Experiment Eleven

by Peter Pringle

  Schatz was discharged from the army on June 15 and officially started work in Waksman’s department on June 30. Whether Waksman’s letter to Dr. Seibert was prompted by a conversation with Schatz after his discharge from the army is not known.

  In any case, Schatz did not begin his experiments testing his candidate antibiotics against Mycobacterium; that was to be the second part of his Ph.D. He started by enriching soil in pots with E. coli, as Boyd Woodruff had done to find actinomycin.

  Schatz’s first experiment, as noted in his 1943 lab notebook, was on June 30 and dealt with a “general survey of the occurrence of antagonistic microorganisms.” He isolated bacteria, fungi, and actinomycetes which might be responsible for destroying E. coli, then tested them by the streak test to see whether they produced the clear zones. On July 23, he noted in his lab notebook, he gave up the soil-enrichment method and instead switched to the agar plate method used by Dr. Kocholaty when he found streptothricin. This meant random testing of soil samples against known disease-causing bacteria. In experiments started at the beginning of August, Schatz noted that “some molds are apparently antagonistic immediately upon isolation, but they seem to lose this property upon repeated culture on artificial media.” In other words, he still hadn’t found anything worthy of isolation and further experiment.

  Every researcher knew that no matter how great the effort, luck was always involved in discovery, in biology especially. Alexander Fleming discovered penicillin in 1928 when a fungus spore fell by a happy accident into one of his petri dishes and he noticed the clear zones of antagonism. Too many researchers went on testing for years and never found a microbe capable of killing off a disease. Only the single-minded, obsessive researcher, the kind willing to give up a full life, the “true devotee to science,” in Einstein’s terms, would have a chance of success. Albert Schatz was such a researcher. Fleming used to describe his microbe experiments as “playing about,” and Schatz knew the feeling well. He loved spotting a likely microbe, one that he thought might make a good candidate to produce an antibiotic. He loved fussing over his precious molds as they blossomed into striking, beautiful sculptures of red, blue, yellow, and gray-green. He was so fascinated by the potential power of his friendly bacteria that it seldom felt like work; even the routine, the drudgery, seemed like play. No great skills were needed, he was the first to admit. At this early stage, it was just about a steady hand and a good eye. The basic techniques, which he had learned quickly, were known as “silly simple” by one of Dr. Waksman’s graduates.

  Schatz already understood that his chances of finding a useful antibiotic were remote at best, and the human TB strain posed a special problem. There was a good reason why others had failed. Of the disease-causing microbes, the deceptively simple cell of Mycobacterium tuberculosis presented one of the greatest challenges.

  Waksman and Schatz working in the laboratory at Rutgers. (Special Collections and University Archives, Rutgers University Libraries)

  Bacteria are divided into two groups on the basis of their reaction to a stain first used in 1884 by a Danish bacteriologist, Hans Christian Gram. Those cells that retain the stain are called Gram-positive; those that don’t, Gram-negative. The distinction is important for diagnostic purposes and is due to a basic difference in the cell structure. Gram-negative cells, which cause typhoid and cholera, have an extra outer layer, making them tougher for antibiotics to penetrate. Penicillin, for example, is effective against Gram-positive bacteria such as Staphylococcus, the common cause of blood poisoning, but not against Gram-negative Salmonella, which can cause typhoid, or Vibrio cholerae, which causes cholera. The TB microbe is even more difficult to penetrate, having an extra layer of protection, a waxy wall preventing the entry of unwanted chemicals.

  Schatz was determined to find antibiotics which destroyed the Gram-negative bacteria and that also destroyed the TB germ. After ten experiments he had no promising results, so that was when he switched to a third method—random selection. This is the least complicated of the three methods, relying to a greater extent than the other methods on chance. He took samples of soil, compost, or stable manure, added tap water to make a liquid mix, then put drops of the mix on a petri dish of agar containing a food microbes are known to thrive on—egg albumin. Then he incubated the dish and watched the microbes grow. By mid-October, Schatz had selected two strains from the gray-green Actinomyces griseus. One strain, 18-16, excreted an antibiotic active against the Gram-negative bacteria E. coli. Another strain came from the culture of the chicken’s throat given him by Jones. It was also active against E. coli. Schatz named it D-1, for Doris. On October 19, Schatz sealed the test tube containing the 18-16 strain and gave it to his mother.

  On October 20, Schatz began isolating his new antibiotic in Experiment 25—the “Collection of Active Material of 18-16.” The entry ends with a note: “Material taken over by Dr. W. & E.B.” E.B. was Elizabeth Bugie, who was working under Dr. Waksman in the upstairs lab. She began testing the strain in different mixes of nutrients to find the best one for producing the new antibiotic. The favorite seemed to be a meat extract.

  It was “impossible to set down in words the excitement that prevailed in the laboratory during those ensuing days,” according to the book Miracles from Microbes: The Road to Streptomycin by Samuel Epstein and Beryl Williams, who had already collaborated on several books for young readers. The book was published by Rutgers University Press in 1946. Schatz did not agree with the Epstein and Williams account—at least of his mood, which was not excitable, he said, because of the work yet to be done. Many years after he wrote in the margin of his copy of the book, “Not true, the results were in vitro tests. Nobody knew the toxicity.” He was right, of course. No one could know, until tests on animals were done, whether his new antibiotic would be toxic to humans like others produced so far in Waksman’s lab.

  Waksman gave a sample of the new antibiotic to Doris Jones in Poultry Pathology for the first animal tests—on chick embryos, still in the egg, infected with fowl typhoid that would normally kill them. Jones injected them with five to ten milligrams of the new antibiotic and many of them hatched. She was so new at this technique that she had never watched them peck their way out of the shells. Then she had to kill them for an autopsy to see if the typhoid bacteria had been completely destroyed. Before the operation, she hosed down the walls of the autopsy room, she recalled later. It was the only method she had of trying to rid the autopsy room of dust-containing bacteria that might have contaminated her experiment. “But I’m paralyzed, I can’t squeeze the scissors through the necks of those little beings,” she wrote. “Dr. Beaudette mocks me. He won’t help. It takes days. Finally, I squeeze and the autopsy proceeds as tears run down.” The new antibiotic had worked; no bacteria were left in the chick’s organs or blood.

  By this point Waksman’s lab must have been buzzing with word of the discovery, and the question was what to call the new antibiotic. The naming of it would be the cause of yet another unresolved disagreement between Waksman and Schultz. Each claimed that they had thought of the name first. Waksman was the first to use the name streptomycin in a document, according to the archives. In a letter, he informed Randolph Major, Merck’s research director, of the discovery on October 28, saying it could “tentatively be designated as streptomycin.” Schatz claims that he was always going to call his discovery streptomycin, but the name does not appear in Schatz’s lab notebook until December 14.

  Toward the end of October Schatz started to test his new antibiotic from the two strains, 18-16 and D-1, against the harmless strains of mycobacteria in the department’s collection. On November 8, he began Experiment 29, which he titled “Bacterial Action of 18-16 Concentration Upon TB.” A week later, on November 18, he started Experiment 30, which was designed to test D-1 on TB. The results were promising, but the tests were again against TB from the department’s collection and, therefore, non-pathogenic. The real test was yet to come.

  AS SCHATZ BEGAN his Exper
iment 30, a distinguished visitor arrived at the Department of Soil Microbiology. He was Dr. William Feldman, a veterinary pathologist at the Mayo Clinic in Rochester, Minnesota, one of the most famous private clinics in America. Its founder, William Worrall Mayo, a British immigrant, had opened the clinic as a frontier practice in 1846. By the 1930s, the Mayo Clinic was well known for its work in trying to find a cure for tuberculosis. Feldman and a colleague, Corwin Hinshaw, were the principal researchers.

  Feldman had immigrated to the United States from Scotland in 1894 at the age of two, and he had grown up in Colorado knowing about TB. His mother had recalled, from her own childhood in Glasgow, the suffering caused by the disease. For his doctorate at Colorado State University, Feldman had taken part in a nationwide effort to eradicate TB in cattle, a cause of often-fatal tuberculosis in children. He had also written a book on tuberculosis in birds. His partner, Hinshaw, was also a Westerner, a zoologist and medical doctor with expertise in parasites and bacteria. He had been at the Mayo Clinic since 1933, working on pulmonary diseases, especially pneumonia. Their mix of disciplines was excellent for testing new drugs on animals; they made a perfect team.

  For more than a decade they had tried a variety of drugs, including compounds of gold and arsenic and the latest sulfa compounds, on guinea pigs infected with the human TB strain. Results from the sulfa drugs were encouraging. They treated about a hundred patients with drugs that showed unmistakable promise. One of the sulfas, Promin, gave the first hint that the waxy outer wall of M. tuberculosis could be breached, but did not destroy the germ. Nothing seemed to halt the onward march of the TB microbe, which nestled in inaccessible places in the lungs of patients suffering from the disease. Feldman felt that he and Hinshaw had a “foot in the door,” as Feldman put it, but their peers thought they were “wasting their time”; the sulfa drugs might halt the infection, but would never completely destroy TB. This only made the two scientists more determined and stubborn.

  They devised more rigorous tests, insisting that virulent and nonvirulent TB germs had different characteristics and that any new drug had to work against the virulent type before they would even consider animal tests. They used guinea pigs because they are highly susceptible to human TB and, once infected, are severely hit by the disease; if a drug worked, it should also work in humans.

  When Feldman heard about Waksman’s new research into antibiotics, he asked for samples, first of the fungus-produced clavacin, which sounded promising. Waksman was cautious. All four of the antibiotics discovered in his lab—actinomycin and streptothricin from the actinomycetes and fumigacin and clavacin from fungi—were too toxic. Even so, he invited Feldman to visit Rutgers.

  Feldman spent several hours talking with Waksman in the upstairs laboratory, offering to carry out “co-operative studies” if and when Waksman’s lab produced a new antibiotic. Waksman said he would let Feldman know. Before they parted, Waksman introduced Feldman to Schatz, but only briefly, and he never mentioned Schatz’s new discovery of streptomycin. Waksman’s loyalty was to his sponsor, Merck.

  ON DECEMBER 14, Schatz began Experiment 33, using extracted streptomycin on an H37 strain of TB, apparently from the strains of uncertain viability Dr. Seibert had sent Waksman back in June. The results were again promising.

  Schatz was astonished to have found not one but two likely candidates so quickly. What the future held for his discovery, however, no one could really say. His chosen two might be eliminated as quickly as the other four because of toxicity. And while he assumed that Dr. Waksman would do all he could to make his discovery a reality and give him full credit for it, sometimes Schatz was not quite so sure.

  Waksman’s permanently wrinkled attire gave the impression of an eccentric academic absorbed in lofty scientific principles and novel ideas, a professor dedicated to pure research. And his passion for the little-known actinomycetes added to the image of unworldly benevolence. However, Schatz and the other researchers knew another side of their professor.

  Selman Waksman was the best-organized, the most practical, and the best-connected professor at the College of Agriculture. Beneath his bonhomie, the twinkle in his bright eyes, the Friday brown-bag lunches, and the informal summer picnics on the Jersey Shore with “kosher” hot dogs wrapped in bacon, there was a traditional European department head who followed a rigid code of rank when it came to his apprentices.

  One unsettling story about Waksman’s relationship with his students had become a legend at Rutgers. Oddly, it involved another Russian Jewish immigrant, Jacob Joffe, a fiery character who was born in Lithuania and had immigrated to America on the eve of World War One. He became one of Waksman’s first graduate students. He finished his dissertation on a bacterium that had the remarkable ability to turn sulfur compounds into sulfuric acid, which could then be used to free up phosphates in the soil, a natural fertilizer. This was a major breakthrough in soil fertility at the time, and Joffe completed his Ph.D. thesis in 1922.

  Waksman cooks “kosher” hot dogs wrapped in bacon at a Department of Soil Microbiology picnic in 1945. (Courtesy Vivian Schatz)

  A few months later, Waksman wrote about these experiments in a scientific paper, putting his name first as the senior author. Joffe was shocked. He believed he was the one, not Waksman, who had discovered the microbe. Though he stayed on at Rutgers, becoming a professor at the college and an authority on soil science, Joffe nourished a dislike for Waksman, never ceasing to complain that, in his view, Waksman had stolen his work.

  As an undergraduate, Schatz had taken a course with Joffe; he had gotten to know him well and had heard him complain that Waksman had not given him due credit. “Joffe would, not in class, but in his office, rant and rave about Waksman,” Schatz later recalled. Some believed Joffe, others Waksman. The disagreement was never resolved, and the incident had left a question mark over Waksman’s apparently amiable stewardship of the Department of Soil Microbiology.

  Most of the time that he worked for Waksman, Schatz never gave the Joffe legend, and what he knew about it, a second thought. But now that he had his own discovery, he began to wonder how Waksman would respond. The two scientists had a close relationship at that point—partly, Schatz was sure, because they were both Jewish, with roots in Russia. Waksman was not only his teacher but had also become a father figure, the male guide and mentor that he had not found in his own childhood growing up on a dirt farm in Connecticut.

  He wrote up his results and gave them to Waksman to check and edit. The first scientific paper announcing streptomycin was published in the Proceedings of the Society for Experimental Biology and Medicine in January 1944. Schatz was thrilled. Waksman had acknowledged the crucial work Schatz had done by putting his name first, then Betty Bugie’s. Waksman’s name came last. It seemed that Schatz had no cause for concern that the Joffe case might be repeated, with him as the loser this time. But he was not privy to the behind-the-scenes struggle over the next stage of the discovery, the race to publish the effects of streptomycin on the deadly human strain of tuberculosis.

  6 • The Race to Publish

  AT THE MAYO CLINIC, WILLIAM FELDMAN received a copy of the three-and-a-half-page paper announcing streptomycin in February 1944. He and Hinshaw read down the list of twenty-two bacteria vulnerable to the new antibiotic and were astonished to find, among the usual test germs, M. tuberculosis. They were also amazed that there was no discussion anywhere in the text explaining the nature of the strain, whether it was the harmless kind or the deadly H37Rv. In reality, it was harmless, as Hinshaw would learn much later. Waksman told him that the strain of Mycobacterium used by Schatz was a non-pathogenic strain from the Department of Soil Microbiology collection. Waksman added that he would “not permit any pathogenic tuberculosis culture in his laboratory for fear of infection to students and technicians.” But this rule was about to change—for Schatz’s experiments in his basement lab. In reading the paper, Feldman also found it odd that Waksman had not mentioned this discovery when Feldman had v
isited Rutgers a few months earlier, though he must have known about it. The lead time on such journal articles was a month at least.

  Feldman was about to contact Waksman when a letter came from the Rutgers professor himself, in which he offered to supply a sample of streptomycin to the Mayo laboratories for guinea pig trials. Feldman accepted immediately, telling Waksman he needed ten grams, which he estimated “from the meager knowledge available” would be enough for a small test—say, four to six guinea pigs. In exchange, Feldman agreed to send Waksman a culture of the virulent human strain, H37Rv, which Schatz needed for his in vitro (test tube) experiments.

  In his basement lab, Schatz cranked up his small stills, frothy brews in glass flasks, to provide Feldman with ten grams. Schatz did not have to be told how to run a still; he had learned the tricks at an early age during Prohibition on the family farm in Connecticut. In the basement lab he now ran stills twenty-four hours a day, sleeping on the lab floor. He had an arrangement with the night watchman that if, on his rounds, he found Schatz asleep and the liquid in the flasks had evaporated below a red line, he was to wake him up. This punishing routine left Schatz permanently exhausted, and one morning he left the lab at about two o’clock and collapsed in the snow outside. Fortunately, the night watchman found him unconscious and called an ambulance to take him to the emergency room. He had pneumonia and spent a week in hospital. Waksman was the only member of the staff who didn’t visit him.