Iron nutrition and physiological responses to iron stress in Nitrosomonas europaea

Nitrosomonas europaea, as an ammonia-oxidizing bacterium, has a high Fe requirement and has 90 genes dedicated to Fe acquisition. Under Fe-limiting conditions (0.2 μM Fe), N. europaea was able to assimilate up to 70% of the available Fe in the medium even though it is unable to produce siderophores. Addition of exogenous siderophores to Fe-limited medium increased growth (final cell mass). Fe-limited cells had lower heme and cellular Fe contents, reduced membrane layers, and lower NH₃- and NH₂OH-dependent O₂ consumption activities than Fe-replete cells. Fe acquisition-related proteins, such as a number of TonB-dependent Fe-siderophore receptors for ferrichrome and enterobactin and diffusion protein OmpC, were expressed to higher levels under Fe limitation, providing biochemical evidence for adaptation of N. europaea to Fe-limited conditions.     

Nitrogen loss caused by denitrifying Nitrosomonas cells using ammonium or hydrogen as electron donors and nitrite as electron acceptor

Cells of the obligately lithotrophic species Nitrosomonas europaea and Nitrosomonas eutropha were able to nitrify and denitrify at the same time when grown under oxygen limitation. In addition to oxygen, nitrite was used as an electron acceptor. The simultaneous nitrification and denitrification resulted in significant formation of the gaseous N-compounds nitrous oxide and dinitrogen, causing significant nitrogen loss. In mixed cultures of N. europaea and various chemoorganotrophic bacteria, the nitrogen loss was strongly influenced by the partners growing under oxygen limitation. Under anoxic conditions, pure cultures of N. eutropha were able to denitrify with molecular hydrogen as electron donor and nitrite as the only electron acceptor in a sulfide-reduced complex medium. The increase of cell numbers was directly coupled to nitrite reduction. Nitrous oxide and dinitrogen were the only detectable end products. In pure cultures of N. eutropha and mixed cultures of N. eutropha and Enterobacter aerogenes, ammonium and nitrite disappeared slowly at a molar ratio of about one when oxygen was absent. However, under these conditions cell growth was not measurable.     

Chemoorganoheterotrophic Growth of <Emphasis Type="Italic">Nitrosomonas europaea</Emphasis> and <Emphasis Type="Italic">Nitrosomonas eutropha</Emphasis>

The ammonia oxidizers Nitrosomonas europaea and Nitrosomonas eutropha are able to grow chemoorganotrophically under anoxic conditions with pyruvate, lactate, acetate, serine, succinate, α-ketoglutarate, or fructose as substrate and nitrite as terminal electron acceptor. The growth yield of both bacteria is about 3.5 mg protein (mmol pyruvate)⁻¹ and the maximum growth rates of N. europaea and N. eutropha are 0.094 d⁻¹ and 0.175 d⁻¹, respectively. In the presence of pyruvate and CO₂ about 80% of the incorporated carbon derives from pyruvate and about 20% from CO₂. Pyruvate is used as energy and only carbon source in the absence of CO₂ (chemoorganoheterotrophic growth). CO₂ stimulates the chemoorganotrophic growth of both ammonia oxidizers and the expression of ribulose bisphosphate carboxylase/oxygenase is down-regulated at increasing CO₂ concentration. Ammonium, although required as nitrogen source, is inhibitory for the chemoorganotrophic metabolism of N. europaea and N. eutropha. In the presence of ammonium pyruvate consumption and the expression of the genes aceE, ppc, gltA, odhA, and ppsA (energy conservation) as well as nirK, norB, and nsc (denitrification) are reduced.     

FERRIC CHELATE REDUCTASE ACTIVITY AS AFFECTED BY THE IRON‐LIMITED GROWTH RATE IN FOUR SPECIES OF UNICELLULAR GREEN ALGAE (CHLOROPHYTA)1

Four species of green algae (Chlorella kessleri Fott et Nováková, Chlorococcum macrostigmatum Starr, Haematococcus lacustris[Girod‐Chantrans] Rostaf., Stichococcus bacillaris Näg.) were grown in iron‐limited chemostats and under phosphate limitation and iron (nutrient) sufficiency. For all four species, steady‐state culture density declined with decreasing degree of iron limitation (increasing iron‐limited growth rate), whereas chl per cell or biovolume increased. Plasma membrane ferric chelate reductase activity was enhanced by iron limitation in all species and suppressed by phosphate limitation and iron sufficiency. These results confirm previous work that C. kessleri uses a reductive mechanism of iron acquisition and also suggest that the other three species use the same mechanism. Although imposition of iron limitation led to enhanced activities of ferric chelate reductase in all species, the relationship between ferric chelate reductase activity and degree of iron limitation varied. Ferric chelate reductase activity in C. macrostigmatum and S. bacillaris was an inverse function of the degree of iron limitation, with the most rapidly growing iron‐limited cells exhibiting the highest ferric chelate reductase activity. In contrast, ferric chelate reductase activity was only weakly affected by the degree of iron limitation in C. kessleri and H. lacustris. Calculation of ferric reductase activity per unit chl allowed a clear differentiation between iron‐limited and iron‐sufficient cells. The possible extension of the ferric chelate reductase assay to investigate the absence or presence of iron limitation in natural waters may be feasible, but it is unlikely that the assay could be used to estimate the degree of iron limitation.