The detection and characterization of bacteria-sized protists in “Protist-free” filtrates and their potential impact on experimental marine ecology

Nuclepore filters of 0.6–1.0m pore size have been used to prepare protist-free water for a number of studies in microbial ecology. This procedure has been called into question by a recent study claiming that a significant portion of bacterial loss in filtrates could be due to uncharacterized predators passing through 0.6m filters. We were unable to directly observe protists in 0.6m filtrates using phase contrast, epifluorescence, or transmission electron microscopy. Using the culture techniques of rice grain enrichment and most probable number, however, we were able to observe and quantify several species of bacterivorous nanoflagellates that developed not only in 0.6m, but also in 0.4m seawater filtrates. The ability of predacious nanoflagellates to squeeze through bacteria-sized pores questions studies of bacterial production and chemical cycling that have assumed protist-free filtrates.     

Intestinal microflora in the deep-sea isopodBathynomus giganteus

Observations with the scanning electron microscope showed that the deep-sea isopodBathynomus giganteus harbors a dense microflora within the digestive tract. The gut microflora is composed of a diversity of microorganisms, including an unusually large bacterial morphotype that predominates almost exclusively in the anterior end of the hindgut. The majority of these large bacteria measured 1.9 m×8.5 m and many reached a size of 2.0 m×10.0 m.     

The Biometric Analysis of Bacterial Cells in Soil

The biometric analysis of bacterial cells in soil by light, fluorescence, and scanning electron microscopy showed that their average size is 0.8 m in diameter, 1.4 m in length, and 0.7 m³ in volume. In soil loci with enhanced microbiological activity (the rhizoplane of plants and the intestinal tract of soil invertebrates), the average size of bacterial cells was found to be 40% smaller than that of cells occurring in other parts of soil. This is the first experimental evidence showing that the metabolic activity of soil bacteria, their concentration, and allometric parameters are related.     

Clonal propagation of Callistemon by tissue culture techniques

Callus development in Callistemon viminalis was readily achieved when axillary buds derived from nodal tissue were placed in a medium containing macro- and micro-nutrients, sucrose (0.06 M), inositol (300 M), nicotinic acid (20 M), pyridoxine hydrochloride (3 M), thiamine hydrochloride (2 M), riboflavin (10 M), cytokinins (5 M) and auxins (0.1 M). The presence of benzylaminopurine (5 M) and p-chlorophenoxyacetic acid (0.1 M) promoted the most vigorous callus development and sprout formation. Rooting of nodal material was rare but occurred readily following the transference of sprouts developed on callus to a basal medium containing sucrose and salts. Root initiation was stimulated, however, by the presence of auxins. Chlorophenoxyacetic acid while stimulating root initiation repressed root growth. Indole butyric acid stimulated both root initiation and shoot growth at concentrations of 0.005 to 0.1 M. The treatment of choice for rooting and shoot growth was the addition of indole butyric acid at a concentration of 0.01 M.     

Saccharomyces cerevisiae 2-mum DNA. An analysis of the monomer and its multimers by electron microscopy.

Summary The non-tandem inverted duplication in the 2-m DNA of Saccharomyces cerevisiae has a length of 0.19 m and is located asymmetrically along the molecule. The majority of the dumb-bell structures that are formed upon denaturation and selfannealing of the 2-m monomer consists of the renatured inverted duplication sequences as double stranded stem and two single stranded loops of 0.67 m±0.06 m (S-loop) and 0.86 m±0.05 m (L-loop) length. Two additional size classes which comprised 5–10% of the measured molecules had contour lengths of around 1.7 m and 2.1 m. The smaller dumb-bells contained two S-loops and the larger dumb-bells contained two L-loops as was shown by heteroduplex mapping with an HindIII fragment from the L-loop. Two models which assume illegitimate or site specific recombination, are presented to explain the generation of double S-loop and double L-loop molecules. At least part of the 4-m and 6- circular molecules present in the yeast supercoiled DNA fraction are shown to be dimers and trimers of 2-m monomers, but often with inverted loop segments most probably due to intramolecular recombination between sequences of the inverted duplication.2-m DNA is used to indicate the supercoiled DNA fraction although in our measurements the average monomeric length is 1.9 mPart of this work has been presented at the Conference: The Genetics and Biogenesis of Chloroplasts and Mitochondria, Munich, August, 1976     

Enumeration of iron-oxidizing bacteria by the membrane filter technique

This work presents a simple methodology to enumerate ferrous-iron-oxidizing bacteria in solution, easily applicable in bioleaching industrial plants, because it does not require expertise or specific equipment. The enumeration is based on bacterial concentration by microfiltration through a membrane filter. The filter containing the bacteria is placed on an agarose plate containing ferrous sulphate for bacterial growth. No difference was observed for the enumeration of Acidithiobacillus ferrooxidans ATCC 19859 when either 0.1 or 0.22 m pore size membrane filters were used. However, when the technique was applied to bacteria present in pregnant leaching solution, the smaller bacteria present in these solutions passed through the 0.22 m pore size membrane. Therefore the number of bacteria could be underestimated if they are monitored and filtered using a filter with pore size greater than 0.1 m. The limit of detection of this technique was one ferrous-iron-oxidizing bacterium in the filtered solutions.     

Micropropagation and ecorestoration of Vanda spathulata,an exquisite orchid

Node explants collected from flowering plants of Vanda spathulata, an endemic and exquisite orchid of Peninsular India and Sri Lanka, were cultured in Mitra medium with combinations of 4.4–88.8 m 6-benzyl adenine (BA) and 0.0–114.2 m indole-3-acetic acid (IAA). Combinations of 44.4 m BA with 17.1 or 28.5 m IAA and 66.6 mM BA with 28.5 or 40.0 m IAA induced maximum formation of 12.6 and 12.1 shoots / node, respectively, in a 6-month period. Subcultured nodal explants produced maximum of 6.1 shoots at combinations of 22.2–44.4 m 21 BA and 5.7–28.5 m IAA. Rooting of shoots occurred in medium containing 75 g l^–1 banana pulp and 5.7 m IAA within 3–9 weeks. Plantlets of 2–5 cm length possessing two to five roots established easily in community pots at 80–90% rates without hardening. Community potted plants introduced into forest segments at Ponmudi and Palode in Southern Western Ghats of India established at a rate of 50–70%.     

Oocysts of Isospora ernsti n. sp. and Isospora blagburni n. sp. (Apicomplexa: Eimeriidae) from the black-capped bulbul,Pycnonotus xanthopygos

Oocysts of Isospora ernsti n. sp. and Isospora blagburni n. sp. are described from the black-capped bulbul Pycnonotus xanthopygos from Lincoln Park Zoo, Chicago, Illinois. The bird came from southwestern Africa seven years earlier. I. ernsti oocysts are ellipsoidal to bluntly ovoid, 28–38 × 23–31m (mean 34 × 28 m) and have a single-layered oocyst wall. Micropyle, oocyst residuum and polar granules are absent. Sporocysts are elongate ovoid, 24–30 × 11–16 m (mean 27×13 m). Stieda and substiedal bodies and sporocyst residuum are present. I. blagburni oocysts are spherical to subspherical. 21–28 × 19–26 m (mean 25 × 23 m) and have a single oocyst wall. Sporocysts are ovoid and 17–23 × 10–13 m (mean 20 × 12 m). Stieda and substiedal bodies and sporocyst residuum are present.     

The spermatozoon of Oikopleura dioica Fol (Larvacea,Tunicata)

Summary The spermatozoon of Oikopleura dioica is about 30 m long, with a spherical head, about 1 m wide, a 3 m long and 1 m wide midpiece, and a 25 m long tail with a tapered end piece. The head contains a nucleus with the chromatin volume limited to about 0.1 m³. A small acrosome is found in an anterior inpocketing, and a flagellar basal body in a posterior inpocketing of the nucleus. The midpiece contains a single mitochondrion with the flagellar axoneme embedded in a groove along its medial surface. The flagellar axoneme has the typical 9 + 2 substructure, and the basal body the typical 9+0 substructure. A second centriole and special anchoring fibres are absent.     

Stimulation of ethylene production by catecholamines and phenylethylamine in potato cell suspension cultures

The catecholamines (50 M dopamine, 50 M norepinephrine and 100 M epinephrine) and phenylethylamine (200 M) were found to stimulate ethylene production in potato suspension cultures. When 100 M amino-oxyacetic acid was added together with epinephrine, ethylene release returned to control levels. The endogenous 1-aminocyclopropane-1-carboxylic acid levels were increased in parallel with the release of ethylene, suggesting that the observed effect probably occurs via regulation of aCC synthase. Our results suggest that there is a link between these naturally occurring monoamines and ethylene in plants.Abbreviations AOA amino-oxyacetic acid - ACC 1-aminocyclopropane-1-carboxylic acid - DA dopamine - NE norepinephrine - E epinephrine - CA catecholamines - PEA phenylethylamine     

Morphology and structural organization of spiracles in femaleArgas (Persicargas) walkerae (Acari: Argasidae)

Scanning electron microscopical investigations of fractures and corrosion casts of the spiracles from femaleA. walkerae ticks revealed a four-part structure, consisting of spiracular plate, ostium and macula forming the external closure, followed by the subostial space and the vestibulum of regulable volume, as well as the atrial chamber as the innermost part from which the main tracheal trunks originate. On the average, the spiracular plate was 158 m long and 188 m at the broadest width. It consisted of a thin, highly perforated external and a thick internal layer, which enclosed the interpedicellar space with numerous stout pedicels. In its posterior region, the spiracular plate was covered by the macula, which was up to 80 m in length and 110 m in width. The interpedicellar cavity opened into the subostial space measuring 95.5 m in length and 159.6m in width, which proceeded into the 112-m long vestibulum. The roof of the vestibulum was flexible and could be everted and inverted. Inverted, the roof formed a quadratic bulge with numerous deep cuticular folds, which confined the lumen of the vestibulum either partially or completely. In corrosion casts, the roof was everted to a length of up to 89.3 m. In the posterior part of the vestibulum, as well as in the initial fourth of the artrial chamber, numerous anvil-, cone-or drop-like cuticular projections were arranged in wedge-like fashion. The atrial chamber was almost spherical with a diameter of 138.4 m. Five main tracheal trunks of different luminal diameter as well as numerous channels opened into the atrial chamber.     

Fine structure of early previtellogenic oocytes in Mugil (Liza) auratus Risso, 1810 (Teleostei,Mugilidae)

Summary Sequential cytological events at the onset of previtellogenesis were studied in oocytes from 12 m to 70 m in diameter of golden grey mullet. The main cytological changes observed (increase in size of cell, nucleus, nucleolus and increase in number of nucleoli, RNP particles, nuage and mitochondria) provide evidence for important synthetic processes in an early preparatory phase of oocyte development.Somatic cells (pre-follicle, follicle and thecal) are also described.     

Modelling of sulfur oxidation from elemental sulfur

An elemental S oxidation model has been developed which combines a maximum S release rate with modifiers for temperature and soil moisture conditions. This model has been combined with a pasture growth and CNSP nutrient cycling model to match S oxidation rate to pasture S demand. In two Southern Australian enviroments, 100m elemental S was superior to 200m particles whilst in Northern Australia the 200m particles were superior. These models can be used to match S release to plant demand.     

Cellular organization of the embryonic lobster heart

Summary The cellular organization of the embryonic heart of the lobster Homarus americanus was examined in 6-week and 6-month-old animals. The heart wall consists of an outer adventitial layer of fibroblast cells and an inner layer of transversely striated myocardial cells. Present in close association with the myocardium are cardiac neurons, hemocytes and so-called storage cells.Adjacent fibroblasts form fasciae adhaerentes and gap junctions. Adherent junctions also occur between fibroblasts and myocardial cells. Intercalated discs and differentiated membrane regions of close apposition (4 nm) occur between adjacent myocardial cells.The cardiac neurons form a ganglion that contains four small and five large somata. Regions of neuropil are present. Motor axons arising from the cardiac ganglion form neuromuscular synapses with the myocardial cells.The storage cells contain large inclusions and form gap junctions with the myocardial cells. They may supply nutritive material to the developing myocardium.The heart at 6 weeks is about 200 m long and 160 m wide. At 6 months, it is about 300 m long and 250 m wide. The myocardium at 6 weeks is one cell layer thick, and the cells are from 2–6 m in maximum width. At 6 months the myocardium is 2–4 cells thick, and the cells are from 6–12 m in width. Therefore, the myocardium grows by an increase in the number and size of the myocardial cells.     

Growth limitation of planktonic bacteria in a large mesotrophic lake

We studied nutrient-limitation of bacterioplankton growth in Lake Constance, a mesotrophic lake, between February and August in 1992. We amended 1-m filtrates with a single nutrient or nutrient combinations at 5 or 10 m final concentration, and the limiting nutrient or nutrient combination was inferred from the assay in which bacterial growth was most stimulated. The following nutrients were added individually or in combination: glucose, amino acids, peptone, and ammonium as C and N sources, and inorganic phosphate. From January until the beginning of the phytoplankton spring bloom in mid-April, C alone was growth-limiting. During the spring bloom a complex growth-limitation pattern occurred; first P was limiting, then for only 1 week C + N together, and thereafter P + C. During the clear-water phase with very low chlorophyll concentrations, P + C together limited bacterial growth again, interrupted by a period when C + N + P shortage caused a triple limitation. Later in the season, P + C were growth-limiting again. The growth efficiency (bacterial biomass produced/substrates used) on the basis of amino acid and carbohydrate used varied between 17 and 35%. The addition of various C and N sources indicated that the growth efficiency strongly depended on the quality of the substrates and the adaptation of the bacterial assemblages, for example, whether C and N originated from amino acids or glucose and ammonium.     

Immunocytochemical study of the hypothalamo-neurohypophysial system

Summary An antiserum cross-reactive against ovine neurophysins-I-II and -III has been used in conjunction with the immunoperoxidase histochemical procedure to localize the cells of the sheep paraventricular (PVN) and supraoptic nuclei (SON). In order to describe the topographical distribution of the SON and PVN a study was made on the serial sections cut (a) transversely from rostral to caudal positions and (b) sagittally from lateral to medial positions of the hypothalamus.The cells of the SON, when examined in the transverse aspect, extended approximately 1900 caudally and when examined in the sagittal plane were contained within a lateral-medial distance of 4830 . In each case the SON cells lay adjacent to the optic chiasm.As sections were cut transversely, the cells of the PVN first appeared in a rostral position defined as 0 and close to the ventral lining of the third ventricle. This general ventral and ventro-lateral distribution of cells maintained up to a caudal distance of approximately 840 . From positions 1260–2310 there was a dramatic dorsal shift of the PVN cells which by this time had also extended laterally. The total rostral-caudal distance occupied by the PVN cells was 3150 . That the lateral-medial distance occupied by the PVN was small (1050 ) was determined on examining the magnocellular nuclei in sagittal section.This work was financed by a grant awarded by the Medical Research Council of New Zealand     

The effect of monoterpenes on astaxanthin production by a mutant ofPhaffia rhodozyma

Summary The total pigment and astaxanthin content ofPhaffia rhodozyma increased with increasing concentrations -pinene up to 500 l -pinene/l. Above this concentration the total pigment and astaxanthin content as well as the biomass production decreased. The addition of 500 l -pinene/l increased the total pigment content from 1652 g/g to 2201 g/g and the astaxanthin content from 1554 g/g to 1883 g/g. A sharp decrease in maximum specific growth rate occurred above 150 l -pinene/l.     

Ultrastructure of Sporothrix schenckii treated with iodine-potassium iodide solution

The ultrastructural changes produced by iodine-potassium iodide solution on yeast cells of Sporothrix schenckii were investigated by transmission electron microscopy in order to clarify the mechanism of oral potassium iodide therapy for sporotrichosis. Yeast cells were dipped with solutions containing various concentrations of iodine. The rate of germination decreased markedly between the range of iodine concentrations from 0.63 g/ml to 5.0 g/ml. No significant ultrastructural changes were seen at the concentration of the iodine of 1.25 g/ml (80% germination) or less. In the concentration of 2.5 g/ml (50% germination), normal cells and degenerated cells coexisted. When the cells were treated with 5.0 g of iodine per ml (0% germination) or more, their interior structures were completely destroyed. It is assumed that iodine treatment of the organism causes rapid destruction in the whole cell.     

Endogenous phosphorylation of rat brain synaptosomal plasma membranes in vitro: Some methodological aspects

The time course of endogenous phosphorylation in vitro of total or separted synaptic plasma membrane proteins (SPM) has been correlated with that of hydrolysis of the phosphate donor (ATP) in the incubation medium. The ATP/SPM ratio in the medium was varied. In a low-ratio medium (7.5 M ATP; 2.2 g SPM/l) a complete hydrolysis of ATP occurred almost instantaneously as was measured by the release of free phosphate in and the disappearance of ATP from the medium. As a consequence, only a very short peak of phosphorylation, followed by dephosphorylation was observed. However, when higher ATP/SPM ratios were used (200 M ATP; 0.4 g SPM/l and 500 M ATP; 0.4 g SPM/l), the incorporation of phosphate into SPM proteins was linear for 20 sec, and the maximum level of phosphate incorporation was increased. Similar results were obtained after separation of³²P-labeled phosphoproteins by slab gel electrophoresis. However, analysis of the autoradiographs obtained fromone SPM preparation under different ATP/SPM ratios revealed dependence of phosphorylation of individual protein bands on the conditions used.     

The contribution of size-fractionated plankton to biomass and primary production of phytoplankton in saline–alkaline ponds

Primary productivity, biomass and chlorophyll-a of size fractionated phytoplankton (<0.22 m, <3 m, <8 m, <10 m, <40 m, <64 m, <112 m and <200 m) were estimated in 6 ponds and 5 experimental enclosures. The results showed that the planktonic algae less than 10 m are important in the biomass and production of phytoplankton in saline–alkaline ponds. The production of size fractionated phytoplankton corresponding to <112 m, <10 m and <3 m in saline–alkaline ponds were 10.5 ± 6.6 , 8.6 ± 5.4 and 0.33 ± 0.1 mgC l^–1 d^–1, respectively. Mean community respiration rate was 1.80 ± 0.73, 1.69 ± 0.90 and 1.38 ± 1.12 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) were 1.61, 8.30 and 0.33 mgC l^–1 d^–1, respectively. The ratio of those to the total phytoplankton production was 15%, 79% and 3%, respectively. The mean respiration rate of the different size groups was 0.11, 0.31 and 1.38 mgC l^–1 d^–1; the ratio of those to total respiration of phytoplankton was 6%, 17% and 77%, respectively. The production of size-fractionated phytoplankton corresponding to <200 m, <10 m and <3 m in enclosures was 2.19 ± 1.63, 2.08 ± 1.75 and 0.22 ± 0.08 mgC l^–1 d^-1, respectively. Mean community respiration rates were 1.25 ± 1.55, 1.17 ± 1.42 and 0.47 ± 0.32 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton was 0.11, 1.86 and 0.22 mgC l^–1 d^–1, respectively. The ratio of those to the total production of phytoplankton was 5%, 85% and 10%, respectively. The mean respiration rate of different size groups were 0.08, 0.72 and 0.46 mgC l^–1 d^–1, the ratio of those to total respiration of phytoplankton was 6%, 57% and 37%, respectively. The concentrations of chlorophyll-a of the phytoplankton in the corresponding size of micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) plankton in the experimental ponds were 19.3, 98.2 and 11. 9 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 15%, 76% and 9%, respectively. The concentrations of chlorophyll-a of phytoplankton micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton in enclosures were 1.7, 34.3 and 3.0 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 4%, 88% and 8%, respectively.