Effect of volume of non-surgical embryo transfer medium on ability of porcine embryos to survive to term

Non-surgical embryo transfer is a promising method for improving efficiency in the pork industry and also for biotechnology applications, such as in vitro embryo production, transgenesis and cloning. Several groups have reported successful piglet production using an artificial insemination catheter or flexible catheter designed for this procedure; however, the efficiency of the technique is still low. The critical points that need to be addressed in order to improve this procedure are (1) the embryo deposition site and (2) volume of transfer medium associated with the embryos; however, the latter has not yet been examined systematically. In the present study, we evaluated the effect of the volume of non-surgical embryo transfer medium on the ability of porcine embryos to survive to term by using a recently produced flexible catheter. The catheter consists of a guide and an injector. Blastocysts 200-230 mum in diameter were collected from donor gilts and transferred to recipient gilts. The time required for the completion of embryo transfer using this catheter was 14.6 +/- 3.9 min. The tip of the injector was determined by laparotomy to be located in a uterine horn 20-30 cm anterior from the branching point of the uterus body. We transferred 17.0-17.3 embryos with different volumes of medium (1.6, 3.2 and 10 ml) into each of 5, 4 and 4 recipients, respectively, and pregnancy was confirmed in 4, 3 and 1 of these recipients, respectively. Three recipients in the 1.6 ml group farrowed a total of 19 piglets (4, 5 and 10 piglets, respectively). These results suggest that successful non-surgical embryo transfer is affected by the volume of transfer medium.     

Evaluation of Single-Incision Laparoscopic Ovariectomy in Standing Mares

Our objective was to compare the invasiveness of single-incision laparoscopic surgery (SILS) and multiport laparoscopic surgery (MLS) for ovariectomy in 5 standing healthy adult Thoroughbred mares. First, laparoscopic ovariectomy was performed by SILS or by MLS on the right paralumbar fossa region in a standing mare. One month after surgery, ovariectomy by the other method was performed on the left side. For surgery evaluation, the surgical time, length of incision, and amount of lidocaine used were recorded and compared between SILS and MLS. Physical examination (body temperature, heart rate, and respiration rate) and hematology (number of leukocytes and hematocrit) were performed before surgery and every day for 7 days after surgery. Similarly, the degrees of swelling, heat, pain, and incisional wound healing were evaluated (grades1-4). The length of incision and amount of local anesthetic for SILS were significantly less than those for MLS. Moreover, the scores for swelling, heat, and pain in the SILS group tended to be lower than those in the MLS group for several days after surgery. We concluded that SILS was less invasive than MLS and is therefore useful for ovariectomy in mares.     

Characteristics of cytochrome P450-dependent metabolism in the liver of the wild raccoon,Procyon lotor

Wildlife is exposed to a wide range of xenobiotics in the natural environment. In order to appropriately assess xenobiotic-induced toxicity in wildlife, it is necessary to understand metabolic capacities. Carnivores, in general, have low metabolic abilities, making them vulnerable to a variety of chemicals. Raccoons (Procyon lotor) in the wild have been found to have high levels of xenobiotics. However, little is known about the metabolic capacity of the cytochrome P450 (CYP) enzymes in this species. Thus, this study used liver samples to investigate the characteristics of CYP enzymes in wild raccoons. In 22 wild raccoons, CYP concentrations in hepatic microsomes were examined. To better understand the properties of CYP-dependent metabolism, in vitro metabolic activity studies were performed using ethoxyresorufin, pentoxyresorufin and testosterone as substrates. In addition, three raccoons were fed commercial dog food in the laboratory for one week, and the effects on CYP-dependent metabolism were investigated. In comparison to other mammalian species, raccoons had very low concentrations of CYP in their livers. In an in vitro enzymatic analysis, raccoons’ ethoxyresorufin O-deethylase (EROD) and pentoxyresorufin O-depentylase (PROD) metabolic capacities were less than one-fifth and one-tenth of rats’, respectively. These results indicate the possible high risk in raccoons if exposed to high levels of environmental xenobiotics because of their poor CYP activity. In this study, the features of CYP-dependent metabolism in wild raccoons are described for the first time.     

Comparative study between full cell and passive impregnation method of wood preservation for laser incised Douglas fir lumber

In this study, the ability of passive impregnation of Douglas fir (Pseudotsuga menziesii Franco) lumber was compared with a conventional full cell method. With an incising density of 10,000 holes/m² carried out by CO₂ laser, square Douglas fir lumber was treated by both passive impregnation and full cell method for 6 h and 50 min. Different incising densities were also used in passive impregnation to examine the optimum incising density. It was observed that there was no significant difference between these two methods regarding absorption of liquid, moisture content after dipping and penetrated area both at cross and longitudinal sections. However, the value was higher in case of full cell method. Regarding different incising densities, absorption and penetration of liquid was similar for 10,000 and 7,500 holes/m² but it was low for 5,000 holes/m². Therefore, it can be concluded that an incising density of 7,500 holes/m² is the optimum for passive impregnation. The absorption of liquid and its distribution in wood indicates that this non-pressure passive impregnation method can be a good preservation method for impermeable lumber and can be an alternative to full cell method.     

Stand-scale transpiration estimates in a Moso bamboo forest: (I) Applicability of sap flux measurements

The applicability of sap flux (F_d) measurements to bamboo forests has not been studied. This study was undertaken to establish an optimal and effective design for stand-scale transpiration (E) estimates in a Moso bamboo forest. To this aim, we validated F_d measurements in Moso bamboos in a cut bamboo experiment. In addition, we analyzed how sample sizes affect the reliability of E estimates calculated from F_d and conducting culm area (A_{S_b}). In the cut bamboo experiments, we found that F_d measurement using a 10 mm probe was a valid means of determining the water-use behavior of a Moso bamboo, although a specific correction was needed. Furthermore, we calculated E from stand A_{S_b} (A_{S_stand}) and mean stand F_d (J_S). Employing Monte Carlo analysis, we examined potential errors associated with sample size in E, A_{S_stand}, and J_S using an original dataset with A_{S_b} and F_d measured for 40 and 16 individuals, respectively. Consequently, we determined the optimal sample size for both A_{S_stand} and J_S estimates as 11. The optimal sample sizes for J_S were almost the same under different vapor pressure deficit and soil moisture conditions. The optimal sample size for J_S at the study site was less than that of a coniferous plantation in the same region probably owing to small individual-to-individual variations in sap flux in the Moso bamboo forest. Our study concludes that sap flux measurements are an applicable technique for assessing water use in Moso bamboo forests.     

Differences in bird species diversities between a natural mixed forest and a coniferous plantation

Bird species diversity and bird species richness were surveyed in a natural mixed forest (mature forest) and in a coniferous plantation (30–40 years old) monthly from 1990 to 1994. These forests were 20 km apart and had similar weather conditions and soil types. There were larch and cedar plantations, and hardwoods along streams in the coniferous plantation. In the natural mixed forest, 55 bird species were found, whereas 40 species were recorded in the coniferous plantation. The species diversity of bird communities was significantly higher in the natural mixed forest than in the coniferous plantation mixed with hardwood trees. This result indicates that the coniferous plantation mixed with hardwood trees, presumed to be a more simplified environment, is likely to be less inhabitable for many bird species than the natural mixed forest, suggesting a relationship between the diversity of the forest environment and the diversity of the bird community. A higher number of bird species was recorded in the small coniferous forest mixed with hardwood trees compared with those in the coniferous plantation. The mixing of broad-leaved trees in a coniferous plantation was suggested to be effective in increasing number of bird species. These tendencies were recorded all through the year during our study. In snow-covering periods, both the diversity and the bird species richness fell in all study sites every year. Snow cover would have greater effect on bird species diversity in the coniferous plantation than in other forest types probably due to diet shortages caused by the snow cover over the forest floor.     

Drying and anatomical characteristics of sugi wood attacked by bacteria during pond storage

Seven species of bacteria were isolated and identified from ponded sugi (Cryptomeria japonica D. Don) logs, and six species showed potent wood-degrading activities. To evaluate the effects of these isolated bacteria on the drying and anatomical characteristics of wood, small fresh blocks of sugi were immersed in water suspensions containing bacteria for 1–7 months. The permeability and drying properties were evidently improved. Most of the encrusting substances adhering to the cell lumens and the pit chambers were removed, and the pit membranes were destroyed. These anatomical changes due to bacterial activity were assumed to improve the permeability of sugi wood.Part of this report was presented at the 40^th annual meeting of the Japan Wood Research Society, Tsukuba, April 1990; the 41^st annual meeting of the Japan Wood Research Society, Matsue, April 1991; and the IUFRO XX World Congress, Tampere, August 1995     

Molecular Monitoring of hrpB-disrupted Mutant of Ralstonia solanacearum in Tomato Plants

A nonpathogenic mutant of Ralstonia solanacearum was produced by the insertion of transposon Tn4431. The mutagenized gene was then cloned from a genomic DNA library by the gene tagging method, using the labeled lux operon located on Tn4431 of pUCD623 as a hybridization probe. From nucleotide sequence analysis of the transposon-inserted genomic clone, the hrpB gene was shown to be disrupted by the inserted transposon. Tomato plants were inoculated with the hrpB-disrupted mutant bacteria, for which multiplication and translocation were then monitored using the colony hybridization method. In addition, the original pathogenic bacteria in which the lux operon had been functionally ligated with the genomic promoter were also used for inoculation and traced by their bioluminescence. Multiplication of the hrpB-disrupted mutant was suppressed initially in the invaded root tissues and then in upper hypocotyl after translocation, suggesting that the pathogenic strain of R. solanacearum overcomes at least two steps of host responses expressed in root and hypocotyl tissues. Thus, our approach for molecular monitoring of the bacteria enabled us to precisely analyze the infection behavior of the pathogenic bacteria in planta. Received 16 April 1999/ Accepted in revised form 10 August 1999     

In Vitro Suppression of Mycelial Growth of Fusarium oxysporum by Extracellular Chitosanase of Sphingobacterium multivorum and Cloning of the Chitosanase Gene csnSM1

A chitosan-degrading bacterium, isolated from field soil that had been amended with chitin, was identified as Sphingobacterium multivorum KST-009 on the basis of its bacteriological characteristics. The extracellular chitosanase (SM1) secreted by KST-009 was a 34-kDa protein and could be purified through ammonium sulfate precipitation, gel permeation column chromatography and SDS polyacrylamide gel electrophoresis. A chitosanase gene (csnSM1) was isolated from genomic DNA of the bacteria, and the entire nucleotide sequence of the gene and the partial N-terminal amino acid sequence of the purified SM1 were determined. The csnSM1 gene was found to encode 383 amino acids, 72 N-terminal amino acid residues were processed to produce the mature enzyme during the secretion process. Germinated microconidia of four formae speciales (lycopersici, radicis-lycopersici, melonis, and fragariae ) of Fusarium oxysporum were treated with SM1. Chitosanase treatment caused morphological changes, such as swelling of hyphal cells or indistinctness of hyphal cell tips and cessation or reduction of mycelial elongation. Received 2 May 2001/ Accepted in revised form 21 June 2001     

Occurrence of New Powdery Mildew on Greenhouse Tomato Cultivars

During a year-round survey on the occurrence of powdery mildew on greenhouse-cultivated tomato plants, the disease was most severe in June and July. All tomato plants (45 commercial cultivars and 11 breeding lines) tested were infected with the pathogen but had different degrees of susceptibility. The pathogen was epiphytic and produced white, round pustules mainly on leaves of tomato plants. The pathogen produced conidia singly on conidiophores and forked appressoria on inoculated tomato leaves and seemed to be an Oidium sp. of Erysiphe polygoni type. Received 18 December 2000/ Accepted in revised form 22 July 2001