Effects Of Iron Limitation Research Articles

Transcriptomic Analysis of the Molecular Response Mechanism of Microcystis aeruginosa to Iron Limitation Stress

Iron is an essential micronutrient for cyanobacteria. It is involved in physiological activities such as photosynthesis, respiration, and the synthesis of pigments. The impact of iron limitation on planktonic algae growth occurs in surface oceans globally, as well as in freshwater ecosystems. However, the molecular and physiological effects and response mechanism of cyanobacteria under iron-limited conditions have not been reported in detail. In this study, the effects of iron limitation on the cell density, chlorophyll content, and photosynthetic activity of Microcystis aeruginosa were determined, and transcriptome sequencing was undertaken. In a severely iron-deficient environment, the cell density and chlorophyll-a content of M. aeruginosa were significantly lower than in the iron-rich group (a 55.42% and 83.51% reduction, respectively). Similarly, the photosynthetic efficiency of M. aeruginosa was also inhibited by iron deficiency, and the maximum photochemical efficiency (Fv/Fm) of the severe iron deficiency group was only 66.72% of the control group. The transcriptome results showed that to cope with the iron-deficient environment, most genes involved in iron absorption and transport in M. aeruginosa were up-regulated. In particular, the fur and perR genes that regulate the iron uptake regulatory protein (Fur) were both up-regulated. Due to the high demand for iron in the photosynthetic electron transport chain of M. aeruginosa, most photosynthesis-related genes were down-regulated, for example, petJ, which regulates iron-containing cytochrome c6. In contrast, most of the genes related to glycolysis and respiration were up-regulated. These changes in gene expression may be a survival strategy for M. aeruginosa to cope with a long-term iron-deficient environment. This study provides insights into the molecular response mechanism of M. aeruginosa under iron limitation stress.

Effects of iron limitation on carbon balance and photophysiology of the Antarctic diatom Chaetoceros cf. simplex

In the Southern Ocean (SO), iron (Fe) limitation strongly inhibits phytoplankton growth and generally decreases their primary productivity. Diatoms are a key component in the carbon (C) cycle, by taking up large amounts of anthropogenic CO2 through the biological carbon pump. In this study, we investigated the effects of Fe availability (no Fe and 4 nM FeCl3 addition) on the physiology of Chaetoceros cf. simplex, an ecologically relevant SO diatom. Our results are the first combining oxygen evolution and uptake rates with particulate organic carbon (POC) build up, pigments, photophysiological parameters and intracellular trace metal (TM) quotas in an Fe-deficient Antarctic diatom. Decreases in both oxygen evolution (through photosynthesis, P) and uptake (respiration, R) coincided with a lowered growth rate of Fe-deficient cells. In addition, cells displayed reduced electron transport rates (ETR) and chlorophyll a (Chla) content, resulting in reduced cellular POC formation. Interestingly, no differences were observed in non-photochemical quenching (NPQ) or in the ratio of gross photosynthesis to respiration (GP:R). Furthermore, TM quotas were measured, which represent an important and rarely quantified parameter in previous studies. Cellular quotas of manganese, zinc, cobalt and copper remained unchanged while Fe quotas of Fe-deficient cells were reduced by 60% compared with High Fe cells. Based on our data, Fe-deficient Chaetoceros cf. simplex cells were able to efficiently acclimate to low Fe conditions, reducing their intracellular Fe concentrations, the number of functional reaction centers of photosystem II (RCII) and photosynthetic rates, thus avoiding light absorption rather than dissipating the energy through NPQ. Our results demonstrate how Chaetoceros cf. simplex can adapt their physiology to lowered assimilatory metabolism by decreasing respiratory losses.

Diurnal regulation of photosynthetic light absorption, electron transport and carbon fixation in two contrasting oceanic environments

Abstract. Understanding the dynamics of marine phytoplankton productivity requires mechanistic insight into the non-linear coupling of light absorption, photosynthetic electron transport and carbon fixation in response to environmental variability. In the present study, we examined the variability of phytoplankton light absorption characteristics, light-dependent electron transport and 14C-uptake rates over a 48 h period in the coastal subarctic north-east (NE) Pacific. We observed an intricately coordinated response of the different components of the photosynthetic process to diurnal irradiance cycles, which acted to maximize carbon fixation, while simultaneously preventing damage by excess absorbed light energy. In particular, we found diurnal adjustments in pigment ratios, excitation energy transfer to reaction centre II (RCII), the capacity for non-photochemical quenching (NPQ), and the light efficiency (α) and maximum rates (Pmax) of RCII electron transport (ETRRCII) and 14C uptake. Comparison of these results from coastal waters to previous observations in offshore waters of the subarctic NE Pacific provides insight into the effects of iron limitation on the optimization of photosynthesis. Under iron-limited, low-biomass conditions, there was a significant reduction of iron-rich photosynthetic units per chlorophyll a, which was partly offset by higher light absorption and electron transport per photosystem II (PSII). Iron deficiency limited the capacity of phytoplankton to utilize peak midday irradiance for carbon fixation and caused an upregulation of photoprotective mechanisms, including NPQ, and the decoupling of light absorption, electron transport and carbon fixation. Such decoupling resulted in an increased electron requirement (Φe,C) and decreased quantum efficiency (ΦC) of carbon fixation at the iron-limited station. In both coastal and offshore waters, Φe,C and ΦC correlated strongly to NPQ, albeit with a significantly different slope. We discuss the implications of our results for the interpretation of bio-optical data and the parameterization of numerical productivity models, both of which are vital tools in monitoring marine photosynthesis over large temporal and spatial scales.

The effects of iron limitation and cell density on prokaryotic metabolism and gene expression: Excerpts from Fusobacterium necrophorum strain 774 (sheep isolate)

Fusobacterium necrophorum is a Gram-negative obligate anaerobe associated with several diseases in humans and animals. Despite its increasing clinical significance, there is little or no data on the relationship between its metabolism and virulence. Previous studies have shown that bacteria grown under iron-limitation express immunogenic antigens similar to those generated in vivo. Thus, this paper describes the relationship between F. necrophorum subsp. necrophorum (Fnn) metabolism and the expression of the encoded putative virulence factors under iron-restricted conditions. At the midlog phase, iron limitation reduced Fnn growth but the cell density was dependent on the size of the inoculum. Preferential utilization of glucose-1-phosphate, d-mannitol and l-phenylalanine; production of 2-hydroxycaproic acid and termination of dimethyl sulphide production were major Fnn response-factors to iron limitation. Ultimately, iron restriction resulted in an increased ability of Fnn to metabolize diverse carbon sources and in the expression of stress-specific virulence factors. Iron starvation in low Fnn cell density was associated with the up-regulation of haemagglutinin (HA) and leukotoxin (lktA) genes (2.49 and 3.72 fold change respectively). However, Fnn encoded Haemolysin (Hly), yebN homologue (febN) and tonB homologue, were down-regulated (0.15, 0.79 and 0.33, fold changes respectively). Interestingly, cell density appeared to play a regulatory role in the final bacteria cell biomass, induction of a metabolic gene expression and the expression pattern virulence factors in Fnn suggesting the role of a cell density-associated regulatory factor. This report suggest that future studies on differential expression of bacterial genes under altered environmental condition(s) should consider testing the effect of cell concentrations as this is often neglected in such studies. In conclusion, iron restriction induces preferential utilization of carbon sources and altered metabolism in Fnn with associated changes in the expression pattern of the virulence factors.

Effects of iron limitation on growth and carbon metabolism in oceanic and coastal heterotrophic bacteria

We investigated the metabolic response to iron (Fe) limitation of two bacterial strains of Alteromonas macleodii, isolated from a coastal and an oceanic marine environment. Bacteria were grown under Fe‐limited and Fe‐replete conditions, and comparative analyses of cellular properties and total proteomes were conducted. Respiration was reduced by a factor of two in both strains, but the growth rate of the oceanic strain was less affected by Fe limitation (reduced by 1.2‐fold) than the coastal strain (reduced by 2‐fold). Fe limitation led to significant changes in the expression of several key enzymes associated with carbon catabolism, specifically those involved in the citric acid cycle, glycolysis, and oxidative phosphorylation. The strain‐specific overall responses to Fe limitation were in part reflected in different metabolic strategies of the carbon metabolism and energy acquisition. Our study provides novel insights on how Fe limitation can affect bacterial heterotrophic metabolism, and how this could influence the coupling of the Fe and carbon cycles in the ocean.

THE EFFECT OF IRON LIMITATION ON THE PHOTOPHYSIOLOGY OF PHAEOCYSTIS ANTARCTICA (PRYMNESIOPHYCEAE) AND FRAGILARIOPSIS CYLINDRUS (BACILLARIOPHYCEAE) UNDER DYNAMIC IRRADIANCE(1).

The effects of iron limitation on photoacclimation to dynamic irradiance were studied in Phaeocystis antarctica G. Karst. and Fragilariopsis cylindrus (Grunow) W. Krieg. in terms of growth rate, photosynthetic parameters, pigment composition, and fluorescence characteristics. Under dynamic light conditions mimicking vertical mixing below the euphotic zone, P. antarctica displayed higher growth rates than F. cylindrus both under iron (Fe)-replete and Fe-limiting conditions. Both species showed xanthophyll de-epoxidation that was accompanied by low levels of nonphotochemical quenching (NPQ) during the irradiance maximum of the light cycle. The potential for NPQ at light levels corresponding to full sunlight was substantial in both species and increased under Fe limitation in F. cylindrus. Although the decline in Fv /Fm under Fe limitation was similar in both species, the accompanying decrease in the maximum rate of photosynthesis and growth rate was much stronger in F. cylindrus. Analysis of the electron transport rates through PSII and on to carbon (C) fixation revealed a large potential for photoprotective cyclic electron transport (CET) in F. cylindrus, particularly under Fe limitation. Probably, CET aided the photoprotection in F. cylindrus, but it also reduced photosynthetic efficiency at higher light intensities. P. antarctica, on the other hand, was able to efficiently use electrons flowing through PSII for C fixation at all light levels, particularly under Fe limitation. Thus, Fe limitation enhanced the photophysiological differences between P. antarctica and diatoms, supporting field observations where P. antarctica is found to dominate deeply mixed water columns, whereas diatoms dominate shallower mixed layers.

Effects of iron limitation on intracellular cadmium of cultured phytoplankton: Implications for surface dissolved cadmium to phosphate ratios

This study compares intracellular Cd content (Cd:C) of cultured marine phytoplankton grown under various Fe levels, with estimated particulate Cd:P ratios derived from regression slopes of Cd versus PO 4 3− relationships from a global dataset. A 66-fold difference in Cd:C ratios was observed among the seven species grown under identical Fe concentrations, with oceanic diatoms having the highest Cd quotas and prymesiophytes the lowest. Interestingly, all species significantly increased their Cd:C ratios under Fe-limitation (on average 2-fold). The global data set also showed that the mean estimated Cd:P ratio of surface water particulates in HNLC (high nutrient low chlorophyll) regions were approximately 2-fold higher than non-HNLC regions. A sequence of events are proposed to explain high Cd:P ratios in HNLC waters. First, the seasonal relief from Fe-limitation in HNLC regions leads to blooms of large chain forming diatoms with high intrinsic Cd:P ratios. These large blooms may, in theory, deplete surface water CO 2 and Zn concentrations, which ultimately, would result in increased Cd uptake. Eventually these blooms will run out of Fe, which has been shown to further increase intercellular Cd via growth biodilution and increased Cd uptake through non-specific Fe(II) transporters. Ultimately, Fe-limited diatoms with enhanced Cd quotas will sink out of surface waters leading to pronounced regional differences in Cd:P ratios between HNLC and non-HNLC waters in the global ocean.

Iron effects on colonization behavior, motility, and enzymatic activity of marine bacteria

Iron availability in the ocean has been shown to affect the growth and production of phytoplankton and free-living bacteria. A large fraction of marine bacteria are specialized in colonizing and living on particles and aggregates, but the effects of iron limitation on these bacteria are not fully known. We conducted laboratory experiments to study the effects of iron availability on particle colonization behavior, motility, and enzymatic activities of 4 strains of marine bacteria. Iron depletion reduced the bacterial particle colonization rate by 1.7%-43.1%, which could be attributed to reduced swimming speeds in 2 of the 4 strains. Protease activity was not affected by iron availability. However, attached bacteria did show higher protease activities than their free counterparts. Our results suggest that iron limitation in the ocean could in some cases reduce bacteria-particle interactions by reducing bacterial motility and colonization rate.

Photosynthetic responses in Phaeocystis antarctica towards varying light and iron conditions

The effects of iron limitation on photoacclimation to a dynamic light regime were studied in Phaeocystis antarctica. Batch cultures were grown under a sinusoidal light regime, mimicking vertical mixing, under both iron-sufficient and -limiting conditions. Iron-replete cells responded to changes in light intensity by rapid xanthophyll cycling. Maximum irradiance coincided with maximum ratios of diatoxanthin/diadinoxanthin (dt/dd). The maximum quantum yield of photosynthesis (F v /F m ) was negatively related to both irradiance and dt/dd. Full recovery of F v /F m by the end of the light period suggested successful photoacclimation. Iron-limited cells displayed characteristics of high light acclimation. The ratio of xanthophyll pigments to chlorophyll a was three times higher compared to iron-replete cells. Down-regulation of photosynthetic activity was moderated. It is argued that under iron limitation cells maintain a permanent state of high energy quenching to avoid photoinhibition during exposure to high irradiance. Iron-limited cells could maintain a high growth potential due to an increased absorption capacity as recorded by in vivo absorption, which balanced a decrease in F v /F m . The increase in the chlorophyll a-specific absorption cross section was related to an increase in carotenoid pigments and a reduction in the package effect. These experiments show that P. antarctica can acclimate successfully to conditions as they prevail in the Antarctic ocean, which may explain the success of this species.

Effects of iron limitation on the respiratory chain and the membrane cytochrome pattern of the Euryarchaeon Halobacterium salinarum.

The effects of iron limitation on the electron transport chain of the extremely halophilic Euryarchaeon Halobacterium salinarum were analyzed. When iron was growth-limiting, the respiratory rates as well as the inhibition pattern of the membranes were significantly different from membranes of iron replete cells. Changes in the availability of iron cause the formation of different respiratory pathways including different entry sites for electrons, different terminal oxidases of the respiratory chain, and drastic changes of the cytochrome composition and of the relative amounts of cytochromes. Under iron-limiting conditions, mainly low-potential cytochromes were measured. EPR spectroscopic studies revealed that the amount of proteins containing iron-sulfur clusters is reduced in membranes under iron-limiting growth conditions. Taken together, our results strongly suggest for the first time an important role of iron supply for the bioenergetics of an Archaeon.

Photoacclimation by the Antarctic flagellate Pyramimonas sp. (Prasinophyceae) in response to iron limitation

In this study we tested the hypothesis that iron limitation suppresses photoacclimation in cultures of the Antarctic flagellate Pyramimonas sp. The cultures were exposed to two different irradiances under iron-rich and iron-poor conditions. Light-harvesting capacity was determined by assessing the pigment composition and measuring in vivo absorption spectra. Light utilization efficiency (α) was determined from photosynthesis versus irradiance curves. The quantum yield of photosynthesis (φm) was calculated using α and the absorption spectra. Iron limitation led to commonly observed changes in cells of Pyramimonas, that is, a decrease in cellular pigment content and a reduction in cellular carbon and nitrogen quota. A reduction in αcell followed a decrease in φm and light-harvesting capacity. Interpretation of the effects of iron limitation was different when considered on a carbon basis. Because iron limitation resulted in a decrease in cellular carbon content, the carbon-specific absorption coefficient was not affected. Consequently, the observed decrease in αC was mainly due to the decrease in φm, showing that iron limitation did not control light utilization via pigment synthesis but exerted control on energy transfer. This is supported by the findings that at high irradiance a shift in pigment ratios within the total pool of violaxanthin, antheraxanthin and zeaxanthin towards zeaxanthin, which is indicative of photoacclimation to high irradiance, was observed for iron-replete cells as well as for iron-depleted cells. In contrast to what is generally hypothesized, the effects of iron limitation were not enhanced at low irradiance. Low irradiance led to an increase in the cellular light-harvesting pigment content. This increase was less pronounced in iron-depleted cells than in iron-replete cells. However, looking at the light-harvesting capacity of the cells on a carbon basis, it was found that iron-depleted cells responded similarly to iron-replete cells. We therefore conclude that the light-harvesting capacity was governed by light conditions and not by iron limitation. In addition to the increase in absorption capacity at low irradiance, an increase in light utilization efficiency was measured, again under both iron-rich and iron-poor conditions. Notably, the relative increase in αC was strongest in iron-depleted cells. Photoacclimation was clearly demonstrated by normalizing α to chl a. For iron-replete cells, αchl was highest at high irradiance. In contrast, for iron-depleted cells a chl was highest at low irradiance. We argue that iron-depleted cells can photoacclimate to low irradiance by a reduction in the ‘package effect’ and reducing growth rates.

Effects of iron limitation and aquatic humic substances on the growth of <i>Microcystis aeruginosa</i>

Effects of iron limitation and aquatic humic substances on the growth of <i>Microcystis aeruginosa</i>

Alterations in cell pigmentation, protein expression, and photosynthetic capacity of the cyanobacteriumOscillatoria tenuisgrown under low iron conditions

To better describe the iron-limited nutrient status of aquatic photosynthetic microorganisms, we examined the effects of iron limitation on pigment content, maximum rates of photosynthetic oxygen evolution, and respiratory oxygen consumption in the filamentous cyanobacterium Oscillatoria tenuis Ag. Within the range of iron (4.2 x 10(-5)-5.1 x 10(-9) M FeCl3), growth rates were not limited by photosynthetic capacity but rather by another, as of yet undetermined, iron-requiring cellular function. We have also investigated membrane proteins that are induced when the cells are grown in low iron medium. Using membrane fractionation techniques we were able to recognize specific proteins localized in the outer membrane and periplasmic space of O. tenuis. The recovery of growth rates at low iron levels occurred in parallel with the induction of these proteins and the production of extracellular siderophores. The additional iron acquired by this high affinity transport system did not reestablish photosynthesis in O. tenuis to the iron-satiated level but did reestablish growth to iron-replete levels. Oscillatoria tenuis appears to invoke an alternate physiology to compensate for iron deficiency.

Effects of Iron Limitation on Photosystem II Composition and Light Utilization in Dunaliella tertiolecta.

The effects of iron limitation on photosystem II (PSII) composition and photochemical energy conversion efficiency were studied in the unicellular chlorophyte alga Dunaliella tertiolecta. The quantum yield of photochemistry in PSII, inferred from changes in variable fluorescence normalized to the maximum fluorescence yield, was markedly lower in iron-limited cells and increased 3-fold within 20 h following the addition of iron. The decrease in the quantum yield of photochemistry was correlated with increased fluorescence emission from the antenna. In iron-limited cells, flash intensity saturation profiles of variable fluorescence closely followed a cumulative one-hit Poisson model, suggesting that PSII reaction centers are energetically isolated, whereas in iron-replete cells, the slope of the profile was steeper and the calculated probability of energy transfer between reaction centers increased to >0.6. Immunoassays revealed that in iron-limited cells the reaction center proteins, D1, CP43, and CP47, were markedly reduced relative to the peripheral light-harvesting Chl-protein complex of PSII, whereas the [alpha] subunit of cytochrome b559 was about 10-fold higher. Spectroscopic analysis established that the cytochrome b559 peptide did not contain an associated functional heme. We conclude that the photochemical conversion of absorbed excitation energy in iron-limited cells is limited by the number of photochemical traps per unit antenna.

Effects of iron limitation on production of a siderophore, outer membrane proteins, and hemolysin and on hydrophobicity, cell adherence, and lethality for mice of Vibrio parahaemolyticus.

Vibrio parahaemolyticus is one of the most important enteropathogens in Taiwan, Japan, and other coastal regions. The pathogenesis of V. parahaemolyticus disease is not clearly understood. The expression of some factors by V. parahaemolyticus in iron-rich and iron-limited media was analyzed. In the clinical hemolytic strains, the production of a siderophore, two outer membrane proteins (77 and 80 kDa), and thermostable direct hemolysin was significantly enhanced in iron-limited culture, and hemolytic activities, cell hydrophobicity, HEp-2 cell adherence, and lethality for mice were also enhanced. The environmental nonhemolytic strain CCRC12958 that was cultured in iron-limited medium exhibited lethal activity for mice, and other factors except hemolysis were also enhanced like the responses of clinical strains were. These results suggested that a virulent factor(s) of V. parahaemolyticus may be induced or enhanced under iron-limited conditions. The iron-regulated factors reported in this paper may be important in the pathogenesis of V. parahaemolyticus disease.

INFLUENCE OF IRON LIMITATION AND NITROGEN SOURCE ON GROWTH AND SIDEROPHORE PRODUCTION BY CYANOBACTERIA

To characterize the mobilization and uptake of iron by cyanobacteria, 14 species were screened for ability to scavenge iron in a competitive system. The cyanobacteria exhibited a range of growth responses to iron limitation which could be separated into three groups, and a representative species from each group was chosen for further study. Effects of iron-limitation on growth and siderophore production of Anacystis nidulans R2, Anabaena variabilis ATCC 29413, and Plectonema boryanum UTEX 581 were determined. Both A. nidulans R2 and A. variabilis showed a reduced rate of growth with decreased available iron concentration (PFe 17–19). Growth rates increased with further reduction in the level of available iron (pFe 20 to pFe 21). The increase in growth rate occurred at the same available iron concentration as the initiation of extracellular siderophore production. In contrast, the growth of P. boryanum decreased with decreasing available iron levels. No siderophore production was detected from P. boryanum cultures. The growth kinetics of siderophore-producing species differ from traditional nutrient-limited growth kinetics and clearly reflect the presence of a high affinity, siderophore-mediated iron transport system in A. nidulans R2 and A. variabilis. Iron-limited growth kinetics more similar to traditional nutrient-limited growth kinetics were found in P. boryanum. The available nitrogen source influenced amount of siderophore produced and concentration of available iron which induced siderophore production. Siderophores were produced at high iron concentrations (pFe 18) when A. variablilis cultures were grown in the absence of combined nitrogen source. When nitrate was supplied to the culture, iron concentrations had to be reduced to pFe 20 before siderophores were produced. Cells grown on nitrogen also produced greater than two times the amount of siderophore compared with nitrate grown cells. This may be indicative of an increased demand for iron by nitrogen fixing A. variabilis Cultures.

Effect of Iron Limitation on the Ultrastructure of Agmenellum Quadruplicatum

Agmenellum quadruplicatum is a unicellular, non-nitrogen-fixing, marine cyanobacterium (blue-green alga). The ultrastructure of this organism, when grown in the laboratory with all necessary nutrients, has been characterized thoroughly. In contrast, little is known of its ultrastructure in the specific nutrient-limiting conditions typical of its natural habitat. Iron is one of the nutrients likely to limit this organism in such natural environments. It is also of great importance metabolically, being required for both photosynthesis and assimilation of nitrate. The purpose of this study was to assess the effects (if any) of iron limitation on the ultrastructure of A. quadruplicatum. It was part of a broader endeavor to elucidate the ultrastructure of cyanobacteria in natural systemsActively growing cells were placed in a growth medium containing 1% of its usual iron. The cultures were then sampled periodically for 10 days and prepared for thin sectioning TEM to assess the effects of iron limitation.