Characterization of denitrifying activity by the alphaproteobacterium, Sphingomonas wittichii RW1

Sphingomonas wittichii RW1 has no reported denitrifying activity yet encodes nitrite and nitric oxide reductases. The aims of this study were to determine conditions under which S. wittichii RW1 consumes nitrite (NO(−)(2)) and produces nitrous oxide (N(2)O), examine expression of putative genes for N-oxide metabolism, and determine the functionality of chromosomal (ch) and plasmid (p) encoded quinol-dependent nitric oxide reductases (NorZ). Batch cultures of wildtype (WT) and a norZ(ch) mutant of S. wittichii RW1 consumed NO(−)(2) and produced N(2)O during stationary phase. The norZ(ch) mutant produced N(2)O, although at significantly lower levels (c.a. 66–87%) relative to the WT. Rates of N(2)O production were 2–3 times higher in cultures initiated at low relative to atmospheric O(2) per unit biomass, although rates of NO(−)(2) consumption were elevated in cultures initiated with atmospheric O(2) and 1 mM NaNO(2). Levels of mRNA encoding nitrite reductase (nirK), plasmid-encoded nitric oxide dioxygenase (hmp(p)) and plasmid-encoded nitric oxide reductase (norZ(p)) were significantly higher in the norZ(ch) mutant over a growth curve relative to WT. The presence of NO(−)(2) further increased levels of nirK and hmp(p) mRNA in both the WT and norZ(ch) mutant; levels of norZ(p) mRNA compensated for the loss of norZ(ch) expression in the norZ(ch) mutant. Together, the results suggest that S. wittichii RW1 denitrifies NO(−)(2) to N(2)O and expresses gene products predicted to detoxify N-oxides. So far, only S. wittichii strains within four closely related taxa have been observed to encode both nirK and norZ genes, indicating a species-specific lateral gene transfer that may be relevant to the niche preference of S. wittichii.