John Y. Sugg

Serologically Reactive Material in Spinal Fluid, Blood, and Urine from a Human Case of Cryptococcosis (Torulosis).∗

Summary Serologically reactive substances (“soluble antigens”) which were readily detectable by precipitation and by complement fixation with Cryptococcus antiserum were found in the spinal fluid, blood serum? and urine of a patient with cryptococcal meningitis. Absorption of the antiserum with solutions of purified Cryptococcus polysaccharide removed its capacity to react with the fluids of the patient. The reactive substances in the patients fluids were relatively heat stable and it is probable that they represent capsular polysaccharides of the fungus with which the patient was infected. Serological reactions of the Quellung type were obtained with the capsules of Cryptococcus cells from the centrifuged sediment of the spinal fluid.

Influence of Sucrose upon Production of Serologically Reactive Material by Certain Streptococci

Summary When grown in sucrose broth some strains of group H streptococci produced large amounts of material reactive with types 2 and 20 antipneumococcus and with antileuconostoc serums; little or none was produced by the same streptococci when grown in dextrose broth. This reactive material was different from the streptococcus antigen involved in the usual Lancefield grouping test; as much of the latter was produced in dextrose as in sucrose broth culture. In addition to indicating an interrelationship of the different Gram-positive cocci the data furnished an example of the influence of a particular carbohydrate upon the capacity of some microörganisms to elaborate a serologically reactive polysaccharide.

THE SEROLOGICAL ACTIVITY OF DEXTRANS

The serological reactivity or antibody-combining properties of dextrans have been studied in our laboratory over a period of years. Studies made in 1941421-g established that natural dextrans elaborated by sucrose broth cultures of Lewonostoc mesenteroides and of certain streptococci, and also dextrans synthesized in vitro by means of cell-free leuconostoc enzymes, possess a distinctive set of serological properties. A prominent feature was the strong capacity to cross-react with types 2 and 20 pneumococcus antiserums and in the case of some dextrans, also to cross-react with type 12 antiserum. These serological properties have since been utilized as a means of recognizing and differentiating dextrans in enzyme-mixtures and in bacterial culture fluids.?-lo The antibody-combining properties of dextrans assumed new significance when the partly hydrolyzed material was introduced in 1944-45 as a plasma volume extender. Our interest was aroused especially because the Swedish authors, failing to recognize our previous studies, claimed immunological inertness as one of the features in favor of dextran as a plasma s ~ b s t i t u t e . ~ ~ ~ ~ Tests of a number of Swedish (Pharmacia) clinical dextrans revealed that all of them possessed serological reactivity of the same qualitative nature as that possessed by native dextrans.16 In later experiments, not yet published, the clinical dextrans of two American manufacturers, as well as the previously studied Swedish products, were found to precipitate and fix complement with leuconostoc and with type 2 and type 20 pneumococcus antiserums. Moreover, the intravenous injection of as little as 0.1 or 0.2 mg. of any of the available clinical, dextrans produced anaphylactic death in guinea pigs passively sensitized with high titered pneumococcus antiserums. In spite of the qualitative similarity in their reactions, however, native and partially hydrolyzed dextrans show obvious quantitative differences in their capacities to give precipitation and complement fixation with types 2 and 20 antipneumococcus serums. FIGURE 1 shows, on a log-log scale, the regions in which a native dextran and a series of dextran fractions (kindly supplied by Dr. B. Ingelman) gave visible precipitation in tests against a highly potent type 2 pneumococcus antiserum. Each dextran was tested in a series of 7 concentrations (0.10 to 1,OOO pg per ml) against a series of 7 dilutions of the antiserum (1 :6 to 1 :400). The points on each curve represent the maximal dilutions of antiserum that gave visible precipitation with the various concentrations of the particular dextran. Points falling on the abscissa represent the instances in which no precipitation was observed with 1 :6 dilution of the serum.

Physical-chemical factors in agglutination of sheep erythrocytes by influenza virus.

Summary The data indicate that agglutination of sheep erythrocytes by influenza virus (WS strain) is influenced by physical-chemical factors within the test systems, and especially by the hydrogen-ion concentration. Over a relatively wide pH range, agglutination of sheea cells is inhibited by substances present in allantoic fluid, normal and infected. The allantoic fluid inhibition is ineffective in the pH range 5.8 to 6.0. When the test systems are adjusted with pH 5.8 buffer, and 1/2% to 1/4% cell suspensions are employed, sheep erythrocytes may be used with essentially the same results as obtained with chicken cells, for the determination of influenza virus content of allantoic fluids, and for the demonstration of antibodies by the agglutination-inhibition test. The data are significant because of the bearing they have upon the mechanism of virus hemagglutination

Significance of Antigenic Differences Among Strains of Influenza A Virus in Reinfection of Ferrets.

Summary Four to 10 weeks following an initial infection with influenza A virus, ferrets were found to be immune to clinical reinfection with that same strain of virus but were susceptible to reinfection with a strain of virus which was antigenically related to but different from the strain used for the original infection.

Inactivation of Hemagglutinating Capacity of Type A Influenza Virus By Trypsin Treatment.

Summary The effect of trypsin on hemagglutinating capacity of 6 type A strains of influenza virus of human origin has been determined under different experimental conditions. With both virus and enzyme in tris-Cl buffer and incubation at 37°C the hemagglutinating capacity of all strains was rapidly inactivated. Minimum concentration of enzyme and minimum time required for inactivation was not the same for all strains. However, with all strains, lowering the incubation temperature or adding sodium or calcium chloride to the trypsin-virus mixtures either reduced the rate of the reaction or completely inhibited it. These changes in test conditions did not in all instances affect all strains to the same extent. With 5 of the 6 strains the effect of added salt appeared to be more closely associated with the concentration than the type of ion present, whereas with the sixth strain the type of ion was a significant factor. The findings illustrate the importance of both the strain examined and the experimental conditions employed in determining the response of type A influenza virus to trypsin treatment.

Further Studies on Serial Passage of a Mixture of Influenza Viruses in Embryonated Eggs.

Summary Undiluted allantoic fluids were used as inocula for serial passage of a mixture of the PR8 and Lee strains of influenza virus in embryonated eggs. Although wide differences were found in the relative concentrations of the two viruses in the various fluids, the mixture was successfully carried through 52 consecutive passages, and each agent was recovered in an apparently pure culture at the end of the experimental period. Since the two viruses have different growth rates and are capable of interfering with the multiplication of each other, the persistence of both agents during serial passage of the mixture suggests that multiplication of the PR8 and of the Lee viruses may not require the same host factors.

Susceptibility of Convalescent Ferrets to Reinfection with Influenza Virus in Absence of Specific Antibodies.

Summary Ferrets which had been infected with either influenza A or influenza B virus were tested for immunity to those 2 viruses on the eighth day of convalescence. They were susceptible to infection when reinocu-lated with the heterologous virus against which they possessed no antibodies, but were immune to infection when reinoculated with the homologous virus against which they possessed antibodies. If the mechanisms of infection with the A and B viruses are the same the results indicate that non-specific cellular refractoriness plays little part in the resistance of ferrets to reinfection with influenza virus.

Effect of Intranasally Administered Immune Serum on Influenza Virus Present in the Mouse Lung.

Summary When mice were infected by the intranasal route with large doses of influenza virus and immune serum was introduced into the lungs 1 hour later, the animals died even though neither the amount of infectious virus in the lungs at the time the serum was given nor the amount subsequently produced was as great as the corresponding amounts found in the lungs of mice that had been infected with small doses of virus and that were protected when immune serum was given 24 hours later. The findings suggest that a large amount of influenza virus introduced directly into the mouse lung may, by itself, be sufficient to kill the animal without any subsequent multiplication of the agent.