Inhibitory effect of penicillin on caecal urease activity in chickens fed on a low protein diet plus urea

1. The effects of dietary penicillin on the urease activities of small intestine, large intestine, caecum and their contents, liver and kidney in chickens fed a diet containing 5 g/kg protein plus urea were examined.

2. About 0.88 of the total urease activity determined was observed in intestinal contents, of which 0.95 of the activity was accounted for by caecal contents, 0.05 by colo‐rectal contents and none by small intestinal contents. Intestinal tissues (caecum included), liver and kidney accounted for 0.03, 0.06 and 0.02, respectively, of the total urease activity.

3. Dietary penicillin decreased urease activity to 0.17 in caecal contents and to 0.05 in colo‐rectal contents of the corresponding control values (P< 0.01). The urease activity of caecal tissue was lowered by penicillin to half that of control activity (P < 0.05) but none of the activities of other tissues were affected.

4. It is concluded that, even when the urease activity is stimulated by dietary urea, 20 mg/kg dietary penicillin can strikingly lower it in the caecum, where most of the urease activity in the chicken body is to be found.     

Laboratory diagnosis of canine GM2-gangliosidosis using blood and cerebrospinal fluid.

In the present study, laboratory techniques were used to diagnose canine GM2-gangliosidosis using blood and cerebrospinal fluid (CSF) that can be collected noninvasively from living individuals. Lysosomal acid beta-hexosaminidase (Hex) was measured spectrofluorometrically using 4-methylumbelliferyl N-acetyl-beta-D-glucosaminide and 4-methylumbelliferyl 7-(6-sulfo-2-acetamido-2-deoxy-beta-D-glucopyranoside) as substrates. Main isoenzymes A and B of Hex in leukocytes were also analyzed using cellulose acetate membrane electrophoresis. GM2-ganglioside in CSF was detected and determined quantitatively by using thin-layer chromatography/enzyme-immunostaining method with anti-GM2-ganglioside antibody. In normal dogs, Hex activities could be determined in leukocytes, serum, and CSF and the total activities were markedly reduced in all the enzyme sources in a dog with Sandhoff disease. Electrophoresis of a leukocyte lysate from a normal dog showed that the Hex A and Hex B were not separated distinctively with formation of a broad band, whereas there were no bands in electrophoresis of a lysate from a dog with Sandhoff disease, showing a deficiency in the total enzyme activity. GM2-ganglioside could be detected and determined quantitatively in as little as 100 microl of canine CSE GM2-ganglioside in CSF in a dog with Sandhoff disease increased to 46 times the normal level. In conclusion, the methods in the present study are useful for diagnosis of canine GM2-gangliosidosis. These techniques enable definitive and early diagnosis of canine GM2-gangliosidosis even if tissues and organs cannot be obtained.     

Pathological analysis of batch safety testing of veterinary vaccines using small laboratory animals

Batch safety tests (BSTs) of veterinary vaccines are conducted using small laboratory animals to assure the safety of vaccines according to several criteria, including clinical signs and change in body weight. Although the latter is used as an evaluation index in BSTs, there have been no reports on the internal changes that affect body weight during the test period. Therefore, we analyzed BST via pathological examination of the tested animals. Here, BSTs were performed for 176 batches using mice and 126 batches using of guinea pigs. Most of the gross findings could be classified into four lesion types (nodules, adhesions, ascites, no apparent signs), with only one vaccine inducing lesions that could not be classified into any of these four types. Histopathological examination revealed that the reactions caused by BST were pyogenic and/or granulomatous inflammation. Nodular or adhesive lesions comprised more severe pyogenic granulomatous inflammation than ascites or cases with no apparent gross lesions. These nodular or adhesive lesions were more frequently induced by vaccines that contained an adjuvant than by vaccines that did not contain an adjuvant. The cases with “exceptional” gross findings histologically presented severe necrosis of the hematopoietic system. Additional testing showed that these “exceptional” lesions were induced when a specific type of light liquid paraffin was injected along with other vaccine additives. Our results show that body weight loss and/or lesions during BST were induced by proinflammatory properties of the tested vaccines and that BST is a sensitive method for detecting unexpected effects of vaccine components.     

Expression of P-glycoprotein and breast cancer resistance protein in three cases of canine lymphoma showing drug resistance

In canine lymphoma, drug resistance is the major factor hindering treatment. In this study, we performed immunohistochemical examination of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), which are considered as transporters related to multidrug resistance in three recurrent canine lymphomas. All cases were negative for both transporters before anticancer drug administration, but became positive after this administration. The expression was confirmed in capillary endothelial cells, such as in brain capillaries acting as the blood-brain barrier (BBB). It is suggested that both transporters expressed on capillary endothelial cells in lymphoma tissue may inhibit the spread of anticancer drugs into tumor tissues from blood, the same as the BBB. Therefore, capillary endothelial cells could act as a blood-tumor barrier, which might be involved in drug resistance in canine lymphoma.     

Immunohistochemical study on the delayed progression of epithelial apoptosis in follicle-associated epithelium of rat Peyer's patch

It is well known that some caspases in apoptosis is involved in determinant of terminal differentiation and maturation of various cells. Our previous study ultrastructurally clarified the differentiation into M cells from immature microvillous epithelial cells and the redifferentiation from M cells to microvillous epithelial cells in the follicle-associated epithelium (FAE) of rat Peyer's patch. In this study, the difference of epithelial apoptosis between the FAE of Peyer's patch and intestinal villi was immunohistochemically investigated in rat jejunoileum. As a result, cleaved caspase-3 was limited to several epithelial cells at the tip of FAE, whereas almost all of the epithelial cells were cleaved caspase-3 positive in intestinal villi. Cleaved caspase-9 was detected only in a few exfoliating or exfoliated epithelial cells of both FAE and intestinal villi. Nuclear DNA-fragmentation was detected only in several epithelial cells of the tip of FAE, while it was expressed from the middle regions in the intestinal villi. The DNase I expression of the epithelial cytoplasm was much weaker in FAE than in intestinal villi. Bcl-x expression was restricted in the apical cytoplasms of epithelial cells in the FAE, whereas it was restricted in whole cytoplasms in villous epithelial cells. These findings suggest that the progression of the apoptotic process in the epithelial cells of FAE is later than in the intestinal villi, so that the possibility of epithelial differentiation might be remained in the FAE, unlike in the intestinal villi.     

Comparison of dendritic cell-mediated immune responses among canine malignant cells

Dendritic cell (DC) vaccination is one of the most attractive immunotherapies for malignancies in dogs. To examine the differences in DC-mediated immune responses from different types of malignancies in dogs, we vaccinated dogs using autologous DCs pulsed with keyhole limpet hemocyanin (KLH) and cell lysate prepared from squamous cell carcinoma SCC2/88 (SCC-KLH-DC), histiocytic sarcoma CHS-5 (CHS-KLH-DC), or B cell leukemia GL-1 (GL-KLH-DC) in vitro. In vivo inductions of immune responses against these tumor cells were compared by the delayed-type hypersensitivity (DTH) skin test. The DTH response against SCC2/88 cells were observed in dogs vaccinated with autologous SCC-KLH-DC, while the response was undetectable against CHS-5 and GL-1 cells in dogs vaccinated with autologous CHS-KLH-DC and GL-KLH-DC. Skin biopsies taken from DTH challenge sites were then examined for immunohistochemistry, and recruitment of CD8 and CD4 T cells was detected at the site where SCC2/88 cells were inoculated in dogs vaccinated with SCC-KLH-DC. By contrast, neither CD8 nor CD4 T cell infiltration was found at the DTH challenge site in the dogs vaccinated with CHS-KLH-DC or GL-KLH-DC. These findings may reflect that the efficacy of immune induction by DC vaccination varies among tumor types and that immune responses could be inducible in squamous cell carcinoma. Our results encouraged further investigation of therapeutic vaccination for dogs with advanced squamous cell carcinoma in clinical trials.     

Development of a plant conveyance system using an AGV and a self-designed plant-handling device: A case study of DIY plant phenotyping

Plant phenotyping technology has been actively developed in recent years, but the introduction of these technologies into the field of agronomic research has not progressed as expected, in part due to the need for flexibility and low cost. “DIY” (Do It Yourself) methodologies are an efficient way to overcome such obstacles. Devices with modular functionality are critical to DIY experimentation, allowing researchers flexibility of design. In this study, we developed a plant conveyance system using a commercial AGV (Automated Guided Vehicle) as a case study of DIY plant phenotyping. The convey module consists of two devices, a running device and a plant-handling device. The running device was developed based on a commercial AGV Kit. The plant-handling device, plant stands, and pot attachments were originally designed and fabricated by us and our associates. Software was also developed for connecting the devices and operating the system. The run route was set with magnetic tape, which can be easily changed or rerouted. Our plant delivery system was developed with low cost and having high flexibility, as a unit that can contribute to others’ DIY’ plant research efforts as well as our own. It is expected that the developed devices will contribute to diverse phenotype observations of plants in the greenhouse as well as to other important functions in plant breeding and agricultural production.     

All-around 3D plant modeling system using multiple images and its composition

In this study, we developed an all-around 3D plant modeling system that operates using images and is capable of measuring plants non-destructively without any contact. During the fabrication of this device, we selected a method capable of performing 3D model reconstruction from multiple images. We then developed an improved SfM-MVS (Structure from Motion / Multi-View-Stereo) method that enables 3D reconstruction by simply capturing images with a camera. The resulting image-based method offers a high degree of freedom because the hardware and software can comprise commercially available products, and it permits the use of one or more cameras according to the shape and size of the plant. The advantages of the image-based method are that 3D reconstruction can be conducted at any time as long as the images are already taken, and that the desired locations can be observed, measured, and analyzed from 2D images and a 3D point cloud. The device we developed is capable of 3D measurements and modeling of plants from a few millimeters to 2.4 m of height using this method. This article explains this device, the principles of its composition, and the accuracy of the models obtained from it.