Application of ELISA-based kit for detecting AOZ and determining its clearance in eel tissues

Furazolidone, an antibacterial drug that was once widely used in the livestock industry and aquaculture, is now prohibited in numerous countries. It is difficult to detect residual furazolidone because it is readily metabolized in animal tissues but, by using and liquid chromatography coupled with tandem mass spectrometry, its metabolite, 3-amino-2-oxazolidinone (AOZ) can be detected. Here we describe the validity of an enzyme-linked immunosorbent assay (ELISA) kit to detect AOZ in Japanese eel Anguilla japonica tissue. ELISA is capable of detecting AOZ at 1.0 μg/kg in an eel sample with excellent accuracy and precision. Our results show that ELISA is suitable for regulatory purposes and for studying the fate of AOZ residues in eel treated with furazolidone. To measure the persistence of AOZ in eel tissues, eels (1.4–6.5g) were immersed in tanks containing 2 and 10 mg furazolidone/L for 3 h, and then maintained in a tank supplying well water for the next 160 days. The half-lives of AOZ, calculated from the linear terminal part of the excretion curve, were 25.0 days in muscle and 21.6 days in liver from fish exposed to 2 mg/L furazolidone. In the eels treated with 10 mg/L furazolidone, by contrast, high levels of AOZ were detected in liver and muscle, but the half-lives of AOZ were similar to those in fish treated with 2 mg/L furazolidone. The half-lives of AOZ in eel tissues were prolonged by the condition of low water temperature.     

Quality evaluation and prediction of Citrullus lanatus by 1H NMR-based metabolomics and multivariate analysis

(1)H NMR spectrometry in combination with multivariate analysis was considered to provide greater information on quality assessment over an ordinary sensory testing method due to its high reliability and high accuracy. The sensory quality evaluation of watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) was carried out by means of (1)H NMR-based metabolomics. Multivariate analyses by partial least-squares projections to latent structures-discrimination analysis (PLS-DA) and PLS-regression offered extensive information for quality differentiation and quality evaluation, respectively. The impact of watermelon and rootstock cultivars on the sensory qualities of watermelon was determined on the basis of (1)H NMR metabolic fingerprinting and profiling. The significant metabolites contributing to the discrimination were also identified. A multivariate calibration model was successfully constructed by PLS-regression with extremely high reliability and accuracy. Thus, (1)H NMR-based metabolomics with multivariate analysis was considered to be one of the most suitable complementary techniques that could be applied to assess and predict the sensory quality of watermelons and other horticultural plants.     

Behavior of four broad-leaved tree species used to revegetate eroded granite hill slopes

Information on primary growth behavior after planting is required for mixed-plantation revegetation using broad-leaved species. To estimate primary growth, especially from the perspective of crown coverage and changing growth rates, we examined the growth and survival of four broad-leaved species that are frequently used in erosion-control plantations in Japan. The species studied were Myrica rubra Sieb. et Zucc., Alnus pendula Matsum., Quercus glauca Thunb., and Q. serrata Thunb. The survival, height, and basal diameter of planted trees were measured over a 4-year period, and crown area was calculated over a 3-year period. We found a negative relationship between relative growth rate (RGR) and survival rate, suggesting that fast growth may be fatal when resources are severely limited. The relative height growth rate (RHGR) of A. pendula was especially high during the early period of the study (1997–1999) and then drastically declined, whereas the opposite tendency was observed in Q. glauca. The results of stem allometry analyses conformed to the specific relationships between height growth and diameter growth of the four species; increases in stem thickness based on height increments were smaller in the pioneer species A. pendula. Between-species differences in coverage per planted tree (mean crown area multiplied by survival rate) were small as a result of the negative relationship between coverage area and survival rate.     

Two-decadal trends in aboveground litterfall and net primary production in self-thinning Pinus banksiana stands in Wood Buffalo National Park,NWT, Canada

We estimated the aboveground net primary production (ANPP) in five self-thinning jack pine (Pinus banksiana) stands in Wood Buffalo National Park, NWT, Canada. The stands (11 to ca.175 years old) were selected to examine the relationship between stand density and tree size and its effect on carbon dynamics. Aboveground litterfall was collected from each stand from 1997 to 2012. Stand biomass was estimated by measuring tree size every 5 years and estimating the individual mass using allometric relationships. ANPP was then estimated by summing the litterfall mass, dead stem mass increment and stand biomass increment. We determined the proportional contribution of each organ to the total litter and the seasonal pattern of needle litterfall. There was a lower turnover rate of aboveground biomass in older stands than younger stands. The ANPP increased in the youngest stand (<30 years old) showed a decreasing trend in stands >50 years old. The maximum ANPP was estimated to be ca. 500?g m^?2 year^?1 in dry matter, which was found in 30–50 year-old stands.     

Effects of heat stress on the endometrial epidermal growth factor profile and fertility in dairy cows

The endometrial epidermal growth factor (EGF) profile is an indicator of uterine function and fertility in cattle. The present study aimed to investigate the effects of heat stress on the endometrial EGF profile and fertility in lactating Holstein cows. The endometrial EGF profiles of 365 cows in the Hokkaido and Kyushu regions were examined between June and September (heat stress period, n = 211) and between October and January (control period, n = 154). EGF profiles were investigated using uterine endometrial tissues obtained by biopsy 3 days after estrus (Day 3). The proportion of cows with an altered EGF profile was higher between June and September than between October and January (41.2 vs. 16.2%, P < 0.05). The effects of rectal temperature on Days 0 and 3 on the endometrial EGF profile were also assessed in cows (n = 79) between June and September in the Kyushu region. A single embryo was transferred to cow on Day 7 to evaluate fertility (n = 67). Regardless of the rectal temperature on Day 3, the proportion of cows with an altered EGF profile was higher (64.1 vs. 30.0%, P < 0.05) and the pregnancy rate after embryo transfer (ET) was lower (26.7 vs. 51.4%, P < 0.05) in cows with a rectal temperature ≥ 39.5°C on Day 0 than in cows with a rectal temperature < 39.5°C on Day 0. The present results indicate that alterations in the endometrial EGF profile induced by an elevated body temperature on Day 0 contributed to reductions in fertility in lactating dairy cows during the heat stress period.     

Development of X-ray computed tomography for live standing cattle

An X‐ray computed tomography (CT) system for live standing cattle was developed for studying the meat yield, carcass composition and so on. The gantry contained three X‐ray tubes and detectors that corresponded to each X‐ray tube. The system was able to operate while the animal remained standing. The scan area had a diameter of 900 mm. The Musculus longissimus and Musculus trapezius areas, and the back fat thickness in the CT image were evaluated and compared to the actual cross‐section of the carcass using eight cattle. The differences among the muscles, and the subcutaneous and intermuscular fat were easily recognized. The correlation coefficient between the CT image and the actual carcass photograph of the M. longissimus area and the back fat thickness was high (r = 0.84, r = 0.93, P < 0.01). The present study demonstrated that muscle, fat and bone can be clearly imaged from a live standing animal using X‐ray equipment.     

Concentrations and specific antibodies to staphylococcal enterotoxin-C and toxic shock syndrome toxin-1 in bovine mammary gland secretions,and inflammatory response to the intramammary inoculation of these toxins

To investigate the pathological role of staphylococcal enterotoxins (SEs) and toxic shock syndrome toxin-1 (TSST-1) in bovine mastitis, the production of SEs and TSST-1 was investigated in staphylococci isolated from 120 mammary gland secretions (MGS, 51 from no clinical sign-mammary glands and 69 from staphylococcal mastitic-mammary glands) collected from dairy farms where staphylococcal mastitis frequently occurred in Miyagi and Yamagata prefectures from 1997 to 1998. Concentrations of these toxins and specific antibody titers in each MGS were also measured. Furthermore, SEC and TSST-1 were inoculated into lactating mammary glands and inflammatory responses were analyzed. A high percentage of staphylococci including Staphylococcus aureus and coagulase-negative staphylococci isolated from both no clinical sign- and mastitic-MGS produced both SEC and/or TSST-1. The concentration of SEC increased with the severity of the mastitis, and was significantly higher (P<0.05) in acute mastitic-than in no clinical signs-MGS. Titers of specific antibodies to TSST-1 in MGS were significantly higher (P<0.05) than those to SEC, regardless of whether or not the cows were lactating or mastitic. Specific antibodies purified from MGS neutralized each toxin in vitro. A significant increase (P < 0.05) in somatic cell counts was induced by the intramammary inoculation of SEC but not TSST-1. These findings indicated that SEC rather than TSST-1 plays an important role in the pathology of staphylococcal bovine mastitis. The inflammatory activity of TSST-1 was probably neutralized by specific antibodies in MGS.     

Localization of inhibin alpha-, betaA- and betaB-subunits and aromatase in ovarian follicles with granulosa theca cell tumor (GTCT) in 6 mares

To clarify the morphological and immunohistochemical characteristics in mares with granulosa theca cell tumor (GTCT), the localization of inhibin subunits (alpha, betaA, betaB) and aromatase in the granulosa cell layers and theca layers in the ovarian follicles were determined by immunohistochemical staining. The follicles were obtained from the ovaries of 6 mares with GTCT and 4 normal mares as controls. Immunohistochemically, inhibin alpha-subunit was localized in the granulosa cells of all follicles showing different sizes in all GTCT cases and betaA- subunit was localized in two GTCT cases in all sized follicles. But inhibin betaB- subunit and aromatase were not localized in GTCT cases. On the other hand, inhibin alpha-, betaA-, and betaB-subunits and aromatase were localized in the large and medium sized follicles, but inhibin betaA- and betaB-subunits and aromatase were not stained in the small sized follicles in normal cases. These findings suggest that some mares with GTCT can secrete dimeric inhibin (inhibin A), but all GTCT cases cannot secrete inhibin B. By the results of aromatase staining it is clear that testosterone is not converted into estradiol due to the lack of aromatase in the GTCT follicles.