Rapid molecular typing of Listeria monocytogenes by pulsed-field gel electrophoresis

Pulsed-field gel electrophoresis (PFGE) is a highly discriminating tool for molecular typing, but the conventional PFGE protocol is time consuming. This paper describes a rapid method of PFGE for Listeria monocytogenes that yields results within 2 days.     

Analysis of bacterial communities in <Emphasis Type="Italic">Nannochloropsis</Emphasis> sp. cultures used for larval fish production

ABSTRACT:   Phytoplankton used in fish hatcheries is mass-cultured in the open air and usually contains large numbers of bacteria. In commercial fish production, the phytoplankton cultures are usually added into the larval rearing tanks; however, the numbers and types of bacteria introduced into the rearing tanks simultaneously are unknown. In this study, the bacterial community structures in Nannochloropsis sp. cultures were analyzed by using fluorescence in situ hybridization (FISH). A direct viable count (DVC)-FISH analysis was also performed as DVC is useful for the detection of actively growing cells. Total numbers of bacteria in Nannochloropsis sp. cultures ranged from 7.72 × 10⁵−2.39 × 10⁶ cells/mL. High proportions of the total bacteria (31.6–53.6%) in the Nannochloropsis sp. cultures showed growth potential. DVC-FISH analysis revealed that α-proteobacteria and the Cytophaga – Flavobacterium cluster were abundant in the bacterial community of actively growing cells. Thus, the high growth potentials of the distinct bacterial communities in Nannochloropsis sp. culture must influence the bacterial communities in larval rearing tanks.     

Effects of growth factors on development of fetal islet B-cells in vitro

To investigate the role of growth factors (epidermal growth factor [EGF], betacellulin, and activin A) in the development of islet B cells of rat fetal pancreatic explants in vitro, pancreases from rat fetuses at day 18 of gestation were cultured for 96 hr, with or without these growth factors. Culture medium was changed every 24 hr, and the level of insulin released in the culture medium was measured. After 72 hr of culture, pancreases were examined histologically. As a result, EGF promoted cell proliferation, but reduced B cell volume. Whereas, betacellulin and activin A inhibited cell division, but promoted increased B cell volume and insulin secretion, especially activin A, which stimulated insulin release in a time dependent manner. These results suggest that EGF, betacellulin, and activin A promote pancreatic cell proliferation, islet B-cell differentiation, and islet B-cell differentiation and functional maturation, respectively, and that EGF, betacellulin, and activin A, in this order, regulate islet B-cell neogenesis.     

Characterization of Andosols in Yakushima Island

We examined soils derived from volcanic ash of Kikai-Akahoya tephra on Yakushima Island, Japan, and classified them according to the Unified Soil Classification System of Japan, 2nd Approximation (USCSJ 2nd) and the World Reference Base for Soil Resources (WRB). Five pedons with horizons showing high (>20%) volcanic glass content were investigated. Soils developed under evergreen broad-leaved forests had high acid oxalate-extractable aluminum (Alo) and acid oxalate-extractable silicon (Sio) concentrations, and low acid oxalate-extractable iron (Fe)/dithionite-citrate-extractable Fe ratio. This indicates a warmer climate and less severe leaching conditions compared with soils developed under coniferous forests dominated by Cryptomeria japonica and grasslands dominated by Pseudosasa owatarii. All soils contained considerable amount of hydroxyl-Al-interlayered 2:1 clay minerals. The surface horizons of the pedons developed under the cool-temperate C. japonica forests contained smectite as a result of podzolization. However, the surface horizon of the pedon developed under cool-temperate P. owatarii grasslands did not contain smectite. All pedons belonged to the Kuroboku soils great group (USCSJ 2nd) and Andosols (WRB). Pedons in mountainous areas did not contain horizons with more than 6?g?kg^?1 of Sio and hence were classified as non-allophanic Andosols. In mountainous areas, it was observed that allophane formation was inhibited by Al leaching due to intense rainfall (>10,000?mm year^?1); Al consumption due to the formation of the Al-humus complex; and Al incorporation into the interlayers of vermiculite. The low soil water pH [pH(H₂O)] and leaching of silicon (Si) in mountainous areas would support these anti-allophanic effects.     

Determination and confirmation of S-RNase genotypes of apple pollinators and cultivars

Summary

We analysed the S-RNase genotypes of 23 crab apple (Malus spp.) pollinators and 102 cultivars of domestic apple (Malus pumila Mill.) by PCR amplification and digestion. Within the 23 pollinators, four pollinators, ‘Hopa’, ‘Jack’, ‘Pink Perfection’ and ‘Profusion B’, each had two unidentified S-RNase alleles. These cultivars should be useful pollinators for all domestic cultivars. Twenty-one of the domestic cultivars exhibited S-genotypes contrary to those expected from their supposed parentage, suggesting that one or both reported parents were wrong. We confirmed many of the S-RNase genotypes by pollination tests.     

Hot water treatment of Japanese pear trees is effective against white root rot caused by <Emphasis Type="Italic">Rosellinia necatrix</Emphasis> Prillieux

Hot water was dripped into the rhizosphere of Japanese pear trees (Pyrus serotina Rehd. grafted on P. betulifolia Bunge.) infested with the white root rot fungus Rosellinia necatrix Prillieux, to destroy the fungus. Isolates of R. necatrix from diseased roots of Japanese pear were vulnerable to water at temperatures above 35°C, and the fungus was eradicated from the colonized substrate when water at 35°C was provided for 3 days. The time required to eradicate R. necatrix decreased exponentially with increasing temperature. Japanese pear trees tolerated a temperature of 45°C without reduction in vigor. Field experiments demonstrated the practical use of hot water drip irrigation (HWD). HWD at 50°C completely destroyed white root rot mycelia on diseased roots, and many rootlets grew after the treatment. HWD at this temperature caused no injury to the trees. HWD of diseased orchard trees was assessed in Takamori and Iida in southern Nagano, Japan. The fungus recurred in two of four trees 28 months after treatment in Takamori and in two of ten trees 16 months after treatment in Iida. The new mycelia emerged on thick roots deep within the soil. Although there is a possibility of recurrence, HWD treatment is a practical control measure for white root rot.     

A novel genotyping technique for distinguishing between Flavobacterium psychrophilum isolates virulent and avirulent to ayu, Plecoglossus altivelis altivelis (Temminck & Schlegel)

We developed a simple genotyping method for Flavobacterium psychrophilum for analysing two single nucleotide polymorphisms (SNPs) in the gyrA gene and to distinguish between isolates that are virulent and avirulent to ayu, Plecoglossus altivelis altivelis (Temminck & Schlegel). The genotyping method is an on/off switch assay and is based on the polymerase chain reaction technique with phosphorothioated primers. We classified 232 isolates from four families of fish (i.e. Plecoglossidae, Osmeridae, Cyprinidae and Salmonidae) into four genotypes (G‐C, A‐T, A‐C and G‐T). The G‐C type isolates exhibited strong pathogenicity to ayu, whereas the A‐T and G‐T types did not show any pathogenicity to this species. The A‐C type exhibited no or weak pathogenicity to ayu. These results indicate that genotyping F. psychrophilum isolates with two SNPs from gyrA can clearly distinguish between isolates potentially harmful to ayu (G‐C type) and those that are potentially not harmful or less harmful (A‐C, A‐T and G‐T type). The on/off switch assay provides a quick, simple, and very powerful DNA genotyping technique for F. psychrophilum isolates.