Abstract
High-resolution imaging with secondary ion mass spectrometry (nanoSIMS) has become a standard method in systems biology and environmental biogeochemistry and is broadly used to decipher ecophysiological traits of environmental microorganisms, metabolic processes in plant and animal tissues, and cross-kingdom symbioses. When combined with stable isotope-labeling—an approach we refer to as nanoSIP—nanoSIMS imaging offers a distinctive means to quantify net assimilation rates and stoichiometry of individual cell-sized particles in both low- and high-complexity environments. While the majority of nanoSIP studies in environmental and microbial biology have focused on nitrogen and carbon metabolism (using 15N and 13C tracers), multiple advances have pushed the capabilities of this approach in the past decade. The development of a high-brightness oxygen ion source has enabled high-resolution metal analyses that are easier to perform, allowing quantification of metal distribution in cells and environmental particles. New preparation methods, tools for automated data extraction from large data sets, and analytical approaches that push the limits of sensitivity and spatial resolution have allowed for more robust characterization of populations ranging from marine archaea to fungi and viruses. NanoSIMS studies continue to be enhanced by correlation with orthogonal imaging and ‘omics approaches; when linked to molecular visualization methods, such as in situ hybridization and antibody labeling, these techniques enable in situ function to be linked to microbial identity and gene expression. Here we present an updated description of the primary materials, methods, and calculations used for nanoSIP, with an emphasis on recent advances in nanoSIMS applications, key methodological steps, and potential pitfalls.
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References
Mayali X, Weber PK, Brodie EL, Mabery S, Hoeprich PD, Pett-Ridge J (2012) High-throughput isotopic analysis of RNA microarrays to quantify microbial resource use. ISME J 6(6):1210–1221
Adamczyk J, Hesselsoe M, Iversen N, Horn M, Lehner A, Nielsen PH, Schloter M, Roslev P, Wagner M (2003) The isotope array, a new tool that employs substrate-mediated labeling of rRNA for determination of microbial community structure and function. Appl Environ Microbiol 69(11):6875–6887
Ouverney CC, Fuhrman JA (1999) Combined microautoradiography-16S rRNA probe technique for determination of radioisotope uptake by specific microbial cell types in situ. Appl Environ Microbiol 65(4):1746–1752
Jehmlich N, Schmidt F, Taubert M, Seifert J, Bastida F, von Bergen M, Richnow H-H, Vogt C (2010) Protein-based stable isotope probing. Nat Protoc 5(12):1957–1966
Murrell JC, Whiteley AS (2011) Stable isotope probing and related technologies. ASM Press, Washington, DC, p 345
Koch BJ, McHugh TA, Hayer M, Schwartz E, Blazewicz SJ, Dijkstra P, van Gestel N, Marks JC, Mau RL, Morrissey EM, Pett-Ridge J, Hungate BA (2018) Estimating taxon-specific population dynamics in diverse microbial communities. Ecosphere 9(1):e02090–e02015
Hillion F, Daigne B, Girard F, Slodzian G (1993) A new high performance instrument: the CAMECA NanoSIMS 50. In: Benninghoven A et al (eds) Secondary ion mass spectrometry: SIMS IX, vol 254-257. John Wiley & Sons, Chichester
Ghosal S, Leighton TJ, Wheeler KE, Hutcheon ID, Weber PK (2010) Spatially resolved characterization of water and ion incorporation in Bacillus spores. Appl Environ Microbiol 76(10):3275–3282
Orphan VJ, House CH, Hinrichs K-U, McKeegan KD, DeLong EF (2001) Methane-consuming archaea revealed by directly coupled isotopic and phylogenetic analysis. Science 293(5529):484–487
Smart K, Kilburn M, Salter C, Smith J, Grovenor C (2007) NanoSIMS and EPMA analysis of nickel localisation in leaves of the hyperaccumulator plant Alyssum lesbiacum. Int J Mass Spectrom 260(2–3):107–114
Stadermann FJ, Walker RM, Zinner E (1999) Nanosims: the next generation ion probe for the microanalysis of extraterrestrial material. Meteorit Planet Sci 34:A111–A112
Guerquin-Kern J-L, Hillion F, Madelmont J-C, Labarre P, Papon J, Croisy A (2004) Ultra-structural cell distribution of the melanoma marker iodobenzamide: improved potentiality of SIMS imaging in life sciences. BioMed Eng. http://www.biomedical-engineering-online.com/content/3/1/10
Kraft ML, Fishel SF, Marxer CG, Weber PK, Hutcheon ID, Boxer SG (2006) Quantitative analysis of supported membrane composition using the NanoSIMS. Appl Surf Sci 252(19):6950–6956
Moreau JW, Weber PK, Martin MC, Gilbert B, Hutcheon ID, Banfield JF (2007) Extracellular proteins limit the dispersal of biogenic nanoparticles. Science 316:1600–1603
Peteranderl R, Lechene C (2004) Measure of carbon and nitrogen stable isotope ratios in cultured cells. J Am Soc Mass Spectrom 15(4):478–485
Wainwright M, Weber PK, Smith JB, Hutcheon ID, Klyce B, Wickramasinghe NC, Narlikar JV, Rajaratnam P (2004) Studies on bacteria-like particles sampled from the stratosphere. Aeorobiologia 20:237–240
Galli Marxner C, Kraft ML, Weber PK, Hutcheon I, Boxer SG (2005) Supported membrane composition analysis by secondary ion mass spectrometry with high lateral resolution. Biophys J 88:2965–2975
Dekas AE, Poretsky RS, Orphan VJ (2009) Deep-sea archaea fix and share nitrogen in methane-consuming microbial consortia. Science 326(5951):422–426
Halm H, Musat N, Lam P, Langlois R, Musat F, Peduzzi S, Lavik G, Schubert CJ, Sinha B, LaRoche J, Kuypers MMM (2009) Co-occurrence of denitrification and nitrogen fixation in a meromictic lake, Lake Cadagno (Switzerland). Environ Microbiol 11(8):2190–2190
Musat N, Halm H, Winterholler B, Hoppe P, Peduzzi S, Hillion F, Horreard F, Amann R, Jørgensen BB, Kuypers MMM (2008) A single-cell view on the ecophysiology of anaerobic phototrophic bacteria. Proc Natl Acad Sci 105(46):17861–17866
Quintana C, Wu TD, Delatour B, Dhenain M, Guerquin-Kern JL, Croisy A (2007) Morphological and chemical studies of pathological human and mice brain at the subcellular level: correlation between light, electron, and NanoSIMS microscopies. Microsc Res Tech 70(4):281–295
Behrens S, Losekann T, Pett-Ridge J, Weber PK, Ng W, Stevenson BS, Hutcheon ID, Relman DA, Spormann AM (2008) Linking microbial phylogeny to metabolic activity at the single-cell level by using enhanced element labeling-catalyzed reporter deposition fluorescence in situ hybridization (EL-FISH) and NanoSIMS. Appl Environ Microbiol 74(10):3143
Finzi-Hart J, Pett-Ridge J, Weber P, Popa R, Fallon SJ, Gunderson T, Hutcheon I, Nealson K, Capone DG (2008) Fixation and fate of carbon and nitrogen in Trichodesmium IMS101 using nanometer resolution secondary ion mass spectrometry (NanoSIMS). PNAS 106:6345–6350
Lechene C, Hillion F, McMahon G, Benson D, Kleinfeld A, Kampf JP, Distel D, Luyten Y, Bonventre J, Hentschel D, Park K, Ito S, Schwartz M, Benichou G, Slodzian G (2006) High-resolution quantitative imaging of mammalian and bacterial cells using stable isotope mass spectrometry. J Biol 5(6):20
Popa R, Weber PK, Pett-Ridge J, Finzi JA, Fallon SJ, Hutcheon ID, Nealson KH, Capone DG (2007) Carbon and nitrogen fixation and metabolite exchange in and between individual cells of Anabaena oscillarioides. ISME J 1(4):354–360
Ghosal S, Fallon SJ, Leighton T, Wheeler KE, Hutcheon ID, Weber PK (2008) Imaging and 3D elemental characterization of intact bacterial spores with high-resolution secondary ion mass spectrometry (NanoSIMS) depth profile analysis. Anal Chem 80(15):5986–5992
Lechene CP, Luyten Y, McMahon G, Distel DL (2007) Quantitative imaging of nitrogen fixation by individual bacteria within animal cells. Science 317:1563–1566
Herrmann A, Ritz K, Nunan N, Clode P, Pett-Ridge J, Kilburn M, Murphy D, O’Donnell A, Stockdale E (2007) Nano-scale secondary ion mass spectrometry – a new analytical tool in biogeochemistry and soil ecology: a review article. Soil Biol Biochem 39:1835–1850
Mueller CW, Weber PK, Kilburn MR, Hoeschen C, Kleber M, Pett-Ridge J (2013) Advances in the analysis of biogeochemical interfaces: NanoSIMS to investigate soil microenvironments. In: Sparks D (ed) Advances in agronomy. Elsevier, Amsterdam
Renslow RS, Lindemann SR, Cole JK, Zhu Z, Anderton CR (2016) Quantifying element incorporation in multispecies biofilms using nanoscale secondary ion mass spectrometry image analysis. Biointerphases 11(2):02A322
Mayali X (2020) NanoSIMS: microscale quantification of biogeochemical activity with large-scale impacts. Annu Rev Mar Sci 12:449–467
Gao D, Huang X, Tao Y (2016) A critical review of NanoSIMS in analysis of microbial metabolic activities at single-cell level. Crit Rev Biotechnol 36(5):884–890
Zhao F-J, Moore KL, Lombi E, Zhu Y-G (2014) Imaging element distribution and speciation in plant cells. Trends Plant Sci 19(3):183–192
Musat N, Musat F, Weber PK, Pett-Ridge J (2016) Tracking microbial interactions with NanoSIMS. Curr Opin Biotechnol 41:114–121
Agüi-Gonzalez P, Jähne S, Phan NT (2019) SIMS imaging in neurobiology and cell biology. J Anal At Spectrom 34(7):1355–1368
Boxer SG, Kraft ML, Weber PK (2009) Advances in imaging secondary ion mass spectrometry for biological samples. Annu Rev Biophys 38:53–74
Nuñez J, Renslow R, Cliff JB III, Anderton CR (2018) NanoSIMS for biological applications: current practices and analyses. Biointerphases 13(3):03B301
Gorman BL, Kraft ML (2019) High-resolution secondary ion mass spectrometry analysis of cell membranes. ACS Publications, Washington, DC
CAMECA. NanoSIMS 50L: scientific publications. https://www.cameca.com/products/sims/nanosims
Pett-Ridge J, Weber PK (2012) NanoSIP: NanoSIMS applications for microbial biology. In: Navid A (ed) Microbial systems biology: methods and protocols. Humana, New York, NY
Dekas AE, Parada AE, Mayali X, Fuhrman JA, Wollard J, Weber PK, Pett-Ridge J (2019) Characterizing chemoautotrophy and heterotrophy in marine archaea and bacteria with single-cell multi-isotope nanoSIP. Front Microbiol 10:2682
Chadwick GL, Otero FJ, Gralnick JA, Bond DR, Orphan VJ (2019) NanoSIMS imaging reveals metabolic stratification within current-producing biofilms. Proc Natl Acad Sci 116(41):20716–20724
Volland J-M, Schintlmeister A, Zambalos H, Reipert S, Mozetič P, Espada-Hinojosa S, Turk V, Wagner M, Bright M (2018) NanoSIMS and tissue autoradiography reveal symbiont carbon fixation and organic carbon transfer to giant ciliate host. ISME J 12(3):714–727
Calabrese F, Voloshynovska I, Musat F, Thullner M, Schlömann M, Richnow HH, Lambrecht J, Müller S, Wick LY, Musat N (2019) Quantitation and comparison of phenotypic heterogeneity among single cells of monoclonal microbial populations. Front Microbiol 10:2814
Braun PD, Schulz-Vogt HN, Vogts A, Nausch M (2018) Differences in the accumulation of phosphorus between vegetative cells and heterocysts in the cyanobacterium Nodularia spumigena. Sci Rep 8(1):1–6
Gross A, Lin Y, Weber PK, Pett-Ridge J, Silver WL (2020) The role of soil redox conditions in microbial phosphorus cycling in humid tropical forests. Ecology 101(2):e02928
Ackerman CM, Weber PK, Xiao T, Thai B, Kuo TJ, Zhang E, Pett-Ridge J, Chang CJ (2018) Multimodal LA-ICP-MS and nanoSIMS imaging enables copper mapping within photoreceptor megamitochondria in a zebrafish model of Menkes disease. Metallomics 10(3):474–485
Hong-Hermesdorf A, Miethke M, Gallaher SD, Kropat J, Dodani SC, Barupala D, Chan J, Domaille DW, Shirasaki DI, Loo JA, Weber PK, Pett-Ridge J, Stemmler TL, Chang CJ, Merchant SS (2014) Selective sub-cellular visualization of trace metals identifies dynamic sites of Cu accumulation in Chlamydomonas. Nat Chem Biol 10:1034–1042
Dawson KS, Scheller S, Dillon JG, Orphan VJ (2016) Stable isotope phenotyping via cluster analysis of NanoSIMS data as a method for characterizing distinct microbial ecophysiologies and sulfur-cycling in the environment. Front Microbiol 7:774
Berry D, Mader E, Lee TK, Woebken D, Wang Y, Zhu D, Palatinszky M, Schintlmeister A, Schmid MC, Hanson BT (2015) Tracking heavy water (D2O) incorporation for identifying and sorting active microbial cells. Proc Natl Acad Sci 112(2):E194–E203
Kopf SH, McGlynn SE, Green-Saxena A, Guan Y, Newman DK, Orphan VJ (2015) Heavy water and 15 N labelling with N ano SIMS analysis reveals growth rate-dependent metabolic heterogeneity in chemostats. Environ Microbiol 17(7):2542–2556
Ploug H, Musat N, Adam B, Moraru CL, Lavik G, Vagner T, Bergman B, Kuypers MMM (2010) Carbon and nitrogen fluxes associated with the cyanobacterium Aphanizomenon sp. in the Baltic Sea. ISME J 4(9):1215–1223
Scheller S, Yu H, Chadwick GL, McGlynn SE, Orphan VJ (2016) Artificial electron acceptors decouple archaeal methane oxidation from sulfate reduction. Science 351(6274):703–707
Dekas AE, Connon SA, Chadwick GL, Trembath-Reichert E, Orphan VJ (2016) Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses. ISME J 10(3):678–692
Green-Saxena A, Dekas AE, Dalleska NF, Orphan VJ (2014) Nitrate-based niche differentiation by distinct sulfate-reducing bacteria involved in the anaerobic oxidation of methane. ISME J 8(1):150–163
Milucka J, Kirf M, Lu L, Krupke A, Lam P, Littmann S, Kuypers MMM, Schubert CJ (2015) Methane oxidation coupled to oxygenic photosynthesis in anoxic waters. ISME J 9(9):1991–2002
Marlow JJ, Steele JA, Ziebis W, Thurber AR, Levin LA, Orphan VJ (2014) Carbonate-hosted methanotrophy represents an unrecognized methane sink in the deep sea. Nat Commun 5(1):1–12
Oswald K, Graf JS, Littmann S, Tienken D, Brand A, Wehrli B, Albertsen M, Daims H, Wagner M, Kuypers MM (2017) Crenothrix are major methane consumers in stratified lakes. ISME J 11(9):2124–2140
Foster RA, Kuypers MM, Vagner T, Paerl RW, Musat N, Zehr JP (2011) Nitrogen fixation and transfer in open ocean diatom-cyanobacterial symbioses. ISME J 5(9):1484–1493
Thompson AW, Foster RA, Krupke A, Carter BJ, Musat N, Vaulot D, Kuypers MMM, Zehr JP (2012) Unicellular cyanobacterium symbiotic with a single-celled eukaryotic alga. Science 337(6101):1546–1550
Adam B, Klawonn I, Sveden JB, Bergkvist J, Nahar N, Walve J, Littmann S, Whitehouse MJ, Lavik G, Kuypers MMM, Ploug H (2016) N2-fixation, ammonium release and N-transfer to the microbial and classical food web within a plankton community. ISME J 10(2):450–459
Berry D, Stecher B, Schintlmeister A, Reichert J, Brugiroux S, Wild B, Wanek W, Richter A, Rauch I, Decker T (2013) Host-compound foraging by intestinal microbiota revealed by single-cell stable isotope probing. Proc Natl Acad Sci 110(12):4720–4725
Carpenter KJ, Weber PK, Davisson ML, Pett-Ridge J, Haverty MI, Keeling PJ (2013) Correlated SEM, FIB-SEM, TEM, and NanoSIMS imaging of microbes from the hindgut of a lower termite: methods for in situ functional and ecological studies of uncultivable microbes. Microsc Microanal 19(06):1490–1501
Tai V, Carpenter KJ, Weber PK, Nalepa CA, Perlman SJ, Keeling PJ (2016) Genome evolution and nitrogen-fixation in bacterial ectosymbionts of a protist inhabiting wood-feeding cockroaches. Appl Environ Microbiol. https://doi.org/10.1128/AEM.00611-16
Ceh J, Kilburn MR, Cliff JB, Raina JB, van Keulen M, Bourne DG (2013) Nutrient cycling in early coral life stages: Pocillopora damicornis larvae provide their algal symbiont (Symbiodinium) with nitrogen acquired from bacterial associates. Ecol Evol 3(8):2393–2400
Pernice M, Dunn SR, Tonk L, Dove S, Domart-Coulon I, Hoppe P, Schintlmeister A, Wagner M, Meibom A (2015) A nanoscale secondary ion mass spectrometry study of dinoflagellate functional diversity in reef-building corals. Environ Microbiol 17(10):3570–3580
Wangpraseurt D, Pernice M, Guagliardo P, Kilburn MR, Clode PL, Polerecky L, Kühl M (2016) Light microenvironment and single-cell gradients of carbon fixation in tissues of symbiont-bearing corals. ISME J 10(3):788–792
Lema KA, Clode PL, Kilburn MR, Thornton R, Willis BL, Bourne DG (2016) Imaging the uptake of nitrogen-fixing bacteria into larvae of the coral Acropora millepora. ISME J 10(7):1804–1808
Kopp C, Domart-Coulon I, Barthelemy D, Meibom A (2016) Nutritional input from dinoflagellate symbionts in reef-building corals is minimal during planula larval life stage. Sci Adv 2(3):e1500681
Yang S-H, Tandon K, Lu C-Y, Wada N, Shih C-J, Hsiao SS-Y, Jane W-N, Lee T-C, Yang C-M, Liu C-T (2019) Metagenomic, phylogenetic, and functional characterization of predominant endolithic green sulfur bacteria in the coral Isopora palifera. Microbiome 7(1):1–13
Samo TJ, Kimbrel JA, Nilson DJ, Pett-Ridge J, Weber PK, Mayali X (2018) Attachment between heterotrophic bacteria and microalgae influences symbiotic microscale interactions. Environ Microbiol 20(2):4385–4400
de-Bashan LE, Mayali X, Bebout BM, Weber PK, Detweiler AM, Hernandez J-P, Prufert-Bebout L, Bashan Y (2016) Establishment of stable synthetic mutualism without co-evolution between microalgae and bacteria demonstrated by mutual transfer of metabolites (NanoSIMS isotopic imaging) and persistent physical association (fluorescent in situ hybridization). Algal Res 15:179–186
Alonso C, Musat N, Adam B, Kuypers M, Amann R (2012) HISH–SIMS analysis of bacterial uptake of algal-derived carbon in the Río de la Plata estuary. Syst Appl Microbiol 35(8):541–548
Leroy C, Jauneau A, Martinez Y, Cabin-Flaman A, Gibouin D, Orivel J, Séjalon-Delmas N (2017) Exploring fungus–plant N transfer in a tripartite ant–plant–fungus mutualism. Ann Bot 120(3):417–426
Lee JZ, Burow LC, Woebken D, Everroad RC, Kubo MD, Spormann AM, Weber PK, Pett-Ridge J, Bebout BM, Hoehler TM (2014) Fermentation couples chloroflexi and sulfate-reducing bacteria to cyanobacteria in hypersaline microbial mats. Front Microbiol 5:61
Nuccio EE, Hodge A, Pett-Ridge J, Herman DJ, Weber PK, Firestone MK (2013) An arbuscular mycorrhizal fungus significantly modifies the soil bacterial community and nitrogen cycling during litter decomposition. Environ Microbiol 15(6):1870–1881
Kaiser C, Kilburn MR, Clode PL, Fuchslueger L, Koranda M, Cliff JB, Solaiman ZM, Murphy DV (2015) Exploring the transfer of recent plant photosynthates to soil microbes: mycorrhizal pathway vs direct root exudation. New Phytol 205(4):1537–1551
Kuga Y, Sakamoto N, Yurimoto H (2014) Stable isotope cellular imaging reveals that both live and degenerating fungal pelotons transfer carbon and nitrogen to orchid protocorms. New Phytol 202(2):594–605
Pett-Ridge J, Firestone MK (2017) Using stable isotopes to explore root-microbe-mineral interactions in soil. Rhizosphere 3:244–253
Hestrin R, Hammer EC, Mueller CW, Lehmann J (2019) Synergies between mycorrhizal fungi and soil microbial communities increase plant nitrogen acquisition. Commun Biol 2(1):1–9
Gorka S, Dietrich M, Mayerhofer W, Gabriel R, Wiesenbauer J, Martin V, Zheng Q, Imai B, Prommer J, Weidinger M (2019) Rapid transfer of plant photosynthates to soil bacteria via ectomycorrhizal hyphae and its interaction with nitrogen availability. Front Microbiol 10:168
Worrich A, Stryhanyuk H, Musat N, König S, Banitz T, Centler F, Frank K, Thullner M, Harms H, Richnow H-H (2017) Mycelium-mediated transfer of water and nutrients stimulates bacterial activity in dry and oligotrophic environments. Nat Commun 8(1):1–9
Bougoure J, Ludwig M, Brundrett M, Cliff J, Clode P, Kilburn M, Grierson P (2014) High-resolution secondary ion mass spectrometry analysis of carbon dynamics in mycorrhizas formed by an obligately myco-heterotrophic orchid. Plant Cell Environ 37(5):1223–1230
Hill PW, Broughton R, Bougoure J, Havelange W, Newsham KK, Grant H, Murphy DV, Clode P, Ramayah S, Marsden KA (2019) Angiosperm symbioses with non-mycorrhizal fungal partners enhance N acquisition from ancient organic matter in a warming maritime Antarctic. Ecol Lett 22(12):2111–2119
Mergelov N, Mueller CW, Prater I, Shorkunov I, Dolgikh A, Zazovskaya E, Shishkov V, Krupskaya V, Abrosimov K, Cherkinsky A (2018) Alteration of rocks by endolithic organisms is one of the pathways for the beginning of soils on Earth. Sci Rep 8(1):1–15
Kopittke PM, Dalal RC, Hoeschen C, Li C, Menzies NW, Mueller CW (2020) Soil organic matter is stabilized by organo-mineral associations through two key processes: the role of the carbon to nitrogen ratio. Geoderma 357:113974
Keiluweit M, Bougoure JJ, Zeglin LH, Myrold DD, Weber PK, Pett-Ridge J, Kleber M, Nico PS (2012) Nano-scale investigation of the association of microbial nitrogen residues with iron (hydr)oxides in a forest soil O-horizon. Geochim Cosmochim Acta 95:213–226
Keiluweit M, Bougoure JJ, Nico PS, Pett-Ridge J, Weber PK, Kleber M (2015) Mineral protection of soil carbon counteracted by root exudates. Nat Clim Chang 5(6):588–595
Morrison KD, Misra R, Williams LB (2016) Unearthing the antibacterial mechanism of medicinal clay: a geochemical approach to combating antibiotic resistance. Sci Rep 6:19043
Londono SC, Hartnett HE, Williams LB (2017) Antibacterial activity of aluminum in clay from the Colombian Amazon. Environ Sci Technol 51(4):2401–2408
Eichorst SA, Strasser F, Woyke T, Schintlmeister A, Wagner M, Woebken D (2015) Advancements in the application of NanoSIMS and Raman microspectroscopy to investigate the activity of microbial cells in soils. FEMS Microbiol Ecol 91(10):fiv106
Pasulka AL, Thamatrakoln K, Kopf SH, Guan Y, Poulos B, Moradian A, Sweredoski MJ, Hess S, Sullivan MB, Bidle KD (2018) Interrogating marine virus-host interactions and elemental transfer with BONCAT and nanoSIMS-based methods. Environ Microbiol 20(2):671–692
Greenwood DJ, Dos Santos MS, Huang S, Russell MR, Collinson LM, MacRae JI, West A, Jiang H, Gutierrez MG (2019) Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages. Science 364(6447):1279–1282
Gates SD, Condit RC, Moussatche N, Stewart BJ, Malkin AJ, Weber PK (2018) High initial sputter rate found for vaccinia virions using isotopic labeling, nanoSIMS, and AFM. Anal Chem 90(3):1613–1620
Stuart RK, Mayali X, Boaro AA, Zemla A, Everroad RC, Nilson D, Weber PK, Lipton M, Bebout BM, Pett-Ridge J (2016) Light regimes shape utilization of extracellular organic C and N in a cyanobacterial biofilm. mBio 7(3):e00650–e00616
Stuart RK, Mayali X, Lee JZ, Everroad RC, Hwang M, Bebout BM, Weber PK, Pett-Ridge J, Thelen MP (2016) Cyanobacterial reuse of extracellular organic carbon in microbial mats. ISME J 10(5):1240–1251
Stuart RK, Mayali X, Thelen MP, Pett-Ridge J, Weber PK (2017) Measuring cyanobacterial metabolism in biofilms with NanoSIMS isotope imaging and scanning electron microscopy (SEM). Bioprotocol 7:e2263
Probst AJ, Weinmaier T, Raymann K, Perras A, Emerson JB, Rattei T, Wanner G, Klingl A, Berg IA, Yoshinaga M, Viehweger B, Hinrichs K-U, Thomas BC, Meck S, Auerbach AK, Heise M, Schintlmeister A, Schmid M, Wagner M, Gribaldo S, Banfield JF, Moissl-Eichinger C (2014) Biology of a widespread uncultivated archaeon that contributes to carbon fixation in the subsurface. Nat Commun 5:5497
Tveit AT, Hestnes AG, Robinson SL, Schintlmeister A, Dedysh SN, Jehmlich N, von Bergen M, Herbold C, Wagner M, Richter A (2019) Widespread soil bacterium that oxidizes atmospheric methane. Proc Natl Acad Sci 116(17):8515–8524
Sheik AR, Muller EE, Audinot J-N, Lebrun LA, Grysan P, Guignard C, Wilmes P (2016) In situ phenotypic heterogeneity among single cells of the filamentous bacterium Candidatus Microthrix parvicella. ISME J 10(5):1274–1279
Nikolic N, Schreiber F, Kiviet DJ, Bergmiller T, Littmann S, Kuypers MM, Ackermann M (2017) Cell-to-cell variation and specialization in sugar metabolism in clonal bacterial populations. PLoS Genet 13(12):e1007122
Gangwe Nana GY, Ripoll C, Cabin-Flaman A, Gibouin D, Delaune A, Jannière L, Grancher G, Chagny G, Loutelier-Bourhis C, Lentzen E (2018) Division-based, growth rate diversity in bacteria. Front Microbiol 9:849
Zimmermann M, Escrig S, Hübschmann T, Kirf MK, Brand A, Inglis RF, Musat N, Müller S, Meibom A, Ackermann M (2015) Phenotypic heterogeneity in metabolic traits among single cells of a rare bacterial species in its natural environment quantified with a combination of flow cell sorting and NanoSIMS. Front Microbiol 6:243
Tsednee M, Castruita M, Salomé PA, Sharma A, Lewis BE, Schmollinger SR, Strenkert D, Holbrook K, Otegui MS, Khatua K (2019) Manganese co-localizes with calcium and phosphorus in Chlamydomonas acidocalcisomes and is mobilized in manganese-deficient conditions. J Biol Chem 294(46):17626–17641
Kessler N, Armoza-Zvuloni R, Wang S, Basu S, Weber PK, Stuart RK, Shaked Y (2020) Selective collection of iron-rich dust particles by natural Trichodesmium colonies. ISME J 14(1):91–103
Newsome L, Lopez Adams R, Downie HF, Moore KL, Lloyd JR (2018) NanoSIMS imaging of extracellular electron transport processes during microbial iron (III) reduction. FEMS Microbiol Ecol 94(8):fiy104
Fleming E, Woyke T, Donatello R, Kuypers MM, Sczyrba A, Littmann S, Emerson D (2018) Insights into the fundamental physiology of the uncultured Fe-oxidizing bacterium Leptothrix ochracea. Appl Environ Microbiol 84(9). https://doi.org/10.1128/AEM.02239-17
Stryhanyuk H, Calabrese F, Kümmel S, Musat F, Richnow HH, Musat N (2018) Calculation of single cell assimilation rates from SIP-NanoSIMS-derived isotope ratios: a comprehensive approach. Front Microbiol 9:2342
Arandia-Gorostidi N, Weber PK, Alonso-Saez L, Moran XAG, Mayali X (2017) Elevated temperature increases carbon and nitrogen fluxes between phytoplankton and heterotrophic bacteria through physical attachment. ISME J 11(3):641–650
Frisz JF, Lou K, Klitzing HA, Hanafin WP, Lizunov V, Wilson RL, Carpenter KJ, Kim R, Hutcheon ID, Zimmerberg J (2013) Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts. Proc Natl Acad Sci 110(8):E613–E622
Smith NS, Boswell RW, Tesch PP, Martin NP (2017) Rf system, magnetic filter, and high voltage isolation for an inductively coupled plasma ion source, Google Patents
Smith N, Tesch P, Martin N, Kinion D (2008) A high brightness source for nano-probe secondary ion mass spectrometry. Appl Surf Sci 255(4):1606–1609
Malherbe J, Penen F, Isaure M-P, Frank J, Hause G, Dobritzsch D, Gontier E, Horréard FO, Hillion FO, Schaumlöffel D (2016) A new radio frequency plasma oxygen primary ion source on nano secondary ion mass spectrometry for improved lateral resolution and detection of electropositive elements at single cell level. Anal Chem 88(14):7130–7136
Cabin-Flaman A, Monnier AFO, Coffinier Y, Audinot J-N, Gibouin D, Wirtz T, Boukherroub R, Migeon H-N, Bensimon A, Jannière L (2011) Combed single DNA molecules imaged by secondary ion mass spectrometry. Anal Chem 83(18):6940–6947
Cabin-Flaman A, Monnier A-F, Coffinier Y, Audinot J-N, Gibouin D, Wirtz T, Boukherroub R, Migeon H-N, Bensimon A, Jannière L (2016) Combining combing and secondary ion mass spectrometry to study DNA on chips using 13C and 15N labeling. F1000Res 5:27429742
Weber PK, Graham GA, Teslich NE, MoberlyChan W, Ghosal S, Leighton TJ, Wheeler KE (2010) NanoSIMS imaging of Bacillus spores sectioned by focused ion beam. J Microsc 238:189–199
Wilson RG, Stevie FA, Magee CW (1989) Secondary ion mass spectrometry: a practical handbook for depth profiling and bulk impurity analysis. Wiley, New York, NY
Polerecky L, Adam B, Milucka J, Musat N, Vagner T, Kuypers MM (2012) Look@ NanoSIMS–a tool for the analysis of nanoSIMS data in environmental microbiology. Environ Microbiol 14(4):1009–1023
Huang W, Hammel KE, Hao J, Thompson A, Timokhin VI, Hall SJ (2019) Enrichment of lignin-derived carbon in mineral-associated soil organic matter. Environ Sci Technol 53(13):7522–7531
Whitman T, Zhu Z, Lehmann J (2014) Carbon mineralizability determines interactive effects on mineralization of pyrogenic organic matter and soil organic carbon. Environ Sci Technol 48(23):13727–13734
Hatton P-J, Remusat L, Zeller B, Brewer EA, Derrien D (2015) NanoSIMS investigation of glycine-derived C and N retention with soil organo-mineral associations. Biogeochemistry 125(3):303–313
Li T, Wu TD, Mazeas L, Toffin L, Guerquin-Kern JL, Leblon G, Bouchez T (2008) Simultaneous analysis of microbial identity and function using NanoSIMS. Environ Microbiol 10(3):580–588
Benn PA, Perle MA (1992) Chromosome staining and banding techniques. In: Rooney DE, Czepulkowski BH (eds) Human cytogenetics: volume I, constitutional analysis: a practical approach. Oxford University Press, New York, NY
Latt SA (1973) Microfluorometric detection of deoxyribonucleic acid replication in human metaphase chromosomes. Proc Natl Acad Sci U S A 49:3395–3399
Manefield M, Whiteley AS, Griffiths RI, Bailey MJ (2002) RNA stable isotope probing, a novel means of linking microbial community function to phylogeny. Appl Environ Microbiol 68:5367–5373
Radajewski S, Ineson P, Parekh NR, Murrell J (2000) Stable-isotope probing as a tool in microbial ecology. Nature 403(10):646–649
Jensen LHS, Cheng T, Plane FOV, Escrig S, Comment A, van den Brandt B, Humbel BM, Meibom A (2016) En route to ion microprobe analysis of soluble compounds at the single cell level: the CryoNanoSIMS. In: European microscopy congress 2016: proceedings. Wiley, New York, NY
Lovrić J, Malmberg P, Johansson BR, Fletcher JS, Ewing AG (2016) Multimodal imaging of chemically fixed cells in preparation for NanoSIMS. Anal Chem 88(17):8841–8848
Gibbin E, Gavish A, Domart-Coulon I, Kramarsky-Winter E, Shapiro O, Meibom A, Vardi A (2018) Using NanoSIMS coupled with microfluidics to visualize the early stages of coral infection by Vibrio coralliilyticus. BMC Microbiol 18(1):1–10
Nunan N, Ritz K, Crabb D, Harris K, Wu K, Crawford JW, Young IM (2001) Quantification of the in situ distribution of soil bacteria by large-scale imaging of thin sections of undisturbed soil. FEMS Microbiol Ecol 37(1):67–77
Kuo J (2007) Electron microscopy: methods and protocols. In: Methods in molecular biology, 2nd edn. Humana, Totowa, NJ
Tippkötter R, Ritz K (1996) Evaluation of polyester, epoxy and acrylic resins for suitability in preparation of soil thin sections for in situ biological studies. Geoderma 69(1–2):31–57
Chandra S, Morrison GH (1992) Sample preparation of animal tissues and cell cultures for secondary ion mass spectrometry (SIMS) microscopy. Biol Cell 74:31–42
Dykstra MJ, Reuss LE (eds) (2003) Biological electron microscopy: theory, techniques and troubleshooting, 2nd edn. Kluwer Academic/Plenum Publishers, New York, NY
Echlin P (1992) Low-temperature microscopy and analysis. Springer, New York, NY
Musat N, Stryhanyuk H, Bombach P, Adrian L, Audinot J-N, Richnow HH (2014) The effect of FISH and CARD-FISH on the isotopic composition of 13C- and 15N-labeled Pseudomonas putida cells measured by nanoSIMS. Syst Appl Microbiol 37(4):267–276
Woebken D, Burow LC, Behnam F, Mayali X, Schintlmeister A, Fleming ED, Prufert-Bebout L, Singer SW, Lopez Cortes A, Hoehler TM, Pett-Ridge J, Spormann AM, Wagner M, Weber PK, Bebout BM (2015) Revisiting N-2 fixation in Guerrero Negro intertidal microbial mats with a functional single-cell approach. ISME J 9(2):485–496
Herrmann AM, Clode PL, Fletcher IR, Nunan N, Stockdale EA, O'Donnel AG, Murphy DV (2007) A novel method for the study of the biophysical interface in soils using nano-scale secondary ion mass spectrometry. Rapid Commun Mass Spectrom 21(1):29–34
Weng N, Jiang H, Wang W-X (2017) In situ subcellular imaging of copper and zinc in contaminated oysters revealed by nanoscale secondary ion mass spectrometry. Environ Sci Technol 51(24):14426–14435
Rogge A, Flintrop CM, Iversen MH, Salter I, Fong AA, Vogts A, Waite AM (2018) Hard and soft plastic resin embedding for single-cell element uptake investigations of marine-snow-associated microorganisms using nano-scale secondary ion mass spectrometry. Limnol Oceanogr Methods 16(8):484–503
Fike DA, Gammon CL, Ziebis W, Orphan VJ (2008) Micron-scale mapping of sulfur cycling across the oxycline of a cyanobacterial mat: a paired nanoSIMS and CARD-FISH approach. ISME J 2(7):749–759
De Gregorio BT, Stroud RM, Nittler LR, Alexander CMOD, Kilcoyne ALD, Zega TJ (2010) Isotopic anomalies in organic nanoglobules from Comet 81P/Wild 2: comparison to Murchison nanoglobules and isotopic anomalies induced in terrestrial organics by electron irradiation. Geochim Cosmochim Acta 74(15):4454–4470
Lehmann J, Liang BQ, Solomon D, Lerotic M, Luizao F, Kinyangi J, Schafer T, Wirick S, Jacobsen C (2005) Near-edge X-ray absorption fine structure (NEXAFS) spectroscopy for mapping nano-scale distribution of organic carbon forms in soil: application to black carbon particles. Global Biogeochem Cycles 29:Art. No. GB1013
Flynn GJ, Keller LP, Jacobsen C, Wirick S (2004) An assessment of the amount and types of organic matter contributed to the Earth by interplanetary dust. Adv Space Res 33:57–66
Gnaser H (1997) Formation of metastable N2- and CO-anions in sputtering. Phys Rev A 56(4):R2518
McMahon G, Saint-Cyr HF, Lechene C, Unkefer CJ (2006) CN− secondary ions form by recombination as demonstrated using multi-isotope mass spectrometry of 13C- and 15N-labeled polyglycine. J Am Soc Mass Spectrom 17(8):1181–1187
Mayali X, Weber PK, Nuccio E, Lietard J, Somoza M, Blazewicz SJ, Pett-Ridge J (2019) Chip-SIP: stable isotope probing analyzed with rRNA-targeted microarrays and NanoSIMS. In: Stable isotope probing. Springer, New York, NY, pp 71–87
Gnaser H (1999) Singly-and doubly-negative carbon clusters in sputtering: energy spectra, abundance distributions and unimolecular fragmentation. Nucl Instrum Methods Phys Res, Sect B 149(1–2):38–52
Weber PK, Bacon CR, Hutcheon ID, Ingram BL, Wooden JL (2005) Ion microprobe measurement of strontium isotopes in calcium carbonate with application to salmon otoliths. Geochim Cosmochim Acta 69(5):1225–1239
Ghosal S, Fallon SJ, Leighton T, Wheeler K, Hutcheon ID, Weber PK (2006) Analysis of bacterial spore permeability to water and ions using NanoSecondary Ion Mass Spectrometry (NanoSIMS). Abstr Pap Am Chem Soc 231:3
Wolfe-Simon F, Blum JS, Kulp TR, Gordon GW, Hoeft SE, Pett-Ridge J, Stolz JF, Webb SM, Weber PK, Davies PCW, Anbar AD, Oremland RS (2011) A bacterium that can grow by using arsenic instead of phosphorus. Science 332(6034):1163–1166
Hauri EH, Papineau D, Wang J, Hillion F (2016) High-precision analysis of multiple sulfur isotopes using NanoSIMS. Chem Geol 420:148–161
Chandra S, Smith DR, Morrison GH (2000) Subcellular imaging by dynamic SIMS ion microscopy. Anal Chem 72:104A–114A
Guerquin-Kern JL, Wu TD, Quintana C, Croisy A (2005) Progress in analytical imaging of the cell by dynamic secondary ion mass spectrometry (SIMS microscopy). BBA-Gen Subjects 1724(3):228–238
Burns MS, File DM, Deline V, Galle P (1986) Matrix effects in secondary ion mass spectrometric analysis of biological tissue. Scan Electron Microscopy 1986(4):1277–1290
Harris WC, Chandra S, Morrison GH (1983) Ion implantation for quantitative ion microscopy of biological soft tissue. Anal Chem 55(12):1959–1963
Phinney D (2006) Quantitative analysis of microstructures by secondary ion mass spectrometry. Microsc Microanal 12(4):352
Decelle J, Veronesi G, Gallet B, Stryhanyuk H, Benettoni P, Schmidt M, Tucoulou R, Passarelli M, Bohic S, Clode P (2020) Subcellular chemical imaging: new avenues in cell biology. Trends Cell Biol 30(3):173–188
Penen F, Malherbe J, Isaure M-P, Dobritzsch D, Bertalan I, Gontier E, Le Coustumer P, Schaumlöffel D (2016) Chemical bioimaging for the subcellular localization of trace elements by high contrast TEM, TEM/X-EDS, and NanoSIMS. J Trace Elem Med Biol 37:62–68
Nomaki H, LeKieffre C, Escrig S, Meibom A, Yagyu S, Richardson EA, Matsuzaki T, Murayama M, Geslin E, Bernhard JM (2018) Innovative TEM-coupled approaches to study foraminiferal cells. Mar Micropaleontol 138:90–104
Kraft ML, Weber PK, Longo ML, Hutcheon ID, Boxer SG (2006) Phase separation of lipid membranes analyzed with high-resolution secondary ion mass spectrometry. Science 313:1948–1951
Wirtz T, Fleming Y, Gysin U, Glatzel T, Wegmann U, Meyer E, Maier U, Rychen J (2013) Combined SIMS-SPM instrument for high sensitivity and high-resolution elemental 3D analysis. Surf Interface Anal 45(1):513–516
Orphan VJ, House CH, Hinrichs K-U, McKeegan KD, DeLong EF (2001) Methane-consuming Archaea revealed by directly coupled isotopic and phylogenetic analysis. Science 293:484–487
Amann RI, Krumholz L, Stahl DA (1990) Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology. J Bacteriol 172(2):762–770
Pernthaler A, Pernthaler J, Amann R (2002) Fluorescence in situ hybridization and catalyzed reporter deposition for the identification of marine bacteria. Appl Environ Microbiol 68(6):3094–3101
Woebken D, Burow LC, Prufert-Bebout L, Bebout BM, Hoehler TM, Pett-Ridge J, Spormann AM, Weber PK, Singer SW (2012) Identification of a novel cyanobacterial group as active diazotrophs in a coastal microbial mat using NanoSIMS analysis. ISME J 6(7):1427–1439
Lemaire R, Webb RI, Yuan Z (2008) Micro-scale observations of the structure of aerobic microbial granules used for the treatment of nutrient-rich industrial wastewater. ISME J 2(5):528–541
Hatzenpichler R, Scheller S, Tavormina PL, Babin BM, Tirrell DA, Orphan VJ (2014) In situ visualization of newly synthesized proteins in environmental microbes using amino acid tagging and click chemistry. Environ Microbiol 16(8):2568–2590
Bradley JP, Dai ZR, Erni R, Browning ND, Graham GA, Weber PK, Smith JB, Hutcheon ID, Ishii H, Bajt S, Floss C, Stadermann FJ, Sandford S (2005) An astronomical 2175 Å feature in interplanetary dust particles. Science 307:244–247
Remusat L, Hatton P-J, Nico PS, Zeller B, Kleber M, Derrien D (2012) NanoSIMS study of organic matter associated with soil aggregates: advantages, limitations, and combination with STXM. Environ Sci Technol 46(7):3943–3949
De Samber B, De Rycke R, De Bruyne M, Kienhuis M, Sandblad L, Bohic S, Cloetens P, Urban C, Polerecky L, Vincze L (2020) Effect of sample preparation techniques upon single cell chemical imaging: a practical comparison between synchrotron radiation based X-ray fluorescence (SR-XRF) and nanoscopic secondary ion mass spectrometry (nano-SIMS). Anal Chim Acta 1106:22–32
Lehmann J, Kinyangi J, Solomon D (2007) Organic matter stabilization in soil microaggregates: implications from spatial heterogeneity of organic carbon contents and carbon forms. Biogeochemistry 85(1):45–57
Wan J, Tyliszczak T, Tokunaga TK (2007) Organic carbon distribution, speciation, and elemental correlations within soil microaggregates: applications of STXM and NEXAFS spectroscopy. Geochim Cosmochim Acta 71(22):5439–5449
Kopp C, Wisztorski M, Revel J, Mehiri M, Dani V, Capron L, Carette D, Fournier I, Massi L, Mouajjah D (2015) MALDI-MS and NanoSIMS imaging techniques to study cnidarian–dinoflagellate symbioses. Zoology 118(2):125–131
Schlüter S, Eickhorst T, Mueller CW (2018) Correlative imaging reveals holistic view of soil microenvironments. Environ Sci Technol 53(2):829–837
Lin S, Henze S, Lundgren P, Bergman B, Carpenter EJ (1998) Whole-cell immunolocalization of nitrogenase in marine diazotrophic cyanobacteria, Trichodesmium spp. Appl Environ Microbiol 64(8):3052–3058
Levenson RM, Borowsky AD, Angelo M (2015) Immunohistochemistry and mass spectrometry for highly multiplexed cellular molecular imaging. Lab Investig 95(4):397–405
Singer SW, Chan CS, Hwang MH, Zemla A, VerBerkmoes NC, Hettich RL, Banfield JF, Thelen MP (2008) Characterization of cytochrome579, an unusual cytochrome isolated from an iron-oxidizing microbial community. Appl Environ Microbiol 74:4454–4462
Gerard E, Guyot F, Philippot P, Lopez-Garcia P (2005) Fluorescence in situ hybridization coupled to ultra small immunogold detection to identify prokaryotic cells using transmission and scanning electron microscopy. J Microbiol Methods 63:20–28
Mayali X, Weber PK, Mabery S, Pett-Ridge J (2014) Phylogenetic patterns in the microbial response to resource availability: amino acid incorporation in San Francisco Bay. PLoS One 9(4):e95842
Mayali X, Weber PK, Pett-Ridge J (2013) Taxon-specific C:N relative use efficiency for amino acids in an estuarine community. FEMS Microbiol Ecol 83(2):402–412
Bryson S, Li Z, Chavez F, Weber PK, Pett-Ridge J, Hettich RL, Pan C, Mayali X, Mueller RS (2017) Phylogenetically conserved resource partitioning in the coastal microbial loop. ISME J 11(12):2781–2792
Smith DF, Kiss A, Leach FE, Robinson EW, Paša-Tolić L, Heeren RM (2013) High mass accuracy and high mass resolving power FT-ICR secondary ion mass spectrometry for biological tissue imaging. Anal Bioanal Chem 405(18):6069–6076
Steele AV, Schwarzkopf A, McClelland JJ, Knuffman B (2017) High-brightness Cs focused ion beam from a cold-atomic-beam ion source. Nano Fut 1(1):015005
Hayes JM (2004) An introduction to isotopic calculations. Woods Hole Oceanographic Institution, Woods Hole, MA, USA. https://www.whoi.edu/cms/files/jhayes/2005/9/IsoCalcs30Sept04_5183.pdf
Legin AA, Schintlmeister A, Jakupec MA, Galanski M, Lichtscheidl I, Wagner M, Keppler BK (2014) NanoSIMS combined with fluorescence microscopy as a tool for subcellular imaging of isotopically labeled platinum-based anticancer drugs. Chem Sci 5(8):3135-3143
Acknowledgments
We thank Ian Hutcheon, for his many years of mentorship, advice, and leadership of the LLNL SIMS group. We also thank our many colleagues and collaborators, with special thanks to Steve Blazewicz, Anne Dekas, Ben Jacobsen, Xavier Mayali, Erin Nuccio, Rhona Stuart, and Dagmar Woebken. Christina Ramon plays a critical role in helping to prepare and organize many of the samples we have discussed. This work was funded in part by awards from the DOE OBER Genomic Science program and LLNL Laboratory Directed Research and Development funding and performed under the auspices of the U.S. Department of Energy at Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344.
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Pett-Ridge, J., Weber, P.K. (2022). NanoSIP: NanoSIMS Applications for Microbial Biology. In: Navid, A. (eds) Microbial Systems Biology. Methods in Molecular Biology, vol 2349. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1585-0_6
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