The growth of staphylococci in human serum

Abstract

and Weatern Infirmary, Q h g o u rOBSERVATIONS by Ekstedt and Nungester (1955) and Ekstedt (1956a and b) have suggested that human serum contains a heat stable, calcium dependent inhibitor which prevents the growth of coagulase-negative staphylococci. Coagulase-positive staphylococci grow well in serum apparently because they can remove the inhibitor. Thus treatment of the serum with heat-killed suspensions of Staphylococcwr aureus or with cell-free preparations of coagulase was found to render it non-inhibitory to the growth of coagulase-negative organisms.It seemed that knowledge gained from studies of this convenient in-vitro system might be relevant to the aggressive action of coagulase and I therefore considered it important to repeat and, if possible, to extend the observations of Ekstedt and Nungester. MATERIALS AND METHODSOrganisms. Twcnty-five strains of staphylococci, 14 coagulaae negative and 11 coagulase positive, were collected mostly from hospital eources. Coagulase tests were performed by the slide technique and also in tub-containing 1 ml. of 20 per cent. human p h a in saline (0.8 per cent. NaCl) solution. The strains were maintained on nutrient agar slopes in screw-capped bijou bottles.Serum specimem. Thirty sera were investigated. Most of them were obtained from the Blood Transfusion Service and had been found to have insufficiently high titres of anti-Rh agglutinins for grouping purposes. They were stored a t -20' C. and were between 2 weeks and 2 months old at the time of investigation. Before use they were Seitz filtered ; this not only sterilised them but made them completely clear, thus aiding the visual detection of growth. To exclude the possibility that a nutritional deficiency of the Bera and serum derivatives might be responsible for the failure of growth, 6 per cent. of nutrient broth was added to every preparation tested for inhibitory activity. This broth was an ox heart infusion broth containing 0.005 per cent. added para-aminobenzoic acid; in 5 per cent. concentration in saline i t supported luxuriant growth of all the strains in the collection. Placmza apecimm. Some plasmas were obtained from expired transfusion stock and these contained approximately 20 per cent. of an anticoagulant mixture consisting of 2 per cent. disodium citrate and 3 per cent. dextrose. Paired specimens of fresh plasma and serum were taken from blood donors. The plasma was obtained by mixing 18 ml. blood with 2 ml. 3.8 per cent. trisodium citrate and removing the red cells by centrifugation. A corresponding 18 ml. of clotted blood was left overnight at room temperature and the clot was scparated by centrifugation the next day; 0.8 ml. of 3.8 per cent. trisodium citrate was then added to each 4 ml. of the serum to make i t comparable with the corresponding plasma.

Staphylococci and Their Classification

“…Further identification of a strain of Staph. aureus can be obtained by determination of phage typability (Parker, 19621, hemolysin pattern (Elek & Levy, 1951), and ability to grow in normal human serum (Fletcher, 1962 Cowan (1939) to distinguish three antigenic types of Staph. aureus.…”

The Classification of Staphylococci and Micrococci from World-wide Sources

“…These tests when used in conjunction with the free and bound coagulase tests (Duthie, 1954) will clearly separate S. aureus from related staphylococci; positive recognition of this species is obtained when a t least two of these tests are positive. Further confirmation of diagnosis can be obtained by determining phage typability , haemolysin pattern (Elek & Levy, 1950) and ability to grow in normal human serum (Fletcher, 1962).…”

Bacteriostatic Activity of Serum Against Staphylococci

“…Similar results were reported by Fletcher (9), who found that the inhibitor is active at an alkaline pH and is resistant to heating at 100 C. This author was unable to confirm the findings of Ekstedt cited in the introduction to this paper. Fletcher (1962) found that many proteinaceous materials, such as peptone or even a pool of amino acids, remove the inhibitor from serum.…”

“…This role of coagulase was, however, not confirmed (23,24) and was also more or less doubtful to other investigators (9,15,18). It is not known whether serum of an animal infected with staphylococci exhibits bacteriostatic activity in vitro.…”