sm80n4-4318

INTRODUCTIONTop The Mediterranean Sea, due to its low nutrient and chlorophyll concentration, is considered one of the most oligotrophic marine systems (, , ). During the stratification period, a severe nutrient depletion causes both phytoplankton and bacterioplankton to be strongly limited in phosphorus or nitrogen (, , ). However, climatic conditions and the geographic location of the Mediterranean favour the reception of nutrients due to a noticeable dust flux from the Saharan desert (). Around 20 to 50 10⁶ t y^–1 () of dust from the Sahara are transported to the Atlantic ocean through the predominant westerly winds and towards the Mediterranean basin influenced by the presence of cyclones (). Large Saharan dust transport events over the Mediterranean Sea occur commonly in spring and summer (). Saharan dust provides soluble nutrients (e.g. nitrogen, phosphorus and iron) to surface waters, so its deposition in marine waters can favour plankton productivity in the ocean (). During the last few years, the effort to understand the impact of dust deposition on the biogeochemistry of the ocean has increased (, ). In fact, field and experimental studies in several aquatic ecosystems of the Mediterranean region have shown that Saharan dust may stimulate both phytoplankton and bacterioplankton growth (, , ). However, little attention has been paid to the effect of anthropogenic-derived particles, which have a mainly European origin in the NW Mediterranean (e.g. ). Anthropogenic aerosols in the Barcelona area can be a major source of nitrogen and phosphorous to the atmosphere (, ). Furthermore, they are much richer than Saharan particles in organic carbon, particularly in black carbon produced by high temperature combustion processes (, ). On the other hand, anthropogenic aerosols tend to contain high amounts of copper, lead and other trace metals, which are known to be toxic to microbiota at high concentrations (, ). Thus, one way or another, an effect of anthropogenic aerosols on marine production is also expected. Much less attention has been paid to the potential impact of aerosols on the quality of dissolved organic matter (DOM), and in particular on the quality of the optically active fraction (CDOM). This fraction is a key parameter regulating the penetration of ultraviolet radiation in the water column, so changes in its concentration can alter both primary and secondary production (). A sub-fraction of CDOM that emits light when excited by UV radiation is called fluorescent dissolved organic matter (FDOM). Fluorometric analyses can be used to characterize this sub-fraction. Emission fluorescence spectra can be collected at different excitation wavelengths represented in excitation-emission matrices (EEMs), and different peaks of humic- and protein-like fluorophores can be distinguished (, ). Usually, peak C and M are associated with humic-like substances, while peak T corresponds to protein-like substances. The fluorescence intensity of these peaks can be used as indicators of biological () and photochemical processes () of the DOM pool. The optical properties of CDOM are sensitive to biological and physical processes and thus provide valuable information not only of the biogeochemical processes in aquatic environments, but also of the origin of organic matter (OM). determined that the organic carbon associated with dust inputs can contribute to the DOM pool in alpine lakes and that the fraction of airborne water-soluble OM can contain chromophoric groups similar to humic-like substances. More recently, reported that the chromophoric components related to the dust inputs significantly affected water transparency to ultraviolet radiation. The study of aerosol influx as a potential source of CDOM is of particular interest in the Mediterranean Sea because these waters are peculiar for having unusually high values of CDOM to chlorophyll ratio in comparison with other marine systems. Our study aimed to 1) quantify the impact of aerosols of different origin (Saharan and non-Saharan) on the CDOM deposition in the surface waters of a NW Mediterranean coastal system and 2) evaluate the posterior chemical transformations in surface waters by means of CDOM optical signatures. To carry out these objectives, we collected weekly to biweekly samples of atmospheric deposition during 23 months for FDOM analyses concurrently with surface water samples in the Barcelona coastal area. Within this time frame we also conducted two aerosol addition experiments with NW Mediterranean coastal waters, in which we evaluated prokaryote and FDOM dynamics in response to both Saharan dust and anthropogenic aerosols. MATERIALS AND METHODS Top Time series sampling We collected samples for atmospheric deposition and seawater analyses over a two-year period (September 2012 - July 2014). For atmospheric deposition, one high-density polyethylene (HDPE) container was filled with 500 mL of sterile artificial seawater and left open on the roof of the Institute of Marine Sciences (ICM-CSIC, Barcelona, 41°23′08″N, 2º11′45.5″E) for about one week in summer and two in winter. Upon collection, subsamples for FDOM were analysed after filtering them through Whatman GF/F filters. The fluorescence intensities measured for sterile seawater at time 0 were subtracted from those measured at the end of the exposure period. Seawater samples were taken monthly 0.5 km offshore of Barcelona (NW Mediterranean, 41°22′55″N, 2°11′58″E). Surface water was collected in 2-L acid-cleaned polycarbonate bottles and subsamples for FDOM were analysed freshly. Aerosol collection for experiments The aerosols used in the experiments were collected on Munktell quartz filters (quality 360) using an MCV CAV-A/mb high-volume air sampler. The sampler operated for 24 h at 30 m³ h^–1. Filter samples for experimental amendments were obtained at different times in January and March 2014 on the roof of the Institute of Marine Sciences in Barcelona (41°23′08″N, 2°11′45.5″E) and on the roof of the Centre for Advanced Studies of Blanes (CEAB, Blanes, 41°40′59.5″N, 2°48′2.6″E). After collection, half of the filters were used for chemical analyses and the other half were employed for the amendment experiments. Collected aerosols tend to be a mix from different sources. The aerosols were classified according to the relative percentage of Saharan dust versus inputs of anthropogenic origin with previous knowledge of the presence of Saharan events based on transport and deposition models and forecasts (www.calima.ws) and on the chemical analyses of the filters. Aerosols of anthropogenic origin tend to have a higher proportion of non-mineral carbon, nitrogen species and phosphorus, while Saharan dust has a higher proportion of silicate and aluminium oxide (Table 1).

Table 1. – Relative percentage of the composition of the different aerosols used in the BLSp and BCNSp experiments, respectively, analysed from filtered air. Abbreviations: A, anthropogenic; S, Saharan; OC, organic carbon; CO₃, carbon trioxide; SiO₂, silicate oxide; Al₂O₃, aluminium oxide; NO₃^–, nitrate; NH₄^+, ammonium; and P, phosphorus.

	Blanes (BLSp)		Barcelona (BCNSp)
	A	S	A	S
OC	31.95%	4.93%	26.38%	6.75%
SiO₂	4.75%	40.64%	13.56%	27.88%
Al₂O₃	1.58%	13.55%	4.51%	9.29%
NO₃^-	11.01%	2.48%	7.81%	2.11%
NH₄⁺	2.12%	0.37%	1.47%	0.52%
P	0.10%	0.08%	0.13%	0.07%

Water sampling and experimental design Our experiments were conducted with water from two locations that differed in the degree of oligotrophy. The water was collected at the Blanes Bay Microbial Observatory (41°40′0″N, 2°48′0″E) on 8 April 2014 and on the Barcelona coast (41°22′55″N, 2°11′58″E) on 12 May 2014. Blanes Bay Microbial Observatory is characterized as an oligotrophic area with an annual mean of 0.63±0.05 μg L^–1 of chlorophyll (). The Barcelona coastal area is less oligotrophic as it receives nutrients from the discharge of two rivers, the Besòs River located in the north of the city and the Llobregat River in the south. The annual average chlorophyll concentration at the Barcelona station is 1.58±1.09 μg L^–1 (). Both experiments were conducted in mid-spring. This season appears to be the ideal period for testing the impact of dust in surface waters of the Mediterranean Sea, because it is a time interval of the year with frequent dust events (, , ). The experiments were termed BLSp and BCNSp for Blanes and Barcelona, respectively. For both, the water was collected from the surface layer (approximately 0.5 m depth) and pre-filtered through a 150-μm nylon mesh to remove the larger zooplankton. The water was then transported to the laboratory in 50-L carboys, which had previously been washed with a dilute solution of sodium hypochlorite and exhaustively rinsed with Milli-Q water and sample water. In the laboratory, the water was distributed in 15-L cylindrical methacrylate containers, which were subjected to experimental conditions in a light and temperature controlled chamber for 7 days for BLSp and for 5 days for BCNSp. Conditions, in duplicate, were anthropogenic particles enrichment (A), Saharan dust enrichment (S) and control (C) without enrichment. Aerosol concentration added in each container was 0.8 mg L^–1. Light conditions were set to 225 μmol photons m^–2 s^–1 inside the containers and the light:dark cycle (13 h:11 h) and temperature (17.5°C) were adjusted to natural conditions. After placing the containers in the experimental chamber, we left them for acclimation before starting the experiment. Because an in situ Saharan event occurred the day before BLSp water collection, we increased the acclimation period (it lasted 45 hours in BLSp in contrast to 19 hours in BCNSp) to prevent the experimental treatment from being masked by a possible response to the in situ input that occurred in the field. An initial sample was taken and aerosols were subsequently added as single doses. Samples for DOM analyses (CDOM, FDOM and dissolved organic carbon (DOC)) were filtered by glass fibre Whatman GF/F (combusted at 450°C for 4 hours) prior to analysis. Samples were taken at 0, 4, 49, 97, and 144 h for BLSp and at 0, 4, 49 and 97 h for BCNSp. Samples for chlorophyll a (Chl a) and bacteria were taken daily. Analytical procedures CDOM measurements CDOM absorption was measured in 10-cm quartz cuvettes using a Varian Cary UV-VIS spectrophotometer equipped with a 10-cm quartz cell. Absorbance was performed between 250 and 750 nm at a constant room temperature of 20°C. Milli-Q water was used as blank. The residual backscattering (colloidal material, fine size particle fractions present in the sample) was corrected by subtracting the mean absorbance calculated in the spectral range 600-750 nm. The absorption coefficient (aCDOM(λ) in m^–1) was calculated as a_CDOM (λ)=2.303 A(λ) / L where A is the absorbance at wavelength λ, L is the optical path length in m, and 2.303 is the factor that transforms decimal logarithms to natural logarithms. FDOM measurements The samples for FDOM were measured immediately after temperature acclimation according to . Single measurements and emission-excitation matrices were performed with an LS-55 Perkin Elmer luminescence spectrometer equipped with a xenon discharge lamp, equivalent to 20 kW. Slit widths were set to 10 nm for emission and excitation wavelengths and scan speed was 250 nm min^–1. Measurements were performed in a 1-cm quartz cell. The EEMs were generated by concatenating 21 synchronous spectra over excitation wavelengths of 250 to 450 nm and emission wavelengths of 300 to 650 nm with an offset between the excitation and emission wavelengths of 50 nm in the first scan and 250 nm in the last scan. Milli-Q water was used as a blank and Raman scattering was corrected by subtracting the Milli-Q water signal. The samples were converted into quinine sulphate units (QSU). The excitation-emission (Ex/Em) wavelengths used for single measurements were described by : Peak C (Ex/Em 340/440 nm) as an indicator of terrestrial-like substances, peak M (EX/Em 320/410 nm) as an indicator of marine-like substances and peak T (Ex/Em 280/420 nm) as an indicator of protein-like substances. Finally, the fluorescence quantum yield at 340 nm, defined as the portion of light absorbed at 340 nm that is re-emitted as fluorescence [Φ(340)], was determined using the ratio of the absorption coefficient at 340 nm and the corresponding fluorescence emission between 400 and 600 nm of the water sample and referred to the quinine sulphate standard (QS) ratio () :

Φ (340) = \frac{F (400 - 600)}{a_{CDOM} (340)} \cdot \frac{a_{CDOM} {(340)}_{QS}}{F {(400 - 600)}_{QS}} Φ {(340)}_{QS}

where aCDOM(340)_QS is the absorption coefficient of the QS standard at 340 nm (in m^–1); F(400-600) and F(400-600)_QS are the average integrated fluorescence spectra between 400 and 600 nm at a fixed excitation wavelength of 340 nm (in QS units) obtained for each sample and the QS standard, respectively (Romera-Castillo et al. 2011Romera-Castillo C., Nieto-Cid M., Castro C.C., et al. 2011. Fluorescence: Absorption coefficient ratio — Tracing photochemical and microbial degradation processes affecting coloured dissolved organic matter in a coastal system. Mar. Chem. 125: 26-38.); Φ(340)_QS is the dimensionless fluorescence quantum yield of the QS standard and equals 0.54 (Melhuish 1961Melhuish W.H. 1961. Quantum efficiencies of fluorescence of organic substances: effect of solvent and concentration of the fluorescent solute. J. Phys. Chem. 65: 229-235.); and a_CDOM(340) is the absorption coefficient of each sample at 340 nm. In this study, the ratio Φ(340) was calculated to add another descriptor of the coloured dissolved organic matter. It has been shown that this ratio increases when microbial transformations dominate in comparison with photobleaching and vice versa (De Haan 1993De Haan H. 1993. Solar UV-light penetration and photodegradation of humic substances in peaty lake water. Limnol. Oceanogr. 38: 1072-1076., Lønborg et al. 2010Lønborg C., Alvarez-Salgado X.A., Martinez-Garcia S., et al. 2010. Stoichiometry of dissolved organic matter and the kinetics of its microbial degradation in a coastal upwelling system. Aquat. Microb. Ecol. 58: 117-126.).

DOC analysis

Samples for DOC were filtered through Whatman GF/F filters using an acid-cleaned glass filtration system. Approximately 10 mL of water was collected in pre-combusted (450°C for 12 h) glass flasks for DOC determination. After acidification with H₃PO₄ (50 μL) to pH<2 the ampoules were heat-sealed and stored in the dark until analysis. DOC was analysed following the high temperature catalytic oxidation (HTCO) technique (Cauwet 1994Cauwet G. 1994. HTCO method for dissolved organic carbon analysis in seawater: influence of catalyst on blank estimation. Mar. Chem. 47: 55-64., Sugimura and Suzuki 1998Sugimura Y., Suzuki Y. 1998. A high temperature catalytic oxidation method for the determination of non-volatile dissolved organic carbon in sea water by direct injection of a liquid sample. Mar. Chem. 24: 105-131., Cauwet 1999Cauwet G. 1999. Determination of dissolved organic carbon (DOC) and nitrogen (DON) by high temperature combustion. In: Grashoff K., Kremling K., Ehrhard M. (eds) Methods of Seawater Analysis. WILEY-VCH, pp. 407-420.) using a Shimadzu TOC-L analyser. The system was calibrated daily with a solution of acetanilide (C₈H₉NO MW=135.17). The DOC concentration was determined by subtracting the blank samples.

Prokaryotic abundance and Chl a determination

Heterotrophic prokaryotic cells were quantified by flow cytometry according to the method of Gasol and del Giorgio (2000)Gasol J.M., Del Giorgio P.A. 2000. Using flow cytometry for counting natural planktonic bacteria and understanding the structure of planktonic bacterial communities. Sci. Mar. 64: 197-224.. Hereafter the term “prokaryote” will be used as synonym of “heterotrophic prokaryote”. Samples (1.8 mL) were fixed with 0.18 mL of a 10% paraformaldehyde and 0.5% glutaraldehyde mixture. Subsamples of 400 µL were stained with SybrGreen deoxyribonucleic acid fluorochrome and left to stain for 15 min in the dark and then ran at low speed (ca. 30 µL min^–1) through a Becton Dickinson FACSCalibur flow cytometer with a laser emitting at 488 nm. As standard, 10 µL per sample of a 10⁶ mL^–1 solution of yellow-green 0.92-µm latex beads was added.

For total Chl a, a 30-mL sample was filtered through Whatman GF/F glass fibre filters and subsequently extracted with acetone (90%) for 24 h at 4°C in the dark. The fluorescence of the extract was measured with a Turner Designs fluorometer (Yentsch and Menzel 1963Yentsch C.S., Menzel D.W. 1963. A method for the determination of phytoplankton chlorophyll and phaeophytin by fluorescence. Deep Sea Res. 10: 221-231.).

RESULTSTop

FDOM time series

In order to evaluate the potential role of atmospheric deposition on the dynamics of coastal FDOM, we calculated its proportion with respect to the in situ seawater concentration for 23 months (September 2012 to July 2014). High contributions to the DOC pool could be found at all times of the year, although the highest was in summer (July 2013). The results revealed that the deposition of humic-like compounds (peak C and M) and protein-like compounds contributed to an increase in FDOM in surface waters that represented between 0.2% and 3% per m³ per day (Fig. 1). The highest increases (>1% per m³ per day) tended to coincide with Saharan dust events, although not always. The annual amount of these compounds deposited per m² ranged from 1.9 to 2.3 times the average in situ concentration per m³. Thus, for a 10 m water column, the annual atmospheric input is between 19% and 23% of the standing stock.

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Fig. 1. – Daily aerosol deposition-derived FDOM flow to the sea surface as a percentage of concentration in Barcelona coastal waters. Humic-like (peak C and peak M) and protein-like (peak T) substances. The arrows indicate the Saharan dust events. The percentages were calculated as the daily average of the deposition considering five different time frames (one to five days).

Microcosm experiments

Prokaryotic abundance and Chl a

The initial abundance of heterotrophic prokaryotic cells was 8.06×10⁵ cells mL^–1 in BLSp and 1.34×10⁶ cells mL^–1 in BCNSp (Fig. 2A, B). In BLSp, we observed a small increase in cell abundance after aerosol addition that continued to a peak after 1 d. This peak was somewhat larger for A and S than in the control. After this peak, prokaryotes decreased over time, more so in aerosol-amended containers than in the control (Fig. 2A). BCNSp also showed a small initial increase in heterotrophic bacteria abundance following aerosol addition in all the treatments. After that, bacteria peaked at day 1 in S and at day 2 in A and showed some fluctuations. Bacteria in C showed more steady concentrations over time. Unlike in BLSp, bacteria showed no clear decreasing trend after peaking and fluctuated around 1.5·10⁶ cells mL^–1 (Fig. 2B).

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Fig. 2. – Heterotrophic prokaryotic abundance over time for the three treatments of BLSp (A) and BCNSp (B) experiments. Aerosols were added at t=4 h in both experiments.

During BLSp, Chl a, which is a proxy for phytoplankton biomass, increased more in A than in S and C, reaching values of 0.51±0.02, 0.74±0.02 and 0.56±0.01 μg L^–1 for C, A and S, respectively (Fig. 2C). In BCNSp, a peak in S and A was observed at day 1 (higher in A) and then Chl a declined to the end of the experiment, reaching a common value of ca. 0.45 μg L^–1 on the last day in all containers, including C (Fig. 2D).

Dynamics of DOC and FDOM

Aerosols did not significantly alter the DOC concentration (Figs 3D and 4D). The largest difference was found after anthropogenic addition in BLSp (Fig. 3D), where it accounted for an increase of about 10% in the total DOC concentration. After the enrichment, DOC concentrations were maintained fairly constant in all conditions of the BLSp experiment. In BCNSp, DOC values did not increase immediately after addition, but an increase at the end of the experiment occurred in all the containers (Fig. 4D). In both experiments, the increase in humic-like (Figs 3A, 3B, 4A and 4B) and protein-like (Figs 3C and 4C) substances was higher in the treatment enriched with anthropogenic particles than in the one enriched with Saharan dust. Humic substances reached values of about 1.5 QSU in A conditions, while in treatments C and S the maximum values were about 0.74 and 0.94 QSU, respectively. Although the increase in humic compounds after addition in treatment A was higher in BLSp, the different groups of OM followed similar patterns in all treatments in both BLSp and BCNSp (Figs 3A, 3B, 3C, 4A, 4B and 4C). After the initial increase due to dust addition, FDOM values were maintained fairly constant during the course of incubation (Figs 3 and 4).

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Fig. 3. – Fluorescence intensities of FDOM peaks during the course of incubation in the BLSp experiment. (A) peak C, (B) peak M, (C) peak T and (D) dissolved organic carbon (DOC). The FDOM peaks are in quinine sulphate units (QSU) and DOC is in µM. The bars indicate the standard deviation.

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Fig. 4. – Fluorescence intensities of FDOM peaks during the course of incubation in the BCNSp experiment. (A) peak C, (B) peak M, (C) peak T and (D) dissolved organic carbon (DOC). The FDOM peaks are in quinine sulphate units (QSU) and DOC is in µM. The bars indicate the standard deviation.

We compared the FDOM matrices before addition (C) with the changes in FDOM that occurred in each experiment after the addition (Fig. 5). In BLSp, the EEM in treatment A showed marked fluorescence peaks in the humic-like and protein-like areas after addition (reaching concentrations around 1.5 QSU), whereas in the S microcosm the increase in fluorescence intensity was minimal (about 0.2-0.3 QSU) and with no defined peak. In BCNSp, the EEM of the water before addition (C condition, Fig. 5) showed two peaks at Ex/Em 280/350 nm and 250/435 nm, corresponding to peak T and peak A (2.0 and 1.3 QSU, respectively). In contrast, the fluorescence in the treatment A after the addition showed two peaks within the range of the marine and terrestrial humic-like substances (peak M and peak C, respectively). These increases were small (about 0.5 to 0.75 QSU) in comparison with BLSp. Finally, the fluorescence alteration after additions in treatment S was low in comparison with initial fluorescence intensities (values only increased by about 0.1-0.3 QSU in the humic-like area, and by about 0.4 QSU in the protein-like area).

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Fig. 5. – Changes in the excitation-emission matrix (EEM) of FDOM after dust addition in the two experiments. Before addition (C) in column A, and after additions of anthropogenic aerosols (A) in column B and Saharan dust (S) in column C. The different peaks are indicated. The fluorescence is expressed in quinine sulphate units (QSU).

DISCUSSIONTop

Atmospheric deposition influence on surface waters

Our two-year data set on deposition constitutes the first time series that evidences the atmospheric impact on FDOM dynamics in Mediterranean surface waters. Interestingly, several of the highest FDOM deposition values coincided with the Saharan dust events (Fig. 1). Although CDOM is present in low concentrations in the Mediterranean (Romera-Castillo et al. 2013Romera-Castillo C., Álvarez-Salgado X.A., Galí M., et al. 2013. Combined effect of light exposure and microbial activity on distinct dissolved organic matter pools. A seasonal field study in an oligotrophic coastal system (Blanes Bay, NW Mediterranean). Mar. Chem. 148: 44-51., Xing et al. 2014Xing X., Claustre H., Wang H., et al. 2014. Seasonal dynamics in colored dissolved organic matter in the Mediterranean Sea: Patterns and drivers. Deep Sea Res. I 83: 93-101.), the CDOM/chlorophyll ratio is exceptionally high in comparison with other areas (Morel and Gentili 2009Morel A., Gentilli B. 2009. The dissolved yellow substance and the shades of blue in the Mediterranean Sea. Biogeosci. 6: 2625-2636., Organelli et al. 2014Organelli E., Bricaud A., Antoine D., et al. 2014. Seasonal dynamics of light absorption by chromophoric dissolved organic matter (CDOM) in the NW Mediterranean Sea (BOUSSOLE site). Deep Sea Res. I 91: 72-85., Pérez et al. 2016Pérez G.L., Galí M., Royer S.J., et al. 2016. Bio-optical characterization of offshore NW Mediterranean waters: CDOM contribution to the absorption budget and diffuse attenuation of downwelling irradiance. Deep-Sea. Res. I. 114: 111-127.). Because FDOM is a part of the CDOM pool, our observations could indicate that atmospheric inputs during Saharan events could significantly contribute to this high CDOM/chlorophyll ratio. Para et al. (2010)Para J., Coble P.G., Charrière B., et al. 2010. Fluorescence and absorption properties of chromophoric dissolved organic matter (CDOM) in coastal surface waters of the northwestern Mediterranean Sea, influence of the Rhône River. Biogeosci. 7: 4083-4103. pointed out that the humic fluorescent components and the salinity had an exceptionally weak correlation and suggested that other processes could influence CDOM distributions. Thus, our results about FDOM deposition during Saharan events could help to explain these anomalies.

Effects of aerosol additions on prokaryotic abundances

Previous studies have shown that prokaryotic abundances increased in response to Saharan dust inputs in oligotrophic systems (Pulido-Villena et al. 2008Pulido-Villena E., Wagener T., Guieu C. 2008. Bacterial response to dust pulses in the western Mediterranean: Implications for carbon cycling in the oligotrophic ocean. Glob. Biogeochim. Cycles 22: 1-12., Reche et al. 2009Reche I., Ortega-Retuerta E., Romera O., et al. 2009. Effect of Saharan dust inputs on bacterial activity and community composition in Mediterranean lakes and reservoirs. Limnol. Oceanogr. 54: 869-879., Marín et al. 2017Marín I., Nunes S., Sánchez-Pérez E.D., et al. 2017. Anthropogenic versus mineral aerosols in the stimulation of microbial planktonic communities in coastal waters of the northwestern Mediterranean Sea. Sci. Total. Environ. 574: 553-568. ). In our experiments, the prokaryotic response to aerosol addition was relatively low. BLSp showed more oligotrophic conditions than BCNSp, as can be seen from the initial bacterial and Chl a concentrations. In BLSp an initial peak response was seen in all treatments and thus cannot be attributed to aerosol addition, whereas the response of chlorophyll took place much later in time. This is in good agreement with the results obtained by Marañon et al. (2010)Marañon E., Fernández A., Mouriño-Carballido B., et al. 2010. Degree of oligotrophy controls the response of microbial plankton to Saharan dust. Limnol. Oceanogr. 55: 2339-2352., they also observed a quicker response of prokaryotes compared with that of chlorophyll when studying oligotrophic areas. The opposite occurred in more eutrophic areas (Teira et al. 2013Teira E., Hernando-Morales V., Martínez-García S., et al. 2013. Response of bacterial community structure and function to experimental rainwater additions in a coastal eutrophic embayment. Est. Coast. Shelf. Sci. 119: 44-53.). In general, the abundance tended to decrease during the course of the incubations, and this decrease was more conspicuous in the enriched treatments than in the control. We attributed this behaviour to the competition for limiting nutrients between phytoplankton and bacteria (Marañon et al. 2010Marañon E., Fernández A., Mouriño-Carballido B., et al. 2010. Degree of oligotrophy controls the response of microbial plankton to Saharan dust. Limnol. Oceanogr. 55: 2339-2352.). However, the response in both experiments was lower than expected. Ridame (2001)Ridame C. 2001. Rôle des apports atmosphériques d’origine continental dans la biogéochimie marine: Impact des apports sahariens sur la production primaire en Méditerranée. PhD thesis, Université Paris, pp. 213., Marañon et al. (2010)Marañon E., Fernández A., Mouriño-Carballido B., et al. 2010. Degree of oligotrophy controls the response of microbial plankton to Saharan dust. Limnol. Oceanogr. 55: 2339-2352. and Herut et al. (2005)Herut B., Zohary T., Krom M.D., et al. 2005. Response of East Mediterranean surface water to Saharan dust: On-board microcosm experiment and field observations. Deep Sea Res. II 52: 3024-3040. also found low prokaryotic stimulation to aerosol inputs in Mediterranean waters. Differences in the microbial responses seemed to be related to the initial environmental conditions (e.g. nutrient availability). Martinez-García et al. (2015)Martinez-García S., Arbones B., García-Martín E.E., et al. 2015. Impact of atmospheric on the metabolism of coastal microbial communities. Est. Coast. Shelf. Sci. 153: 18-28. also pointed out the importance of initial conditions to explain the variety of microbial responses when examining the effect of rainwater additions in experiments performed with NW Iberian Peninsula shelf waters. In our experiments, the quality of the added particles could be another factor explaining the differences in microbial response between the two experiments. Even if the A and S aerosols were collected during non-Saharan and Saharan events, respectively, the OM composition of the particles differed between locations-experiments. The aerosols collected at the Blanes site induced a higher increase in FDOM in the experimental waters than in the aerosols collected in Barcelona (especially during non-Saharan events). As the weight of total aerosol added was the same between treatments and experiments, these differences indicated local peculiarities related to the quality of the particles. However, more data are needed to investigate the causes of these differences in quality and whether or not they are persistent.

Transformations of DOM optical properties after dust additions

In BLSp, the humic-like fractions of OM increased after the addition of aerosols, and this increase was more conspicuous in treatment A than in treatment S. In fact, peak C/DOC ratios in QSU/µmol C L^–1 were 153% higher in A and 33% higher in S than in the control. However, FDOM compounds showed no variation during the incubation time. In BCNSp, FDOM values were also higher after the addition. However the increases in FDOM/DOC ratio after additions did not differ significantly between A and S. In both BLSp and BCNSp, we observed that DOC tended to increase in all treatments during the time of incubation. This increase was larger in BCNSp than in BLSp, which is in accordance with a high activity of phytoplankton (Fig. 2C, D). The EEMs confirmed that both anthropogenic and Saharan aerosols contained fluorescence organic substances (Fig. 5), as has been previously reported by Mladenov et al. (2011)Mladenov N., Sommaruga R., Morales-Baquero R., et al. 2011. Dust inputs and bacteria influence dissolved organic matter in clear alpine lakes. Nat. Commun. 2: 405..

Regarding the fluorescence quantum yield at 340 nm [Φ(340)], we observed a similar increase in both experiments after the addition was performed, reaching values of about 0.65% (Fig. 6). The quantum yield decreases with light exposure and increases with microbial activity (Romera-Castillo et al. 2011Romera-Castillo C., Nieto-Cid M., Castro C.C., et al. 2011. Fluorescence: Absorption coefficient ratio — Tracing photochemical and microbial degradation processes affecting coloured dissolved organic matter in a coastal system. Mar. Chem. 125: 26-38.). Therefore, the increase in quantum yield values observed after the enrichment could indicate a rapid, although low, bacterial response. In fact, we observed that low values of Φ(340) coincided with low bacterial abundance and vice versa. The values of Φ(340) obtained in our experiments were within the range of other data reported previously from field studies in the Mediterranean (Ferrari 2000Ferrari G. 2000. The relationship between chromophoric dissolved organic matter and dissolved organic carbon in the European Atlantic coastal area and in the West Mediterranean Sea (Gulf of Lions). Mar. Chem. 70: 339-357., Romera-Castillo et al. 2011Romera-Castillo C., Nieto-Cid M., Castro C.C., et al. 2011. Fluorescence: Absorption coefficient ratio — Tracing photochemical and microbial degradation processes affecting coloured dissolved organic matter in a coastal system. Mar. Chem. 125: 26-38.), thus indicating that our induced changes in optical characteristics of OM were within the range of variations occurring in nature.

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Fig. 6. – Fluorescence quantum yield at 340 nm [Φ(340)] before and after anthropogenic and Saharan dust addition. A, BLSp experiment; B, BCNSp experiment. The [Φ(340)] is expressed in percentage (%).

CONCLUSIONSTop

A two-year time series data shows, for the first time, the influence of atmospheric deposition on the temporal dynamics of FDOM in Mediterranean surface waters. This time series data evidences an increase in the FDOM flux during Saharan events. Our experimental results revealed that aerosol deposition induced an increase in the proportion of FDOM in comparison with DOC. The refractory character of the OM added with aerosols was confirmed from the negligible utilization of this fraction within a short time period (days). Thus, considering our in situ results showing how dust deposition increases the coloured DOC content in surface waters, together with the experimental findings that corroborate the low biological utilization of this coloured fraction, we conclude that the atmospheric deposition could help to explain the exceptionally high CDOM/ chlorophyll values found in the Mediterranean Sea.

ACKNOWLEDGEMENTSTop

We thank Raquel Gutiérrez for her help in the processing of data from atmospheric deposition. We also thank J. Caparros for the analysis of dissolved organic carbon. This study was supported by the projects ADEPT (CTM2011-23458), DOREMI (CTM2012-342949) and ANIMA (CTM2015-65720-R MINECO/FEDER, UE). E.D. Sánchez-Pérez would like to thank the Consejo Nacional de Ciencia y Tecnologia (CONACyT) for their financial support through a PhD fellowship. I. Marín thanks the FPI Spanish scholarship programme for its support (BES 2012-052976).

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