Genus Babesia

In this genus the trophozoites multiply by binary fission in the erythrocytes, forming pairs, or by schizogony, forming tetrads. A "blepharoplast" from which a rhizoplast arises has been described in the trophozoites. However, Rudzinska and Trager (1960) did not report seeing either of these structures in electron micrographs of Babesia rodhaini from the mouse.

If present, the blepharoplast and rhizoplast may betray a flagellate origin for the group (Dennis, 1932). However, Reichenow (1953) thought that it originated from the amoebae. Another possibility is that it is related to Plasmodium. This is suggested by Rudzinska and Trager's (1960) finding in B. rodhaini of structures composed of concentric membranes (possibly representing primitive mitochondria) similar to those they had previously seen in Plasmodium berghei, and by their observation that B. rodhaini apparently engulfs bits of host cell cytoplasm by phagotrophy like Plasmodium.

There are two opposing schools of thought as to the speciation of this genus. One breaks it up into several genera or subgenera, each with a number of species (e.g., Sergent et al., 1945; Antipin et al., 1959), while the other prefers a single genus with a relatively small number of species, each of which may include several strains (e.g., Wenyon, 1926; Neitz, 1956). The second system seems preferable. The taxonomy of the group has been discussed by Reichenow (1953), Poisson (1953) and Laird and Lari (1957) in addition to the above authors.

Synonyms of Babesia include Piroplasma, Achromaticus, Nicollia, Nuttallia, Smithia, Rossiella, Microbabesia, Babesiella, Francaiella, Luhsia, Pattonella, Rangelia, and Gonderia in part.

For practical purposes, one can divide the genus into 2 groups of species, large forms more than 3 u long and small forms less than 3 u long. In general, infections with the large forms can be successfully treated with trypan blue, while infections with the small ones cannot.

Babesia and babesiosis occur in most parts of the world where there are ticks, except in countries such as the United States where they have been wiped out by a concerted effort. They are most important in the tropics, where, along with the trypanosomoses, they often dominate the livestock disease picture. However, they also occur in the temperate zone. Bovine babesiosis nearly reaches the Artie circle in Norway, and Thambs-Lyche (1943) reported that it was increasing in that country.

Babesiosis was once an extremely important disease of cattle in the United States, but it has now been eliminated, and the only domestic animal species left in this country is B. canis, which occurs in dogs in Florida, Virginia and Texas. However, Babesia is still important in livestock in Central and South America. It occurs in most of Europe, being especially important in the countries bordering the Mediterranean Ocean. It is one of the most important diseases of livestock in the Middle East, thruout Africa, and also in parts of India and the Far East. Its importance in the USSR, and especially in its southern part, is attested by the fact that 61% of the protozoan section of Antipin et al/s (1959) textbook on veterinary parasitology is devoted to it and a related disease, theileriosis. It also occurs in Australia.

Veterinarians and livestock owners in the United States today do not know what it is to have to contend with babesiosis, but other parts of the world are not so fortunate. The disease is of great economic importance in the tropics and subtropics; indeed, Curasson (1943) believed that it was no exaggeration to say that the babesioses are the most formidable diseases of livestock in these regions and that they are taking a more and more important place in the animal disease picture as we discover new manifestations of their activity.

Among recent discussions of babesiosis and its manifestations are those by Curasson (1943), Sergent et al. (1945), Muromtseva and Dobrokhotova (1955), Henning (1956), Malherbe (1956) and Antipin et al. (1959).

Life Cycle

The trophozoites of Babesia occur in the erythrocytes, where they multiply by binary fission or by schizogony. In some species, two trophozoites are formed, which break out of the erythrocytes and enter new red cells, while in others tetrads composed of 4 trophozoites are formed. Some authors place the latter in a separate genus, Nuttallia. The formation of more than 4 trophozoites by schizogony has also been described in the erythrocytes (Dschunkowsky, 1937; Ivanic, 1942; Delpy, 1946), but most workers (e.g., Reichenow, 1953) consider that it is merely simulated by repeated binary fissions or by multiple invasion of a host cell.

The above asexual cycle continues indefinitely, the animals sometimes remaining infected for life.

Babesia is transmitted by ticks. The discovery of this fact - by Smith and Kilborne (1893) for B. bigemina of cattle - was a milestone in the history of parasitology, since it was the first demonstration that an arthropod was the vector of any disease.

Dennis (1932) described sexual reproduction of B. bigemina from cattle in the tick, Boophilus annulatus, and Petrov (1941) did the same for B. bovis in Ixodes ricinus. However, Regendanz and Reichenow (1933) denied its existence in the life cycle of B. canis from the dog in Dermacentor reticulatus, and Regendanz (1936) and Muratov and Kheisin (1959) found no evidence of sexual reproduction in B. bigemina in Boophilus microplus and B. calcaratus, respectively, nor could Polyanskii and Kheisin (1959) for B. bovis in Ixodes ricinus. It is likely that Dennis and Petrov may have been misled by trying to draw an analogy with the life cycle of Plasmodium. Pending final settlement of the question, however, both accounts are given below.

According to Regendanz and Reichenow (1933), most of the B. canis ingested by the female tick die in its intestine. Some of them become vermiform and enter the intestinal epithelial cells, coming to lie against the basal membrane, and grow into large amoeboid forms. These then multiply by a series of binary fissions, producing more than 1000 individuals in 2 to 3 days. These lie together loosely at first, but finally fill the whole host cell. They then become vermiform and pass into the body cavity.

The vermiform stages are broadly rounded at the anterior end and pointed posteriorly, about 16 u long, and have a gliding motion. They enter the ovary, where they penetrate the eggs. Here they round up and divide a few times, forming very small round individuals. They do not develop further in the larval tick which hatches from the egg, but when it molts they enter the salivary glands and continue their development. This first occurs in the nymphal stage, but is much more active in the adults, both male and female. The parasites undergo a series of binary fissions and enter the cells of the glandular acini. Here they multiply further, becoming smaller and filling the whole host cell, so that it finally contains thousands of minute parasites. These become vermiform, break out of the host cell, come to lie in the lumen of the gland, and are injected into the host when the tick sucks blood. The developmental process in the salivary glands takes 2 to 3 days.

The tick larvae are not able to infect new hosts. The nymphs may do so, but generally it takes so long for the parasites to reach the salivary glands that most transmission, in this species at least, is by the adults.

Regendanz (1936) found that the development of Babesia bigemina from cattle in the intestinal wall of Boophilus microplus corresponded completely with that of B. canis in Dermacentor reticulatus. After numerous binary fissions, the protozoa turn into the motile, vermiform stage and enter the developing eggs of the female tick, where development continues. He found no evidence of sexual stages.

Bovine species of Babesia in avian erythrocytes

Muratov and Kheisin (1959) described a similar process for B. bigemina in Boophilus calcaratus, except that they said that schizogony occurs. They studied only the stages in the females and their eggs. On the day after the tick drops from its host, the protozoa begin to reproduce in its intestine by binary fission or by schizogony, producing club-shaped forms which penetrate into the epithelial cells of the intestine. Here they develop and undergo atypical multiple fission, characterized by asynchronous segmentation, into amoeboid or round agamonts. These become club- or cigar-shaped, penetrate other intestinal cells and repeat the asexual cycle. The dividing stages in the intestinal cells are up to 30 to 45 u in diameter and produce about 250 daughter parasites.

Some of the club-shaped stages enter the body cavity and divide further. They penetrate all the organs of the female, including the ovary, and continue to divide. In the ovary they enter the eggs and divide by binary or multiple fission just as in the intestine, producing round or amoeboid agamonts which turn into club-shaped stages. Their number increases as the eggs develop, and they are distributed thruout the organs of the developing larvae. Muratov and Kheisin found no evidence of copulation or sexual reproduction.

Polyanskii and Kheisin (1959) found essentially the same pattern for B. bovis in Ixodes ricinus. They said that it reproduces by binary fission or schizogony in the tissues of the tick and in the eggs of infected females, and found no stages of sexual reproduction or sporogony.

Quite a different process was described by Dennis (1932) for B. bigemina in Boophilus annulatus. According to him, when a female tick ingests blood, most of the parasites in the blood are destroyed, but some of them turn into vermiform bodies about 6 u long which he considered to be gametes and which he called isogametes because they all look alike. They move actively by bending or gliding. Two of them unite to form a motile, club-shaped zygote or ookinete 7 to 12 u long. The ookinetes pass thru the intestinal wall, enter the ovaries and then the eggs. Here they round up to form sporonts 7.5 to 12 u in diameter. The sporonts grow, and then divide by multiple fission into 4 to 32 amoeboid sporoblasts. The nuclei of the sporoblasts divide repeatedly, forming small, multinucleate, amoeboid sporokinetes which are distributed thruout the tissues of the developing tick embryo. The sporokinetes vary in shape, being round, elongate, club- or ribbon-shaped, and may be as much as 15 u long. They contain a varying number of granular nuclei 0.4 u in diameter. In the course of the embryonal development of the tick, the sporokinetes multiply, probably by plasmotomy. All the tissues of the tick may be invaded, and sometimes the cytoplasm of a host cell is almost entirely supplanted by the parasites, particularly in the salivary glands. Toward the end of the tick's development, some sporokinetes produce sporozoites, which are the infectious stage. Others produce them only after the larva has hatched. The sporozoites resemble minature trophozoites; they are piriform and have a blepharoplast. They are particularly numerous in the salivary glands, in the coelenchymatous tissue at the base of the legs and around the viscera. They are inoculated into the blood with the saliva when the tick feeds.

Petrov (1941) described a similar process for B. bovis in Ixodes ricinus. According to him, the isogametes fuse in the tick's intestine to form an ookinete which passes thru the intestinal wall and enters a developing ovum. Here it rounds up, forms sporoblasts, and these in turn form sporozoites which pass to the salivary glands. The larvae, nymphs and adults of the succeeding generation can all transmit the parasite.

It should be said that Reichenow (1953) considered some of the stages described by Dennis to be normal, intracellular symbionts (cf. Buchner, 1953; Koch, 1956) rather than Babesia.

In the life cycles described above, the adult tick picks up the infection, but does not transmit it. This is done by the next generation. Babesia can also be transmitted by different stages in the same generation; it can be picked up by a larval tick and transmitted by the nymph, or it can be picked up by the nymph and transmitted by the adult. The occurrence of this stage-to-stage transmission depends upon both the species of Babesia and the species of tick. Neitz (1956) has assembled information on this subject, and it is given below in the discussion of the individual species.

The life cycle in the tick in stage-to-stage transmission was studied carefully by Shortt (1936) for B. vogeli (B. canis) of the dog in Rhipicephalus sanguineus in India. He saw no evidence of sexual stages. After the nymph has taken a blood meal, the parasites do not multiply in the gut epithelium, but in the phagocytes next to the hypodermis in the body cavity. Here they reproduce by multiple fission to form what Shortt called pseudocysts - clumps of up to 200 organisms contained within the envelope of the parasitized host cell. These are fully developed about 7 days after the nymph has left its host. They are 14 to 35 u in diameter. The stages within these pseudocysts are at first more or less spherical and 1.7 to 3.3 u in diameter. They become club-shaped in 4 to 8 more days, at which time they measure about 9 by 2 u. The club-shaped stages then break out of the host cell and migrate to the muscles and muscle-sheaths. They penetrate the cells, round up, and divide by repeated binary fissions to form a large number of relatively small, ovoid or slightly elongate parasites about 1.2 u long. This stage is reached about 20 days or more after the nymph has fed. This phase of the life cycle corresponds to that which takes place in the eggs of the adult.

The muscles remain unchanged during metamorphosis. When the adults begin to feed on a dog, the parasites migrate to the salivary glands and enter their cells. Development then continues as described by Reichenow and Regendanz (1933) for B. canis. The parasites multiply by repeated binary fissions to form large numbers of spherical or ovoid infective stages about 1.9 u or less in diameter.

Pathogenesis

Babesiosis is a highly pathogenic disease in most hosts. It is unusual in that the death rate is much higher in adults than in young animals.

The various species of Babesia cause a similar disease in different hosts. In most cases there are fever, malaise and listlessness. Affected animals do not eat, or eat little. There is severe anemia, and destruction of the erythrocytes is accompanied by hemoglobinuria. The mucous membranes become pale, and icterus develops. The spleen is greatly enlarged, with soft, dark red pulp and prominent splenic corpuscles. The liver is enlarged and yellowish brown. The lungs may be slightly edematous. There may be diarrhea or constipation, and the feces are yellow except in very early or peracute cases. Affected animals lose condition, become emaciated, and often die.

The signs of babesiosis may vary markedly from this typical picture, however. As Malherbe (1956) said, "Anybody with extensive experience of these diseases ... is forcibly struck by the deviate and protean manifestations of the disease picture as it is encountered from time to time. There is almost no guise under which the disease does not masquerade at some time or another, and it is therefore no accident that the majority of South African veterinarians have a pronounced attachment to their microscopes". Malherbe remarked on the similarity of the clinical and pathological manifestations of babesiosis to those of malaria, stating that "in spite of the differences in the life cycle of the parasites, their effect on the body is capable of exactly similar potentialities".

Death, if it occurs, is due to organic failure which, in turn, is due not only to the destruction of erythrocytes with resultant anemia, edema and icterus, but also to the clogging of the capillaries of various organs by parasitized cells and free parasites (Malherbe and Parkin, 1951; Malherbe, 1956). The stasis resulting from this sludging (Knisely el al., 1947) causes degeneration of the endothelial cells of the small blood vessels, anoxia, accumulation of toxic metabolic products, capillary fragility, and eventually perivascular escape of erythrocytes and macroscopic hemorrhage. Purpura may result, the great majority of such cases in dogs being due to babesiosis. The signs of the disease depend in part on the location where the most serious stasis takes place. Cerebral babesiosis similar to cerebral falciparum malaria may occur. Gilles, Maegraith and Andrews (1953) described liver damage in B. canis infections, beginning with early damming of the blood in the sinusoids around the central vein, thru centrilobular atrophy and degeneration of the hepatic cells, to necrosis of the cells. Kidney damage is also present.

Immunity

Cattle which have recovered from an attack of babesiosis due to B. bigemina remain infected for life, and are immune to reinfection. This type of immunity, due to continuing low-grade infection, is known as premunition. Premunition in cattle due to species other than B. bigemina, and in sheep, swine and dogs, lasts up to 2 years. Premunized animals do not show signs of disease except under stress of one sort or another. For instance, an attack of foot and mouth disease may reactivate babesiosis in cattle, and distemper may do the same in dogs.

The spleen plays an important role in maintaining immunity, and it is a common observation that splenectomy is often followed by a severe or fatal relapse in premunized animals. In addition, splenectomized animals are much more susceptible to infection with Babesia and much more seriously affected than normal ones.

Calves, foals, young pigs and kids are much less seriously affected by babesiosis than are adult animals. This is the reason that cattle can often be raised in highly endemic areas without being seriously affected, whereas imported animals usually die. The native cattle were infected as calves and are premunized. Lambs and puppies, however, are highly susceptible. There is no cross-immunity between the different species of Babesia.

Treatment

The treatment of babesiosis has been reviewed by Goodwin and Rollo (1955), Carmichael (1956) and Richardson and Kendall (1957), among others. There is an interesting relationship between the chemotherapy of babesiosis and that of trypanosomosis. Many of the compounds effective against Trypanosoma are also effective against Babesia. This may perhaps indicate a phylogenetic relationship, but I hasten to warn that a similar line of reasoning was once used to suggest a relationship between the trypanosomes and the spirochetes.

Nuttall and Hadwen (1909) introduced the first effective drugs, the azonaphthalene dyes, trypan red and trypan blue. The latter is still used in some areas. It is the sodium salt of ditolyl diazo-bis-8-amino-1-naphthol-3, 6-disulfonic acid. It must be given intravenously, since abscessation and sloughing follow subcutaneous injection. It stains the tissues blue-green for several months after injection. It does not eliminate all parasites, so that recovered animals are premunized.

The acridine derivative, acriflavine (trypaflavine, gonacrine, flavin, euflavin) was introduced by Stephan and Esquibel (1929). It is a mixture of 2, 8-diamino-10-methylacridinium chloride with a small amount of 2, 8-diaminoacridinium chloride. It, too, is still being used, especially against B. equi in South Africa and in cattle in North Africa. It does not eliminate all parasites, and recovered, treated animals are premunized.

The quinoline derivative, acaprin (Acapron, Pirevan, Babesan, Piroparv, Zothelone, Piroplasmin) was introduced by Kikuth (1935) and also by Carmichael (1935). It is 6, 6'-di-(N-methylquinolyl) urea dimethosulfate. It is administered subcutaneously. In large doses it eliminates all parasites, but in small ones it leaves some so that recovered animals are premunized (Kikuth, 1938). It affects the parasympathetic nervous system, and may cause alarming reactions, including salivation, vasodilation, sweating, copious urination, diarrhea, panting, a drop in blood pressure and even collapse and death. Adrenaline and calcium gluconate can be given as antidotes. To avoid such reactions, the drug is often given in 2 or 3 divided doses a few hours apart. Dogs are much more sensitive than cattle. Animals showing reactions usually recover rather quickly. Despite these reactions, acaprin is still one of the most widely used drugs for treating babesiosis in all animals thruout the world.

Lourie and York (1939) found that a number of aromatic diamidines were effective against Babesia. Adler and Tchernomoretz (1940) found that stilbamidine (4, 4'-diamidinostilbene) was effective against B. bigemina, and B. ovis, and it is also used for B. canis and B. caballi (Daubney and Hudson, 1941). Propamidine (4, 4'-diamidino-1, 3-diphenoxypropane) has been used against B. canis in dogs (Carmichael and Fiennes, 1941). Pentamidine (lomidine; 4,4'-diamidino-1, 5-diphenoxypentane) is used quite widely, especially in North Africa, for babesiosis in all animals. Phenamidine (4, 4'-diamidinodiphenyl ether) was introduced by Carmichael (1942) for canine babesiosis and is now used in cattle and other animals as well. Berenil (4, 4'-diamidino diazoaminobenzene diaceturate) was introduced by Bauer (1955), and is effective against babesiosis in cattle, dogs and other animals. Amicarbalide (M&B 5062A; 3, 3'-diamidinocarbanilide diisethionate) was introduced by Ashley, Berg and Lucas (1960). Preliminary studies indicate that it is effective against babesiosis in cattle (Beveridge, Thwaite and Shepherd, 1960; Lucas, 1960).

The diamidines are injected subcutaneously or intramuscularly, depending upon the compound. Many of them tend to cause a fall in blood pressure, but it soon returns to normal. Subcutaneous injection of concentrated solutions may cause irritation. Transitory swelling of the face and lips which is anaphylactic in nature sometimes occurs with phenamidine.

Prevention and Control

Since babesiosis is transmitted by ticks, prevention and control depend primarily on tick elimination. This can be done by regular dipping, which should be carried out on an area basis for livestock, at least. Dogs and riding horses can be treated individually.

Artificial premunization of young animals has been practiced with a good deal of success, especially in North Africa (Sergent et al., 1945). A mild strain of the organism is ordinarily used. This practice is not necessary if the animals are raised in an endemic area where they will all become naturally infected at an early age, but it is worthwhile in areas where only a certain proportion of the animals become infected or for animals which are destined to be shipped to endemic areas later on.