Pueruli of Panulirus longipes bispinosus (Crustacea, Decapoda, Palinuridae) stranded on the beach of Kuroshima Island, Ryukyu Archipelago, southern Japan

ABSTRACT: Stranded organisms were observed on the sandy beach of Kuroshima Island, Ryukyu Archipelago, in the early morning of 28 January 1993. Stranded organisms of 525 individuals were classified into 74 species. These organisms included 13 individuals of Panulirus pueruli, which were identical to P. longipes bispinosus. Kuroshima Island is located near Ishigakijima and Iriomote islands, Ryukyu Archipelago, where coral reefs develop. Coral reefs are common and develop in the Ryukyu Archipelago, southern Japan, with the exception of Yakushima and Tanegashima islands. Panulirus japonicus in Japanese waters has not been found in coral reef regions where P. longipes bispinosus are mainly found. The present study describes and identifies to species level puerulus specimens that were stranded on the beach of Kuroshima Island.     

The effect of depopulation and restocking on reproductive and growth performances on Japanese commercial swine farms

This study compared the reproductive and growth performances of pigs before and after depopulation and restocking after a foot-and-mouth disease outbreak in Japan. Data for the time period before and after depopulation and restocking were obtained from three farrow-to-finish farms. As a result of depopulation and restocking, hygiene levels were improved, and common infectious diseases, such as porcine reproductive and respiratory syndrome and Aujeszky’s disease, remained undetected on the farms. Compared with before depopulation, reproductive and growth performances improved after depopulation; the number of total pigs born was higher, the postweaning mortality rate was lower, and the age at slaughter was lower (P<0.05). In summary, depopulation and restocking improved the reproductive and growth performances of pigs.     

Isolation of Pathogenicity Mutants of Fusarium oxysporum f. sp. melonis by Insertional Mutagenesis

Restriction enzyme-mediated integration (REMI) mutagenesis was used to isolate mutants of Fusarium oxysporum f. sp. melonis impaired in pathogenicity. The race 2 strain Mel02010 was transformed with linearized pSH75, conferring resistance to hygromycin B, with or without the enzyme used to linearize the plasmid. Addition of restriction enzymes did not affect the transformation frequency. A total of 2929 REMI transformants were tested for pathogenicity to three melon cultivars, Amus, Ogon 9 and Ohi. The race 2 strains are pathogenic to Amus and Ogon 9, but not to Ohi. Of 43 transformants with reduced pathogenicity on susceptible melon cultivars, 12 mutants were examined in detail for pathogenicity, vegetative growth and integrative mode of pSH75. The levels of pathogenicity varied among these mutants. Two mutants (B48 and B137) almost completely lost pathogenicity to both susceptible cultivars, and the others had reduced pathogenicity. Mutants B48, B241, B886 and X36 were also impaired in vegetative growth. Mutant B809 was a biotin auxotroph. By DNA gel blot analysis, nine mutants were found to contain a single copy of the transformation vector. These mutants may thus be useful in isolating genes involved in pathogenicity. Received 22 December 2000/ Accepted in revised form 16 April 2001     

Development of a shuttle polymerase chain reaction for the detection of bovine herpesvirus 2

Three different polymerase chain reaction (PCR) protocols were evaluated for their ability to detect bovine herpesvirus 2 (BoHV-2): single-step PCR with 3 reaction stages (denaturation, annealing and extension), 2 reaction stages (denaturation and annealing/extension; shuttle PCR), and semi-nested PCR with 3 reaction stages. All the PCR protocols showed the same sensitivity (detection limit of 0.4 TCID(50)). A non-specific band sometimes appeared in mock cell DNA at annealing temperatures below 64 degrees C. The shuttle PCR was found to be superior to the other protocols under consideration because of the speed of its application. Furthermore, no non-specific band was detected in DNAs of eight other DNA viruses. Thus, the shuttle PCR seems to be an excellent diagnostic tool for BoHV-2 infections.     

Calcium ions are involved in egress of Babesia bovis merozoites from bovine erythrocytes

Bovine babesiosis is a livestock disease known to cause economic losses in endemic areas. The apicomplexan parasite Babesia bovis is able to invade and destroy the host’s erythrocytes leading to the serious pathologies of the disease, such as anemia and hemoglobinuria. Understanding the egress mechanisms of this parasite is therefore a key step to develop new therapeutic strategies. In this study, the possible involvement of Ca²⁺ in the egress of B. bovis merozoites from infected erythrocytes was investigated. Egress was artificially induced in vitro using calcium ionophore A23187 and thapsigargin to increase Ca²⁺ concentration in the cytosol of the parasite cells. The increased intracellular Ca²⁺ concentration following these treatments was confirmed using live cell Ca²⁺ imaging with confocal laser scanning microscopy. Based on our findings, we suggest a Ca²⁺ signalling pathway in the egress of B. bovis merozoites.     

Weak activity of UDP-glucuronosyltransferase toward Bisphenol analogs in mouse perinatal development

Bisphenol A (BPA) is a widely used industrial chemical that disrupts endocrine function. BPA is an endocrine disrupting chemical (EDC) that has been demonstrated to affect reproductive organ development, brain development, metabolic disease and post-natal behavior. Accordingly, Bisphenol analogs, Bisphenol F (BPF, bis (4-hydroxyphenyl) methane) and Bisphenol AF (BPAF, 4,4-hexafluoroisopropylidene) diphenol) are used as replacements for BPA. BPA is mainly metabolized by UDP-glucuronosyltransferase (UGT), UGT2B1, but this effective metabolizing system is weak in the fetus. In the present study, we demonstrated that hepatic UGT activity toward BPAF was very weak, in comparison with BPA and BPF, in the fetus, pups and dams. Conversely, hepatic UGT activity toward BPF was very weak in the fetus and newborn pups, and was increased to the same level as BPA post-partum. In conclusion, BPAF possibly tends to accumulate in the fetus, because of weak metabolism during the perinatal period, suggesting that the metabolism of individual Bisphenol analogs requires assessment to properly gauge their risks.     

Mitigation of nitrous oxide (N2O) emission from swine wastewater treatment in an aerobic bioreactor packed with carbon fibers

Mitigation of nitrous oxide (N₂O) emission from swine wastewater treatment was demonstrated in an aerobic bioreactor packed with carbon fibers (CF reactor). The CF reactor had a demonstrated advantage in mitigating N₂O emission and avoiding NOx (NO₃ + NO₂) accumulation. The N₂O emission factor was 0.0003 g N₂O‐N/gTN‐load in the CF bioreactor compared to 0.03 gN₂O‐N/gTN‐load in an activated sludge reactor (AS reactor). N₂O and CH₄ emissions from the CF reactor were 42 g‐CO₂ eq/m³/day, while those from the AS reactor were 725 g‐CO₂ eq/m³/day. The dissolved inorganic nitrogen (DIN) in the CF reactor removed an average of 156 mg/L of the NH₄‐N, and accumulated an average of 14 mg/L of the NO₃‐N. In contrast, the DIN in the AS reactor removed an average 144 mg/L of the NH₄‐N and accumulated an average 183 mg/L of the NO₃‐N. NO₂‐N was almost undetectable in both reactors.     

Histone H4 Modification During Mouse Spermatogenesis

The core histone is composed of four proteins (H2A, H2B, H3 and H4). Investigation of the modification patterns of histones is critical to understanding their roles in biological processes. Although histone modification is observed in multiple cells and tissues, little is known about its function in spermatogenesis. We focused on the modification patterns of histone H4 during murine spermatogenesis. We demonstrated that the individual N-terminal sites of H4 show different modification patterns during the differentiation of male germ cells. The methylation pattern varied depending on the residues that were mono-, di-, or tri-methylated. All the H4 modifications were high during the meiotic prophase, suggesting that histone H4 modification plays an important role during this stage of spermatogenesis. Elongating spermatids showed increased acetylation of histone H4, which may be associated with a histone-to-protamine substitution. Our results provide further insight into the specific relationship between histone H4 modification and gene expression during spermatogenesis, which could help to elucidate the epigenetic disorders underlying male infertility.     

Microdroplet In Vitro Fertilization Can Reduce the Number of Spermatozoa Necessary for Fertilizing Oocytes

Successful in vitro fertilization (IVF) in mice has been achieved using spermatozoa at concentrations specifically optimized for the experimental conditions, such as species and source of spermatozoa. Although IVF in mice is mostly performed using about 80–500 µl drops, it is expected that the number of spermatozoa used for insemination can be reduced by decreasing the size of the IVF drops. The present study was undertaken to examine the extent to which the number of spermatozoa used for IVF could be reduced by using small droplets (1 µl). We devised the experimental parameters using frozen–thawed spermatozoa from C57BL/6 mice in anticipation of broader applications to other mouse facilities. We found that as few as 5 spermatozoa per droplet could fertilize oocytes (1 or 3 oocytes per droplet), although the fertilization rates were low (13–15%). Practical fertilization rates (> 40%) could be achieved with frozen-thawed C57BL/6J spermatozoa, which are sensitive to cryopreservation, when 20 sperm per droplet were used to inseminate 3 oocytes. Even with spermatozoa from a very poor quality suspension (10% motility), about 25% of oocytes were fertilized. Our calculations indicate that the number of inseminated spermatozoa per oocyte can be reduced to 1/96–1/240 by this method. In two separate embryo transfer experiments, 60% and 47%, respectively, of embryos developed to term. Our microdroplet IVF method may be particularly advantageous when only a limited number of motile spermatozoa are available because of inadequate freezing-thawing or genetic reasons.