Development of a multiplex PCR and PCR-RFLP method for serotyping of Avibacterium paragallinarum

Avibacterium (Haemophilus) paragallinarum (A. paragallinarum) is a causative agent of infectious coryza in chickens and is classified into three serovars by agglutination tests. In an effort to identify the serovars easily, PCR and PCR-RFLP were employed. As the target gene for PCR, the hypervariable region of HMTp210, which encodes the HA antigen, was used. PCR using primer sets around the hypervariable region amplified 0.8, 1.1 and 1.6 kbp fragments for serovars A, B and C, respectively. Alternatively, the 1.6 kbp fragments were amplified with another primer pair encompassing the hypervariable region and was subjected to digestion with Bgl II, which resulted in the detection of serovar-specific digestion patterns. These results indicate that PCR and PCR-RFLP using the hypervariable region of HMTp210 are alternative methods to identify the serovar of A. paragallinarum.     

Germination stimulatory activity of bacterial butenolide hormones from Streptomyces albus J1074 on seeds of the root parasitic weed Orobanche minor

Damage caused by Orobanchaceae root parasitic weeds is a substantial agricultural problem for global food security. Many studies have been conducted to establish practical methods of control, but efforts are still required for successful management. Seed germination of root parasitic weeds requires host-derived germination stimulants including strigolactones (SLs). Studies on SLs have revealed that a butenolide ring is the essential moiety for SL activity as a germination stimulant. Interestingly, recent studies have revealed that butenolide hormones regulate the biosynthesis of secondary metabolites and mediate communication in actinomycete bacteria. Because of the structural similarity between SLs and the bacterial butenolides, we evaluated the germination stimulatory activity of butenolides isolated from Streptomyces albus J1074 on root parasitic weeds. These butenolides were found to specifically induce seed germination of Orobanche minor. Our findings contribute to understanding the molecular mechanisms of germination stimulant perception and to the development of a method for their biological control.     

Antioxidative activity of green tea treated with radical initiator 2, 2'-azobis(2-amidinopropane) dihydrochloride

This study investigated the antioxidative activity of green tea extract, and a green tea tannin mixture and its components, under conditions of radical generation using the hydrophilic azo compound, 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) to generate peroxyl radicals at a constant and measurable rate in the cultured renal epithelial cell line, LLC-PK(1), which is susceptible to oxidative damage. Treatment with AAPH decreased cell viability and increased the formation of thiobarbituric acid-reactive substances. However, green tea extract, and the tannin mixture and its components, comprising (-)-epigallocatechin 3-O-gallate (EGCg), (-)-gallocatechin 3-O-gallate (GCg), (-)-epicatechin 3-O-gallate (ECg), (-)-epigallocatechin (EGC), (+)-gallocatechin (GC), (-)-epicatechin (EC), and (+)-catechin (C), showed protective activity against AAPH-induced cellular damage. The tannin mixture and its components exhibited higher antioxidative activity than the green tea extract. Furthermore, EGCg and GCg had higher activity than EGC and GC, respectively. In particular, EGCg exerted the most significant cellular protective activity against AAPH. These results indicate that green tea tannin may inhibit cellular loss and lipid peroxidation resulting from the peroxyl radical generated by AAPH, and that the chemical structure of tannin is also involved in the activity, suggesting that the O-dihydroxy structure in the B ring and the galloyl groups are important determinants for radical scavenging and antioxidative potential.     

Comparison of threshold algorithms for automatic image processing of rice roots using freeware ImageJ

High-throughput phenotyping of crop root system architecture using digital image analysis requires costly special software (WinRHIZO). Novel freeware, ImageJ, has also been developed for the similar purpose, but the presetting of the threshold to create root binary images makes this method time-consuming. This study compares the 16 algorithms available in the ImageJ for processing of rice root images. Among the algorithms, the Triangle algorithm proved to be the best binary method, where the coefficient of correlation between ImageJ-estimated and WinRHIZO-estimated root lengths is extremely high (r = 0.986). However, using the Triangle algorithm, ImageJ overestimated the rice root length compared with WinRHIZO. By multiplying the values obtained using ImageJ by 2/3, the estimates closely corresponded to those estimated by WinRHIZO. The correspondence of root lengths estimated using WinRHIZO and ImageJ with Triangle algorithm was valid for roots of various morphologies, and for rice plants grown in uplands, rainfed lowlands, and irrigated lowlands. This report proposes the completely automated estimation of rice root length using freeware ImageJ with the appropriate threshold algorithm for image processing.     

Radiation use efficiency,N accumulation and biomass production of high-yielding rice in aerobic culture

The concept of aerobic culture is to save water resource while maintaining high productivity in irrigated rice ecosystem. This study compared nitrogen (N) accumulation and radiation use efficiency (RUE) in the biomass production of rice crops in aerobic and flooded cultures. The total water input was 800–1300 mm and 1500–3500 mm in aerobic culture and flooded culture, respectively, and four high-yielding rice cultivars were grown with a high rate of N application (180 kg N ha⁻¹) at two sites (Tokyo and Osaka) in Japan in 2007 and 2008. The aboveground biomass and N accumulation at maturity were significantly higher in aerobic culture (17.2–18.5 t ha⁻¹ and 194–233  kg N ha⁻¹, respectively) than in flooded culture (14.7–15.8 t ha⁻¹ and 142–173 kg N ha⁻¹) except in Tokyo in 2007, where the surface soil moisture content frequently declined. The crop maintained higher N uptake in aerobic culture than in flooded culture, because in aerobic culture there was a higher N accumulation rate in the reproductive stage. RUE in aerobic culture was comparable to, or higher than, that in flooded culture (1.27–1.50 g MJ⁻¹ vs. 1.20–1.37 g MJ⁻¹), except in Tokyo in 2007 (1.30 g MJ⁻¹ vs. 1.37 g MJ⁻¹). These results suggest that higher biomass production in aerobic culture was attributable to greater N accumulation, leading to higher N concentration (N%) than in flooded culture. Cultivar differences in response to water regimes were thought to reflect differences in mainly (1) early vigor and RUE under temporary declines in soil moisture in aerobic culture and (2) the ability to maintain high N% in flooded culture.     

Presence of a Temperature Gradient Among Genital Tract Portions and the Thermal Changes Within These Portions Over the Estrous Cycle in Beef Cows

The aim of the present study was to describe the temperature of the different portions of the female genital tract and their relation to rectal temperature and to investigate the effect of steroid hormones profiles on these variables over the estrous cycle in cattle. Four nonpregnant Japanese Black cows were investigated daily over two successive estrous cycles using a digital thermometer with a long probe and rounded-end sensor to record the temperature of the rectum (RT), vagina (VT), cervix (CT), uterine body (UBT) and uterine horns (UHT). Blood samples were collected immediately before temperature recording to assay peripheral levels of progesterone (P₄) and estradiol-17β (E₂). Moreover, transrectal ultrasonography was carried out after temperature recording to monitor the ovulatory follicle and track ovulation. During the experiment, the ambient temperature and relative humidity were recorded for further calculation of the temperature humidity index (THI). The temperature within the genital tracts in these cows progressively increased towards the uterine horns from the vagina. The VT, CT, UBT and UHTs were significantly higher in association with peripheral P₄ concentrations greater than 4 ng/ml (mid-luteal phase) when compared with lower peripheral P₄ concentrations. The VT was more significantly (P<0.01) correlated to the CT, UBT and UHTs than RT. In conclusion, a temperature gradient was present among the vagina, cervix and uterus over the estrous cycle, and changes in peripheral P₄ concentrations were associated with the thermal variations within these portions. The VT could be more beneficial than RT in monitoring temperature of deeper portions of the female genital tract in bovine.     

Quantitation of tocotrienol and tocopherol in various rice brans

Rice bran is abundant in bioactive compounds including tocotrienol (T3, unsaturated vitamin E). T3 has been reported about its potential functionalities (i.e., antiangiogenic effect), so much attention has been paid on usability of rice bran T3. Hence, we developed a rapid screening method for T3-rich rice bran by one-step equilibrium direct solvent extraction followed by normal phase high-performance liquid chromatography (HPLC). The method gave high-extraction rate of rice bran T3 and tocopherol (above 90%), and the determination of vitamin E by HPLC was completed within 15 min. Using the method, an average of total T3 content in 109 kinds of rice bran samples was 830 mug/g dry wt. Kouchi-Akamai, Joushuu, and Wataribune were found as the T3-rich rice bran varieties (1350-1430 microg T3/g dry wt). According to T3 ratio against total vitamin E (wt %), the average ratio was 61%. Hirayama, Moritawase, and Kaneko were found as the varieties having the highest T3 ratio (80-86%). Since T3 content in Koshihikari rice bran (the leading variety in Japan) was a little above the average, we cross-fertilized Koshihikari with T3-rich varieties and found that T3 content or ratio in F1 was improved compared with Koshihikari. The varieties found rich in T3 could be used for nutraceutical purpose.