DNA–DNA hybridization in blackflies (Diptera: Simuliidae)

This paper is a study of base sequence relationships in blackflies by DNA–DNA hybridization. The species used are: Simulium venustum, S. tuberosum, S. vittatum, Cnephia dacotensis, and Prosimulium multidentatum. The midpoints of thermal transition and the buoyant density in CsCl were determined for the DNA of each species and the percent G + C estimated. With the filter method of hybrid formation, maximal yield was obtained at 60 °C; 24 h was found to be a practical length of time for incubation. When the amount of DNA on filter was varied, the homologous reaction reached a plateau at about 50% of the input radioactive DNA; the heterologous reaction reached a plateau at a lesser value in a species-dependent manner. The results indicate a close relationship between S. venustum and S. tuberosum, a more distant one between these species and S. vittatum, and an intermediate one with C. dacotensis. Prosimulium multidentatum shared very little homology with members of the other genera. A precise quantitative evaluation of the results is difficult owing to the complexity of interacting factors. Interpretation would be aided by knowledge of genome sizes and intragenome repetition frequencies. Appropriate studies are underway.

CROSS-REACTIONS AMONG GROUP A STREPTOCOCCI

Strains of four streptococcal types, 33, 41, 43, 52, and a nontypable strain, Ross, cross-reacted in precipitin and bactericidal tests. The homologous reactions, which determined the type, afforded the major protection and developed promptly and regularly in the serum of rabbits during immunization. The associated cross-reactions, on the other hand, appeared in the serum of certain rabbits only, were often not as strong as the associated homologous reactions, and required for their presence a longer period of immunization than the homologous reactions. Agar gel analysis of the homologous precipitin reactions revealed, as would be expected, reactions of serological identity, while those cross-reactions which were strong enough to test in this way formed bands of precipitate which joined with spur formation on the side of the homologous reaction. These experiments and others referred to in the text suggest that cross-protection, as demonstrated in bactericidal tests, is sufficiently widespread to be a factor in streptococcal immunity, if a corresponding protection occurs in vivo. Thus, streptococcal infection with one of the cross-reacting strains might confer, in addition to strong homologous protection, a certain amount of cross-protection.

The antigenicity of sera of man and animals in relation to the preparation of specific precipitating antisera

Specific precipitating antisera can be prepared in rabbits by the repeated inoculation of alum-precipitated serum.High titre of sensitivity to the homologous serum is obtained only with a complete loss of specificity.By absorption of the non-specific antiserum with heterologous sera the specificity can be restored.Two types of reactions can be observed in such non-specific antisera: (1) the homologous reaction which occurs with the sera of closely related animals, and (2) the heterologous reaction which is attributed to the presence of antigens which are widespread among mammalian sera.Cross-immunization experiments have demonstrated the presence of specific antigenic material in the serum of indvidual species.A schematic representation of the antigenic arrangement of mammalian sera is suggested.I am indebted to the Colonial Medical Research Committee for their help in this investigation and for the financial help provided by the Colonial Office.I would like to acknowledge my indebtedness to Prof. P. A. Buxton, F.R.S., for his help and encouragement and in providing and housing the chimpanzee used in these experiments, to Dr C. H. N. Jackson and Mr W. A. Hilton for their valued help in the field in collecting a number of sera from East Africa, and also to Dr E. Hindle, F.R.S., for supplying me with materials from animals which had died at the Zoological Gardens, London. I wish to record my thanks to Mr E. Pope, for his valued technical assistance throughout these experiments.

THE SEROLOGICAL SPECIFICITY OF PARTICULATE COMPONENTS DERIVED FROM VARIOUS NORMAL MAMMALIAN ORGANS

1. Particles derived from filtrates of organ suspensions by high speed centrifugation were serologically active as shown by agglutination and complement fixation techniques. Particles from brain, liver, lung, kidney, heart muscle, spleen, testicle, and pancreas of various species have been studied. 2. All particles showed a certain degree of organ specificity with the exception of pancreas. Cross-reactions occurred between the particles from various organs from one species, which were more marked when complement fixation technique was employed than by the agglutination test. However, agglutination always appeared earlier and was stronger, and complement fixation was positive in higher dilutions of antigen in the presence of homologous antiserum than with heterologous antisera. 3. The cross-reactions did not depend on the occasional precipitins for serum and the agglutinins for the red cells of the species from which the particles were derived, nor did they bear a relation to Wassermann and Forssman antibodies present in some of the sera. 4. The organ specific differentiation of the particles from various organs could more clearly be demonstrated by two means: The antiserum could be diluted in such a way that only the homologous reaction still showed a positive result while the cross-reactions had become negative; or the cross-reacting antibodies could be absorbed by heterologous particles and the homologous reaction was still more or less intact. 5. In addition to the organ specific differentiation, most particles were found to exhibit species specificity. While the particles derived from kidney, lung, testicle, and heart muscle aggregated only in the presence of the antiserum against the corresponding organ particles from the homologous species, brain particles reacted with brain antisera against both homologous and heterologous species alike. Absorption of brain particle antisera with brain preparations from a heterologous species removed all antibodies. Liver particle preparations showed an intermediate position in that all liver preparations with the exception of rabbit liver particles were aggregated by any liver particle antiserum. However, absorption with liver particles from a heterologous species left a distinct species specific reaction in the serum. 6. The antigens involved are all destroyed by heating to 100° C. for a few minutes with the exception of brain particles, which after 20 minutes at 100° C. still gave complement fixation almost to the same strength as the untreated controls. 7. Alcoholic and ether extracts of brain reacted with the brain particle antisera only. All alcoholic or ether extracts of other organs gave no complement fixation. None of the various other organ particle antisera tested contained antibodies for these extracts. 8. The relationship between the heat-stable and the alcohol-soluble brain particle antigen studied by absorption technique revealed that there were two antigens present, both organ specific and independent of the species, the one alcohol- and ether-soluble, the other not soluble in these solvents but heat stable. Some of the sera showed besides a few species specific antibodies. 9. Preliminary evidence has been gathered to show that no iso-immunization could be obtained with any one of the organ particles. As far as cytotoxic activity of the sera is concerned only the kidney particle antisera have been studied for nephrotoxins; these failed to reveal any such activity in the mouse.