Obligate sulfide-dependent degradation of methoxylated aromatic compounds and formation of methanethiol and dimethyl sulfide by a freshwater sediment isolate, Parasporobacterium paucivorans gen. nov., sp. nov

Abstract

Methanethiol (MT) and dimethyl sulfide (DMS) have been shown to be the dominant volatile organic sulfur compounds in freshwater sediments. Previous research demonstrated that in these habitats MT and DMS are derived mainly from the methylation of sulfide. In order to identify the microorganisms that are responsible for this type of MT and DMS formation, several sulfide-rich freshwater sediments were amended with two potential methyl group-donating compounds, syringate and 3,4,5-trimethoxybenzoate (0.5 mM). The addition of these methoxylated aromatic compounds resulted in excess accumulation of MT and DMS in all sediment slurries even though methanogenic consumption of MT and DMS occurred. From one of the sediment slurries tested, a novel anaerobic bacterium was isolated with syringate as the sole carbon source. The strain, designated Parasporobacterium paucivorans, produced MT and DMS from the methoxy groups of syringate. The hydroxylated aromatic residue (gallate) was converted to acetate and butyrate. Like Sporobacterium olearium, another methoxylated aromatic compound-degrading bacterium, the isolate is a member of the XIVa cluster of the low-GC-content Clostridiales group. However, the new isolate differs from all other known methoxylated aromatic compound-degrading bacteria because it was able to degrade syringate in significant amounts only in the presence of sulfide.

FIG. 1

Scanning electron micrograph of a logarithmic-phase culture of strain SYR1 grown on syringate (5 mM). The photograph clearly shows the double-rod morphology of strain SYR1.

FIG. 2

Time courses of growth of strain SYR1 on syringate and sulfide. Symbols: ▴, syringate; ▪, MT; ♦, DMS; ●, acetate; ×, optical density at 660 nm (OD₆₆₀).

FIG. 3

Time courses of volatile sulfur compounds (VSC) (A) and syringate and volatile fatty acids (VFA) (B) of a sulfide-rich culture of strain SYR1 and time courses of syringate and VOSC of a sulfide-free culture of strain SYR1 (C). Arrows indicate pulsewise additions of sodium sulfide. Symbols: ▪, sulfide; ♦, MT; ▴, DMS; □, syringate; ⋄, acetate; ▵, butyrate.

FIG. 4

Time courses of cell suspensions incubated under sulfide-rich (closed symbols) and sulfide-free (open symbols) conditions. Triangles, syringate; diamonds, methyl groups (MT plus DMS).

FIG. 5

Phylogenetic tree based on a distance matrix prepared from an alignment of a partial 16S rRNA sequences (1,478 bp) of P. paucivorans and other selected closely related bacteria. Bootstrap values indicate the percentage of occurrence of 100 bootstrap trees. Only values above 80 are given. Reference sequences were from the GenBank and EMBL databases: X95624, Ruminococcus hydrogenotrophicus; X94966, Ruminococcus productus; X85101, Ruminococcus obeum; AF116854, Sporobacterium olearium; Z49863, Sporobacter termitidis; X77215, Holophaga foetida; X77216, Pelobacter acidigallici; X96954, Acetobacterium woodii; X71858, Clostridium polysaccharolyticum; X71853, Clostridium populeti; AF028351, Clostridium indolis; X71855, Clostridium xylanolyticum; AF067965, Clostridium methoxybenzovorans; X73449, Clostridium sphenoides; X71848, Clostridium celerecrescens; AF028349, Clostridium fusiformis; AF202259, Eubacterium oxidoreducens. Sporobacter termitidis was used as the outgroup. Bar, 10 base substitutions per 100 bases. Names of MT- or DMS-producing microorganisms are underlined.