Developing a national food defense guideline based on a vulnerability assessment of intentional food contamination in Japanese food factories using the CARVER+Shock vulnerability assessment tool.

The awareness of food terrorism has increased following the September 11, 2001 terrorist attacks in New York City, United States, and many measures and policies dealing with this issue have been established worldwide. Suspected deliberate food-poisoning crimes have occurred in Japan, although they are not regarded as acts of food terrorism. One area of concern is that the small- to medium-sized companies that dominate Japan's food industry are extremely vulnerable to deliberate food poisoning.

[Tentative food defense guidelines for food producers and processors in Japan].

Objectives: With increasing global interest in intentional food contamination, expert meetings have been held by the G8, while the U.S. government has proposed policies for preventing food terrorism and intentional contamination.

Identification of the sperm motility-initiating substance in the newt, Cynops pyrrhogaster, and its possible relationship with the acrosome reaction during internal fertilization.

Motility initiation is a key event during internal fertilization of female-stored sperm, although the underlying mechanisms remain unclear. In internally fertilizing urodeles, quiescent sperm initiate motility on the surface of the egg-jelly, a thick extracellular matrix that accumulates around the egg in oviduct. By immunizing mice with egg-jelly extracts, we successfully generated an alpha34 monoclonal antibody (mAb) which neutralized sperm motility-initiating activity in the egg-jelly of the newt, Cynops pyrrhogaster, in a dose-dependent manner.

Two states of active spermatogenesis switch between reproductive and non-reproductive seasons in the testes of the medaka, Oryzias latipes.

Seasonal change in spermatogenesis was examined in the restricted spermatogonium-type testes of a teleost, Oryzias latipes. Histological observation revealed that the number of each stage of germ cells during most of the non-reproductive season, from October to January (O-J period) was nearly half of that during the reproductive season, from May to July (M-J period), except for type B spermatogonia (B-gonia), which was actually equal. As a result, the ratio of primary spermatocytes (P-cytes) to B-gonia was remarkably small in the O-J period.

Identification of egg-jelly substances triggering sperm acrosome reaction in the newt, Cynops pyrrhogaster.

Our previous studies have shown that the acrosome reaction (AR) occurs in egg-jelly of the Japanese newt, Cynops pyrrhogaster. This is analogous to the substances of echinoderms but distinct from those of many other vertebrates derived from the egg envelope or its derivative, the zona pellucida. To identify the AR-inducing substances in newt egg jelly, a monoclonal antibody (mAb) was generated against the jelly by screening the culture supernatants to find the one that best neutralized the AR-inducing activity of the jelly substance.

Egg jelly of the newt, Cynops pyrrhogaster contains a factor essential for sperm binding to the vitelline envelope.

The acrosome reaction of newt sperm is induced at the surface of egg jelly and the acrosome-reacted sperm acquire the ability to bind to the vitelline envelope. However, because the substance that induces the acrosome reaction has not been identified, the mechanism by which the acrosome-reacted sperm bind to the vitelline envelope remains unclear. We found here that a Dolichos biforus agglutinin (DBA) specifically mimicked the acrosome reaction immediately upon its addition in the presence of milimolar level Ca(2+).

The outermost layer of egg-jelly is crucial to successful fertilization in the newt, Cynops pyrrhogaster.

The significance of egg-jelly layers in internal fertilization was evaluated in the newt, Cynops pyrrhogaster. In this species, six egg-jelly layers, J1, J2, J3, J4, J5 and the outermost J6 layers, are accumulated on the surface of the fertilizable eggs in pars convoluta of the oviduct. When a large number of sperm (about 6 x 10(5)) were placed on eggs having different numbers of jelly layers, all the eggs were fully fertilized, although many of the eggs developed abnormally.

Ordered progress of spermiogenesis to the fertilizable sperm of the medaka fish, Oryzias latipes, in cell culture.

Spermiogenesis is significant for producing sperm with equipment for achieving fertilization. Although multiple events occur in a particular order during spermiogenesis, it is unclear how the timing of those events is controlled. In the present study, we found that primary spermatocytes obtained from the spermatogenic testes of Oryzias latipes synchronously differentiated into sperm without contact with somatic cells in culture.

Characteristics of sperm motility induced on the egg-jelly in the internal fertilization of the newt, Cynops pyrrhogaster.

Most urodeles undergo internal fertilization and sperm are directly inseminated onto the surface of egg-jelly. Feature of sperm motility induced on the egg-jelly was examined in the newt, Cynops pyrrhogaster. When sperm were directly inseminated onto an egg-jelly, sperm motility was immediately induced on its surface.

CpZPC, a newt ZPC molecule, localizes to the inner surface of the egg envelope.

Zona pellucida-3 is an essential molecule for the binding of sperm to the egg envelope and for induction of the acrosome reaction in mice. Its homologous molecules, ZPCs, have been widely identified in the eggs of many vertebrates, except for urodeles. In this study, to investigate the participation of ZPC in newt fertilization, we cloned the cDNA of newt ZPC from the ovary of Cynops pyrrhogaster by reverse transcription polymerase chain reaction (RT-PCR).

The urodele egg-coat as the apparatus adapted for the internal fertilization.

Fertilization is a significant event for reproducing offspring. It is achieved under a species-specific environment, which influences the conditions to assure the successful fertilization in some cases. Several studies about the basic mechanism of fertilization suggest that the fertilization mechanism is modified among species to be suited for the fertilization environment.

Egg transport in the coelom of the newt cynops pyrrhogaster. I. The ovulated egg is transported to the ostium by the ciliary movement of coelomic epithelial cells.

We investigated the mechanism of egg transport in the newt not only by inserting various conditioned eggs into the recipient's body but also by placing them on the coelomic epithelia of the opened body cavity in the adult female newt. Most of the inserted coelomic eggs were oviposited, while 4 of 14 inserted de-jellied uterine eggs and 3 of 10 inserted de-jellied fertilized eggs were oviposited. The coelomic eggs placed on the coelomic epithelia were transported toward the ostium and entered the ostium.

The activity for the induction of the sperm acrosome reaction localises in the outer layers and exists in the high-molecular-weight components of the egg-jelly of the newt, Cynops pyrrhogaster.

Localisation of the acrosome reaction inducing activity in egg-jelly was examined in the newt, Cynops pyrrhogaster. The jelly has six layers: the J0, J1, J2, J3, J4 and st layers. Jelly was mechanically dissected and placed on a Millipore filter.

Egg-jelly structure promotes efficiency of internal fertilization in the newt, Cynops pyrrhogaster.

Egg-jelly is composed of a network of fibrous components and contains substances regulating the sperm-egg interaction. Many studies on the latter have been conducted, whereas the role of the egg-jelly structure in fertilization has not yet been fully assessed. In this study, we examined the fertilization efficiency in the presence and absence of the structure around the egg of the newt, Cynops pyrrhogaster, using a gelatin gel system.

Activation of Xenopus Eggs by an Extract of Cynops Sperm: (fertilization/egg-activation/polyspermy block/sperm protease).

An extract obtained from Cynops sperm induced the activation of both Cynops and Xenopus eggs with accompanying changes in the potential of the egg membrane that were quite similar to those caused by the Cynops sperm. The activation-inducing properties of the extract were abolished by treatment with proteinase K or by heating (60°C, 15 min) and were associated with a protease activity against peptidyl Arg-MCA substrates. The activation of Xenopus eggs by the extract was inhibited by those substrates, or by protease inhibitors, aprotinin or leupeptin.

Cloning of Protamine cDNA of the Medaka (Oryzias latipes) and Its Expression during Spermatogenesis: (protamine/gene expression/spermatogenesis/in situ hybridization/medaka).

Protamines or sperm specific basic proteins are highly basic low molecular weight proteins that substitute histones in the chromatin of sperm during spermatogenesis. They condense sperm DNA into a highly compact, stable and inactive complex. In this study, cDNA of protamine of the medaka, Oryzias latipes, was cloned to elucidate the molecular mechanisms involved in spermatogenesis.

Time Sequence of Early Events in Fertilization in the Medaka Egg: (micropyle/exocytosis/sperm movement/calcium release/medaka egg).

The time sequence of early events in fertilization was examined in eggs of the medaka Oryzias latipes. The mean time after insemination required for sperm attachment to the egg surface through the micropyle depended on sperm concentrations. It was 3 ± 1 sec with a range from 1 to 6 sec after insemination when concentration of spermatozoa was high (about 2 × 10 /ml at 23°-25°C).

Fertilization of newt coelomic eggs in the absence of jelly envelope material.

Fertilization ofCynops pyrrhogaster (Japanese newt) coelomic eggs was studied in the absence of jelly envelope material or synthetic high polymer. An undiluted sperm fluid from the vas deferens fertilized coelomic eggs in the absence of the jelly envelope material or synthetic high polymer. The fertilized eggs developed beyond gastrulae and formed tail bud embryos.

Fertilization of the eggs ofCynops pyrrhogaster (japanese newt) after immersion in water.

Cynops pyrrhogaster oviducal eggs with and without jelly envelopes (jelly egg and dejellied egg respectively) were immersed in water, and then inseminated artificially. After 1 h of immersion in water, more than half the dejellied eggs were fertilized and developed, but no jelly eggs developed. The rapid decrease in the ability of jelly eggs to be fertilized after immersion in water is not due to a deficiency in the egg.

Immunocytochemical location of vitellin in the egg of the silkworm,Bombyx mori, during early developmental stages.

Vitellin was purified from eggs of the silkworm,Bombyx mori, by a new method in which vitellin was extracted from isolated yolk granules. The purified vitellin had a molecular weight of 540,000. An antibody against purified vitellin was prepared in rabbits.

IMMUNOCHEMICAL STUDY ON THE SPECIES SPECIFICITY OF SPERM-BINDING PROTEIN FROM THE SURFACE OF THE SEA URCHIN EGG.

Immunological species specificity of sperm-binding protein from eggs of the 4 sea urchin species, Hemicentrotus pulcherrimus, Pseudocentrotus depressus, Anthocidaris crassispina and Temnopleurus toreumaticus, was examined by means of double immuno-diffusion technique in agar. Ca-soluble fraction of sperm-binding protein which is considered to be responsible for initial sperm-egg bonding at fertilization, has species-specific antigenic component. Correlations in antigenic constituents among the 4 species are described.