Journal of General Microbiology (1971), 63, 211-220
Printed in Great Britain
A Simple Chemically Defined Medium for the Production of Phase I Bordetella pertussis
By D. W. STAINER AND M. J. SCHOLTE
Connaught Medical Research Laboratories, University of Toronto,
Toronto, Ontario, Canada
(Accepted for publication 9 July 1970)
SUMMARY
A simple, chemically defined medium is described consisting of sodium glutamate, proline, cystine, salts, and growth factors, which is suitable for the large-scale production of phase I Bordetella pertussis. More than 30 x 10" organisms/ml. were produced in 48 to 72 h. growth in shake flasks and fer- mentors. The cultures were detoxified by the addition of 0.14% formalin to yield vaccines which were non-toxic to mice and guinea pigs and had good mouse-protective antigen levels. The antigenic stability was satisfactory after storage of the final vaccines at elevated temperatures.
INTRODUCTION
Hornibrook (1939) first described the use of a liquid medium for the propagation of phase I Bordetella pertussis, and since that time many modifications of his original formulation have been published (Verwey & Sage, 1945: Wilson, 1945; Cohen & Wheeler, 1946; Verwey, Thiele, Sage & Schuchardt, 1949; Sutherland & Wilkinson, 1961). These liquid media have consisted of a casein hydrolysate to which were added various salts, growth factors, and either starch, charcoal, or anionic resins. Studies on the growth requirements of B. pertussis have resulted in the publication of several formulae for chemically defined media (Ungar et al. 1950; Jebb & Tomlinson, 1955, 1957; Wilson, 1963; Goldner, Jakus, Rhodes & Wilson, 1966), and an excellent review of the problem by Rowatt (1957). In a series of metabolic studies on B. pertussis in our laboratories, Wilson (1963), Goldner et al. (1966), Vajdic, Goldner & Wilson (1966), used a synthetic medium in rocked L-tubes and obtained good growth of phase I organisms. The composition of their optimal medium is given in Table 1. This is a completely defined medium apart from the liver co-enzyme preparation, but would be unsuitable for large-scale production due to its relative complexity. This paper describes further modifications which have resulted in greatly improved yields of phase I bacteria. The properties of vaccines prepared from these cultures were also examined.
METHODS
Seed culture preparation and growth conditions
The strain of Bordetella pertussis used in most of our investigations was one of the Connaught Laboratories' routine vaccine production strains, obtained originally from Dr Pearl L. Kendrick, and designated no. 18334. Two other strains were also used in the large-scale production experiments, Kendrick's no. 10536, and a strain obtained [end of text]