Abstract
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is routinely used as a rapid and cost-effective method for pathogen identification in clinical settings. In comparison, its performance in other microbiological fields, such as environmental microbiology, is still being tested, although isolates of environmental microbes are essential for in-depth in vivo studies of their biology, including biotechnological applications. We investigated the applicability of MALDI-TOF MS for the identification of bacterial isolates from a highly oligotrophic environment — Dinaric Karst caves, which likely harbor specific microorganisms. We cultured bacteria from the shell surface of the endemic mussel Congeria jalzici, one of the three known cave mussels in the world that lives in the Dinaric karst underground. The bacterial isolates were obtained by swabbing the shell surface of mussels living in microhabitats with different amounts of water: 10 air-exposed mussels, 10 submerged mussels, and 10 mussels in the hygropetric zone. A collection of 87 pure culture isolates was obtained, mostly belonging to the phylum Bacillota (72%), followed by Pseudomonadota (16%), Actinomycetota (11%), and Bacteroidota (1%). We compared the results of MALDI-TOF MS identification (Bruker databases DB-5989 and version 11, v11) with the results of 16S rDNA-based phylogenetic analysis, a standard procedure for bacterial identification. Identification to the genus level based on 16S rDNA was possible for all isolates and clearly outperformed the results from MALDI-TOF MS, although the updated MALDI-TOF MS database v11 gave better results than the DB-5989 version (85% versus 62%). However, identification to the species-level by 16S rDNA sequencing was achieved for only 17% of isolates, compared with 14% and 40% for the MALDI-TOF MS databases DB-5989 and v11 database, respectively. In conclusion, our results suggest that continued enrichment of MALDI-TOF MS libraries will result with this method soon becoming a rapid, accurate, and efficient tool for assessing the diversity of culturable bacteria from different environmental niches.
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Data Availability
All results obtained within this study are available in Supplementary Table S1, including the accession numbers of 16S rRNA gene sequences that are freely available in the GenBank database. MALDI-TOF MS spectra of bacterial colonies are available upon request.
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Acknowledgements
The authors are thankful to Sanja Puljas for participating in collecting of the samples; Lidija Buzuk, Lidija Šver, Nives Marčina, Karla Orlić, and Dunja Šikić for help with the laboratory work; and Prof. Rup Lal for advice on 16S rDNA phylogenetic analysis.
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Conceptualization: Ana Bielen and Sandra Hudina. Methodology: Ana Bielen, Snježana Kazazić, Ana Šimatović, and Nikolina Udiković-Kolić. Formal analysis and investigation: Ana Bielen, Ivana Babić, Marija Vuk Surjan, Snježana Kazazić, Ana Šimatović, and Zrinka Mesić. Writing—original draft preparation: Ana Bielen and Ivana Babić. Writing—review and editing: Ana Bielen, Ivana Babić, Jasna Lajtner, Nikolina Udiković-Kolić, Zrinka Mesić, and Sandra Hudina. Supervision: Ana Bielen.
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Bielen, A., Babić, I., Vuk Surjan, M. et al. Comparison of MALDI-TOF mass spectrometry and 16S rDNA sequencing for identification of environmental bacteria: a case study of cave mussel-associated culturable microorganisms. Environ Sci Pollut Res 31, 21752–21764 (2024). https://doi.org/10.1007/s11356-024-32537-1
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DOI: https://doi.org/10.1007/s11356-024-32537-1