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Schistosomal complement-fixation test
298
CORRESPONDENCE
TUBERCULOSIS AND MALNUTRITION
SIR,--Dr. Barlovatz (Transactions, 1959, 53, 214) states that he has found no evidence that plenty of food or vitamins materially improve the results of treatment of tuberculous Africans with antibiotics. This is contrary to our experience in Tanga Province (Tanganyika) during 1958 when over 300 patients were being treated. As is common in the tropics the majority did not attend until the disease was well advanced and their nutritional state was often poor. Many cases did not improve as well as expected whilst receiving daily out-patient antibiotic treatment under supervision. On admission, with the same treatment but with an adequate diet, these patients almost invariably did much better. Physical rest probably did not play a large part in the change as most of them were not working at home. It was considered desirable to admit most patients for " feeding up," treatment of concomitant parasitic infestations and for educative purposes. The period of detention was usually from 6 to 12 weeks. About a quarter of the patients were treated as out-patients entirely. The main criteria for admission were poor general condition, poor social circumstances, failure to improve as out-patients and suspicion of failure to take the drugs regularly. I believe that neglect of reasonable nutrition and physical rest in the early stages of treatment may lead to poor results. It appears probable that a large proportion of T.B. treatment in Africa must be domiciliary. I consider it unwise to start large-scale domiciliary treatment schemes without adequate numbers of beds for those who need them. I think there is a place for hostel accommodation where patients could be provided with simple accommodation and a fair diet ; and, in addition, their drugs given under supervision. I am, etc.,
D. R. W. HADDOCK.
125, Sussex Road, Southport, Lancashire. 3rd April, 1959.
SCHISTOSOMAL COMPLEMENT-FIXATIONTEST
SIR,--Dr. Schofield (Transactions, 1959, 53, 64) draws some useful conclusions about the increasing incidence of negative results with increasing duration of infection, or more probably repeated reinfection. It should be emphasized however that the test was positive in 82 per cent. of all Schistosoma mansoni infections of which the possible duration did not exceed 10 years (the figure for S. haematobium was higher than this) ; and as far as is known the test does not give false positive results provided that a good antigen is used. This letter refers to the difficulty of obtaining satisfactory antigens. During the period covered by Dr. Schofield's analysis we were fortunate in having the use of the antigen obtained by Sir Neil Fairley in 1927-1931 from livers of Planorbis exustus naturally infected with S. spindale, which has been stored dry in powder form at 0-5°C. without detectable loss of potency ; the alcoholic extract is also stable for very long periods;
CORRESPONDENCE
299
a small quantity of antigen recently sent to us from the same source near Bombay gave identical results to the old stock. Supplies of these antigens however are now exhausted apart from small reserves kept for the standardization of other antigens, and heavily infected snails from natural sources are not readily obtainable. In attempting to find a laboratory substitute we have been generously supplied with a variety of experimental antigens at different times by Dr. D. G. Davey, Dr. O. D. Standen and Dr. J. Newsome ; the dried powdered antigens have been made up as 1 per cent. extracts in absolute alcohol (30,000 cercariae/ml, approx.) in the same manner as Fairley's antigen, against which they have been titrated with pooled schistosomal sera, and as controls W.R. positive non-schistosomal sera. Some of the results are summarized as follows :
Antigen
Source
S. spindale/Planorbis: various batches
(1927-31) S. spindale/Planorbis (1956) Uninfected Planorbis liver S. rnansoni/Australorbis (2 batches) S. mansoni cercariae (clean, 5 batches) S. mansoni cercariae contaminated with
snail faeces (5 batches) mansoni cercariae (Lurie's method) mansoni ova mansoni miracidia mansoni adult worm
S. S. S. S.
Natural Laboratory
Dilution of Titre of 1% alcoholic schistosomal sera extract
Titre of control i
sera
~+ ~+ Neg. at ~+ ~+ + ~to
Neg. at
~+ ~+ ~+ k+ Neg. at
Neg. at
i'+ ~+ Neg. at
It will be seen that the only antigens which gave a satisfactory titre with schistosomal sera and a negative reaction with control sera were the naturally infected snail livers and the cercariae contaminated with snail faeces; these latter were quite heavily contaminated, were softer in consistency than clean cercariae and gave a yellow-green colour in solution. They have been put into routine use during the last 2 years and given results indistinguishable from those of the original infected snail liver antigen. By contrast Lurie and de Meillon (1952), using S. boris cercariae, cleaned them thoroughly in the expectation that clean cercariae would produce a better antigen than crude liver-cercaria preparations ; using an antigen dilution of ] to ~ they obtained complement fixation with schistosomal monkey sera at ]. This is in line with my own result with S. mansoni cercariae prepared according to their method, but it is the weakest of all the cercarial antigens. Attempts to improve the pure cercarial antigens by the addition of faeces, snail liver or cholesterol, have all been unsuccessful, but there is still a possibility that the intimate admixture of some factor in snail liver or faeces with cercariae at the time they are extracted with alcohol may enhance the solubility of the active principle of the antigen. The other possibility is that the principle, being water-soluble, is lost during washing of the cercariae, although Dr. Standen denies that his antigens are washed. If naturally infected snail livers make good antigen, and laboratory bred cercariae plus faeces are equally good, laboratory infected snail livers should be good also. If either of the reasons postulated for the failure of clean cercariae is correct, it is likely that the failure to date of laboratory infected snail
300
CORRESPONDENCE
livers is due to over destruction of the snail liver by hyper-infection, or that the antigen is lost by washing of the livers. Chaffee et al. (1954) use a saline extract of adult schistosome worms as an antigen but they take elaborate precautions to preserve the antigen during extraction at low temperature after a preliminary extraction with ether ; and then the solution of antigen in saline is not stable. I cannot account for the complete failure of my own adult worm antigen. The antigens prepared from live ova or miracidia of S. mansoni showed some species specificity in favour of S. mansoni as against S. haematobium. None of the other antigens tested showed any species specificity. Small modifications in the technique of the complement-fixation test have been made from time to time without noticeable effect on the results, but the quality of the antigen is all important ; even if W.R. positive serum is used as a control, weak antigens which have to be used at a dilution of 1 in 10 or less are not in my experience altogether reliable. Different batches of antigen of the same type prepared at different times in different laboratories give remarkably uniform titres provided the technique of their preparation is the same in essentials. I am, etc., D. S. RIDLEY. Hospital for Tropical Diseases, London. 4th April, 1959. REFERENCES :
CHAFFEE,E. F., BAUMAN,P. M. & SHAPILO,J. J. (1954). Amer. J. trop. Med. Hyg., 3, 905. LURIE, H. I. & DE MEXLLON,B. (1952). S. Aft. IVied.J., 26, 1005.
RANGE OF Plasmodium gonderi Sm,--Garnham, Lainson and Cooper (Transactions, 1958, 52, 509), have extended the geographical range of Plasmodium gonderi of mangabeys and drills to include the Cameroons. This parasite was previously known only from the West of the Belgian Congo. I should like to report briefly the presence of P. gonderi in Cercocebus fulginosus from Liberia. The blood of 19 C. fulginosus was examined ; six were found to be infected with Hepatocystis kochi only, one was found to be infected with P. gonderi only, and two animals harboured both parasites. One animal having a light infection of P. gonderi was splenectomized. The subsequent heavy infection was studied by myself and by Professor Garnham who, happily, was visiting Liberia at the time. The parasite was typical of P. gonderi. Attempts to infect Anopheles gambiae failed. One point worthy of note is the chronicity of the infection. The splenectomized mangabey has maintained high infections of the order of 10,000 to 25,000 parasites/c.mm. for 2 months at the time of writing. Similarly an intact mangabey infected by blood inoculation showed a similar intensity of infection for 42 days before any decrease was noted.
CORRESPONDENCE
TUBERCULOSIS AND MALNUTRITION
SIR,--Dr. Barlovatz (Transactions, 1959, 53, 214) states that he has found no evidence that plenty of food or vitamins materially improve the results of treatment of tuberculous Africans with antibiotics. This is contrary to our experience in Tanga Province (Tanganyika) during 1958 when over 300 patients were being treated. As is common in the tropics the majority did not attend until the disease was well advanced and their nutritional state was often poor. Many cases did not improve as well as expected whilst receiving daily out-patient antibiotic treatment under supervision. On admission, with the same treatment but with an adequate diet, these patients almost invariably did much better. Physical rest probably did not play a large part in the change as most of them were not working at home. It was considered desirable to admit most patients for " feeding up," treatment of concomitant parasitic infestations and for educative purposes. The period of detention was usually from 6 to 12 weeks. About a quarter of the patients were treated as out-patients entirely. The main criteria for admission were poor general condition, poor social circumstances, failure to improve as out-patients and suspicion of failure to take the drugs regularly. I believe that neglect of reasonable nutrition and physical rest in the early stages of treatment may lead to poor results. It appears probable that a large proportion of T.B. treatment in Africa must be domiciliary. I consider it unwise to start large-scale domiciliary treatment schemes without adequate numbers of beds for those who need them. I think there is a place for hostel accommodation where patients could be provided with simple accommodation and a fair diet ; and, in addition, their drugs given under supervision. I am, etc.,
D. R. W. HADDOCK.
125, Sussex Road, Southport, Lancashire. 3rd April, 1959.
SCHISTOSOMAL COMPLEMENT-FIXATIONTEST
SIR,--Dr. Schofield (Transactions, 1959, 53, 64) draws some useful conclusions about the increasing incidence of negative results with increasing duration of infection, or more probably repeated reinfection. It should be emphasized however that the test was positive in 82 per cent. of all Schistosoma mansoni infections of which the possible duration did not exceed 10 years (the figure for S. haematobium was higher than this) ; and as far as is known the test does not give false positive results provided that a good antigen is used. This letter refers to the difficulty of obtaining satisfactory antigens. During the period covered by Dr. Schofield's analysis we were fortunate in having the use of the antigen obtained by Sir Neil Fairley in 1927-1931 from livers of Planorbis exustus naturally infected with S. spindale, which has been stored dry in powder form at 0-5°C. without detectable loss of potency ; the alcoholic extract is also stable for very long periods;
CORRESPONDENCE
299
a small quantity of antigen recently sent to us from the same source near Bombay gave identical results to the old stock. Supplies of these antigens however are now exhausted apart from small reserves kept for the standardization of other antigens, and heavily infected snails from natural sources are not readily obtainable. In attempting to find a laboratory substitute we have been generously supplied with a variety of experimental antigens at different times by Dr. D. G. Davey, Dr. O. D. Standen and Dr. J. Newsome ; the dried powdered antigens have been made up as 1 per cent. extracts in absolute alcohol (30,000 cercariae/ml, approx.) in the same manner as Fairley's antigen, against which they have been titrated with pooled schistosomal sera, and as controls W.R. positive non-schistosomal sera. Some of the results are summarized as follows :
Antigen
Source
S. spindale/Planorbis: various batches
(1927-31) S. spindale/Planorbis (1956) Uninfected Planorbis liver S. rnansoni/Australorbis (2 batches) S. mansoni cercariae (clean, 5 batches) S. mansoni cercariae contaminated with
snail faeces (5 batches) mansoni cercariae (Lurie's method) mansoni ova mansoni miracidia mansoni adult worm
S. S. S. S.
Natural Laboratory
Dilution of Titre of 1% alcoholic schistosomal sera extract
Titre of control i
sera
~+ ~+ Neg. at ~+ ~+ + ~to
Neg. at
~+ ~+ ~+ k+ Neg. at
Neg. at
i'+ ~+ Neg. at
It will be seen that the only antigens which gave a satisfactory titre with schistosomal sera and a negative reaction with control sera were the naturally infected snail livers and the cercariae contaminated with snail faeces; these latter were quite heavily contaminated, were softer in consistency than clean cercariae and gave a yellow-green colour in solution. They have been put into routine use during the last 2 years and given results indistinguishable from those of the original infected snail liver antigen. By contrast Lurie and de Meillon (1952), using S. boris cercariae, cleaned them thoroughly in the expectation that clean cercariae would produce a better antigen than crude liver-cercaria preparations ; using an antigen dilution of ] to ~ they obtained complement fixation with schistosomal monkey sera at ]. This is in line with my own result with S. mansoni cercariae prepared according to their method, but it is the weakest of all the cercarial antigens. Attempts to improve the pure cercarial antigens by the addition of faeces, snail liver or cholesterol, have all been unsuccessful, but there is still a possibility that the intimate admixture of some factor in snail liver or faeces with cercariae at the time they are extracted with alcohol may enhance the solubility of the active principle of the antigen. The other possibility is that the principle, being water-soluble, is lost during washing of the cercariae, although Dr. Standen denies that his antigens are washed. If naturally infected snail livers make good antigen, and laboratory bred cercariae plus faeces are equally good, laboratory infected snail livers should be good also. If either of the reasons postulated for the failure of clean cercariae is correct, it is likely that the failure to date of laboratory infected snail
300
CORRESPONDENCE
livers is due to over destruction of the snail liver by hyper-infection, or that the antigen is lost by washing of the livers. Chaffee et al. (1954) use a saline extract of adult schistosome worms as an antigen but they take elaborate precautions to preserve the antigen during extraction at low temperature after a preliminary extraction with ether ; and then the solution of antigen in saline is not stable. I cannot account for the complete failure of my own adult worm antigen. The antigens prepared from live ova or miracidia of S. mansoni showed some species specificity in favour of S. mansoni as against S. haematobium. None of the other antigens tested showed any species specificity. Small modifications in the technique of the complement-fixation test have been made from time to time without noticeable effect on the results, but the quality of the antigen is all important ; even if W.R. positive serum is used as a control, weak antigens which have to be used at a dilution of 1 in 10 or less are not in my experience altogether reliable. Different batches of antigen of the same type prepared at different times in different laboratories give remarkably uniform titres provided the technique of their preparation is the same in essentials. I am, etc., D. S. RIDLEY. Hospital for Tropical Diseases, London. 4th April, 1959. REFERENCES :
CHAFFEE,E. F., BAUMAN,P. M. & SHAPILO,J. J. (1954). Amer. J. trop. Med. Hyg., 3, 905. LURIE, H. I. & DE MEXLLON,B. (1952). S. Aft. IVied.J., 26, 1005.
RANGE OF Plasmodium gonderi Sm,--Garnham, Lainson and Cooper (Transactions, 1958, 52, 509), have extended the geographical range of Plasmodium gonderi of mangabeys and drills to include the Cameroons. This parasite was previously known only from the West of the Belgian Congo. I should like to report briefly the presence of P. gonderi in Cercocebus fulginosus from Liberia. The blood of 19 C. fulginosus was examined ; six were found to be infected with Hepatocystis kochi only, one was found to be infected with P. gonderi only, and two animals harboured both parasites. One animal having a light infection of P. gonderi was splenectomized. The subsequent heavy infection was studied by myself and by Professor Garnham who, happily, was visiting Liberia at the time. The parasite was typical of P. gonderi. Attempts to infect Anopheles gambiae failed. One point worthy of note is the chronicity of the infection. The splenectomized mangabey has maintained high infections of the order of 10,000 to 25,000 parasites/c.mm. for 2 months at the time of writing. Similarly an intact mangabey infected by blood inoculation showed a similar intensity of infection for 42 days before any decrease was noted.