Morphological features of the spermatic cord in the musk shrew (Suncus murinus) with special reference to extratesticular Leydig cells

Morphological features of the testicular artery and vein in the spermatic cord of the musk shrew (Suncus murinus) were evaluated by light microscopy, transmission electron microscopy, corrosion cast technique combined with scanning electron microscopy and immunohistochemistry. The vascular architecture in the spermatic cord of the musk shrew was simple. The testicular artery in the musk shrew was straight and accompanied by 1 to 3 branches of testicular vein. The testicular vein was also straight and anastomosed with each other in some points along its length, but it did not form a delicate pampiniform plexus. In the middle and distal portions of the spermatic cord, the tunica adventitia of the artery and vein was joined together to form a single connective tissue septum. Clusters of cells were found in this connective tissue septum in the middle portion of the cord. These cells were located close to the arterial wall and nerve endings, but they did not appear inside of neurium. They showed several typical characteristics similar to Leydig cells, and they were positive for 3beta hydroxysteroid dehydrogenase (HSD) antibody. Ultrastructural and immunohistochemical studies also indicated that the cells in cluster found in the vascular wall of the musk shrew spermatic cord may be equivalent to Leydig cells in testes. These extratesticular Leydig cells had characteristics of the active steroid-producing cell and seemed to be another source of testosterone.     

Invasion and colonization of mature apple fruit by Erwinia amylovora tagged with bioluminescence genes

Invasion and colonization of mature apple fruit by a transformant of Erwinia amylovora tagged with bioluminescence genes from Vibrio fischeri was examined. The transformant was deposited on cut surfaces of fruit stems, wounds on the shoulders and calyces, injured fruit-bearing twigs of harvested apple fruit, and cut fruit flesh. After incubation in closed stainless steel or plastic boxes at 25°C, fruit were periodically observed with a two-dimensional luminometer. The presence of the transformant in luminous areas was confirmed by isolating it on selective media. E. amylovora, when deposited in fruit stems: (1) can invade mature as well as immature apple fruit; (2) vertically and horizontally spreads and colonizes along vascular bundles, increasing its population; (3) reaches the calyx end and the flesh just under the exocarp within 3–4 days after inoculation; (4) when deposited on cut fruit flesh, irrespective of its maturity, can easily increase its population and survive 2–4 weeks or more at 25°C; and (5) even at the time of fruit maturation, can migrate within twigs rapidly and reaches the abscission layers between fruit-bearing twigs and fruit stems.     

Incidence of thiophanate-methyl resistance in Cercospora kikuchii within a single lineage based on amplified fragment length polymorphisms in Japan

We collected 247 isolates of Cercospora kikuchii from soybean seeds with typical purple stain symptoms from 15 prefectures in Japan. Of the 247 isolates, 93 were sensitive to thiophanate-methyl, a benzimidazole used to control this soybean disease; the remaining 154 were highly resistant to the fungicide. To examine genetic variability among the population of 247 isolates, we developed amplified fragment length polymorphism (AFLP) markers. An AFLP primer pair generated DNA fingerprint polymorphisms among the sample isolates, and with the unweighted pair-grouping method to cluster arithmetic means of the similarity coefficients among all pairs of the fingerprint patterns, the isolates were divided into four lineages (I to IV). Of the 247 isolates, 225 belonged to lineage I, including all isolates that were resistant to thiophanate-methyl. To determine whether the resistance of these isolates was related to mutations in the β-tubulin gene, we amplified partial nucleotide sequences of the gene from 29 representative isolates, including 12 that were resistant to thiophanate-methyl, by means of the polymerase chain reaction. The resistant isolates had identical nucleotide sequence with a one-step change at codon 198, in which the amino acid glutamic acid had been replaced by alanine. The evidence thus suggests that thiophanate-methyl resistance might have arisen in lineage I, the largest of the four lineages. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under accession numbers AB214511 to AB214515     

Production and Immunohistochemical Characterization of Monoclonal Antibodies Against Ovarian Antigen of the Common Tree Shrew (Tupaia glis)

To study the ovarian function of the common tree shrew (Tupaia glis), monoclonal antibodies (MAb) against its ovarian antigen were produced. Several positive hybridomas were cloned and MAb-T2C9 (IgG) was examined for reactivity. Strong immunoreactivity of MAb-T2C9 was localized within the cytoplasm of luteal cells and in the theca interna cells of Graafian follicles. MAb-T2C9 reacted also with testicular interstitial cells and adrenocortical cells, except those of the zona glomerulosa. Similar reactions were seen in the goat and sheep. Western blotting analysis of the ovine corpus luteum after reaction with MAb-T2C9 revealed a single positive band of ?60kDa. These findings suggest that MAb-T2C9 recognized protein molecules related to steroid synthesis.     

Surveillance of avian influenza viruses in Northern pintails (Anas acuta) in Tohoku District, Japan

Among winter migratory waterfowl, Northern pintails (Anas acuta), in one of the largest flocks in Tohoku district, northeast Japan, were surveyed for influenza A viruses at five wintering sites in three prefectures, viz., Aomori, Akita, and Miyagi. A total of 38 influenza A viruses were isolated from 2066 fecal samples collected during November 2006 through March 2007. The overall isolation rate was 1.84%. Eleven different subtypes were isolated, including nine H5N2, seven H6N8, seven H10N1, four H4N6, three H6N1, three H11N9, and one each of H1N1, H6N2, H6N5, H10N9, H11N1. Only the H4N6 subtype was detected during two successive months, November and December, from Lake Ogawara of Aomori prefecture. One wintering site, Lake Izunuma of Miyagi prefecture, was negative for virus isolation throughout the study period. During the sampling period, the highest virus isolation rate was in December (4.90%) followed by November (2.18%), January (0.91%), and February (0.30%). Virus isolation was negative for samples collected in March 2007. These results suggest that influenza viruses are introduced by Northern pintail when they migrate into Japan, but the viruses are not maintained in the flocks, most likely because the birds are not breeding during the winter. We believe that this relatively large data set creates a strong foundation for future studies of avian influenza virus (AIV) prevalence, evolution, and ecology in wintering sites, along with the role of Northern pintails in the spread of AIV during their migration from northern Russia and Asia to Japan.     

Distribution of two groups of melon landraces and inter-group hybridization enhanced genetic diversity in Vietnam

To understand the genetic diversity and differentiation of Vietnamese melon (Cucumis melo L.), we collected 64 landraces from the central and southern parts of the country and assessed molecular polymorphism using simple sequence repeat and random amplified polymorphic DNA markers. The Vietnamese melon was divided into seven cultivar groups, namely “Dua le”, “Dua vang”, “Dua bo”, “Dua gang-andromonoecious”, “Dua gang-monoecious”, “Dua thom”, “Montok”, and the weedy-type melon “Dua dai”. Among these, Dua le, Dua vang, Dua bo, and Dua gang-andromonoecious are cultivated on plains and they formed cluster II along with the reference accessions of Conomon and Makuwa. Based on genetic distance, Dua le and Dua vang were regarded as Makuwa and Dua bo and Dua gang-andromonoecious as Conomon. In contrast, Dua thom and Montok are cultivated in highlands, and they formed cluster III along with landraces from the southern and eastern foot of the Himalayas. Dua gang-monoecious which is commonly cultivated in the southern parts of Vietnam, exhibited the greatest genetic diversity, as explained by its possible origin through the hybridization between Dua gang-andromonoecious and Montok. Genetic differences in melon landraces between plains and highlands and hybridization between these two geographical groups have contributed to the enhancement of genetic diversity in Vietnamese melon.     

Effect of extracts from <Emphasis Type="Italic">narezushi</Emphasis>, a fermented mackerel product,on cholesterol metabolism in Wistar rats

Narezushi extract was separated into peptide and nonpeptide fractions by ion-exchange column chromatography. The narezushi extract and fractions were administered to rats in a diet enriched with lipid and cholesterol for 30 days. In the narezushi extract and nonpeptide fraction groups, increases in triglyceride, total cholesterol, and low-density lipoprotein-cholesterol levels in the plasma and accumulation of total lipids and triglyceride in the liver were suppressed, while both lipid and cholesterol fecal excretion were increased. In the peptide fraction group, these effects were also observed, except for the suppressing effect on liver lipid accumulation. Narezushi extract administration tended to increase fecal bile acids and promoted the activity of cholesterol 7α-hydroxylase, the rate-limiting enzyme in the synthesis of bile acid from cholesterol in the liver. However, the activity of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the rate-limiting enzyme in the cholesterol synthesis system in the liver, decreased due to regulation by the feedback of lipid transportation from diet to the liver. These results suggest that both the increase in cholesterol and bile acid fecal excretion and the promotion of cholesterol 7α-hydroxylase activities are related to the hypocholesterolemic effects of narezushi extract. Amino acids and organic acids, which are abundantly contained in the nonpeptide fraction, seemed to have more intensive hypocholesterolemic effects than peptides existing in the peptide fraction.     

Mass-processing method for aging otoliths of walleye pollock <Emphasis Type="Italic">Theragra chalcogramma</Emphasis> using black resin

ABSTRACT:   The reliability of a mass-processing aging method for walleye pollock Theragra chalcogramma was examined. The method is a half-otolith method, in which 10–20 otoliths are embedded simultaneously in black resin and the mass-processed half plane section is viewed under reflected light (black-resin method). Seasonal periodicity of otolith growth was confirmed by the monthly observations of the percent of translucent zone at the otolith edge, and interannual variation in age distribution detected the progression of dominant year-classes. These results support the assumption that the translucent zone is formed once a year. The black-resin method was compared to the break–burn method, which is the standard technique for this species. The estimated ages of these methods were not biased throughout the age range examined. The coefficient of variation in the black-resin method (4.9–5.7%) was similar to those of the break–burn method (3.5–5.5%). These results suggest that the black-resin method is reliable as an aging method for walleye pollock.