Ultrasensitive chemosensory responses by a protozoan to epinephrine and other neurochemicals.

A behavioral assay was developed based on differential tendency of a protozoan to attach to an agar gel containing the test substance. The heterotrophic marine dinoflagellate Crypthecodinium (Gyrodinium) cohnii responded negatively (less tendency to attach) to epinephrine at concentrations above 5 X 10(-15)M and to norepinephrine at concentrations above 5 X 10(-9)M. Response to choline as choline H2 citrate, choline bitartrate, and choline chloride was negative above 10(-7)M, but response to the choline analog carbachol was positve (greater tendency to attach) in the range 5 X 10(-6) to 5 X 10(-4)M. Other responses to neurochemicals at comparable concentrations were: dopa, betaine, and glycine--positive; L-glutamic acid, tryptophan, putrescine, and taurine--negative. Serotonin was inert, responses to tyrosine and gamma-aminobutyric acid were variable, and phenylalanine (6 X 10(-3)M) and 5-hydroxytryptophan (5 X 10(-4)M) were negative only at the highest concentrations tested.

Utilization of Fe3+ by the inshore colorless marine dinoflagellate Crypthecodinium cohnii.

A Puerto Rican isolate of the colorless dinoflagellate Crypthecodinium cohnii was grown in a defined marine medium. Fe was added as Fe(NH4)2(SO3)2 - 6H2O (2.0 mg%), FeCl3 - 6H2O (1.0 mg%) or a particulate slurry prepared from FeCl3 + KOH, along with varying concentrations of several chelators. Heavy growth at pH 7.5-7.7 occurred with salicylhydroxamic acid, aurintricarboxylic acid, EDTA, NTA, and humic acid; and at pH 7.9-8.1 with SHAM and ATA. Moderate growth occurred at pH 7.5-7.7 with sulfosalicylic acid, dipicolinic acid, pyrocatecholdisulfonic acid, hexanohydroxamic acid, L-histidine, and at pH 7.9-8.1 with 1-naphthohydroxamic acid, EDTA, NTA. Slight growth occurred at pH 7.5-7.7 with benzohydroxamic acid, 1-naphohydroxamic acid, 2.6-dipicolinic acid N-oxide, salicylic acid, rhodotorulic acid, Na oxalate, EDDHA, sorbohydroxamic acid, gamma-pyrone-2, 6-decarboxylic acid, and at pH 7.9-8.1 with hexanohydroxamic acid, benzohydroxamic acid. Some ecological and physiological implications are discussed.

Naked Amoebas and Bacteria in an Oil-Impacted Salt Marsh Community.

Populations of soil amoebas were monitored in two salt marshes in Staten Island, NY for 2 years. One site, Gulfport Reach on the Arthur Kill, has been highly impacted by numerous oil spills. In particular, in 1990 a massive no. 2 fuel oil spill from a ruptured pipe flooded the area; its sediments had total petroleum hydrocarbon (TPH) concentrations in the range 800-46,000 ppm. A reference site 11 km away, on the Atlantic coast, had low TPH levels. Amoeba population densities were in general higher in the impacted sediments. In laboratory microcosm experiments, sediment samples from unimpacted sites were treated with added fresh (unweathered) hydrocarbons (no. 2 fuel oil) and cultured; these also yielded higher amoeba numbers than untreated controls. Four distinct amoeba morphotypes were monitored. Changes in population levels of total amoebas were correlated in the two sites, particularly for morphotype 2 (r = 0.83). The ratios of total amoebas to total bacterial numbers were also correlated (r = 0.85) between the sites. This suggests the amoebas may function as generalists, and that their trophic relation to bacterial prey is not much affected by the presence of petroleum hydrocarbons, but rather may reflect regional parameters such as ambient temperature or other physical factors.

Unity and diversity in biochemistry.

A consideration of A Biochemical Phylogeny of the Protists, by M.A. Ragan and D.J. Chapman (Academic Press, 1978) and Biochemical Adaptation to Environmental Change, ed. by R.M.S. Smellie and J.F. Pennock (the Biochemical Society, 1976), within some thoughts on biochemical evolution.

Chemosensory responses of Acanthamoeba castellanii: visual analysis of random movement and responses to chemical signals.

A visual assay slide chamber was used in conjunction with time-lapse videomicroscopy to analyze chemotactic behavior of axenically grown Acanthamoeba castellanii. Data were collected and analyzed as vector scatter diagrams and cell tracks. Amebas responded to a variety of bacterial products or potential bacterial products by moving actively toward the attractant. Responses to the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP), lipopolysaccharide, and lipid A were statistically significant (P < or = 0.03), as was the response to fMLP benzylamide (P < or = 0.05). Significant responses to cyclic AMP, lipoteichoic acid, and N-acetyl glucosamine were also found. Chemotactic peptide antagonists, mannose, mannosylated bovine serum albumin, and N-acetyl muramic acid all yielded nonsignificant responses (P > 0.05). There was no single optimal concentration for response to any of the attractants tested, and amebas responded equally over the range of concentrations tested. Pretreatment of amebas with chemotactic peptides, bacterial products, and bacteria reduced the directional response to attractants. Amebas that had been grown in the presence of bacteria appeared more responsive to chemotactic peptides. Treatment of amebas with trypsin reduced the response of cells to chemotactic peptides, though sensitivity was restored within a couple of hours. This suggests the ameba membrane may have receptors, sensitive to these bacterial substances, which are different from the mannose receptors involved in binding bacteria to the membrane during phagocytosis. The rate of movement was relatively constant (ca. 0.40 microns/s), indicating that the locomotor response to these signals is a taxis, or possibly a klinokinesis, but not an orthokinesis. Studies of the population diffusion rate in the absence of signals indicate that the basic population motility follows the pattern of a Levy walk, rather than the more familiar Gaussian diffusion. This suggests that the usual mathematical models of ameboid dispersion may need to be modified.

Chemosensory responses of a protozoan are modified by antitubulins.

Modification of a behavioral response of a marine dinoflagellate to chemical cues is described. Negative response to choline was modified by the antitubulins vincristine, vinblastine, griseofulvin, and trifluralin, but not by colchicine. Positive responses to 3,4-dihydroxyphenylalanine were unaffected by these drugs.

Chemosensory responses by the heterotrophic marine dinoflagellateCrypthecodinium cohnii.

Chemosensory responses by the colorles inshore marine dinoflagellateCrypthecodinium cohnii were observed in quadrant-divided Petri plates containing an agar layer + liquid overlay. A suspension of organisms in salt solution was poured onto this and allowed to stand 3 hr. A differential tendency of the cells to become firmly attached or embedded in the substratum was observed when various substances were incorporated in the gel. A positive response (tendency to attach) occurred with: α-L-fucose, dimethyl-ß-propiothetin, betaine, sucrose, glycine, L-alanine, hemin, and fructose; negative response: formalin, glutathione, acid hydrolyzed agar, protamine SO4, L-glutamic acid, lactose, glutamine, taurine, L-aspartic acid, putrescine 2 HCl, choline citrate, choline bitartrate, K citrate, and choline HCl. γ-Aminobutyric acid was negative or positive dependeng on concentration. Dead or immotile cells did not become attached. The following compounds elicited no response: α-D-fucose, dimethyl acetothetin chloride, cyclic AMP, and glucose.

A mathematical model of pattern formation by swimming microorganisms.

Bioconvection in suspensions of Tetrahymena pyriformis and Crypthecodinium cohnii is described and 2 new patterns, the toroid and the cat's-eye, which appear in shallow suspensions of C. cohnii, are reported. Except in very dense cultures, bioconvection does not arise unless the depth of the suspensions or the mean concentration exceed certain critical values, other things being equal. A mathematical model describing the hydrodynamics of suspension of negatively geotactic microorganisms is described which predicts the existence of critical depths and concentrations. The equations presented admit solutions describing the "polka-dot" patterns seen at low organism concentration in suspensions slightly deeper than the critical value. The discussion here is limited to the case of fairly dilute suspensions, but the basic approach can be applied also to richer cultures.

CHEMOSENSORY RESPONSES TO AMINO ACIDS AND CERTAIN AMINES BY THE CILIATE TETRAHYMENA: A FLAT CAPILLARY ASSAY.

An assay for chemosensory responses by the ciliate Tetrahymena thermophila is described that uses glass capillaries with a rectangular cross-section (inner dimensions, 20 x 2 x 0.2 mm). These have optical and geometrical properties permitting convenient observation of cell behavior within the capillaries. Washed cells, starved for 12 h, accumulated preferentially in capillaries containing L-methionine, L-leucine, L-cysteine, L-histidine, L-histamine, cimetidine, agmatine, and berenil at concentrations of 10-3 M or less. They avoided capillaries containing tripelennamine, diphenhydramine, and pentamidine at these concentrations. It is argued that the actual response thresholds are much lower than the concentrations put into the capillaries, since cells respond to the gradient of the diffusing chemical. L-Isoleucine, itself inert, blocked the response to L-leucine but not to L-methionine, L-cysteine, or L-histidine. L-Ethionine and 1-homocysteine caused accumulation but not L-cysteine or DL-cystathionine. L-Cystine did not block the response to L-cysteine. Cells accelerated when entering a capillary where accumulation occurred. On reaching the interior they swam more slowly and uniformly, and with fewer turns or stops than in control capillaries lacking the chemical signal, or when outside of the capillaries. Cells were inhibited from leaving both control and test capillaries, possibly because of accumulated wastes or secretions in the surrounding medium.