Identification of the medicinal off-flavor compound formed from ascorbic acid and (E)-hex-2-enal

A test apple beverage made up of apple juice (20%), high-fructose corn syrup (11.5%), citric acid (0.43%), trisodium citrate (0.02%), apple-odor flavor (0.1%), and ascorbic acid (0.02%) was stored at 40 °C and then analyzed for the change of odor in the beverage. Although no thermoacidophilic bacteria (TAB) were detected, a medicinal off-flavor was perceived after the 8 weeks of storage. Model experiments on the ingredients of the test apple beverage revealed that the off-flavor compound had been formed by ascorbic acid and (E)-hex-2-enal. Synthesis and NMR (1H, 13C, HMQC, and HMBC) analyses identified the compound as 6-propylbenzofuran-7-ol. The odor quality, retention index (RI), and mass spectrum of synthetic 6-propylbenzofuran-7-ol were identical with those of the medicinal odor compound from the test apple beverage. Sensory evaluation revealed the recognition thresholds for medicinal odor were 31.4 ppb in water and 24.0 ppb in apple beverage, and the detection thresholds were 19.6 ppb in water and 8.6 ppb in apple beverage, respectively. The quantified concentration of 6-propylbenzofuran-7-ol formed in test apple beverage was 90 ppb, approximately. This concentration was well above the odor threshold, so it was concluded that the compound was the source of the medicinal off-flavor.     

Change of plasma insulin-like growth factor-1 (IGF-1) concentration with early growth in Japanese beef cattle

The aim of the present study was to examine the change in plasma insulin‐like growth factor‐1 (IGF‐1) concentration with early growth, changes of bodyweight (BW) and relative dairy gain (RDG) in the pre‐ (PRW) and postweaning periods (POW) in Japanese beef cattle, and relationships with metabolites. A total of 33 calves, 22 Japanese black, 6 Japanese shorthorn and 5 of their crossbreed were studied. Insulin‐like growth factor‐1 and metabolite (glucose, triacylglycerol, nonesterified fatty acid) levels in the plasma, from jugular vein blood taken every month, were measured along with BW. Insulin‐like growth factor‐1 in POW increased dramatically with increase of BW (P < 0.05), and the correlation was positive at 0.52 (P < 0.01). Glucose levels correlated significantly with BW, RDG and IGF‐1 (P < 0.01). Metabolic required calorie correlated positively with IGF‐1 (P < 0.01). Also, correlations of BW in POW, with BW and RDG in PRW were positive (P < 0.01). Growth in PRW would be influenced by maternal effects, while active self‐secretion of IGF‐1 in POW might contribute to POW growth. These factors suggested that to increase growth in PRW, maintaining enough maternal effect and IGF‐1 level in POW, was important for establishing better growth after weaning.     

Safety and efficacy of the cell-associated vaccine prepared by an attenuated infectious laryngotracheitis virus.

The safety and efficacy of the cell-associated (C-A) vaccine prepared by chicken embryo fibroblast (CEF) cells infected with the tissue-culture-modified strain of infectious laryngotracheitis (ILT) virus were studied in chickens. Over seventy percent of chickens inoculated with the C-A vaccine by the subcutaneous (S.C.) or intramuscular (I.M.) route at 1 day of age was protected against challenge with a virulent strain of ILT virus without any clinical signs. Chickens vaccinated with the C-A vaccine at 1 day of age acquired immunity within 6 days after vaccination, and the protection rate maintained more than 60% until 10 weeks post-vaccination. The C-A vaccine was invariably effective for chickens at various age. There was no evidence that the development of immunity was hindered by further vaccination with Newcastle disease and infectious bronchitis combined live vaccine. In addition, the C-A vaccine was safe when chickens were inoculated with 10 doses. In the field trials of the C-A vaccine, no adverse reaction was observed, and over 65% of vaccinated chickens was protected against the challenge of the virulent ILT virus at 8 weeks after vaccination.     

Precise, simple screening for resistance in potato varieties to common scab using paper pots

Resistance to common scab pathogen Streptomyces turgidiscabies of seven potato varieties was compared in the field with a newly developed paper pot method. Seedlings raised in soil in paper pots containing inocula at 1 × 10³ to 10⁷cfu/g soil were transplanted into a scab-free field and grown for 3 months. The disease severity of the seven varieties in the field trials differed in iteration and from year to year, even though their resistance levels were approximately similar at the expected levels. With the paper pot method, the seven varieties had different resistance levels, which were almost completely consistent with the results of the field trials, at more than 1 × 10⁵cfu/g soil. Significant differences in disease severity between resistant and susceptible varieties were observed (P = 0.05) for 2 years, and the resistance level of the varieties was elucidated.     

Isolation and characterization of staphylococci from external auditory meatus of dogs with or without otitis externa with special reference to Staphylococcus schleiferi subsp. coagulans isolates

Staphylococci were isolated from the external auditory meatus in 14 (48.3%) of 29 dogs affected with otitis externa (OE dogs) and 28 (68.3%) of 41 dogs without OE (non-OE dogs). Twenty-two OE isolates were identified as belonging to 12 species, and 42 non-OE isolates were identified as belonging to 13 species. The predominant species found in both OE and non-OE isolates were S. intermedius, and S. epidermidis. Thirty-eight (59.4%) of 64 isolates were resistant to one or more of the 17 antimicrobial agents tested. Resistance to PCG and ABPC was most frequent. S. schleiferi subsp. coagulans, a recent etiologic agent of canine OE, was isolated from OE and non-OE dogs. All of the 5 S. schleiferi subsp. coagulans isolates showed typical characteristics. No clear difference in the extracellular enzyme or toxin profiles, nor in the PFGE patterns, was demonstrated between the OE and non-OE isolates of S. schleiferi subsp. coagulans. A new PCR primer set specific for 16S rDNA was designed to identify strains of S. schleiferi subsp. coagulans. The amplified fragment was detected in all of the 5 isolates as well as the type strain GA 211 (=JCM 7470) and a reference strain GA 11, but was not detected in any strains of the related species, S. aureus, S. intermedius and S. hyicus. The PCR may allow a simple, rapid and precise identification of S. schleiferi subsp. coagulans, in addition to the standard tube test for free coagulase.     

Fine surface structure of bovine acrosome-intact and reacted spermatozoa observed by atomic force microscopy

The atomic force microscope (AFM) provides nanometer resolution, topographic data of the natural surface structure of materials. We studied the topology of the surface structure of bovine sperm heads during the acrosome reaction by AFM. In addition, we numerically analyzed the areas of the median sagittal plane of the sperm heads. Bovine frozen-thawed spermatozoa were washed, capacitated by heparin, and incubated with lysophosphatidylcholine (LPC) to induce the acrosome reaction, smeared on a cover glass, air-dried, and observed with AFM using the dynamic force (tapping) mode. AFM analysis of spermatozoa showed the clear surface structure of acrosomes, equatorial segments, postacrosomal regions and necks. Although AFM images of spermatozoa capacitated by heparin had complete acrosomes, most spermatozoa treated with LPC had no acrosomal caps as shown by AFM. These observations coincided with those obtained by light microscopy after staining with naphthol yellow S and erythrosin B. Furthermore, numerical analysis of AFM images indicated that areas of the median sagittal plane of the anterior portions of acrosome-reacted sperm heads (2679 +/- 616 pixels) were approximately 40% less than those of intact heads (4535 +/- 174 pixels, P<0.05). These results indicate that AFM can usefully observe and numerically analyze the fine surface structures of bovine spermatozoa.     

Magnetic resonance imaging of alveolar echinococcosis experimentally induced in the rat lung

Pulmonary alveolar echinococcosis (AE) caused by the metacestode of Echinococcus multilocularis is a lethal zoonosis and is a lesion secondarily induced by hematogenous dissemination from hepatic AE lesions. In the present study, a hematogenous pulmonary AE model was experimentally induced in rats by the injection of echinococcal larval tissue homogenate to the tail vein, and then the pathological and diagnostic aspects of pulmonary AE were examined by magnetic resonance imaging (MRI). Histological primary, mature and degenerated AE lesions were observed 5, 18 and 50 weeks after injection, respectively. These lesions were discriminated as signal-void, hypointense and hyperintense regions in T1-weighted MRI (T1WI), respectively. The change in signal intensity in T1WI might reflect the content of proteinaceous fluid as a result of AE cyst degeneration. Western blot analysis of sera with antibodies of two epitopes (Em18 and Em16) of E. multilocularis provided evidence for AE infection in the early stage. T1WI in combination with Western blot analysis could possibility become definitive and early signs of hematogenous pulmonary AE infection.     

Histopathological characteristics of Ito cells and Kupffer cells in the feline liver

The histopathological characteristics of Ito cells and Kupffer cells were investigated in the liver of 21 cats (age range: 6 months -18 years) autopsied in our laboratory during 2003. Immunohistochemical examinations were performed using antibodies against lysozyme, desmin and alpha-smooth muscle actin. No Kupffer cells reacted with the antibody against lysozyme. However, macrophages in the lung and spleen showed a positive reaction with the antibody. This finding suggests a possibility that the amount of lysozyme in the Kupffer cells of feline liver is comparatively small. On the other hand, large vacuole-laden cells were observed in the hepatic perisinusoid of some feline cases, and these cells showed a positive reaction with antibodies against desmin and alpha-smooth muscle actin. These cells could be Ito cells with large lipid vacuoles. This conclusion was supported by electron microscopic observation and oil red O staining. However, no such large vacuole-laden perisinusoidal cells were detected in the liver of young cats less than 2 years old. The present study revealed the histopathological features of Kupffer cells and Ito cells in the feline liver.     

Expression and subcellular localization of GSE protein in germ cells and preimplantation embryos

We previously identified a novel gonad-specific expression gene (Gse) and investigated its expression during gametogenesis in the mouse testis and ovary. In this study, we generated a polyclonal antibody to GSE protein and determined the profiles of the protein's expression in germ cells and preimplantation embryos in detail using immunocytochemical and immunofluorescence staining. In a Western blot analysis, the anti-GSE antibody recognized long and short isoforms (approximately 27.6 kDa and 23.1 kDa) of the protein in the mouse testis and the long isoform in the ovary. In the mouse testis, GSE protein was expressed in spermatocytes I in the pachytene stage, round spermatids, and elongated spermatids. In the mouse ovary, the protein was located in the cytoplasm and nucleus of all oocytes regardless of the stage of the ovarian follicles. In preimplantation embryos from the pronuclear to blastocyst stage, however, GSE protein was mainly detected in the nuclei of cells. At the blastocyst stage, the protein was confirmed to have accumulated in the inner cell mass (ICM), whereas it had mostly disappeared from the trophectoderm (TE). These findings suggest that GSE protein may play a role in the establishment of nuclear totipotency and may be associated with early lineage specification.     

A cytolethal distending toxin gene-based multiplex PCR assay for detection of Campylobacter spp. in stool specimens and comparison with culture method

In this study, we evaluated the applicability of cytolethal distending toxin (cdt) gene-based species-specific multiplex PCR for the direct detection and identification of Campylobacter jejuni, C. coli and C. fetus from stool specimens of patients with gastroenteritis in comparison to culture methods. A total of 711 stool specimens were examined for the isolation or detection of campylobacters by using Skirrow's selective agar culture plates, a filtration method and the multiplex PCR assay. Forty-one and 36 C. jejuni strains were isolated by culture and filtration methods, respectively. In addition, 2 and 3 C. coli strains were isolated by Skirrow and the filtration methods, respectively. However, when the multiplex PCR was employed, the cdtB genes of C. jejuni and C. coli were detected in 45 and 4 stool samples, respectively, and 9 C. jejuni PCR-positive samples by multiplex PCR were negative by culture method. Sequence analysis of the PCR products obtained from 8 stool specimens from which campylobacters were not isolated by culture method but the sequences exactly matched with that of the cdtB gene of C. jejuni strain 81-176. None of the remaining stool samples which were culture negative for campylobacters produced any amplicon. Stool samples were defined as Campylobacter-positive if detected by any method. The sensitivity of the multiplex PCR was 83%, which was higher than Skirrow (74%) and filtration method (66%). These data indicate that cdtB gene-based multiplex PCR is a rapid and more sensitive method to identify the most important species of Campylobacter for human diseases. (248).