A Glimpse at Siderophores Production by Anabaena flos-aquae UTEX 1444

Abstract

In this study, a strain of Anabaena flos-aquae UTEX 1444 was cultivated in six different concentrations of iron (III). Cultures were extracted with organic solvents and analyzed using our dereplication strategy, based on the combined use of high-resolution tandem mass spectrometry and molecular networking. The analysis showed the presence of the siderophores’ family, named synechobactins, only in the zero iron (III) treatment culture. Seven unknown synechobactin variants were present in the extract, and their structures have been determined by a careful HRMS/MS analysis. This study unveils the capability of Anabaena flos-aquae UTEX 1444 to produce a large array of siderophores and may be a suitable model organism for a sustainable scale-up exploitation of such bioactive molecules, for the bioremediation of contaminated ecosystems, as well as in drug discovery.

A Four-Step Platform to Optimize Growth Conditions for High-Yield Production of Siderophores in Cyanobacteria

“…Combining this information with those coming from our previous study [ 31 ], a new set of cultures was prepared, in which the concentration of Fe 3+ , supplemented as ferric ammonium citrate in FWBG11, has been varied from 0 µM to 5 μM. Taking into account that µM quantities of citrate are necessary to promote synechobactin production ( Figure 3 ), sodium citrate has been supplemented to each of the above cultures in the amount necessary to compensate for the citrate removed with the ferric ammonium salt.…”

“…We have previously reported that Anabaena flos aquae UTEX 1444 produces a cluster of chemically related siderophores (including schizokinen, rhizobactin 1021, synechobactin, and aerobactin) in a condition of iron limitation [ 31 , 32 ]. Till now, the yield of these siderophores from Anabaena flos aquae has been low [ 31 ]. Schizokinen is one of the most structurally well-defined hydroxamate siderophores and is produced by some Anabaena species in freshwater [ 33 ].…”

“…As shown in Figure 1 C, detailed analysis of the promoter region of Iuc of Anabaena flos aquae confirmed in this strain the presence of the Fur responsive element. One of the strategies commonly used to increase the yield of in vitro produced siderophore is to cultivate cyanobacteria in iron-depleted media [ 31 ]. However, it is notoriously difficult to determine for each specific cyanobacteria the optimal Fe 3+ concentration since iron is essential for many other bacterial processes.…”

“…We thus maintained fixed these parameters and modified growth media varying Fe 3+ and citrate concentration. Till now [ 31 ], the culture medium used for siderophore production was Fe 3+ deprived and contains 31 μmols/L of citric acid. We thus verified the effect of different iron and citrate concentration on the yields of siderophore produced.…”

“…We thus verified the effect of different iron and citrate concentration on the yields of siderophore produced. As first, six cultures (no added citrate, 0 mM (final (citrate) 31 μM, #1), 0.1 mM ((citrate) 131 μM, #2), 1 mM ((citrate) 1.031 mM #3), 10 mM ((citrate) 10.031 mM, #4), 100 mM ((citrate) 100.031 mM #5), STD ((citrate) 31 μM in the presence of Fe 3+ , #6) were extracted and analysed through LC-HRMS and molecular networking [ 31 ]. In the comprehensive molecular network of Anabaena #1-6 cultures’ extracts ( Figure 3 and Figure S2 ) [ 43 ], the arrangement of the cluster containing schizokinen and the known synechobactins A and C14 showed an uneven distribution of such siderophores in the different cultures, with a clear prevalence in those with the lowest concentrations of citrate (31 to 131 µmol of citrate/L) and in the absence of Fe 3+ .…”