Loop-mediated isothermal amplification for the rapid, sensitive, and specific detection of the O9 group of Salmonella in chickens

In the present study, the loop-mediated isothermal amplification (LAMP) assay was developed to amplify the fragments of the O9 Salmonella-specific insertion element and evaluated in the laboratory for its potential use in a field situation, such as poultry farms. Among the bacteria tested, a positive reaction was observed only for 128 strains of 6 serovars of the O9 group Salmonella, such as Enteritidis (SE) and Pullorum. The detection limit of the LAMP assay was 10(3)CFU/ml, which was more sensitive than that of the polymerase chain reaction (PCR) assay with the same target gene (10(6)CFU/ml). The final results were obtained within 30 min for the LAMP assay, while the PCR assay needed a total of 120 min. When the LAMP assay was applied to the enrichment broth mixed with cecal dropping samples either spiked with SE in vitro or excreted by SE-inoculated hens, the results were comparable to those of the conventional plating method including 2 separate enrichments. In conclusion, the LAMP assay developed in the present study is an effective method for the specific detection of the O9 group Salmonella serovars, including SE.     

Preparation of Photocrosslinked Fish Elastin Polypeptide/Microfibrillated Cellulose Composite Gels with Elastic Properties for Biomaterial Applications

Photocrosslinked hydrogels reinforced by microfibrillated cellulose (MFC) were prepared from a methacrylate-functionalized fish elastin polypeptide and MFC dispersed in dimethylsulfoxide (DMSO). First, a water-soluble elastin peptide with a molecular weight of ca. 500 g/mol from the fish bulbus arteriosus was polymerized by N,N′-dicyclohexylcarbodiimide (DCC), a condensation reagent, and then modified with 2-isocyanatoethyl methacrylate (MOI) to yield a photocrosslinkable fish elastin polypeptide. The product was dissolved in DMSO and irradiated with UV light in the presence of a radical photoinitiator. We obtained hydrogels successfully by substitution of DMSO with water. The composite gel with MFC was prepared by UV irradiation of the photocrosslinkable elastin polypeptide mixed with dispersed MFC in DMSO, followed by substitution of DMSO with water. The tensile test of the composite gels revealed that the addition of MFC improved the tensile properties, and the shape of the stress–strain curve of the composite gel became more similar to the typical shape of an elastic material with an increase of MFC content. The rheology measurement showed that the elastic modulus of the composite gel increased with an increase of MFC content. The cell proliferation test on the composite gel showed no toxicity.     

Assessment of environmental nitrogen pollution load potential from sewage treatment water in the Tedori River Alluvial Fan Area, Japan

An evaluation of the nitrogen pollution load potential (NPLP) arising from several sewage treatment systems in the Tedori River Alluvial Fan Area was evaluated by determining the removal percentage of nitrogen as a result of sewage treatment and amounts of nitrogen lost from the different treatment systems. These data were applied to the entire area resulting in an estimated total NPLP of 734 t year⁻¹ (10³ kg year⁻¹), of which 548 t year⁻¹ discharges into the Japan Sea and so does not contribute to the pollution of the environment considered here. The residual 186 t year⁻¹ of the NPLP has a high probability of causing groundwater or downstream pollution, with 58% of the NPLP being contributed from septic tanks not connected to public sewage systems. Rural sewage system accounted for about 18% of the NPLP. Material directly flowing into the middle stream of the Tedori River together with sewage water from small companies and untreated sewage water from local family dwellings made up about 3–12% of the NPLP. To improve water quality due to sewage effluent, it is essential to increase the number of connections to public sewage systems from septic tanks.     

Analysis of the nitrogen pollution load potential from farmland in the Tedori River Alluvial Fan Areas in Japan

A nitrogen balance study was carried out by setting up a test paddy in order to estimate the nitrogen pollution load potential (NPLP) from farmland in the Tedori River Alluvial Fan Areas and the load from the entire area under consideration was estimated using the cropping record and fertilizer application rate (FAR). The total NPLP was estimated to be 261 tons/year and the load from the paddy 79 tons/year for 5,704 ha, which would translate to an intensity of 14 kg/ha, while the load from the vegetable field was estimated to be 118 tons/year for 215 ha, which would equate to an intensity of 549 kg/ha. The pollution loads for the vegetables were significantly greater than those for the rice. The load for the barley was 57 tons/year for 261 ha (216 kg/ha) and that for the orchards was 23 tons/year for 64 ha (359 kg/ha). The estimation of soybean load was a negative 15 tons/year for 717 ha (−21 kg/ha), which meant that the nitrogen in the yield was greater than the FAR. The results also confirmed the yield absorption ratio in relation to the FAR. The load from the paddy for the entire area was also estimated using the percolation rate and the water quality load underneath the farmland was estimated to be 89 tons/year.     

Goblet cell hyperplasia and muscular layer thickening in the small intestine of a cynomolgus monkey

We report here the interesting case of a 5-year-old male cynomolgus monkey with goblet cell hyperplasia and thickening of the muscular layer throughout the small intestine without exhibiting any clinical symptoms. Necropsy examination showed diffuse thickening of the intestinal wall from the jejunum to the ileum, with an appearance likened to a rubber tube. Histopathologically, marked thickening was observed in both the mucosal and muscular layers in the jejunum and ileum, and slight thickening was observed in the duodenum. Goblet cell hyperplasia with extension of the circular folds and villi was prominently observed. The mucosal surface was covered with a thick mucus layer containing desquamated mucosal epithelial cells, and both the inner and outer muscular layers were markedly thickened due to smooth muscle hypertrophy. Neither macroscopic nor histopathological examination identified any causative factors, such as infection, enteritis and intestinal stenosis, or obstruction that may have caused development of this lesion. Given these observations, this case may simply be considered of spontaneous goblet cell hyperplasia and muscular layer thickening in the small intestine of a cynomolgus monkey.     

INTRAOPERATIVE SPINAL ULTRASONOGRAPHY IN DOGS: NORMAL FINDINGS AND CASE-HISTORY REPORTS

Intraoperative spinal ultrasonography was performed in cervical and lumbar spine of 2 and 5 normal dogs, respectively, following ventral slot technique or dorsal or hemilamenectomy. The dura was hyperechoic, while the parenchyma was hypoechoic. The subarachnoid space was anechoic. An echogenic line was present in the center of the spinal cord, as seen in human. Pulsation of the spinal cord was noted during M-mode imaging. Clinical findings of one dog with thoracolumbar disk herniation and one with thoracic vertebral fracture/subluxation confirmed the usefulness of intraoperative spinal ultrasonography for real time evaluation of spinal canal spatial abnormalities (mass lesion and degree of spinal cord compression on scanning planes) and spinal cord motion. Follow-up ultrasound examinations were possible from 6 days postoperatively.     

Presence of surfactant proteins in the uteri and placentae of pregnant mares

Immunohistochemical investigations of the expression of surfactant protein A (SP-A) and surfactant protein D (SP-D) in the uterine and placental tissues of 13 pregnant mares were performed using anti-horse monoclonal primary antibodies. Strong positive reactions for both SP-A and SP-D were observed in the trophoblasts in the microcotyledons of the placentae at 182 to 314 days of gestation; in uterine glandular epithelial cells, faint-to-weak reactions were observed during gestation. This study describes, for the first time, the changes in the SP-A and SP-D expression levels in the endometrium of mares during gestation; the SP-A and SP-D expression levels increased after the second trimester of gestation.     

Juvenile bird acanthocephalans recovered incidentally from raccoon dogs (Nyctereutes procyonoides viverrinus) on Yakushima Island, Japan

Helminth parasites were collected from 14 raccoon dogs (Nyctereutes procyonoides viverrinus) and 2 Japanese weasels (Mustela itatsi sho) on Yakushima Island, Kagoshima Prefecture (Kyushu), Japan. In addition to trematodes and nematodes reported separately, everted cystacanths of 3 bird acanthocephalans (Mediorhynchus robustus in 5 animals, Porrorchis leibyi in 2 animals, and Sphaerirostris lanceoides in 1 animal) were collected from the raccoon dogs introduced recently on this World Natural Heritage Area. A cystacanth of P. leibyi was collected also from a weasel indigenous to this island that were killed by traffic accidents. Recovery of everted cystacanths of M. robustus from the intestine of raccoon dogs was remarkably frequent (36%), suggesting that this acanthocephalan should be highly prevalent in passerine hosts on Yakushima Island, and the animals might vigorously take insect hosts or paratenic hosts. Furthermore, the present report is the second on M. robustus after its first record in Japan (Aichi Prefecture, Honshu) as M. garruli from a Eurasian jay (Garrulus glandarius japonicus) by Yamaguti in 1939. Porrorchis leibyi has never been recorded in Japan.