A single-cell view on the ecophysiology of anaerobic phototrophic bacteria

Abstract

Quantitative information on the ecophysiology of individual microorganisms is generally limited because it is difficult to assign specific metabolic activities to identified single cells. Here, we develop and apply a method, Halogen In Situ HybridizationSecondary Ion Mass Spectroscopy (HISH-SIMS), and show that it allows simultaneous phylogenetic identification and quantitation of metabolic activities of single microbial cells in the environment. Using HISH-SIMS, individual cells of the anaerobic, phototropic bacteria Chromatium okenii, Lamprocystis purpurea, and Chlorobium clathratiforme inhabiting the oligotrophic, meromictic Lake Cadagno were analyzed with respect to H 13 CO3 ؊ and 15 NH4 ؉ assimilation. Metabolic rates were found to vary greatly between individual cells of the same species, showing that microbial populations in the environment are heterogeneous, being comprised of physiologically distinct individuals. Furthermore, C. okenii, the least abundant species representing Ϸ0.3% of the total cell number, contributed more than 40% of the total uptake of ammonium and 70% of the total uptake of carbon in the system, thereby emphasizing that numerically inconspicuous microbes can play a significant role in the nitrogen and carbon cycles in the environment. By introducing this quantification method for the ecophysiological roles of individual cells, our study opens a variety of possibilities of research in environmental microbiology, especially by increasing the ability to examine the ecophysiological roles of individual cells, including those of less abundant and less active microbes, and by the capacity to track not only nitrogen and carbon but also phosphorus, sulfur, and other biological element flows within microbial communities.anaerobic phototrophs ͉ nanoSIMS

Carbon, nitrogen and O2 fluxes associated with the cyanobacterium Nodularia spumigena in the Baltic Sea

“…For each individual cell, we recorded simultaneously secondary ion images of 12 C, 13 C, 12 C 14 N and 12 C 15 N using four electron multipliers. The measurements and the image and data processing were performed as described previously (Musat et al, 2008;Ploug et al, 2010).…”

A single-cell view of ammonium assimilation in coral–dinoflagellate symbiosis

“…Metabolic studies on coral-dinoflagellate symbioses generally involve the unnatural culture of both symbiotic partners separately or the separation of algal and host tissue fractions, an experimental procedure, which is often incomplete and hampered by crosscontamination (Yellowlees et al, 2008). In this context, recent advances in a high-resolution isotopic imaging technology, the sub-micrometer scale ion microprobe technique referred to as NanoSIMS provides direct imaging and quantification of the metabolic exchanges within an intact symbiosis following stable isotope labeling (Lechene et al, 2006(Lechene et al, , 2007Kuypers, 2007;Musat et al, 2008;Dattagupta et al, 2009;Foster et al, 2011). Nano-SIMS has recently been successfully applied to reefbuilding corals in order to image trace-elemental distributions in different ultra-structural components of both fossil and living corals (Meibom et al, 2004(Meibom et al, , 2008Clode et al, 2007;Reynaud et al, 2007;Stolarski et al, 2007;Houlbreque et al, 2009;Brahmi et al, 2010).…”

Detection and Capturing of 14C Radioactively-Labeled Small Subunit rRNA from Mixed Microbial Communities of a Microbial Mat Using Magnetic Beads

“…For example, stable isotope probing (SIP) technique involved the separation of heavy DNA (i.e. 13 C or 15 N labeled) from natural samples by density-gradient centrifugation after incubation with isotope-labeled substrate [7,8], followed by sequencing and identification of key microorganisms involved in the assimilation of this substrate. The technique provided more sensitive results when performed on RNA or polar-lipid-derived fatty acids [9][10][11].…”

“…The isotope measurements in single microbial cells became possible with the help of isotopic measurement using high resolution ion microprobe, a technique known as NanoSIMS [12]. The coupling of this technique with halogen in situ hybridization (HISH-SIMS) allowed the simultaneous phylogenetic identification and quantification of metabolic activities in single bacterial cells in the environment [13]. Unlike stable isotopes, the use of radioactive isotopes was very limited although they are more sensitive, most likely because of the difficulty to handle them and the strict requirements of special precautions as well as the high cost of radioactively labeled compounds.…”