It is now generally accepted that the increased rate of weight gain in chicks receiving dietary antibiotics depends largely, if not entirely, on consequent modification of the microbial population of the chick's alimentary tract. Fundamental investigation of the phenomenon, therefore, calls for the maintenance of chicks free from indigenous organisms so that a defined flora can be introduced as desired. We deal here with experiments on the rearing of chicks in the Gustafsson (1948, 1959) germ-free apparatus and the effects of dietary penicillin on chick growth in germ-free or conventional environments. E X P E R I M E N T A LChicks. The chicks were produced on our own premises from Light Sussex hens crossed with Rhode Island Red cocks, so that the sexes were distinguishable at hatching by down colour. For much of this work the hens were housed in batteries and artificially inseminated twice weekly, fouling of the egg-shells being thus avoided as far as possible. On some occasions the hens were flock-mated in fold units on grass; some of the eggs from these birds were soiled, and only the cleanest were selected for the production of germ-free chicks.Gerrn-free apparatus. Three of the small light-weight tanks (see P1. I ) designed by Gustafsson (1959) and supplied by Wojidkow and Co. A. B., Malmo, Sweden, were used. The tank consists essentially of a stainless steel box, 34 in. long, 18 in. wide and 20 in. deep, covered by a plate-glass window. Into one of the sides a pair of heavy rubber sleeves is sealed, to which can be fitted rubber gloves of any suitable pattern. A third sleeve is provided on the opposite side. At one comer of the tank is a stainlesssteel U-trap to be filled with germicide, through which objects can be passed into or out of the tank. The air supply is sterilized by passing it through a steel cylinder containing granulated carborundum heated electrically to 3 50"; it is cooled by passing through a tube along the side of the tank. The exhaust air is similarly treated in order to destroy any organisms that may, deliberately or accidentally, have been introduced into the apparatus. In the experiments described here air was passed into the tanks at the rate of 7l./min during the hatching period, and subsequently the flow was increased to 10 l./min. Positive pressure was always maintained in the tanks. M. E. COATES AND OTHERS I963The apparatus was originally designed for rats; small modifications had to be made to meet the more exacting environmental requirements of young chicks. During the hatching period a relative humidity of about 65 % is optimal and was achieved by dripping water slowly into the air-intake tube as it entered the air sterilizer. After hatching, ordinary atmospheric air was supplied through the sterilizer until the birds were 2-3 weeks old, by which time humidity in the tanks began to rise because of the chicks' respiration and of evaporation from droppings and water troughs. At this timethe air entering the tanks was dehumidified bypassing it over a freezing coil at -j' bef...

I. Chicks were reared in germ-free and conventional environments on diets devoid of one of each of the following vitamins: riboflavine, pantothenic acid, vitamin Be, vitamin B,,, folic acid, nicotinic acid, thiamine or biotin. When marked signs of deficiency began to appear the birds were killed. The livers and caecal contents were removed and assayed microbiologically for the vitamin omitted from the diet.2. Negligible quantities of the omitted vitamin were detected in the caecal contents of the germ-free birds, but considerable amounts were found in corresponding samples from conventional birds. There was little evidence that signs of deficiency were less severe in the conventional birds.3. It was concluded that all the vitamins tested were synthesized by microbial action in the gut of conventional birds but that the birds derived little benefit from the synthesized vitamins, with the possible exception of folic acid.4. There were indications that the requirements for thiamine and pantothenic acid were higher in conventional than in germ-free chicks.Experiments with conventional animals have provided many indications that some vitamins are produced in the alimentary tract as a result of microbial activity. For instance, in coprophagous animals such as the rat and the guinea-pig omission from the diet of vitamins of the B complex does not result in signs of deficiency unless coprophagy is prevented or antibacterial agents (e.g. sulphonamides) are given in the diet. The vitamin requirement of chickens reared in contact with their excreta is partly satisfied by ingestion of their own droppings. Inclusion of low levels of antibiotics in the diet of chicks or rats 'spares' the need for certain vitamins, an effect probably mediated through the intestinal microflora.The present investigation was undertaken to determine the extent of vitamin synthesis in the alimentary tract of the chick, and of the availability of the vitamins to the bird. MATERIALS A N D METHODS ChicksChicks of the Rhode Island Red x Light Sussex cross were used. For the experiment on vitamin B,, deficiency, eggs were taken from hens given a diet low in vitamin B,, in order to reduce the amount carried over from the dam to the chick. For the other experiments eggs were taken from hens given a normal breeders' ration. Each experiment was done on comparable groups of germ-free and conventional chicks.The eggs were incubated in a commercial incubator for 18 days, then candled and treated by the disinfection process described by Coates, Fuller, Harrison, Lev & Suffolk (1963). Half were immediately introduced into Gustafsson isolators and half were replaced in the conventional incubator to hatch. After hatching, the germ-free As far as possible males and females were distributed evenly throughout the experimental groups. A total of from twelve to thirty-two birds was used on each treatment. In each IOO g diet the salt mixture provided CaCO, 1.5 g, K,HPO, 1.61 g, CaHPO,. zH20 1.37 g, NaCI 0.837 g, MnSO,.qH,O 25 mg, KI 4 mg, CuSO,. 5H20 1.5 mg, ZnCI21*z5 m...

Procaine penicillin [penicillin (procaine salt)] added to a good practical ration improved the growth of chicks in a laboratory used for poultry for ten years. Chicks from the same batch in two other laboratories, where birds had not been kept before, grew equally well on the ration with and without penicillin, and growth was the same as that on the penicillinsupplemented diet in the usual chick laboratory. The growth depression in the absence of dietary penicillin was not due to differences in management or to recognizable disease. It is suggested that it is due to an ‘infection’ hitherto undescribed and shown to be transmissible and counteracted by penicillin.

Growth depression was induced in chicks and a collection of aerobic and anaerobic bacteria was isolated from the crop and caecum. The collection of bacteria was tested in gnotobiotic chickens for its ability to depress growth with and without a bacteria-free faecal filtrate. None of the anaerobic bacteria depressed growth. Aerobic bacteria always depressed growth but only in the case of Streptococcus faecium was this statistically significant. The faecal filtrate also depressed growth. A statistically significant effect was obtained when faecal filtrate was combined with any of the following treatments: (a) the total collection of bacteria, (b) the aerobic group, (c) the anaerobic group, (d) Strep. faecalis var. [iquefaciens, (e) Strep. faecium, (f) the unclassified streptococci, and (9) a group containing the lactobacilli and coliform organisms. The largest growth depression was obtained in chickens dosed with Sfrep. faecium and faecal filtrate.

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