Fluorescence imaging of local membrane electric fields during the excitation of single neurons in culture.

The spatial distribution of depolarized patches of membrane during the excitation of single neurons in culture has been recorded with a high spatial resolution (1 micron2/pixel) imaging system based on a liquid-nitrogen-cooled astronomical camera mounted on an inverted microscope. Images were captured from rat nodose neurons stained with the voltage-sensitive dye RH237. Conventional intracellular microelectrode recordings were made in synchrony with the images. During an action potential the fluorescence changes occurred in localized, unevenly distributed membrane areas, which formed clusters of depolarized sites of different sizes and intensities. When fast conductances were blocked by the addition of tetrodotoxin, a reduction in the number and the intensities of the depolarized sites was observed. The blockade by tetrodotoxin of voltage-clamped neurons also reduced the number of depolarized sites, although the same depolarizing voltage step was applied. Similarly, when a voltage-clamped neuron was depolarized by a constant-amplitude voltage step, the number of depolarized sites varied according to the degree of activation of the voltage-sensitive channels, which was modified by changing the holding potential. These results suggest that the spatial patterns of depolarization observed during excitation are related to the operations of ionic channels in the membrane.     

Gluconobacter thailandicus sp. nov., an acetic acid bacterium in the alpha-Proteobacteria

Four strains of acetic acid bacteria were isolated from flowers collected in Thailand. In phylogenetic trees based on 16S rRNA gene sequences and 16S-23S rDNA internal transcribed spacer (ITS) region sequences, the four isolates were located in the lineage of the genus Gluconobacter and constituted a separate cluster from the known Gluconobacter species, Gluconobacter oxydans, Gluconobacter cerinus, and Gluconobacter frateurii. In addition, the isolates were distinguished from the known species by restriction analysis of 16S-23S rDNA ITS region PCR products using three restriction endonucleases Bsp1286I, MboII, and AvaII. The DNA base composition of the isolates ranged from 55.3-56.3 mol% G+C. The four isolates constituted a taxon separate from G. oxydans, G. cerinus, and G. frateurii on the basis of DNA-DNA similarities. Morphologically, physiologically, and biochemically, the four isolates were very similar to the type strains of G. oxydans, G. cerinus, and G. frateurii; however, the isolates were discriminated in their growth at 37 degrees C from the type strains of G. cerinus and G. frateurii, and in their growth on L-arabitol and meso-ribitol from the type strain of G. oxydans. The isolates showed no acid production from myo-inositol or melibiose, which differed from the type strains of the three known species. The major ubiquinone homologue was Q-10. On the basis of the results obtained, Gluconobacter thailandicus sp. nov. was proposed for the four isolates. The type strain is isolate F149-1(T) (=BCC 14116(T)=NBRC 100600(T)=JCM 12310(T)=TISTR 1533(T)=PCU 225(T)), which had 55.8 mol% G+C, isolated from a flower of the Indian cork tree (Millingtonia hortensis) collected in Bangkok, Thailand.     

Streptomyces endocoffeicus sp. nov., an endophytic actinomycete isolated from Coffea arabica (L.)

An aerobic, non-motile, Gram-stain positive actinomycete, designated strain CA3R110^T, was isolated from the surface-sterilised root of Coffea arabica L. collected from Lampang Province, Thailand. 16S rRNA gene sequence analysis indicated that strain CA3R110^T was a member of the genus Streptomyces and showed the closest similarities to Streptomyces buecherae AC541^T (99.2%), followed by Streptomyces rapamycinicus NRRL B-5491^T (99.1%), Streptomyces luteoverticillatus NBRC 3840^T (99.1%), Streptomyces coerulescens NBRC 12758^T (99.1%), and Streptomyces iranensis HM 35^T (99.0%). Strain CA3R110^T contained LL-diaminopimelic acid in cell peptidoglycan, MK-9(H₆), and MK-9(H₈) as major menaquinone, iso-C_16:0, iso-C_15:0, C_16:0 as major fatty acids. Diphosphatidylglycerol, phosphatidylethanolamine, hydroxy-phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositolmannoside were detected in the cell. The chemotaxonomic characteristics possessed the typical properties of the genus Streptomyces. A low digital DNA–DNA hybridization (<?55.7%) and average nucleotide identity-blast (ANIb) (<?92.2%) values revealed that strain CA3R110^T could be distinguished from any known Streptomyces species. With the differences in phenotypic and genotypic data, strain CA3R110^T represents a novel species of genus Streptomyces, for which the name Streptomyces endocoffeicus sp. nov. is proposed. The type strain is CA3R110^T (=?TBRC 11245^T?=?NBRC 114296^T).

     

Patterns and possible environmental controls of skeletogenesis of Porites lutea,South Thailand

Eleven fringing reef sites were investigated over a distance of about 50 km in the Phuket Region. There is a wide range in exposure to wave energy, and also water turbidity across the area. Annual increments of growth of shallow-water reef-front colonies of Porites lutea were calculated for the period November 1984 November 1986 using seasonal fluorescent banding (revealed with ultraviolet light) and Alizarin staining. Measurement of linear extension rate, skeletal bulk density, calcification rate, polyp numbers per unit area and colony surface morphology were made and compared. Linear extension rate and skeletal bulk density are inversely related within and between reef sites. Linear extension rate decreases and bulk density increases along a gradient of increasing hydraulic energy of the setting. Calcification (the product of linear extension rate and bulk density), although varying slightly from site to site, does not appear to relate to any obvious environmental inshore-offshore gradient. Skeletal bulk density is the most sensitive discriminator between reef sites, and we suggest that hydraulic energy of the setting is the main control on these spatial variations in skeletogenesis.     

Mannose transport in Amphora coffeaeformis var. perpusilla (Bacillariophyceae).

Amphora coffeaeformis var. perpusilla is able to concentrate but not metabolize mannose. The sugar, which can be accumulated against a concentration gradient, is taken up only after a short lag. Preincubation with glucose allows immediate mannose uptake. Substances that reduce the ATP content of the diatom cells also inhibit mannose uptake as do sugars of similar stereochemical configuration. Mannose appears to act as a non-metabolizable analogue of glucose in these respects.     

ISOLATION AND PHYSIOLOGICAL STUDIES ON THREE ISOLATES OF AMPHORA (BACILLARIOPHYCEAE)1

Three clones of the diatom Amphora were euryhaline, able to grow autotrophically at 160 lx (0.001 ly/min) and heterotrophically on glucose and fructose. Furthermore 2 clones grew on glutamate and feast extract. Light-limited growth of individual clones was stimulated by glycerol, galactose, lactate, acetate, aspartate and asparagine, although mannose torn inhibitory at low and high light levels. The half-saturation constant for growth of A. coffeaefomis var. perpusilla Grunow (Cleve) with glucose was 25 μM. Heterotrophic growth rate of this organism became saturated with respect to glucose at 0.5 mM.     

Purification and properties of carotene 15,15'-dioxygenase of rabbit intestine

Carotene 15,15'-dioxygenase, which oxidizes carotenoids to retinal, has been purified up to 200-fold from rabbit intestine by ammonium sulfate fractionation, heat treatment, and acetone precipitation. With beta-apo-10'-carotenol as the substrate, the purified enzyme has a pH optimum of 7.8, a K(m) of 6.7 x 10(-5) m, and a V(max) at 37 degrees C of 9 nmoles of retinal/mg protein/hr. The purified enzyme is inhibited by ferrous ion-chelating agents such as alpha,alpha'-dipyridyl and o-phenanthroline, and by sulfhydryl-binding agents such as iodoacetamide, N-ethylmaleimide, and p-chloromercuribenzoate. The latter inhibitory effects are reversed by reduced glutathione. The cleavage of beta-apo-10'-carotenol is competitively inhibited by its acetylenic analog, 15,15'-dehydro-beta-apo-10'-carotenol. The enzyme is present in the intestinal mucosa of several mammals, the chicken, the tortoise, and a freshwater fish, but it is absent from cat intestinal tissue.