Posidonia oceanica (L.) Delile in its westernmost biogeographical limit (northwestern Alboran Sea): Meadows characterisation, phenology and flowering events

Mateo-Ramírez A, Urra J, Rueda J, Marina, Bañares-España E, García Raso E. (2016) Posidonia oceanica (L.) Delile in its westernmost biogeographical limit (northwestern Alboran Sea): Meadows characterisation, phenology and flowering events. Front. Mar, Sci. Conference Abstract: XIX Iberian Symposium on Marine Biology Studies. doi: 10.3389/conf.FMARS.2016.05.00055Posidonia oceanica is a Mediterranean endemic seagrass species that forms meadows covering ca. 2.5–4.5 millions of hectares, representing ca.25 % of the infralittoral and shallow circalittoral (down to 50m) bottoms of the Mediterranean. This seagrass is considered a habitat-engineer species and provides an elevated number of ecosystem services. In addition the Marine Strategy Framework Directive (MSFD, 2008/56/EC) includes seagrass like elements to evaluate the “Good Environmental Status” of the European coasts. Information about their phenological characteristic and structure of the meadows is needed for indicator estimations in order to establish their conservation status. The studied meadows are located in the westernmost limit of the P. oceanica distribution (North-western Alboran Sea) in the vecinity of the Strait of Gibraltar, an Atlantic-Mediterranean water transition area. Four sites were selected from East to West: Paraje Natural de Acantilados de Maro-Cerro Gordo (hereafter Maro), Special Area of Conservation “Calahonda” (hereafter Calahonda), Site of Community Importance Estepona (hereafter Estepona) and Punta Chullera (hereafter Chullera) where P. oceanica present their westernmost meadows. Phenological data were recorded from mid November to mid December in P. oceanica patches located at 2 – 3 m depth. At each site three types of patches (patch area 2 m2, large patches) were sampled. At each patch and site, 3 quadrants of 45 x 45 cm were sampled for shoot and inflorescences density measurements. In each quadrant, 10 random shoots were sampled for shoot morphology (shoot height and number of leaves). Shoot and inflorescences densities were standardized to squared meters. All the studied P. oceanica meadows develop on rocks and they present a fragmented structure with a coverage ranging between ca. 45% in Calahonda and Estepona and ca. 31% in Maro. The meadows of Chullera are reduced to a few small - medium patches with areas ranging between 0.5-1.5 m2 (Fig. 1). The meadows of Chullera and Estepona presented similar values of shoot density (ca. 752 – 662 shoots m-2, respectively) and leaf height (ca. 25 cm). Similarly, the Calahonda and Maro meadows also showed similar values of shoot density (ca. 510 – 550 shoots m-2, respectively) but displaying lower values than those of sites located closer to the Strait of Gibraltar. Regarding patch sizes and leaf height, the longest leaves (ca. 25 cm) were found in medium and large patches, but the number of leaves per shoot were higher in the small and the medium size patches (ca. 6.3 leaves per shoot). Flowering was only detected at the Calahonda meadows with maximum values of ca. 330 inflorescences m-2 (115.2 ± 98.2 inflorescences m-2, n= 9; mean ± SD) (Fig.1). Inflorescence density was not significant different among patches of different sizes. In the Alboran Sea and unlike the studied meadows, extensive beds of P. oceanica occur at the National Park of Cabo de Gata (northeastern Alboran Sea), but from east to west (Strait of Gibraltar), meadows are gradually fragmenting and their depth range decrease from 30m to 2m depth between Cabo de Gata and Chullera, respectively. Probably, the Atlantic influence and the characteristic oceanographic conditions of the Alboran Sea (i.e., higher turbidity, higher water turbulence) represent a developmental limiting factor for P. oceanica at higher depths. Similarities between the meadows located closer to Strait of Gibraltar (Chullera and Estepona) were detected as well as between those more distant (Calahonda and Maro). The first ones showed higher values of shoot densities and leaf heights than the formers, which could be relating to the higher hydrodynamic exposure found at Chullera and Estepona meadows. Regarding flowering events, sexual reproduction in P. oceanica is not common in different locations of the Mediterranean Sea. The available information seems to indicate that flowering represent an irregular event and it is related to high seawater temperature. In fact, the flowering episodes that occurred in Calahonda in November 2015, match with the warmest year ever recorded. This is the third flowering event registered in these meadows located close to the westernmost distributional limit of P. oceanica (Málaga, Alboran Sea), which could indicates that these meadows presents a healthy status. Furthermore, the absence of significant differences in relation to inflorescence density between patches of different sizes may be indicating that the fragmentation does not necessarily influence on the flowering of this seagrass species.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Environmental change rate and dispersion pattern modulate the dynamics of evolutionary rescue of the cyanobacterium Microcystis aeruginosa

The rate of biodiversity loss is so high that some scientists affirm that we are being witnesses of the sixth mass extinction. In this situation, it is necessary to ask the following question: can the organisms be able to resist the environmental changes that are taking place? Recent studies have shown the possibility of a population recovering from a stress situation through evolutionary rescue (ER) events. These events depend on the size of the population, its previous history and the rate of the environmental change. The aim of this work is to add more knowledge about the ER dynamics creating stress situations with selective agents (sulphur and salinity) and using the toxic cyanobacterium Microcystis aeruginosa as a model organism. The experiments are based on exposing populations to severe stress and analyze the effect of previous dispersal events and deterioration rates on the occurrence of ER events among populations. The model consists in three different rates of environmental change (constant, slow and fast; under salinity stress we only used the first two treatments) and three dispersal models (isolated, local or global). In total, 324 and 720 populations were exposed to stressful conditions caused by sulphur and salinity, respectively. The results showed that the dispersal modes and the environmental deterioration rates modulated the occurrence of ER events. It has been observed that dispersal favours ER events for both selective agents. Regarding the rate of environmental change, we observed an increase of ER events under constant changes in the populations exposed to sulphur stress. However, ER events were higher when there was previous deterioration (i.e., slow environmental change rate) under saline stress. As a conclusion, ER events in M. aeruginosa depend on selective agent, being the probability higher for salinity than for sulphur. Thus, it could be hypothesized that general conclusions in ER studies must take into account the selective agent.This work has been financially supported by the projects CGL2014- 53682-P (Ministerio de Economía y Competitividad) and CGL2017-87314-P (Ministerio de Economía, Industria y Competitividad), and the Universidad de Málaga, Campus de Excelencia Internacional Andalucía Tech

The limit of sulphide adaptation in the sulphide-sensitive cyanobacterium "Microcystis aeruginosa"

Libro de resúmenes de ponencias y comunicaciones. XXVII Congreso Nacional de Microbiología 2019, p 276 , ISBN 978-84-09-02644-9Although sulphide has played an important role in the evolution of photosynthesis, it produces a lethal effect on most photosynthetic organisms due to its redox activity on certain enzymes, inhibiting oxygenic photosynthetic and respiratory electron transport. However, cyanobacteria vary in sulphide tolerance, showing different degrees of sulphide resistant oxygenic photosynthesis and even some taxa have the capacity to perform anoxygenic sulphide-dependent photosynthesis. Microcystis aeruginosa is a sulphide-sensitive species. However, it is known that genetic adaptation of aquatic photosynthetic microorganisms to selective agents can be rapidly achieved, even at lethal levels, as the consequence of single mutations. The aim of this work was to determine the maximum sulphide concentration to which this sulphide-sensitive species is able to adapt in order to shed light on the process of sulphide adaptation in cyanobacteria. We used three M. aeruginosa strains, Ma1Vc, Ma5Vc and MaAVc, isolated from a non-sulphureous environment, and whose lethal doses were 0.10, 0.16 and 0.20 mM sulphide, respectively. To study the adaptation of these strains to sulphide a modified ratchet experiment was carried out [...]. The M. aeruginosa strains reached different limits of sulphide adaptation. Ma5Vc and Ma1Vc strains adapted up to 0.40 mM sulphide, i.e. 2,5-fold and 4-fold their initial lethal doses, respectively. However, the MaAVc strain, which showed an initial higher sulphide tolerance, adapted up to only 0.27 mM. Sulphide-lethal dose and photosynthetic performance of the resistant strains obtained in the ratchet experiment were characterized. The lethal dose of the M. aeruginosa Ma1Vc and Ma5Vc mutant strains was 0.40 and 0.72 mM sulphide, respectively. These resistant strains showed lower growth and photosynthetic rates than wild-type ones in the absence of sulphide, which indicates the physiological cost of the mutation conferring sulphide resistance.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Characterization of the cyanobacterium Oscillatoria sp. isolated from extreme sulphureous water from Los Baños de la Hedionda (S Spain)

Los Baños de la Hedionda (Málaga, S Spain) is a natural sulphureous spa (150-200 µM sulphide). Although this high sulphide levels can affect the photosynthetic process, there are numerous photosynthetic microorganisms inhabiting the spa. Among them, we isolated a strain of the cyanobacterium Oscillatoria sp., a genus well known by its tolerance to sulphide. Objectives Firstly, to analyze the photosynthetic characteristics and growth rate of the isolated strain, as well as the effect of the presence of sulphide in both processes. Secondly, to determine the limit of genetic adaptation of this strain to sulphide. Methods The resistance of the isolated strain to sulphide was studied by analyzing the effect of increasing sulphide levels (up to 1600 µM) on photosynthetic performance and growth. The limit of genetic adaptation was explored using an evolutionary experimental design named as ratchet protocol. This design allows discerning the maximum capacity of genetic adaptation of Oscillatoria sp. to the exposure of increasing doses of sulphide Conclusions The strain showed maximum growth rates at 200 µM sulphide although reduced rates can be found up to 800 µM sulphide. A significant increase in resistance was achieved in all derived populations during the ratchet experiment (surviving at sulphide concentrations higher than 2 mM). Moreover, they showed different evolutionary potential to adapt to sulphide, depending on historical contingency.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech.Spanish Ministry of Science and Innovation through CGL2014-53682-P project. Predoctoral State Grant from Scientific and Technical Research and Innovation Plan, Spanish Ministry of Economy, Industry and Competitiveness I+D+i ECC/1402/2013, 201

Disentangling mechanisms involved in the adaptation of the cyanobacterium Microcystis aeruginosa to the extreme sulphureous water from Los Baños de la Hedionda (S Spain)

Backgrounds Los Baños de la Hedionda (Málaga, S Spain) is a natural sulphureous spa where sulphide can reach a concentration of 150-200 µM. Although this ion has biocide properties, including inhibition of the photosynthetic process, a rich flora can be found in this extreme environment. Objectives To study the adaptation mechanisms allowing resistance of photosynthetic microorganisms to these sulphureous waters Methods For this purpose, a modified Luria–Delbrück fluctuation analysis was carried out. The adaptation to La Hedionda waters of three different strains of the cyanobacterium Microcystis aeruginosa (Kützing) Kützing (isolated from a non-sulphureous freshwater reservoir) was analyzed in order to find out if it was achieved by a physiological adaptation process (acclimation) or by the selection of rare spontaneous mutations (genetic adaptation). Conclusions Several resistant strains were obtained after 6 weeks of cultivation with La Hedionda waters. The fluctuation analysis showed that genetic adaptation was the phenomenon that allowed resistant M. aeruginosa cells from the three strains to survive, with similar mutation rates in the order of magnitude of 1 mutant resistant cell per 106-107 cell division-1. It could be hypothesized that this cyanobacterium could adapt to sulphureous environment by the selection of favoured mutants.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech.Spanish Ministry of Science and Innovation through CGL2014-53682-P project. Predoctoral State Grant from Scientific and Technical Research and Innovation Plan, Spanish Ministry of Economy, Industry and Competitiveness I+D+i ECC/1402/2013, 201

Changes in the growth rate of Chlamydomonas reinhardtii under long-term selection by temperature and salinity: Acclimation vs. evolution

We investigated the roles of acclimation and different components involved in evolution (adaptation, chance and history) on the changes in the growth rate of the model freshwater microalga Chlamydomonas reinhardtii P. A. Dang. exposed to selective temperature and salinity. Three C. reinhardtii strains previously grown during one year in freshwater medium and 20 °C were exposed to 5 °C temperature increase and a salinity of 5 g L−1 NaCl. Cultures under each selective scenario and in combination (increase of salinity and temperature), were propagated until growth rate achieved an invariant mean value for 6 months (100–350 generations, varying as a function of scenario and strain). The changes of the growth rate under increased temperature were due to both adaptation and acclimation, as well as history. However, acclimation was the only mechanism detected under salinity increase as well as in the selective scenario of both temperature and salinity, suggesting that genetic variability would not allow survival at salinity higher than that to which experimental populations were exposed. Therefore, it could be hypothesized that under a global change scenario an increase in salinity would be a greater challenge than warming for some freshwater phytoplankton.This work was financially supported by the project CGL2017-87314-P (Ministerio de Economía, Industria y Competitividad, Spain) and the project PID2020-118045-GB-I00 (Ministerio de Ciencia e Innovación, Spain). Funding for open access charge: Universidad de Málaga/CBUA

The limit of the genetic adaptation to herbicides in freshwater phytoplankton and the adaptation photosynthetic cost

Background: One of the most important anthropogenic impacts on freshwater aquatic ecosystems is the continuous increase of herbicide concentrations, which impacts on the structure of phytoplankton communities. Objectives: The maximum adaptation of two green microalgae (Dictyosphaerium chlorelloides and Chlamydomonas reinhardtii) and a cyanobacterium (Microcystis aeruginosa) to two widely used herbicides: glyphosate (N-(phosphonomethyl)glycine) and diuron (C9H10Cl2N2O, 3-(3,4-diclorofenil)- 1,1-dimetilurea) was explored. Additionally, the cost of the herbicide resistance on photosynthesis and growth rate was evaluated. Methods: We used an eco-evolutionary approach (ratchet protocol) to explore the maximum genetic adaptation. We characterised the photosynthetic performance by oxygen production and PSII chlorophyll a fluorescence. Results: A dose of 1 μM diuron or 40 ppm glyphosate completely inhibited the growth of M. aeruginosa and D. chlorelloides, whereas C. reinhardtii growth was completely abolished at 2 μM diuron or 90 ppm glyphosate. However, an increase in resistance to both herbicides was achieved in certain populations during the ratchet experiment. Microcystis aeruginosa and D. chlorelloides were able to adapt up to 8 μM diuron and 80 ppm glyphosate, whereas C. reinhardtii adapted up to twice these herbicide concentrations. The photosynthetic performance was generally lower in the resistant than in the wild-type strains in the three species. These results suggest that increasing concentrations of these herbicides in freshwater bodies could induce the selection of herbicideresistant mutants in phytoplankton communities but showing lower primary production than original populations.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Detection of the maximum resistance to the herbicides diuron and glyphosate, and evaluation of its phenotypic cost, in freshwater phytoplankton

One of the most important anthropogenic impacts on freshwater aquatic ecosystems close to intensive agriculture areas is the cumulative increase in herbicide concentrations. The threat is especially relevant for phytoplankton organisms because they have the same physiological targets as the plants for which herbicides have been designed. This led us to explore the evolutionary response of three phytoplanktonic species to increasing concentrations of two herbicides and its consequences in terms of growth and photosynthesis performance. Specifically, we used an experimental ratchet protocol to investigate the differential evolution and the limit of resistance of a cyanobacterium (Microcystis aeruginosa) and two chlorophyceans (Chlamydomonas reinhardtii and Dictyosphaerium chlorelloides) to two herbicides in worldwide use: glyphosate and diuron. Initially, the growth rate of M. aeruginosa and D. chlorelloides was completely inhibited when they were exposed to a dose of 0.23 ppm diuron or 40 ppm glyphosate, whereas a higher concentration of both herbicides (0.46 ppm diuron or 90 ppm glyphosate) was necessary to abolish C. reinhardtii growth. However, after running a ratchet protocol, the resistance of the three species to both herbicides increased by an adaptation process. M. aeruginosa and D. chlorelloides were able to grow at 1.84 ppm diuron and 80 ppm glyphosate and C. reinhardtii proliferated at twice these concentrations. Herbicide-resistant strains showed lower growth rates than their wild-type coun- terparts in the absence of herbicides, as well as changes on morphology and differences on photosynthetic pigment content. Besides, herbicide-resistant cells generally showed a lower photosynthetic performance than wild-type strains in the three species. These results indicate that the introduction of both herbicides in freshwater ecosystems could produce a diminution of primary production due to the selection of herbicide-resistant mutants, that would exhibit ...This work was financially supported by the project CGL2017-87314- P (Ministerio de Economía, Industria y Competitividad, Spain). The acquisition of the FlowCAM by the University of Málaga was co-financed by the 2008-2011 FEDER program for scientific-technique infrastructure (Proposal number: UNMA08-1E005). Funding for open access charge: Universidad de Málaga/CBUA

Notas corológicas del macrofitobentos de Andalucía (España). X

New records for the macrophytobenthos of Andalusia (Spain). X Palabras clave: Andalucía, corología, macroalgas marinas. Key words. Andalusia, geographical distribution, seaweeds