The Spirochætes

The Spirochætes are long, narrow, wavy, thread-like organisms, with a firm yet flexible outer covering or periplast. There is a diffuse nucleus internally in the form of bars or rodlets of chromatin distributed along the body. In some forms there is a membrane or crista present (fig. 53), which in the past was compared with the undulating membrane of a trypanosome, but the membrane of a spirochæte does not undulate. Progression is very rapid, corkscrew-like and undulatory movements occurring simultaneously.

The genus Spirochæta was founded by Ehrenberg in 1833 for an organism which he discovered in stagnant water in Berlin. Ehrenberg named the organism Spirochæta plicatilis. According to Zuelzer (1912) S. plicatilis does not possess a membrane or crista, but an axial filament. S. gigantea has been described by Warming from sea-water.

Spirochætes occur in the crystalline style and digestive tract of many bivalve molluscs. The first molluscan spirochæte to be studied was that of the oyster, named by Certes (1882) “Trypanosoma” balbianii (fig. 53). Similar spirochætes, probably belonging to the same species, occur in various species of Tapes and in Pecten (the scallop). S. balbianii has rounded ends (fig. 53). Other spirochætes occur in freshwater mussels (Anodonta spp). S. anodontæ, studied by Keysselitz (1906) and by Fantham (1907), has pointed ends. Gross (1911) suggested the generic name Cristispira for molluscan spirochætes, because they possess a well-marked membrane or “crista,” which appears to be absent from S. plicatilis, according to Zuelzer’s researches.

Schaudinn in 1905 founded the genus Treponema for the parasite of syphilis (T. pallidum), discovered by him and by Hoffmann. According to Schaudinn the Treponemata have no membrane or crista. The pathogenic agent of yaws or frambœsia, discovered by Castellani, is also placed in the genus Treponema, as T. pertenue.

There remain the blood spirochætes. It is somewhat disputed as to whether these organisms possess a membrane. The present writer considers that they have a slight membrane or crista. The name of the genus in which to place the blood-inhabiting forms is somewhat uncertain and disputed. Various generic names given to them are Spirochæta, Treponema, Spiroschaudinnia (Sambon) and Borrelia (Swellengrebel). Included in this division are the causal agents of relapsing or recurrent fever. These Protists will be named, for description, Spirochætes without prejudice as to the ultimate correct generic name.

It is sometimes made a matter of argument as to whether the spirochætes are Protozoa or Bacteria. Such arguments are somewhat unprofitable. Morphologically the spirochætes are like the Bacteria in possessing a diffuse nucleus. They differ from Spirillum, an undoubted bacterial genus, in being flexible and not possessing flagella. Molluscan spirochætes, however, may appear to have flagella if their membrane becomes frayed or ruptured, when the myonemes therein (fig. 53), becoming separated, form apparent threads or flagella (Fantham, 1907–08).

Again, the mode of division of spirochætes has been used as a criterion of their bacterial or protozoal affinity. They have been stated to divide transversely, longitudinally, and by “incurvation,” or bending on themselves in the form of a U, “a form of transverse fission.” The present writer believes that they divide both transversely and longitudinally, and that there is a periodicity in their mode of division at first longitudinal (when there are few spirochætes in, say, the blood) and then transversely (when spirochætes are numerous in the blood). Some authors consider that longitudinal division is explained by “incurvation.”

The spirochætes of relapsing fever show a remarkable periodic increase and decrease in numbers in the blood. They are transmitted by ticks or by lice. They react to drugs (e.g., salvarsan or “606”) rather like trypanosomes, and—like Protozoa, but unlike Bacteria—they are cultivated with difficulty. These and other criteria have been used to endeavour to determine whether they are Protozoa or Bacteria. The present writer believes that they are intermediate in character, showing morphological affinities with the Bacteria and physiological and therapeutical affinities with the Protozoa. The group Spirochætacea, as an appendix to the Protozoa, has been created for them by the present writer (Jan., 1908). Others have placed them in the Spirochætoidea of the Bacteria or with the Spirillacea. Doflein (1909) called them Proflagellata. Further discussion is unnecessary, as they are undoubtedly Protista (see p. ).

There is no true conjugation, sex or encystment in spirochætes, but morphological variation may occur. They may agglomerate.

The Spirochætes form an interesting chapter in the evolution of parasites. There are free living forms, parasitic forms in the guts of both vertebrates and invertebrates, and blood-inhabiting forms. These probably represent the order of evolution of parasitism. The blood-inhabiting forms are pathogenic to warm-blooded hosts.

We must now consider the blood Spirochætes and the Treponemata (organisms of syphilis and of yaws).

THE SPIROCHÆTES OF THE BLOOD.

There are at least two important human parasites included hereunder:—

(a) Spirochæta recurrentis (=S. obermeieri), (b) Spirochæta duttoni.

More is known of the life-cycle of Spirochæta duttoni, and it will be convenient to consider that first.

Spirochæta duttoni, Novy and Knapp, 1906.

The specific name duttoni was also given, independently, to this parasite in 1906 by Breinl and Kinghorn.

S. duttoni is the pathogenic agent of African tick fever in man, prevalent in the Congo State and other parts of Africa. The full-grown organism is about 16 µ to 24 µ long, and has pointed ends. It is 0·25 µ to 0·5 µ broad. P. H. Ross and Nabarro were among the earliest to see a spirochæte in the blood of patients in Uganda. It is transmitted by the tick, Ornithodorus moubata.

In the blood of the patient some of the spirochætes may show, after staining, lighter and darker portions (chromatin dots) and evidence of the possession of a very narrow membrane (fig. 54). The mode of division has already been discussed. Periodicity in the direction of division was first described by Fantham and Porter, (1909). Just before the crisis in African tick fever, Breinl has stated that S. duttoni becomes thinner in the spleen and bone-marrow and rolls up into skein-like forms, which are surrounded by a thin “cyst” wall (probably the periplast). Such occur in apyrexial periods. Inside the cyst the spirochæte breaks up into granules. Balfour and Sambon have described somewhat similar rolled up forms, breaking into granules, inside the red blood cells of Sudanese fowls in the case of S. granulosa (possibly only a variety of S. gallinarum). The intracorpuscular stage is not definitely established.

The granule phase, however, is an essential one in the invertebrate transmitter (fig. 54c). In 1905, Dutton and Todd proved experimentally that O. moubata transmitted S. duttoni. They fed ticks, obtained from Congo native huts in which infected persons lived, on monkeys and the latter became infected. Dutton and Todd also found the offspring of infected ticks to be capable of transmitting the infection to experimental animals. They concluded that O. moubata was a true intermediate host.

A little later in 1905, Koch stated that spirochætes from the gut of the tick penetrated the gut wall and tissues and found their way into the eggs in the ovary. Koch figured tangled masses of spirochætes as occurring in the tick eggs. He found ticks infective to the third generation. He thought that the infection was spread by the salivary fluid of the tick, in the act of biting. (This is now known to be incorrect.) Markham Carter (1907) corroborated Koch’s work on the spirochætes in the tick eggs, and they have been seen since by Kleine and Eckard (1913).

Sir William Leishman, in 1909–10, found that at ordinary temperatures the salivary glands of infected ticks (O. moubata) were not themselves infective, and that the infection occurred by way of the ticks’ excretion. The spirochætes (contained in the ticks’ excrement) found their way into the vertebrate host through the wound made by biting. While feeding, ticks pass large quantities of clear fluid from the coxal glands; in this fluid an anticoagulin occurs. Some of the ticks also pass thick, white Malpighian secretion, that is, excrement, towards the end of the feed. Leishman, using experimental monkeys, showed that if infected ticks were interrupted while feeding, then no infection resulted in the monkeys. If, however, the ticks were allowed to finish their feed, and the Malpighian secretions were passed, then the experimental monkeys became infected. Fantham and Hindle (1911), independently, have repeated the experiments with mice.

Leishman’s methods and results may be summarized thus: Saline emulsions of the organs of infected ticks were made, after the organs had been most carefully dissected out. The ticks were first kept for several days at certain constant temperatures, such as 24° to 25° C. or blood heat, 37° C. The saline emulsions of the organs were inoculated, separately, into experimental animals, and the results recorded:—

At 24° C. At 37° C.

Salivary glands Negative Positive

Malpighian tubules Positive Positive

Gut and contents Positive Positive

Excrement Positive Positive

Genital organs Positive Positive

Coxal fluid is usually negative; thick, white excrement from Malpighian tubes is positive.

When the ticks were incubated at 21° to 24° C. no spirochætes, as such, were seen in the organs, except perhaps in the gut, where they often disappeared in a few days. When the ticks were previously incubated at 35° to 37° C. for two to three days, spirochætes, as such, reappear in the gut, organs and hæmocœlic fluid. The infection proceeds, not from the salivary gland, but from the infective excrement, that is, from the thick, white material voided by the tick while feeding, usually towards the end of the meal. This Malpighian excrement passes into the wound caused by the bite, being greatly aided by the clear and more limpid coxa fluid, which bathes the under surface of the tick’s body, and mixes with and carries the infective excrement into the wound. Ticks remain infective for a long time.

The spirochætes in the gut of infected ticks divide by a process of multiple transverse fission into granules, which are composed of chromatin (fig. ). These granules—sometimes known as coccoid bodies—are capable of multiplication. Leishman first found them in clumps inside the cells of the Malpighian tubules (cf. fig. 55).

To summarize, when spirochætes are ingested by a tick, some of them pass through the gut-wall into the hæmocœlic (body) fluid. They then bore their way into the cells of various organs (fig. 55a) and break up into coccoid bodies. In this manner the granules find their way into the ovaries and ova, thus explaining how the young ticks are born infected. Inoculation of these chromatinic granules usually produces infection. Infective granules are also seen in the rudiments of the Malpighian tubules of embryo ticks. Bosanquet and Fantham (1911), independently, have shown that molluscan spirochætes also break up into similar granules or coccoid bodies. Gross has also demonstrated multiple transverse fission in molluscan forms. Marchoux and Couvy (1913) and Wolbach (1914) consider the granules or coccoid bodies to be degeneration products. This is unlikely (see below).

Schuberg and Manteufel have found that certain O. moubata, perhaps 30 per cent. of the specimens of a given neighbourhood, may acquire a natural active immunity against infection with S. duttoni.

S. duttoni, or a closely allied form (by some termed S. novyi), occurs in Colombia, and is spread by the tick Ornithodorus turicata. In Panama a similar spirochæte is probably spread by O. talaje.

Spirochæta gallinarum, Stephens and Christophers, 1905
(= Spirochæta marchouxi, Nuttall, 1905).

This Spirochæte, which occurs in fowls and is pathogenic, is transmitted by the tick Argas persicus. It is about 10 µ to 20 µ long. There is a pathogenic spirochæte known to occur in geese, named by Sakharoff (1891) S. anserina, and found in Caucasia. This may be the same as S. gallinarum, in which case the name S. anserina will have priority. These organisms cause fever, diarrhœa, anæmia and death. The life history of the avian pathogenic spirochætes has been studied by Balfour, by Hindle and by Fantham. It is essentially similar to that of S. duttoni.

Marchoux and Couvy (1913) consider that the “fragmentation of the chromatin” in spirochætes is a process of degeneration. Working with A. persicus and S. gallinarum, they state that a large number of the spirochætes ingested by the Argas almost immediately pass through the wall of the alimentary canal and appear in the hæmocœlic fluid. Marchoux and Couvy consider that Leishman’s granules may be found in the Malpighian tubules of various Arachnids. They found spirochætes in the cephalic glands of infected Argas. They consider that spirochætes remain as wavy spirochætes within the tick, if they are to be infective, though the spirochætes may become so thin as to be invisible! The latter argument is obviously weak, and it was never asserted that all granules in the Malpighian tubules of infected ticks were derived from spirochætes. With dark-ground illumination small, refractile spirochætal granules may be seen to grow into spirochætes. The granule phase of spirochætes has recently been discussed by Fantham (1914).

Spirochæta recurrentis, Lebert, 1874.

Syn.: Spirochæta obermeieri, Cohn, 1875.

This organism was discovered by Obermeier (1873) in cases of relapsing fever in Berlin. Short forms 7 µ to 9 µ long, and longer (probably adult) forms, 16 µ to 19 µ, are found in the blood. The width is 0·25 µ. Parasites 12 µ or 13 µ long are often observed.

The spirochæte is found in the blood during febrile attacks and relapses, but not during intervening periods. It can be inoculated into monkeys, rats and mice. It can live in the bed-bug, Cimex lectularius, and Nuttall has succeeded in transmitting S. recurrentis from mouse to mouse by the bites of the same bug. The French investigators Sergent and Foley (1908–9) in Algeria, and Nicolle, Blaizot and Conseil (1912) in Tunis, have shown experimentally that S. recurrentis (var. berbera) is transmitted by lice. The latter workers also demonstrated the method of infection that commonly occurs, namely, by the scratching of the skin and crushing of lice containing spirochætes on the excoriated surface of the body.

Lice as transmitting agents for relapsing fever were indicated by Mackie in 1907. An epidemic among Indian school children furnished the materials. It was noted that out of 170 boys, 137 were infected, and the boys were very verminous. Among the girls, 35 out of 114 suffered, and few lice were found on them. Twenty-four per cent. of the lice taken from the boys contained spirochætes as compared with 3 per cent. of those from the girls. As the epidemic died out among the boys, the lice also became fewer, and an increase in the number of cases among the girls coincided with an increase in the number of lice. Spirochætes were found in the gut, Malpighian tubules and genital organs of the lice. Mackie thought that infection of the patients was brought about by the regurgitation of the spirochætes when the lice fed, but proof of this was lacking.

In 1912, Nicolle, Blaizot and Conseil, working in Tunis and using chiefly an Algerian strain of relapsing fever spirochætes (sometimes called S. berbera), showed by direct experiments that infection by means of the bites of Pediculus vestimenti and P. capitis was untenable. As many as 4,707 infected lice were fed on one man, and 6,515 on another occasion were allowed to bite a man after they had fed on a monkey heavily infected with spirochætes, yet no infection of the man followed. Examination of the lice showed that the spirochætes left the gut soon after they were ingested, and passed into the body cavity, which swarmed with spirochætes. The contents of the alimentary tract and the fæces of the lice alike were uninfective. The spirochætes did not reappear in the gut till eight days after an infective feed, but some persisted as late as the nineteenth day when kept at 28° C.

It was noted that the irritation due to the lice caused scratching, and that thereby lice became crushed on to the skin. An emulsion was made of two infected lice and rubbed on to the slightly excoriated skin of one of the above workers. Infection followed five days later. A drop of emulsion placed on the conjunctiva of the human eye produced spirochætosis after an incubation of seven days. The body contents of such lice, then, produce infection when they reach the blood by any excoriated or penetrable surface. The stages leading up to infection in nature briefly are: The irritation due to the louse bites causes scratching, and the lice are crushed on to the skin. The slight abrasion is quite sufficient to permit the entry of the parasite. The louse bite alone is harmless. Infection by way of the eye is quite probable in Africa, remembering the constant trouble due to sand, dust, insects, etc., resulting in frequent touching of the eyes.

The spirochætes occur in the body fluid of the lice and can pass in it to the adjacent organs. Thus they probably find their way into the genital organs, and into the eggs of the lice. Eggs laid twenty to thirty days after the parent became infected have retained the infection, and the larvæ issuing from such eggs must have contained some form of spirochætes, for an emulsion of either the eggs or the larvæ produced spirochætosis when inoculated into monkeys. Further details regarding the spirochætosis in the eggs of the lice and in the larvæ are needed. Hereditary infection, however, has been demonstrated, but is not very common. Sergent and Foley (1914) state that the spirochæte possesses a very small and virulent form which it assumes during apyrexial periods in man and during a period following an infecting meal in the louse. Nicolle and Blanc (1914) find that the organisms are infective in the louse just before they reappear as spirochætes. Nicolle and Blaizot found that female lice were more susceptible to spirochætes than males, four times as many females as males being infected.

Tictin (1897) found S. recurrentis in bugs recently fed on patients, and infected a monkey with the fluids of crushed bugs. Karlinski (1902) found the spirochætes in bed-bugs in infected houses. There is some other evidence to show that bugs may transmit the spirochæte in Nature. Further researches are needed regarding the relationship of bed-bugs and human spirochætosis.

Multiplication of S. recurrentis is by longitudinal and transverse division (including so-called “incurvation”), and the organism forms small, ovoid bodies (“coccoid” bodies) in the same way as S. duttoni.

S. recurrentis is the cause of European relapsing fever, and a number of possible varieties of it are associated with relapsing fevers in other parts of the world. Such spirochætes only differ by biological reactions, such as acquired immunity tests. They include:—

S. rossii, the agent of East African relapsing fever; S. novyi, the agent of North American relapsing fever; S. carteri, the agent of Indian relapsing fever; S. berbera, the agent of North African and Egyptian relapsing fever.

Other Human Spirochætes are:—

S. schaudinni. This organism, according to Prowazek, is the agent of ulcus tropicum. It varies in length from 10 µ to 20 µ.

S. aboriginalis has been found in cases of granuloma inguinale in British New Guinea and Western Australia. It also occurs in dogs, and may not be truly parasitic.

S. vincenti. This spirochæte is 12 µ to 25 µ in length, tapers at both ends and has few coils. It has been associated with angina vincenti. It often occurs in company with fusiform bacilli.

S. bronchialis, found by Castellani in 1907 in cases of bronchitis in Ceylon. The parasites are delicate, but show morphological variation. This organism is important and has since been found in the West Indies, India, Philippine Islands and various parts of Africa, such as the Anglo-Egyptian Sudan, Uganda and West Africa. It has recently been the subject of research by Chalmers and O’Farrell, Taylor, and Fantham.

S. phagedenis was found by Noguchi in a ten days old ulcerated swelling of the labium. The organism shows much variation in size, being 4 µ to 30 µ in length.

S. refringens (Schaudinn, 1905) occurs in association with Treponema pallidum in syphilitic lesions, but is non-pathogenic. It is 20 µ to 35 µ long and 0·5 µ to 0·75 µ broad, being larger than T. pallidum and more easily stained.

Various spirochætes have also been notified in vomits, chiefly in Australia; others from the human intestinal tract, e.g., S. eurygyrata; S. stenogyrata (Werner); S. hachaizæ (Kowalski), in cholera motions; S. buccalis (Cohn, 1875) and S. dentium occurring in the human mouth and in carious teeth (S. dentium, Koch, 1877, being the smaller); S. acuminata and S. obtusa found by Castellani in open sores in cases of yaws.

Animal spirochætes of economic importance include:—

S. anserina, highly pathogenic to geese.

S. gallinarum (= S. marchouxi) in fowls. (See p. .)

S. theileri in cattle and S. ovina in sheep also occur in Africa; their pathogenicity is not clear.

S. laverani (= S. muris), occurring in the blood of and pathogenic to mice, is probably the smallest spirochæte from the blood, being only 3 µ to 6 µ long.

Numerous spirochætes have been recorded from the guts of various mammals, birds, fishes, amphibia and insects.

Cultivation of Spirochætes.—Cultures of spirochætes have been made with little success or with great difficulty until comparatively recently, when Noguchi (1912) devised a means whereby he has cultivated most of the pathogenic spirochætes as well as some Treponemata.

Noguchi has now cultivated S. duttoni, S. recurrentis, S. rossii, S. novyi and S. gallinarum from the blood; S. phagedenis from human phagedænic lesions; S. refringens and spirochætes from the teeth.

His method is as follows:—

A piece of fresh, sterile tissue, usually rabbit kidney, is placed in a sterile test-tube. A few drops of citrated blood from the heart of an infected animal, e.g., rat or mouse, is added, and about 15 c.c. of sterile ascitic or hydrocœle fluid is poured quickly into the tube. Some of the tubes are covered with a layer of sterile paraffin oil, others are left uncovered. The tubes are incubated at 37° C. The best results are obtained if the blood is taken from an animal forty-eight to seventy-two hours after it has been inoculated, that is, before the spirochætes reach their maximum multiplicative period in the blood. The presence of some oxygen seems indispensable for these blood spirochætes, and they fail to develop in vacuo or in an atmosphere of hydrogen.

For subcultures, 0·5 c.c. of a culture is added to the medium instead of citrated blood, and it is useful to add a little fresh, normal blood, either human or from an animal, such as a rat.

Noguchi found that the events in cultures were:—

S. duttoni, maximum multiplication on the eighth to ninth day; disintegration beginning on the tenth day, spirochætes disappeared after about the fifteenth day. No diminution of virulence was found at the ninth day.

S. rossii (= S. kochi). Maximum development on the ninth day, after which the virulence diminishes. The incubation period is also prolonged.

S. recurrentis (= S. obermeieri). Maximum growth on the seventh day.

S. novyi.—Maximum development on the seventh day. It is more difficult to grow than the preceding forms.

All the above spirochætes showed undoubted longitudinal division and transverse division was observed in part.

S. gallinarum can be cultivated as above, but transverse division was usual here. Maximum growth occurred in the culture about the fifth day.