Waksman first became interested in actinomycetes in 1915 as a student at Rutgers. For the next several decades, he studied their occurrence and abundance in soil, their taxonomy, their role in soil processes such as the decomposition of plant and animal residues and the formation of humus, and their relationship to bacteria and fungi. In particular, Waksman and his students calculated the antagonistic effects that acinomycetes had on bacteria and fungi, establishing in the end that perhaps half of all the actinomycetes found in the soil had the capacity to inhibit the growth of other microorganisms.

Early on in the research on the actinomycetes at Martin Hall on the Cook campus at Rutgers, Waksman and his colleagues knew of Streptomyces griseus, the organism that yielded the streptomycin strain, but it would not be tested for its antibiotic producing properties for several decades.² It was also known that the actinomycetes had toxic effects upon bacteria and fungi.

In 1924, after several years devoted to soil research, Waksman and his wife spent six months in Europe, the first time they had returned since they immigrated to the United States. Part of the trip was sentimental, a return to the town of their birth. This was, of course, the early years of the Russian Revolution when much had been destroyed and little yet rebuilt. Waksman wrote of his impressions: "The greatest misery that one could ever magine, the greatest catastrophe that the peoples in Russia had ever gone through; the greatest experiment in social and political relations of men can hardly express what we have seen…"³ The Waksmans spent ten days in Priluka, listening to family members and friends describe their plight: "Each one suffered to an extreme. Many cried like children before their father; they came to pour out before us all their sufferings."⁴

But science was the main reason for the trip. In his autobiography, Waksman referred to the trip as a "grand scientific tour" the purpose of which was to assess his career as a soil microbiologist. "There was no question in my mind," he wrote, "concerning the role of microorganisms in soil processes, but there was a certain question that was continuously arising as to whether I was headed in the right direction."⁵ To get answers, Waksman visited important laboratories in France, Italy, Germany, and Scandinavia to discuss methods and research with the leading scientists working in the fields of soil biology and chemistry. He returned to the United States stimulated by what he learned and convinced of the need for a comprehensive treatise on soil microbiology, which he published in 1927 as the Principles of Soil Microbiology.

In the 1920s and 1930s, Waksman continued his studies on actinomycetes in the soil, which "resulted in the isolation of numerous new species, development of a system for the generic classification of this group of organisms, and a better understanding of their metabolic processes."⁶ Over time, he gradually became convinced that these microorganisms could "exert a considerable influence upon the activities of fungi and bacteria in the soil."⁷ Still, in these years, his major research activity focused on soil microbes and not disease-producing organisms. Two events occurred in 1939 that forced a change in his approach. One was the start of World War II which suggested the need for new agents to control infectious diseases and epidemics certain to arise, especially in tropical theaters. The second event, "a particular stimulus," was the work of René Dubos, Waksman's former student, who isolated tyrothricin, which destroyed disease-producing bacteria.⁸

Dubos had shown that it was possible to find bacteria that inhibited the growth of other bacteria. This represented a significant change in the traditional approach to combating infectious disease. The pioneers in the study of infectious diseases, Koch, Lister, and others, had emphasized the necessity of avoiding contamination by soil and other non-sterile matter. Now a researcher was using a soil-derived agent to combat infectious disease. It was logical that Waksman, given the background of his work on soil microorganisms, would be spurred by this conceptual breakthrough to search for agents active against pathogenic bacteria. Unlike Dubos, however, Waksman concentrated on fungi and, especially, actinomycetes.

Waksman approached the search for antibiotics in a novel and systematic way, unlike the chance discovery of penicillin by Fleming, who observed an accidental contamination of a bacterial pathogen by an airborne mold. Waksman and his students screened by looking for growth inhibition zones surrounding single colonies of a series of isolated soil microbes on agar plates growing under a variety of culture conditions. They then proceeded to test the inhibition on specifically targeted pathogenic bacteria. This was painstaking work, as thousands of cultures of different microbes were isolated and then tested for antibacterial activity, but which only a small percentage demonstrated. These were then further tested to discover which would successfully yield microbe-fighting substances in sufficient quantities and then which were not too toxic for therapeutic use. Waksman's screening protocols were to be very successful, yielding around twenty new natural inhibitory agents, with most coming from the actiniomycetes. In fact, it was Waksman who suggested what has become the common term - antibiotics - for these therapeutic agents.⁹

The first agent isolated in 1940 under the initial screening program was actinomycin by Boyd Woodruff, a Waksman graduate student, who demonstrated that the methodology would yield antibiotic-producing cultures. Actinomycin was active against a broad range of bacteria and even showed promise of attacking a tuberculosis strain, but it proved too toxic for therapeutic use in humans.¹⁰ Two years later, Woodruff isolated streptothricin, an antibiotic which exhibited activity against both gram-positive and gram-negative bacteria.¹¹ The researchers were initially excited about streptothricin because, as Waksman said in his Nobel acceptance speech, it "gave promise of filling the gap left by penicillin in the treatment of infectious diseases due to gram-negative bacteria." In addition to being active against gram-positive and gram-negative bacteria, initial tests of streptothricin showed that it was not toxic to animals. However, pharmacology studies demonstrated that streptothricin had a delayed toxic effect in animals and thus could not be used for therapeutic purposes in humans.¹²

The partial success of streptothricin indicated that Waksman and his students were on the right track. They needed to find a variant that inhibited pathogenic organisms - the easy part - without actually killing host - the hard part. The breakthrough came in 1943 when Albert Schatz joined the team, and continuing the general research approach of cross streaking pioneered by Woodruff, found two strains of Steptomyces griseus that produced streptomycin. One of these was found by chance when Doris Jones, another Waksman student, tested the tracheal flora of healthy chickens and noted zones of antagonism on several plates.¹³ The cultures were given to Schatz and from one of them he isolated the active strain of S. griseus, which produced an antibiotic that inhibited both gram-negative and gram-positive bacteria. This was significant since penicillin had no effect on gram-negative bacteria. Even more exciting to the researchers was that streptomycin exhibited activity in vitro activity against Mycobacterium tuberculosis, the Great White Plague.¹⁴