Pathological findings of Japanese Black Cattle living in the restricted area of the Fukushima Daiichi Nuclear Power Plant accident, 2013–2016

Fifty‐one Japanese black cattle from four farms in the evacuation zone of the Fukushima nuclear accident were examined pathologically during the period, 2013–2016. We found no evidence of a radiation effect on pathological findings in any of these autopsy cases, although nine cases (3.7%) of enzootic bovine leukosis (EBL) and three cases (1.2%) of goiter were diagnosed. Estimated integrating dose of external exposure in EBL cases ranged from a maximum of 1200 mSv to a minimum of 72 mSv. Clinically, five cases showed wobble, dysstasia or paralysis. Exophthalmos was observed in three cases. Macroscopically, enlarged lymph nodes, multiple irregular masses of intra‐abdominal and intrapelvic adipose tissue, diffuse thickening of the abomasal submucosa with ulceration, and numerous white nodules of myocardium were observed. Histologically, neoplastic lymphoid cells were extensively proliferated and infiltrated in the lesions. Lymphoid tumor cells showed mature or large lymphoblastic appearance. Immunohistochemically, tumor cells were positive for BLA‐36, CD20 and CD5, and negative for CD3. Three cases showed diffuse enlargement of the thyroid gland with no mass or nodule lesion. Histologically, diffuse proliferation of follicular epithelium was observed with preservation of normal structures. There were no malignant findings such as cellular atypia or invasion to capsule.     

Radioactive cesium and potassium in cattle living in the ‘zone in preparation for the lifting of the evacuation order’ of the Fukushima nuclear accident

To contribute to the reconstruction of livestock industry in Fukushima, radioactive cesium (¹³⁴Cs, ¹³⁷Cs) and potassium (⁴°K) were measured in various tissues of beef cattle living in an area where the evacuation order will be lifted in the near future. Radioactive cesium concentration was less than 100 Bq/kg in most of the samples. Skeletal muscles and kidney had the highest concentrations of radioactive cesium, whereas the liver was lowest among samples, excepting blood. Radioactive cesium concentration in the sirloin, tenderloin and top round was significantly higher than that in the neck muscle. Radioactive cesium concentration in the urine was not correlated with that in the blood, but the relationship became proportional when corrected with urinary ⁴°K. Distribution of ⁴°K was similar but not identical to that of radioactive cesium. These results suggest that it will be possible to resume livestock production in this area after the decontamination measures are completed and the evacuation order is lifted. Contamination level of living cattle can be estimated not only by blood samples but also by urine samples. If 50–100 Bq/kg of cesium is detected in the neck muscle at a slaughterhouse, the cattle should be reinspected using a sample from different muscles.     

Pathological characteristics of thyroid glands from Japanese Black Cattle living in the restricted area of the Fukushima Daiichi Nuclear Power Plant accident

To study the effect of ionizing radiation on thyroid glands, 66 Japanese Black cattle residing in the restricted area of the Fukushima Daiichi Nuclear Power Plant in 2013–2017 were pathologically examined. There were no clinical symptoms of thyroid disease in these cattle. Three cases of goiter and seven of atrophy were found in two among the four farms examined. Cases of goiter exhibited normal morphological structure without mass or nodule formation in thyroid glands. Cellular atypia or capsular invasion of the follicular epithelium was absent. The estimated integrated dose of external radiation in goiter cases ranged from maximum 797 mSv to minimum 24 mSv. All lobules in the seven atrophic thyroid glands were affected, but pathological findings, such as inflammatory cell infiltration or stromal fibrosis, were not observed. The estimated integrated dose of external radiation in atrophic thyroids ranged from maximum 589 mSv to minimum 8 mSv. Immunohistochemical analysis of anti‐nitroguanosine and the TUNEL method in goiter and atrophic thyroid glands did not reveal any positive findings. The present study indicates that there was no significant relationship between a radiation effect and pathological findings in any thyroid glands.     

Unusual “erythroid loops” in canine blood smears after viper‐bite envenomation

Abstract: Northern Italy is a habitat for many species of viper; Vipera aspis is responsible for most reported bites of humans and animals. Five dogs of different breeds and ages were presented by their owners to a veterinary clinic in northern Italy between September 2004 and August 2007 with a history of being bitten by a viper within the past 2 hours. On physical examination, all of the dogs were depressed, had pale mucous membranes, and had a painful area consistent with a recent viper bite wound on the nose (2), distal front leg (2), or lip (1). Hemoglobinemia and hemoglobinuria were observed in plasma and urine from all dogs. CBCs were done at the time of presentation, daily for 4 days, and 10 days after presentation in all dogs with the exception of one dog that died after 2 days. All dogs had an acute decrease in HCT within 24 hours of presentation; all dogs had neutrophilia, 3/5 had a mildly toxic left shift, and 4/5 had thrombocytopenia. On Diff‐Quik‐stained blood smears, moderate numbers of echinocytes, spherocytes, and erythrocyte ghosts were observed. In addition, moderate numbers of unusual erythrocyte membrane‐like structures (“erythroid loops”) were observed. The loops were annular in shape and sometimes disrupted, appearing as thin pale blue bands. Erythrocyte morphologic abnormalities decreased by day 3 and were no longer observed on day 10. The unique appearance of the erythroid loops together with evidence for intravascular hemolysis and other erythrocyte morphologic changes suggest they may be a consequence of erythrocyte lysis. Echinocytes, spherocytes, and erythrocyte ghosts are known to result from the action of phospholipase in viper venom; however, erythroid loops have not been reported previously and their exact mechanism of formation is unknown.     

24‐Hour Kinetics of Cardiac Troponin‐T Using a “High‐Sensitivity” Assay in Thoroughbred Chuckwagon Racing Geldings after Race and Associated Clinical Sampling Guidelines

Background

A “high‐sensitivity” cardiac troponin‐T (hscTnT) assay recently has been validated for use in horses and is a specific biomarker of myocardial damage. Postexercise release kinetics of cTnT utilizing the hscTnT assay have yet to be established in horses.

Objectives

To determine: (1) cTnT release kinetics in racing Thoroughbreds after a high‐intensity 5/8th mile Chuckwagon race; (2) the effects of age on pre‐ and postrace cTnT concentrations; and (3) sampling guidelines for clinicians evaluating horses presenting after exercise.

Animals

Samples were obtained from 38 Thoroughbred geldings aged 5–16 years before racing and immediately, 2, 3, 4, 6, 12, and 24 hour postrace.

Methods

Prospective, observational study with convenience sampling. A fifth‐generation hscTnT assay was used for plasma sample analysis, and concentrations were compared at all time‐points. Correlations were determined between cTnT concentrations and age. Biochemistry analysis was performed to assess rhabdomyolysis, renal failure, and exercise‐induced dehydration.

Results

All horses with measureable cTnT concentrations had significant postexercise increases in cTnT with a median peak (8.0 ng/L) at 3‐hour postrace. All horses had peak postexercise cTnT concentrations 2‐ to 6‐hour postrace ≤ the 99th percentile upper reference limit of 23.2 ng/L, after which all cTnT concentrations decreased until returning to baseline by 12–24 hours. There was no correlation over time between cTnT concentrations and age.

Conclusions and Clinical Importance

In racing Thoroughbreds completing short‐duration, high‐intensity Chuckwagon races, cTnT concentrations are expected to be increased 2‐ to 6‐hour postrace and to decrease by 12–24 hours while remaining ≤23.2 ng/L throughout. This study contributes to establishing guidelines for clinical use of the hscTnT assay in exercising horses.     

Effect of synchronization of donor cells in early G1‐phase using shake‐off method on developmental potential of somatic cell nuclear transfer embryos in cattle

In this study, we compared the developmental ability of somatic cell nuclear transfer (SCNT) embryos reconstructed with three bovine somatic cells that had been synchronized in G0‐phase (G0‐SCNT group) or early G1‐phase (eG1‐SCNT group). Furthermore, we investigated the production efficiency of cloned offspring for NT embryos derived from these donor cells. The G0‐phase and eG1‐phase cells were synchronized, respectively, using serum starvation and antimitotic reagent treatment combined with shaking of the plate containing the cells (shake‐off method). The fusion rate in the G0‐SCNT groups (64.2 ± 1.8%) was significantly higher than that of eG1‐SCNT groups (39.2 ± 1.9%) (P < 0.05), but the developmental rates to the blastocyst stage of SCNT embryos per fused oocytes were similar for all groups. The overall production efficiency of the clone offspring in eG1‐SCNT groups (12.7%) per recipient cow was higher than that in G0‐SCNT groups (3%) (P < 0.05). The mean birth weight of cloned calves and the average calving score in the G0‐SCNT groups (48.1 ± 3.4 kg and 3.3 ± 0.3, respectively) was significantly higher (P < 0.05) than those of eG1‐SCNT groups (37.2 ± 2.1 kg and 2.3 ± 0.2, respectively). Results of this study indicate that synchronization of donor cells in eG1‐phase using the shake‐off method improved the overall production efficiency of the clone offspring per transferred embryo.     

The “Tears of the Virgin” at Lakes Entrance,southeast Australia were made by the intertidal barnacle Chthamalus antennatus (Cirripedia: Thoracica) and cyanobacteria

Curious eroded depressions, most resembling an eye shedding an elongate tear, are found in gently sloping, intertidal, carbonate‐rich arenite outcropping on the sea coast near Lakes Entrance, Victoria, southeast Australia. The depressions, known locally as “Tears of the Virgin,” are evidently formed by multiple generations of a barnacle, Chthamalus antennatus Darwin, 1854 in association with cyanobacteria. While the round part of a depression offers the barnacle a modicum of protection from impacts during high tides, it is also partially inhabited by cyanobacteria, which extend into and tend to fill the elongate tear. As such, this appears to be the first case of mutualism between a higher invertebrate and cyanobacteria, with the cyanobacteria reducing the barnacle's risk of desiccation while receiving metabolic wastes from it during low tides. It is also the first record of a balanomorph barnacle eroding calcareous arenite beneath its shell, the net effect of which would be expected to reduce its adhesion to the substrate. However, the siliceous residue, resulting from the barnacle's dissolution of the more than 80% of the calcite‐rich sedimentary rock, is sequestered in delicate folds on the inside of the shell wall as it grows. A brief review of cirripedes capable of excavation includes the first photographic documentation of excavation of a mollusc shell by a verrucomorph.     

Discrimination of “grazing milk” using milk fatty acid profile in the grassland dairy area in Hokkaido

Milk produced by the grazing system, referred to as “grazing milk” contains many components required for human health. The milk fatty acid (FA) profile is strongly associated with the diet on the farms. In the present study, based on the FA profile of farmer's bulk milk, we determined how to discriminate between milk produced on grazing and on a confinement system. A field survey was conducted four times (grazing and confinement season) in the Konsen (29 farms) and Okhotsk (25 farms) area in Hokkaido. Farmer's bulk milk samples and details of feeding management were collected and the FA profile of milk was measured. Milk produced during the grazing season contained less C16:0 and cis‐9 C16:0, and more C18:0, cis‐9 C18:1, trans‐11 C18:1, cis‐9,12 C18:2, cis‐9,trans‐11 C18:2 and cis‐9,12,15 C18:3 than milk produced during the confinement season. Discrimination analysis using 16 FA revealed that almost all milk samples were discriminated correctly (confinement season: 90% correct and 10% borderline, grazing season: 88% correct, 9% borderline and 3% incorrect). For farmers that were categorized incorrectly and were considered borderline in the grazing season, the dependency on pasture was low compared with that for farmers correctly discriminated. Therefore, to claim “grazing milk”, a high dependency on pasture is required for grazing dairy farmers.     

“In vitro” capacitation and further progesterone‐induced acrosome exocytosis are linked to specific changes in the expression and location of threonine phosphorylation of boar spermatozoa

The aim of this study was to determine if the achievement of the “in vitro” capacitation (IVC) status and subsequent progesterone‐induced “in vitro” acrosome exocytosis (IVAE) was accompanied with overall changes in threonine phosphorylation (pThre) of boar spermatozoa. For this purpose, mono‐ and bi‐dimensional Western blot analyses as well as immunocytochemistry studies against pThre were performed in boar sperm subjected to IVC and subsequent IVAE. Mono‐dimensional Western blot in non‐capacitated samples showed that launching of IVC did induce an overall increase in signal intensity in all observed bands that was followed by a subsequent decrease afterwards. Bi‐dimensional Western blot analysis showed the presence of four main signal protein clusters. The attainment of IVC induced an overall decrease in the number and intensity of spots of Clusters A, B and C and a concomitant increase in the intensity of spots of Cluster D. The IVAE launching caused a rapid increase in the intensity of spots of Clusters B, C and D, which was followed by a subsequent decrease of the intensity together with a concomitant pI displacement of Cluster C. Finally, immunocytochemistry showed that the pThre signal of non‐capacitated cells was located at the whole sperm. The IVC did not induce prominent changes in this location. In contrast, the induction of IVAE caused the appearance of an additional an intense acrosome and tail pThre signal that subsequently decreased. In conclusion, our results indicate that IVC and further IVAE induced specific changes in the intensity and appearance of pThre protein phosphorylation which were linked to changes of specific protein characteristics as pI. These results support, thus, the existence of a specific role of pThre in IVC/IVAE of boar sperm.     

Transitional cell carcinoma of the urinary bladder in a 12‐year‐old Belgian Warmblood gelding

A 12‐year‐old Belgian Warmblood gelding was examined for haematuria and dysuria of 24 h duration. Cystoscopy revealed an intraluminal multinodular soft tissue mass originating from the dorsal bladder neck. Histopathological examination of biopsies identified transitional cell carcinoma. The bladder mass was surgically debulked via a temporary perineal urethrotomy. The horse commenced treatment with oral piroxicam. Follow‐up examination 18 months post operatively revealed no evidence of tumour recurrence. Neoplasia of the equine bladder is uncommon and this case describes the successful short‐term outcome of treatment of a transitional cell carcinoma by surgical debulking and oral piroxicam.     

Intralesional 5‐fluorouracil (5‐FU) for the treatment of eyelid squamous cell carcinoma in 5 horses

Eyelid squamous cell carcinoma in equine patients often presents a therapeutic challenge to practitioners due to the generally large area affected upon presentation. Surgical excision can be curative if wide enough margins are achieved, but this is not often attainable without enucleation. Other alternatives have been examined including cryotherapy, radiotherapy, brachytherapy, intralesional chemotherapy and photodynamic therapy. Intralesional chemotherapy using cisplatin, mitomycin‐C and bleomycin have been shown to be successful in treating eyelid squamous cell carcinomas but may be prohibitive to some owners due to the cost of therapy. To the authors' knowledge, this is the first case series to illustrate the effectiveness of intralesional 5‐fluoruracil in treating large equine eyelid squamous cell carcinomas. Macroscopic reduction in the size of the affected areas was noted in each case together with regression of clinical signs associated with the mass. This, coupled with the relative inexpensive nature of the procedure, makes this technique an attractive therapy for either primary treatment of eyelid squamous cell carcinoma or as a cytoreductive technique prior to surgical excision.     

Analytical validation of the Sysmex XT‐2000iV for cell counts in canine and feline effusions and concordance with cytologic diagnosis

Background: The Sysmex XT‐2000iV is a hematology analyzer that combines laser and impedance technology. Its usefulness for determining cell counts in canine and feline intracavitary effusions has not yet been studied. Objectives: The objectives of this study were to evaluate the analytical performance of the Sysmex XT‐2000iV for cell counts in effusions from dogs and cats, and to assess correlation with an impedance counter and concordance with diagnoses based on cytologic findings. Methods: Effusions (43 pleural, 23 peritoneal, 6 pericardial) were analyzed from 32 dogs and 34 cats. Total nucleated cell count (TNCC), HCT, and RBC count were determined on the Sysmex and compared with those obtained on an impedance counter (Hemat 8, SEAC). Imprecision, linearity, and limit of detection were determined for the Sysmex. An algorithm was designed using quantitative and qualitative data from the Sysmex to classify the effusions and the results were compared with diagnoses based on cytologic findings. Results: Intra‐assay and interassay coefficients of variation on the Sysmex were variable. Linearity of TNCC was ≥0.993 for dogs and cats, with the exception of effusions from cats with feline infectious peritonitis, which had delta (Δ) TNC values >3.0. In comparison with the Hemat 8, a proportional error was found for TNCC on the Sysmex. Effusion classification based on the algorithm was concordant with that obtained by cytologic examination in 43/72 (60%) samples. Discordant results usually were due to the misclassification of cells with similar morphology (such as mesothelial and carcinoma cells) in Sysmex scattergrams. Conclusion: The Sysmex XT‐2000iV provides a precise and accurate TNCC and has moderate concordance with cytologic findings for classifying canine and feline effusions. Although microscopic examination of effusions is necessary to achieve an accurate diagnosis, the Sysmex can provide preliminary information that may be helpful to cytopathologists.     

Effects of supplementation with green tea by‐products on growth performance,meat quality,blood metabolites and immune cell proliferation in goats

Forty‐eight castrated male goats were used to determine the effects of feeding green tea by‐products (GTB) on growth performance, meat quality, blood metabolites and immune cell proliferation. Experimental treatments consisted of basal diets supplemented with four levels of GTB (0%, 0.5%, 1.0% or 2.0%). Four replicate pens were assigned to each treatment with three goats per replicate. Increasing dietary GTB tended to linearly increase the overall average weight gain and feed intake (p = 0.09). Water holding capacity, pH and sensory attributes of meat were not affected by GTB supplementation, while cooking loss was reduced both linearly and quadratically (p < 0.01). The redness (linear; p = 0.02, quadratic; p < 0.01) and yellowness (quadratic; p < 0.01) values of goat meat were improved by GTB supplementation. Increasing dietary GTB quadratically increased protein and decreased crude fat (p < 0.05), while linearly decreased cholesterol (p = 0.03) content of goat meat. The proportions of monounsaturated fatty acid, polyunsaturated fatty acid (PUFA) and n‐6 PUFA increased linearly (p < 0.01) and n‐3 PUFA increased quadratically (p < 0.05) as GTB increased in diets. Increasing dietary GTB linearly increased the PUFA/SFA (saturated fatty acid) and tended to linearly and quadratically increase (p ≤ 0.10) the n‐6/n‐3 ratio. The thiobarbituric acid‐reactive substances values of meat were lower in the 2.0% GTB‐supplemented group in all storage periods (p < 0.05). Dietary GTB linearly decreased plasma glucose and cholesterol (p < 0.01) and quadratically decreased urea nitrogen concentrations (p = 0.001). The growth of spleen cells incubated in concanavalin A and lipopolysaccharides medium increased significantly (p < 0.05) in response to GTB supplementation. Our results suggest that GTB may positively affect the growth performance, meat quality, blood metabolites and immune cell proliferation when supplemented as a feed additive in goat diet.     

Sensitive detection of the c‐KIT c.1430G>T mutation by mutant‐specific polymerase chain reaction in feline mast cell tumours

Here, we describe the establishment of mutant‐specific polymerase chain reaction (PCR) for detection of a c‐KIT c.1430G>T mutation in feline mast cell tumours. Several mutations in feline c‐KIT have been identified, with the c.1430G>T mutation accounting for a significant portion of feline mast cell tumour mutations. The c.1430G>T mutation in c‐KIT exon 9 was detected in 15.7% (11 of 70) of samples by mutant‐specific PCR but in only 7.1% (5 of 70) by PCR–restriction fragment length polymorphism (RFLP) in the genomic DNA isolated from 70 formalin‐fixed paraffin‐embedded sections or cells collected by fine needle aspiration. Mutant‐specific PCR showed remarkably higher detection rate than did PCR–RFLP. DNA sequence analysis did not always yield identical results to those of mutant‐specific PCR, suggesting heterogeneity of tumour cells. Mutant‐specific PCR is a valid and efficient screening tool for detection of the c‐KIT c.1430G>T point mutation in feline mast cell tumours compared with PCR–RFLP and sequencing analysis.     

Evaluation of the global DNA methylation in canine mast cell tumour samples by immunostaining of 5‐methyl cytosine

Cutaneous mast cell tumours (MCT) are the most common skin tumour in dogs, and to our knowledge, there are no previous studies regarding the global methylation of these tumours. DNA hypomethylation and hypermethylation have been described in several tumours and both mechanisms can lead to carcinogenesis. The purpose of this study was to evaluate the global DNA methylation in canine MCT. A total of 48 MCT samples were classified in grades 1, 2 and 3 or high‐grade or low‐grade. Monoclonal antibodies were used for the immunohistochemical detection of the 5‐methylcytosine. The immunostained nuclei were classified in strong, weak or negative pattern, and these were quantified in five distinct microscopic fields (40× objective) in each slide. The results showed that global DNA hypomethylation was predominant in grade 3, high‐grade, less differentiated MCT. These epigenetic changes in neoplastic mast cells warrant further detailed investigation aiming the establishment of tumour epigenetic therapies.