No abstract

In experimental immunology, it frequently is of interest to obtain quantitative estimates of the virulence of strains of microorganisms as evidenced by their ability to produce fatal infections in experimental animals. It is recognized that, in the broader sense, virulence is not a specific intrinsic attribute of an organism but is the result of a combination of a number of factors pertaining to the organism, the affected host, and the conditions (known or unknown) under which the pathological state is established. Nonetheless, when suitable laboratory animals are employed, and when the combination of experimental conditions is propitious, it can be demonstrated with some consistency that even closely related strains of organisms differ in their capability of producing experimental infections. For the purpose of this presentation, such differences in capability can be considered as differences in virulence of the organisms regardless of the true nature of the experimental infective process.Quantitative estimates of virulence have been employed by various workers in studying the relationship of this property to antigenic constitution, bacterial dissociation, and various epidemiological phenomena. Furthermore, it popularly is considered that the immunogenic activity of a strain of bacteria is directly related to its virulence. As a consequence, such tests are employed extensively as a control technique in routine production of bacterial vaccines and in searching for immunogenically superior strains.Mice have been widely accepted as the laboratory animals of choice for determining the virulence of strains of Salmonella typhosa and the testing procedures employed have been of two main types. The first, based on the intraperitoneal injection of graded doses of the organisms suspended in physiological saline, is utilized almost exclusively by British workers. The second, incorporating the use of hog gastric mucin as a suspending medium, has been employed by most American workers since the original reports of the technique by Nungester et al.(1) and of its application to typhoid by Rake (2). A third method, which was reported by Norton and Dingle (3) to show promise, is based on the intracerebral injection of the organisms without the aid of an adjuvant. This latter method has been used successfully in determining the virulence of strains of Hemophilus pertussis by Kendrick and her coworkers (4, 5) and as an experimental infective technique with Shigella dysenteriae by Dubos et al.

A study of the effect of graded immunizing doses of typhoid vaccine and graded challenge doses of S. typhosa in 5 per cent mucin on the degree of gradation of response (survival or death) elicited in successive groups of mice is reported. In the range of doses employed the effect of graded immunizing doses was markedly greater than was the effect of graded challenge doses. Statistically the difference exceeded the 0.1 per cent level of significance. It was concluded that the use of graded immunizing doses was preferable to the use of graded challenge doses as a basis for the immunological assay of typhoid vaccine, since with the former there was obtained (a) a greater significance of differences in response of groups given graded doses even with smaller fold increments in successive doses, (b) a greater slope of the dosage-response curve which permitted estimation of ED50 values with smaller standard errors, and (c) an effect on response apparently less dependent on the sex of mice used.

The need for an efficient, practical laboratory method for determining the potency of tvphoid vaccines long has been recognized by immunologists concerned with the development, production, and assay of the product. The need in relation to production and control in routine manufacture of typhoid vaccine is mainly for a testing method which will permit the ready demonstration that an individual lot, or successive lots, of vaccine equal or exceed an established level of potency. Researchers, on the other hand, require an assay method which, in addition to the foregoing, permits quantitative estimation of potency, i. e., which permits numerical expression of potency in absolute or relative terms with an estimable degree of uncertainty. Only by the use of such a method is the investigator enabled to detect or evaluate differences or changes in potency as effected by experimental treatments or conditions of interest. Methods early used in the determination of the potency of typhoid vaccines were based upon parenteral administration of the products to rabbits and subsequent in viro determination of the agglutinin response (15), or the actual protection of such immunized rabbits against subacute infection following injection of living, virulent Salmonella typhosa (18). It is beyond the scope of this report to discuss or criticize these tests in detail and it is considered sufficient to state that such methods have proved to be neither adequately definitive nor reproducible to warrant their continued use. Investigation and establishment by Siler and his associates at the Army Medical School (22) of the mouse-protection test for assessing the antibody response of humaDs to typhoid immunization stimulated

The determination of values, even repeated measurements of a certain dimension of a single inanimate article, is subject to errors or lack of precision. Likewise, in the mass production of articles designed to be alike, determination of a certain dimension or attribute of each article produced yields a series of values differing from unit to unit. If these values are plotted as a frequency distribution, they describe a pattern characteristic of the process under consideration. Every process, no matter now rigorous the attempt to control it, is subject to inherent variability, and the pattern of variability for each process is unique unto itself. When any change from the basic pattern occurs, some new source of variability must have entered the process. The methods of quality control are based on this principle. Prior to World War II, Shewhart (1931, 1939) and Pearson (1935) studied the application of statistical methods to problems pertaining to the control of variability of manufactured articles and introduced' the basic techniques of modern quality control. The urgent demand for testing procedures combining accuracy and economy, occasioned by the accelerated production schedules of