The Fundamentals of Bacteriology

PART II.
PHYSIOLOGY.

CHAPTER VI.
GENERAL CONDITIONS FOR GROWTH.

OCCURRENCE.

Bacteria are probably the most widely distributed of living organisms. They are found practically everywhere on the surface of the earth. Likewise in all surface waters, in streams, lakes and the sea. They occur in the air immediately above the surface, since they are carried up mechanically by air currents. They cannot fly of themselves. There is no reason to believe that any increase in numbers occurs to an appreciable extent in the air. The upper air, for example, on high mountains, is nearly free from them. So also is the air over midocean, and in high latitudes. As a rule, the greater the amount of dust in the air, the more numerous are the bacteria. Hence they are found more abundantly in the air in cities and towns than in the open country. The soil is especially rich in numbers in the upper few feet, but they diminish rapidly below and almost disappear at depths of about six feet unless the soil is very porous and open, when they may be carried farther down. Hence the waters from deep wells and springs are usually devoid of these organisms. In the sea they occur at all levels and have been found in bottom ooze dredged from depths of several miles. It is perhaps needless to add that they are found on the bodies and in the alimentary tract of human beings and animals; on clothing, utensils; in dwellings, stables, outhouses, etc. From one-fourth to one-half of the dry weight of the feces of animals and men is due to the bacteria present. The urine is practically free from them in health.

While bacteria are thus found nearly everywhere, it is an entirely mistaken idea to suppose that all are injurious to man. As a matter of fact, those which are dangerous are relatively few and are for the most part found only in close association with man. Most bacteria are harmless and the vast majority are beneficial or even essential to man’s existence on the earth. These facts must be constantly borne in mind, and it is hoped that the pages which follow will make them clear.

In order that any organism may thrive there are a number of general environmental conditions which must be fulfilled. These conditions vary more or less for each kind of organism. Bacteria are no exception to this general rule. These conditions may be conveniently considered under the general heads of moisture; temperature; light; oxygen supply; osmotic pressure; action of electricity; of Röntgen and radium rays; pressure; mechanical vibration; and chemical environment, including the reaction of the medium, the effect of injurious chemicals, and especially the food requirements of bacteria. For each of these conditions there is a maximum, meaning the greatest amount of the given condition which the organism can withstand, a minimum, or the least amount, and an optimum or that amount which is most favorable for development. Further, there might be distinguished a maximum for mere existence and a lower maximum for development; also a minimum for mere existence and a higher minimum for development. These maxima, minima, and optima for bacteria have been determined with exactness for only a very few of the general conditions and for comparatively few kinds.

MOISTURE.

The maximum moisture is absolutely pure water, and no organism can thrive in this alone owing to the factor of too low osmotic pressure and to the further factor of absence of food material. There are many bacteria which thrive in water containing only traces of mineral salts and a large class whose natural habitat is surface water. These “water bacteria” are of great benefit in the purification of streams. They are as a class harmless to men and animals. Some of the disease-producing bacteria like Bacterium typhosum (of typhoid fever) and Vibrio choleræ (of Asiatic cholera) were undoubtedly originally water bacteria, and it is rather striking that in these diseases conditions are induced in the intestine (diarrheas) which simulate the original watery environment. The minimum moisture condition is absolute dryness, and no organism can even exist, not to say develop, in such a condition since water is an essential constituent of living matter. Some bacteria and especially most spores may live when dried in the air or by artificial means for months and even years, while some are destroyed in a few hours or days when dried (typhoid, cholera, etc.). The optimum amount of moisture has not been determined with any great accuracy and certainly a rather wide range in percentage of water is permissible with many, though a liquid medium is usually most favorable for artificial growth. The “water bacteria” have been mentioned. In the soil a water content of 5 to 15 per cent. seems to be most suitable for many of the organisms which aid in plant growth. In animals and man the organisms infecting the intestinal tract prefer a high percentage of moisture as a rule, especially those causing disease here. Those found on the surface of the body (pus cocci) need a less amount of water, while those invading the tissues (tuberculosis, black-leg, etc.) seem to be intermediate in this respect. In artificial culture media a water content of less than 30 per cent. inhibits the growth of most bacteria.

As a general rule those bacteria which require the largest percentage of water are most susceptible to its loss and are most readily killed by drying. The typhoid and cholera organisms die in a few hours when dried, while pus cocci and tubercle bacilli live much longer.

TEMPERATURE.

The temperature conditions for bacterial existence and growth have been determined more accurately than any of the other general conditions. The maximum for existence must be placed at or near 100° since it is known that all bacteria including spores may be killed by boiling in time. Nevertheless, certain forms have been reported as thriving in hot springs where the water temperature was 93°. This is the highest known temperature for development. The minimum for existence lies at or near the absolute zero (-273°) since certain organisms have been subjected to the temperature produced by the sudden evaporation of liquid hydrogen (-256° to -265°) and have remained alive. Whether they could withstand such temperatures indefinitely is not known. The minimum for development is near the freezing-point of water, since reproduction by division has been observed in the water from melting sea-ice at a temperature of -1.5°. Thus bacteria as a class have a range for existence of about 373° (-273° to +100°) and for development of 94.5° (-1.5° to +93°) certainly much wider ranges than any other group of organisms.5

The optimum temperature for development varies within rather wide limits for different organisms. In general it may be stated that the optimum temperature is approximately that of the natural habitat of the organism, though there are exceptions. The optimum of the “hot spring” bacteria just mentioned is apparently that of the springs (93° in this case). Many soil organisms are known whose optimum is near 70° (a temperature rarely, if ever, attained in the soil), but only when grown in air or oxygen; but is very much lower when grown in the absence of oxygen. Many other soil organisms exhibit very little difference in rate or amount of growth when grown at temperatures which may vary as much as 10° or 15°, apparently an adaptation to their normal environment. The disease-producing organisms show much narrower limits for growth, especially those which are difficult to cultivate outside the body. For example, the bacterium of tuberculosis in man scarcely develops beyond the limits of 2° or 3° from the normal body temperature of man (37°), while the bacterium of tuberculosis in birds grows best at 41° to 45°, the normal for birds, and the bacterium of so-called tuberculosis of cold-blooded animals at 14° to 18°.

Those bacteria whose optimum temperature is above 40° are sometimes spoken of as the “thermophil” bacteria. The fixing of the “thermal death-point” that is, the minimum temperature at which the bacteria are killed is a matter of great practical importance in many ways and numerous determinations of this have been made with a great many organisms and by different observers. The factors which enter into such determinations are so many and so varied that unless all the conditions of the experiment are given together with the time of application, the mere statements are worthless. It may be stated that all young, actively growing (non-spore-containing) disease-producing bacteria, when exposed in watery liquids and in small quantities are killed at a temperature of 60° within half an hour. It is evident, that this fact has very little practical application, since the conditions stated are rarely, if ever, fulfilled except in laboratory experiments. (See Sterilization and Pasteurization, Chapter XIII.)

LIGHT.

Speaking generally, it can be said that light is destructive to bacteria. Many growing forms are killed in a few hours when properly exposed to direct sunlight and die out in several days in the diffuse daylight of a well-lighted room. Even spores are destroyed in a similar manner, though the exposure must be considerably longer. Certain bacteria which produce colors may grow in the light, since the pigments protect them. Some few kinds, like the sulphur bacteria, which contain a purplish-red pigment that serves them to break up H₂S, need light for their growth. Since disease-producing bacteria are all injuriously affected by light, the advantage of well-lighted habitations both for men and animals is obvious.

OXYGEN SUPPLY.

Oxygen is one of the constituents of protoplasm and is therefore necessary for all organisms. This does not mean that all organisms must obtain their supply from free oxygen, however, as animals and plants generally do. This fact is well illustrated by the differences among bacteria in this respect. Some bacteria require free oxygen for their growth and are therefore called aërobic bacteria or aërobes (sometimes strict aërobes, though the adjective is unnecessary). Others cannot grow in the presence of free oxygen and are therefore named anaërobic bacteria or anaërobes (strict is unnecessary). There are still other kinds which may grow either in the presence of free oxygen or in its absence, hence the term facultative anaërobes (usually) is applied to them. The distinction between facultative aërobe and facultative anaërobe might be made. The former means those which grow best in the absence of free oxygen, though capable of growing in its presence, while the latter term means those which grow best in the presence of free oxygen, but are capable of growing in its absence. The amount of oxygen in the atmosphere in which an organism grows may be conveniently expressed in terms of the oxygen pressure, i.e., in millimeters of mercury. It is evident that the maximum, minimum and optimum oxygen pressures for anaërobic bacteria are the same, namely, 0 mm. Hg. This is true only for natural conditions, since a number of anaërobic organisms have been gradually accustomed to increasing amounts of O, so that by this process of training they finally grew in ordinary air, that is, at an oxygen pressure of about 150 mm. Hg. (Normal air pressure is 760 mm. Hg. and oxygen makes up one-fifth of the air.) The minimum O pressure for facultative anaërobes is also 0 mm. Hg. Some experiments have been made to determine the limits for aërobes, but on a few organisms only, so that no general conclusions can be drawn from them. To illustrate: Bacillus subtilis (a common “hay bacillus”) will grow at 10 mm. Hg. pressure but not at 5 mm. Hg. It will also grow in compressed oxygen at a pressure of three atmospheres (2280 mm. Hg.), but not at four atmospheres (3040 mm. Hg.), though it is not destroyed.

Parodko has determined the oxygen limits for five common organisms as follows:

These few instances do not disclose any general principles which may be applied either for the growth or for the distinction of aërobes or facultative anaërobes.

It has been shown that compressed oxygen will kill some bacteria but this method of destroying them has little or no practical value. Oxygen in the form of ozone, O₃, is rapidly destructive to bacteria, and this fact is applied practically in the purification of water supplies for certain cities where the ozone is generated by electricity obtained cheaply from water power. The same is true of oxygen in the “nascent state” as illustrated by the use of hypochlorites for the same purpose.

It was stated (p. 74) that certain thermophil bacteria in the soil have an optimum temperature for growth in the air which is much higher than is ever reached in their natural habitat and that they grow at a moderate temperature under anaërobic conditions. It has been shown that if these organisms are grown with aërobes or facultative anaërobes they thrive at ordinary room temperature. These latter organisms by using up the oxygen apparently keep the tension low, and this explains how such organisms grow in the soil.6

OSMOTIC PRESSURE.

Like all living cells bacteria are very susceptible to changes in the density of the surrounding medium. If placed in a medium less concentrated than their own protoplasm water is absorbed and they “swell up”; while if placed in a denser medium, water is given off and they shrink (plasmoptysis or plasmolysis). Should these differences be marked or the transition be sudden, the cell walls may even burst and the organisms be destroyed. If the differences are not too great or if the transition is made gradually, the organisms may not be destroyed, but will either cease to grow and slowly die out, or will show very much retarded growth, or will produce abnormal cell forms. This is illustrated in the laboratory in attempting to grow bacteria on food material which has dried out. A practical application of osmotic effects is in the use of a high percentage of sugar in preserving fruits, etc., and in the salting of meats. Neither the cane-sugar nor the common salt themselves injure the bacteria chemically, but by the high concentration prevent their development. In drying material in order to preserve it there are two factors involved: first, the loss of water necessary for growth and second, the increased osmotic pressure.

In a medium of greater density diffusion of water is outward from the cell and this will continue until an equilibrium is established between cell contents and medium. Food for the organism must be in solution and enter the cell by diffusion. Therefore, growth ceases in a medium too dense, since water to carry food in solution does not enter the cell.

ELECTRICITY.

Careful experimenters have shown that the electric current, either direct or alternating, has no direct destructive effect on bacteria. In a liquid medium the organisms may be attracted to or repelled from one or the other pole or may arrange themselves in definite ways between the poles (galvanotaxis), but are not injured. However, electricity through the secondary effects produced, may be used to destroy bacteria. If the passage of the electric current increases the temperature of the medium sufficiently, the bacteria will be killed, or if injurious chemical substances are formed (ozone, chlorine, acids, bases, etc.), the same result will follow (see Ozone, pp. 77 and 157).

RADIATIONS.

Röntgen or x-rays and radium emanations when properly applied to bacteria will destroy them. The practical use of these agents for the direct destruction of bacteria in diseases of man or animals is restricted to those cases where they may be applied directly to the diseased area, since they are just as injurious to the animal cell as they are to the bacteria, and even more so. Their skilful use as stimuli to the body cells to enable them to resist and overcome bacteria and other injurious organisms or cell growths is an entirely different function and will not be considered here.

PRESSURE.

Hydrostatic pressure up to about 10,000 pounds per square inch is without appreciable effect on bacteria as has been shown by several experimenters and also by finding living bacteria in the ooze dredged from the bottom of the ocean at depths of several miles.

Pressures from 10,000 to 100,000 pounds show variable effects. Some bacteria are readily killed and others, even non-spore formers, are only slightly affected. The time factor is important in this connection. The presence of acids, even CO₂, or organic acids, results in the destruction of most non-spore formers.

MECHANICAL VIBRATION.

Vibrations transmitted to bacteria in a liquid may be injurious to them under certain circumstances. Some of the larger forms like Bacillus subtilis may be completely destroyed by shaking in a rapidly moving shaking machine in a few hours. Bacteria in liquids placed on portions of machinery where only a slight trembling is felt, have been found to be killed after several days. Reinke has shown that the passing of strong sound waves through bacterial growths markedly inhibits their development.