Heavy metal tolerance in Stenotrophomonas maltophilia

Abstract

Stenotrophomonas maltophilia is an aerobic, non-fermentative Gram-negative bacterium widespread in the environment. S. maltophilia Sm777 exhibits innate resistance to multiple antimicrobial agents. Furthermore, this bacterium tolerates high levels (0.1 to 50 mM) of various toxic metals, such as Cd, Pb, Co, Zn, Hg, Ag, selenite, tellurite and uranyl. S. maltophilia Sm777 was able to grow in the presence of 50 mM selenite and 25 mM tellurite and to reduce them to elemental selenium (Se(0)) and tellurium (Te(0)) respectively. Transmission electron microscopy and energy dispersive X-ray analysis showed cytoplasmic nanometer-sized electron-dense Se(0) granules and Te(0) crystals. Moreover, this bacterium can withstand up to 2 mM CdCl(2) and accumulate this metal up to 4% of its biomass. The analysis of soluble thiols in response to ten different metals showed eightfold increase of the intracellular pool of cysteine only in response to cadmium. Measurements by Cd K-edge EXAFS spectroscopy indicated the formation of Cd-S clusters in strain Sm777. Cysteine is likely to be involved in Cd tolerance and in CdS-clusters formation. Our data suggest that besides high tolerance to antibiotics by efflux mechanisms, S. maltophilia Sm777 has developed at least two different mechanisms to overcome metal toxicity, reduction of oxyanions to non-toxic elemental ions and detoxification of Cd into CdS.

Figure 1. ESEM-EDX and TEM-EDX observations.

Microscopic observations and representative energy-dispersive X-ray spectra of electron-dense particles of S. maltophilia Sm777 cells grown in ten fold-diluted TSB medium solidified with 15 g.l⁻¹ agar, and supplemented with metals. (A & B) Colony shape, TEM-EDX micrographs and spectra of S. maltophilia Sm777 grown in the presence of selenite (10 mM) and tellurite (1 mM). (C) Colony shape, ESEM-EDX observation and analysis of cells grown in the presence of CdCl₂ (500 µM). Arrows on micrographs indicate the presence of intracellularly localized electron-dense particles of Se and Te, and arrows on spectra indicate metal-specific peak detected.

Figure 2. Soluble thiols analysis.

HPLC analysis of nonprotein thiols in S. maltophilia Sm777 grown in TSB/10 without (A) or supplemented (B) with 500 µM of CdCl₂. The arrow indicates cysteine peack. N-acetyl-L-cysteine (NAC) was used as an internal standard.

Figure 3. Extended X-ray Absorption Fine Structure (EXAFS) Spectroscopy.

(A) Comparison between experimental and calculated EXAFS of S. maltophilia Sm777 strain cells. (B) Modeling of Cd dimers and trimers.