Characterization of the Eyespot Regions of "Blind"Chlamydomonas Mutants after Restoration of Photophobic Responses

Chlamydomonas reinhardtii exhibits photophobic and positive and negative phototactic responses that can be defined for cell populations using computerized cell tracking and motion analysis. Mutants CC-2359 and FN68 are pigment deficient mutants that are blocked in carotenoid synthesis and lack these photo responses. In particular, neither mutant exhibits flash-induced photophobic responses to visible light stimuli to which wild-type gametic cells exhibit a strong response, with several behavioral stages. Upon addition of all-trans retinal to these mutants, the photophobic responses are restored with minor quantitative differences from wild-type populations. Using both light and electron microscopy, we have compared the ultrastructural characteristics of wild-type C. reinhardtii to those of both mutants. As previously described, wild-type cells contain an eyespot consisting of 2–4 layers of pigmented granules encased within thylakoid membranes, located between the distal extremities of the flagellar root. This structure is also visible as an orange-red spot in light microscopy. The photoreceptor is thought to be concentrated in the plasma membrane above the eyespot. The mutant, CC-2359, lacks this eyespot as seen by both light and electron microscopy, even when the photophobic response has been restored. FN68-like mutants studied earlier by Morel-Laurens and Feinlieb and others contain an eyespot which can be seen only by electron microscopy. In FN-68, the eyespot generally has the same dimensions as in wt cells, differing mainly in pigment granule appearance. Consistent with these findings, several laboratories have shown that the full range of phototactic responses can be reconstituted in FN68 and CC-2359, but that negative phototaxis requires a significantly stronger light stimulus in the latter strain. We confirm the suggestion that the eyespot is not necessary for the photophobic response, and is not critical for the appropriate assembly and function of the photophobic response receptor in the membrane. Furthermore, the locus of reconstitution of the functional receptor is not the eyespot. Because of the definitive demonstration of the absence of the eyespot in CC-2359, however, the eyespot may play a role in negative phototaxis.     

REFLECTIVE PROPERTIES OF THE STIGMA IN MALE GAMETES OF ECTOCARPUS SILICULOSUS (PHAEOPHYCEAE) STUDIED BY CONFOCAL LASER SCANNING MICROSCOPY1

The reflection properties of the stigma in male gametes of Ectocarpus siliculosus (Dillw.) Lyngbye were investigated using confocal laser scanning microscopy in the epireflection contrast mode. The complex reflection pattern obtained after optical xy (horizontal) and xz (vertical) sectioning was consistent with stigma ultrastructure as revealed by serial thin sections. The intensity and pattern of the reflection signal varied with the orientation of the cell/stigma to the incident laser light. Maximal reflection occurred only in approximately normal orientation of the stigma to the light source. Focusing of reflected light from an elongated concave depression of the stigma on the region of the flagellar swelling was observed in xy and xz sections of living and fixed gametes. The results indicate the importance of mechanisms (focusing) other than quarter-wave interference reflection in signal amplification by the eyespot of flagellate algae.     

GEOTAXIS/PHOTOTAXIS AND BIOCHEMICAL PATTERNS IN HETEROCAPSA (=CACHONINA) ILLDEFINA (DINOPHYCEAE) DURING DIEL VERTICAL MIGRATIONS

Two separate experiments with Heterocapsa (= Cachonina ) illdefina Herman et Sweeney, one with and the other without water volume replacement, were performed in a 250-L laboratory mesocosm (45-cm diameter × 150-cm height) to examine how diel vertical migration (DVM) relates to taxis sign and strength and to cellular biochemical state. Although only the cell population grown with water volume replacement maintained a division per day over the course of the experiment, periodic measurements during both experiments demonstrated that cells aggregating at the surface during the light period generally were deficient in all measured biochemical constituents compared to cells obtained from a midcolumn depth. More specifically, H. illdefina cells that aggregated at the surface during the light period in both experiments exhibited weakened positive geotaxis but strengthened positive phototaxis and were very deficient in lipid and free amino acid compared to midcolumn cells. Cells sampled at midcolumn during the light period exhibited similar but weaker taxes changes compared to surface samples, and geotaxis strength was inversely correlated with cell diameter, cellular DNA and protein content, and RNA/DNA ratio. In comparison, published data on Gymnodinium breve Davis, a harmful algal bloom species, showed that cells aggregating at the surface during the light period generally exhibited weakened negative geotaxis and strengthened positive phototaxis and were very deficient in lipid and chl a compared to midcolumn cells. Although the persistent tendency toward negative geotaxis was weaker in midcolumn subpopulations throughout the day, its strength was inversely correlated with cell diameter and cellular lipid content. The combined results for both species support a revised conceptual model of optimized DVM in autotrophic marine dinoflagellates incorporating generalized expressions of taxis and biochemical state of individual cells.     

OBSERVATIONS ON THE PECULIAR DIURNAL MIGRATION OF A RED TIDE DINOPHYCEAE IN TROPICAL SHALLOW WATERS1

The dinoflagellate Peridinium cf. quinquecorne Abé forms red tide-like blooms in eutrophic shallow waters in the Philippines. The organism moves into a distinct near-surface layer when intensive solar radiation occurs, but only during the incoming tide. Shortly before high tide, regardless of light levels, the dinoflagellates seem to disappear. Simple experiments show that once intensive radiation has been reduced Peridinium quinquecorne moves out of the water column and attaches itself to solid objects away from the light. The morphology of the organism, especially as related to attachment, was studied through SEM. Its high swimming velocity and the reaction to radiation and tidal changes are described. The possibility that, superimposed on its reaction to light, this dinoflagellate may follow intrinsic tide-dependent oscillations is discussed.     

Euglena: The Photoreceptor System for Phototaxis*

SYNOPSIS. The photoreceptor structures (eyespot-paraflagellar body-flagellum) for Euglena phototaxis were investigated by electron microscopy. The paraflagellar body—the photoreceptor—is a highly ordered crystalline lamellar structure. Optical diffraction of the electron micrographs and resulting filtered images of the paraflagellar body suggest that it is formed of rods in a helical arrangement. The action spectra for phototaxis, the in situ spectrum by microspectrophotometry of the paraflagellar body, and flavin analysis of the organism indicate that the photoreceptor molecule is a flavoprotein. The phototaxis action spectrum is similar to the spectrum for O₂ evolution and implies that similar molecules participate in the photo-processes. As a result, a photochemical scheme is suggested in which a photo-excited flavin and a cytochrome participate in the photoprocess. The photochemistry and photoreceptor structures for Euglena phototaxis are likened to a photoneuro sensory cell.     

Photoresponses of second-instar Chaoborus larvae

The diel vertical migration of Chaoborus larvae varies with larval instar. Although light is involved in the control of vertical migration the contribution of larval photoresponses is unknown. In order to describe ontogenetic changes in larval photoresponses we measured photoresponses of second-instar Chaoborus punctipennis larvae in the laboratory. The response spectrum of these larvae had peaks in sensitivity at 420 and 620 nm with a wide plateau of lower sensitivity from 460 to 600 and 640 to 680 nm. Dark adapted larvae were positively phototactic at intensities from 10^?7 to 10¹ Wm^?2 at 420 nm. The level of response decreased somewhat above 10^?4 Wm^?2, and above 10^?2 Wm^?2 a small proportion of larvae shifted to a negative phototaxis. At 420 nm the threshold intensity was about 10^?7 Wm^?2 for positive phototaxis and 10^?2 Wm^?2 for negative phototaxis. Light adaptation increased the threshold intensity for positive phototaxis. The differences in larval photoresponses between second- and fourth-instar larvae suggests that the young instars are adapted to the photoenvironment of the water column and older larvae are adapted to avoid the water column except at very low light intensities. These predictions match the diel distribution of these larvae.     

Effects of Solar Radiation on Photoorientation,Motility and Pigmentation in a Freshwater Cryptomonas

The effects of solar radiation on motility, photoorientation and pigmentation have been studied in a freshwater Cryptomonas species. The diaphototactic orientation performed by the cells is impaired within about 90 min of solar radiation. Likewise, the percentage of motile cells within the population and the average velocity of the swimming cells decreases within about the same exposure time. This effect is not due to a thermal stress but rather seems to be caused by the solar UV-B component, since decreasing short wavelength UV radiation by means of an artificial ozone filter or UV cut-off filters increased the tolerated exposure time. Solar radiation also bleached the photosynthetic pigments of the cells as shown by absorption difference spectra.     

Surveys of stem base diseases and fusarium ear diseases in winter wheat in England, Wales and Scotland, 1989–1990

An investigation into the incidence of stem base diseases and the pathogens associated with them was undertaken on winter wheat samples collected at growth stages 31 and 73–75 in 1989 and 1990. Symptoms at growth stage 31 were classified into 10 different types based on visual characteristics. Although Pseudocercosporella herpotrichoides was associated mainly with eyespot lesions and Rhizoctonia spp. with sharp eyespot lesions, a significant number of isolates of each pathogen were taken from lesions classified as fusarium. Different types of fusarium lesion on stem bases at both growth stages were not associated with any one particular Fusarium or Microdochium species. 98% of P. herpotrichoides isolates obtained from eyespot lesions at growth stage 31 in 1989, and 87% of those obtained in 1990 were identified as the sub-species P. herpotrichoides var. acuformis, Microdochium nivale was more common than any of the Fusarium species at both growth stages, and was particularly prevalent in samples from Scotland at the earlier growth stage in 1989. Isolations from the top internodes at growth stage 73–75 indicated that systemic infection by Fusarium species, although present in some stems, was of little importance. Fusarium ear blight affected 0.4% of ears in 1989 and 0.5% in 1990. Glume spot lesions on the ears in 1990 were predominantly associated with Fusarium poae.     

MEASURING PHOTOSYNTHETIC ACTION SPECTRA OF NATURAL PHYTOPLANKTON POPULATIONS1

The photosynthetic response, defined as the initial slope of the photosynthesis-irradiance curve, was determined spectrally (every 25 nm from 400 to 675 nm; 25 nm half-maximum bandpass) for natural phytoplankton populations from High Arctic, Grand Banks and Sargasso Sea waters, as well as for populations living in the lower margin of sea ice off Newfoundland, All spectra were similar in shape with a maximum at 425–450 nm, a broad shoulder to 550 nm, a valley from 600 to 650 nm and a rise at 675 nm. The error resulting from the use of spectrally averaged initial slope to predict photosynthesis under different optical and fluid dynamical conditions at sea is discussed.     

OCCURRENCE AND ABUNDANCE OF GREEN-FLUORESCING DINOFLAGELLATES IN SURFACE WATERS OF THE NORTHWEST ATLANTIC AND NORTHEAST PACIFIC OCEANS1

We have cultured green fluorescing heterotrophic dinoflagellates whose continuous green fluorescence is due to an unidentified compound, probably a flavin, that excites with blue (～460 nm) light and emits green (～535 nm) light. No evidence of bioluminescence was found, but we note that compounds with similar fluorescence characteristics have been associated with bioluminescence in other taxa. These cells, all naked gymnodinoids, are widespread and abundant in the Northwest Atlantic and Northeast Pacific Oceans (10³–10⁵ L^?1). They comprise 4–100% of the total heterotrophic dinoflagellate component which, in turn, is usually equivalent magnitude to the phototrophic naked dinoflagellate component of the phytoplankton community.     

High Speed Cinemicrography of the Direct Photophobic Response of Euglena and the Mechanism of Negative Phototaxis*

SYNOPSIS. The shock reaction of Euglena gracilis strain Z to a sudden increase in light intensity (the “direct photophobic response”) was examined by high speed cinemicrography. The response is expressed as a turning reaction toward the dorsal side of the cell, after a transduction time of 0.1–0.5 sec after the onset of stimulation. Transduction times, turning rates, and flagellar beat frequencies were measured by analyzing the filmed sequences. The experimental data are consistent with a mechanism of directional homeostasis in negative phototaxis that is based upon shading of the photoreceptor by the cell's posterior end.