Ontogeny of testicular inhibin and activin in ducks: An immunocytochemical analysis

The ontogeny of testicular inhibin/activin in ducks was investigated. Testicular localization of three inhibin/activin subunits (α, β_A and β_B) was determined in embryonic and newly hatched ducks from 12 days of incubation to 1 day of age, in immature ducks and in adult ducks. In the duck embryonic testis, positive α‐subunit immunostaining was first detected in the Leydig cells and Sertoli cells on day 15 of incubation, whereas β_A‐subunit and β_B‐subunit immunostaining were found in Sertoli cells and primary germ cells on day 18 of incubation. In 1 month old ducks, intense staining of α‐subunit was present in the seminiferous epithelium consistent with localization in Sertoli cells and primary germ cells, and the immunostaining of the β_A‐ and β_B‐subunit was also present in Sertoli cells and primary germ cells. Specific immunostaining with inhibin/activin α‐, β_A‐ and β_B‐subunits antisera occurred in Sertoli cells in the adult duck testes. In conclusion, it was shown that, in the duck testis, the majority of α‐, β_A‐ and β_B‐subunits are colocalized in Sertoli cells with a certain degree of staining in germ cells and the α‐subunit is present in Leydig cells of embryonic testes before day 18 of incubation. These results indicate that Sertoli cells and possibly germ cells in the embryonic testes of late stage of incubation and newly hatched ducks, immature ducks and mature ducks may produce bioactive inhibin dimers, inhibin A and inhibin B, as a possible regulator of follicle‐stimulating hormone secretion. Free inhibin/activin subunits and their dimers may also play an autocrine/paracrine role in the development of the testis and spermatogenesis. Furthermore, early onset of the α‐subunit in duck testes indicates that it may have an autocrine/paracrine effect on steroid hormones, which is important for sex differentiation.     

Induction of Systemic Resistance in Cucumber against Several Diseases by Plant Growth-promoting Fungi: lignification and Superoxide Generation

Five fungal isolates (Trichoderma, Fusarium, Penicillium, Phoma and a sterile fungus) from zoysiagrass rhizosphere that promote plant growth were tested for their ability to induce systemic resistance in cucumber plants against Colletotrichum orbiculare. Roots of cucumber plants were treated with these fungal isolates using barley grain inocula (BGI), mycelial inocula (MI) or culture filtrate (CF). Most isolate/inoculum form combinations significantly reduced the disease except BGI of Trichoderma. These fungal isolates were also evaluated for induction of systemic resistance against bacterial angular leaf spot and Fusarium wilt by treatment with BGI. Penicillium, Phoma and the sterile fungus significantly reduced the disease incidence of bacterial angular leaf spot. Phoma and sterile fungus protected plants significantly against Fusarium wilt. Roots treated with CFs of these fungal isolates induced lignification at Colletotrichum penetration points indicating the presence of an elicitor in the CFs. The elicitor activity of CFs was evaluated by the chemiluminescence assay using tobacco callus and cucumber fruit disks. The CFs of all isolates elicited conspicuous superoxide generation. The chemiluminescence activity of the CF of Penicillium was extremely high, and its intensity was almost 100-fold higher than that of other isolates. The chemiluminescence activity was not lost following treatment with protease or autoclaving or after removal of lipid. The MW 12,000 dialyzed CF fraction was highly effective in eliciting chemiluminescence activity. Chemiluminescence emission from cucumber fruit disks treated with Penicillium was the same as that obtained from tobacco callus, except that the lipid fraction also showed a high activity. Both the MW 12,000 fraction and the lipid fraction induced lignification in the epidermal tissues of cucumber hypocotyls.     

Development and promotion of laborsaving application technology for paddy herbicides in Japan

Total labor time on paddy rice has been decreasing year by year with the development and introduction of appropriate herbicides, especially ‘one‐shot' herbicides. However, in the case of application of granules, it is still difficult for a farmer to apply herbicides while carrying heavy power backpack sprayers. New formulation recipes and different application technology such as a throw‐in type formulation on jumbo granules or flowable has improved heavy workloads in comparison with granule application by using heavy power backpack sprayers. In addition, other advantageous points such as application volume and elimination of the drift problems of this new application technology were introduced and confirmed. As a result of this introduction, these formulations were used in 830 000 ha and reached 30% of the total treatment area in paddy rice in 2000.     

Biocontrol ofRhizoctonia damping-off of cucumber by non-pathogenic binucleateRhizoctonia

Three isolates of binucleateRhizoctonia (BNR) were tested for biological control of damping-off of cucumber seedlings caused byRhizoctonia solani AG 2-2 and AG 4. BNR isolates L2 (AG Ba) and W1 and W7 (AG A) provided protection of 58 to 71% against virulent isolate C4 of AG 4 and 64 to 75% protection against virulent isolate RH 65 of AG 2-2. Varying protection was provided to the seedlings by the BNR isolates against the virulentR. solani from the two AGs depending on their combination. The BNR isolates did not vary in providing protection to the seedling when tested against virulent C4 when both isolates were inoculated using three different methods,viz. in water agar, combination of water agar and soil and using soil alone. Protection of 58 to 71 % was provided by the isolates when inoculation was done on the hypocotyl using water agar, 62.8 to 75% using the combination of water agar and soil, and 75 to 85% when inoculation of both isolates was done in soil. Pre-incubation of BNR W7 or delayed inoculation of C4 (from 0.5 day to longer duration) using the different methods provided an increased protection to the seedlings to give complete inhibition of damping-off disease. Simultaneous inoculation of both BNR W7 and C4 using the three methods failed to provide protection to the seedlings. Among the BNR isolates, BNR W7 showed plant growth promotion in terms of significant increase in plant height (P=0.01) and fresh weight (P=0.05).     

Isolation and purification of a protective protein antigen of Erysipelothrix rhusiopathiae

The rapid growth and high survival rate of Erysipelothrix rhusiopathiae was determined using a culture of the bacterium in tryptic soy broth supplemented with 0.3% Tris-hydroxymethyl aminomethane and 0.1% Tween 80 (TT-TS broth). High concentrations of 64, 66 and 43 kDa proteins, which are associated with protection against E. rhusiopathiae infection in mice, were obtained by alkaline treatment of whole cells using 0.05-1 N NaOH. The supernatant of alkaline treated cells (alkaline extract; AE) was stable at alkaline or neutral pH. However, aggregates appeared at neutral pH in the absence of sodium dodecyl sulphate (SDS). A high yield of 64, 66 and 43 kDa proteins was obtained from strain Agata (serovar 5). The proteins were eluted from gel bands following SDS-polyacrylamide gel electrophoresis (SDS-PAGE) of the AE from strain Agata and designated P64 and P43. The amounts of P64 and P43 isolated were 0.7 and 0.3 mg/16 g of wet bacteria, respectively. In a mouse protection test, 50% protective doses (PD50) of P64 and P43 were 0.58 and 0.63 microgram, respectively. Upon Western blotting of the AE, both anti-P64 and anti-P43 antibodies reacted with the 64 and 43 kDa proteins. From these results, it is suggested that P64 is the most effective protective antigen and that P43 (43 kDa protein) is a degradation product of P64. Therefore, the 64 kDa structural proteins are associated with the induction of a protective activity against E. rhusiopathiae infection in mice.     

Ultrastructure of the thyroid gland of the one-humped camel (Camelus dromedarius)

Follicular epithelium of the thyroid gland of the one-humped camel was examined by light and electron microscopy. It consisted of a single type of epithelial cell which varied from flattened to columnar in shape. Follicular epithelial cells were characterized by the presence of markedly dilated cisternae of rough-surfaced endoplasmic reticulum, well developed Golgi apparatus, abundant small vesicles (150 nm to 200 nm in diameter) in the apical cytoplasm, and electron-dense colloid droplets measuring from 250 nm to 1600 nm. Follicular epithelial cells frequently showed apocrine secretion into colloidal lumens. Apocrine protrusions with smooth surface were dome-like or balloon-like structures and contained a fine granular matrix. These findings indicate that the morphological features of the follicular epithelial cells of the thyroid gland of the camel are essentially similar to those of mammals except for the presence of apocrine secretion, which is unique to the camel.     

Morphology and size of embryos recovered from superovulated cows.

A total of 691 normal embryos were recovered from 183 superovulated donor cows on the 5th and 6th days after the first insemination, and were examined for their morphology and size in relation to their developmental stage. There was no significant difference in the thickness of the zona pellucida, the diameter of the cell mass, and the overall diameter of the embryos among zygotes, 2-, 4-, 8- and 16-cell embryos, and morulae. In the blastocyst stage, however, the diameter of the cell mass and the overall embryo diameter were significantly greater and the zona pellucida was significantly thinner than in the earlier-stage embryos. The volume of the blastomere significantly decreased from zygote to morula in proportion to the increase in the number of blastomeres. The volume of the cell mass of 2-cell embryos was decreased by about 30% compared with that of zygotes and no increase in the volume of the cell mass was observed during the progression from 2-cell stage to morula. The diameter of the cell mass and the overall diameter of morulae recovered on the 6th day after the first insemination were significantly greater than those of morulae recovered on the 5th day.     

Fine structure of atrial natriuretic peptide(ANP)-granules in the atrial cardiocytes in the pig, cattle and horse.

In the pig, cattle and horse, the right and left atria and ventricles were examined by immunohistochemistry, and the right atrial and auricular cardiocytes were studied by transmission electron microscopy. Moreover, ANP-granules in the cardiocytes were analyzed by ultrastructural morphometry. Immunohistochemically, the most intensely ANP-reacted cardiocytes were localized in the right auricle, particularly more prominent in the pig and cattle than in the horse. Ultrastructurally, ANP-granules were located principally in the perinuclear region associated with the Golgi apparatus and throughout the sarcoplasmic layers. They were classified into 2 types, A and B. A-granule had a conspicuous electron-dense core enveloped by a membrane. B-granule had a fibrillogranular core enveloped by an indistinct membrane. By TEM equipped with a goniometer stage, a part of the limiting membrane of B-granules became visible as the stage tilt progressed. By ultrastructural morphometry, the number of each type of granules was significantly greater in the pig and cattle than in the horse. In the pig and cattle, the total number of both types of granules in the auricular cardiocytes was significantly greater than that in the atrial ones. The diameter of each type of granules was significantly larger in the pig and cattle than in the horse. The diameter of A-granules was significantly larger than that of B-granules.