Production of offspring from vacuum-dried mouse spermatozoa and assessing the effect of drying conditions on sperm DNA and embryo development

Freeze-dried sperm (FD sperm) are of great value because they can be stored at room temperature for long periods of time, However, the birth rate of offspring derived from FD sperm is low and the step in the freeze-drying process particularly responsible for low offspring production remains unknown. In this study, we determined whether the drying process was responsible for the low success rate of offspring by producing vacuum-dried sperm (VD sperm), using mouse spermatozoa dried in a vacuum without being frozen. Transfer of embryos fertilized with VD sperm to recipients resulted in the production of several successful offspring. However, the success rate was slightly lower than that of FD sperm. The volume, temperature, and viscosity of the medium were optimized to improve the birth rate. The results obtained from a comet assay indicated that decreasing the drying rate reduced the extent of DNA damage in VD sperm. Furthermore, even though the rate of blastocyst formation increased upon fertilization with VD sperm, full-term development was not improved. Analysis of chromosomal damage at the two-cell stage through an abnormal chromosome segregation (ACS) assay revealed that reduction in the drying rate failed to prevent chromosomal damage. These results indicate that the lower birth rate of offspring from FD sperm may result from the drying process rather than the freezing process.     

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.     

Selenium Behavior in Open Bulk Precipitation, Soil Solution and Groundwater in Alluvial Fan Area in Tsukui, Central Japan

A monitoring study was carried out in an alluvial fan area in Tsukui, Central Japan during the study period of 1999–2003, in order to explain selenium (Se) behaviors in ecosystem combined with air, soil and groundwater. Monthly Se concentrations in open bulk precipitation (rainfall+aerosol, gaseous deposition and etc.), soil solution (collected by porous ceramic-cup) and groundwater ranged from 0.1 to 1.4 μg L^?1 (volume-weighted average: 0.34 μg L^?1), 0.21 to 1.0 μg L^?1 (0.48 μg L^?1) and 1.6 to 2.4 μg L^?1 (2.2 μg L^?1), respectively. Se concentration in open bulk precipitation was negatively correlated with the rainfall amount. Se concentration in soil solution significantly increased with DOC concentration in soil solution. Besides, despite atmospheric Se input and rainfall to the grassland study area, Se concentration in soil solution and groundwater received no significant effect from the rainfall amount, pH, Se, DOC, SO₄ ^2?, NO₃ ^? and EC in rainfall. Even though Se concentrations in groundwater were significantly correlated with soil solution volume, Se, DOC and NO₃ ^? and groundwater level, the result of multiple regression analyses (MRA) indicated that the groundwater Se was negatively influenced by groundwater level, which depended on groundwater recharge. Se was transported into the groundwater through the groundwater recharge that largely increased in this alluvial fan study area after heavy rain.     

Effect of brown seaweed lipids on fatty acid composition and lipid hydroperoxide levels of mouse liver

Brown seaweed lipids from Undaria pinnatifida (Wakame), Sargassum horneri (Akamoku), and Cystoseira hakodatensis (Uganomoku) contained several bioactive compounds, namely, fucoxanthin, polyphenols, and omega-3 polyunsaturated fatty acids (PUFA). Fucoxanthin and polyphenol contents of Akamoku and Uganomoku lipids were higher than those of Wakame lipids, while Wakame lipids showed higher total omega-3 PUFA content than Akamoku and Uganomoku lipids. The levels of docosahexaenoic acid (DHA) and arachidonic acid (AA) in liver lipids of KK-A(y) mouse significantly increased by Akamoku and Uganomoku lipid feeding as compared with the control, but not by Wakame lipid feeding. Fucoxanthin has been reported to accelerate the bioconversion of omega-3 PUFA and omega-6 PUFA to DHA and AA, respectively. The higher hepatic DHA and AA level of mice fed Akamoku and Uganomoku lipids would be attributed to the higher content of fucoxanthin of Akamoku and Uganomoku lipids. The lipid hydroperoxide levels of the liver of mice fed brown seaweed lipids were significantly lower than those of control mice, even though total PUFA content was higher in the liver of mice fed brown seaweed lipids. This would be, at least in part, due to the antioxidant activity of fucoxanthin metabolites in the liver.     

Concomitant inhibition of nitrogen fixation and N-Phenyl-1-Naphthylamine uptake activity of bacteroids of a Bradyrhizohium japonicum strain by nitrate

Abstract

Effect of the treatment of soybean nodules with nitrate on the permeability of the outer membrane of bacteroids of Bradyrhizobium japonicurn strain 138NR was examined using the hydrophobic fluorescent probe N-phenyl-1-naphthylamine (NPN). The incorporation of NPN into the outer and inner membranes of the bacteroid cells isolated from nodules of soybean (Glycine max L. Merro cv. Tamahomare) was followed by the measurement of fluorescence at 30°C. The NPN-uptake activity of the bacteroids was expressed by two parameters, the initial rate of NPN-uptake, k, and the final fluorescence intensity, F _max. Acetylene reduction activity (ARA) of nodules, k, and F _max decreased exponentially during three weeks following the 20 d period after planting (DAP), resulting in high correlations between logarithmic transforms of k-values or F _max per bacteroid and those of ARA per bacteroid. Application of 20 mM KNO₃ to the nodulated roots at 24 DAP inhibited ARA of the roots by 85% of the control after 4 d of treatment. There were concomitant decreases in k and F _max by 75% and 65% of the control, respectively. These results suggest that treatment of soybean nodules with nitrate results in some changes in the permeability of the outer membrane of bacteroids with a concomitant decline of N₂-fixing activity of bacteroids.     

Domestication and cereal feeding developed domestic pig‐type intestinal microbiota in animals of suidae

Intestinal microbiota are characterized by host‐specific microorganisms, which have been selected through host‐microbe interactions under phylogenetic evolution and transition of feeding behavior by the host. Although many studies have focused on disease‐related intestinal microbiota, the origin and evolution of host‐specific intestinal microbiota have not been well elucidated. Pig is the ideal mammal model to reveal the origin and evolution of host‐specific intestinal microbiota because their direct wild ancestor and close phylogenetic neighbors are available for comparison. The pig has been recognized as a Lactobacillus‐type animal. We analyzed the intestinal microbiota of various animals in Suidae: domestic pigs, wild boars and Red river hogs to survey the origin and evolution of Lactobacillus‐dominated intestinal microbiota by metagenomic approach and following quantitative PCR confirmation. The metagenomic datasets were separated in two clusters; the wild animal cluster being characterized by a high abundance of Bifidobacterium, whereas the domesticated (or captured) animal cluster by Lactobacillus. In addition, Enterobacteriaceae were harbored as the major family only in domestic Sus scrofa. We conclude that domestication may have induced a larger Enterobacteriaceae population in pigs, and the introduction of modern feeding system further caused the development of Lactobacillus‐dominated intestinal microbiota, with genetic and geographical factors possibly having a minor impact.     

Isolation of multiple drug-resistant enteric bacteria from feces of wild Western Lowland Gorilla (Gorilla gorilla gorilla) in Gabon

Prevalence of drug-resistant bacteria in wildlife can reveal the actual level of anthropological burden on the wildlife. In this study, we isolated two multiple drug-resistant strains, GG6-2 and GG6-1-1, from 27 fresh feces of wild western lowland gorillas in Moukalaba-Doudou National Park, Gabon. Isolates were identified as Achromobacter xylosoxidans and Providencia sp., respectively. Minimum inhibitory concentrations of the following 12 drugs—ampicillin (ABPC), cefazolin (CEZ), cefotaxime (CTX), streptomycin (SM), gentamicin (GM), kanamycin (KM), tetracycline (TC), nalidixic acid (NA), ciprofloxacin (CPFX), colistin (CL), chloramphenicol (CP) and trimethoprim (TMP)—were determined. Isolate GG6-2 was resistant to all antimicrobials tested and highly resistant to CTX, SM, TC, NA and TMP. Isolate GG6-1-1 was resistant to ABPC, CEZ, TC, CL, CP and TMP.     

Decaying toxic wood as sodium supplement for herbivorous mammals in Gabon

African rainforest harbors herbivores at high density. However, because plants and soils typically lack in some essential minerals, rainforest is not always a suitable habitat for herbivores. How they fulfill the mineral requirements is therefore an important question to animal ecology and conservation. Although large marshes, called ‘bais’, are often mentioned as efficient mineral-resource, little information on other sodium resources has still been available. Our laboratory works and field surveys found that a peculiar item, decaying wood stumps of Anthostema aubryanum, played as a major sodium resource for herbivores in Moukalaba-Doudou National Park, Gabon. When A. aubryanum is alive, the sodium content of its bark is low and its latex is toxic. Sodium is accumulated in decaying stumps (mean=1,343 mg/kg dry matter). Eight herbivores visited stumps to ingest the dead wood. Fecal sample analysis revealed that western lowland gorillas, a species most-frequently using the stumps, consumed large amount of the dead wood as regular food. Our findings suggest that decaying A. aubryanum is critical sodium-resources and is a key species for herbivores in our study area. Importance of the A. aubryanum may be particularly large there, because it is a limited sodium-rich material that is available year round. Our study site is known as the site where the densities of several herbivores are among the highest at Central Africa. The relatively high herbivores density in our study site may partly depend on decaying A. aubryanum as sodium resources.     

Diluted isoflurane as a suitable alternative for diethyl ether for rat anaesthesia in regular toxicology studies

Despite its explosive properties and toxicity to both animals and humans, diethyl ether is an agent long used in Japan in the anaesthesia jar method of rat anaesthetises. However, in response to a recent report from the Science Council of Japan condemning diethyl ether as acceptable practice, we searched for an alternative rat anaesthesia method that provided data continuous with pre-existing regular toxicology studies already conducted under diethyl ether anaesthesia. For this, we examined two candidates; 30% isoflurane diluted with propylene glycol and pentobarbitone. Whereas isoflurane is considered to be one of the representatives of modern volatile anaesthetics, the method of propylene glycol-diluted 30% isoflurane used in this study was our modification of a recently reported method revealed to have several advantages as an inhalation anaesthesia. Intraperitoneal pentobarbitone has long been accepted as a humane method in laboratory animal anaesthesiology. These 2 modalities were scrutinized in terms of consistency of haematology and blood chemistry with previous results using ether. We found that pentobarbitone required a much longer induction time than diethyl ether, which is suspected to be the cause of fluctuations in several haematological and blood chemical results. Conversely, only calcium ion concentration showed a slight difference from traditional results in the case of 30% isoflurane. Additionally, serum prolactin and corticosterone levels indicated that 30% isoflurane induced less stress than ether, confirming that 30% isoflurane can both provide results consistent with diethyl ether, while at the same time remove its disadvantages. As such 30% isoflurane appears to be a strong alternative anaesthetic agent for future regular toxicology studies in Japan.     

Environmental characteristics of spawning and nursery grounds of Japanese sardine and mackerels in the Kuroshio and Kuroshio Extension area

The sustainable use of marine resources requires understanding the surrounding ecosystem and elucidating mechanisms of variation. However, we still lack a comprehensive understanding of environmental variation in the spawning and nursery grounds of important fisheries species Japanese sardine (Sardinops melanostictus) and mackerels (Scomber japonicus and Scomber australasicus) in the northwest Pacific. Here, we investigate detailed physical, chemical, and biological environment variations in the spawning and nursery grounds along the Kuroshio and Kuroshio Extension area from intensive investigation in spawning season (April) of 2013. We found similar water mass property and copepod community in the egg‐rich Kuroshio area and the larvae‐rich downstream Kuroshio Extension area, indicating environmental variability is small during transportation and development processes. The egg‐rich northern Izu Islands region showed high copepod abundance, although low nutrient and chlorophyll concentrations were observed. Eggs were scarce or absent in the second survey 10 days after abundant eggs were observed in the region, along with differences in water property and copepod community. This indicates that not only the location but also the specific water characteristic and copepod community are a determining factor for spawning. Indicator communities of copepod found in our study (indicator community of transportation process from spawning ground, of non‐spawning ground, and of reproductive area in the Kuroshio Extension area) would be a key factor for recruitment prediction.