Picocianobatteri nel Mar Nero profondo: i misteri di un ambiente meromittico

Responsabili di progetto

Cristiana Callieri, Snejana Moncheva

Accordo

BULGARIA - BAS - Bulgarian Academy of Sciences

Bando

CNR-BAS 2016-2018

Dipartimento

Scienze del sistema terra e tecnologie per l'ambiente

Area tematica

Scienze del sistema Terra e tecnologie per l'ambiente

Stato del progetto

Nuovo

Proposta di ricerca

Picocyanobacteria are autotrophic microorganisms with cell size < 2µm, considered as the main primary producers in the oceans and in many large lakes (Scanlan 2012, Callieri et al. 2012). Their size and adaptation ability are the reasons of their success on our planet, where they evolved colonizing aquatic environments (Sanchez-Baracaldo et al. 2005). Picocyanobacteria fix inorganic carbon through photosynthesis, constituting the energy source for their consumers, i.e. flagellates, ciliates and zooplankton. They play a key role in the organic carbon cycle, supplying energy and matter to higher level of the trophic chain. Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus numerically dominate vast tracts of the oceans and contribute a significant proportion of the global primary production (Scanlan 2012). Some clusters of the Synechococcus marine lineage are phylogenetically near to some of freshwaters (Callieri et al. 2013), showing the extreme plasticity of these microorganisms in the colonization of different habitats. The presence in the euphotic layers of the Black Sea (up to 50m) of Synechococcus (Uysal 2006) and to a lesser extent of Prochlorococcus (Mella-Flores et al. 2011) has been observed. Nevertheless data on their presence in the deep zone from 50 to 1000m has been not yet described. Black sea (maximum depth: 2245m) represents an interesting environment for the study of microbial communities as it includes heterogeneous zones: a superficial layer with nitrogen limitation, an oxicline where chemical features abruptly change and an anoxic deeper layer with the presence of hydrogen sulphide (Oguz et al. 2000). Furtheremore, its salinity is low as it receives freshwaters from Danube and from other important rivers and it is a relatively closed system. The Bulgarian Academy of Sciences (BAS) of Varna and the University of Sofia are studying the biogeochemistry of the west basin of Black Sea using profilers for the measurements of main chemical, physical and biological parameters from the surface to 1000m. During this campaign they found unusual profiles with an unexpected increase of chlorophyll-a in the deeper layers. Some of these samples have been analysed in our laboratories of the CNR Institute of Ecosystem Study (ISE) of Verbania. Here we verified the presence of picocyanobacteria in samples from deep layers of the Black Sea through flow cytometry and epifluorescence microscopy. We detected the characteristic Synechococcus mark in orange vs red fluorescence at 488nm excitation. The microscopic check visualised phycoerytrin-rich cells in division. The presence of Synechococcus in the meso- and bathypelagic oxygenated zone of the oceans has been described recently (Sohrin et al. 2011) but this was attributed to vertical or lateral sedimentation of the cells. This cannot be the case for the Black Sea where a pronounced chemocline segregates the anoxic monimolimnion. Therefore, we decided to start a cooperation project to investigate the presence of picocyanobacteria in the deep zone of the Black Sea to isolate the strains and to quantify constitutive and functional genes indicative of their activity in the dark, anoxic zone, with the presence of nitrite, nitrate and hydrogen sulphate. Our hypothesis is that these small cyanobacteria could carry out activities alternative to photosynthesis in the dark, anoxic layers of the Black Sea as for example nitrogen fixation and fermentation. Synechococcus is able to switch on a variety of genes involved in the fermentation when in dark anaerobic conditions (Steunou et al 2006). This can provide it the energetic support for fixing N2, also if Synechococcus lacks heterocysts, particularly if it grows in anoxic condition (Fay 1992). In the Black Sea, in the absence of oxygen, N2 comes from nitrite reduction (operated for example by anammox of Planctomycetes phylum) (Ulloa et al 2012). Therefore, the denitrifiers could act in syntrophy with picocyanobacteria fuelling the N2. To test this hypothesis we plan to operate both on natural samples and isolates, quantifying by qPCR the transcripts (mRNA) encoding for enzymes involved in nitrogen fixation and fermentation. The cooperation with the BAS Institute of Varna, with their infrastructures and experience in the measurement of profiles of chlorophyll and chemical physical parameters in the Black Sea is fundamental for the execution of this project. The Black Sea is a unique environment for studying the presence of picocyanobacteria in extreme conditions and is a transitional habitat between oceans and lakes.
References
Callieri C, et al. 2013. FEMS Microbiol. Ecol. 85: 293-301./ Callieri C, et al. 2012. In: Ecology of Cyanobacteria: Their Diversity in Time and Space. Springer Publishers: 229-269./ Fay P. 1992. Microbiol. Rev. 56: 340-373./. Mella-Flores et al. 2011. Biogeoscience 8: 2785-2804./ Oguz et al. 2000. Global Biogeochemical Cycles 14:1331-1352./ Sánchez-Baracaldo P, et al. 2005. Geobiology 3:145-165./ Steunou et al 2006. PNAS 103:2398-2403. / Scanlan D. 2012. In: Ecology of Cyanobacteria: Their Diversity in Time and Space. Springer Publishers: 503-533./ Sohrin et al. 201. AME 64:1-14./ Ulloa et al. 2012. PNAS 109:15996-16003./ Uysal 2006. Deep Sea Res.53:1976-1987

Obiettivi della ricerca

We aim to study cyanobacterial populations of the picoplanktonic fraction (0.2-2µm) discovered in the deep anoxic layer of the Black Sea to understand their role in this euxinic environment (anoxic and with H2S, from Pontus Euxinus: Black Sea). If we will corroborate the presence of living cyanobacteria cells in the bathypelagic zone this would represent the first finding of autotrophic cells in a hostile environment, dark and anoxic. Therefore, our first aim is to ascertain the presence of Synechococcus or other picocyanobacteria in the Black Sea, to isolate them, and to understand what they are doing there, by measuring the expression of specific genes. Our research plan develops in tree steps: 1) to obtain profiles of chemical, physical, and biological parameters, and of concurrent picocyanobacteria numbers and clades at different depths from surface to 1000m in the western Black Sea; 2) to isolate and characterize picocyanobacteria from superficial and deep layers by 16S rDNA and rpoC1gene sequencing; 3) to test the hypothesis of the role of picocyanobacteria as nitrogen fixers in the bathypelagic, anoxic zone of Black Sea (evaluating the expression by RTqPCR on specific genes eg. nifHDK, nitrogenase) fuelled by fermentation (pflAB and acs). The discovery of a cyanobacteria with the capacity to shift from photosynthesis to fermentation and nitrogen fixation would be new in the sea and would open new understanding of the deep marine system.

Ultimo aggiornamento: 19/04/2024