In the following notes no attempt has been made to enter into the pathology or etiology of the diseases described, as only isolated cases have been available for investigation.

I am much indebted to the Council of the Marine Biological Association of the United Kingdom for their kindness in granting me a table at their Plymouth Laboratory.

During the winter of 1908, when working in the Wellcome Laboratories at Khartoum, Dr Balfour proposed that I should examine a number of parasites both of plants and animals in order to ascertain if the former were infested with any other species of parasite; in other words, to study what has been termed hyper-parasitism.

In an earlier paper (1908, pp. 255–257), we gave a full account of a paper by Miyajima (1907) wherein this author described experiments in which he states that he succeeded in cultivating Theileria (Piroplasma) parva. According to Miyajima, he had no difficulty in cultivating the parasite when he added the blood of cattle (containing Theileria) to ordinary bouillon in the proportion of 1 : 5 to 1 : 10, the cultures being maintained at 20–30° C. Miyajima states that trypanosomes appeared in his cultures after an interval of 3 days and underwent vigorous multiplication reaching “the maximum after the tenth to fourteenth day.” Miyajima's description of the supposed process of development is obscure, and it is difficult to understand how the diminutive intra-corpuscular Theileria can develop into a Trypanosoma. We refer the reader who desires particulars regarding these experiments to our paper already cited. In our paper we stated that “a certain amount of scepticism” appeared justified until Miyajima's results had been extended and confirmed. The warning appears to have come too late in respect to Woodcock's recent review of the Haemoflagellates in Ray Lankester's Treatise on Zoology (1909, Part I. fasc. 1, p. 260), for Woodcock appears to accept Miyajima's remarkable discovery as authentic and conclusive.

All attempts hitherto made to infect different species of animals with P. canis have failed, and the remarkable fact remains that this parasite is apparently only communicable to dogs.

1. Regarding attempts at immunization by means of immune serum. The experiments of Nocard and Motas indicated that protective substances occur in the blood of dogs which have recovered from European piroplasmosis. These authors claimed that immune serum destroys the parasites and that it exerts a protective and curative effect upon the disease. Similar experiments conducted by Robertson in Cape Colony gave contrary results to those of Nocard and Motas. In the absence of further evidence no conclusions can therefore be drawn from these conflicting results. Two of Robertson's dogs which received injections of hyperimmune serum every day after they showed fever, died but no parasites could be found in their blood at death. This result suggests that a partial immunity may have been obtained by the treatment.

2. Regarding attempts at immunization by means of inoculations with blood containing dead parasites. Of the five dogs which we attempted to immunize in the manner we have described, three died of acute piroplasmosis; one died without parasites in its blood although parasites had been previously found; one dog died on the 36th day from chronic piroplasmosis. The absence of parasites in the blood of one dog at autopsy and the occurrence of chronic piroplasmosis in another dog may or may not indicate a partial acquisition of immunity. In any case the experiments afford no evidence that practical results are likely to follow further investigation of this character.

3. The duration of immunity following recovery is undetermined since there are no experiments to prove that animals are immune after the parasites have completely disappeared from their blood. In the socalled “immune,” “recovered,” or “salted” dogs, the animals harbour parasites in small numbers for weeks, months or years, consequently such dogs are suffering from a mild chronic form of piroplasmosis. Whilst subject to this mild form of the disease dogs usually escape acute infection when reinoculated with the parasite.

4. The parasites of piroplasmosis canis may persist in the blood of apparently recovered dogs for a considerable length of time, 6 months to 2 years, and so long as they are present in the blood the latter remains fully virulent for clean dogs. Consequently there is no evidence that the African Piroplasma canis becomes modified in its virulence during the course of chronic piroplasmosis. According to Nocard and Motas the European P. canis does become modified in its virulence in dogs suffering from chronic piroplasmosis. It should be noted that the European disease is a milder affection than the African.

5. The passage of the African P. canis through two series of dogs, that is through upwards of 90 animals in the course of two and a half years, has shown that the parasite may be communicated by inoculation from dog to dog for an indefinite period. No evidence was obtained that P. canis is in any way modified in its virulence by passage through dogs. In a number of dogs which were inoculated with post-mortem blood the disease developed more slowly than usual but nevertheless led to a fatal issue.

6. The onset of fever after inoculation with virulent blood may precede or succeed the appearance of P. canis in the peripheral circulation. Occasionally the disease may run its course to a fatal termination without the appearance of febrile symptoms.

1. Trypanblau injected subcutaneously into dogs a day before or a day after they have been inoculated with blood containing Piroplasma canis effectually prevents the development of piroplasmosis by destroying the parasites at the onset of infection.

2. Trypanblau given by the mouth is ineffective, since it exerts no apparent influence either upon the parasite or upon the course of the disease.

3. Tryparosan, when injected subcutaneously or when given by the mouth, has no effect upon the parasite and is ineffective as a remedy against piroplasmosis in the dog.

1. Trypanblau promises to be an efficient remedy for bovine piroplasmosis, since it exerts a direct and obvious effect upon the parasites.

2. The effect of the drug upon Piroplasma bovis is similar to that which it produces upon the canine parasite. The dividing forms are the first to disappear, and after a few hours the pyriform parasites also disappear from the peripheral circulation; the parasites which are detected in the blood after a few hours appear degenerated and rounded or irregular; within nine to 45 hours or less all the parasites have disappeared from the blood.

3. As in canine piroplasmosis the disappearance of the parasites from the blood may be temporary. The parasites also disappear and reappear in small numbers (after two to 11 days) in animals undergoing natural recovery. In three treated animals the parasites reappeared in exceedingly small numbers after five to six days; in two they had not reappeared after 16 and 18 days respectively. The animals show no symptoms and progress towards recovery.

4. It remains to be detercnined (1) how long the blood of treated cows may contain parasites after the apparent recovery, (2) if the parasites in such recovered animals are altered in virulence, (3) if the parasites are capable of infecting ticks.

5. The experiments were conducted on nine cows, of which four served as controls and five were treated with trypanblau. Of the controls two suffered from haemoglobinuria, and one of these died of piroplasmosis; the two other controls had no haemoglobinuria and were very mild cases. All of the treated cows had haemoglobinuria and recovered. In four of the treated cows haemoglobinuria occurred before treatment began.

6. As might be expected, the drug exerts a more rapid effect when injected intravenously. The parasites disappear more slowly after subcutaneous injection of the drug. (Judging from our recent experiments on dogs, the giving of the drug per os promises to be without effect. See This Journal, p. 231.)

7. Although doses of 150–200 c.c. of a saturated watery solution of the dye were used, it is probable that smaller doses will prove efficient. The drug appears to produce no ill-effects upon cattle.

8. The drug, being a dye, has the disadvantage of colouring the tissues, more especially the subcutaneous connective tissues. How long the colouration persists remains to be determined. In any case this disadvantage can scarcely weigh in the balance as against saving the life of the animal, especially when used for breeding purposes.

9. We hope that experiments, which are about to be conducted in the field in Africa and elsewhere, will demonstrate the value of the remedy in practice.

10. Trypanblau and similar drugs should be given a trial in the treatment of Carçeag in sheep and Biliary Fever in horses.

The morphology of the spirochaetes has proved in recent years a fruitful source of controversy. One has only to look through some of the vast literature on the subject to be convinced that the time has not yet come for drawing final conclusions, so varied and so contradictory are the observations by different well-accredited authors.

It has been stated that in the stomach of the bed-bug the spirochaetes of relapsing-fever may frequently be observed to lose their normal regular curves and become more or less “worm-like.” This is of interest to those who oppose the view that spirochaetes are rigid spirals, a view which, after gradual abandonment by most observers, has lately been again advanced by Novy and Knapp in their study on S. recurrentis.

There seems little reason to doubt that most blood-sucking invertebrates harbour in their digestive tract natural flagellate parasites, which have erroneously been described by various authors as part of the life-cycles of blood-parasites from vertebrates. Among arthropods such natural flagellates have now been observed in ticks, lice, bugs, mosquitoes, biting and non-biting flies, and fleas; and the list of such hosts steadily grows.

It would appear from the data presented that the favourite location for the common rat fleas of this vicinity, Ceratophyllus fasciatus, Bosc., and Loemopsylla cheopis, Roth., is about the hind quarters of the rat, while Ctenopsyllus musculi (Dugès), Wagner, prefers the region of the head and neck.

The same regional distribution of rat fleas was found in the case of the guinea-pigs.

Squirrel fleas are most numerous on the hind quarters.

These observations, while they are not to be insisted upon as throwing any special light upon the regional distribution of buboes in naturally infected plague rats, are of particular interest in showing the very constant preference of Ct. musculi for the region of the head and neck.

In regard to the other species of fleas it should be borne in mind that the skin areas are not of equal dimensions; that of the head and neck being smallest; that of the hind quarters being the largest; and the skin area of the fore quarters being between these in size.

The above-mentioned considerations bearing upon the influence of insect prevalence and the influence of meteorological conditions on the prevalence of trypanosome infections in rats may be summarized as follows:

(1) There are definite seasonal variations in the prevalence of trypanosome infections in M. rattus and in M. decumanus.

(2) No definite correlation is forthcoming between the seasonal prevalence of T. lewisi and the seasonal prevalence of rat fleas. The seasonal prevalence of the transmitting insects is apparently a factor of subsidiary importance in the causation of the trypanosome prevalence.

(3) The evidence adduced above indicates that the dominant factor determining the seasonal prevalence of trypanosome infections is the atmospheric temperature, the optimum temperature approximating to 79° F. It is probable that temperature operates by influencing a developmental cycle of the trypanosomes in the transmitting insects.

(4) It is possible that in addition to temperature the atmospheric humidity may play a part by influencing the direct or mechanical transference of the infection.