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Raman imaging of microbial colonization in rock—some analytical aspects

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Abstract

Raman imaging allows one to obtain spatially resolved chemical information in a nondestructive manner. Herein, we present analytical aspects of effective in situ and in vivo Raman imaging of algae and cyanobacteria from within their native rock habitats. Specifically, gypsum and halite inhabited by endolithic communities from the hyperarid Atacama Desert were analyzed. Raman imaging of these phototrophic colonization reveals a pigment composition within the aggregates that helps in understanding some of their adaptation strategies to survive in this harsh polyextreme environment. The study is focused on methodical aspects of Raman imaging acquisition and subsequent data processing. Point imaging is compared with line imaging in terms of their image quality, spatial resolution, spectral signal-to-noise ratio, time requirements, and risk of laser-induced sample alteration. The roles of excitation wavelength, exposure time, and step size of the imaging grid on successful Raman imaging results are also discussed.

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Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author upon a reasonable request.

References

  1. Delhaye M, Dhamelincourt P. Raman microprobe and microscope with laser excitation. J Raman Spectrosc. 1975;3:33–43.

    Article  CAS  Google Scholar 

  2. Markwort L, Kip B, Dasilva E, Roussel B. Raman imaging of heterogeneous polymers – a comparison of global versus point illumination. Appl Spectrosc. 1995;49:1411–30.

    Article  CAS  Google Scholar 

  3. Schlücker S, Schaeberle MD, Huffmann SW, Levin IW. Raman microspectroscopy: a comparison of point, line, and wide-field imaging methodologies. Anal Chem. 2003;75:4312–8.

    Article  Google Scholar 

  4. Puppels GJ, Grond M, Greve J. Direct imaging Raman microscope based on tunable wavelength excitation and narrow-band emission detection. Appl Spectrosc. 1993;47:1256–67.

    Article  CAS  Google Scholar 

  5. Baranski R, Baranska M, Schulz H. Changes in carotenoid content and distribution in living plant tissue can be observed and mapped in situ using NIR-FT-Raman spectroscopy. Planta. 2005;222:448–57.

    Article  CAS  Google Scholar 

  6. Schulz H, Baranska M, Baranski R. Potential of NIR-FT-Raman spectroscopy in natural carotenoid analysis. Biopolymers. 2005;7:212–21.

    Article  Google Scholar 

  7. Agarwall UP. Raman imaging to investigate ultrastructure and composition of plant cell walls: distribution of lignin and cellulose in black spruce wood (Picea mariana). Planta. 2006;224:1141–53.

    Article  Google Scholar 

  8. Gierlinger N, Schwaninger M. Chemical imaging of poplar wood cell walls by confocal Raman microscopy. Plant Physiol. 2006;140:1246–54.

    Article  CAS  Google Scholar 

  9. Häninen T, Kontturi E, Vuorinen T. Distribution of lignin and its coniferyl alcohol and coniferyl aldehyde groups in Picea abies and Pinus sylvestris as observed by Raman imaging. Phytochemistry. 2011;72:1889–95.

    Article  Google Scholar 

  10. Gierlinger N, Keplinger T, Harrington M. Imaging of plant cell walls by confocal Raman microscopy. Nat Protoc. 2012;7:1694–708.

    Article  CAS  Google Scholar 

  11. Gierlinger N, Keplinger T, Harrington M, Schwanninger M. Raman imaging of lignocellulosic feedstock. In: van de Ven T, Kadla J, editors. Cellulose biomass conversion 3. Rijeka: INTECH; 2013. p. 159–92.

    Google Scholar 

  12. Ji Z, Ma JF, Zhang ZH, Xu F, Sun RC. Distribution of lignin and cellulose in compression wood tracheids of Pinus yunnanensis determined by fluorescence microscopy and confocal Raman microscopy. Ind Crop Prod. 2013;47:212–7.

    Article  CAS  Google Scholar 

  13. Gowen AA, Feng Y, Gaston E, Valdramidis V. Recent applications of hyperspectral imaging in microbiology. Talanta. 2015;137:43–54.

    Article  CAS  Google Scholar 

  14. Wang A, Korotev RL, Jolliff BL, Ling Z. Raman imaging of extraterrestrial materials. Planet Space Sci. 2015;112:23–34.

    Article  Google Scholar 

  15. Marshall CP, Olcott MA. (2013). Raman hyperspectral imaging of microfossils: potential pitfalls. Astrobiology. 2013;13:920–31.

    Article  CAS  Google Scholar 

  16. Emry JR, Olcott Marshall A, Marchall CP. Evaluating the effects of autofluorescence during Raman hyperspectral imaging. Geostand Geoanal Res. 2015;40:29–47.

    Article  Google Scholar 

  17. Hofmann A, Bolhar R, Orberger F. Cherts of the Barberton greenstone belt, South Africa: petrology and trace-element geochemistry of 3.5 to 3.3 Ga old silicified volcanoclastic sediments. S Afr J Geol. 2013;116:297–322.

    Article  CAS  Google Scholar 

  18. Schopf JW, Kudryavtsev AB, Walter MR, Van Kranendonk MJ, Williford KH, Kozdon R, et al. Sulfur-cycling fossil bacteria from the 1.8-Ga Duck Creek Formation provide promising evidence of evolution’s null hypothesis. Proc Natl Acad Sci U S A. 2015;112:2087–92.

    Article  CAS  Google Scholar 

  19. Foucher F, Westall F. Raman imaging of metastable opal in carbonaceous microfossils of the 700-800 Ma old Draken formation. Astrobiology. 2013;13:57–67.

    Article  CAS  Google Scholar 

  20. Foucher F, Lopez-Reyes G, Bost N, Rull-Perez F, Rüβmann P, Westall F. Effect of grain size distribution on Raman analyses and the consequences for in situ planetary missions. J Raman Spectrosc. 2013;44:916–25.

    Article  CAS  Google Scholar 

  21. Westall F, Foucher F, Bost N, Bertrand M, Loizeau D, Vago JL, et al. Biosignatures on Mars: what, where, and how? Implications for search for Martian life. Astrobiology. 2015;15:998–1029.

    Article  Google Scholar 

  22. Vítek P, Ascaso C, Artieda O, Wierzchos J. Raman imaging in geomicrobiology: endolithic phototrophic microorganisms in gypsum from the extreme sun irradiation area in the Atacama Desert. Anal Bioanal Chem. 2016;408:483–92.

    Article  Google Scholar 

  23. Vítek P, Ascaso C, Artieda O, Casero MC, Wierzchos J. Discovery of carotenoid red-shift in endolithic cyanobacteria from the Atacama Desert. Sci Rep. 2017;7:11116.

    Article  Google Scholar 

  24. Rooney JS, Tarling MS, Smith SAF, Gordon KC. Submicron Raman spectroscopy mapping of serpentinite fault rocks. J Raman Spectrosc. 2017;49:279–86.

    Article  Google Scholar 

  25. Mosca S, Artesani A, Gulotta D, Nevin A, Goidanich S, Valentini G, et al. Raman mapping and time-resolved photoluminescence imaging for the analysis of a cross-section from a modern gypsum sculpture. Microchem J. 2018;139:500–5.

    Article  CAS  Google Scholar 

  26. Rousaki A, Botteon A, Colombo C, Conti C, Matousek P, Moens L, et al. Development of defocusing micro-SORS mapping: a study of a 19th century porcelain card. Anal Methods. 2017;9:6435–42.

    Article  CAS  Google Scholar 

  27. Lauwers D, Brondeel P, Moens L, Vandenabeele P. In situ Raman mapping of art objects. Phil Trans R Soc A. 2016;374:20160039.

    Article  Google Scholar 

  28. Wierzchos J, DiRuggiero J, Vítek P, Artieda O, Souza-Egipsy V, Škaloud P, et al. Adaptation strategies of endolithic chlorophototrophs to survive the hyperarid and extreme solar radiation environment of the Atacama Desert. Front Microbiol. 2015;6:934.

    Article  Google Scholar 

  29. Wierzchos J, Ascaso C, McKay CP. Endolithic cyanobacteria in halite rocks from the hyperarid core of the Atacama Desert. Astrobiology. 2006;6:415–22.

    Article  Google Scholar 

  30. Vítek P, Edwards HGM, Jehlička J, Ascaso C, De los Ríos A, Valea S, et al. Microbial colonization of halite from the hyper-arid Atacama Desert studied by Raman spectroscopy. Phil Trans R Soc A. 2010;368:3205–21.

    Article  Google Scholar 

  31. Vítek P, Jehlička J, Ascaso C, Mašek V, Gómez-Silva B, Olivares H, et al. Distribution of scytonemin in endolithic microbial communities from halite crusts in the hyperarid zone of the Atacama Desert, Chile. FEMS Microbiol Ecol. 2014;90:351–66.

    PubMed  Google Scholar 

  32. Vítek P, Jehlička J, Edwards HGM, Hutchinson I, Ascaso C, Wierzchos J. Miniaturized Raman system and the detection of traces of life in halite from the Atacama Desert: some considerations for the search for life signatures on Mars. Astrobiology. 2012;12:1095–9.

    Article  Google Scholar 

  33. Artieda O, Davila A, Wierzchos J, Buhler P, Rodríguez-Ochoa R, Pueyo J, et al. Surface evolution of salt-encrusted playas under extreme and continued dryness. Earth Surf Process Landf. 2015;40:1939–50.

    Article  Google Scholar 

  34. Robinson CK, Wierzchos J, Black C, Crits-Christoph A, Ma B, Ravel J, et al. Microbial diversity and the presence of algae in halite endolithic communities are correlated to atmospheric moisture in the hyper-arid zone of the Atacama Desert. Environ Microbiol. 2015;17:299–315.

    Article  Google Scholar 

  35. Lee E. Imaging modes. In: Zoubir A, editor. Raman imaging, techniques and applications. Springer series in optical sciences 168. Berlin: Springer; 2012. p. 1–37.

    Google Scholar 

  36. Prats-Mateu B, Gierlinger N. Tip in-light on: advantages, challenges, and applications of combining AFM and Raman microscopy on biological samples. Microsc Res Tech. 2017;80:30–40.

    Article  CAS  Google Scholar 

  37. Zhang X, Chen S, Ling Z, Zhou X, Ding D-Y, Kim YS, et al. Method for removing spectral contaminants to improve analysis of Raman imaging data. Sci Rep. 2016;7:39819.

    Google Scholar 

  38. Bonnier F, Mehmood A, Knief P, Meade AD, Hornebeck W, Lambkin H, et al. In vitro analysis of immersed human tissues by Raman microspectroscopy. J Raman Spectrosc. 2011;42:888–96.

    Article  CAS  Google Scholar 

  39. Nasdala L, Beyssac O, Schopf JW, Bleisteiner B. Application of Raman-based images in the Earth sciences. In: Zoubir A, editor. Raman imaging, techniques and applications. Springer series in optical sciences 168. Berlin: Springer; 2012. p. 145–87.

    Chapter  Google Scholar 

  40. Foucher F, Guimbretiére G, Bost N, Westall F. Petrographical and mineralogical applications of Raman mapping. In: Raman spectroscopy and applications, Chapter 8, Intech, 2017; pp. 163–180.

  41. Vítek P, Veselá B, Klem K. Spatial and temporal variability of plant responses cascade after PSII inhibition: Raman, chlorophyll fluorescence and infrared thermal imaging. Sensors. 2020;20:1015.

    Article  Google Scholar 

Download references

Acknowledgments

We wish to thank the anonymous reviewer for his/her thorough critical approach when reviewing the manuscript and valuable comments, which helped to improve and polish the present study.

Funding

This study was supported by the Czech Republic Ministry of Education, Youth and Sports under the Funding Programme INTER-COST, grant number LTC18036 (COST Action NEUBIAS, CA15124), and by grant PGC2018-094076-B-I00 from MCIU/AEI (Spain) and FEDER (UE). The work of P. Vítek on the development of the imaging possibilities of Raman spectroscopy was supported by SustES – Adaptation strategies for sustainable ecosystem services and food security under adverse environmental conditions (CZ.02.1.01/0.0/0.0/16_019/0000797).

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Authors and Affiliations

  1. Global Change Research Institute of the Czech Academy of Sciences, Bělidla 986/4a, 603 00, Brno, Czech Republic

    Petr Vítek

  2. Museo Nacional de Ciencias Naturales, CSIC, c/ Serrano 115 dpdo., 28006, Madrid, Spain

    Carmen Ascaso, M. Cristina Casero & Jacek Wierzchos

  3. Departamento Biología Vegetal, Ecología y Ciencias de la Tierra, and IACYS, Universidad de Extremadura, 10600, Plasencia, Spain

    Octavio Artieda

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  1. Petr Vítek
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  2. Carmen Ascaso
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  4. M. Cristina Casero
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  5. Jacek Wierzchos
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Correspondence to Petr Vítek.

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Vítek, P., Ascaso, C., Artieda, O. et al. Raman imaging of microbial colonization in rock—some analytical aspects. Anal Bioanal Chem 412, 3717–3726 (2020). https://doi.org/10.1007/s00216-020-02622-8

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  • DOI: https://doi.org/10.1007/s00216-020-02622-8

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