Читать книгу The Fundamentals of Bacteriology - Charles Bradfield Morrey - Страница 11

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The essential structures which may by appropriate means be distinguished in the bacterial cell are cell wall and cell contents, technically termed protoplasm, cytoplasm. The cell wall is not so dense, relatively, as that of green plants, but is thicker than the outer covering of protozoa. It is very similar to the cell wall of other lower fungi. Diffusion takes place readily through it with very little selective action on substances absorbed as judged by the comparative composition of bacteria and their surrounding medium.

Cytoplasm.—The cytoplasm according to Bütschli and others is somewhat different and slightly denser in its outer portion next to the cell wall. This layer is designated the ectoplasm, as distinguished from the remainder of the cell contents, the endoplasm. When bacteria are suddenly transferred from a given medium into one of decidedly greater density, there sometimes results a contraction of the endoplasm, due to the rapid diffusion of water. This phenomenon is designated plasmolysis (Fig. 17), and is similar to what occurs in the cells of higher plants when subjected to the same treatment. This is one of the methods which may be used to show the different parts of the cell just described.

If bacteria are suddenly transferred from a relatively dense medium to one which is of decidedly less density, it occasionally happens that water diffuses into the cell and swells up the endoplasm so much more rapidly than the cell wall that the latter ruptures and some of the endoplasm exudes in the form of droplets on the surface of the cell wall. This phenomenon is called plasmoptysis. Students will seldom observe the distinction between cell wall and cell contents, except that in examining living bacteria the outer portion appears more highly refractive. This is chiefly due to the presence of a cell wall, but is not a proof of its existence.

Fig. 17.—Cells of bacteria showing plasmolysis. The cell substance of three of the cells in the middle of the chain has shrunk until it appears as a round black mass. The cell wall shows as the lighter area.

Fig. 18.—Vacuoles in the bacterial cell. The lighter areas are vacuoles.

Nucleus.—Douglas and Distaso3 summarize the various opinions with regard to the nucleus in bacteria as follows:

1. Those who do not admit, the presence of a nucleus or of anything equivalent to it. (Fischer, Migula, Massart).

2. Those who consider that the entire bacterial cell is the equivalent of a nucleus and contains no protoplasm. (Ruzicka).

3. Those who admit the presence of nuclein but say that this is not morphologically differentiated from the protoplasm as a nucleus. (Weigert).

4. Those who consider the bacterial protoplasm to consist of a central endoplasm throughout which the nuclein is diffused and an external layer of ectoplasm next to the cell wall. (Bütschli, Zettnow).

5. Those who say that the bacterial cell contains a distinct nucleus, at least in most instances. These authors base their claims on staining with a Giemsa stain. (Feinberg, Ziemann, Neuvel, Dobell, Douglass and Distaso).

That nucleoproteins are present in the bacterial cell in relatively large amounts is well established. Also that there are other proteins and that the protoplasm is not all nuclein.

Some workers as noted above have been able to demonstrate collections of nuclein by staining, especially in very young cells. In older cells this material is in most instances diffused throughout the protoplasm and can not be so differentiated.

The following statement probably represents the generally accepted view at the present time:

A nucleus as such is not present in bacterial cells, except in a few large rare forms and in very young cells. Nuclein, the characteristic chemical substance in nuclei, which when aggregated forms the nucleus, is scattered throughout the cell contents and thus intimately mingled with the protoplasm, and cannot be differentiated by staining as in most cells.

The close association of nuclein and protoplasm may explain the rapid rate of division of bacteria (Chapter VIII, p. 91).

The chemical composition of the bacterial cell is discussed in Chapter VII.

In addition to the essential parts just described the bacterial cell may show some of the following accidental structures: vacuoles, capsules, metachromatic granules, flagella, spores.

Vacuoles.—Vacuoles appear as clear spaces in the protoplasm when the organism is examined in the living condition or when stained very slightly (Fig. 18). During life these are filled with liquid or gaseous material which is sometimes waste, sometimes reserve food, sometimes digestive fluids. Students are apt to confuse vacuoles with spores (p. 47). Staining is the surest way to differentiate (Chapter XIX, p. 209). If vacuoles have any special function, it is an unimportant one.

Fig. 19.—Bacteria seen within capsules.

Fig. 20.—Metachromatic granules in bacteria. The dark round spots are the granules. The cells of the bacteria are scarcely visible.

Capsule.—The capsule is a second covering outside the cell wall and probably developed from it (Fig. 19). It is usually gelatinous, so that bacteria which form capsules frequently stick together when growing in a fluid, so that the whole mass has a jelly-like consistency. The term zoöglœa was formerly applied to such masses, but it is a poor term and misleading (zoön = an animal) and should be dropped. The masses of jelly-like material frequently found on decaying wood, especially in rainy weather, are in some cases masses of capsule-forming bacteria, though a part of the jelly is a product of bacterial activity, a gum-like substance which lies among the capsulated organisms. When these masses dry out, they become tough and leathery, but it is not to be presumed that capsules are of this consistency. On the contrary, they are soft and delicate, though they certainly serve as an additional protection to the organism, doubtless more by selective absorption than mechanically. Certain bacteria which cause disease form capsules in the blood of those animals which they kill and not in the blood of those in which they have no effect (Bacterium anthracis in guinea pig’s blood and in rat’s blood). The presence of capsules around an organism can be proved only by staining the capsule. Many bacteria when stained in albuminous fluids show a clear space around them which appears like a capsule. It is due to the contraction of the fluid away from the organism during drying.

Metachromatic Granules.—The term “metachromatic” is applied to granules which in stained preparations take a color different from the protoplasm as a whole (Fig. 20). They vary widely in chemical composition. Some of them are glycogen, some fat droplets. Others are so-called “granulose” closely related to starch but probably not true starch. Others are probably nuclein. Of many the chemical composition is unknown. They are called “Babes-Ernst corpuscles” in certain bacteria (typhoid bacillus). Since they frequently occur in the ends of cells the term “polar granules” is also applied. Their presence is of value in the recognition of but few bacteria (“Neisser granules” in diphtheria).

Flagellum.—A flagellum is a very minute thread-like process growing out from the cell wall, probably filled with a strand of protoplasm. The vibrations of the flagella move the organism through the liquid medium. Bacteria which are thus capable of independent movement are spoken of as “motile bacteria.” The actual rate of movement is very slight, though in proportion to the size of the organism it may be considered rapid. Thus Alfred Fischer determined that some organisms have a speed for short periods of about 40 cm. per hour. This is equivalent to a man moving more than 200 miles in the same time.

It is obvious that bacteria which can move about in a liquid have an advantage in obtaining food, since they do not need to wait for it to be brought to them. This advantage is probably slight.

Fig. 21.—A bacterium showing a single flagellum at the end—monotrichic.

Fig. 22.—A bacterium showing a bundle of four flagella at the end—lophotrichic.

An organism may have only one flagellum at the end. It is then said to be monotrichic (Fig. 21) (μόνος = alone, single; τριχος = hair). This is most commonly at the front end, so that the bacterium is drawn through the liquid by its motion. Rarely it is at the rear end. Other bacteria may possess a bundle of flagella at one end and are called lophotrichic (Fig. 22) (λοφος = tuft). Sometimes at approaching division the flagella may be at both ends and are then amphitrichic (Fig. 23) (αμφι = both). It is probable that this condition does not persist long, but represents the development of flagella at one end of each of a pair resulting from division of an organism which has flagella at one end only. In many bacteria the flagella arise from all parts of the surface of the cell. Such bacteria are peritrichic (Fig. 24) (περι = around). The position and even the number of the flagella are very constant for each kind and are of decided value in identification.

Fig. 23.—A bacterium showing flagella at each end—amphitrichic.

Fig. 24.—A bacterium showing flagella all around—peritrichic.

Flagella are too fine and delicate to be seen on the living organism, or even on bacteria which have been colored by the ordinary stains. They are rendered visible only by certain methods which cause a precipitate on both bacteria and flagella which are thereby made thick enough to be seen (Chapter XIX, p. 210). The movement of liquid around a bacterium caused by vibrations of flagella can sometimes be observed with large forms and the use of “dark-field” illumination.

Flagella are very delicate and easily broken off from the cell body. Slight changes in the density or reaction of the medium frequently cause this breaking off, so that preparations made from actively motile bacteria frequently show no flagella. For this reason and also on account of their fineness the demonstration of flagella is not easy, and it is not safe to say that a non-motile bacterium has no flagella except after very careful study.

The motion of bacteria is characteristic and a little practice in observing will enable the student to recognize it and distinguish between motility and “Brownian” or molecular motion. Dead and non-motile bacteria show the latter. In fact, any finely divided particles suspended in a liquid which is not too viscous and in which the particles are not soluble show Brownian motion or “pedesis.” This latter is a dancing motion of the particle within a very small area and without change of place, while motile bacteria move from place to place or even out of the field of the microscope with greater or less speed. There is a marked difference in the character of the motion of different kinds of bacteria. Some rotate around the long axis when moving, others vibrate from side to side.

Among the higher thread bacteria there are some which show motility without possessing flagella. Just how they move is little understood.

Spores.—Under certain conditions some bacterial cells undergo transformations which result in the formation of so-called spores. If the process is followed under the microscope, the changes observed are approximately these: A very minute point appears in the protoplasm which seems to act somewhat like the centrosome of higher cells as a “center of attraction” so that the protoplasm gradually collects around it. The spot disappears or is enclosed in the collected protoplasm. This has evidently become denser as it is more highly refractive than before. In time all or nearly all of the protoplasm is collected. A new cell wall is developed around it which is thicker than the cell wall of the bacterium. This thickened cell wall is called the “spore capsule.” Gradually the remnants of the former cell contents and the old cell wall disappear or dissolve and the spore becomes “free” (Fig. 25).

Fig. 25.—The smaller oval bodies in the middle of the field are free spores.

If the spore is placed in favorable conditions the protoplasm absorbs water, swells, the capsule bursts at some point, a cell wall is formed and the bacterium grows to normal size and divides, that is, it is an active growing cell again. This process is called “germination” of the spore. The point at which the spore capsule bursts to permit the new cell to emerge is characteristic for each kind of bacterium. It may be at the end when the germination is said to be polar (Fig. 26). It may be from the middle of one side which gives equatorial germination (Fig. 27). Rarely it is diagonally from a point between the equator and the pole, which type may be styled oblique germination. In one or two instances the entire spore swells up, lengthens and becomes a rod without any special germination unless this type might be designated bi-polar.

Fig. 26.—Spores showing polar germination. The lighter part of the two organisms just below A and B is the developing bacterium. In the original slide the spore was stained red and the developing bacterium a faint blue.

Fig. 27.—A spore showing equatorial germination. The spore in the center of the field shows a rod growing out of it laterally. In the original slide the spore was stained red and the developing bacterium blue.

Fig. 28.—Spores in the middle of the rod without enlargement of the rod. The lighter areas in the rods are spores.

Fig. 29.—Spores in the middle of the rod with enlargement of the rod around them. The lighter areas in the rods are spores.

Spores are most commonly oval or elliptical in shape, though sometimes spherical. A spore may be formed in the middle of the organism without (Fig. 28) or with (Fig. 29) a change in size of the cell around it. If the diameter through the cell is increased, then the cell with the contained spore becomes spindle-shaped. Such a cell is termed a “clostridium.” Sometimes the spore develops in the end of the cell either without (Fig. 30) or with enlarging it (Fig. 31). In a few forms the spore is placed at the end of the rod and shows a marked enlargement. This is spoken of as the “plectridium” or more commonly the “drumstick spore” (Fig. 32). The position and shape of the spore are constant for each kind of bacteria. In one or two instances only, two spores have been observed in a single organism.

Fig. 30.—Spores in the end of the rod with no enlargement of the rod around them. The lighter areas in the rods are spores.

Fig. 31.—Spores in the end of the rod with enlargement of the rod, A, A, A, A.

Fig. 32.—Drumstick spores at the end of the rod.

The fact that the protoplasm is denser and the spore capsule thicker (the percentage of water in each is decidedly less than in the growing cell) gives the spore the property of much greater resistance to all destructive agencies than the active bacterium has. For example, all actively growing cells are destroyed by boiling in a very few minutes, while some spores require several hours’ boiling. The same relation holds with regard to drying, the action of chemicals, light, etc. That the coagulation temperature of a protein varies inversely with the amount of water, it contains, is shown by the following table from Frost and McCampbell, “General Bacteriology”:

Egg albumin	plus	50 per cent. water	coagulates at	56°
Egg albumin	plus	25 per cent. water	coagulates at	74–80°
Egg albumin	plus	18 per cent. water	coagulates at	88–90°
Egg albumin	plus	6 per cent. water	coagulates at	145°
Egg albumin	dry	water	coagulates at	160–170°

This resistance explains why it happens that food materials boiled and sealed in cans to prevent the entrance of organisms sometimes spoil. The spores have not been killed by the boiling. It explains also in part the persistence of some diseases like anthrax and black leg in pastures for years. From the above description it follows that the spore is to be considered as a condensation of the bacterial protoplasm surrounded by an especially thick cell wall. Its function is the preservation of the organism under adverse conditions. It corresponds most closely to the encystment of certain protozoa—the ameba for example. Possibly the spore represents a very rudimentary beginning of a reproductive function such as is gradually evolved in the higher thread bacteria, the fission yeasts, the yeasts, the molds, etc. Its characteristics are so markedly different, however, that the function of preservation is certainly the main one.

It must not be supposed that spores are formed under adverse conditions only, because bacteria showing vigorous growth frequently form spores rapidly. Special conditions are necessary for their formation just as they are for the growth and other functions of bacteria (Chapters VI and VII).