Fluorescence quenching in four unicellular algae with different light-harvesting and xanthophyll-cycle pigments

We examined the relationship between non-photochemical quenching (NPQ) and xanthophyll de-epoxidation in the unicellular algae Euglena gracilis, Ochromonas danica, Phaeodactylum tricornutum, and Dunaliella tertiolecta. Generally, low-light-grown algae had a smaller pool of xanthophyll-cycle pigments per chlorophyll than medium-light-grown grown cells, but they developed more NPQ during exposure to high light. Thus, lumen acidification was apparently lower in medium-light-grown cells in spite of the exposure to a photon flux density (PFD) three times the growth PFD. In darkness Dunaliella maintained a relatively large content of de-epoxidized xanthophylls, and NPQ developed without concomitant de-epoxidation in response to a 5-min exposure to high light. Violaxanthin de-epoxidation that occurred during longer exposures to light did not cause a further rise in NPQ in Dunaliella. In Ochromonas, NPQ and xanthophyll de-epoxidation increased simultaneously during a 15-min exposure to high light. A further rise in NPQ was not accompanied by xanthophyll de-epoxidation. In Phaeodactylum, the rise in NPQ and de-epoxidation were nearly linearly related during a 60-min exposure to high light. NPQ recovered quickly after darkening in these three algae and no significant photodamage occurred. In Euglena no xanthophyll-conversions and no quickly reversible NPQ occured in response to high light, suggesting that photodamage occurred. Dunaliella has similar light-harvesting and xanthophyll-cycle pigments as higher plants but the relationship between NPQ and DPS during the exposure to high light was different from the linear relationship that is commonly observed in plants. Conversely, Phaeodactylum, which has different light-harvesting and xanthophyll-cycle pigments, had a relationship similar to that in plants.     

Nigericin insensitive post-illumination reduction in fluorescence yield in Dunaliella tertiolecta (chlorophyte)

Cells of the green alga Dunaliella tertiolecta grown in a light/dark cycle were exposed to high light for about 15 min. In light, energy-dependent quenching reduced fluorescence emission and decreased PS II efficiency. Within 3 minutes after darkening fluorescence quenching largely relaxed. However, PS II fluorescence emission decreased again after further darkening. F_o and F_m decreased to the same relative extent and the PS II efficiency was not reduced. This Reduction in Fluorescence yield in Darkness, termed RFD for the purpose of this paper, lasted about 20 min. The deepoxidation state of xanthophylls remained unchanged during and after the 15-min exposure to high light. We show that RFD is insensitive to the uncoupler nigericin and thus unrelated to energy-dependent quenching. RFD correlated with a reduction of the PQ pool after darkening and low levels of far red or blue light (430 nm more than 460 nm) prevented RFD. This is in contrast to observations in higher plants, where a post-illumination reduction of the PQ pool causes and increase in F_o (Groom et al. (1993) Photosynth Res 36: 205–215). Changes in the adenylate energy charge were not correlated with RFD. Antimycin A and cyanide, both inhibitors of the PQ-oxidase, caused an increase in RFD whereas SHAM, an inhibitor of the chloroplastic glycolate-quinone oxidoreductase, caused a decrease. Low CO₂ concentrations, known to increase the oxygenase activity of Rubisco and to generate glycolate and P-glycolate in light, caused an increase in RFD. We propose that accumulated glycolate and P-glycolate reduce the PQ pool in darkness, leading to the formation of RFD. During RFD, 77 K fluorescence emission from PS II was more reduced than that from PS I, thus resembling a state I, state II transition. However, the reduction in fluorescence yield during RFD is much larger than the reduction previously attributed to state transitions and it is unclear whether RFD and state transitions are identical. The formation and relaxation of RFD increased with higher temperatures and the extent of RFD was largest at the growth temperature (25°C). RFD has to be taken into account when fluorescence is measured after darkening as it may be mistaken for energy-dependent quenching.Abbreviations F_o fluorescence, measured when PS II traps are open - F_o difference between F_o and F_o - F_m fluorescence, measured when PS II traps are temporarily closed - F_m difference between F_m and F_m - FR far red - PFD photosynthetically active photon flux density - PQ plastoquinone - RFD reduction in fluorescence in darkness - SHAM salicylhydroxamic acid - Q_A primary quinone acceptor of PS II     

Rapid fluorescence-based screening for Wolbachia endosymbionts in Drosophila germ line and somatic tissues

Wolbachia is a globally distributed bacterial endosymbiont present in arthropods and nematodes. The advent of sensitive PCR-based approaches has greatly facilitated the identification of Wolbachia-infected individuals and analysis of population infection levels. Here, a complementary visual fluorescence-based Wolbachia screening approach is described. Through the use of the fluorescent dye Syto-11, Wolbachia can be efficiently detected in various Drosophila tissues, including ovaries. Syto-11 also stains Wolbachia in other insects. Because Wolbachia is inherited through the maternal germ line, bacteria reside in the ovaries of flies in infected populations. An advantage of this staining approach is that it informs about Wolbachia titer as well as its tissue and cellular distribution. Using this method, the infection status of insect populations in two central California locations was determined, and variants with unusually low or high Wolbachia titers were isolated. In addition, a variant with ovarioles containing both infected and uninfected egg chambers was identified. Syto-11 staining of Cardinium- and Spiroplasma-infected insects was also analyzed.     

The Molecular Biology of Chloroplasts and Mi-tochondria in Chlamydomonas. Edited by J.-D. Rochaix,M. Goldschmidt-Clermont and S. Merchant. Advances in Photosynthesis,Vol. 7, 1998

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