Production of transgenic and non-transgenic clones in miniature pigs by somatic cell nuclear transfer

Miniature pigs have been recognized as valuable experimental animals in various fields such as medical and pharmaceutical research. However, the amount of information on somatic cell cloning in miniature pigs, as well as genetically modified miniature pigs, is much less than that available for common domestic pigs. The objective of the present study was to establish an efficient technique of cloning miniature pigs by somatic cell nuclear transfer. A high pregnancy rate was achieved following transfer of parthenogenetic (3/3) and cloned (5/6) embryos using female miniature pigs in the early pregnancy period as recipients after estrus synchronization with prostaglandin F2 alpha analog and gonadotrophins. The production efficiency of the cloned miniature pigs using male and female fetal fibroblasts as nucleus donors was 0.9% (2/215 and 3/331, respectively). Cloned miniature pigs were also produced efficiently (7.8%, 5/64) by transferring reconstructed embryos into the uteri of common domestic pigs. When donor cells transfected with the green fluorescent protein (GFP) gene were used in nuclear transfer, the production efficiency of the reconstructed embryos and rate of blastocyst development were comparable to those obtained by non-transfected cells. When transfected cell-derived reconstructed embryos were transferred to three common domestic pig recipients, all became pregnant, and a total of ten transgenic cloned miniature pigs were obtained (piglet production efficiency: 2.7%, 10/365). Hence, we were able to establish a practical system for producing cloned and transgenic-cloned miniature pigs with a syngeneic background.     

Effects of co‐supply ratios of swimming crab Portunus trituberculatus zoeae and Artemia on survival and growth of East Asian common octopus Octopus sinensis paralarvae under an upwelling culture system

Despite recent advances in culture technology for East Asian common octopus Octopus sinensis paralarvae using upwelling systems, securing suitable feed for the paralarvae is an unresolved issue. The zoea of the swimming crab Portunus trituberculatus is a good candidate for paralarval feed because of the high fecundity of the adult females. To investigate the effects of supplying P. trituberculatus zoeae and their feeding method on paralarvae, we cultured paralarvae with supplying different combination ratios of zoeae and Artemia (10:0, 7:3, 5:5, 3:7 and 0:10), and with or without supplementing rotifers using small‐scale (3‐L) upwelling systems. Paralarval survival rate and growth were improved when zoeae were supplied as the main feed, but reduced when the proportion of Artemia exceeded half the whole preys. Supplementing rotifers did not affect the paralarval survival and growth. Subsequently, paralarvae were cultured by supplying zoeae (partially augmented by Artemia) using three large (1‐kl) upwelling systems to assess their feeding effectiveness in juvenile octopus production. Paralarvae could be cultured at high survival rates of 77.1 ± 5.1% to reach benthic juveniles at 23 days after hatching. In conclusion, supplying P. trituberculatus zoeae augmented with Artemia under an upwelling culture system has great potential for juvenile octopus production.     

Assessment of decay risk of airborne wood-decay fungi

The decay risk of airborne wood-decay fungi was investigated by using an air sampler. Japanese cedar disks 7.8 cm in diameter and about 3 mm in thickness with moisture content of about 100% were placed in a “BIOSAMP” air sampler and exposed to 1000 l air. Air sampling was carried out from June to September at the same sampling site in Tsukuba, Japan. The exposed disks were then incubated for 16 weeks in a damp container kept at 26° ± 2°C. During the incubation period, wood mass loss ranged from −15 to 807 mg with a mean mass loss of 244 mg. Factors affecting mass loss were explored. Wood moisture content and ratio of heartwood area proved to be significant factors. In addition, six weather factors were found to influence mass loss. Disks that were sampled on a cloudy day showed significantly higher mean mass loss compared to those sampled on a sunny day. Subculturing of filamentous fungi from 16-week incubated disks suggested one-third of the isolated fungi produced ligninolytic enzymes.     

Effect of drying method as a pretreatment on CUAZ preservative impregnation in Japanese cedar logs

Green Japanese cedar logs 2 m in length and 18 cm in diameter were dried to a mean moisture content of less than 30% by either air drying or kiln drying. Dried logs were impregnated with copper azole (CUAZ) solution according to Japanese Industrial Standard (JIS) A9002. Preservative absorption was calculated from the log weight before and after preservative impregnation. Impregnated logs were then dried in the air and cut at the center to determine preservative penetration. The penetration area was determined after visualizing the preservative with chrome azurol S. The visualized area indicated that the preservative solution penetrated into the sapwood portion of almost all the air-dried logs. However, the kiln-dried logs did not show full penetration into the sapwood portion. The visualized area of some kiln-dried sapwood showed a penetration value of less than 80%, which is the minimum requirement set by the Japanese Agricultural Standard (JAS) for sawn timber. Statistical analysis showed that penetration in the air-dried sapwood was significantly better than that in the kiln-dried sapwood. It was concluded that air drying is more favorable than kiln drying as the predrying method for CUAZ impregnation. On the other hand, preservative absorption was not affected by the drying method.     

Natural Resistance of Two Plantation Woods Populus × canadensis cv. and Cunninghamia lanceolata to Decay Fungi and Termites

Natural durability of two plantation woods, Chinese fir and I-214 poplar, was investigated thoroughly by three testing methods, namely an accelerated laboratory decay test, a fungus cellar test and a field test. Aft er the decay test using Postia placenta and Trametes versicolor, Chinese fir and the I-214 poplar showed 34% and 69% of mass loss, respectively, indicating they should be classified as slightly durable and non-durable wood. This conclusion was confirmed by the fungus cellar test and the field test. Like the performance in the decay test, I-214 poplar showed no resistance to termites either in the laboratory or in the field,whereas Chinese fir would be classified as moderately resistant.     

Temporal changes in distribution and composition of N from labeled fertilizer in soil organic matter fractions

Variations in the amount and composition of immobilized nitrogen (N) in major soil organic matter fractions were investigated in a 730-day soil incubation experiment using ¹⁵N-labeled urea and ¹⁵N nuclear magnetic resonance spectroscopy with the cross polarization/magic angle spinning (¹⁵N CPMAS NMR) method. After 730 days, 24.7% of the applied N was recovered from the soil as organic N. The urea-derived N recovered from humic acids and humin decreased from 11.2 and 33.8% of the applied amount after 14 days to 1.6 and 20.4% after 730 days, respectively. When these values were corrected for the microbial biomass (MB) N, they ranged from 9.0 to 1.2% and 28 to 18%, respectively. The proportion of urea-derived N recovered from fulvic acids was low, ranging between 0.4 and 5.8% (with MB N) or 5.6% (without MB N) of the applied amount, whereas that from water-soluble nonhumic substances (WS-NHS; NHS in the fulvic acid fraction) remained high, 28–33% of the applied amount after correction for the contribution of MB N up to day 365, and decreased to 0.9% thereafter. The ¹⁵N CPMAS NMR spectra of humic acids, fulvic acids, and humin showed the largest signal at −254 to −264 ppm, corresponding to peptide/amide N. The proportions of heterocyclic, peptide/amide, guanidine/aniline, and free amino N in the urea-derived humic acid N were 3–7, 83–90, 5–7, and 2–4%, respectively. More than 80% loss of the urea-derived humic acid N did not markedly alter their composition. No time-dependent variations were also observed for the proportions of respective N functional groups in humin N, which were 3–5, 71–78, 12–17, and 6–10% in the same order as above. These results suggest the greater importance of physical stability than structural variation for the initial accumulation of organic N in soil.     

Response of microbial activity and microbial community composition in soils to long-term arsenic and cadmium exposure

Arsenic (As) and cadmium (Cd) in soils can affect soil microbial function and community composition and, therefore, may have effects on soil ecosystem functioning. The aim of our study was to assess the effects of long-term As and Cd contamination on soil microbial community composition and soil enzyme activities. We analyzed soils that have been contaminated 25 years ago and at present still show enhanced levels of either As, 18 and 39 mg kg⁻¹, or Cd, 34 and 134 mg kg⁻¹. Soil without heavy metal addition served as control. Polymerase chain reaction (PCR) followed by denaturing gradient gel electrophoresis (DGGE) showed that bacterial community composition in As and Cd contaminated soils differed from that in the control soil. The same was true for the microbial community composition assessed by analysis of respiratory quinones. Soil fungi and Proteobacteria appeared to be tolerant towards As and Cd, while other groups of bacteria were reduced. The decline in alkaline phosphatase, arylsulphatase, protease and urease activities in the As- and Cd-contaminated soils was correlated with a decrease of respiratory quinones occuring in Actinobacteria and Firmicutes. Xylanase activity was unaffected or elevated in the contaminated soils which was correlated with a higher abundance of fungal quinones, and quinones found in Proteobacteria.     

Altered pesticide use on transgenic crops and the associated general impact from an environmental perspective

The large-scale commercial cultivation of transgenic crops has undergone a steady increase since their introduction 10 years ago. Most of these crops bear introduced traits that are of agronomic importance, such as herbicide or insect resistance. These traits are likely to impact upon the use of pesticides on these crops, as well as the pesticide market as a whole. Organizations like USDA-ERS and NCFAP monitor the changes in crop pest management associated with the adoption of transgenic crops. As part of an IUPAC project on this topic, recent data are reviewed regarding the alterations in pesticide use that have been observed in practice. Most results indicate a decrease in the amounts of active ingredients applied to transgenic crops compared with conventional crops. In addition, a generic environmental indicator -- the environmental impact quotient (EIQ) -- has been applied by these authors and others to estimate the environmental consequences of the altered pesticide use on transgenic crops. The results show that the predicted environmental impact decreases in transgenic crops. With the advent of new types of agronomic trait and crops that have been genetically modified, it is useful to take also their potential environmental impacts into account.     

Linear relation between the amount of respiratory quinones and the microbial biomass in soil

A linear relationship was observed between the amount of total respiratory quinones and the microbial biomass measured by a fumigation-extraction method in 15 soil samples regardless of the significant differences in the composition of the quinone profiles, with one exception in a soil amended with a very high application rate of farmyard manure. It is suggested that the amount of total respiratory quinones can be used as an indicator of the microbial biomass in soil.     

Risk factors for canine tail chasing behaviour in Japan

The aim of this study was to investigate the risk factors for tail chasing behaviour that occurs when a dog spins in tight circles to chase its tail, sometimes biting it. The behaviour is a sign of canine compulsive disorder (CD). A questionnaire about tail chasing behaviour and general information about the animals was used to collect data on seven breeds of pet dogs. The data were gathered at a dog event and at veterinary practices. To determine which variables were associated with tail chasing behaviour, stepwise multiple regression analyses were performed. Regardless of cohort, 'breed' and 'source of acquisition' were significantly associated with tail chasing behaviour. Using a chi-square test, the association between 'source of acquisition' and the behaviour was examined separately in two breeds (Shiba inu and Dachshund) that had the largest number of individuals chasing their tails accompanied by biting and/or growling at them. This factor showed a significant and consistent association across the two breeds. With respect to the risk factors of 'breed' and 'source of acquisition', high percentages of Shiba inu and dogs originating from pet stores were included in the group chasing their tails with biting and/or growling. The results suggest that distinct risk factors exist for tail chasing behaviour and such factors appear to be regulated by both genetics and the environment.