Pernasal and throat swabs taken on 106 newborn babies showed that 99% harboured coagulase-positive Staph. aureus by the time they left the hospital nursery. When the strains isolated from thirty-four of them were phage typed and tested for penicillin sensitivity, it was found that thirty-three (97%) carried at least one penicillin-resistant strain. Phage typing of their subsequent cultures, taken as they became older, demonstrated that they retained these strains for very long periods. Of the twenty-one still under observation at 6 months of age, eleven (53%) were carrying one or more of their original hospital strains, and an additional three (14%) still were consistently yielding an untypable strain believed to be identical to that acquired in hospital. Similarly, of those remaining under observation for the last 6 months of their first year, at least 50% continued to carry their hospital strains.

These babies frequently retained the hospital strains in their throats longer than in their noses. Although their nasal cultures often became negative after the first few months of life, the original hospital strains continued to be recovered from the throats. This would indicate that the throat culture is more accurate than the nasal culture in determining staphylococcal carriers among this age group. Antibiotic-resistant staphylococci in the infant throat may have considerable epidemiological significance, since babies tend to widely distribute their saliva.

The observations suggest that the staphylococci acquired at birth become a normal component of the upper respiratory flora, and may thus prevent new strains of staphylococci from becoming established later. This may explain why newborn babies retain the antibiotic-resistant staphylococci of the hospital much longer than do adult patients. Antibiotic-resistant strains among the general non-hospital population will be increased steadily by babies born in hospitals.

Of thirty-six babies delivered at home 72% acquired Staph. aureus in their noses and throats within their first fortnight of life. The carrier rate among the twenty-eight who were subsequently followed was 65% during the first fortnight period. Only 18% acquired penicillin-resistant strains, in contrast to the previously observed hospital-born group, 97% of whom acquired strains resistant to penicillin. Phage typing demonstrated that the home-born babies who acquired Staph. aureus during their first fortnight carried the same strains throughout the 15–25 week observation period. As in the hospital-born babies, many of them tended to lose the strains from the nose while retaining them in the throat. Strains having phage patterns of Groups I and II were most common among the home-delivered babies, whereas among the hospital-born babies Groups I, III, or untypable strains were most frequent. There was no evidence that the penicillin sensitive strains acquired from the home are carried for shorter periods than the penicillin-resistant strains of the hospital. Babies failing to acquire Staph. aureus within the first fortnight tended either to acquire it in the throat alone, or to remain non-carriers. It is suggested that babies who do not become carriers within the first two months of life are unlikely to do so later.

I owe thanks to many people: to the late Prof. Sir Alexander Fleming for generously extending the laboratory facilities of The Wright-Fleming Institute; to Prof. Robert Cruickshank for patient guidance and invaluable suggestions; to Dr R. E. O. Williams for providing materials and laboratory space for the phage typing, and to Dr Joan E. Rippon for instruction in the phage typing technique; to Dr Agnes H. W. Smythe for the co-operation of the Marylebone and Lisson Grove Child Welfare Centres, and to Mrs Eastham and Miss MacGilley who took the swabs there; to Dr W. Howard Hughes, Dr W. G. Booth, and Dr K. Hart for arranging co-operation of the Ealing Public Health Department; to the Ealing midwives who collected the swabs from the domiciliary-delivered babies; to members of the Wright-Fleming Institute technical staff.

Also, I am grateful to the following organizations for personal grants while conducting the study: The United States Educational Commission in the United Kingdom for a Fulbright Scholarship, 1951–53; The University of California for a grant from the Student Aid Fund; Sigma Delta Epsilon for a Fellowship Award.

A streptomycin-resistant mutant of a virulent strain of Salm. typhimurium was less virulent to mice than the parent strain. The difference in virulence was only observed when mice were infected with small numbers of organisms. Although the mutant strain caused fewer deaths than the parent strain it remained capable of establishing persistent infection in the majority of animals.

The significant difference in the behaviour of the two strains in vivo was that after clearance from the blood multiplication in the tissues began more rapidly with the virulent strain and was progressive, whereas multiplication of the mutant was delayed, was less in degree and was apparently suppressed within a few days of infection.

The mutant strain was apparently identical antigenically with the parent strain, and was equally toxic. The mice dying of infection with this strain did so when the bacterial population had reached a level similar to that in the terminal stage of infection with the virulent strain.

The essential difference between the two strains was the speed and probability of attaining a critical final population. The mutant strain of Salm. typhimurium, both in vitro and in vivo, had a slower growth rate than the parent strain. A possible hypothesis to explain the importance of this finding in relation to the outcome of infection has been discussed.

I am indebted to the Medical Research Council for the provision of a grant towards the expenses of this investigation and to Mr T. Norris and Miss Italia S. Alderton for technical assistance.

Mice surviving infection with a strain of Salm. typhimurium of reduced virulence developed a progressive resistance to reinfection with virulent strains.

The degree of resistance was greater than that of vaccinated mice, although the primary infection had not caused any significant degree of natural selection or higher serum titres of O antibody.

The previously infected animals responded to reinfection with a virulent strain of Salm. typhimurium by gradually restraining bacterial growth. Many of the survivors eradicated the reinfecting strain.

The possible reasons for the difference between vaccinated mice and survivors of previous infection have been discussed.

I am indebted to the Medical Research Council for the provision of a grant towards the expenses of this investigation and to Mr T. Norris and Miss Italia S. Alderton for technical assistance.

For the purpose of determining the immunogenic potency of polio virus, relatively large amounts of concentrated virus material were prepared which had titres of the order of 1010 T.C.I.D.50 per ml. These were obtained by pervaporating large quantities of tissue culture fluid containing approximately 106.5 T.C.I.D.50 per ml.

We wish to acknowledge the valuable assistance received from Dr P. D. Winter and his staff, of the Poliomyelitis Research Foundation, in carrying out the tissue culture titrations of these materials.

The growth curve of rabbit skin-adapted vaccinia virus in the chick chorioallantoic membrane incubated in Hanks' solution showed a drop in titre of virus for about 10 hr. followed by growth. At least 25% of virus, sometimes more, remained infective. A similar fall in titre was observed in heated membranes in which the virus did not grow and this occurred also when membranes, either normal or heated, were infected and disintegrated before incubation.

The growth curve of virus in minced chick-embryo was similar to that in chorioallantoic membrane.

Virus in cell suspensions prepared from chick embryo and incubated in a nutrient medium showed only a small loss of infectivity before growth in some experiments and rarely dropped below 65–70 % of the original titre in others.

These results throw considerable doubt on the view that loss of infectivity preceding growth of vaccinia virus should be interpreted as an essential part of a growth cycle.

A study has been made of the properties of soluble antigen in the brains of infant mice infected intracerebrally with the Flury strain of rabies virus.

Soluble antigen is produced at the same time as infective virus, and reaches a high concentration in a period of 2–3 days.

It can be partially purified by precipitation at pH 4·3. It is partially resistant to the action of trypsin, RNAse and DNAse. It is relatively stable at pH 6–10.

Experimental results suggest that the soluble antigen remains antigenically active after heating at 56° C. and treatment with 0·5% phenol or 0·35% formal-dehyde, but that such heating markedly reduces the ability to stimulate formation of neutralizing antibody.

Rabbits and mice appear to differ in the production of neutralizing antibody following immunization against soluble antigen in which residual live virus was inactivated by heat, phenol or formaldehyde.

It is suggested that this difference may depend on the different susceptibility to traces of incompletely inactivated virus remaining in the immunizing antigens.

The authors are grateful to Miss T. Madsen for her assistance in some aspects of this work. Dr N. Sapeika kindly made available facilities for the in vitro anaphylaxis experiments.

Financial assistance was received from the Nkana-Kitwe and Chingola Poliomyelitis Research Funds.

The Mahoney, Type I, poliomyelitis virus undergoes two ‘cycles’ of multiplication in the CNS of suckling mice—the first within 48 hr. after intracerebral inoculation, and the second approximately 10 days later. The former may not be associated with paralysis whilst the latter usually is.

By to-and-fro passage in suckling mouse brain and monkey kidney-tissue culture there is a gradual increase in the amount of virus obtained during the early multiplication cycle in suckling mouse brains and associated with this is the occurrence of early paralysis in 80% or more of these mice. The virus capable of causing early paralysis in infant mice is gradually lost by direct mouse-to-mouse passage.

The occurrence of two variants is postulated, one of these apparently nonparalytic and multiplying selectively in suckling brains.

The author would like to thank the late Prof. M. van den Ende for his stimulating discussions and advice, Dr P. K. Olitsky of New York for his constant interest, and Mr T. C. Norcott for his able technical assistance.

The behaviour of sheep cells sensitized with tetanus toxins and toxoids in the presence of antitoxin has been investigated.

Sensitized sheep cells were agglutinated in the presence of tetanus antitoxin. The agglutinin titres of such direct agglutinin systems were roughly proportional to the antitoxin content of the sera with some marked exceptions. The discrepancies were associated with the presence of non-specific agglutinins (antibodies other than specific antitoxin) and with the immunization history of the horses providing the sera.

When sensitized cells were used as indicators of the presence of free antitoxin in toxin or toxoid-antitoxin mixtures, the correlation with in vivo tests was quite good at the L+ level of test, but not so reliable at the 0·01 L+ level of test.

Sensitized cells preserved with formalin were found to be more reliable and the results with such materials were more reproducible than those obtained with freshly prepared sensitized cells.

Cells sensitized with different concentrations of antigen gave different titres by direct agglutination with simple dilution of antitoxin; this difference was very much less in haemagglutination inhibition tests.

The LA dose (the volume of toxin or toxoid which when mixed with one unit of antitoxin gave a + + ± end-point) varied according to the sensitivity of the suspension used and the purity of the components of the reacting mixtures.

Prozones (zones of agglutination inhibition in the region of antibody excess) were found to be associated with the immunization history of the horses from which the sera were obtained. In general the more thorough the immunization the longer the prozones found. The length of prozones was proportionately increased with decrease in the concentration of sensitizing antigen. Possible reasons for the behaviour of certain antitoxins in these tests have been discussed.

I am indebted to Prof. M. Raynaud of the Pasteur Institute, Garches, for the supply of tetanus toxoids of high purity.

I am very grateful to Mr Charles Newman for considerable technical assistance.

Quantitative studies have shown that the haemolytic activity of canine complement was in all essentials similar to guinea-pig complement. Canine serum which had been heated at 56° C. markedly enhanced the haemolytic activity of canine complement. The complement-enhancing power of heated canine serum was found to be increased with the concentration of serum up to 20%. Further concentration up to 50 % serum dilution, or the use of undiluted serum, resulted in a reduction of this effect.

Serum aliquots were either pro-complementary or anti-complementary to cn, depending on whether they were heated at 56 or 62° C.

Factors that possibly affect the complement-enhancement were investigated and evidence is presented which indicates that the electric charge on the protein particle is probably responsible for either the pro-complementary or anti-complementary serum activity; the mechanism of this, however, still remains obscure.

The present work shows also that canine complement can be used instead of guinea-pig complement for testing canine sera with virus antigen by the complement-fixation reaction, an advantage of this being that the use of canine complement permits the testing of canine sera which are anti-complementary to guinea-pig complement.

The data obtained are discussed in relation to canine complement and its specific activity.

The authors are indebted to Professor W. I. B. Beveridge for his constructive advice, to Dr R. R. A. Coombs for his helpful comments on the complement activity, and to Dr Christine E. Rice of the Animal Diseases Research Institute, Canada, who has generously supplied the zymosan.

Miss W. Barr and Miss M. Baker gave fine technical assistance throughout all the experiments. Miss M. Lambourne assisted in preparing the manuscript and graphs.

In the course of salmonella epizootics in guinea-pigs due to Salm. enteritidis and Salm. typhi-murium, respectively, salmonella conjunctivitis was widespread, and conjunctival swabbing was found to be a more effective procedure for detecting the spread of infection than the examination of rectal swabs. Eye-swabbing with segregation of eye-positive guinea-pigs and their cage-mates was apparently successful in controlling two Salm. enteritidis epizootics.

Later experimental observations showed that the conjunctival route was a far more effective one for producing systemic salmonella infection in guinea-pigs than the oral route. The 50% spleen infecting dose by the conjunctival route was of the order of 100 organisms, while a dose of 100,000,000 organisms of the same strain of Salm. enteritidis was ineffective by mouth. Blood-borne invasion from the primary conjunctival lesion occurred by the lymphatic route. Gut infection of guinea-pigs infected experimentally throughout the eye was secondary to spleen infection and probably occurred by excretion of salmonellae through the gall-bladder.

The importance of the conjunctival route in natural salmonella infections of guinea-pigs was confirmed by goggling experiments, which showed that, except in animals with complicating conditions such as pregnancy, effectively goggled guinea-pigs were highly resistant to infection in environments heavily contaminated with Salm. enteritidis which produced high infection rates in ungoggled control animals.

I am very grateful to my technician, Mr G. Sandys, for his enthusiastic help, particularly with the goggling experiments.

1. The membrane piece technique for in vitro titrations of the infectivity of influenza virus is described. Rectangles of shell, about 8 × 25 mm., with the chorio-allantoic membrane still attached (membrane pieces) are cut from thirteenth-day fertile eggs. One piece in a test-tube with glucose-buffered salt solution forms an individual assay unit. Five or more tubes are inoculated with each virus dilution. After incubation at 37° C. for 72 hr., with agitation for the first 24 hr. the fluid in each tube is tested for haemagglutinins. From the results at each dilution, an estimate of the 50% membrane piece (MP50) infectivity titre is obtained.

2. Six hundred assay units, with pieces cut from twenty eggs, can be set up by two workers in 1 hr. and used for titration of between three and twenty-four individual virus preparations, depending on the reliability desired for the 50% end-point estimates.

3. With the D.S.P. and PR 8 strains of influenza A virus, the MP50 titres parallel the EID50 titres from egg titrations, but are eight times and twenty times lower, respectively. The MP50: EID50 ratio is the same for various preparations of the same strain, including standard allantoic fluid and chorio-allantoic membrane virus, incomplete virus, and inactivated (heated) allantoic fluid virus. Preliminary experiments with Lee influenza B virus show that slightly different experimental conditions are required, and the MP50 titres are about fifty times less than the EID50 titres.

4. Consistent results have been obtained on titration of samples of the same virus preparation on a number of occasions over a period of several months.

5. A large number of membrane pieces can be used to test each virus dilution, and sampling variations in the MP50 estimates thus made quite small. Statistical data on the reliability of a 50 % titration result, and on the minimum significant differences between two end-points, are given for different values of n, the number of membrane pieces used to test each virus dilution, and of d, the log dilution step.

We are grateful to Mr J. Collins for invaluable technical assistance, and also to Miss I. Allen for help with the computations.