1. In the course of the examination of sputum from cases of lobar pneumonia, observations have been made on the incidence of the chief types of pneumococci. In the district from which the material was obtained, there was an apparent local diminution in the number of cases of lobar pneumonia due to Type II the figures were 326 per cent. of Type II cases in the period 1920–22, and only 74 per cent. in the period 1924–27. The incidence of Type I was approximately the same in the two periods, the percentages being 3O6 and 343.
2. Several different serological varieties of pneumococci have been obtained from the sputum of each of several cases of pneumonia examined at various stages of the disease. This has occurred most frequently in cases of pneumonia due to Type I, and in two instances four different types of Group IV were found in addition to the chief types. The recovery of different types is facilitated by the inoculation of the sputum (preserved in the refrigerator), together with protective sera corresponding to the various types in the order of their appearance.
3. Two interesting strains of Group IV pneumococci have been obtained from pneumonic sputnm.
One was an R strain which produced typical rough colonies, yet preserved its virnlence for mice and its capacity to form soluble substance. This R pneumococcus developed a large capsnle in the mice, which died of a chronic type of septicaemia. A strain producing smooth colonies was obtained from it in the course of a prolonged series of passage experiments.
The second strain, which was proved not to be a mixture, agglutinated specifically with the sera of two different types. In the peritoneal cavity of the mouse the specific soluble substance of each type was produced.
4. A method of producing the S to R change through ageing of colonies on chocolate blood medium containing horse serum is described. After two to three days' incubation small rough patches appear in the margins of the smooth colonies, and from these pure R strains can be isolated.
5. It has been shown that the R change is not equally advanced in the descendants of virulent pneumococci which have been exposed to the action of homologous immune serum. Some R strains form traces of soluble substance in the peritoneal cavity of the mouse; these revert readily to the virulent S form and, in addition, are able to produce active immunity. Others show no evidence of S antigen; spontaneous reveraion takes place with difficulty, if at all, and they are incapable of producing active immunity. The stronger the immune serum used, the more permanent and complete is the change to the R form.
6. Restoration of virulence to an attenuated R strain, with recovery of the S form of colony and of the original serological type characters may be obtained by passage through mice. The change from the R to the S form is favoured by the inoculation of the R culture in large doses into the subcutaneous tissues; but the most certain method of procuring reversion is by the inoculation of the R culture, subcutaneously into a mouse, together with a large dose of virulent culture of the same type killed by heat. Incubation of such a mixture in vitro does not induce reversion.
7. Reversion of an R strain to its S form may occasionally be brought about by the simultaneous inoculation of virulent culture of another type, especially when this has been heated for only a short period to 60° 0., e.g. R Type II to its S form when inoculated with heated Type I culture.
8. Type I antigen appears to be more sensitive to exposure to heat than Type II antigen, since the former loses the power to cause reversion when heated to 80° C., whereas Type II culture remains effective even after steaming at 100°C.
9. The antigens of certain Group IV strains appear to be closely related to that of Type II, and are equally resistant to heat. Steamed cultures of these Group IV strains cause the R form derived from Type II to revert to its S form, while they fail to produce reversion of the R form derived from Type I.
10. The inoculation into the subcutaneous tissues of mice of an attenuated R strain derived from one type, together with a large dose of virulent culture of another type killed by heating to 60° C., has resulted in the formation of a virulent S pneumococcus of the same type as that of the heated culture.
Thp newly formed S strain may remain localised at the seat of inoculation, or it may disseminate and cause fatal septicaemia.
The S form of Type I has been produced from the R form of Type II, and the R form of Type I has been transformed into the S form of Type II.
The clear mucinous colonies of Type III have been derived both from the R form of Type I and from the R form of Type II, though they appear to be produced more readily from the latter. The newly formed strains of Type III have been of relatively low virulence, and have frequently remained localised at the subcutaneous seat of inoculation.
Virulent strains of Types I and TI have been obtained from an R strain of Group IV.
11. Heated R cultures injected in large doses, together with small doses of living R culture have never caused transformation of type, and only rarely produced a reversion of the R form of Type II to its virulent S form.
12. The results of the experiments on enhancement of virulence and on transformation of type are discussed and their significance in regard to questions of epidemiology is indicated.
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