Functional changes in beta-lactoglobulin by conjugation with cationic saccharides

Bovine beta-lactoglobulin (beta-LG) was conjugated to each of three cationic saccharides [glucosamine (GlcN), chitopentaose (CPO), and chitosan (CHS)] by means of a water-soluble carbodiimide or by the Maillard reaction in an effort to improve the functional properties of beta-LG. The molar ratios of beta-LG to the cationic saccharide in the beta-LG-GlcN, beta-LG-CPO, and beta-LG-CHS conjugates were 2:1, 2:5, and 2:1, respectively. Fluorescence studies indicated that the conformation around Trp had changed in each conjugate and that the surface of each of the conjugates was covered with a saccharide chain. Structural analysis using monoclonal antibodies indicated that the conformation around (15)Val-(29)Ile (beta-sheet region) in beta-LG-GlcN and beta-LG-CPO had changed but that in beta-LG-CHS was maintained, whereas the conformation around (125)Thr-(135)Lys (alpha-helix region) in the conjugates had changed. The emulsifying activity of beta-LG was improved by conjugation with CPO or CHS, and aggregation of beta-LG was suppressed by conjugation with CHS. Reduction of the antigenicity and immunogenicity of beta-LG was achieved by conjugation with CHS.     

Identification and phylogenetic analysis of Babesia parasites in domestic dogs in Nigeria

The present study examined the presence of Babesia parasites in 104 domestic dogs in Nigeria. Sequentially, Babesia parasites infecting domestic dogs underwent genetic and phylogenetic analyses. The results of nested PCR based on the Piroplasmida 18S rRNA gene illustrated that 13.5% (14/104) of the samples were positive. The obtained positive samples determined the nucleotide sequences of the 18S rRNA genes. In the genetic and phylogenetic analyses, four of five nucleotide sequences were similar to Babesia canis rossi, and one sample exhibited a close similarity to a Babesia sp. isolated from a raccoon in Hokkaido, Japan. The present study revealed the widespread presence of B. canis rossi among domestic dogs in Nigeria.     

Chronic otitis externa with heat shock protein 70-positive intranuclear inclusion bodies in the ceruminous gland epithelium of a Chihuahua dog

A Chihuahua dog showed persistent itching in the right ear canal. Anti-inflammatory medicines and prednisolone were ineffective and total ear canal ablation was performed. Histological diagnosis was chronic otitis externa. Eosinophilic intranuclear inclusion bodies (Cowdry type A and full-type) were occasionally observed in the ceruminous gland epithelium. The inclusion bodies were negative for nucleic acid and ultrastructurally composed of fibrous structures (approximately 10 nm in width). Viral infection was initially suspected, but polymelase chain reaction tests did not detect the expected viral genes. Immunohistochemistry revealed that the inclusion bodies were positive for heat shock protein 70 (HSP70), suggesting that these bodies could be protein aggregates including HSP70. The etiology of this lesion has not been elucidated, but chronic inflammation may influence the cytoplasm-to-nuclear transportation of HSP70. To the best of our knowledge, this is the first report of canine chronic otitis externa with HSP70-positive intranuclear inclusion bodies.     

Development and evaluation of pyramiding lines carrying early or late heading QTLs in the indica rice cultivar ‘IR64’

The heading date is an important trait for determining regional and climatic adaptability in rice. To expand the adaptability of the indica rice cultivar ‘IR64’, we pyramided multiple early or late heading quantitative trait locus (QTLs) in the ‘IR64’ genetic background by crossing previously developed near-isogenic lines (NILs) with a single QTL for early or late heading. The effects of pyramiding QTLs were observed in three different climatic zones of the Philippines, Madagascar, and Japan. The early heading pyramiding lines (PYLs) headed 6.2 to 12.8 days earlier than ‘IR64’ while the late heading PYLs headed 18.8 to 27.1 days later than ‘IR64’. The PYLs tended to produce low grain yield compared to ‘IR64’. The low yield was not improved by combining SPIKE, which is a QTL that increases the number of spikelets per panicle. Conversely, ‘IR64-PYL(7+10)’ carrying Hd5 and Hd1 headed earlier, produced more tillers, and more panicles per m² than ‘IR64’, and mitigated the yield decrease in early heading. These results suggest that the effects of pyramided QTLs on heading date were consistent across various environments and PYLs could be used to enhance the adaptation of ‘IR64’ in other rice growing environments.     

The effect of tractor logging on forest soil compaction and growth ofShorea selanica seedlings in Indonesia

The hardness in soil was investigated in four forest areas in Indonesia after a crawler tractor passed over it several times. The hardness in soil, expressed by bulk density, was measured at various depth and various distances from the main track. Recovery of soil hardness was also examined. The bulk density increased markedly by the first and second pass of the tractor, but did not change after the fifth pass. The bulk density was highest just under the main and the log track, and decreased with an increase in distance from the track (0.5 to 1.5 m). The greatest increase in bulk density was observed at the depth of 0–5 cm. The soil hardness tended to show recovery within 9 years after logging, though not completely. Growth and root penetration of seedlingsShorea selanica decreased with the increase of soil hardness. We considered that the 2 tractor passes that caused bulk density of 1.3 g/cm³ is critical for the growth ofS. selanica’s seedling.     

Partial cloning of 17B-HSD1 from the Nile tilapia ovary and its expression pattern during spawning cycle

A 548 bp partial cDNA fragment of 17β-hydroxysteroid dehydrogenase (17β-HSD1) was obtained by RT-PCR from the ovary of Nile Tilapia. The expression of 17β-HSD1 was high from 0 to 11 days after spawning, but there was a sharp decline at the day of spawning (day 14) indicating its involvement in ovarian cycle.     

Artificially induced tetraploid masu salmon have the ability to form primordial germ cells

Diploid gametes generated with tetraploid animals are a stepping stone to improving chromosome manipulation techniques. However, artificially induced tetraploid individuals generally die soon after hatching. Diploid gametes could be induced by in vivo cultures of tetraploid primordial germ cells (PGCs) through germ-line chimera. In the present study, characteristics of PGCs were studied in inviable tetraploid masu salmon, Oncorhynchus masou. Histological observation of tetraploid embryos revealed that the same or smaller numbers of PGCs were observed and they migrate into the genital ridges as did diploid PGCs during gonadogenesis. By whole-mount in situ hybridization using vasa messenger RNA (mRNA), 4–35 vasa-positive signals were detected in a pair of genital ridges of tetraploids. By cytological observation of genital ridge cell suspensions, several large round cells were observed, some of which extended pseudopodia. They also contained large nuclei and round granules in their cytoplasm, characteristics of PGCs. As the results suggest that inviable artificial tetraploids have PGCs, we expect to achieve diploid gamete production through surrogate propagation and tetraploid fish production.     

Sex change in coral reef fish

Gonadal differentiation can take many forms in fish, ranging from gonochorism, where individuals directly develop as male or female and finally possess only testis or ovaries at sexual maturation, to hermaphroditism where the same individuals can produce mature male and female gametes at some time in their lives. Hermaphrodite fish are, thus, an excellent model for studying the plasticity of sex determination and differentiation in vertebrates. We have shown that sex steroids play a principal role in sex differentiation and sex change in fish. Our laboratory implements several fish models that undergo sex change from female to male or male to female or in both directions. In this review, we will briefly discuss recent advances in our understanding of the mechanism of sex change in coral reef fish.     

Molecular cloning and gene expression of the riboflavin-binding protein in the Nile tilapia, Oreochromis niloticus

Tilapia riboflavin-binding protein (RfBP) cDNA was isolated by library screening. Even though it shows only 31～35% similarity to the reported RfBPs, tilapia RfBP is conserved in amino acid residues that are known to be essential for its protein function. The exclusive expression of RfBP in the brain-pituitary-gonadal axis, as revealed by both RT-PCR and Northern blot, suggested a possible role for RfBP in fish reproduction.     

Gonadal morphogenesis and sex differentiation in cultured chub mackerel,Scomber japonicus

The timing of primordial germ‐cell (PGC) migration with regard to the gonadal anlagen, gonad formation and sex differentiation was examined histologically in the chub mackerel (Scomber japonicus) at 5–190 days post hatching (dph). At 5 dph, PGCs appeared on the peritoneal epithelium surface or in the mesentery, on the dorsal side of the abdominal cavity. By 10 dph, stromal cells around the PGCs proliferated. The gonadal primordium was formed by 15 dph. The gonadosomatic index was 0.01% at 30 dph and increased thereafter (0.32% in females and 0.04% in males at 160 dph). Ovarian differentiation occurred at 30–40 dph, indicated by ovarian cavity formation (elongation and fusion of the upper and lower ovarian edges). Meiosis was subsequently initiated. A few meiotic oocytes surrounded the cavity at 50 dph; most were in the perinucleolus stage at 60 dph and attained a diameter of 60–70 μm at 190 dph. Testicular differentiation occurred at 30 dph, indicated by the formation of the sperm duct primordium. Spermatogonia gradually proliferated, developing into spermatocytes at the chromatin–nucleolus stage (after 90 dph) and subsequently into spermatids and spermatozoa (160 dph). These data could aid the development of seeding and cell‐engineering technologies for scombrid fish.