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Bacillus amyloliquefaciens strains isolated from moisture-damaged buildings produced surfactin and a substance toxic to mammalian cells

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Abstract

Fungicidic Bacillus amyloliquefaciens strains isolated from the indoor environment of moisture-damaged buildings contained heat-stable, methanol-soluble substances that inhibited motility of boar spermatozoa within 15 min of exposure and killed feline lung cells in high dilution in 1 day. Boar sperm cells lost motility, cellular ATP, and NADH upon contact to the bacterial extract (0.2 μg dry wt/ml). Two bioactive substances were purified from biomass of the fungicidal isolates. One partially characterized substance, 1,197 Da, was moderately hydrophobic and contained leucine, proline, serine, aspartic acid, glutamic acid and tyrosine, in addition to chromophore(s) absorbing at 365 nm. In boar sperm and human neural cells (Paju), the compound depolarized the transmembrane potentials of mitochondria (ΔΨ m) and the plasma membrane (ΔΨ p) after a 20-min exposure and formed cation-selective channels in lipid membranes, with a selectivity K+:Na+:Ca2+ of 26:15:3.5. The other substance was identified as a plasma-membrane-damaging lipopeptide surfactin. Plate-grown biomass of indoor Bacillus amyloliquefaciens contained ca. 7% of dry weight of the two substances, 1,197 Da and surfactin, in a ratio of 1:6 (w:w). The in vitro observed simultaneous collapse of both cytosolic and mitochondrial ATP in the affected mammalian cell, induced by the 1,197-Da cation channel, suggests potential health risks for occupants of buildings contaminated with such toxins.

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Abbreviations

RP-HPLC:

Reversed-phase high-performance liquid chromatography

BLM:

Black lipid membrane

DAD:

Diode-array detector

ΔΨ m :

Mitochondrial membrane potential

ΔΨ p :

Plasma membrane potential

JC-1:

5,5′,6,6′-Tetrachloro-1,1′,3,3′-tetraethylbenz-imidazolo carbocyanine iodide

AM:

Calcein acetoxymethyl ester

PI:

Propidium iodide

MALDI-TOF-MS:

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry

ESI-IT-MS:

Electrospray ionization ion trap mass spectrometry

EC50 :

Endpoint concentration which caused ≥50% change in the viability parameters

FCCP:

Carbonyl cyanide 4-trifluoromethoxyphenylhydrazone

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Acknowledgments

The study was supported by grants from Academy of Finland grant 50733, the grant for Center of Excellence “Microbial Resources,” Magnus Ehrnrooth Foundation and the Finnish Medical Society. We would like to thank Leif C. Andersson for collaboration, Anu Harju for growing Paju cells, the Viikki Science Library for excellent information services, Hannele Tukiainen, Leena Steininger and Tuula Suortti for many kinds of help.

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Authors and Affiliations

  1. Department of Applied Chemistry and Microbiology, University of Helsinki, P.O. Box 56 (Biocenter), Helsinki, 00014, Finland

    Raimo Mikkola, Maria A. Andersson, Pavel Grigoriev, Vera V. Teplova, Nils-Erik L. Saris & Mirja S. Salkinoja-Salonen

  2. Institute of Cell Biophysics, Russian Academy of Sciences, Pushchino, Moscow Region, 142290, Russia

    Pavel Grigoriev

  3. Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, Pushchino, Moscow Region, 142290, Russia

    Vera V. Teplova

  4. Department of Biological Sciences, Louisiana State University, Baton Rouge, LA, USA

    Frederick A. Rainey

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  1. Raimo Mikkola
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Correspondence to Raimo Mikkola.

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Mikkola, R., Andersson, M.A., Grigoriev, P. et al. Bacillus amyloliquefaciens strains isolated from moisture-damaged buildings produced surfactin and a substance toxic to mammalian cells. Arch Microbiol 181, 314–323 (2004). https://doi.org/10.1007/s00203-004-0660-x

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