When staphylococcal strains from hospital patients are plotted in a frequency curve according to the size of the growth-free zone in a disk test for sensitivity to penicillin-G, this curve is trimodal. This implies that, besides the sensitive, there are two populations of resistant staphylococci: the moderately and the highly resistant, In a hospital-wide survey of coagulase or mannite positive staphylococci isolated in North Carolina Memorial Hospital between October 1964 and October 1965 it was inferred that: (1) there was a statistically highly significant difference between the distributions of the moderately and the highly resistant strains over duration of hospital stay prior to the time of isolation; (2) the highly resistant strains were seldom introduced from outside; and (3) there was no significant association between use of penicillin-G by a patient harbouring a strain and the frequency with which such a strain was found to be highly resistant to this drug. The latter finding is in contrast to significant associations found when the penicillin resistance is moderate, or when the drug used is other than penicillin-G. It is concluded that a fall in the incidence of staphylococcal strains highly resistant to penicillin-G in patients who did not receive other antibiotics may well be a valid index of the efficacy of programmes to control hospital infection. If so, there is a practical merit in adopting the routine of distinguisning between two groups of penicillin-resistant staphylococci, which behave, epidemiologically, as two distinct bacterial populations.

A modification of the growth-inhibition test for identifying Mycoplasma species is described. The modification simplifies the screening of a large number of strains for one species. Experiments showed that it was effective and specific when used to identify M. mycoides var. mycoides. Its use in studying the epidemiology of contagious bovine pleuropneumonia is discussed.

This work was financed in part by the United States of America Agency for International Development under the terms of the CCTA/AID Joint Project 16 for research on contagious bovine pleuropneumonia.

We wish to thank Dr R. H. Leach for the various bovine mycoplasma cultures.

Laboratory aspects of a programme of sewer swabbing during and after the Aberdeen typhoid fever outbreak, 1964, have been described. Results were discussed and reasons suggested for the relative insensitivity of methods employed in the isolation of Salmonella typhi. It was tentatively concluded that generalized sewer swabbing in a large city was not likely to be very helpful in tracing undetected excreters of S. typhi during and immediately after a major typhoid fever outbreak. The cost of the laboratory side of this investigation, for staff salaries, equipment, media, etc., was somewhat over £7000.

The authors wish to express thanks to Dr I. A. G. MacQueen, M.O.H., Aberdeen, and his staff including Mr H. B. Parry, Chief Sanitary Inspector and staff, for willing and helpful co-operation throughout this programme. Grateful acknowledgement is also made to Dr B. Moore of the Public Health Laboratory, Exeter; to Dr E. S. Anderson of the Enteric Reference Laboratory, London, for ‘phage typing’ the numerous isolations and to Dr Joan Taylor of the Salmonella Reference Laboratory, London, for assistance in typing Salmonella spp. isolated. Especial thanks are due to our laboratory technicians and other staff for the willing support given during the investigation.

Influenza vaccines used in trials appeared to stimulate a small increase of saline-reactive anti-A1 antibodies in group O volunteers. There was no evidence that haemolysins were elicited by adjuvant vaccine. It is unlikely that influenza vaccines would cause ABO haemolytic disease in infants.

The mechanism of spread of an epidemic of variola minor (alastrim) occurring in 1956, in Vila Guarani, a semi-rural school district of the City of São Paulo, Brazil, is examined. Cases were grouped according to time and space relationships corresponding to either a household or a school class, as once variola minor was introduced into a household or a school class spread was independent of occurrences in neighbouring social units. The constant occurrence of a single introduction in school classes and households implied that the chance for transmission of the disease outside the homes or the school was very small. The secondary attack rate was higher in households than in school classes. Twenty households with cases had more pre-school children and school children and a lower vaccination level than twenty-two neighbouring households without cases but also including one or more persons who were known to have had personal contacts with patients.

Examination of the chain of contagion clearly shows that spread in the school was directly and indirectly responsible for the majority of cases in the epidemic, for the peak of the epidemic curve and for the extent of topographical spread of the epidemic. This predominant role of the school was accentuated by incomplete preventive measures (vaccination campaign and isolation of patients) applied late in the epidemic and it is attributed to the following facts, (a) There was a single school in the district, and school children were thus concentrated, (b) The school was the only place of the district where persons living in different neighbourhoods had regular, repeated contacts, (c) The great majority of school children had had no previous variola and many had no vaccination scar, (d) The social activities of school children gave them the highest potential as transmitters of variola minor.

Two hundred and twenty-two infants and fifty-one staff were examined for carriage of Staphylococcus pyogenes and Gram-negative bacilli, and the presence of these bacteria in the ward environment was also investigated. Staphylococci were phage-typed and tested for sensitivity to antibiotics. Assessment of clinical infection caused by these bacteria was made.

Infant carriage of Staphylococcus pyogenes was maximal on the sixth day, 73·1 % of umbilical swabs yielding significant growth. Carriage of Gram-negative bacilli was maximal on the fifth day, 31·7% of umbilical swabs being positive. Where special nursing was enforced, i.e. premature baby unit, isolation of Gram-negative bacilli was very infrequent.

The low nasal carriage of staphylococci seen in infants and nursing staff is attributed to traces of penicillinase-resistant penicillins in the ward air.

Staphylococcus pyogenes was recovered from 23 % of ward surfaces, and constituted 0·7% of the air-borne flora. In contrast, Gram-negative bacilli were recovered from 4·7 % of surfaces, and composed only 0·15% of the air-borne flora.

Phage group I constituted 73·0, 44·9 and 41·2% of strains from infants, staff and ward environment respectively.

Antibiograms were grouped into three categories, A, B and C. Category A (sensitive to chloramphenicol, erythromycin and methicillin but penicillin-resistant) composed 79·0% of infants' strains, being found mostly in group I, but was rare among staff strains.

Of 54 specimens taken from 40 infants showing evidence of infection, Staphylococcus pyogenes was isolated from 16, of which 14 were category A. Gram-negative bacilli were isolated 7 times. Only 21 infants required antibiotic therapy (9·3%) and none were infected with Gram-negative bacilli—this contrasts with the frequency of these bacilli in nursery infection today.

Ward air is considered to be the main depot of Staphylococcus pyogenes. Infant ‘dispersers’ are probably the principal source of the air-borne staphylococci. Infants and staff abstract from the air those staphylococci which become their colonizing strains. No transfer is thought to occur from the staff to the infants. Transmission of Gram-negative bacilli is similar in that the infants are the principal source and the air the main depot of these bacteria. Nasal colonization by Gram-negative bacilli is low, this is possibly due to poor colonizing properties. Infant umbilical carriage almost certainly results from endogenous faecal spread.

We are grateful to Dr J. C. Gould, Director, Central Microbiological Laboratories, Edinburgh, for his continued interest and advice, and to Mr J. C. Ferguson and Mrs M. M. Winton for skilled technical assistance. This study would not have been completed without the co-operation of Sisters McCartney and Struthers and their nursing staff.

Mycoplasma pneumoniae is a common respiratory pathogen in the south of England. Of 1150 sera from normal people, 19% had antibody against this organism. Antibody was found with significantly greater frequency in patients with chronic bronchitis.

Live type 4 adenovirus contained in enteric-coated tablets was given to 2628 military recruits. In two trials, the enteric adenovirus vaccine reduced febrile and afebrile adenovirus illness by about 90% or more. Moreover, a decided protective effect against subclinical adenovirus infection as evidenced by seroconversion was found.

The antibody-stimulating effect of enteric adenovirus vaccine was not an adequate measure of its protective capacity. The neutralizing antibody response of recruits given vaccine was poor in comparison with the antibody response to naturally acquired infection with type 4 adenovirus. The latter response was related to the severity of infection.

We are indebted to Colonel Dr B. J. W. Beunders, Head of the Division of Preventive Medicine of the Royal Netherlands Army, for granting us facilities to conduct this study and to Misses W. T. C. J. Smulders, E. M. J. C. Vallas, J. E. M. Dusseldorp and M. M. J. P. Keeris for their valuable technical assistance.

Two hundred and ninety-three milk ring test-positive individual milk samples were examined by whey complement fixation and agglutination tests and the results were compared with the isolation of B. abortus by cultures made from the milk.

There was a closer correlation between whey CF tests and isolation of the organism than between whey AG tests and isolation. Brucella organisms were not isolated from 116 MRT-positive milk samples when the whey CF titre was < 1/10; whereas 9/104 (9%) isolations were made from milk when the whey AG titre was < 1/20.

Because of its closer correlation with culture results it is suggested that the whey CF test would be valuable in an eradication programme for the detection of infected udders.

The limitations of the milk ring test for the examination of individual milk samples are emphasized and it should be used only as a preliminary screening test. In the individual milk samples examined, 32% of the MRT-positives showed no evidence of infection by either the whey AG, whey CF, or culture tests.

1. When mice are infected intraperitoneally with Langat virus only a small proportion develop clinical encephalitis, but all mice have substantial titres of virus in the brain and also incontrovertible histological evidence of encephalitis.

2. When specific antibody is given intraperitoneally or intravenously to mice during the first 3 days after intraperitoneal infection with Langat virus, the viraemia (normally maximal during this period) is depressed, the production of antibody is depressed or delayed, and the incidence of clinical encephalomyelitis is increased significantly.

3. Specific antibody given intraperitoneally or intracerebrally before infection, protects the animals from encephalitis.

4. These findings are discussed in terms of the histology of the central nervous system of the affected mice.

We are very grateful to Miss S. J. Illavia, B.Sc., and to Miss G. E. Fairbairn for their skilled technical assistance; and to Mr S. Peto of the Microbiological Research Establishment for statistical advice.

This work was made possible by a generous grant from the Wellcome Trust and the Endowment Funds of St Thomas's Hospital

Genetic marker tests were performed on 61 strains of poliovirus isolated during the Blackburn outbreak from cases or suspected cases and their contacts. The results were correlated with the epidemiological data and good agreement was found for the serological and dextran inhibition tests. The reproductive capacity temperature marker, however, showed inhibition at 39·8° C. of strains otherwise judged to be of naturally occurring type. These strains could be differentiated from those of vaccine origin because they grew readily at 39·2° C. This feature also distinguished the strains isolated in Blackburn from those obtained in the nearby town of Hyde in a simultaneous outbreak.

Dr J. Ardley, Medical Officer of Health, Blackburn, kindly provided a key which enabled the contacts of cases to be traced; and Dr L. Robertson, Public Health Laboratory, Preston, made available his extensive records as well as the many strains isolated in his laboratory.

The growth of vaccinia and cowpox in RK13 cells was studied by measurement of infective virus production, gel diffusion and haemagglutination. The effects of virus growth inhibitors on the normal course of infection were then followed. Although the results with the two viruses were essentially similar some differences were detected between them. Inhibitors of DNA synthesis permitted production of many virus-specific soluble antigens. Compounds which inhibit DNA function and compounds that affected energy-yielding reactions had more dramatic effects. The different results obtained suggested that the synthesis of virus-specific materials was sequential and a possible part of the sequence is suggested.

The results of a survey of the foot-and-mouth disease carrier state in three localities of an FMD enzootic area of Botswana are presented. The collection, storage and handling of the samples is mentioned and the isolation of virus on calf thyroid monolayer tissue cultures is described. Type SAT 3 virus was isolated from up to 20% of the animals sampled at periods between 7 and 12 months after natural infection. Sera were tested for all the animals from which virus was recovered and compared with similar numbers of sera from virus-negative animals. No correlation was found between the carrier state and serum antibody titres.

Antigenic studies are described which indicated not only an antigenic variation of some carrier viruses isolated from the outbreak strain but also antigenic differences between strains of carrier virus from different animals in the same herd. There was no evidence at the time of sampling of the persistence as carrier virus of the attenuated vaccine strains of FMDV used previously in the cattle sampled.

The results suggest that transference of carrier virus from animal to animal may occur in the absence of clinical infection.

Two groups of calves were dosed orally with 104, 105, and 106S. typhimurium at approximately 2 days of age and at 14–21 days of age. No obvious clinical signs were observed with this strain, but younger calves excreted the salmonellas in the faeces for longer periods than older animals. The younger animals also excreted more salmonellas per gram of faeces in the first week following dosing. These observations may explain why salmonella cross-infection is likely to occur where very young calves are congregated. Following cessation of excretion of salmonellas in the faeces and subsequent slaughter of calves, examination of all the mesenteric, caecal, and colic lymph nodes showed these to be the most useful sites for recovery of salmonellas.

Provided good nutrition and hygienic conditions prevail, calves retained on their farm of origin for longer periods are more likely to recover from neonatal salmonella infections.

1. Irradiation in a whole body dose of 200 rads or more increased the sensitivity of mice to intraperitoneal infection with Langat virus so that the LD 50 was increased to about the intracerebral LD 50.

2. In mice given 500 rads before infection: (a) viraemia was prolonged by about 5 days; (b) the IgM response was depressed; (c) the IgG response was delayed by about 3 days and depressed in titre; (d) virus concentration in the brain rose continuously until death on about the tenth day while in the controls it reached a peak on the fifth day then subsided; (e) histological changes in the CNS were delayed and minimal even at death; (f) irradiated mice died with little evidence of paralysis while the controls died with severe paralysis.

3. In irradiated mice, protection was observed when antibody was administered on the third day following infection. Antibody given on the 3 days after infection to control mice aggravated the disease.

4. The results in this and the preceding paper are discussed in relation to the pathogenesis of encephalitis. It is concluded that neuronal damage is caused both by virus multiplication in neurones and by damage superimposed by inflammatory changes with associated oedema and hypoxia. The inflammatory changes appear to be due to an allergic reaction to virus-antibody complexes formed in the circulation and in the central nervous system.

We are grateful to Miss S. J. Illavia, B.Sc., and Miss G. E. Fairbairn for their skilled technical assistance; to the Department of Radiotherapy at St Thomas's Hospital for providing time and staff to help with the irradiation experiments; and to Mr S. Peto of the Microbiological Research Establishment for statistical advice.

This work was made possible by a generous grant from the Wellcome Trust and the Endowment Funds of St Thomas's Hospital.

Two hundred and sixty-four samples of sera obtained from normal healthy persons belonging to different age groups were examined for haemagglutination-inhibiting antibodies against reovirus types 1–3 to assess the prevalence of reovirus infection in the northern part of India.

Reovirus infection appeared as early as 6 months to 1 year of age. With reovirus types 1 and 2, the maximum incidence of 67 and 48% respectively occurred by the age of 10–20 years, whereas with type 3 infection the maximum incidence (73%) was reached by the age of 25 years. The incidence of reovirus type 2 infection in all the age groups was remarkably low in this series. A drop in the incidence of reovirus type 2 infection was also noticed after the age of 20 years.

The difference in geometric mean titre of antibody in different ages against reovirus type 2 was highly significant, suggesting probably that most of the infection occurred by the age of twenty years. The difference in the geometric mean value of the titres of antibody against types 1 and 3 was not significant in different age groups. The levels of antibody against types 1 and 3 in all the age groups were almost the same, suggesting that infection, specially with reovirus type 3, was occurring in all the age groups even beyond 20 years of age.

The authors wish to express their sincerest thanks to Dr Leon Rosen, Pacific Research Section, National Institute of Allergy and Infectious Diseases, Honolulu, Hawaii, for the supply of prototype strains of reoviruses, to Drs B. K. Aikat, Director Professor of Pathology, R. N. Chakravarti, Department of Experimental Medicine, O. N. Bhakoo, and Sucheta Thukral, Department of Paediatrics, Post- Graduate Institute of Medical Education and Research, Chadigarh, for their invaluable help and co-operation, to Prof. P. V. K. Iyer, Department of Mathematics, Panjab University, and to Mr H. D. Gupta, Biostatistician, Post-Graduate Institute of Medical Education and Research, Chadigarh for their statistical analysis of the results.

The authors gratefully acknowledge the helpful criticism and suggestion of Dr C G. Pandit, Ex-Director, Indian Council of Medical Research, during the preparation of the paper.

The authors acknowledge also the technical assistance of Mr Jagmohan Lai Pipat, Mr Gurmeet Singh Khatra, and Mr Amar Singh Saini.

This work was partly supported by the Research Grant of the Indian Council of Medical Research sanctioned to one of the authors (S. R. P.).

The capacity of certain strains of Streptococcus pyogenes to produce opacity in aged horse serum has been studied. Cells from all stages of the growth cycle are able to produce opacity. Maximal activity is reached towards the end of the exponential phase of growth.

Examination of cell fractions obtained by mechanical breakage and differential centrifugation suggested that the cell-bound activity is predominantly associated with the membrane fraction. Extraction with sodium deoxycholate yields a soluble fraction of high activity.

There is considerable strain variation in heat stability of the serum opacity factor. Cell-bound activity is often quite resistant to heat, whereas extracted activity is less stable.

Low concentrations of divalent cations have an activating effect, whereas high concentrations inhibit the serum opacity reaction. High concentrations of uni-valent cations are without effect on the cell-free enzyme but have an activating effect on the cell-bound enzyme.

For both the cell-bound and the cell-free enzyme the pH optimum was 5·8.

Although sensitive to trypsin and pepsin, the serum opacity factor appears to be resistant to streptococcal proteinase. Its activity is destroyed by formaldehyde and by periodate but is unaffected by a number of reducing agents.

Pre-heating of the serum or the addition of iodoacetate did not affect the serum opacity reaction. The enhanced cholesterol esterification previously described with fresh serum appears to be a secondary reaction. Even when isolated by relatively gentle methods, α-lipoprotein serves as a substrate only in the presence of crystalline serum albumin.

Sheep, after infection with louping ill virus and after re-infection with the same strain of virus 19–21 months later, were bled at intervals and their sera examined for neutralizing and haemagglutinin-inhibiting antibodies. Each antibody type was measured by the constant serum/variable virus and constant virus/variable serum methods. The persistence of each type of antibody and its significance in epidemiological studies is discussed. The relationship of antibody levels in ewes and their lambs was also examined.

1. From the Municipal abattoir, specimens of liver, spleen, lymph node, surface meat, bile and faeces from a sheep and a bovine, were examined fortnightly for 14 months for the presence of possible pathogenic bacteria. The results suggest that slaughtering procedures are satisfactory.

2. Offal, consisting mainly of tripe and intestine, is eaten in large quantities by the Bantu population and is both nutritious and economical. The high incidence of salmonella isolations in the tripe (48%) and intestines (29%), and faecal E. coli and Cl. welchii, show that this commodity is distributed in an inadequately cleansed condition.

3. Faecal E. coli was isolated from 86% of samples of sour milk collected from street pedlars.

4. Dog faeces collected from the township pavements yielded 21% salmonellas, and faeces from fowls sold live by shopkeepers 14%.

5. From the offal specimens, S. typhimurium (23%) and S. london (18%) were the salmonella types most frequently isolated. S. dublin was isolated on only one occasion.

6. Throughout the survey no shigellas were isolated.

7. The significance of the Cl. welchii, coagulase positive Staphylococcus aureus and B. cereus isolations from the various specimens tested is not known.

8. It is emphasized that although there is a definite need for improved treatment of the offal before distribution to the consumer, the resulting increase in cost must not be such as to deprive the population of this important source of protein.

We wish to thank the Director of this Institute, Professor J. H. S. Gear, for permission to publish this paper, Dr P. J. Meara, Director of the City of Johannesburg Abattoir and Livestock Market Department for facilities granted, and Mr C. J. Franklin for collecting all the abattoir specimens. Our thanks are also due to Dr A. Smith, Medical Officer of Health, and Dr I. H. F. Spencer of the Johannesburg City Health Department for help and advice; to Mr A. J. MacG. Bryce for collecting all the specimens from the Soweto township; to Mrs Leonie Jonk for typing the salmonella organisms and Mr R. G. Robinson for investigating the micro-organisms' sensitivity to antibiotics.

The purpose of surveys of amoebae or other organisms is the estimation of the carrier rate in a population, that is, of the proportion of persons who carry a given parasite. As a rule, the tests do not infallibly detect the presence of the organism in a known carrier. Thus simple estimation from a single examination of each member of a sample is not applicable.