Rea nulyzing health effects of the Gulf War hen the troops who served in operations Desert Shield , and Desert Storm in 1991 returned 113m the Persian Gulf, some of them showed signs of unusual illnesses. Fatigue, debilitating pain, blurred vision, acute hair loss in unusual patterns, unexplained fevers, recurrent rashes, and dizziness were among the symptoms they reported. They blamed these illnesses on agents they might have been exposed to during their tours of duty. A congressional hearing was held in September 1992 to examine possible causes for what was quickly, if unofficially, becoming known as “Persian Gulf War Syndrome.” Some potential causes included petroleum exposures from the oil well fires in Kuwait and from heavy routine pesticide and petroleum use in the base camps, post-traumatic stress disorder (PTSD), parasitic infections, and adverse reactions from the drugs the soldiers were required to take to guard against possible poisoning by nerve or mustard gas. After the 1992 hearing, Congress voted to support the creation of a health registry for Gulf War veterans that was modeled after the Agent Orange registry set up for Vietnam vets. TWO years later, however, no one disease or chemical agent has yet emerged as the clear cause of any of these illnesses, many of which have persisted. Is there any objective way to find out what happened to these veterans? In an attempt to answer this question, the Department of Defense (DOD), the Department of Veterans Affairs, the Environmental Protection Agency (EPA), and
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2 =)
Many Veterans believe the PeT’Sian Gulf War ’el them a legacy Of Unusual health problems. Examining the possible Causes has brought U p new challengesfor analytical chemists
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the National Institutes of Health (NIH) convened a Technology Assessment Workshop in April to hear testimony on the current state of investigations on Persian Gulf War Syndrome. Speakers from various agencies and universities presented overviews of illnesses reported during and after the Gulf War, the evidence of chemical and disease exposures, and current aspects of diagnosing or defining the possible syndrome($. The workshop panelists, who were neither military personnel nor federal agency researchers, were supposed to determine whether evidence existed for a distinct syndrome resulting from service in the war. If they found confirmation of a syndrome, their goal was to ascertain the symptoms that define it, the plausible causes and their biological effects, and the direction of future research. Much of what the speakers presented was similar to what had been reported at the congressional hearing in 1992. However, some of the new details that surfaced-particularly in the discussions of a novel viscerotropic form of leishmaniasis that was detected, the troops’ daily routine exposures, and the alarms for chemical warfare agents that went off during the war-reopened the possibility of acquiring hard analytical data to support or reject these events as factors in the veterans’ illnesses. Panel recommendations
After hearing a day and a half of testimony, panelists deliberated in closed sessions and presented a draft statement of their findings. The panel concluded, “No single disease or syndrome is apparent but rather multiple illnesses with overlapping symptoms and causes.” One of the key problems that emerged from the workshop, according to the statement presented by the panel, was that “Few data are available regarding the troops who served in the Persian Gulf and their exposures during that period of service . . . . Data collection that was carried out was initiated only after a considerable delay.” Because of the lack of hard evidence for the incidence of symptoms in veterans relative to a nonexposed group, the panel declined to put out a working case definition, but they specified viscerotropic
the firefighters-some of whom were monitored during service in 1991-with the same kinds of debilitating symptoms the ill veterans reported. “We were more impressed by the report of the environmental conditions in the camps-the tent heaters that ran on diesel or leaded fuel, the fuel the troops sprayed on the roads to keep down the dust, the burning of human waste, and so on,” says Green. In some areas, the troops used up to 30,000 gallons a day of diesel fuel just to Leishmaniasis Viscerotropic leishmaniasis, a rare and po- spray down the roads-fuel was extremely cheap and plentiful in Saudi Arabia, and tentially deadly form of the normally cutawater was not. In addition, much of the neous disease, was confirmed by parasite identification in 31 veterans. In this form of military’s water supply for showers and other nondrinking uses was salt water leishmaniasis, the first physical signs that had been desalinated by reverse osmay be only vague flu-like symptoms that mosis and chlorinated. appear long after infection. The parasite The water also contained some benhides in the tissues, so the usual blood, zene. Exposure tests for people taking bone marrow, and skin tests may not showers in benzene-contaminated water pick it up. “The underlying prevalence is likely to be substantially higher than re- have demonstrated that blood levels of benzene rise 60%after a 20-min,hot shower ported,” the panel stated. The panel with water contaminated at the part-perurged the agencies to develop new assay methods based on immune response to billion level because of skin absorption and vapor inhalation. The NIH workshop isolated leishmaniasis antigens; cytokine profile analyses; and amplification of para- panel stated that this type of retrospective occupational exposure study, in which site DNA markers, which may be present environmental conditions in the camps in very low numbers (as few as 5-10 cop are simulated and measured to model exies) in bone marrow. posures, could prove useful in conjunction with patient questionnaires to estimate inPetroleum exposures dividuals’ chronic exposures to petroGreen says that despite the drama of the oil well fires, this source of exposure to chemicals, pesticides, spray paint, and other harmful substances. petrochemicals did not appear to affect
leishmaniasis and PTSD as two conditions that could be attributed to service in the Gulf War region. “We’re not saying that they were the probable causes of most of the veterans’ symptoms,” explains panel chairperson Gareth Green of Harvard University. “It’s just that these two are treatable now, and we don’t want to limit the veterans’ ability to be examined and treated for them in the meantime.”
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The IMS-based “chemical agent monitor.”
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Table 1. Chemical warfare agent detectors used in the Persian Gulf War Detector
Analyte(s)
Method
M8A1 alarm
Tabun, Sarin, Soman, VX (nerve agents)
ionization
CAM
Tabun, Sarin, VX, sulfur mustard, nitrogen mustard
ion mobility spectrometry
Sensitivity (mg/m3 air)
0.1-0.2
Response time
c 2 min
I -
< 0.1
The expected nerve agents, with a structure similar to that of organophosphate pesticides, work by inhibiting chopossibilities presented at the workshop was that the troops were exposed to chem- linesterase activity. The nerve agent filter disk was impregnated with acetylchoical warfare agents such as nerve and mustard gases, contrary to all the US. mil- linesterase, and indoxyl acetate was used as the color indicator. Kirkwood says that itary reports that were made public up in addition to the M256A1 cholinesterase through November 1993. Two levels of test, similar assay “tickets” produced in chemical monitoring were used by the France were obtained and gave identical U.S. troops. Although the “M8A1” ionization alarms set out at the perimeters of negative results when used in the field. each military base went off frequently For the blister agent (e.g., mustard) during the war, none of the colorimetric test, an ampule of 4-(4-nitrobenzyl)assay-based “M256A1” detection kits were pyridine with mercuric cyanide catalyst reported to have confirmed the alarms. was emptied onto a silica gel filter disk and exposed to the air. K,CO, was added. If an alarm sounded, the troops put on Mustard vapor turned the disk bluish protective gear and an M256A1 detection kit was placed at the alarm site by a desig- purple; phosgene oxime turned it reddish purple. According to Robert Eckhaus, a nated chemical officer, says Charles Kirkchemical engineer with the Army Chemiwood of the Army’s Chemical School at Ft. McClellan, AL. The detection kits con- cal and Biological Defense Command, the test for mustard agents has been used for tained disposable cards, each with four filter paper-based colorimetric spot tests to more than 40 years in one form or andetect chemical warfare agents and difother. A separate test for Lewisite used a ferentiate them as nerve gases, blister solid tablet of Michler’s thioketone reagents, Lewisite (a specific blister agent), agent, which was rubbed on a card. Afor blood agents (see Table 1). The cards ter 10 min, the tan mark from the tablet turned green if Lewisite was present. were exposed to the air for 10 min, then ampules of appropriate reagent mixTo test for the presence of blood-active tures were added to each filter disk for agents such as HCN and CNCl, ampules color development. of barbituric acid and sodium hypochlo-
Chemical warfare agents One of the most intriguing, if unlikely,
e 1 min
rite (bleach) were opened onto the blood agent test filter. After exposure to the air, CNCl would react with the acid to turn the filter pink or blue depending on the concentration. HCN was detected as CNCl after reaction with bleach. The United States also issued a “chemical agent monitor’’ (CAM) based on ion mobility spectrometry for detecting nerve and blister agents (see photo). Designed for military use by a British firm as an offshoot of an explosives detector project, the CAM had a small radioactive ionization source and used an electric field to accelerate molecular ion clusters down a flight tube. The time-of-flight analog peaks were converted and displayed on an eight-bar LCD; the number of bars indicated the concentration of selected compounds. In addition, 60 mobile laboratories drove ahead of the military units to determine the presence or absence of nuclear, biological, and chemical warfare agents in each new region. They contained the M8A1, M256A1, and CAM detectors and were also equipped with quadrupole mass spectrometers operating in selected ion monitoring (SIM) mode (referred to as MM-1 detectors by the military) to monitor for nerve, blister, and blood agents.
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The Czech report
During the war, no detections of chemical warfare agents were reported by U.S. troops. Last November, however, the Army released a statement acknowledging the validity of a Czech report, made during the Gulf War, that that country’s chemical warfare agent monitors had detected the presence of a nerve agent on Jan. 19,1991, near Hafr a1 Batin. Also, a liquid mustard agent was identified in a wet patch of sand outside King Khaled Military City on Jan. 24,1991. At the NIH meeting, Joseph Corriveau of the U.S. Army Foreign Science and Technology Center said the Army investigative team that went to the Czech Republic last October to review the evidence found the Czech report “credible,” although no air or sand samples had been kept for independent verification. The Czech detectors were an automatic chemical alarm and a semi-automatic chemical detector run by a mobile lab. Both detectors were based on cholinesterase activity, said Corriveau. The automated air monitor consisted of a reel of paper tape that was wetted a little at a time with solutions of cholinesterase and phenol red substrate and exposed to the air. If the air was clean, the tape turned yellow, but if a cholinesterase inhibitor was present, the tape remained red. An optical detector tracked the reaction over time. Interferences included organophosphate pesticides. The semi-automaticdetector worked on the same enzyme activity principle as the automatic alarm, said Corriveau, but it also performed several chemical spot tests to detect structural features such as organophosphate, tertiary amine, fluorine, and isopropyl groups. On Jan. 19,1991, the Czech alarms detected a nerve agent twice in 40 min at levels of 0.05 mg/m‘-too low to cause o b vious characteristic symptoms (beyond narrowed pupils and runny noses) in any personnel it might have affected. The semiautomatic detectors confirmed the presence of a substance that was positive for cholinesterase suppression, was an organophosphate, had no tertiary amines, and had both a fluorine and an isopropyl group, indicating that the substance was Sarin. “I don’t know of any pesticide with these features,” said Corriveau. Five days
probably in Iraq. Between the two points, he reasoned, the more concentrated gases should have caused deaths or serious injuries, but neither the DOD nor other countries’ troops reported any. Anecdotal evidence given by two disabled veterans who testified at the NIH workshop contradicted that argument, however. One said he witnessed a Scud missile being blown up over the area near Hafr a1 Batin around Jan. 20 and had seen numerous dead camels, sheep, and goats near the road the next day. The other veteran, a veterinarian by training, said the total livestock deaths reported in the region during the war topped 16,000. The workshop panel’s statement said only, “Exposure to chemical and biological warfare agents remains controversial. Many veterans report that exposures occurred. . . . The preliminary report of the DOD’s ‘Defense Science Board,’ which was specifically charged to evaluate chemical and biologic warfare, was not provided to the panel.” Green says the panel restricted its official comments because “The dead animals are an unresolved issue. We were not presented with the full battery of information. The Defense Science Board is supposed to release its own report shortly, but for a lot of these issues, either we’re not getting the data or it wasn’t measured.’’ At this point, little biological evidence is likely to remain for any low-level expoor was it just undergoing electronic sures to chemical warfare agents. Neverdrift-I had no real measure for that.” theless, several European researchers Although the cholinesterase tickets in have begun to examine the possibility of the M256A1 kits had no positive or negative controls, he adds, the tickets would assaying veterans’ DNA samples for the presence of mustard gas adducts as a turn color only if the test was negative. way to confirm or deny allegations of expo“If you think about it, there was a margin of safety because if the test failed, it failed sure. DNA adduction indicates chronic or nonrecent exposure to an agent as well as to the positive,”he says. At the NIH workshop, a spokesman for its carcinogenic potential. MS or immunothe Defence Intelligence Agency said, “In assay methods may be able to detect very low levels of adducts. both of the Czech incidents, there was absolutely no evidence of [Iraqi] military Green says that doing retrospective action, such a s bomb fragments” that analyses on some of the environmental factors in the Gulf War may be more succould have accounted for the detection of nerve or mustard agents. “The U.S. cessful at this late date. For example, soil samples might be acquired from a numfound nothing a few hours later.” ber of areas in the Gulf. “There wasn’t He pointed out that for the Czech demuch emphasis on the role of ground-up tectors to respond to such a low level of surface dust due to traffic around the nerve and mustard gases, there should logically have been a release of the poison bases,” he adds. “That is something else that could be looked at.” gases from a more concentrated source Deborah Noble somewhere north of the detection sites,
later, Czech detectors recorded three positive tests for liquid mustard agent. If the Czech detectors didn’t sense a false alarm, were the American detectors not working? Kirkwood, who was serving as an Army Reserve chemical officer near King Khaled Military City at the time of the Czech detections, says it’s unlikely that all the detectors would fail at the same time. “We had more than 12 M8Als running on a continual basis. There was never more than one that sounded, and in most cases, I was able to identify the cause.” Interferences ranged from a truck idling near the affected alarm to drums of waste from the latrines being burned with fuel. Kirkwood says that when he couldn’t find a tangible reason for the alarm, “I wondered was it getting a whiff of something
ti
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