Sfax Solar Saltern Research Articles

Hydrolytic enzyme screening and carotenoid production evaluation of halophilic archaea isolated from highly heavy metal-enriched solar saltern sediments.

This paper aimed to screen the enzymatic activities and evaluate the carotenoid production level of twenty-two halophilic archaea isolated from Sfax solar saltern sediments. The molecular identification performed by sequencing the 16S rRNA genes showed that all strains have a high similarity degree (99.7-100%) with Halobacterium salinarum NRC-1. The strains were screened for the presence of eight hydrolase activities using agar plate-based assays. The most detected enzyme was gelatinase (77.27% of total strains), followed by protease (63.63%) and amylase activities (50%). The carotenoid production yields of the strains ranged between 2.027 and 14.880mg/l. The UV-Visible spectroscopy of pigments revealed that it was a bacterioruberin type. When evaluated and compared to the standard β-carotene, the antioxidant capacities of these pigments showed a scavenging activity of more than 75% at a concentration of 5μg/ml for three strains (AS16, AS17, and AS18). Then a sequence of one-step optimization processes was performed, using the one-factor-at-a-time approach, to define the optimum conditions for growth and carotenoid production of the highest carotenoid producing strain (AS17). Different environmental factors and nutritional conditions were tested. Variations in these factors were found to deeply influence growth and carotenoid production. A maximum carotenoid production (16.490mg/l), higher than that of the control (14.880mg/l), was observed at 37°C, pH 7, 250g/l of salinity, with 80% air phase in the flask at 110rpm, in presence of light and in culture media containing (g/l) 10, yeast extract; 7.5, casamino acid; 20, MgSO4; 4, KCl; and 3, trisodium citrate.

Genomic analysis of heavy metal-resistant Halobacterium salinarum isolated from Sfax solar saltern sediments

The draft genome sequences of five archaeal strains, isolated from Sfax solar saltern sediments and affiliated with Halobacterium salinarum, were analyzed in order to reveal their adaptive strategies to live in hypersaline environments polluted with heavy metals. The genomes of the strains (named AS1, AS2, AS8, AS11, and AS19) are found to contain 2,060,688; 2,467,461; 2,236,624; 2,432,692; and 2,428,727 bp respectively, with a G + C content of 65.5, 66.0, 67.0, and 66.2%. The majority of these genes (43.69–55.65%) are annotated as hypothetical proteins. Growth under osmotic stress is possible by genes coding for potassium uptake, sodium efflux, and kinases, as well as stress proteins, DNA repair systems, and proteasomal components. These strains harbor many genes responsible for metal transport/resistance, such as: copper-translocating P-type ATPases, ABC transporter, and cobalt-zinc-cadmium resistance protein. In addition, detoxification enzymes and secondary metabolites are also identified. The results show strain AS1, as compared to the other strains, is more adapted to heavy metals and may be used in the bioremediation of multi-metal contaminated environments. This study highlights the presence of several commercially valuable bioproducts (carotenoids, retinal proteins, exopolysaccharide, stress proteins, squalene, and siderophores) and enzymes (protease, sulfatase, phosphatase, phosphoesterase, and chitinase) that can be used in many industrial applications.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00792-022-01273-0.

Behavior Assessment of Moderately Halophilic Bacteria in Brines Highly Enriched with Heavy Metals: Sfax solar saltern (Tunisia), A Case Study

This study, based on complementary descriptive and analytical approaches, focuses on the behavior of moderately halophilic Bacteria toward selected heavy metals (Zn, Cd, Cu and Pb) in Sfax solar saltern brines (Tunisia). The brines metal concentrations (mg/L) varied from 0.065 to 1.57 for Zn, 0.002–0.034 for Cd, 0.006–0.064 for Cu and 0.002–0.128 for Pb. Analyzed metals have been ranged in the following order: Zn > Pb > Cu > Cd. Computed enrichment factors (EFs) showed a significant contamination of brines due the impact of industrial particulate fallouts highly enriched with heavy metals. Multivariate statistical analysis results, concerning the principal component analysis, the hierarchical cluster analysis and the multiple regression analysis revealed that the salinity, the temperature, TOC, Pb and Cd have a negative significant influence on Bacteria growth. Only pH, O2, Zn and Cu were significant positive factors associated with their growth density. The analysis of variance applied to growth parameters of nine bacterial strains affiliated to genera Halomonas, Cobetia, Idiomarina, Marinobacter and Salinivibrio, already isolated from the brines, showed that pH values, temperature and NaCl concentrations have different effects on growth rate (p values < 0.05). The optimum growth of moderately halophilic bacteria was obtained at NaCl = 50 g/L, pH = 7 and T = 35 °C. The Minimum Inhibitory Concentrations (MICs) of such strains revealed their highly resistance to Pb and Cd, while response to Zn and Cu was variable. The order of toxicity was found to be Cu > Zn > Cd > Pb. Halomonas sp. K22 and Salinivibrio costicola were the most resistant strains.

Resistance of a Halobacterium salinarum isolate from a solar saltern to cadmium, lead, nickel, zinc, and copper.

The current study focuses on the tolerance of a strain of Halobacterium salinarum isolated from Sfax solar saltern (Tunisia) towards cadmium (Cd), lead (Pb), nickel (Ni), zinc (Zn), and copper (Cu) by using agar dilution methods in complex and minimal media. The results showed the least inhibitory metals based on Minimum Inhibitory Concentrations (MICs) were lead (MIC = 4.5mM), cadmium (MIC = 4mM), and nickel (MIC = 2.5mM) in complex medium. The MICs of these metals were more inhibitory (MIC < 2mM) in the other tested media. The archaeal strain revealed a high sensitivity for copper and zinc, with MICs below 0.5mM for both metals. Growth kinetics in complex and minimal media showed the strain to be more sensitive to the metals in liquid media than in solid media. The growth kinetic assays indicated the presence of selected heavy metals resulted in a lower growth rate and lower total cell mass relative to the control. Despite that cadmium and lead are nonessential and have no nutrient value, they were the most tolerated metals by H. salinarum strain. In addition, pigment intensity in the strain was inhibited by the presence of the heavy metals relative to the control.

Metal Contamination and Resistance of Superficial Sediment’s Prokaryotic Flora in Extreme Environments: Case of Sfax Solar Saltern (Tunisia)

The current study focuses on the distribution of heavy metals in 17 superficial sediments of Sfax solar saltern ponds (Tunisia) and their impacts in prokaryotic flora. The results showed significant accumulation of Pb, Zn, Cu, Ni, and Cd in sediments. Maxima of Cd, Pb, and Ni concentrations were respectively more than 500, 70 and 3 times higher than those recorded in the most worldwide heavy metals contaminated solars. Complementary statistical approaches based on correlation matrix, principal component analysis (PCA), hierarchical cluster analysis (HCA) and multiple linear regression models highlight the relationship between the Bacteria and extremely halophilic Archaea and the other studied parameters (salinity, heavy metals and TOC). It was demonstrated that Bacteria and Archaea have different behaviors toward these parameters. Only, Archaea seemed to be sensitive to selected heavy metal concentrations. Their inter-pond variability rates were mainly conditioned by ‘Pb, Ni and TOC’ variability rates. The increase rate of Archaea was associated with an increase of Pb, Ni and TOC rates (and vice versa). However, Bacteria's variability rate dosent shows any dependence to all studied parameters. One could presume a good resistance of Archaea to Pb and Ni, probably reinforced by organic matter. Archaeal strains isolated from the most contaminated ponds were shown tolerant to high concentrations of Pb, Ni and Cd up to 2.5–4.5 mM. For Cu and Zn, the Archaea resistance levels were not, however, exceeded 1 mM.

Pollution characteristics and health risk assessment of heavy metals in dry atmospheric deposits from Sfax solar saltern area in southeast of Tunisia

Concentrations of six heavy metals such as Fe, Pb, Cu, Cd, Ni and Zn in dry atmospheric deposits weekly collected through 20 sample sites from Sfax solar saltern during two successive sampling campaigns, selected from a long time monitoring, have been examined in order to evaluate their contamination levels as well as their human health risk; such concentrations (expressed in mg/kg) have shown that Fe varied from 7006.24 to 7856.45, Pb from 8.64 to 344.45, Cu from 9.33 to 298.67, Cd from 0.16 to 85.24, Ni from 6.02 to 150.02, and Zn from 12.23 to 624.57. During the study period, average concentrations (mg/kg) have been 7315.99, 103.08, 82.34, 15.93, 46.21 and 142.39, for Fe, Pb, Cu, Cd, Ni and Zn, respectively. Except for Fe, the other concentrations in dry atmospheric deposits have recorded the highest level during the first campaign especially in the sites which are close and more exposed to emissions of local pollutant industries, as well as nearby main road, resulting from high exposure frequencies. Statistical approaches, such as principal component analysis and hierarchical cluster analysis have been applied to all data, revealing an affinity between analyzed metals resulting from their origins, and confirming the influence of exposure frequencies on the spatial variability of heavy metal concentrations. Human health risk assessment has revealed that ingestion of heavy metals is the main exposure pathway for the local population. Computed Hazardous Quotient and Hazardous Index have been higher for children than for adults, thus indicating that the former may be subjected to potentially higher health risk than the latter especially during the first campaign. Calculated carcinogenic risks through ingestion and dermal contact, as well as the total carcinogenic risk for the selected heavy metals, have shown that cancer could occur more probably through ingestion than dermal contact, for children than adult, and during the first campaign (during C1: average values CRing = 8.72 × 10-4 and CRder = 1.40 × 10-6 for child; average values CRing = 5.61 × 10-5 and CRder = 2.84 × 10-6 for adult) than the second one (during C2: average values CRing = 1.59 × 10-4 and CRder = 2.54 × 10-7 for child; average values CRing = 1.02 × 10-5 and CRder = 5.19 × 10-7 for adult). The total calculated carcinogenic risk through all the sites have infrequently signaled high to very high carcinogenic risk in the first campaign (average CRA = 8.73 × 10-4 for child and CRA = 5.89 × 10-5 for adult) and occasionally exceeded the safe range for the local population in the second one (average CRA = 1.59 × 10-4 for child and CRA = 1.07 × 10-5 for adult).

Abiotic stress resistance, plant growth promotion and antifungal potential of halotolerant bacteria from a Tunisian solar saltern

The uses of halotolerant bacteria isolated from naturally saline habitats have the potential to be useful crop protection agents for plants in stressful conditions. These beneficial microbes generate several plant growth regulators and bioactive molecules, which enhance plant protection from adversities, such as plant pathogens, salts and metals stresses. In this study, 15 halotolerant bacterial strains endowed with important antimicrobial activities were isolated from Sfax solar saltern (Tunisia). All of these strains were characterized by biochemical and molecular tools aiming to investigate their in-vitro and in-vivo antifungal potentialities, plant growth promotion capabilities and metal tolerance abilities under saline stress condition. The 16S rRNA gene sequencing showed that the isolated strains were affiliated to different phylum and three species were described for the first time as plant growth promoting strains (Idiomarina zobelli FMH6v, Nesterenkonia halotolerans FMH10 and Halomonas janggokensis FMH54). The tested strains exhibited several potentialities: to tolerate high salt and heavy metal concentrations, to produce biosurfactants, exopolysaccharides and extracellular hydrolytic enzymes, to form biofilms and to liberate plant promoting substances. Eight strains were able to protect tomatoes fruits from the proliferation of the fungal disease caused by Botrytis cinerea and six strains improved plant vigor indexes. Principal component analysis showed an important correlation between in-vitro and in-vivo potentialities and two strains Bacillus velezensis FMH2 and Bacillus subtilis subsp. spizizenii FMH45 were statistically considered as the most effective strains in protecting plants from fungal pathogens attack and promoting the growth of tomatoes seedlings under saline and multi heavy-metals stress conditions.

Assessment of heavy metals contamination and their potential toxicity in the surface sediments of Sfax Solar Saltern, Tunisia

Concentrations of six heavy metals such as Fe, Pb, Zn, Cu, Ni and Cd have been examined in 20 surface sediment samples from Sfax solar saltern in order to evaluate their contamination levels; such concentrations (expressed in mg/kg of dry weight) have shown that Fe varied from 8750 to 8889.1, Pb from 18.98 to 233.46, Zn from 39.92 to 574.89, Cu from 13 to 98, Ni from 17.47 to 160.92, and Cd from 4.86 to 37.42. Importantly, the highest metal concentrations—except for Fe—have been more significant in sites frequently exposed to the industrial plumes of the local pollutant sources on the one hand and in sites often overwhelmed by high tide marine’s water draining industrial waste from the port area on the other hand. Calculated enrichment factors have shown a spatial distribution in consistency with that related to concentrations. Complementary statistical approaches based on principal component analysis and hierarchical cluster analysis have proved that Fe is natural and other analyzed metals are anthropogenic. The geoaccumulation index has shown different contamination and toxicity levels, which have been confirmed by the study of mean-effect range medium-quotient, demonstrating a high probability of toxicity ranging from 49 to 76%, especially at sites with the highest metal concentrations. It has been suggested by the potential ecological risk index that the combined ecological risk of anthropogenic metal differ from one site to another, which highly significant in the case of Cd.

Evaluation of the biotechnological potential of a novel purified protease BS1 from Bacillus safensis S406 on the chitin extraction and detergent formulation

An extracellular alkaline stable protease BS1 from a new bacteria strain, Bacillus safensis S406, isolated from the Sfax solar saltern, was purified and characterized. The enzyme was purified to homogeneity by ammonium sulfate precipitation, Sephadex G-75 gel filtration, Mono-Q anion-exchange chromatography and ultrafiltration, with a 12.70-fold increase in specific activity and 20.29% recovery. The enzyme has a molecular weight of 29kDa and appeared as a single band on native-PAGE. The optimum pH and temperature values of its proteolytic activity were pH 11.0 and 60°C, respectively. BS1 was tested for the deproteinization of shrimp wastes to extract chitin. An enzyme-protein ratio of 10U/mg of proteins allows to eliminate 93% of protein linked to the chitin after 3h hydrolysis at 45°C. Being very active in alkaline conditions, the potential application of BS1 in laundry formulation was investigated. The enzyme showed high stability in the presence of non-ionic surfactants and some commercial liquid and solid detergents, suggesting its eventual use in detergent formulations.

Antagonistic interactions and production of halocin antimicrobial peptides among extremely halophilic prokaryotes isolated from the solar saltern of Sfax, Tunisia.

Thirty-five extremely halophilic microbial strains isolated from crystallizer (TS18) and non-crystallizer (M1) ponds in the Sfax solar saltern in Tunisia were examined for their ability to exert antimicrobial activity. Antagonistic assays resulted in the selection of eleven strains that displayed such antimicrobial activity and they were further characterized. Three cases of cross-domain inhibition (archaea/bacteria or bacteria/archaea) were observed. Four archaeal strains exerted antimicrobial activity against several other strains. Three strains, for which several lines of evidence suggested the antimicrobial activity was, at least in part, due to peptide/protein agents (Halobacterium salinarum ETD5, Hbt. salinarum ETD8, and Haloterrigena thermotolerans SS1R12), were studied further. Optimal culture conditions for growth and antimicrobial production were determined. Using DNA amplification with specific primers, sequencing and RT-PCR analysis, Hbt. salinarum ETD5 and Hbt. salinarum ETD8 were shown to encode and express halocin S8, a hydrophobic antimicrobial peptide targeting halophilic archaea. Although the gene encoding halocin H4 was amplified from the genome of Htg. thermotolerans SS1R12, no transcript could be detected and the antimicrobial activity was most likely due to multiple antimicrobial compounds. This is also the first report that points to four different strains isolated from different geographical locations with the capacity to produce identical halocin S8 proteins.

Characterization of halo-alkaline and thermostable protease from Halorubrum ezzemoulense strain ETR14 isolated from Sfax solar saltern in Tunisia.

A total of 54 halophilic strains were isolated from crystallizer TS18 (26 strains) and non-crystallizer M1 (28 strains) ponds and screened for their ability to produce protease, amylase, and lipase activities. Enzymatic assays allowed the selection of thirty-two active strains, among them, the ETR14 strain from TS18 showed maximum protease production yields and therefore, selected for further analysis. The results from 16S rRNA gene sequence analysis revealed that the strain belonged to Halorubrum ezzemoulense (Hrr. ezzemoulense) species. Further results indicated that optimum growth and protease production yields were obtained with 10-15% NaCl concentrations in the DSC-97 medium. The enzyme was also able to maintain high levels of protease activity at salt concentrations of up to 25%. While readily available carbon sources were noted to significantly reduce protease production, the combination between yeast extract and peptone enhanced protease excretion, which reached a maximum of 284 U ml(-1) at the end of the exponential growth phase. The enzyme exhibited optimum activity at pH 9 and 60 °C. The halophilic protease retained 87% of its initial activity after 1 h incubation at 70 °C and showed high stability over a wide range of pH, ranging from 7 to 10. This protease exhibited good temperature, pH, and salinity tolerance, which distinguishes it from other proteases previously described from other members of the holoarchaea genera and makes it a promising candidate for application in various industries.

The effect of environmental factors on the fatty acid composition of copepods and Artemia in the Sfax solar saltern (Tunisia)

The biochemical composition and abundance variation of zooplankton (copepods and Artemia salina) were determined in four ponds of increasing salinity (A5, A16, C41 and M2) in the Sfax solar saltern (Tunisia). The zooplankton community was dominated by copepods in the ponds A5, A16 and C41. The pond M2 was marked by the presence of only Artemia salina. Our results showed the dominance of total saturated fatty acids (SFA), which made up 57%–95% of total fatty acids (TFA). SFA 16:0 and 18:0 dominate in all ponds. A. salina showed the highest amounts of the total monounsaturated (MUFA) and polyunsaturated fatty acids (PUFA), this indicates that this species could be employed in hatcheries and used as food source for some aquarium species. Fatty acids of herbivory, proportion of all diatom markers to all flagellate markers (D/F), were negatively correlated with the total zooplankton (r = −0.998, p < 0.05). A. salina was negatively correlated with a biomarker for carnivory polyunsaturated fatty acids/saturated fatty acids (PUFA/SFA) (r = −0.959, p < 0.05). The dietary quality of zooplankton seems to be dependent on food availability in the four studied ponds.

Impacts of restoration of an uncontrolled phosphogypsum dumpsite on the seasonal distribution of abiotic variables, phytoplankton, copepods, and ciliates in a man-made solar saltern

The restoration of an uncontrolled phosphogypsum landfill was investigated for its effects on the seasonal distribution of phytoplankton, ciliates, and copepods. Sampling was carried out monthly from September 2007 to August 2008 at four ponds of increasing salinity (A1, 41psu; A5, 46psu; A16, 67psu; and C31, 77psu) in the Sfax solar saltern (southeastern Tunisia). Physicochemical and biological analyses were carried out using standard methods. Results showed drastic reduction of phosphate input and greater diversity of phytoplankton, ciliates, and copepods than before restoration. Pennate diatoms and new ciliates, considered bio-indicators of less-stressed marine ecosystems, proliferated in the A1 pond for the first time after restoration. Copepods appeared to feed on a wide range of prey. Economically, removal of the 1.7 million m(3) of phosphate improved the quality of the site's salt production, enabling the salt company to receive the quality ISO 9001 accreditation.

Spatial and seasonal prokaryotic community dynamics in ponds of increasing salinity of Sfax solar saltern in Tunisia

The spatial and seasonal dynamics of the halophilic prokaryotic community was investigated in five ponds from Sfax solar saltern (Tunisia), covering a salinity gradient ranging from 20 to 36%. Fluorescence in situ hybridization indicated that, above 24% salinity, the prokaryotic community shifted from bacterial to archaeal dominance with a remarkable increase in the proportion of detected cells. Denaturing gradient gel electrophoresis (DGGE) profiles were rather similar in all the samples analyzed, except in the lowest salinity pond (around 20% salt) where several specific archaeal and bacterial phylotypes were detected. In spite of previous studies on these salterns, DGGE analysis unveiled the presence of microorganisms not previously described in these ponds, such as Archaea related to Natronomonas or bacteria related to Alkalimnicola, as well as many new sequences of Bacteroidetes. Some phylotypes, such as those related to Haloquadratum or to some Bacteroidetes, displayed a strong dependence of salinity and/or magnesium concentrations, which in the case of Haloquadratum could be related to the presence of ecotypes. Seasonal variability in the prokaryotic community composition was focused on two ponds with the lowest (20%) and the highest salinity (36%). In contrast to the crystallized pond, where comparable profiles between autumn 2007 and summer 2008 were obtained, the non-crystallized pond showed pronounced seasonal changes and a sharp succession of "species" during the year. Canonical correspondence analysis of biological and physicochemical parameters indicated that temperature was a strong factor structuring the prokaryotic community in the non-crystallizer pond, that had salinities ranging from 20 to 23.8% during the year.

Impact of a Mixed “Industrial and Domestic” Wastewater Effluent on the Southern Coastal Sediments of Sfax (Tunisia) in the Mediterranean Sea

Concentrations of Zn, Cu, Cr, Cd, Pb and Fe in surface marine sediments of the southern coast of Sfax were studied in order to understand current metal contamination at the outlet of a mixed industrial and domestic wastewater effluent discharged into the seawater since 1953. We found that heavy metal concentrations (mg/kg dry weight) in surface marine sediments varied from 42 to 391 for Zn, 15 to 44 for Cu, 44 to 137 for Cr, 6 to 56 for Cd, 19 to 59 for Pb and 52344 to 54543 for Fe. The computed metal enrichment factors (EF) in the sediments showed that EF of Zn ranged from 0.49 to 9.13, EF of Cu from 0.35 to 3.04, EF of Cr from 0.3 to 3.05, EF of Cd from 0.63 to 9.37 and EF of Pb from 0.43 to 3. All heavy metals were characterized by maxima EF values greater than 1.5, indicating a considerable metallic pollution of the study area. The PCA analysis revealed two distinct groups: (i) the first group (G 1 ) is representative of Fe selected as natural tracer and (ii) the second group (G 2 ) is articulated around Zn, Cu, Cr, Cd and Pb. These metals which did not show any correlations with Fe of the first group imply a significantly impact of the selected wastewater effluent. The calculation of the geoaccumulation index (I geo ) showed that all analyzed metals (except for Fe) can be considered as moderate to extreme pollutants. The microbial analysis of seawater samples and also of both water and sediment samples collected from the Sfax solar saltern (placed at the vicinity of the outlet of the selected effluent) showed that no indicator germs of contamination (total coliforms, faecal coliforms and faecal streptococci) were detected.

Novel prokaryotic diversity in sediments of Tunisian multipond solar saltern

The phylogenetic diversity of a sediment microbial community from two ponds having different salinities, 150–200 g/l (M2) and 250–300 g/l (TS38), of an Sfax (Tunisia) solar saltern, was investigated using 16S rRNA clone libraries. The 16S rRNA genes from 135 bacterial clones and 105 archaeal clones were sequenced and phylogenetically analyzed. 32 operational taxonomic units (OTUs) were generated for Bacteria and 64 for Archaea. The bacterial community in M2 sediment was affiliated only with Bacteroidetes, while that in TS38 sediment was dominated by clones affiliated with Bacteroidetes, (gamma, alpha, delta) Proteobacteria and unclassified bacteria; these represented 56.52, 26.08, 4.34, 4.34 and 8.7% of the OTUs, respectively. In the M2 and TS38 sediments, 44.44 and 43.47% of the bacterial OTUs, respectively, were novel. All archaeal sequences fell into the Euryarchaeota phylum. In both sediments, 38.46 and 72.55% of the OTUs had less than 97% 16S rRNA sequence identity, representing novel OTUs. Two sequences, retrieved from TS38 sediment, were found to be affiliated with the candidate division MSBL-1 defining two OTUs. The sediment phylogenetic study revealed the presence of a highly diverse microbial population in highly salty media.

Biochemical adaptation of phytoplankton to salinity and nutrient gradients in a coastal solar saltern, Tunisia

The distribution of protein and carbohydrate concentrations of the particulate matter (size fraction: 0.45–160 μm) was studied, from 22 January 2003 to 02 December 2003, in three ponds of increasing salinity in the Sfax solar saltern (Tunisia). The coupling of N/P: DIN (DIN = NO 2 − + NO 3 − + NH 4 +) to DIP (DIP = PO 4 3−) with P/C: protein/carbohydrates ratios along salinity gradient allowed the discrimination of three types of ecosystems. Pond A1 (mean salinity: 45.0 ± 5.4) having marine characteristics showed enhanced P/C ratios during a diatom bloom. N/P and P/C ratios were closely coupled throughout the sampling period, suggesting that the nutritional status is important in determining the seasonal change in the phytoplankton community in pond A1. In pond A16 (mean salinity: 78.7 ± 8.8), despite the high nitrate load, P/C ratios were overall lower than in pond A1. This may be explained by the fact that dinoflagellates, which were the most abundant phytoplankton in pond A16 might be strict heterotrophs and/or mixotrophs, and so they may have not contributed strongly to anabolic processes. Also, N/P and P/C ratios were uncoupled, suggesting that cells in pond A16 were stressed due to the increased salinity caused by water evaporation, and so cells synthesized reserve products such as carbohydrates. In pond M2 (mean salinity: 189.0 ± 13.8), P/C levels were higher than those recorded in either pond A1 or A16. N/P and P/C were more coupled than in pond A16. Species in the hypersaline pond seemed paradoxally less stressed than in pond A16, suggesting that salt-tolerant extremophile species overcome hypersaline constraints and react metabolically by synthesizing carbohydrates and proteins.

Abundance and biomass of prokaryotic and eukaryotic microorganisms coupled with environmental factors in an arid multi-pond solar saltern (Sfax, Tunisia)

The distribution of abundance and biomass of prokaryotes, flagellates, ciliates and phytoplankton, were studied in five ponds of increasing salinity in the Sfax solar saltern (Tunisia) coupled with environmental factors. The results showed that abundance of eukaryotic microorganisms decreased with increasing salinity of the ponds whereas prokaryotes (heterotrophic bacteria and Archaea) were abundant in the hyper-saline ponds. Phototrophic picoplankton was found in a large range of salinity values (70 and 200‰). Phototrophic non-flagellated nanoplankton which dominated in the first sampled pond was substituted by phototrophic flagellated nanoplankton in the other ponds. Heterotrophic nanoplankton dominated in the crystallizer pond but its quantitative importance declined in the less saline ponds. Diatoms and dinoflagellates were the major contributors to phytoplankton abundance in the first ponds (&gt;90% of total abundance). Ciliated protozoa were found in all the ponds except in the crystallizer in which prokaryotes proliferated. Oligotrichida and Heterotrichida were the most abundant ciliate groups. Overall, species richness decreased with salinity gradient. We propose a simplified diagram of the Sfax saltern's food web showing the dominant role of the microbial loop along the salinity gradient.

Coupling changes in fatty acid and protein composition ofArtemia salinawith environmental factors in the Sfax solar saltern (Tunisia)

The biochemical composition and biometry of different Artemia salina stages were determined in four ponds of increasing salinity (M1, M2, M3 and B1) in the Sfax solar saltern (Tunisia). Results showed the dominance of saturated fatty acids, which made up 48 to 57% of total fatty acids (FAs). Polyunsaturated fatty acids (PUFAs) 22:6(n-3) docosahexaenoic acid (DHA) and 20:5(n-3) eicosapentaenoic acid (EPA) represented on average only 3.1 and 4.0% of total FAs respectively. A. salina nauplii, cysts and metanauplii, in ponds M1, M2 and B1 respectively, were found to have optimal DHA/EPA ratios (>2) for use as live feed for invertebrate and fish larvae. Significant inter-pond variation in DHA/EPA levels was also recorded. The predominant FAs in Artemia were negatively correlated with both temperature and salinity. FA and protein contents were strongly affected by high temperatures (>30 ◦ C) and probably by food sources (e.g., Dunaliella salina )( r = 0.9, n = 27). The density of Artemia salina was positively correlated with protein content in pond B1. The high DHA/EPA ratios (1 to 3.3) found in this study indicate that Sfax Artemia could be a valuable food source for larvae in large marine hatcheries and also for some aquarium species.