A simultaneous investigation into certain epidemiological aspects of scarlet fever was carried out over a 2-year period at Oxford, Cambridge and Cardiff. The results obtained at the three centres are compared and contrasted.

Nose and throat swabs were taken from 648 patients on admission to hospital; group A haemolytic streptococci were demonstrated in the swabs of 75·3 %. Eight days after admission 81·9 % had given positive swabs.

Serological typing showed that 56·9 % of strains were types 1–4. The only other common types were types 11 and 8/25.

The average length of stay in hospital was estimated for all cases, divided into groups according to whether or not they had suffered from complications or been cross-infected. The average for the whole series was 30·6 days; this was increased to 33·4 days for all complicated cases and to 37·6 days for all those cross-infected. It was longest (42·3 days) when there were both complications and cross-infections.

The incidence of cross-infection with fresh serological types of haemolytic streptococci was noted both in hospital and for the first 3 weeks after discharge. In the whole series 20·1 % of cases were cross-infected in hospital. The cross-infection rate was highest at Cardiff (27·9 %) and lowest at Oxford (13·5 %). Of the 123 cross-infected cases, complications occurred in 23·6%.

A total of ninety-eight patients (16·4 %) suffered from complications; in twenty-three of these (23·4%) complications were attributed to cross-infecting strains.

Nose and throat swabs were taken from all patients shortly before discharge; 60·3% were still carrying haemolytic streptococci.

Data were obtained regarding the rate at which cases became free of haemolytic streptococci. By the 10th week of the disease only 3·2 % of those who had not been cross-infected were carrying streptococci. Cross-infected patients were slower in becoming negative; at the 10th week 25·8% were still carriers.

There were fourteen return cases (return case rate 2·1%) following the return home of eleven primary cases (infecting case rate 1·7 %).

Swabbing of home contacts at the time of the patient's admission to hospital showed that one or more contacts of 25·5% of cases carried in their naso-pharynx the same type of haemolytic streptococci as that infecting the patient; 12·8% of the contacts swabbed were positive. During the first 3 weeks after discharge 24·6 % of contacts, that is one or more contacts of 55·5% of cases, carried a type that had been harboured by the patient while in hospital.

It was noted that in a particular area the common types that gave rise to scarlet fever also commonly caused other streptococcal infections of the upper respiratory tract. Type 12, however, though frequently found in other streptococcal infections and in healthy carriers, appeared only rarely to cause scarlet fever.

It is suggested that the low incidence of complications, less intimate relationship between complications and cross-infections and low return case rate found in this investigation, as compared to earlier observations made in England, are associated with lowered virulence and invasive powers of the strains of Str. pyogenes responsible for the much milder scarlet fever now prevalent.

The bacteriological findings confirm the wisdom of modern fever hospital practice of early discharge from hospital. It is suggested that discharge within 3 weeks should be the universally adopted rule.

1.Phase variation of the somatic antigen I in Bact. paratyphosum A and of the component XII2 in Bact. typhosum and British strains of Bact. enteritidis has been confirmed.

2. Strains of Bact. enteritidis isolated in India by blood culture from cases of invasive disease in man fall into two serological types, neither of which undergoes demonstrable somatic phase variation.

3. The probable mechanism of this type of variation is discussed in the light of experimental findings.

4. The effect which phase variation may have on the clarity of end-points in agglutinin titrations and on the serological standardization of Salmonella O-suspensions is demonstrated.

5. The variation is briefly discussed from the points of view of Salmonella philogeny and of human Salmonella infection.

I am indebted to the D.M.S. in India for permission to publish this paper.

1. Attention is called to the widespread dissatisfaction with the standard of reproducibility attained with the existing disinfectant tests.

2. It is suggested that this lack of reproducibility is inherent in any 100% mortality test.

3. The trend in all recent disinfectant developments appears to be towards greater specificity than was the case with the older products.

4. Where a comparison is being made to ascertain the true relative efficiency of two or more disinfectants, this possible specificity necessitates their examination under a wider range of conditions than has been catered for in the past.

5. A technique is suggested for use in those cases where a more limited examination will provide all the information that is required. It employs a simple culture medium, uses a measured quantity for the transfer from the medication tube, and makes use of a compensated photo-electric circuit to determine the amount of growth produced by the organisms which have survived the contact with the disinfectant.

I wish to acknowledge my grateful thanks to the directors of Messrs Cooper, McDougall and Robertson Ltd. for permission to publish this paper, and to my colleagues of the Cooper Technical Bureau for assistance in various directions.

When individual responses in a biological assay show considerable variation associated with the values of a concomitant variate, covariance analysis may be used in order to adjust the mean responses and to improve the precision of the assay. Usually this is preferable to the choice of an adjustment which involves an arbitrary assumption about the effect of variations in the concomitant variate on the measured response. Published accounts of the process are open to certain theoretical objections, though they may be sufficiently exact for most practical purposes.

The present paper describes a method of calculating the relative potency, and its precision, which may be a little more laborious, but which is in full accord with standard statistical practice. The computations are illustrated on data from a prolactin assay by the pigeon crop-gland technique, in which the final crop-gland weight showed a positive correlation with the body weight at the start of the assay. The results are compared with those obtained either from the unadjusted crop-gland weights or from these weights expressed as proportions of body weights. The covariance method leads to a more precise estimate of the potency of the test preparation than do either of the others; there is evidence, however, that the increase in precision will not necessarily be large unless the correlation between the response and the concomitant variate is very close.

In a final section, the full statistical tests of assay validity in the covariance analysis are described; these are lengthy, and fortunately are required only when the validity is in considerable doubt.

The methods of adjustment have been described in this paper with respect to an assay depending upon parallel regression lines of responses on the logarithms of doses. They may be adapted for use with ‘slope-ratio’ assays (Bliss, 1946; Finney, 1945; 1948; Wood & Finney, 1946), in which the regression of response on dose itself is linear. So far the need for adjusting for concomitant variation in these assays seems not to have arisen, and discussion of computational details may be postponed until the need is felt.

1. A method is described for estimating the disinfecting power of chemical substances on small cylindrical blood-clots of standard dimensions (about 1·5 mm. in diameter) containing a measured number of bacteria (Staph. aureus). It is capable of giving guidance for the use of disinfectants under the described conditions.

2. No practicable disinfectant regularly sterilized heavily infected clots (400,000 cocci) in 5 min. at 17–20° C.

3. In 30 min. the following were the only substances that were uniformly successful with heavily infected clots: 2·5% iodine in aqueous KI solution; 50% Chloros, containing about 5% chlorine; 70% alcohol containing 0·1% acriflavine. (The first of the three was tested on a larger scale than the others, and its potency is therefore the more firmly established.)

4. Reduction of the number of cocci in the clot increased the range of effective disinfectant solutions. Thus with 4000 cocci per clot regular clot disinfection was achieved in 30 min. with the following additional solutions: ethyl alcohol 70 and 50%; Chloros 25%; phenol 2·5% and lysol 1·25%. A few tests indicated that Zephiran (and probably CTAB) at 1 or 2% concentration would also pass this less exacting test.

The studý of the Fusiformis necrophorus culture ‘132’ showed that new pure LI cultures could be isolated from it, which have already been maintained for one year and undergone more than 100 passages without reverting to their original type. The new cultures resemble the old L 1 strains from Streptobacillus moniliformis in every respect except for their strict anaerobiosis. It has been observed that swellings of the bacteria develop into large bodies not distinguishable from the elements of which the pure L 1 culture consists.

In the light of the new observations on Fusiformis necrophorus which are in full agreement with those of Dienes & Smith it is still not possible to disprove the symbiosis theory, for the newly established L 1 cultures have not reverted so far. Secondly, though it has been seen that L 1 forms develop from swellings of bacteria, the possibility that a parasite might behave in a similar way cannot be excluded.

On the other hand Dienes's opinion of the bacterial nature of the L 1 component cannot be disproved either. However, we know yet too little about the normal developmental cycle of bacteria to be able to decide whether the L 1 form is an independent entity or whether it may be a normal stage through which bacteria have to pass from time to time or whether it may represent a ‘loss-mutant’ which some bacteria might produce under not yet known conditions. If in the future the bacterial nature of the L1 could be convincingly proved we would be faced with the fact that bacteria can assume a form in which they grow as a thin shapeless slime containing nuclear structures arranged as bodies, rings or very finely dispersed particles—not the dumbbell pattern so characteristic for young bacteria of all kinds—and that further very small parts of the material (filterable granules?) are able to reproduce the L1 culture which at least can go on for many years without reverting into the bacterium from which it was derived.

The work began with efforts to isolate a type II Vi phage from local sewage. When this attempt failed, typing material was obtained from Dr J. Craigie and Dr A. Felix.

The typing method was applied to 495 strains of typhoid bacilli, the technique laid down by Craigie being followed throughout.

It was found that in South Africa the distribution of the various types of typhoid bacilli is different from that obtaining in other countries where this typing method has so far been applied.

During further investigations of type F strains the fact came to light that by means of a simple biochemical test type F strains can be subdivided into two sub-types, which subdivision will be useful in further epidemiological work. No such biochemical subdivision was found possible in any other type.

It did not prove possible to type all South African strains with the available phage preparations. Some of these strains must represent new types. Workers in other countries have had similar experiences and it will become necessary to compare all these unusual strains directly with one another in order to reach uniformity in nomenclature. It has already been possible to co-operate with workers in England, and they have been assisted by my finding that a strain, unique in England and supposed to have been brought there by a carrier, who had typhoid fever in South Africa 40 years ago, was endemic in South Africa.

During the course of the work, two outbreaks of typhoid fever occurred in which the typing method proved most useful, not only for linking up a particular carrier with a particular outbreak, but also for excluding an already known carrier who was under grave suspicion.

I have to thank Dr Craigie and Dr Felix for supplying material, and Prof. Pijper, of the Pretoria University, for placing laboratory facilities at my disposal. I thank all others for cultures received for typing.

The details are given of the epidemiology of S. thompson infection in a flock of pullets which survived an outbreak of this disease when chicks, and also of a flock of adult birds suspected of being carriers of the organism. The results of blood testing and bacteriological examinations of cloacal swabs, random faecal samples and eggs from both flocks indicated that:

(1) Many chicks which survived an outbreak of S. thompson continued to carry the organism for some months without showing any symptoms. In most cases the organism was harboured in the intestines and was excreted intermittently in the faeces. On one occasion S. thompson infected the gall bladder, and the bacilli were excreted in the faeces for at least 18 months after the outbreak had occurred.

(2) The common method of egg infection was by the contamination of the shell with infected faeces. Under conditions of incubation the bacilli penetrated the egg shell and infected the yolk. Although there was little penetration of the organism under storage conditions, the bacilli on infected shells remained viable for at least 21 days.

(3) The common methods of spreading infection in a hatchery were from:

(a) The contact of infected and non-infected egg shells.

(b) The handling of eggs before and during incubation.

(c) The contact of egg shells with infected incubators.

(d) The ingestion and inhalation of infected fluff and incubator debris at hatching time.

(e) The ingestion of food and water contaminated with infected faeces from survivor chicks.

For the control of S. thompson infection in poultry, the following procedures have been recommended:

(1) The production of agglutinins by carrier birds was not a reliable indication of infection. In known infected flocks, however, the detection of carriers by blood testing and by the examination of cloacal swabs is of value. For such a test an alcoholized antigen is preferable to a heat-treated broth antigen, and a titre of 1/20 or more should be regarded as an indication that a bird is infected.

(2) Only clean eggs should be used for hatching. Dirty egg shells should be cleaned by scraping or brushing and not by wiping with a damp cloth.

(3) Fertile eggs should be stored in a cool, dry atmosphere for as short a period as possible before incubation.

(4) Eggs should be fumigated in the incubator with formaldehyde vapour, not later than 24 hr. after they have been set.

We wish to record our appreciation of the assistance received from Mrs E. Sidery in the bacteriological examination of eggs and the preparation of agglutination tests.

Presence of thermolabile agglutination-inhibiting substances in strains of the genus Shigella has been demonstrated by numerous authors. Archer (1942) used as antigen for agglutination tests with S. alkalescens cultures which had been killed at 100° C., since a thermolabile factor which prevents agglutination by the usual technique had been found present in the organism. The same method was employed by Mendes Silva (1943) in agglutination tests with S. ambigua. Braun & Unat (1943) found in S. paradysenteriae (Flexner) a labile antigen designated ‘O1’ which inhibits O-agglutination of the living bacteria. Schuetze (1944) reported that the insensitivity of S. dysenteriae (Shiga) to agglutination is abolished by heating at 100° C. for 30 min. Weil, Black & Farsetta (1944) reported that boiling for 1 hr. renders inagglutinable strains of S. paradysenteriae (Flexner) fully agglutinable. Recently, Weil et al. (1946) reported that one of the Sachs types contains an inhibiting substance, a heat labile antigen and a heat stable antigen. He observed inhibition of agglutination of living bacteria in many Flexner types, S. alkalescens and S. ambigua. Olitzki, Shelubsky & Koch (1946) reported that there is present in S. dysenteriae (Shiga) a labile substance ‘I’ which inhibits the agglutination of thd living bacteria. They were able to demonstrate a labile antigen in toxic saline extracts but not in the intact bacteria. The present paper reports a method for isolation of this antigen in the form of a relatively concentrated preparation.

The numerical aspects of bacterial growth have hitherto been investigated chiefly in fluid media and the laws governing the development of bacterial populations in fluid cultures are known in some detail.

The effect of penicillin alone, and with sulphonamide upon various Gram-negative bacteria, has been investigated.

S. typhi, S. paratyphi B and strains of Proteus vulgaris were inhibited by penicillin in concentrations of 8–20 units/c.c. Certain coliforms were inhibited by 30–128 units.

Organisms which were inhibited to some extent by penicillin and sulphonamide acting separately were inhibited under experimental conditions at lower concentrations when both agents acted together. The mechanism of this reaction has been discussed.

A proportion of the insensitive coliform strains were identified as penicillinase producers.

It has been shown that in all cultures of B. proteus and B. coli a large-cell type with two to four ‘chromosomes’ develops first and produces, after a few hours of growth, by sudden subdivision the smallest unit, a generation of small cells, each with one nuclear structure only. This cell type, singly or in filaments, persists until the end of the growing period. On a ‘swarming plate’ of B. proteus a number of these biphasic growing periods may follow each other.

‘Fusion cells’, in which the chromatinic structures have contracted into a longitudinally arranged bar, are formed when young cultures are stored in the cold.

In a cytological investigation of three branching and two non-branching strains grown on Loewenstein medium, it was found that avian tubercle bacilli contain chromatinic material which gives a positive Feulgen reaction and is readily stainable with Giemsa's solution after treatment of the fixed bacteria with hydrochloric acid.

Growing filamentous forms of both ‘bacterial’ and ‘mycelial’ strains from 1 to 2 day old cultures contain variable numbers of irregularly spaced, more or less spherical chromatinic bodies which vary in staining in the same bacillus, some being red, others purple. During the third or fourth day the chromatinic material in the bacteria increases very much until most of it is fused into an almost homogeneous deeply stained column. In the non-branching strains the filamentous forms with high chromatin content soon break up into small mono-or binucleate elements, and the same holds true for the ‘straight’ filamentous forms which are also present in cultures of branching strains. The ‘mycelial’ forms, on the other hand, disintegrate at this time (fourth or fifth day of cultivation), and it is uncertain whether they contribute (by partial fragmentation) to the masses of small mono- or binucleate forms which are the predominant element in old cultures of all the strains investigated.

The chromatinic structures of avian tubercle bacilli have the same staining properties as those of ordinary non-acid-fast bacteria but differ from them in their behaviour during the early development of the bacilli.

Apparatus for determination of the penetration of particulate airborne material through the nose is described. The penetration of aerosols of liquid particles produced by a spray has been measured in rats, rabbits, goats, sheep and men. The penetration decreased with increase in the rate of flow through the nose.

Particles of the order of I μ diameter penetrate the nose at all rates of flow, while very few particles larger than 15 μ can penetrate except at low rates of flow.

The change by which a strain of B. dysenteriae Flexner becomes resistant to sulphonamides is similar in character to the change in B. coli mutabile from white (non-lactose-fermenting) to red (lactosefermenting).

The highest proportion of variants are found in papillae, while no variants are found in the growing margin of colonies.

Reversion does not take place during growth on sulphonamide-free media.

The variation is a direct and heritable response to a chemical stimulus and in adaptation to this part of the environment.

The variation described by Reiner Müller of B. typhosus on rhamnose and of certain coliform bacteria on arabinose may also be of the same nature.

Strains of B. dysenteriae Flexner resistant to sulphanilamide or sulphaguanidine 1/10,000 are only partially resistant to sulphathiazole 1/100,000 and to sodium sulphapyridine 1/10,000.

Technique. The size of (uncrowded) colonies on a plate containing 1/10,000 sulphanilamide or sodium sulphacetamide is a useful criterion of resistance or non-resistance, but a resistant strain must be able to grow on 1/5000 Sa and will not grow on 1/5000 Ssac.

The best culture medium for these tests is Lab. Lemco bouillon with agar, but without peptone.

In concluding I wish to express my thanks for their help to Prof. H. R. Dean, Prof. C. H. Browning, Drs E. T. C. Spooner, A. MacDonald, J. E. McCartney and R. D. Stuart. The sulphonamides I received through the kindness of the British Pharmacopoeial Commission and of Messrs May and Baker.

A review is given of some of the literature concerned with bacterial infections and immunity in invertebrates and cold-blooded vertebrates. Among the former, insects have been most fully studied, although a certain amount is known of the immunological reactions of marine invertebrates. Diseases of insects are mainly generalized, bacteraemic conditions, and the great majority are caused by Gramnegative cocco-bacilli. Diseases of cold-blooded vertebrates also are usually generalized, but bacteria from a very wide variety of groups have been isolated from them.

It is probable that this resemblance in the infections of these widely separated groups of animals is due to the fact that, unlike mammals and birds, with whose reactions we are more familiar, the regulation of their temperature and the constitution of their body fluids is not exact. This toleration of wide variations of their own physical and chemical constitution must reduce their sensitivity to the changes produced by infection, and hence decrease the likelihood of a local reaction, designed to confine the invading organism to the immediate region of its point of entry, i.e. a local inflammation.

The production of humoral antibodies appears to be almost universal in the animal kingdom, although greatly affected by changes in temperature.

The effect of temperature upon the balance between host and parasite is also discussed. This question bears upon the problem of occasional pathogenesis by saprophytes. Even among mammalian pathogens the borderline between parasite and saprophyte is an indistinct one, especially in such cases as Proteus and Pseudomonas pyocyanea, and even those species which are usually regarded as exclusively parasitic may readily be constrained to adopt a saprophytic existence on artificial culture. Where cold-blooded animals are concerned the boundary is even more difficult to draw, and it is possible that under suitable conditions, bacteria which are normally saprophytes may be capable of causing infection. This point will be impossible of proof until more sensitive means of definition of bacterial species are discovered.

Considerable errors arose when the plate count method was used to determine the number of bacteria on dehydrated vegetables. Bacteria probably occurred on the surface of the vegetables only.

Plate counts tended to be greater when suspensions were prepared at higher temperatures over the range 10–40° C. Counts often increased by 20% and sometimes by 50% in the short period allowed in the standard procedure (25 min.), probably owing to multiplication of bacteria. Changes in the volume of suspensions, caused by rehydration of vegetables, led to small errors for which allowance could be made. Larger variations in count arose from sampling errors through differences in the number of bacteria on individual strips and, to a lesser extent, through imperfect distribution of bacteria in suspensions. The relation of plate counts to the actual number of bacteria on the vegetables was uncertain.

However, the error in counting a single sample (— 65% to + 150%) was appreciably smaller than that in counting successive samples from the same factory (about fivefold) and this in turn was less than the variation between different factories (up to a thousandfold). The plate count could therefore be used with some confidence as a general guide to the state of factory hygiene.

The author is grateful for assistance given by Dr M. Stephenson, F.R.S., of the Biochemical Laboratory, Cambridge, and by Dr M. Ingram, Dr H. G. Wager and others of the Low Temperature Research Station, Cambridge.

There have been groups of observers both in this country and in Germany who have been entirely sceptical about the value of the prophylactic application of antiseptics to wounds and who seem to incline to the opinion that such medication is likely to be more harmful than useful; e.g. Colebrook in a special report to the Medical Research Council (1928) stated that ‘the ordinary “antiseptics” have a higher affinity for the leucocytes than for the microbes, and by combining with the former forfeit their microbicidal potential’. Further, he stated that with the exception of optochin and neosalvarsan, substances which have been tried as antiseptics are so rapidly fixed on serum proteins, blood cells, and the fixed tissues, that they fail to impart bactericidal power to the serum. Fleming (1924, 1931) in a number of ingenious experiments showed that injury to leucocytes is the most likely outcome of antiseptic applications to wounds, and has argued that they are more likely to be harmful than useful for that reaon; later (1938), he pointed out the superiority of the sulphonamides to the older antiseptics as tested by his ‘slide cell’ technique and in penicillin he has discovered an antiseptic which in respect of a high ratio of bactericidal potency to its toxicity to mammalian tissues approaches the ideal. This has given the clinical results which would be expected on theoretical grounds. There are, however, limitations to the use of penicillin as a general prophylactic antiseptic application which depend on (a) its failure to destroy some types of bacteria which cause suppuration, (b) its relative lability, (c) its present cost. There is therefore ground for further careful assessment of the best prophylactic antiseptic. Fleming (1940) has in fact admitted that there may be a place for proflavine in the interim dressing of wounds pending surgical treatment. The German opinion on the question of the prophylactic use of antiseptics seems to have been considerably influenced by the observations of Schimmelbusch, who had demonstrated that mice inoculated on a scarified area of the tail with anthrax bacilli could not be saved when the tail was amputated at a point proximal to the site of inoculation even 10 min. after the wound had been infected. It was concluded on this account that the penetration of bacteria from a wound into the deeper tissues was so rapid that local treatment of the wound was obviously futile. Schiemann & Wreschner (1922) and Weise (1922), however, who record these observations of Schimmelbusch, made similar observations with streptococci but showed that with this micro-organism not only tail amputation but also local application of antiseptics was often successful in saving mice so infected. They did not dispute the rapid penetration of the bacteria but concluded that in the case of the streptococcus the body could deal with limited numbers of these micro-organisms, if the local lesions from which they were reaching the general circulation were eliminated. Further, Browning (1943) in a discussion of the antiseptic action of the amino acridine compounds made an extensive survey of the literature on the influence of these substances on leucocytes and concluded that in view of the wide variations in results the particular technique adopted must be an important factor. Hence he suggests there is need for much caution in accepting such results as an indication of what will actually happen in the animal body.

1. Whole cultures of Bact. coli grown under carefully controlled conditions have been subjected to the action of heat at temperatures ranging from 47 to 55° C. and the survivor curves determined.

2. The occurrence of excessive variation between replicate plates of counts made when the mortality exceeds 95 % was very much less evident than when phenol was the lethal agent.

3. At temperatures of 35° C. and below, disinfection of the cultures was never complete, as a permanent population of cells became established. In some cases the numbers of heat-resistant cells reached nearly 0·01 % of the original population, but great fluctuations were observed.

4. The death-rate during the active part of the disinfection was not constant but, in general, increased with time. In the faster disinfections this increase was difficult to detect and the logarithmic death-rate appeared to be virtually constant, but the increase is nevertheless believed to have been present in all cases.

5. The data for all the experiments have been combined to give a composite disinfection curve from which it was concluded that the death-rate was low at first but rose to a maximum at which it remained constant until the mortality had reached at least 99·99%.

6. This finding led to the decision to treat all the experiments as if the regression of log survivors on time had been linear between mortalities of 90 and 99·99%. The standard errors of the calculated regression coefficients were small, so that this method of treating the experimental data appears to be justified.

The authors wish to express their gratitude to Messrs I.C.I. (Pharmaceuticals) Ltd. for their valuable financial assistance, which has enabled one of us (H. E. F. D.) to devote his full time to this work.

From the faeces of a carrier a strain of Salm. paratyphi B was grown repeatedly which—even under conditions most favourable for bacterial motility—was completely non-motile despite its being richly flagellated and optimally Hagglutinable. The strain has in all subcultures, so far, for more than 1 year retained its peculiarity, viz. absence of the motor function of the flagella which morphologically and serologically appears to be normally developed.

This new bacterial variant, the non-motile H form, is considered to be the result of a loss variation which—possibly related to a somatic degradation of the bacilli—affects the genetic factor responsible for the function or the development of the centre for the motility of the flagella.