By David N. Leff

What subversive forces are dumping toxins into groundwater and poisoning wells all over America?

The answer, to paraphrase Pogo, is: "We have met that enemy, and it is us."

For decades, dry cleaners poured their dirtied trichloroethylene down the drain -- and out into the water table. Gas-station operators used the highly toxic chlorinated hydrocarbons for cleaning off brake pads, and didn't care where they dumped the residue. In industry, these highly toxic solvents degrease machinery and electronic parts and serve as paint thinners and removers, antifreeze and refrigerants. Farmers apply them as insecticides and weed-killers, often ignoring where the run-off ends up.

Among the myriad of hazardous waste sites targeted by the Environmental Protection Agency for clean-up are more than 7,000 military bases, drenched in the grimly stable solvents and jet fuels.

From biotechnology's beginnings, 20 years ago, "bioremediation" counted on training and turning loose genetically engineered pit-bull bacteria to chew up and degrade these hazardous chemicals. Molecular bioremediators are still counting.

The June 6 Science reports "Isolation of a bacterium that reductively dechlorinates tetrachloroethene [same as perchloroethylene = PCE] to ethene." Its senior author, Cornell University microbiologist Stephen Zinder, told BioWorld Today: "That was the first bacterial culture ever isolated that could take PCE all the way to ethylene."

That is, the novel bacterium yanked all four of PCE's chlorine-atom teeth, reducing it from CHCl 2 CHCl 2 to zero-chlorine ethylene (H2C:CH2). That colorless gas is the third highest-volume chemical produced in the U.S. Besides the plastics industry, antifreeze, refrigerants and anesthetics, ethylene (a.k.a. ethene) helps put the blush of ripeness on fruits.

"Ethylene is not basically toxic to humans or animals," Zinder noted, "so that complete conversion from PCE was sort of big."

All previous bioremediators, Zinder continued, never took the degradation any further than to vinyl chloride (CH2:CHCl). Anent the partial transformation, he quotes the paper's co-senior author, environmental engineer James Gossett, who observed: "They took a suspected human carcinogen, PCE, and made a known human carcinogen out of it -- vinyl chloride."

Zinder and Gossett, who are on the faculty at Cornell, in Ithaca, N.Y., are joint discoverers and developers of the chlorine-devouring bacterium, which bears no name but "Strain 195." It surfaced 12 years ago in sludge from the city of Ithaca's old sewage treatment plant. "That mixed bacterial culture," Zinder observed, was "a microbe-rich relic from the bad old days, when dry-cleaning solvents and industrial degreasing agents were flushed down the drain."

Grooming 195 From Trace Agent To Top Degrader

He recalled: "In those first crude cultures, we could see trace amounts of PCE being biodegraded at a very slow rate. The next step was to get the culture to the point where the metabolism of PCE was a large portion of the whole culture's metabolism, involving much larger amounts of PCE relative to the other contaminants."

Gossett's lab, Zinder recounted, "increased the PCE dosage added to the culture, squirting it in every other day." With years of such tender loving care, Strain 195 began to grow by itself. Over the decade following its discovery in 1985, it battened on its chlorine-rich diet of PCE to become, numerically, the dominant organism in the Ithaca culture. The researchers showed that the source of reductive dechlorination electrons fueling 195's chlorine-only appetite was hydrogen.

One outstanding puzzler that Zinder and Gossett have put on a far-back burner is: What did Strain 195 live on before chlorinated solvents were invented?

Zinder's lab more recently made the discovery that potent antibiotics, such as penicillin and vancomycin, bounced harmlessly off the mystery microbe's coccoid (spherical) cell wall while decimating other organisms in the mix. Among 195's other unique features was its minute size -- 0.5 microns in diameter -- half that of other coccoids, such as Staphylococcus or Streptococcus.

But superficial resemblances gave no clue to the anaerobic, lone-ranger bug's place in the bacterial family tree. "I thought it looked like a member of the archaea," Zinder observed, "but it's not in them. It's clearly in the eubacteria, the normal bacteria, but related to no known genus or species. Strain 195 occupies a fairly deep branch of the tree all by itself."

Comparative taxonomy doesn't work, he pointed out, "because there's not much we can compare it to, as 195 grows only on these chlorinated ethylenes. And using molecular sequencing, it didn't seem to cluster with any known organism." So far, the group hasn't isolated any genes from Strain 195, except for one 16S RNA sequence.

"It's a difficult organism to handle, in terms of the standard genetic techniques," Zinder pointed out. "As of now, anything we might do in that direction is going to be very exploratory."

Polluting Binge Seems To Be Subsiding

Recently, the Cornell microbiologists looked in on Ithaca's new, modern sewage plant. "Knowing what we know now," Zinder recalled, "we expected to get the toxic-waste degrader out really quickly. In fact, we couldn't find any there. People are apparently not dumping the stuff down the drains any more. They're much more careful about it, which I think is true of everybody these days."

The Cornell team has a U.S. Air Force contract to help plan the clean-up of two polluted air fields. One is a deactivated B-52 base in Plattsburgh, N.Y.; the other, a still-operational facility at Fallon, Nev., where the U.S. Navy trains fighter pilots.

"In Plattsburgh," Zinder said, "there's good evidence that these chlorinated pollutants are going away on their own, by natural attenuation -- intrinsic bioremediation.

"In sites like Plattsburgh Air Force Base," he explained, crash-and-rescue personnel used to pour expired jet fuel into pits partly full of discarded trichlor, then practiced putting the fires out. Which seemed like a good idea at the time. The quicker they quenched the fires, the more solvent was left over and washed down into the water table."

He continued: "Now these sites are full of wells that people have drilled to monitor the cultures. When you look down in there you see reduction products -- dichloroethylene, vinyl chloride, ethene -- indicating that the actual biodegradation process is occurring in situ at the site.

"That in general is where the bioremediation field is moving today," Zinder said. "Now there's interest in monitoring sites to see that the solvents are going away on their own, or maybe accelerating the process by adding nutrients with sources of reducing power. This is called accelerated intrinsic bioremediation in situ."

Injecting cultures of Strain 195 into the process, Zinder suggested, "is something that may happen in Fallon, because we're doing a pilot study there. We're going to be adding organic compounds. They have actually set up raceways for dosing with several different ones. At some point," he observed, "if we don't feel the dechlorination is going well down there, we might consider adding the Strain 195 organism." *