Later in the day I selected an earth-worm which had not been exposed to strongyle infection, but which was in a moribund condition. In the intestine there were several free nematoids and also several psorosperms of the genus Monocystis, so well illustrated by E. Ray Lankester. As to the nematoids, which were filariform, they neither corresponded in size nor structure with my strongyle embryos.
At 1 p.m. on the 29th I renewed my examination of the larva removed from the fern-pinnule. It showed a further stage of growth, the male character of the reproductive organs having become apparent. The now tolerably well-formed vas deferens had pushed the chylous intestine on one side, whilst a series of caudal rays, five on either side, supported two narrow membranous wings, which represented the lateral lobes of the hood of the adult strongyle.
At 1.30 p.m. I submitted the intestinal contents of four fresh earth-worms removed from my garden to microscopic examination, but no nematoids were found in any one of them.
About 2 p.m. I removed another large and active strongyle larva that had been reared on another fern-pinnule. It was of the same size as that previously described, but was in the act of changing its skin. It was then put aside along with the other worm under the glass shade.
At 3 p.m. I intended to have examined one or more of the earth-worms placed in the open-mouthed phial, but all had escaped and buried themselves in the fern-mould out of reach.
At noon on the 30th I renewed my examination of the two large larvæ whose developmental changes I had been instrumental in producing from the time of their escape from the egg-coverings. I saw no reason to doubt that the sequence of changes thus far noticed referred to the species of parasite under consideration. Both larvæ were active, but the moulting one had now completed its ecdysis. Its sexual distinctiveness had become yet more pronounced by the formation of two rather short and stout spicules, the point of the tail displaying a very minute awl-shaped projection. The lateral membranes had not visibly increased in size. One of these larvæ, the first under observation, now perished from the injuries sustained during inspection.
Again, and later in the day, I sought to clear up any doubts that might still suggest themselves respecting the source of these larvæ, by once more submitting the intestinal contents of two fresh and uninfected earth-worms to careful scrutiny. In the first worm no parasite could be found, and in the second only one minute nematoid; its organisation, which was sexually incomplete, neither corresponded with my strongyle embryos, nor, so far as I could judge, with Goeze’s Ascaris minutissima microscopica (the Anguillula lumbrici of Diesing and others), nor with Dujardin’s Dicelis filaria. It was a very long and narrow creature, but I lost it whilst attempting to secure an accurate measurement. I should say it was about 1/50th of an inch in length, and not more than 1/1500th in breadth. I made a rough outline sketch of it.
In view of further observations I now placed five more earth-worms in the jar containing strongyle embryos, and I also placed six others in the phial which contained coarser mould, and only a comparatively small number of the original strongyle embryos. The phial was closed with a cork and half buried in the fern-mould of one of my larger Wardian fern-pans. Before this transfer was made I again took an opportunity of ascertaining by microscopic evidence that the embryos lodged in the coarse and fine mould had none of them made the slightest advance in organisation. The worms placed in the jar immediately proceeded to bury themselves.
At noon on the 1st of November I sought to get further results from the only large free larva which now remained to me (for the fern-pinnules on which the larvæ were originally placed had dried up and no third specimen could be discovered). Structurally the larva presented no advance. It therefore appeared to me necessary to place it under new conditions in view of exciting further progress towards sexual maturity and adult growth. To transfer it to the bronchus of a living calf would, of course, have been the crucial experiment, but the hopelessness of getting any satisfactory result from this solitary transfer deterred me from the attempt. On a larger scale, with many larvæ, a positive issue would of course prove decisive. Accordingly, the only thing I could do, in partial imitation of nature, was to try and induce some further changes by placing the larva in human saliva, kept warm artificially. As a first step I immersed the creature in a little of the secretion added to the glass slide, when it immediately displayed very lively movements, such as could only be fitly described as frantic. This encouraged me to replace the slide under one of the fern shades without applying any additional heat. I then left it.
At 12.30 p.m. I selected three of the eleven worms lodged in the infested earth, namely, two from the jar and one from the closed phial, and made a microscopic examination of their respective intestinal contents. In one of the worms from the jar I found several embryos clearly referable to my strongyles, their structure showing scarcely any advance upon that exhibited by the embryos in the mould itself. The weather was now excessively cold and the larvæ were motionless; nevertheless, the application of warmth showed that they were by no means dead. The fæcal matter obtained from the worm that had lived in the phial displayed an immense number of infusoriæ (Bacteria) which rushed about rapidly over the field of the microscope. No other signs of life were detected.