Ubiquity and diversity of dissimilatory (per)chlorate-reducing bacteria

Environmental contamination with compounds containing oxyanions of chlorine , such as perchlorate or chlorate [(per)chlorate] or chlorine dioxide, has been a constantly growing problem over the last 100 years. Although the fac t that microbes reduce these compounds has been recognized for more than 50 gears, only six organisms which can obtain energy for growth by this metab olic process have been described, As part of a study to investigate the div ersity and ubiquity of microorganisms involved in the microbial reduction o f (per)chlorate, we enumerated the (per)chlorate-reducing bacteria (ClRB) i n very diverse environments, including pristine and hydrocarbon-contaminate d soils, aquatic sediments, paper mill waste sludges, and farm animal waste lagoons. In all of the environments tested, the acetate-oxidizing ClRB rep resented a significant population, whose size ranged from 2.31 x 10(3) to 2 .4 x 10(6) cells per g of sample. In addition, we isolated 13 ClRB from the se environments. All of these organisms could grow anaerobically by couplin g complete oxidation of acetate to reduction of (per) chlorate. Chloride wa s the sole end product of this reductive metabolism. All of the isolates co uld also use oxygen as a sole electron acceptor, and most, but not all, cou ld use nitrate. The alternative electron donors included simple volatile fa tty acids, such as propionate, butyrate, or valerate, as well as simple org anic acids, such as lactate or pyruvate, Oxidized-minus-reduced difference spectra of washed whole-cell suspensions of the isolates had absorbance max ims close to 425, 525, and 550 nm, which are characteristic of type c cytoc hromes. In addition, washed cell suspensions of all of the ClRB isolates co uld dismutate chlorite, an intermediate in the reductive metabolism of (per )chlorate, into chloride and molecular oxygen. Chlorite dismutation was a r esult of the activity of a single enzyme which in pure form had a specific activity of approximately 1,928 mu mol of chlorite per mg of protein per mi n. Analyses of the 16S ribosomal DNA sequences of the organisms indicated t hat they all belonged to the alpha, beta, or gamma subclass of the Proteoba cteria, Several were closely related to members of previously described gen era that are not recognized for the ability to reduce (per)chlorate, such a s the genera Pseudomonas and Azospirllum. However, many were not closely re lated to any previously described organism and represented new genera withi n the Proteobacteria. The results of this study significantly increase the limited number of microbial isolates that are known to be capable of dissim ilatory (per)chlorate reduction and demonstrate the hitherto unrecognized p hylogenetic diversity and ubiquity of the microorganisms that exhibit this type of metabolism.