CONDITIONS AFFECTING THE GROWTH OF BACTERIA Food Requirements.—The supply of food is obviously the most important condition. It has been seen that milk can support the life of bacteria at or near its optimum height, it is in fact as admirable a food for the majority of bacteria as it is for human beings. From this it might appear that bacteria should be consid ered as animals rather than plants if it were not for the fact that the food of plants is in chemical essence the same as that of animals. Every living cell be it plant or animal requires certain food ingredients the chief of which are sugars or other carbo hydrates, proteins or other nitrogenous substances, phosphates, sulphates, chlorides, calcium, magnesium, potassium, sodium and traces of other materials. The only real difference between plants and animals is in the power of the plant to utilize energy from the sun by which it is able to build up its necessary carbo-hydrates from the carbon dioxide of the atmosphere. Bacteria are non green plants and in consequence are unable to invoke the aid of the sun in this direction and are therefore, with two exceptions, de pendent upon some other source of carbon. The exceptions are the nitrifying bacteria and certain sulphur bacteria which obtain the necessary energy for the assimilation of carbon dioxide by the oxidation of ammonia and sulphides respectively. One kind of bacterium is known, Bacillus oligocarbophilus, which can ob tain its carbon from carbon monoxide, and one other Bacillus rnethanicus can make use of marsh gas for this purpose. All other kinds require some sugar or other material like the higher alcohols which can readily be converted into sugar in order to build up their body substance. Most bacteria require a source of nitrogen in combination with organic matter; this requirement is met in milk and in beef extract by the proteins in which these materials are rich, but in using beef extract as an artificial culture medium it has become usual to add a certain amount of peptone as this is an easily digestible ingredient and gives the organisms every encouragement to make rapid initial growth. The nitrifying bacteria, on the other hand, can make no use of such combined forms of nitrogen but are able to build up their own protein from such simple molecules as ammonia and nitrous acid. Other or ganisms, the so-called nitrogen-fixers, can even make use of elemental nitrogen which they assimilate from the air. The re quirements of bacteria in phosphorus, sulphur and other inorganic ingredients is so small that the amount necessary for bacterial development is almost universally present in nature. When these have to be incorporated in artificial culture media they are usually supplied in the form of potassium and sodium phosphates, cal cium and magnesium sulphates and sodium chloride with traces of ferric chloride.
Experiments on soil have shown that there is no growth of bacteria when the water content is 2 to 3%, but bacteria become active at 4 to 5% and reach their optimum, depending on the character of the soil, at 25 to 4o%, in fact, at approximately one-half the water-holding capacity of the soil.
The importance of moisture for bacterial growth will be clearly seen if it is realized that bacteria have no mouth parts and that all their food must be imbibed in a soluble form by the process of diffusion through the cell wall ; without sufficient moisture therefore the inflow of food and the outflow of excreta becomes impossible. A very small percentage of water, however, is capable of preventing death of organisms by desiccation. That which is held hygroscopically by substances in the air-dry condition is as a rule sufficient ; for instance, a fluid suspension of bacteria can be dried down on cotton wool or on filter paper and the bacteria will remain alive for upwards of i 2 months, whereas the same suspension dried down on glass may be dead in 24 hours. As stated earlier the spores of bacteria are considerably more resistant to drying than are the vegetative cells.
For every organism there are certain cardinal points of temperature; there is first the minimum point below which growth is impossible ; this varies with individuals, but for the majority it lies between 5° and 6° C. ; some marine bacteria and certain soil types are active below o° C., but this is ex ceptional. The optimum, depending largely on the usual habitat of the organism, is around 25° C. for soil organisms and about 37° C. for animal parasites. The maximum, above which again no growth is possible, for many organisms lies between 38° C. and 48° C. Death of the organism does not as a rule occur at the maximum temperature for growth, but at a point some ten or 15 degrees higher. The actual death temperature can only be con sidered in relation to time, a low temperature acting for a long time will produce the same effect as a high temperature applied for a short time. A thermal death point standard has been chosen arbitrarily by bacteriologists and represents the lowest tempera ture which, when applied for exactly ten minutes, will destroy every individual in a fluid suspension of the organism. Tempera tures below the minimum prevent bacterial growth but there ap pears to be no equivalent thermal death point at low temperatures. It is known that typhoid and other disease germs survive quite a long inclusion in ice, and instances are known where typhoid epi demics have been spread by the vendor of ice cream. J. Mac Fadyan and S. Rowland found that organisms frozen in liquid air and even in liquid hydrogen at —252° C. were still capable of de velopment when restored to normal incubation temperature.
Everyone knows that if hay be stacked in too moist a condition it is liable to ferment, begin to steam and finally become so hot that it bursts into flame ; but it was only with the advance of bacteriological knowledge that the explanation of this phenomenon was found. There has been dis covered in soil and manure heaps a group of bacteria whose rela tions to temperature are entirely different from those stated above. These bacteria are all of the sporing kind and seldom in the or dinary way of things do they get a chance to germinate, for their minimum temperature for growth is a point above the maxi mum for most organisms and even above the thermal death point of many; e.g., Bacterium ludwigii has its minimum at so° C., reaches its optimum at 55° C.-57° C. and its maximum at 80° C. Fermentation in the haystack is started by moulds, yeasts and bacteria, and heat is thereby developed which enables these ther mophilic organisms to become active carrying the temperature to 80° C. Chemical oxidation then becomes so rapid that the spon taneous ignition point of the gases is reached and the stack bursts into flame. The writer well remembers seeing potatoes in a stor age clamp cooked to that desirable floury condition by the action of thermophilic bacteria. The source of heat in this case was de rived from the rotting of the po tatoes in the lower region of the clamp, though the potatoes above this region were sound but cooked, for the temperature reg istered on a thermometer with its bulb nine inches below the sur face of the clamp was 70° C. and steam was rising to a height of io or i2ft. above the clamp.
It is remarkable to find in organisms, which under the microscope ap pear exactly alike, such differ ences in thermal behaviour as be tween the thermophilic and the ordinary forms of bacteria. It must in fact be a very peculiar form of protoplasm which will re main alive at 80° C. No less re markable perhaps are the differ ences which bacteria of various kinds exhibit in their relation to oxygen. All races of human be ings, and in fact all other animals, make use of oxygen in the same way, yet with bacteria some are known to be aerobic, i.e., they f unction only in presence of air while for others precisely the re verse holds and the smallest quantity of oxygen prevents their growth ; others again, and in this class fall the majority, can tolerate both conditions. These are spoken of among bacteriologists as fac ultative anaerobes. The fact that many bacteria can exist in the anaerobic condition often renders food in hermetically sealed tins unfit for human consumption. The decomposition of organic matter is entirely different in presence or in absence of oxygen. In the former case the process is that of decay in which the breakdown is ultimately complete, the carbon being oxidized to carbon di oxide, the hydrogen to water, the phosphorus and sulphur to phos phates and sulphates, while the nitrogen may be given off as am monia or may be oxidized to the form of nitrates. In the latter case the decomposition is putrefactive, the result being only a partial breakdown of the organic matter with the accompaniment of very offensive smells and the formation of poisonous sub stances, the ptomaines, such as putrescene and cadaverine. Ptomaine poisoning is usually the result of anaerobic bacterial ac tion; Bacillus botulinus, the bacillus which is the cause of botu lism, is perhaps the worst offender.
The development of bacteria is favoured by darkness, many bacteria which are actively motile in the dark become sluggish when moved into the light and in direct sunlight bacteria exposed in thin layers, as for instance on the surface of an agar plate culture, rapidly die ; some organisms under these conditions survive only for ten or 15 minutes. Experi ments on the bacterial formation of nitrate in ammonia solution exposed in shallow dishes to the two conditions showed the rela tive speed of action in the dark to that in the light to be as 86 to 19. It is common knowledge that sunlight is regarded as one of the most powerful agencies in the destruction of pathogenic germs. The part of sunlight which possesses this lethal effect is that at the ultra-violet end of the spectrum. Experiments with ultra-violet light from a Cooper-Hewitt mercury-vapour lamp, of the type now used for treating rheumatism, have shown that Bacillus coli exposed at a distance of one foot from the lamp survives the action of the light for only three minutes.
In order that bacteria may be cul tivated, substances of antiseptic nature must be absent. Anti septic surgery has now given place to aseptic surgery wherein the hands of the surgeon and nurses and every instrument used are sterilized before and during the operation. Antiseptics still have considerable use in industry and in the household, where carbolic acid and chloride of lime are extensively used. Chlorine is a very powerful antiseptic and in the form of sodium hypochlorite is used at the waterworks of large towns to reduce the number of bacteria in the water-supply. It is interesting to note that, whereas, in the main, antiseptics behave similarly towards all organisms, differences exist as between one organism and another in the con centration of poison which they can withstand ; e.g., Bacillus typhosus will not grow in bouillon containing as little as one part of formalin in i 5,000, while Bacillus coli, an organism in many respects closely related to it, develops vigorously in a concentra tion of one part in 3,000. Moreover there have been isolated from soil some organisms which are not only insensitive to the poisonous action of phenol and toluene but can actually utilize these sub stances as their source of carbon.
The reaction of the culture medium, that is, the amount of free acid or free alkali present in it, is of the greatest importance for bacterial develop ment. A culture medium which is neither acid nor alkaline is said to be neutral and is best adapted to the growth of many kinds of bacteria. There are numerous organisms that are favoured in their growth by a faintly acid medium and others again that prefer the medium to be slightly alkaline. It seems rather to depend upon the way in which they have been brought ap; i.e., upon the reac tion of the medium in which the organisms normally exist. Many of the common inhabitants of milk, a medium that as we have seen readily becomes acid, grow better in artificial culture media whose H. ion concentration is on the acid side of neutrality, whereas as a rule the animal parasites, whose normal habitat (blood) is slightly alkaline, make better initial growth on media whose H. ion concentration is on the alkaline side of neutrality.
In a few instances bacteria will develop vigorously in distinctly acid media ; a common example of this kind is the familiar "mother of vinegar," which gelat inous material when examined by the microscope is revealed as a colony of bacteria adhering so closely together that they are difficult to separate.
Such a mass is termed a zoogloea and forms a remarkable phenomenon in the life-history of certain bacteria.
A zoogloea may occur as a mem brane on the surface of the medium, or as irregular clumps or branched masses (sometimes several inches across) submerged in it. Such growths represent a resting condition, the various ele ments being glued together, as it were, by their enormously thickened and mucilaginous cell-walls. Such thickened cell-walls are called capsules and the organisms in which this thickening occurs are said to be capsulated. Under favourable conditions the elements in the zoogloea again become active, move out of the matrix and distribute themselves in the surrounding medium to grow and multiply as before. If the zoogloea is formed on a solid substratum it may become firm and horny; the subsequent im mersion in water softens it and the organisms become active as described.
