in older dogs. However, diagnosis is complicated by necrosis and hemorrhage, which can mimic a number of benign processes. Currently, there is no consensus about the number of sections pathologists should examine to rule out hemangiosarcoma. To answer this question, we examined 413 histopathologic sections from 50 cases of canine hemangiosarcoma (mean: 8.1 sections per case; range, 5-14). Each section had the presence or absence of hemangiosarcoma determined by 2 board-certified anatomic pathologists. Then, 100 Monte Carlo simulations were performed, randomly selecting sections from each case 10 000 times and the results averaged. These simulations suggest that examination of 5 sections from a spleen with hemangiosarcoma yields a 95.02% chance of diagnosing hemangiosarcoma, while examination of 10 sections yields a 98.59% chance of diagnosis when hemangiosarcoma is in fact present. The data emphasize the need to submit the entire spleen for histopathologic examination in suspected cases of hemangiosarcoma and suggest that 5 sections obtained by a trained individual are likely sufficient for diagnosis.

Prevalence of CD20+ cutaneous epitheliotropic T-cell lymphoma in dogs: a retrospective analysis of 24 cases (2011-2018) in the USA.

BACKGROUND: Limited information is known about the baseline frequency of canine CD20 positive (+ ) cutaneous epitheliotropic T-cell lymphoma. A single canine case report has been published. OBJECTIVES: To characterize the baseline frequency of CD20+ staining in cases of canine cutaneous epitheliotropic T-cell lymphoma; to assess its values as a prognostic indicator. ANIMALS: Skin biopsies from 24 client-owned animals diagnosed with cutaneous epitheliotropic lymphoma were assessed. METHODS AND MATERIALS: A retrospective review of medical records from 2011 to 2018. Clinical histories and previous histological and immunohistochemical slides were collected from 24 dogs and additional immunohistochemical staining was performed as needed to assess cases of cutaneous epitheliotropic T-cell lymphoma for CD3, CD20, CD79a and PAX5 staining characteristics. Staining characteristics were assessed for their statistical relationship to median survival time. RESULTS: Overall median survival time was 189 days following the onset of clinical signs and 99 days following definitive diagnosis; 54% of cases had CD20+ staining. There was no statistically significant correlation between staining characteristics and median survival time from onset of signs (P = 0.54) or from diagnosis (P = 0.61). CONCLUSIONS AND CLINICAL SIGNIFICANCE: Canine cutaneous epitheliotropic lymphoma has a higher instance of CD20 positivity than documented previously. This indicates that CD20 staining may be unsuitable to differentiate T-cell and B-cell cutaneous epitheliotropic lymphoma. This also may have therapeutic implications with the advent of canine CD20 monoclonal antibody therapies.

FOXA2 depletion leads to mucus hypersecretion in canine airways with respiratory diseases.

Because of exposure to environmental pollutants, infectious agents, and genetic predisposition, companion animals develop respiratory illnesses similar to those in humans. Older dogs of smaller breeds develop canine infectious respiratory disease, chronic bronchitis, and chronic obstructive pulmonary disease, with chronic lung infection, airway goblet cell hyperplasia and metaplasia, and mucus hypersecretion. Excessive mucus clogs airways, reduces gas exchanges, disables the mucociliary clearance, and reduces drug penetration. The Forkhead box protein A2 (FOXA2) is a key transcriptional regulator that maintains airway mucus homeostasis. Prior studies have shown that FOXA2 expression is frequently depleted in diseased human airways. Unfortunately, FOXA2 depletion has not been examined in dogs. Our current study indicated that both single bacterial infection by Pseudomonas aeruginosa and Bordetella bronchiseptica and polymicrobial infection by viral/bacterial pathogens depleted FOXA2 in canine airways, resulting in goblet cell hyperplasia and metaplasia and excessive mucus production. Furthermore, P. aeruginosa virulence factor pyocyanin activated the antagonistic STAT6 and epidermal growth factor receptor signalling pathways to inhibit FOXA2. Unravelling the mechanism of FOXA2 inactivation will hasten the development of non-antibiotic therapeutics to improve mucociliary clearance of pathogens in canine airway.

Comparing dermoscopy and histological examination of normal equine skin.

BACKGROUND: Dermoscopy is a noninvasive diagnostic technique that allows visualization of structures of the superficial dermis not visible with the naked eye. HYPOTHESIS/OBJECTIVES: To assess the usefulness and applicability of dermoscopy for evaluation of healthy equine skin. ANIMALS: Twelve healthy horses from a research herd. METHODS: Five regions (cheek, lateral neck, dorsum, flank and abdomen) were examined with contact dermoscopy using both nonpolarized and polarized light at both 17-fold and 24-fold magnification. These findings were compared to histological features of skin biopsies cut both longitudinally and transversely. RESULTS: Using a hand-held dermatoscope with nonpolarized light, epidermal ridges were observed. Using polarized light, follicular openings and distinctly separate epidermal openings of sweat gland ducts were observed in some but not all individuals. Similarities were noted between histological and dermoscopic results. CONCLUSIONS: Although not ideal for visualizing many structures in the superficial dermis of healthy equine skin, dermoscopy allowed visualization of epidermal ridges, hair shafts in the infundibular portion of the hair follicles and sweat gland duct openings. Dermoscopy could potentially be useful in the evaluation of diseases affecting the sweat glands, epidermis and hair shaft.

Anesthetic effects and body weight changes associated with ketamine-xylazine-lidocaine administered to CD-1 mice.

Anesthesia for mice is commonly performed through the injection of parenteral agents via the intraperitoneal (IP) route. Variability in anesthetic sensitivities has been noted in mice resulting in inconsistencies in anesthetic depth and/or mortality. Anesthetic protocols that improve consistency and safety are needed. The objectives of this study were to assess the effects of intraperitoneal (IP) ketamine (95 mg/kg) and xylazine (7 mg/kg) alone or combined with lidocaine at 4, 8, or 16 mg/kg on time to loss (LRR) and return (RRR) of righting reflex, duration of immobilization and loss of pedal withdrawal response (PWR), body weight and histopathology in CD-1 mice. In a prospective, randomized trial, 36 male CD-1 mice, 4-6 weeks of age were randomly assigned to 5 groups: saline (SA, n = 4); ketamine-xylazine (KX, n = 8); ketamine-xylazine-lidocaine 4 mg/kg (KXL4, n = 8); ketamine-xylazine-lidocaine 8 mg/kg (KXL8, n = 8); ketamine-xylazine-lidocaine 16 mg/kg (KXL16, n = 8). Two mice in each group were euthanized at day 2 post-injection and the remaining mice were euthanized at day 11 post-injection. After IP injection, LRR and RRR, duration of immobilization and loss of PWR, body weight and histopathology were evaluated. LRR occurred sooner in mice receiving KXL16 compared with KX, with median (range) times of 78 (62-104) and 107 (91-298) seconds, respectively. Loss of PWR occurred in 1, 5, 4, 6 mice for groups KX, KXL4, KXL8, and KXL16 respectively. Median (range) duration of absent PWR was longer in mice receiving KXL16 at 13 (0-30) minutes, compared to KX at 0 (0-9) minutes. Duration of immobilization and RRR were not different between groups. Weight loss occurred 2 days following anesthesia but was not different between groups. Weight gain was significantly greater in all lidocaine groups 11 days post-injection compared to KX. No mortality or histopathologic abnormalities were observed in any group. Lidocaine administered with ketamine and xylazine shortens the onset of anesthesia in mice and improves anesthetic depth without prolonging recovery time.