The experimental platform was composed of submerged enclosures (1.2 m diameter and 2 m depth) which allowed the isolation of up to 2,000 L and the simulation of
UVBR and temperature selleck increases in order to study the responses of pelagic communities to these manipulated factors simultaneously. The regulations of UVBR and temperature are performed with high frequency monitoring following the in situ temperature and natural incident UVBR (see details in supplementary data; full description in Nouguier et al. [25]). Four enclosures, filled with lagoon surface-water at random, were used as incubators for the 2 L experimental bags (UV-permeable sterile Whirl Pack® polyethylene bags incubated at subsurface) in which microbial communities were isolated. The factorial experimental
design constituted eight different treatments HMG-CoA Reductase inhibitor (each being tested in three replicates): C: control, C + Nut: control with nutrient addition, UV: UVBR increase (+20%), UV + Nut: UVBR increase (+20%) and nutrient addition, T: temperature increase Ruboxistaurin cost (+3°C), T + Nut: temperature increase (+3°C) and nutrient addition, TUV: temperature (+3°C) and UVBR (+20%) increases, TUV + Nut: temperature (+3°C) and UVBR increases (+20%) and nutrient addition (Figure 1). Figure 1 Crossed factorial experimental design conducted to assess the effects of the three regulatory factors: (Temperature, UVB radiation and nutrient increases). In order to fill the 24 Whirl Pack bags, 100 L subsurface lagoon water was pumped and pre-filtered through 6-μm-pore-size
polycarbonate membranes (47 mm in diameter) in order to isolate the smallest planktonic fraction. This water sample (<6 μm) was equally distributed into 24 sterile Whirl Pack® polyethylene bags. 12 of these experimental bags received nutrients addition at time zero, while the others were kept without nutrient addition. The set bags which represented the enriched Alanine-glyoxylate transaminase nutrient conditions were obtained by addition of a mixture of leucine (C and N) and phosphate in order to maintain a substrate C:N:P molar ratio close to that of marine bacteria [26] as described in Bouvy et al.[24]. The bags with and without nutrient addition exhibited concentrations of 0.20 μM and 0.07 μM of PO4, respectively. The two levels of P concentration mimicked natural fluctuations in coastal lagoon waters. These concentrations were chosen to be relevant to phosphorus concentrations recently measured in Thau lagoon (a general decrease over the past 30 years has led to low values of soluble reactive phosphorus: i.e. from 3 μM to undetectable values (<0.03 μM in winter) [27]). Since nutrients usually refer to inorganic nutrients, it should be noted that in this study, “nutrients” actually refer to “nutrients and organic source of C and N”.