Atrazine [2-chloro-4-(ethylamino)-6-(isopropylamino)-1,3,5-triazine)] is a widely used s-triazine (i.e., symmetric triazine) herbicide for the control of broad-leaf weeds. Approximately 800 million pounds were used in the United States between 1980 and 1990. As a result of this widespread use, for both selective and nonselective weed control, atrazine and other s-triazine derivatives have been detected in ground and surface water in several countries.
Numerous studies on the environmental fate of atrazine have shown that atrazine is a recalcitrant compound that is transformed to CO.sub.2 very slowly, if at all, under aerobic or anaerobic conditions. It has a water solubility of 33 mg/l at 27.degree. C. Its half-life (i.e., time required for half of the original concentration to dissipate) can vary from about 4 weeks to 57 weeks if in soils at low concentration (i.e., less than about 2 parts per million (ppm)). High concentrations of atrazine, such as those occurring in spill sites, have been reported to dissipate even more slowly.
As a result of its widespread use, atrazine is often detected in ground water and soils in concentrations exceeding the maximum contaminant level (MCL) of 3 .mu.g/l (i.e., 3 parts per billion (ppb)), a regulatory level that took effect in 1992. Point source spills of atrazine have resulted in levels as high as 25 ppb in some wells. Levels of up to 40,000 mg/l (i.e., 40,000 parts per million (ppm)) atrazine have been found in the soil of spill sites more than ten years after the spill incident. Such point source spills and subsequent runoff can cause crop damage and ground water contamination.
There have been numerous reports of the isolation of s-triazine-degrading microorganisms (see, e.g., Behki et al., J. Agric. Food Chem., 34, 746-749 (1986); Behki et al., Appl. Environ. Microbiol., 59, 1955-1959 (1993); Cook, FEMS Microbiol. Rev., 46, 93-116 (1987); Cook et al., J. Agric. Food Chem., 29, 1135-1143 (1981); Erickson et al., Critical Rev. Environ. Cont., 19, 1-13 (1989); Giardina et al., Agric. Biol. Chem., 44, 2067-2072 (1980); Jessee et al., Appl. Environ. Microbiol., 45, 97-102 (1983); Mandelbaum et al., Appl. Environ. Microbiol., 61, 1451-1457 (1995); Mandelbaum et al., Appl. Environ. Microbiol., 59, 1695-1701 (1993); Mandelbaum et al., Environ. Sci. Technol., 27, 1943-1946 (1993); Radosevich et al., Appl. Environ. Microbiol., 61, 297-302 (1995); and Yanze-Kontchou et al., Appl. Environ. Microbiol., 60, 4297-4302 (1994)). Many of the organisms described, however, failed to mineralize atrazine (see, e.g., Cook, FEMS Microbiol. Rev., 46, 93-116 (1987); and Cook et al., J. Agric. Food Chem., 29, 1135-1143 (1981)). While earlier studies have reported atrazine degradation only by mixed microbial consortia, more recent reports have indicated that several isolated bacterial strains can degrade atrazine. For example, we previously reported the isolation of a pure bacterial culture, identified as Pseudomonas sp. strain ADP (Mandelbaum et al., Appl. Environ. Microbiol., 61, 1451-1457 (1995); Mandelbaum et al., Appl. Environ. Microbiol., 59, 1695-1701 (1993); and Mandelbaum et al., Environ. Sci. Technol., 27, 1943-1946 (1993)), which degraded a high concentration of atrazine (&gt;1,000 .mu.g/ml) under growth and non-growth conditions. See also, Radosevich et al., Appl. Environ. Microbiol., 61, 297-302 (1995) and Yanze-Kontchou et al., Appl. Environ. Microbiol., 60, 4297-4302 (1994). Pseudomonas sp. strain ADP (Atrazine Degrading Pseudomonas) uses atrazine as a sole source of nitrogen for growth. The organism completely mineralizes the s-triazine ring of atrazine under aerobic growth conditions. That is, this bacteria is capable of degrading the s-triazine ring and mineralizing organic intermediates to inorganic compounds and ions (e.g., CO.sub.2).
Little information is available concerning the genes and enzymes involved in the metabolism of s-triazine compounds. Although genes that encode the enzymes for melamine (2,4,6-triamino-s-triazine) metabolism have been isolated from a Pseudomonas sp. strain, and that encode atrazine degradation activity from Rhodococcus sp. strains, to date there have been no reports identifying the genes encoding atrazine dechlorination.