Effects of acrosomal conditions of frozen-thawed spermatozoa on the results of artificial insemination in Japanese Black cattle

The purposes of this study were to examine the relationship between male artificial insemination (AI) fertility and sperm acrosomal conditions assessed by new and conventional staining techniques and to identify possible reproductive dysfunctions causing low conception rates in AI using frozen-thawed spermatozoa with poor acrosomal conditions in Japanese Black bulls. We investigated individual differences among bulls in the results concerning (1) acrosomal conditions of frozen-thawed spermatozoa as assessed by not merely peanut agglutinin-lectin staining (a conventional staining technique) but also immunostaining of acrosomal tyrosine-phosphorylated proteins (a new staining technique), (2) routine AI using frozen-thawed spermatozoa as assessed by pregnancy diagnosis, (3) in vivo fertilization of frozen-thawed spermatozoa and early development of fertilized eggs as assessed by superovulation/AI-embryo collection tests and (4) in vitro fertilization of frozen-thawed spermatozoa with oocytes. The percentages of frozen-thawed spermatozoa with normal acrosomal conditions assessed by the abovementioned staining techniques were significantly correlated with the conception rates of routine AI, rates of transferable embryos in superovulation/AI-embryo collection tests and in vitro fertilization rates. These results are consistent with new suggestions that the distribution of acrosomal tyrosine-phosphorylated proteins as well as the acrosomal morphology of frozen-thawed spermatozoa are AI fertility-associated markers that are valid for the prediction of AI results and that low conception rates in AI using frozen-thawed spermatozoa with poor acrosomal conditions result from reproductive dysfunctions in the processes between sperm insemination into females and early embryo development, probably failed fertilization of frozen-thawed spermatozoa with oocytes.     

Multi-locus sequence typing of Salmonella enterica subsp. enterica serovar Enteritidis strains in Japan between 1973 and 2004

Salmonella enterica subsp. enterica serovar Enteritidis (S. Enteritidis) was responsible for a worldwide pandemic during the 1980s and 1990s; however, changes in the dominant lineage before and after this event remain unknown. This study determined S. Enteritidis lineages before and after this pandemic event in Japan using multilocus sequence typing (MLST). Thirty S. Enteritidis strains were collected in Japan between 1973 and 2004, consisting of 27 human strains from individual episodes, a bovine strain, a liquid egg strain and an eggshell strain. Strains showed nine phage types and 17 pulsed-field profiles with pulsed-field gel electrophoresis. All strains had homologous type 11 sequences without any nucleotide differences in seven housekeeping genes. These MLST results suggest that S. Enteritidis with the diversities revealed by phage typing and pulsed-field profiling has a highly clonal population. Although type 11 S. Enteritidis may exhibit both pleiotropic surface structure and pulsed-field type variation, it is likely to be a stable lineage derived from an ancestor before the 1980s and/or 1990s pandemic in Japan.     

Cryptic occurrence of <Emphasis Type="Italic">Chattonella marina</Emphasis> var. <Emphasis Type="Italic">marina</Emphasis> in mangrove sediments in Probolinggo,East Java Province,Indonesia

Mangrove forests and adjacent creeks are known to be highly productive estuaries, which are partly supported by benthic microalgae that grow on the sediments. During surveys investigating the microalgal floras of mangrove swamps in the eastern part of Java Island, a mud sample unexpectedly included large numbers of a notorious fish killer, Chattonella-like motile cells, and its resting cysts. These motile cells were established as clonal cultures for further identification and physiological tests. The cysts were examined through palynological and molecular biological means. Identification based on light microscopy and ribosomal RNA gene sequences confirmed that these cells and cysts were Chattonella marina var. marina. While the strains were genetically identical to the temperate strains isolated from Japan and China, temperature experiments showed that the Indonesian strains possessed a high maximum quantum yield of photosystem II even after exposure to 34 °C, a temperature at which the Japanese strain could not survive. Salinity experiments showed adaptation of the strains to a salinity of 15. These findings, together with the discovery of populations of cysts in the mangrove sediment, highlight the tough and unique nature of the Indonesian strains, which are likely adapted to wide fluctuations of temperature and salinity in mangrove swamps, and pose a potential risk to fisheries in Indonesia.     

Phytoplankton surveys off the southern Myanmar coast of the Andaman Sea: an emphasis on dinoflagellates including potentially harmful species

A detailed list of dinoflagellate species for the Mergui Archipelago, off southern Myanmar, was compiled for the first time. This was based on surveys conducted in two seasons in pre- and post-monsoons, and on observations for both plankton and cyst assemblages. In the plankton samples, 57 and 26 dinoflagellate species were recorded, respectively, in the pre- and post-monsoons. Among the list from plankton and cyst samples, harmful species were found: red-tide species including Prorocentrum spp. and Alexandrium affine, and causative species of paralytic shellfish poisoning, e.g., Alexandrium tamiyavanichii and Gymnodinium catenatum, and of diarrhetic shellfish poisoning, e.g., Dinophysis spp. Therefore, although no incidences have been reported yet, attention must be paid to these harmful algal bloom (HAB) events in Myanmar where exploitation of fishery resources is drastically increasing. Other dinoflagellate compositions were also unique and may relate to the oceanographic system in this region. In the pre-monsoon, rather rare oceanic species of Ornithocercus spp. were found concurrently with neritic species, probably due to the extensive southwesterly current from the Indian Ocean. Throughout the seasons, diverse species of Protoperidinium were predominately recorded in both planktonic and sediment samples, indicating richness of prey organisms probably sustained by upwelling systems.     

A newly devised electric field screen for avoidance and capture of cigarette beetles and vinegar flies

A bifunctional electric field screen was proposed to physically exclude insect pests from warehouses. The screen consists of insulated iron wires (ICW) arranged in parallel and two earthed conductor nets placed on both sides of the ICW. A negative charge (0.1-8.0 kV) was applied to the insulated wires with a voltage generator to polarize an insulator sleeve used to cover the wire, negatively on the outer surface and positively on the inner conductor wire surface of the sleeve. The negative surface charge of the ICW caused an electrostatic induction in the earthed nets and an opposite charge on the net surfaces facing the ICW. An electric field formed in a space between the ICW and the earthed net, and the field strength increased in direct proportion to increasing voltages applied to the ICW. Adults of the test insects (cigarette beetle (Lasioderma serricorne) and vinegar fly (Drosophila melanogaster)) reaching the outer surface of the earthed net were deterred from entering the inside of the charged screen, whereas all insects immediately passed through the screen when the ICW was not charged. This avoidance was directly proportional to the increase in the voltage. In addition, the capability of the screen to capture insects that enter inside the screen was proven by introducing insects into the space between the ICW and the earthed net. Strong capture was observed when the ICW was negatively charged with more than 4.1 kV, under which conditions a short-term electric current (peaking at 0.3-0.6 μA, for 3 min) occurred transiently. This electric current was due to the release of electricity from the insects, giving a net overall positive charge to the insects, which therefore were attracted more strongly to the negatively charged ICW. A test using an attractant-set chamber showed that the insects were completely prevented from passing through the charged screen, in contrast to a rapid transfer of all insects when the screen was not charged. Thus, the present results show that the described screen is a promising physical tool for controlling insect pests in warehouses.     

Tryptophan synthesis block-associated sugar response, a physiological marker for rapid selection of Arabidopsis thaliana transformants, marked with feedback-inhibition-insensitive anthranilate synthase gene

The feedback-insensitive anthranilate synthase (AS) gene was used as a selection marker for transformants of Arabidopsis thaliana . The mutant gene ( mAS1-2 ) was constructed by substituting nucleotide at the effector-binding site of the intrinsic AS gene via PCR-mediated site-directed mutagenesis and flanked with the myrosinase promoter pyk10 to drive its expression during initial root elongation. This inducible gene cassette was first introduced into Agrobacterium tumefaciens and then delivered into A. thaliana by floral-dip inoculation. 5-methyltryptophan (5-MT) inhibited AS and suppressed seedling growth of wild type plants as a result of tryptophan starvation. With the addition of sucrose (10 mg/ml), 5-MT inhibited cotyledon opening and caused anthocyanin to accumulate in juvenile seedlings. The present mutant reversed the tryptophan starvation caused by 5-MT and blocked subsequent sugar responses. The sugar responses were detected in non-transformed plants grown on a selection medium containing 10 mg/ml of sucrose and 10 μg/ml of 5-MT after 3 days of incubation. Thus, true transformants could be selected after a short incubation, compared to the conventional kanamycin-selection method that did not eliminate all non-transformed plants.     

Identifying the origin of Corbicula clams using trace element analysis

The trace element contents of Corbicula clam shells collected from Japan, Russia, China, and the Republic of Korea were analyzed to determine their geographic origin. The crushed shells were decomposed with nitric acid–hydrogen peroxide, and the concentrations of 14 elements (Li, Mg, V, Mn, Co, As, Rb, Mo, Ba, Ce, Pb, U, Sr, and Ca) were measured by inductively coupled plasma mass spectrometry and inductively coupled plasma optical emission spectrometry. Some of the elements identified in samples displayed a geographic trend. The average content of manganese in Japanese samples was twice that of Russian samples. Conversely, the arsenic content in Japanese samples was approximately half of that in Russian samples. Linear discriminant analysis was applied to the data from Japanese and Russian samples, and a discriminant model was constructed. The discriminant model was used to determine the geographic origin of Corbicula clams produced in Japan, with 89.8 % of those identified as Japanese and 92.2 % of those identified as Russian being classified correctly. Therefore, trace element analysis of the shells of Corbicula clams is a useful technique for the identification of their country of origin.     

水稻种胚愈伤组织增殖力的形态学和组织学研究

对高愈伤组织增殖力品种Aikoku和低愈伤组织增殖力品种Moritawase进行了形态学和组织学的观察。在相同的培养条件下2品种表现出不同的形态和组织学特征,证实了基因型对愈伤组织增殖的作用。Aikoku的愈伤组织颜色淡黄、增殖较快,而Moritawase的愈伤组织则呈褐色且增殖速度慢。组织学的观察表明,Aikoku的种胚盾片上皮组织以及由上皮细胞起源的愈伤组织的细胞分裂活性高于Moritawase而且在上述2个品种中还观察到不同的细胞分化类型,在Aikoku的愈伤组织中观察到了带有表皮状结构的圆形细胞块,而在Moritawase的愈伤组织中却观察到了根状结构。在本研究中还观察到Aikoku中起源于中胚轴和盾片维管束的愈伤组织表现出与盾片上皮细胞起源愈伤组织不同的细胞学特征。