Effects of cooling rate on post-thaw motility and fertility of Japanese pearl oyster <Emphasis Type="Italic">Pinctada fucata martensii</Emphasis> spermatozoa

The post-thaw motility and fertility of Japanese pearl oyster sperm show large variances, even among sperm samples obtained from the same individuals. This study aimed to clarify the factors that cause such differences. Spermatozoa were diluted 50 times with diluent comprising 10 % methanol, 18 % fetal bovine serum, and 72 % seawater, and dispensed into 0.25 ml straws. A total of 59 straws were cooled, one by one, at 11 different heights from the surface of liquid nitrogen (LN) to −50 °C, and then immediately immersed in LN. After thawing the straws, the relationships between the cooling rate and the post-thaw motility and post-thaw fertility of the spermatozoa were examined. Both the post-thaw motility and the post-thaw fertility showed a sharp peak when the straws were cooled at around −20 °C/min. There was a strong correlation between post-thaw motility and fertility (P < 0.001). There was a large difference in the cooling rates and the post-thaw motilities and fertilities of the spermatozoa, even between straws cooled at the same height. These results indicate that the optimum range for the cooling rate of oyster spermatozoa is quite narrow, and the method of cooling straws at a fixed distance from the LN surface is unsuitable for the cryopreservation of Japanese pearl oyster spermatozoa.     

Secretome analysis of the basidiomycete <Emphasis Type="Italic">Phanerochaete chrysosporium</Emphasis> grown on ammonia-treated lignocellulosic biomass from birch wood

Ammonia pretreatment is a promising technique for enhancing enzymatic saccharification of lignocellulosic biomass. However, an enzymatic cocktail suitable for the breakdown of pretreated biomass samples is still being developed. The basidiomycete Phanerochaete chrysosporium is a well-studied fungus with regard to bioconversion of lignocellulosic biomass. In the present work, we analyzed proteins secreted by P. chrysosporium grown on untreated and ammonia-treated birch wood meal. Fungal growth, xylanase activity, and extracellular protease activity increased in the media containing the ammonia-treated biomass; however, cellulase production decreased compared to that observed in the untreated biomass. Secreted extracellular proteins were separated by two-dimensional electrophoresis and identified by liquid chromatography ion–trap mass spectrometry. Fifty-five spots corresponding to secreted proteins were chosen for further analysis. In the culture with ammonia-treated biomass, the relative concentration of a xylanase belonging to glycoside hydrolase (GH) family 11 increased, while acetyl xylan esterases belonging to carbohydrate esterase family 1 decreased. Moreover, GH family 10 xylanases were promoted proteolysis in the culture of ammonia-treated biomass, leading to the loss of family 1 carbohydrate-binding modules. These results indicated that P. chrysosporium produced enzymes related to the recognition of structural changes on xylan with de-acetylation and introduction of nitrogen by ammonia pretreatment of birch wood meal.     

The quality after culture in vitro or in vivo of porcine oocytes matured and fertilized in vitro and their ability to develop to term

The quality of porcine blastocysts produced in vitro is poor in comparison with those that develop in vivo. We examined the quality of in vitro‐matured and fertilized (IVM/IVF) oocytes, their abilities to develop to blastocysts under in vivo and in vitro conditions, and the potential of the embryos to develop to term after transfer. IVM/IVF oocytes were either transferred and the embryos recovered on Days 5 and 6 (100% and 87.5%, respectively) (‘ET‐vivo’ embryos), or cultured in vitro for 5 or 6 days (‘IVC’ embryos). The proportion of blastocysts differed significantly between the two groups on Day 5 (20.6% and 8.0%, respectively), but not on Day 6 (23.8% and 21.2%, respectively). The mean number of cells in ET‐vivo blastocysts on Days 5 or 6 was significantly higher (72.8 and 78.7, respectively) than that in IVC blastocysts (22.1 and 39.7, respectively). When IVM/IVF oocytes and IVC blastocysts on Day 6 were transferred, all (three and three, respectively) developed to piglets (16 and 16, respectively), without any difference in the rates of development to term (2.1% and 2.6%, respectively). These data suggest that, although blastocyst production differs between the two culture conditions, IVM/IVF oocytes possess the same ability to develop to term.     

Land cover change detection based on satellite data for an arid area to the south of Aksu in Taklimakan desert

An experiment is made to detect the land-cover change in the area located to the south of Aksu in the northern Taklimakan desert through analyses of satellite data pixel by pixel basis. The analyzed data are those observed in the late summer and early autumn of 1973, 1977, 1993 and 1995. As a parameter of land-cover, SAVI (Soil Adjusted Vegetation Index) derived from the data of Landsat MSS and JERS-1 OPS (Optical Sensor) is used. The result indicates the increase of vegetation in the oasis areas, confluent area of the Yarkant and Kashgar Rivers and around reservoirs while little change occurs in the desert area. The 1973 satellite image shows the abundant flow in the Yarkant River while the river is almost dried up in the satellite images of later years. The trend of the decrease in the Hotan River flow is recognized although not so dramatic as that of the Yarkant River.     

Effect of sampling size on the determination of accurate pesticide residue levels in Japanese agricultural commodities

The uncertainty in pesticide residue levels (UPRL) associated with sampling size was estimated using individual acetamiprid and cypermethrin residue data from preharvested apple, broccoli, cabbage, grape, and sweet pepper samples. The relative standard deviation from the mean of each sampling size (n = 2(x), where x = 1-6) of randomly selected samples was defined as the UPRL for each sampling size. The estimated UPRLs, which were calculated on the basis of the regulatory sampling size recommended by the OECD Guidelines on Crop Field Trials (weights from 1 to 5 kg, and commodity unit numbers from 12 to 24), ranged from 2.1% for cypermethrin in sweet peppers to 14.6% for cypermethrin in cabbage samples. The percentages of commodity exceeding the maximum residue limits (MRLs) specified by the Japanese Food Sanitation Law may be predicted from the equation derived from this study, which was based on samples of various size ranges with mean residue levels below the MRL. The estimated UPRLs have confirmed that sufficient sampling weight and numbers are required for analysis and/or re-examination of subsamples to provide accurate values of pesticide residue levels for the enforcement of MRLs. The equation derived from the present study would aid the estimation of more accurate residue levels even from small sampling sizes.     

Molecular detection of Anaplasma phagocytophilum in cattle and Ixodes persulcatus ticks

The tick-borne pathogen, Anaplasma phagocytophilum (A. phagocytophilum), the causative agent of human granulocytic anaplasmosis (HGA), is increasingly becoming a public health concern as an aetiological agent for emerging infectious disease. We found A. phagocytophilum infection in a pooled sample of field-collected Ixodes persulcatus (I. persulcatus) ticks from one district in Hokkaido, Japan. Thus, to further investigate the prevalence in field-collected ticks, we used PCR assays targeting the A. phagocytophilum gene encoding 44 kDa major outer membrane protein (p44) for screening of I. persulcatus ticks and samples from cattle from pastures. Out of the 281 I. persulcatus ticks, 20 (7.1%) were found to harbor A. phagocytophilum DNA. The infection rate for A. phagocytophilum in cattle was 3.4% (42/1251). In future studies, it will be necessary to investigate effects of the infection in order to understand its pathogenesis of A. phagocytophilum in domestic animals.     

Symbiotic efficiency and compatibility of native rhizobia in northern Thailand with different soybean cultivars

Symbiotic efficiency and compatibility of 81 isolates of native bradyrhizobia from irrigated areas in northern Thailand with four soybean cultivars and one cowpea cultivar were evaluated under laboratory conditions. Effectiveness and / or compatibility of the tested isolates were compared with those of a standard strain (Bradyrhizobium japonicum CB 1809) by using plants grown on plastic seed bags. Effectiveness of the isolates was also estimated using uninoculated control plants grown in a nitrogen-free solution. Nodulation of a wide range of host plants by the majority of the tested isolates was observed, which agreed well with the results of our previous field experiment (Shutsrirung et al. 2002: Soil Sci. Plant Nutr., 48, 491–499). Up to 75% of the tested isolates induced a higher growth efficiency than that of the uninoculated control in association with one of the tested cultivars, Black soybean. Comparision with uninoculated control plants, enable to estimate the proportion of the tested isolates leading to effective growth promotion (E + e) of each cultivar, namely, Black soybean (local Thai cultivar), 75%; Cowpea, 82%; SJ5 (commercial Thai cultivar), 33%; Bragg (US cultivar), 33%; and Improved Pelican (US cultivar), 9%. These results indicated that although isolates with a high infectiveness with both “Asian-type” and “US-type” soybeans could be found, a high frequency of isolates leading to inefficient nodules was observed in the US cultivar, suggesting the presence of genetic differences in the soybean cultivars that express high-preference (efficient nodules) or low-preference (inefficient nodules) for a certain group of tested isolates. Based on the results of this laboratory experiment together with our previous field experiment, native rhizobial populations in the irrigated area of northern Thailand could be separated into three groups; Group 1: rhizobium strains showing a high effectiveness with only Asian cultivars, Group 2: strains showing a high effectiveness with only US origin cultivars, and Group 3: strains showing a high effectiveness with both Asian and US origin cultivars. The majority of the native rhizobial populations belonged to Group 1. The isolates in Group 3 may display a high potential for manipulating useful rhizobial inoculant.     

Cloning and characterization of a NADP+-malic enzyme gene from Bradyrhizobium japonicum USDA110

Malic enzymes have been considered to play a key role in energy metabolism for nitrogenase reaction in bacteroids. To elucidate the physiological role of the malic enzymes in Bradyrhizobium japonicum bacteroids, a putative malic enzyme gene Bjtme1 was cloned by polymerase chain reaction (PCR) using degenerated primers from conserved regions of the protein sequences of bacterial malic enzymes and draft sequence data of the Bradyrhizobium japonicum USDA110 genome sequence project. To confirm the characteristics of the Bjtme1 gene, the protein encoded by this gene was over-expressed using a pET32a(+) system and it exhibited a NADP⁺-malic enzyme (EC 1.1.1.40) activity, indicating that Bjtme1 was the gene of the NADP⁺-malic enzyme. This is the first report on the cloning and characterization of the NADP⁺-malic enzyme gene from B. japonicum, and the gene structure was compared with that of NADP⁺-malic enzyme genes of other rhizobia.     

Abstracts of Nipon Dojo-Hiryogaku Zasshi

We investigated changes in radiocesium concentrations in a Japanese chestnut (Castanea crenata Sieold &; Zucc.) orchard in Ibaraki prefecture for 3 years after the Tokyo Electric Power Company’s Fukushima Daiichi nuclear power plant accident in March 2011. The radiocesium concentrations in the aboveground organs of Japanese chestnut trees were almost the same, while the concentration in the roots was the lowest among all the organs investigated. The concentration of radiocesium decreased exponentially for 3 years in nuts, leaves and current shoots. The radiocesium concentrations in soils were higher in the surface layer, and the trend of an annual decrease in radiocesium in the soils was similar to that of the natural decay of radiocesium. The transfer factor of radiocesium from soils to nuts of Japanese chestnut decreased annually. These results suggest that radiocesium adhered directly to the aboveground organs of Japanese chestnut trees in March 2011, and that the accumulation of radiocesium in nuts is mainly due to radiocesium transfer from the branches and trunk to nuts several years after the nuclear power plant accident.