The Regulation of Pea (Pisum sativum L.) Symbiotic Nodule Infection and Defense Responses by Glutathione, Homoglutathione, and Their Ratio.

In this study, the roles of glutathione (GSH), homoglutathione (hGSH), and their ratio in symbiotic nodule development and functioning, as well as in defense responses accompanying ineffective nodulation in pea (Pisum sativum) were investigated. The expression of genes involved in (h)GSH biosynthesis, thiol content, and localization of the reduced form of GSH were analyzed in nodules of wild-type pea plants and mutants sym33-3 (weak allele, "locked" infection threads, occasional bacterial release, and defense reactions) and sym33-2 (strong allele, "locked" infection threads, defense reactions), and sym40-1 (abnormal bacteroids, oxidative stress, early senescence, and defense reactions). The effects of (h)GSH depletion and GSH treatment on nodule number and development were also examined. The GSH:hGSH ratio was found to be higher in nodules than in uninoculated roots in all genotypes analyzed, with the highest value being detected in wild-type nodules. Moreover, it was demonstrated, that a hGSHS-to-GSHS switch in gene expression in nodule tissue occurs only after bacterial release and leads to an increase in the GSH:hGSH ratio. Ineffective nodules showed variable GSH:hGSH ratios that correlated with the stage of nodule development. Changes in the levels of both thiols led to the activation of defense responses in nodules. The application of a (h)GSH biosynthesis inhibitor disrupted the nitrogen fixation zone in wild-type nodules, affected symbiosome formation in sym40-1 mutant nodules, and meristem functioning and infection thread growth in sym33-3 mutant nodules. An increase in the levels of both thiols following GSH treatment promoted both infection and extension of defense responses in sym33-3 nodules, whereas a similar increase in sym40-1 nodules led to the formation of infected cells resembling wild-type nitrogen-fixing cells and the disappearance of an early senescence zone in the base of the nodule. Meanwhile, an increase in hGSH levels in sym40-1 nodules resulting from GSH treatment manifested as a restriction of infection similar to that seen in untreated sym33-3 nodules. These findings indicated that a certain level of thiols is required for proper symbiotic nitrogen fixation and that changes in thiol content or the GSH:hGSH ratio are associated with different abnormalities and defense responses.

First observation of microcystin- and anatoxin-a-producing cyanobacteria in the easternmost part of the Gulf of Finland (the Baltic Sea).

The aim of this study was to obtain the first data on the occurrence and distribution of potentially toxic cyanobacteria and cyanotoxins in the Russian Easternmost part of the Gulf of Finland of the Baltic Sea. Studied samples were collected from 2012 to 2017 and three independent approaches - HPLC-HRMS, PCR and light microscopy were applied for cyanotoxins analysis and detection of toxigenic cyanobacteria. Aphanizomenon flos-aquae Ralfs ex Born. et Flah., Planktothrix agardhii (Gom.) Anag. et Kom., Microcystis aeruginosa (Kütz.) Kütz. and Dolichospermum spp. dominated among cyanobacteria in collected samples. In 2012-2013 during research cruises, microcystins concentrations varied from below detection levels to low (0.01-0.6 µg L-1) values. In the autumn of 2015 and 2017, during cyanobacterial bloom events very high concentrations of microcystins (dissolved up to 49 µg L-1, intracellular up to 466 µg g-1) and dissolved anatoxin-a (1.4 µg L-1) were detected. The evaluated toxin profile was represented by most common arginine-containing variants of microcystins (MC-LR, MC-RR, MC-YR) and their desmethylated forms. Leucine-containing congeners (MC-LF; MC-LY; MC-LW for the biomass sample from the coast of Komarovo, 2015) were found at low concentrations. In environmental DNA from bloom samples, we identified mcy genes regions responsible for MC biosynthesis that are specific for Dolichospermum, Microcystis, and Planktothrix. This study is the first molecular evidence the ability of Microcystis aeruginosa and Planktothrix agardhii from the Gulf of Finland to produce microcystins. On the basis of the obtained data of genus-specific PCR and microscopy, we suppose the presence of anatoxin-a-producing Apanizomenon flos-aquae population in the phytoplankton of Russian part of the Gulf of Finland.

Dolichospermum and Aphanizomenon as neurotoxins producers in some Russian freshwaters.

Last decades, cyanobacterial blooms have been commonly reported in Russia. Among the boom-forming species, potential toxin producers have been identified. The aim of this paper was to study the presence of neurotoxic compounds - saxitoxins and anatoxin-a - in water bodies from different regions of Russia. We also made attempts to identify the neurotoxin-producing genera. The good convergence of the results obtained by light microscopy, PCR and LC-MS/MS analyses indicated the presence of active neurotoxin producing species in all investigated water bodies. Saxitoxin was detected in phytoplankton from 4 water bodies in Central European Russia and West Siberia, including lake and reservoirs used as a source for potable water. The water bodies differed with the respect of saxitoxin producers which belonged to Aphanizomenon and/or Dolichospermum genera. For the first time, we obtained quantitative data on the intracellular saxitoxin concentration in Russian freshwaters using LC-MS/MS. Anatoxin-a was detected only in lakes of Northwestern Russia. In the eutrophic shallow Lower Suzdal Lake, Aphanizomenon was the stated anatoxin-a-producing genus. In the large shallow artificial hypertrophic Sestroretskij Razliv Lake, it was very likely that both dominant species - Aphanizomenon flos-aquae and Dolichospermum planctonicum - were anatoxin-a producers.