Microbial thiocyanate utilization under highly alkaline conditions

Three kinds of alkaliphilic bacteria able to utilize thiocyanate (CNS-) at pH 10 were found in highly alkaline soda lake sediments and soda soils. The first group included obligate heterotrophs that utilized thiocyanate as a nitrogen source while growing at pH 10 with acetate as carbon and energy so urces. Most of the heterotrophic strains were able to oxidize sulfide and t hiosulfate to tetrathionate. The second group included obligately autotroph ic sulfur-oxidizing alkaliphiles which utilized thiocyanate nitrogen during growth with thiosulfate as the energy source. Genetic analysis demonstrate d that both the heterotrophic and autotrophic alkaliphiles that utilized th iocyanate as a nitrogen source were related to the previously described sul fur-oxidizing alkaliphiles belonging to the gamma subdivision of the divisi on Proteobacteria (the Halomonas group for the heterotrophs and the genus T hioalkalivibrio for autotrophs). The third group included obligately autotr ophic sulfur-oxidizing alkaliphilic bacteria able to utilize thiocyanate as a sole source of energy. These bacteria could be enriched on mineral mediu m with thiocyanate at pH 10. Growth with thiocyanate was usually much slowe r than growth with thiosulfate, although the biomass yield on thiocyanate w as higher. Of the four strains isolated, the three vibrio-shaped strains we re genetically closely related to the previously described sulfur-oxidizing alkaliphiles belonging to the genus Thioalkalivibrio, The rod-shaped isola te differed from the other isolates by its ability to accumulate large amou nts of elemental sulfur inside its cells and by its ability to oxidize carb on disulfide. Despite its low DNA homology with and substantial phenotypic differences from the vibrio-shaped strains, this isolate also belonged to t he genus Thioalkalivibrio according to a phylogenetic analysis. The heterot rophic and autotrophic alkaliphiles that grew with thiocyanate as an N sour ce possessed a relatively high level of cyanase activity which converted cy anate (CNO-) to ammonia and CO2. On the other hand, cyanase activity either was absent or was present at very low levels in the autotrophic strains gr own On thiocyanate as the sole energy and N source. As a result, large amou nts of cyanate were found to accumulate in the media during utilization of thiocyanate at pH 10 in batch and thiocyanate-limited continuous cultures. This is a first direct proof of a "cyanate pathway" in pure cultures of thi ocyanate-degrading bacteria. Since it is relatively stable under alkaline c onditions, cyanate is likely to play a role as an N buffer that keeps the a lkaliphilic bacteria safe from inhibition by free ammonia, which otherwise would reach toxic levels during dissimilatory degradation of thiocyanate.