Soil Organisms and Disease

FEW OF THE bacteria and other micro-organisms that cause human and animal diseases survive long in the soil. Once introduced, they "are inhibited or killed by antagonistic organisms, which produce active chemical substances known as antibiotics.

This purifying effect of the soil has been recognized since the early days of modern bacteriology, that is, since about the latter part of the nineteenth century. But because antibiotics vary greatly in chemical nature, selective antibacterial properties, toxicity to animals, and activity, it is only in recent years that a systematic attempt has been made to use them to control infections in man and animals. The most promising antibiotics now known are penicillin, tyrothricin, and streptomycin.

Only comparatively few groups of soil organisms have so far been examined for their antibacterial potentialities. Nothing is known yet of the ability of soil organisms to combat viruses and other infectious agents for which no adequate methods of control are now available. The field for further investigation is broad, therefore, and offers promise of potential practical developments.

Pasteur was the first to observe, in 1877, that the presence of certain "common bacteria” in the culture of an anthrax organism brought about considerable modification of the pathogenic properties of the latter. He suggested, therefore, the possibility of utilizing "common bacteria” for therapeutic purposes. When soon afterward, in 1881, the gelatin-plate method for counting and isolating bacteria was introduced by Koch, the attention of the medical bacteriologist was focused upon the soil as a habitat of disease-producing bacteria and as a possible source of infections and epidemics. The soil was analyzed for the total number of bacteria and for the presence of organisms capable of causing infections and epidemics.  The results obtained were, however, entirely negative; they did not justify the fear that human pathogens may multiply or even remain in a viable state for any length of time in the soil. On the contrary, it soon became established that the great majority of bacteria causing the common infections, such as diphtheria, anthrax, typhoid, cholera, dysentery, undulant fever, various staphylococci, and even the tuberculosis organism rapidly disappears from the soil.

More detailed and more recent investigations of the problems of soil pollution by intestinal infections revealed that the rate of destruction of typhoid, dysentery, and colon bacteria in the soil depends upon a number of factors. Chief among them are the moisture content of the soil and its reaction, and the nature and abundance of its microbiological population.

In most soils, a large proportion of such disease-producing bacteria was found to die out within 10 days. It was also found that various other pathogenic organisms that cause some of the most deadly human and animal scourges, namely, leprosy, pneumonia, bubonic plague, influenza, cattle mastitis, cattle abortion, and many human and animal virus infections, that constantly find their way into the soil in large numbers, also disappear there sooner or later.

As a result of these and numerous other studies, no one now ever raises the question concerning the role of the soil as a carrier of the great majority of disease-producing organisms or as the cause of severe or even of minor epidemics and infections. One must, of course, exclude from consideration certain spore-forming bacteria, responsible for such infections as tetanus, gas gangrene, and anthrax, or capable of producing toxic substances when they find their way into improperly sterilized foods, such as botulinus. These organisms are of only minor importance and are readily subject to control.

It was quite logical, therefore, that the question should have been raised as to what becomes of all the bacteria excreted by infected individuals. This was expressed by the writer and H. B. Woodruff in 1940 as follows: “If one considers the period for which animals and plants have existed on this planet and the great numbers of disease-producing microbes that must have thus gained entrance into the soil, one can only wonder that the soil harbors so few bacteria capable of causing infectious diseases in man and in animals. One hardly thinks of the soil as a source of epidemics.”

Several theories have been proposed in order to explain the rapid disappearance from the soil of disease-producing bacteria and other micro-organisms. The theories can be grouped into six categories:
   The soil offers an unfavorable environment for the growth of disease-producing micro-organisms.
   The soil is lacking in a sufficient or in a proper food supply for the growth of such micro-organisms.
   The disease-producing bacteria are destroyed in the soil by various predaceous agents, such as protozoa and other animals.
   The bacteria are destroyed in the soil by specific bacteriophages.    The soil-inhabiting micro-organisms acting as antagonists are responsible for the destruction of the pathogenic bacteria and other organisms that find their way into the soil.    These antagonists form specific toxic or antibiotic substances which destroy the pathogenic bacteria, the viruses, and the other disease-producing agents.

It has gradually become recognized that the effectiveness of the soil as a purifier depends upon its nature, to a considerable extent. Actually, the soil exerts a double action upon the contaminating bacteria. In the first place, it removes them by physical adsorption. Heavy loams or clay soils, incidentally, are far more efficient in removing the bacteria from sewage or from contaminated soils than light, sandy, porous soils. Secondly, soil removes bacteria by biological destruction. Different soils show differences in abundance of antagonistic micro-organisms. As a result of early studies on the survival of the cholera organism, some soils are now recognized as “cholera-immune” or “cholera-destroying" soils.

W.D. Frost made an exhaustive study of the destruction of the typhoid organism by antagonistic micro-organisms. He observed that when typhoid bacteria are added to the soil they are rapidly destroyed:  98 percent of the cells were killed in 6 days; in the course of a few more days all the cells tended to disappear entirely. When conditions were not very favorable to the development of the antagonistic organisms, the bacteria survived not only for many days, but even for months.

He visualized clearly the antagonistic effects of micro-organisms: “Bacteria in nature occur almost invariably in mixed cultures. Their association may be without effect on the various species, or it may effect them in various ways. They may offer mutual or one-sided aid, and thus live in a symbiotic relation. They may, on the other hand, offer mutual or one-sided injury. i. e., they may exert an antagonism on one another.”

This antagonism results not only in the inhibition of growth of the pathogenic organisms, but in their actual destruction. The effect of the saprophytic micro-organisms—those that feed on decaying organic material—upon the pathogenic bacteria was found to be due not to the exhaustion of the food supply, to the action of proteolytic enzymes, or to a change in reaction, but to the production of specific agents, designated as “antagonistic substances” which are thermostable in nature.

Dr. Frost also suggested that there was no evidence that would lead one to believe that the antagonistic substances exist as such in the soil, but rather that the antagonistic organism produces substances which are responsible for the destruction of the pathogens. The rapidity of the death-rate of E. typhosa as the result of the presence in the soil of an antagonist was found to depend on the period of preliminary cultivation of the antagonist, which Frost designated as an “antibiont.”

It thus became definitely established that the survival of Eberthella typhosa in manure and in soil depends upon the development of saprophytic micro-organisms; these produce specific substances that are antagonists to the pathogenic bacteria and bring about their rapid destruction.

Similar observations were made for the survival of Escherichia coli.  This organism is rapidly crowded out by other microbes in manure piles; the addition of 9 million Escherichia coli and 13 million Aerobacter aerogenes cells to a soil resulted in reductions to 6,000 and 25,000 cells, respectively, in 106 days; in 248 days, both organisms had completely disappeared from the soil. Gradually evidence began to accumulate that tended to indicate that the occurrence of coliform bacteria in the soil depends entirely on the degree of pollution; soils relatively free from pollution contain none or only a small number of coliform bacteria.

The organism that causes malta fever, Brucella melitensis, is closely related to that responsible for brucellosis and contagious abortion in cattle. It survived in sterile tap water 42 days and in unsterile water only 7 days; it survived 69 days in dry sterile soil and only 20 days in unsterile manured soil. The cholera and diphtheria organisms, as well as other disease-producing bacteria, were found to disappear rapidly in the soil. The same was true of the bacillus causing tuberculosis.

The great importance of soil micro-organisms in the destruction of disease-producing bacteria has thus become definitely established. The nature of this process aroused considerable speculation. In the early days of microbiology, when the antagenistic relations of micro-organisms were first observed, the terms “antibiosis” and “struggle for existence” were used as virtual synonyms. Such interpretations would have been justified when applied to micro-organisms only in those cases when one organism secretes enzymes that actually destroy the other organism. The production by one microbe of a substance that interferes with the life of another microbe, without necessarily benefiting its producer by such an effect, could hardly be thus interpreted.

The numerous interrelations among micro-organisms in natural environments include: (a) Favorable effects, which range from the consumption of oxygen by an aerobe, thereby making conditions favorable to an anaerobe, to the production by an organism of growth-promoting substances necessary for the growth of another; (b) strict symbiosis, which benefits both participants; (¢) unfavorable effects, which include the undesirable effect of an acid produced by one organism upon the growth of another, specific nutrient competition, and the phenomenon of antagonism or antibiosis when one organism produces substances exerting an unfavorable effect upon the growth of anotner. The phenomena of metabiosis, or the living together of two organisms without any apparent effect of one upon the other, fall in a separate category.

G. Papacostas and J. Gaté suggested that the application of the Darwinian concept to the interactions among micro-organisms be completely abandoned. However, in an attempt to clarify or simplify inhibitive phenomena, they applied the term “antibiosis” to mixed cultures (in vitro) and “antagonism’ to mixed infections (in vivo) ; such designations tended to suggest that the interactions among micro-organisms in the test tube are distinctly different from those in the animal body.

Recent evidence in the field of antibiotics does not bear out this distinction. In differentiating between these two terms, the suggestion that the term “antagonism” be applied to the complex unfavorable effects of one living system upon another, when the mechanism involved is not yet clearly understood; and that the term “antibiosis” be used to describe the phenomena of specific selective activities of chemical substances, or antibiotic agents, produced by one organism upon another, appears to have greater justification.

Gradually there evolved the concept of antibiotics, or those agents that are produced by micro-organisms and that have the capacity of inhibiting the growth and destroying bacteria and other micro-organisms. More important yet is the practical application of some of these antibiotics in. the control of human and animal infections. The action of antibiotics against bacteria and other micro-organisms, as distinct from the common antiseptics and disinfectants, is selective in nature, that is, the antibiotics act upon some bacteria and not at all, or to only a limited extent, upon other bacteria. Some act readily upon fungi and others do not. Some are readily soluble in water and others are not. Antibiotics also vary greatly in their toxicity to animals. Because of these characteristics, certain antibiotics have remarkable chemotherapeutic properties and can be used for the control of various bacterial diseases in man and in animals.

The most important property of antibiotics is their selective action upon bacteria and other cells of lower and higher forms of life. Because of this, they may be active upon bacteria, without affecting the host cells.  Although some 100 antibiotics have now been isolated, only 3 have so far found definite practical application. This small percentage of agents having chemotherapeutic potentialities is due to several factors: Some substances are too toxic to animal tissues; some leave undesirable after-effects in the animal body; some are inactivated by blood or tissue constituents of the body; some are not very active and are inferior to others that possess greater activity.

The organisms that produce the three important antibiotics, tyrothricin, penicillin, and streptomycin, represent. typical soil forms, namely, Bacillus brevis, Penicillium notatum-chrysogenum, and Streptomyces griseus, respectively. Although these organisms may also be found in other substrates, the soil may be considered as their natural habitat.

Tyrothricin is produced by a group of aerobic spore-forming bacteria belonging to the B. brevis group. Although many other soil bacteria, notably strains of the spore formers B. mycoides, B. subtilis and B. mesentericus, as well as various non-spore-formers (Pseudomonas aeruginosa), are also capable of producing antibacterial substances, these differ greatly in their chemical composition, antibacterial action, and in vivo activity. A number of antibiotics have now been isolated from soil bacteria, in addition to tyrothricin. It is sufficient to mention subtilin, simplexin, bacillin, pyocyanase, and pyocyanin. Tyrothricin is the best known. It is a polypeptide, or rather a group of polypeptides, several of which have been crystallized, notably gramicidin, tyrocidine, and gramicidin S. Tyrothricin is not soluble in water, but is soluble in alcohol.  It acts largely against gram-positive bacteria, but since it is hemolytic, it can be used only for topical and not for parenteral administration. It is utilized for the treatment of a variety of infections caused by gram-positive bacteria, such as various streptococci and staphylococci, that cause carbuncles and other skin eruptions, sinus infections, and cattle mastitis.

Penicillin is produced by a large number of fungi belonging to the genera Aspergillus and Penicillium. For manufacturing purposes, however, only certain strains of P. notatum and P. chrysogenum are used. The ability to form penicillin is characteristic of both of these species, although different strains vary greatly as regards the quantitative production and the chemical nature of the penicillin type. These fungi are widely distributed in the soil. In 1916, for example, strains of P. notatum and P. chrysogenum were isolated from a variety of soils collected from New Jersey, Louisiana, Colorado, North Dakota, and Puerto Rico.  Penicillin is active only against certain bacteria. The soil contains many organisms capable of destroying penicillin, and it is produced on artificial media only under special conditions of culture. It is highly improbable therefore that penicillin is formed in a natural soil and, even if traces of it are produced, that it is of any significance in the survival of the fungi producing it.

The utilization of specific strains of fungi for the production of penicillin became the high mark of chemotherapy during the Second World War. Many infections caused by various gram-positive and certain gram-negative bacteria were brought under control. It is sufficient to mention the various staphylococcal and streptococcal diseases, the pneumococcal, meningococcal, gonococcal, and clostridial infections; as well as syphilis, anthrax, actinomycosis, and a variety of others. Penicillin is nontoxic and can be used liberally in the treatment of these and other infections.

Streptomyecin is produced by certain strains of an organism known as Streptomyces griseus, belonging to the actinomycetes. As opposed to that of penicillin production, the property of forming streptomycin is characteristic not of the genus Streptomyces, nor of the species of S. griseus, but only of certain strains of this organism. Other species of the genus Streptomyces are also capable of producing antibiotics, but these are markedly different from streptomycin in their physical and chemical properties, antibacterial action, and in vivo activity. The closest antibiotic to streptomycin is streptothricin, produced by S. lavendulae, another soil species.  This substance is much more toxic to animals than is streptomycin.

Streptomycin is active against a variety of gram-negative and gram-positive bacteria not affected by penicillin, including Mycobacterium tuberculosis. It is not very toxic to animals and can be administered parenterally for the treatment of a number of infections, including those of the urinary tract, tularemia, pertussis, typhoid, and tuberculosis. It can also be used orally for the elimination of certain bacteria in the digestive system. It is not active against viruses and fungi and has only a limited effect on anaerobic bacteria.

THE AUTHOR
Selman A. Waksman is a microbiologist at the New Jersey Agricultural Experiment Station, New Brunswick, N. J.