Crystal structures and structural stabilities of the disulfide bond-deficient soybean proglycinin mutants C12G and C88S

The constituent subunits of seed storage protein 11S globulin have two disulfide bonds that are common among 11S globulins from legume and nonlegume seeds. In the case of the A1aB1b subunit of soybean 11S globulin, glycinin, Cys12-Cys45 and Cys88-Cys298 are observed by X-ray crystallography. The significance of these two disulfide bonds for structural stability was investigated by mutagenesis of Cys12 to Gly and of Cys88 to Ser. The disulfide bond-deficient mutants C12G and C88S could form the correct conformations identical to that of the wild-type proglycinin except in the vicinities of the mutation sites C12 and C88 as shown by their crystal structures. Thermal stability monitored by differential scanning calorimetry of the mutants indicated that the contribution of these disulfide bonds to the thermal stability of proglycinin A1aB1b is low, although there is a small difference in the extent of the contribution between the two disulfide bonds (Cys12-Cys45 > Cys88-Cys298). The contribution of Cys88-Cys298 to the resistance of proglycinin A1aB1b to proteinase digestion is higher than that of Cys12-Cys45. Possible effects of structure on the different properties of C12G and C88S are discussed.     

Use of an automatic cell-counting system with LED illumination for enumeration of marine bacteria

ABSTRACT:   The concentration of aquatic bacteria is basic information required to evaluate the status of environments and to assess bacterial contribution to material cycles. However, the standard direct counting method using epifluorescence microscopy (EFM) is tedious and there is variation in the counts among workers. Here an automatic counting system that consists of Bioplorer (BP) and image analysis has been applied to marine bacteria. BP is composed of a light-emitting diode (LED) illuminant, an optical unit, a driving stage and a charge-coupled device camera. In combination with fluorescent labeling and simplified membrane filtration, bacteria are enumerated automatically. The reproducibility, sensitivity and accuracy of the system were tested for natural marine bacteria, in comparison with EFM and flow cytometry (FCM). The counts obtained by BP showed good correlation with those obtained by EFM and FCM methods. The counts were significantly higher in inshore and oceanic samples, indicating high sensitivity with low background noise. Considering its reproducibility, objectivity, ease of use and compact size, BP can be used as a routine tool for counting aquatic bacteria in substitution for EFM or FCM.     

A morphological study off a thyroid carcinoma in a medaka, Oryzias latipes (Temminck & Schlegel)

A large mass protruding beyond the left operculum was found in an adult male medaka, Oryzias latipes (Temminck & Schlegel). Histologically, the mass was mainly composed of thyroid follicles measuring up to 292 μm in diameter. The follicular epithelium consisted of cuboidal or squamous cells. The follicles lacked colloidal material in the lumen. The thyroid tissue infiltrated into the kidney. Electron microscopically, the tumour cells were characterized by dilation and vesiculation of rough endoplasmic reticulum, which extended throughout the cytoplasm, and an irregular outline of the nucleus. Mitotic figures and partly degenerate cells were frequently observed. Based on the above features, the tumour was diagnosed as a follicular carcinoma of the thyroid, the first reported case in the medaka.     

A new method for evaluation of the resistance to rice kernel cracking based on moisture absorption in brown rice under controlled conditions

We developed and evaluated the effectiveness of a new method to detect differences among rice cultivars in their resistance to kernel cracking. The method induces kernel cracking under laboratory controlled condition by moisture absorption to brown rice. The optimal moisture absorption conditions were determined using two japonica cultivars, ‘Nipponbare’ as a cracking-resistant cultivar and ‘Yamahikari’ as a cracking-susceptible cultivar: 12% initial moisture content of the brown rice, a temperature of 25°C, a duration of 5 h, and only a single absorption treatment. We then evaluated the effectiveness of these conditions using 12 japonica cultivars. The proportion of cracked kernels was significantly correlated with the mean 10-day maximum temperature after heading. In addition, the correlation between the proportions of cracked kernels in the 2 years of the study was higher than that for values obtained using the traditional late harvest method. The new moisture absorption method could stably evaluate the resistance to kernel cracking, and will help breeders to develop future cultivars with less cracking of the kernels.     

Immune response to Echinococcus multilocularis infection in the mouse model: a review.

Echinococcus multilocularis is a cestode helminth which, along with E. granulosus, E. oligarthus and E. vogeli is a causative agent of hydatid disease in man. In the intermediate host (including man), cysts formed by the metacestode (larval stage) develop in the internal organs, causing functional impairment which often leads to the death of the host. In this review larval E. multilocularis infection in mice, the most popular experimental intermediate host, is examined, and the immune response to the organism is described in detail. Evidence is presented which suggests that cell-mediated immunity (CMI) plays a large role in suppression of larval growth. Congenitally athymic nude mice, and mice treated to remove thymocytes had high susceptibility to infection, while mice strains assessed as having high helper T lymphocyte function showed resistance to infection. The degree of antibody response shown by the host does not correlate with the susceptibility to E. multilocularis. Infection with E. multilocularis is accompanied by immunosuppression, manifested by inhibition of effector cell chemotaxis and receptor expression, suppressor macrophage and lymphocyte activity, decline in helper T-lymphocyte activity and immune-complex deposition.     

Research on targeting sources of alveolar echinococcosis in Japan

Echinococcus multilocularis is a fatal zoonotic parasite in the Northern Hemisphere. Recently, it has become endemic in many countries in Asia, especially in the northern island of Hokkaido in Japan. The increasing threat of public health due to alveolar echinococcosis has compelled researches for sensitive diagnosis and effective control. This paper reviews on the epidemiology, diagnosis and control of echinococcosis specifically in Japan. International collaborative responses by researchers and government initiatives such as mandatory reporting system for veterinarians who diagnose echinococcosis in dogs are presented. Successful control measures in Japan using anthelmintic fortified baits for foxes are described. Assessment of prevalence rates during control campaigns is analyzed favoring the use of intravital diagnosis rather than the traditional necropsy method from hunting or trapping activities of wild foxes. The novel concept of “endogenous development” by local resident volunteers towards sustainable control of echinococcosis is stressed.     

Molecular cloning and tissue distribution of uncoupling protein 1 (UCP1) in plateau pika (Ochotona dauurica)

Uncoupling protein 1 (UCP1) is present exclusively in brown adipose tissue, and contributes to body temperature control during cold exposure. We cloned UCP1 cDNA of plateau pika (Ochotona dauurica), a small, non-hibernating, diurnal lagomorph that inhabits in relatively cold climates and at high altitudes in Mongolia and in northern China. The nucleotide sequence of pika UCP1 was highly homologous to UCP1 of other species, and the deduced amino acid sequence had some common domains for UCP, including six mitochondrial carrier protein motifs and a putative purine-nucleotide binding site. RT-PCR and Western blot analyses revealed that both UCP1 mRNA and protein were expressed exclusively in the interscapular adipose tissue. These results suggest that pika UCP1 contributes to heat production in brown adipose tissue, as do those in other species.     

The use of tetracycline in anthelmintic baits to assess baiting rate and drug efficacy against Echinococcus multilocularis in foxes

Anthelmintic (praziquantel) baiting of wild red foxes against Echinococcus multilocularis infection was studied in a highly epizootic suburban area of Otaru, Hokkaido (the northern island of Japan) during the summer and autumn in the years 1999-2004. Acceptance of baits containing the biomarker tetracycline (TC) was evaluated. The prevalence of E. multilocularis infection in foxes before baiting (1999-2000) was 58% (88/153), whereas in the fourth year of bait distribution year (2004), it decreased to 11% (5/45). Analysis of TC marking in the teeth of foxes showed that 39% (77/195) of those captured after baiting were estimated to have consumed baits in the year of capture. Importantly, more juvenile (56%, 49/87) than adult foxes (26%, 28/108) were marked, indicating efficient baiting of juveniles, which tended to have a higher worm burden of E. multilocularis. Of 77 marked foxes, E. multilocularis and Alaria alata (monitored as the second indicator species of deworming) were not detected in 70 (90%) and 76 (99%) foxes, respectively. The results suggest effective deworming by bait consumption. However, it was also demonstrated that 9% of the marked foxes were infected or re-infected after bait consumption, suggesting high infection pressure and the importance of frequent baiting.