Medical Hypotheses 5: 1237-1249, 1979 THE NAEGLERIAL CAUSATION OF RHEUMATOID DISEASE AND MANY HUMAN CANCERS. A NEW CONCEPT IN MEDICINE. R. Wyburn-Mason, 2 Hillbrow, Richmond Hill, Richmond, Surrey, England. ABSTRACT Man and terrestrial animals live in an environment containing free- living amoebae on the surface soil, in pools, fresh water lakes, rivers and streams. They form cysts, which float in the air and which are continually inhaled and found in the nasopharynx and their trophozoites are present in human and animal faeces. Amoebae of the genus, Naegleria, have been demon- strated in all human tissues, both healthy and in larger numbers in those taken from cases of rheumatoid disease, in all human cancers and in the unaffected tissues of cancer patients. They can be killed in vitro by a series of different anti-amoebic substances and treatment of active cases of rheumatoid disease by any of these, either causes cessation of disease activity or a temporary exaggeration of symptoms followed by their lessening or disappearance (Herxheimer reaction), indicating the presence of an amoeba in the affected tissues as the causative organism of the inflammation in this disease in subjects genetically sensitive to the organism. Every internal organ may be involved in the inflammatory response in cases of rheumatoid disease and this also ceases with the above treatments. Many of these internal lesions are premalignant, so that infection with the organism either in sensitive subjects or with pathogenic species, appearsto be the primary cause of cancer in many cases. The presence in the body of Naegleria represents the source of the constant antigenic stimulation thought to be responsible both for rheumatoid disease and for the development of lymphomata and myelomatosis. Key Words AMEBA AMEBIASIS ARTHRITIS, RHEUMATOID NEOPLASMS PRECANCEROUS CONDITIONS PROTOZOAN INFECTIONS SJOGREN'S SYNDROME 1237 INTRODUCTION- The considerations which follow are a resume of the author's recently published monograph (1) and statements made can be confirmed by reference to this. The existence of free-living amoebae, not requiring a host for their conti‘nued existence, has been known at least since the last century. They are found on the surface soil in most parts of the world, preferring warm moist conditions, and they proliferate in warm stagnant pools, lakes and at the bottom of rivers, particularly in the regions where a warm effluent enters. Pathogenic free-living amoebae are readily isolated from chlorinated swi'mming pools, potable waters, sewage and nasal and throat cavities. They have been found in the domestic water supply in South Australia and recently in the warm water of the Roman Baths at Bath, England. There are probably about 300 different species known at present, and they have been divided into two main genera, the Acanthamoebae and Naegleria. The trophozoites of these differ in various details, such as shape, size, number of pseudopodia and vacuoles,rate of movement, mode of nuclear division and in antigenic content. In inimical conditions they form hollow spherical cysts of 9-27 microns in diameter. The Naegleria cysts have a smooth surface with fenestrations and readily form in vitro, but never in the tissues, while Acanthamoebae readily encyst both in vitro and in the tissues. Naegleria develop into biflagellate forms in distilled water and organisms of both genera are soluble in one per cent deoxycholate (bile salts). Cysts of free-living amoebae are found in the air in most parts of the world other than those covered by ice and can readily be observed on agar plates exposed to air for 10 minutes or more. These organisms often contaminate tissue cultures. Most free-living amoebae prefer warm surround- ings and they tend to migrate from cooler to warmer conditions, a property known as thermotropism possessed by many parasites of warm-blooded animals. Apart from their presence in the nose and throat, trophozoites of these organisms are often found in human faeces and that of most animals and in sewaye. It is obvious that all living terrestrial animals and plants and many in fresh water live in a world surrounded by many species of free- living amoebae, which certainly pass into the respiratory passages as cysts or trophozoites and which are also present as trophozoites in the bowel: since they are found in the faeces. As the organisms possess thermotropic properties, it would be unreasonable to suppose that once they had entered the orifices of the body, they would not migrate through the mucosae to the warmth of the body tissues and, if pathogenic, induce an inflammatory reaction in susceptible subjects. Free-living amoebae as a cause of human disease Pathogenic Naegleria have been recovered from the nasopharynx and bronchus in patients with or without fever and respiratory symptoms. Since 1970 one species of Naegleria (N. fowleri) has been shown to be the cause of primary amoebic meningo-encephalitis, an uniformally fatal condition. An amoebic infection of the eye with a free-living amoeba, probably of the Acanthamoeba genus, was described in 1974 and a case of multiple brain abscessescontaining a free-living amoeba, also probably Acanthamoeba, who suffered from long-standing Hodgkin's disease of the lung has also been reported. Apart from these cases free-living amoebae have been considered non-pathogenic in man. One of the features stressed by all workers on 1238 infection with all amoebae, either in man or induced by inoculation of cultures into experimental animals, is that the organisms usually cannot be identified in the tissues by ordinary staining techniques, although they are known to be there. They are usually mistaken for macrophages, polymorphs or lymphocytes. Their presence can be shown, however, by the use of immunofluorescent staining techniques using antisera prepared against various species of Naegleria and Acanthamoebae. Introduction of cultures of Acanthamoebae or Naegleria into experi- mental animals, such as monkeys and guinea pigs, produces a chronic wasting disease resembling carcinomatosis, lymphadenopathies, splenomegaly and infiltration of lymphocytes into various tissues and especially vascular lesions typified by formations like periarteritis nodosa, arteritis and thromboses and marked intimal proliferation and by pyelitis. The lymph node lesions eventually change to a granulomatous reaction with lymphocytes, plasma cells, intense reticul urncell proliferation and multi-nucleated cell formation. The liver shows lesions in the portal triads. Recently it has been shown by using indirect fluorescent antibody tests that the sera of all living humans, including the new-born, contain anti- bodies against free-living amoebae, either Naegleria or Acanthamoeba (2). This shows that every human, and probably other animals also, have been or are presently infected with free-living amoebae to produce such an antibody response, though the organisms may not have been observed in the tissues. The discovery that FREE-LIVING PROTOZOA ARE NOT PURELY NON-PATHOGENIC, BUT ARE ABLE TO INFECT MAN AND ANIMALS HAS REVOLUTIONIZED THE VERY CONCEPT OF PARASITOLOGY and shows that all humans contain foreign antigens in their plasma and tissues, in the case of the new-born evidently resulting from transplacental passage of either the organisms or their antibodies into the foetus in utero. In 1922 several groups of workers in California described the presence of an amoeba in the bone marrow in cases of rheumatoid disease. This could be grown in cultures, but subsequent work, though confirming its amoebic nature, failed to verify that this was E. Histolytica as postulated, though its nature was never elucidated. Furthermore, amoebae have been demonstrated in every soft tissue of the body (3), while Acanthamoebae have been cultured from a liver abscess in man. A new method of recovery of Naegleria from human tissues and other _.sources. In view of these observations the author attempted to induce an organism to migrate out of fresh, minced, unfixed human tissue obtained at autopsy, biopsy or operation using the property of thermotropism. The minced tissue was placed on a membrane filter with pores of 0.5-1.0 m in diameter, which itself rested on a zinc gauze diaphragm anchored half w s-'y down a funnel. The tissue was cooled to O°C by placing a glass beaker containing ice on top of it. The lower half of the funnel and its stem were filled with Ringer's solution reaching to the level of the zinc gauze diaphragm and wetting the membrane filter. The spout of the funnel was closed with rubber tubing and a clip. To the Ringer's solution was added 20 units of penicillin and 40 units of streptomycin per ml. The temperature of the Ringer's solution was maintained at 37OC by a water-bath reaching to the level of the gauze diaphragm. The whole apparatus was sterilized before use 1239 and the introduction of the minced material and the experiment carried out in sterile conditions. In the above circumstances there existed a temperature differential between the minced tissue at O°C and the Ringer's solution at body temperature of 370C. If a contractile organism was present in the tissues, it might migrate through the filter under thermotropic influences into the Ringer's solution, which is, in fact, what occurred. The organism was found in the Ringer's solution when this was run off into a sterile centrifuge tube and centrifuged at 150 revolutions per minute. When placed on slides, the deposit showed an amoeba-like organism, usually brown stained from uptake of cell debris, about 30 /urn in diameter, showing no motility and being impossible to culture. It was when discovered that centrifuging killed the organism. If this was avoided and the Ringer's solution replaced by "amoeba saline" and later this was run off on to Noble agar plates carpeted with a live strain of E.coli, the organism could be cultured.