Detection of the isiA Gene across Cyanobacterial Strains: Potential for Probing Iron Deficiency

Abstract

The use of isiA expression to monitor the iron status of cyanobacteria was investigated. Studies of laboratory cultures of the cyanobacterium Synechocystis sp. strain PCC 6803 showed that isiA expression is dependent on the organism's response to iron deficiency; isiA expression starts as soon as a decline in the rate of growth begins. isiA expression is switched on at concentrations of iron citrate of less than 0.7 M. A PCR method was developed for the specific amplification of the iron-regulated isiA gene from a variety of cyanobacteria. After we developed degenerate primers, 15 new internal isiA fragments (840 bp) were amplified, cloned, and sequenced from strains obtained from algal collections, from new isolates, and from enriched field samples. Furthermore, isiA expression could be detected by means of reverse transcription-PCR when enriched field samples were exposed to restricted iron availability. These results imply that determining the level of ironregulated isiA expression can serve to indicate iron deficiency in cyanobacterial samples of differing origins from the field.Iron as an important micronutrient of phytoplankton had been neglected for a long period before Martin and coworkers (28-30) revived its consideration in their approach to defining the basic pattern of plankton productivity in the open oceans. Subsequent research dealt with iron as the crucial nutrient in high-nitrate, low-chlorophyll regions of the open oceans and its ecological importance (1,12,16,22,43). Some evidence of iron-limited phytoplankton development also exists for coastal upwelling regions (13). Owing to the complex chemical behavior of iron, the bioavailable iron concentration cannot easily be manipulated in oxygen-rich waters by iron enrichment and iron depletion procedures. Trace-metal precipitation and adsorption to particles as well as ion-exchange processes (for a review, see reference 39) are only a few of the potential effects of these procedures that can hamper the desirable effects of fertilization and depletion experiments. Although these are methods that provide support for the characterization of the concentration of bioavailable iron in water (42), the great number and variety of variously efficient mechanisms of organismic iron sequestration (6, 14, 43) limit the interpretation of the corresponding analyses. Thus, the use of molecular markers to detect the physiological state of iron limitation in microalgae without the use of artificial manipulations of water chemistry would represent an important achievement in proving iron limitation in parts of phytoplankton.Besides producing results of general interest, the use of molecular approaches became necessary to go beyond the limits of conventional methodology in aquatic ecology. Scanlan et al. (36) introduced the phosphate-binding protein PstS as a potential diagnostic marker for investigating phosphate stress in photosynthetic picoplankton. For iron limitation, flavodoxin accumulation was used as a biochemical marker and its detection in single cells pr...

Detection of Prochlorothrix in Brackish Waters by Specific Amplification of pcb Genes

“…Specific amplification of the pcbC gene was performed with primers pcbC-fw (5Ј-GTA ATA TCC GCC TCG TAG AC-3Ј) and pcbC-rev (5Ј-CTA ACC GTC AGA CCT TAA CC-3Ј). The PCR program and all other procedures have been described previously (13). Sequencing of cloned fragments was done at least in triplicate by using a capillary sequencer (Beckman-Coulter).…”

Phytoplankton diversity and photosynthetic acclimation along a longitudinal transect through a shallow estuary in summer

“…Even when all of the DFe represents bioavailable iron, the minimum concentrations in the estuary (0.6 µmol l -1 ) can induce the iron starvation response of the moderately salinity-resistant freshwater cyanobacterium Synechocystis sp. PCC 6803 (Geiß et al 2001a). In autumn, iron fertilisations of phytoplankton from the Darss-Zingst estuary generally reduced stress to PSII (Pilkaitytë et al 2004).…”

Investigations on cyanobacterial diversity in a shallow estuary (Southern Baltic Sea) including genes relevant to salinity resistance and iron starvation acclimation

“…Under iron‐depleted conditions, isiA transcription is highly (50–100‐fold) induced in many cultured strains (Leonhardt and Straus, 1992; Ghassemian and Straus, 1996; Vinnemeier et al ., 1998; Geiß et al ., 2001a). First investigations in a model cyanobacterium indicated gene induction at the onset of iron‐depleted growth, and isiA expression also could be detected in cyanobacterial phytoplankton after enrichment in artificial growth medium without iron source (Geiß et al ., 2001b). The isiA gene has neither been amplified nor has its expression ever been detected in phytoplankton samples growing under changing physical and chemical conditions.…”