Researches involving dilution egg-counts, recently carried out by members of the staff of this Institute, have shown that the error variance, i.e. that between parallel counts, approximates more or less closely to the mean count. The following examples illustrate this point:—

The counts were all made by the same technique, and it is fair to total them in order to observe the average effect. This agreement between variance (or mean square) and mean is a feature of the Poisson distribution, and it has been assumed that dilution counts are in fact distributed according to the terms of that series, in all cases where the technique is adequate. Reasonable agreement between mean and error variance becomes, indeed, a criterion of the accuracy of the sampling technique. The purpose of the present note is to enquire a little further into the theoretical basis of this assumption, doubts having arisen as to whether high counts (say, up to 100) can be expected to conform to the Poisson series.

The essence of the McMaster (Gordon & Whitlock) technique is that one counts the eggs lying under a centimetre square engraved on a coverglass which is supported 0·15 cm. above a slide, i.e. the eggs contained in 0·15 cc. of faecal suspension. Since the suspending liquid is a half-saturated solution of salt, the eggs float up into one optical plane immediately below the coverglass, this making for ease in counting.

Amongst biological strains or races of the.stem eelworm, Anguillulina dipsaci, some are comparatively polyphagous and are capable of causing disease in a number of different host plants whilst others are restricted to one or two hosts. The strain causing “ tulip root” in oats is usually regarded as a restricted one being Confined to oats or, as in Yorkshire, to oats and beans, in which infection takes place in the seedling stage and persists during the greater part of the life of the plant. Whether the parasite will attack other agricultural plants is a matter of some practical importance and since in a crop rotation a seeds mixture for ley production may follow oats, it was decided to test whether the parasite would infect plants comprising a typical seeds mixture. It was also hoped to determine whether any of the grasses or clovers would be more or less permanently affected and thus serve as reservoir hosts for the parasite and so enable it to tide over the period between successive oat crops. The present paper deals with pot experiments set up to test this matter as a result of which it has been found that the oat strain of the parasite is capable of causing serious injury to the young seedlings of a number of grasses and clovers of a seeds mixture.

In a previous paper, Peters & J. Leiper (1940) pleaded that, in anthelmintic experiments, the statistical problem of variation should be considered, not only so as to assess the significance of treatment differences after the results have come to hand, but also with a view to the prior construction of an experimental design capable of giving reliable answers to the questions posed. Reference was made to the “Enormous expenditure of time, labour and money” on anthelmintic experiments, the results of which usually remain undigested and of unknown significance.

While the paper was passing through the press a symposium on phenothiazine as an anthelmintic, led and summarized by Taylor & Sanderson (1940), was published in the Veterinary Record. The symposium deals with “Work carried out in various parts of the country and co-ordinated through the Agricultural Research Council” it covers some 150 horses, 300 to 400 sheep, 70 to 80 goats, and a few cattle, pigs, and dogs. The drug was found to be remarkably efficient against horse strongyles and the stomach-worms of ruminants, and in varying degree less efficient against other worms.

Extensive tables of egg-counts, worm-counts, and weights of animals are published in the symposium. It was usual to count eggs or weigh animals once before treatment and several times afterwards. The repeated after-treatment determinations are made in order to detect a general trend in egg-counts or weights: in other words there is no attempt to measure and make allowance for the very considerable upand- down variation from day to day.

The earlier work of this Institute on phenothiazine in lambs (Peters et al, 1941; J. Leiper et al, 1941) using egg-counts, worm-counts, and lamb-weights as criteria of anthelmintic efficacy, has revealed conflicting results as between one criterion and another. Thus, the lowest dosage rate used (0·15 gm. per Kg.) led to a reduction in the egg-count, whilst even the highest rate (1 gm. per Kg.) led to no reduction in the worm-count: by an irony of chance the worm-counts were lowest in the untreated controls in both experiments. In the first experiment weights were not affected by the drug, and in the second the only significant effect was a set-back in those lambs given tablets. Numerous other workers have reported increased weights due to the drug, in long-term experiments; ours were too short in duration to show such increases, yet the danger of reinfestation precluded any extension of the time, the lambs being kept outside on infective pastures. The most obvious explanation of the egg- and worm-count results appears to be that the drug has a temporary inhibiting effect on egg-laying, which would presumably return to normal in a few weeks; but here again a prolonged experiment increases the liability to reinfestation.

These being the relevant circumstances, the present experiment was designed to take account of them in the following ways. In the first place the lambs were kept inside throughout the experiment, their stalls being cleaned out thoroughly twice a week to obviate reinfestation.

A total dose of 30 gm. of phenothiazine, given as tablets in one dose of 15 gm. and three of 5 gm. to three goats and six sheep known to be worm-free, had no effect on the weights of the animals in comparison with equal numbers of worm-free controls, over a period of 15 weeks from the first dose.

It is concluded that weights can validly be used as a criterion of anthelmintic efficacy; they not only constitute an obvious economic criterion, but also are statistically more sensitive than egg-counts when subjected to analysis of covariance.

In an experiment previously reported (1941) we were able to show little anthelmintic effect against the gastro-intestinal nematodes of sheep after using phenothiazine powder in single doses up to 0·3 gm. per Kg. body weight. In view of our experience and its contrast with that of many" other workers, we felt it desirable to test a much larger dose, to compare it with another brand of the drug, and (because of the very large variation in worm burden from lamb to lamb) to use not less than four lambs in each treatment group.

Accordingly, 12 lambs from a flock known to be naturally infested were purchased last November, and these were divided at random into three groups as follows: A, untreated controls; B, given compressed 5 gm. tablets of phenothiazine (recently placed on the market by a well-known firm) at the rate of 5 gm. per 5 Kg. body weight; and C, given phenothiazine powder in water as a drench, at the same dosage rate, with the addition to each dose of 1 gm. sodium glycocholate as a wetting agent. This powder was from the same batch as that used in our previous experiment. In each case the full amount was given as a single dose.

The same three criteria of efficacy were used as before: three eggcountings and lamb-weighings before and after treatment, and post mortem worm-counts. Eggs were counted by the same modified McMaster technique, the individual counts representing the eggs in 0·005 gm. of faeces, but on this occasion only three cells were counted per sheep per day.

Hook dimensions of Multiceps glomeratus show that the hooks are capable of reaching full adult size in the coenurus stage.

The available material showed that M. serialis hooks were slightly larger than those of M. glomeratus in the adult stage; in the coenurus stage the differences were not significant.

Statistically, the dimensions measuring the length of the blade of the hook are the most satisfactory.

There are significantly large variances in most cystic hook dimensions from one cyst to another, and from one scolex to another within a single cyst, but not from one group of scolices to another. It follows that in biometric work it is important to sample several scolices from each cyst, and several cysts of any one species, but that groups can be disregarded.

The variability of most hook measurements is in the region of 5%; i.e. the standard deviation is about 5% of the mean dimension.

The danger of extensive generalisation from limited data should be guarded against. Thus, while the species variance was significantly larger than the cyst variance in the case of adult hooks in the material available, it is quite possible that this would no longer have been the case if material from other sources had been available.

Enzymes hydrolysing olive oil and ethyl butyrate (probably lipases and esterases) have been extracted from the intestines of Strongylus edentatus and Ascaris lumbricoides (pig strain).

Maximum enzyme activity was obtained at pH 8·0.

A. lumbricoides enzymes were assisted by sodium glycocholate. Sodium taurocholate had a retarding effect on the enzymes from both parasites.

Enzymes extracted from A. lumbricoides were activated by the presence of NaHCO3 solutions whereas S. edentatus enzymes were not activated by this salt.

Esterase action, relative to lipase action, was 2½ times greater in S. edentatus than in A. lumbricoides.

Tryptic-like enzymes have been extracted from the intestines and intestinal contents of Ascarts lumbricoides and Strongylus edentatus. The action of these enzymes, in relation to hydrogen ion concentration, has been examined.

The parasite trypsins resembled pancreatic trypsin in that the relative production of “free acid” and “formaldehyde acid” was similar, but though acting only on the alkaline side of the isoelectric points of the substrate proteins, optimum action on gelatin, blood-albumin and casein was obtained at pH 6·2.

The amount of protease extracted from a given weight of S. edentatus tissue was far greater than that from a similar amount of A. lumbricoides. Thus Strongylus digested 4·9 to 8·3 times as much gelatin, 12·5 to 40·9 times as much casein and 2·5 to 5·2 times as much blood-albumin as Ascaris.

Spectroscopic examination of the process of digestion of oxyhaemoglobin showed, first, the formation of reduced haemoglobin and then the formation of haematin. These changes took place most rapidly at higher hydrogen ion concentrations.

In the spring of 1941 a male patient died at Peterborough and the autopsy which followed showed that death was due to pontine haemorrhages. As the brain also carried a coenurus, it was submitted to me for further examination. I am much indebted to Dr. David Fulton, M.D., M.R.C.S., L.R.C.P., pathologist and physician to the Peterborough and District Memorial Hospital, and to Dr. L. P. Garrod, M.R.C.S., F.R.C.P., pathologist to St. Bartholomew's Hospital, London, through whose kindness and good offices this interesting specimen was made available to me.

The changes in protein digestion, crude fibre digestion and the chief urinary constituents found in four rats after experimental infection with Trichinella spiralis are detailed.

Diarrhoea and anorexia occurred during the period 8 to 12 days after infection when protein digestion fell to its lowest point. It is suggested that this was due to antiproteases secreted by the adult parasites and to mechanical damage in the intestinal mucosa caused by their movements.

Urinary N rose immediately after infection in “non-resistant” rats. This was followed by a period of decreased N output after which the excretion rate rose steeply.

The urea output also rose immediately after infection in “non-resistant” rats. During the period 4 to 12 days from the time of infection urea excretion fell. Thereafter there was a great increase in its rate of output. Most of these changes have been attributed to toxins elaborated by the adult parasites for massive larval invasion of the tissues probably did not occur till after the experiments were terminated.

Ammonia excretion rose as urinary urea decreased. The fall in urea N was not compensated by the rise in ammonia N + the fall in dietary N intake. The ammonia output returned rapidly to normal suggesting that the excess ammonia was not produced in response to acidosis when the urea excretion rate rose.

Creatine excretion showed a marked fall during the period 4 to 12 days after infection. Urinary creatine and creatinine rose steeply following this period. Possible reasons for these variations are discussed.

In the body cavity of a female specimen of the fly, Sepsis cynipsea L., (Diptera-Cyclorrhapha), caught at this Institute in August, 1930, the writer found eight adult nematode parasites; 3 females and 5 males. The fly was dissected in Ringer's solution in which the worms became actively motile when set free from the host. At the time of dissection it was seen that the fly's ovaries were in an undeveloped state and there is no doubt that this was due to the presence of the parasites within the body cavity. Beyond noting the fact that the worms seemed to be of a mermithid type they were not investigated further at the time but were fixed in hot 70% glycerine alcohol, to which a trace of Nile Blue sulphate solution was added, and were set aside. Although a good number of the flies were dissected during the month of-August no other example was found to be infected with this species of nematode and, as the worms have not been found again since 1930 in casual specimens of the fly captured and dissected here, it seems desirable to describe them since they appear to be new to science. Structurally they bear some resemblance both to Aproctonema entomophagum Keilin, 1917 and to Tetradonema plicans Cobb, 1919 but differ from these species in several anatomical features. Their systematic relationships are discussed in detail later on.

Debray and Maupas (1896) described a disease of broad beans, Vicia Faba L., occurring in and around Algiers, N. Africa, caused by the stem eelworm, Anguillulina dipsaci, the adults of which were considerably larger than from any other host plant attacked by this species; the females being 1·758 mm. to 2·216 mm., and the males 1·716 mm. to 2·016 mm. long. As far as the writer is aware, from the time their paper appeared to the present time no further record of such giant forms of A. dipsaci has been published. During the past few years the writer has encountered similar large forms in diseased broad bean stems, first in material originating in Portugal and on two later occasions in bean plants grown under ordinary field conditions in this country, and it seems desirable to put on record a short account of his observations.

The occurrence of the chrysanthemum eelworm, Aphelenchoides ritzema-bosi (Schwartz, 1911), in an unusual situation, namely within the tissues of tomato fruits, calls for brief report since, though well known as a leaf parasite of a good number of plants, it has not, to our knowledge, been reported previously from a fruit.

Towards the end of October, one of us (S.H.B.) visited a grower who complained of having a number of tomato fruits from which the calyx tissue came away at the slightest touch. These fruits then rotted deeply towards the centre. Two such fruits were taken from the store for examination in the laboratory. One of them was attacked by the fungus Didymella Lycopersici whilst the other was found to contain eelworms. This fruit was sent by S.H.B. to T.G. with a request for a determination of the species of eelworm present and an opinion on whether it was a parasitic species or not.

On receipt of the fruit the senior author cut it in two by a cut passing along the longitudinal axis from the calyx region and through the central fleshy placenta. It was then seen that the discoloured diseased area was about ¼ in. deep by about ⅝ in. broad and occupied much of the solid fleshy base of the fruit. On teasing up a small portion of the diseased tissues in water and examining under the binocular microscope, considerable numbers of small eelworms were found together with numerous eggs and larvae in various stages of growth.