Comparison between immunohistochemistry and genetic clonality analysis for cellular lineage determination in feline lymphomas

Clinical application of genetic clonality analysis by PCR has been available in feline lymphoma. However, it is in under dispute whether the analysis can be applied for cellular lineage determination. To evaluate utility of the analysis for lineage determination, the results between immunohistochemistry and the genetic analysis were compared. Based on the result with immunohistochemistry, sensitivity of the primer for IgH was 89% and specificity was 75%. Sensitivity of the primer for TCRγ was 25% and specificity was 100%. Cross-lineage IgH rearrangement was found in 25% of T-cell lymphomas. The present study suggests that the genetic analysis had better be applied as not a definite but an adjunctive tool for lineage determination in feline lymphomas.     

Ultrastructural and histochemical properties of the olfactory system in the japanese jungle crow, Corvus macrorhynchos

Although it has been commonly believed that birds are more dependent on the vision and audition than the olfaction, recent studies indicate that the olfaction of birds is related to the reproductive, homing, and predatory behaviors. In an attempt to reveal the dependence on the olfactory system in crows, we examined the olfactory system of the Japanese jungle crow (Corvus macrorhynchos) by histological, ultrastructural, and lectin histochemical methods. The olfactory epithelium (OE) of the crow occupied remarkably a small area of the nasal cavity (NC) and had the histological and ultrastructural features like other birds. The olfactory bulb (OB) of the crow was remarkably small and did not possess the olfactory ventricle. The left and right halves of the OB were fused in many cases. In the lectin histochemistry, soybean agglutinin (SBA) and Vicia villosa agglutinin (VVA) stained a small number of the receptor cells (RCs) in the OE and the olfactory nerve layer (ONL) and glomerular layer (GL) on the dorsocaudal region of the OB. Phaseolus vulgaris agglutinin-E (PHA-E) stained several RCs in the OE and the ONL and GL on the ventral region of the OB. These results suggest that 1) the crow has less-developed olfactory system than other birds, and 2) the dedicated olfactory receptor cells project their axons to the specific regions of the OB in the crow.     

In-straw cryoprotectant dilution for bovine embryos vitrified using Cryotop

The aim of this study was to develop an in-straw dilution method suitable for 1-step bovine embryo transfer of vitrified embryos using the Cryotop vitrification-straw dilution (CVSD) method. The development of embryos vitrified using the CVSD method was compared with those of embryos cryopreserved using in-straw vitrification-dilution (ISVD) and conventional slow freezing, outside dilution of straw (SFODS) methods. In Experiment 1, in vitro-produced (IVP) embryos cryopreserved using the CVSD method were diluted, warmed and exposed to the dilution solution at various times. When vitrified IVP embryos were exposed to the dilution solution for 30 min after warming, the rates of embryos developing to the hatched blastocyst stage after 72 h of culture (62.0-72.5%) were significantly lower (P<0.05) than those of embryos exposed to the solution for 5 and 10 min (82.4-94.3%), irrespective of supplementation with 0.3 M sucrose in the dilution solution. In Experiment 2, the rate of embryos developing to the hatching blastocyst stage after 48 h of culture in IVP embryos cryopreserved using the SFODS method (75.0%) was significantly (P<0.05) lower than those of embryos cryopreserved using the CVSD and ISVD methods (93.2 and 97.3%, respectively). In Experiment 3, when in vivo-produced embryos that had been cryopreserved using the CVSD, ISVD and SFODS methods and fresh embryos were transferred to recipient animals, no significant differences were observed in the conception and delivery rates among groups. In Experiment 4, when IVP embryos derived from oocytes collected by ovum pick-up that had been cryopreserved using the CVSD and ISVD methods and fresh embryos were transferred to recipient animals, no significant differences were observed in the conception rates among groups. Our results indicate that this simplified regimen of warming and diluting Cryotop-vitrified embryos may enable 1-step bovine embryo transfer without the requirement of a microscope or other laboratory equipment.     

Establishment of cell line and in vivo mouse model of canine Langerhans cell histiocytosis

A cell line named FB‐LCH01, derived from a dog diagnosed with Langerhans cell histiocytosis (LCH), was established and characterized. FB‐LCH01 had C‐shaped nucleoli, characterized by modal chromosome aberrations. The original tumour cells as well as established FB‐LCH01 cells were immunopositive for human leukocyte antigen‐DR, Iba‐1 and E‐cadherin, and immunonegative for CD163 and CD204, suggesting Langerhans cell origin. Furthermore, the characteristics of FB‐LCH01 were compared with those of two canine histiocytic sarcoma cell lines (PWC‐HS01 and FCR‐HS02) established previously. Expression of E‐cadherin was detected only in FB‐LCH01, but not in PWC‐HS01 and FCR‐HS02. All (n = 9) the severe combined immunodeficiency mice inoculated with the FB‐LCH01 cells developed subcutaneous tumour masses after 3 weeks. Eight of nine mice also developed metastatic lesions in the lymph nodes (8/8; 100%), lung (5/8; 62.5%), stomach (5/8; 62.5%), heart (4/8; 50%), pancreas (4/8; 50%), kidney (3/8; 37.5%), skin (3/8; 37.5%) and bone marrow (1/8; 12.5%). Tumour cells were pleomorphic and round‐ to polygonal‐shaped with prominent anisocytosis and anisokaryosis. The xenotransplanted tumour cells maintained the immunohistochemical features of the original tumour with persistent E‐cadherin expression at injection site and some visceral organs. In conclusion, the established cell line as well as the mice xenotransplant model in this study reflect the nature of canine LCH and may serve as promising models for investigating the patho‐tumorigenesis and therapy of the disease.     

Synergistic Antifungal Activity of Chitinolytic Enzymes and Prodigiosin Produced by Biocontrol Bacterium, Serratia marcescens Strain B2 against Gray Mold Pathogen, Botrytis cinerea

Serratia marcescens strain B2 is an effective biocontrol agent against gray mold of cyclamen, caused by Botrytis cinerea Persoon. Strain B2 has strong antifungal activity against B. cinerea in vitro. The culture filtrate was found to contain different types of chitinolytic enzyme activity, including endochitinase and chitobiase activity. Four chitinolytic enzymes were detected among the extracellular proteins of strain B2. Two major enzymes, a 58-kDa endochitinase and a 98-kDa chitobiase, were purified by chromatography and electrofocusing, respectively. Both enzymes inhibited spore germination of B. cinerea. And a red pigment, prodigiosin, extracted and purified from the bacterial cells, also inhibited spore germination. When prodigiosin and the chitinolytic enzymes were applied in concert, a synergistic inhibitory effect was observed. S. marcescens strain B2 has multiple modes of action against the pathogen. Received 14 February 2001/ Accepted in revised form 31 May 2001     

Karyotypes of three shrew species (Soriculus nigrescens,Episoriculus caudatus and Episoriculus sacratus) from Nepal

The karyotypes of three shrew species (Mammalia, Soricomorpha, Soricidae) from Nepal were examined for the first time. Based on the karyotypes and overall differences in skull size, the current Episoriculus caudatus appeared to include two distinct species: the larger E. caudatus and the smaller Episoriculus sacratus. Episoriculus sacratus has three subspecies, E. s. soluensis in Nepal and Sikkim, E. s. umbrinus in Assam, Myanmar and the Yunnan Province of China, and E. s. sacratus in the Sichuan Province of China. Soriculus nigrescens had a diploid chromosome number (2n) and fundamental number (FN), including two X chromosomes, of 64 and 92, respectively, consisting of 11 metacentric or submetacentric, two subtelocentric and 18 acrocentric pairs of autosomes, a metacentric X and an acrocentric Y chromosome. Episoriculus sacratus soluensis had 2n = 74 and FN = 126, consisting of 12 metacentric or submetacentric, 13 subtelocentric and 11 acrocentric pairs of autosomes, a submetacentric X and an acrocentric Y chromosome. Episoriculus caudatus had 2n = 60 and FN = 118, consisting of 19 metacentric or submetacentric, nine subtelocentric and one acrocentric pair of autosomes, a subtelocentric X and an acrocentric Y chromosome. The karyotypes of these three species are characterized by their large 2n and FN values compared with other Soricidae.     

Lime-nitrogen application reduces N2O emission from a vegetable field with imperfectly-drained sandy clay-loam soil

Abstract

We studied the effect of lime-nitrogen (calcium cyanamide, CaCN₂) application on the emission of nitrous oxide (N₂O) from a vegetable field with imperfectly-drained sandy clay-loam soil. Lime-nitrogen acts as both a pesticide and a fertilizer. During the decomposition of lime-nitrogen in the soil, dicyandiamide (DCD), a nitrification inhibitor, is formed, and as a result lime-nitrogen application may mitigate N₂O emission from the soil. The study design consisted of three different nitrogen-application treatments in field plots with a randomized block design. The nitrogen application treatments were: CF (chemical fertilizer), LN (all nitrogen fertilizer applied as lime-nitrogen), and CFD (chemical fertilizer containing DCD). Soil nitrification activity was lower in the LN and CFD plots than in the CF plots, and nitrification was inhibited for a longer period in the LN plots than in the CFD plots. In the LN plots, N₂O emission was lower than those of other treatments from 20 to 40 days after fertilization, a period when large peaks of N₂O emission were observed after rainfall in the CF and CFD plots. Cumulative N₂O emission over 63 days in the CF plots (mean ± standard deviation: 30.2 ± 14.4 mg N₂O m^?2) and CFD plots (24.3 ± 10.8 mg N₂O m^?2) was significantly higher than that in the LN plots (10.7 ± 1.2 mg N₂O m^?2; P < 0.05). Our results suggested that lime-nitrogen application decreased N₂O emission by inhibiting both nitrification and denitrification.     

Nitrous oxide flux from komatsuna (Brassica rapa) vegetated soil: a comparison between biogas digested liquid and chemical fertilizer

Biogas production generates digested slurry as a by-product. It can be used as fertilizer especially after its conversion into digested liquid. A microcosm-based study was conducted in order to compare the effects of single application of digested liquid or chemical fertilizer on N₂O flux and crop yield of komatsuna vegetable. Analysis revealed that digested liquid-treated soils released almost equal cumulative N₂O (0.43 g?N m^?2) compared to chemical fertilizer (0.39 g?N m^?2). The uncropped soils treated with the digested liquid and chemical fertilizer released more N₂O compared to corresponding cropped soils. The N₂O emission factor and soil mineral N contents were similar for the digested liquid and chemical fertilizer-treated soils. Plant biomass in the first crop after digested liquid application was significantly higher (5.59 g plant^?1) than that after applied chemical fertilizer (4.78 g plant^?1); but there was no significant difference for the second crop. Nitrogen agronomic efficiency was improved by the digested liquid compared to chemical fertilizer. This study indicates that cumulative N₂O flux was similar after application of the digested liquid and chemical fertilizer, while the overall yield from both croppings was increased in the digested liquid-treated soil compared to chemical fertilizer-treated soil.     

Micro‐elevations in paddy fields affect the efficacy of mechanical weeding: Evaluation of weeding machines to control Monochoria vaginalis in herbicide‐free farming

Differences in local topography (micro‐elevation) within a paddy field that constitute a source of variability in agronomical indicators have not been considered thoroughly as a block factor in weed studies. This study investigated and evaluated the performance of weeding machines (weeders) in two herbicide‐free paddy fields that contained micro‐elevations. The plant density of Monochoria vaginalis, a typical and harmful paddy weed unless controlled with herbicides, was used as the indicator of the efficacy of the weeders. Among the three weeders that were tested, one suppressed M. vaginalis dramatically at low elevations and the others were less sensitive to micro‐elevation. For comparison across the fields, micro‐elevations at weed sampling locations were converted to the initial depth of water by using the records of hydrographs that had been set in each field. The relationship between the initial depth of water and the plant density of M. vaginalis was very clear with the use of the elevation‐sensitive weeder. Moreover, this relationship was valid, even with the less‐sensitive weeders. The finding that the greater the depth of water, the less the plant density was significant, even for M. vaginalis, a difficult aquatic paddy weed that was controlled with any of the weeders tested. Thus, micro‐elevation within a paddy field needs to be treated as a crucial block factor in weed‐sampling studies. A coarse survey of the level of a field and the installation of a hydrograph are recommended for a clear analysis of the background of weed control practices.     

A canine case with cystic meningioma showing miraculous reduction of the cystic lesion

A 12-year-old spayed female Labrador retriever was presented with forebrain signs. Brain MRI revealed a huge cystic lesion with the thickened falx in the frontal region. The brain parenchyma surrounding the lesion showed significant signs of a mass effect and also increased intracranial pressure. However, the dog suddenly became lucid after about two weeks, and an MRI scan one month after the initial study revealed a dramatically shrunken cystic lesion. The dog survived for over a year until it was euthanized for other reasons, and the brain lesion was diagnosed as a cystic meningioma histologically. To the authors’ knowledge, this is the first report that described the reduction of the cystic lesion of a cystic meningioma in dogs.