AN 1NTRACORPUSCULAR PARASITE IN THE BLOOD O F COTTUS BUBALIS AND COTl'UX XCORPI US.

By HERBERT HENKY, M.D., B.S.(Lond.).

(YLATE XVI.)

THIS parmite was first Pound in blood films prepared from specimens of Cottus babalis and Cottus scorpius caught in the rock pools of Port Erin Bay during September 1909, and a brief verbal description of it was given a t it meeting of the I'athological Society of Great Britain, held a t Guy's Hospital in January 1910 (2). Inasniuch, however, as its appearance did not conform with that of any recorded piscine blood parasite, a published description was withheld in the hope of obtaining Curt her material .

Of the Port Erin fish only five out of twenty-six were found to be infected, and ill one of these five the parasite occurred side .by side with a hmuogregarine, H. cotti (Brumpt and Lebailly, 1904 '). It was found again in three out of thirty-seven specimens of Cottus caught in Bobin Hood's Bay during the summer of 191 1, but in none of these spccinieus was the hzeriiogregarine found (Henry, 1913 3). Of the Plymouth fish, four out of tweuty-three were found to harbour the hzmogregariiie, but throughout this series there was no infection with the Port Erin parasite. Hence, in the examination of eighty-six sample of Cottzis obtained from different sources, six individuals were found to be infected with €X cotti, eight with the parasite which is the subject of this communication, and but one individual showed the presence of both parasites.

In the case of two of the Port Erin fish, only a few minutes' exauiination of a blood filui is necessary to discover an infected corpuscle, but iu the reruaining three, where the infection is slight, the search iiiay have to be continued for fifteen to twenty minutes before finding a single parasite. In the Robin Hood's Bay fish, on the other hand,

[lieceived for publication, July 27, 1913.1 Communicated to the P;ithological Society 01 Gieat Biitsiii and Ixcland, June 27-28, 1913.

INTRACORPUSCULAR PARASITBS IN PISB. 225

the infection is so scanty that it would certainly have been overlooked altogether but for one's previous experience with the Port Erin fish.

As a rule an infected corpuscle harbours only one parasite (Plate XVI. Figs. 1 to 15), but infection with two or three parasites is not uncommon (Figs. 1 6 to 30). Occasionally four parasites may be seen in the same host cell, and once five parasites were found occupying the same corpuscle. Infected corpuscles show no alteration in size or in shape, and there is no apparent loss of haemoglobin, for the cytoplasm stains evenly and with the same tilietorial affinities as does that of the surrounding normal corpuscles. Moreover, the nucleus of the host cell is unaltered either in size or in position or in its staining reactions.

The parasite is an irregularly roulld or oval body which lies embedded in the protoplasm of the red blood corpuscle. It vmies in length from 2 to 4.5 p, the average being 3.4 p, and has a breadth of 1 to 3 p, the average being 2.2 p. I have never seen it inside or even impinging on the red cell nucleus.

There has been no opportunity of examining fresh blood, so that there is no description of the parasite in its living state. In air-dried films, fixed in absolute methyl alcohol and stained with Leishman or Giemsa, only the periphery of the parasite is visible. Of this stained margin, one to two-thirds is apparently taken up with chromatin, the remaining portion being pale blue. The chromatinic material occurs as an irregular flattened band which is often broken up into granules of various sizes. These granules may be confluent, but not in- frequently the chain is broken up into isolated portions, which seem to wander round the margin of the parasite like beads on a string. In overstnined films the blue part of the periphery may be entirely masked by the eosin taken on by the cytoplasm of the host cell, and in like circumstances even the chromatin granules iiiay be indistinct. If, however, in place of a Ronianowsky stain, one makes use of Unna's polychrome niethylene-blue or of Rorrel blue, then the pyotoplasm of the corpuscle takes on a pale sea-green tint, against which the periphery of the parasite is sharply diflerentiated into the two portions above mentioned-the pale blue portion and the deeper chromatin portion. I n Gienisa and in Leishman preparations the body of the parasite seems to be unstained, for the centre is slightly paler as contrasted with the more deeply staining corpuscular proto- plasm which surrounds it. It would appear to be embedded between two layers of red cell cytoplasm, which together are thinner and therefore stain less deeply than the rest of the Corpuscle cytoplasm. This feature is more evident in microphotograplis than in specimens viewed microscopically, and it is therefore not improbable that the body actually stains a faint blue, which is not appreciable to the eye and which becomes obvious only by the use of suitable colour screens.

Another distinguishing feature of these bodies is their variation in

226 HERBERT HENRY:

contour. Both the blue and the chromatinic portions of the periphery show many depressions or protrusions, so that the bodies assume an extraordinary variety of forms (Plate XVI.).

I n summing up, then, one niay give to the parasite the following distinctive features, namely :-

1. It is an oval or rounded globular body which exhibits great plasticity, and hence assumes many different shapes in the corpuscle cytoplasm.

2. Only the periphery of the parasite is readily stainable, and of this periphery one-third to two-thirds is taken up with ;I thin band of chromatin, which is often broken up into irregular beads and fragments.

At first one experienced considerable difficulty in reaching any satisfactory conclusion with regard to the nature of this parasite. Professor Minehin, whom I have to thank very cordially for examining films submitted to him, expressed the guarded opinion that it might represent the young form of a hamaniceba-like organism. Its obviously amceboid character seemed to favour such an assumption, but it differs in one particular from all the known parasites of hzmamceba type in that it does not produce melanin. Further, no forms suggestive of either schizogony or of sporont formation, such as one finds in hurnan malaria, have ever been seen. One was faced, then, with two possible explanations as to its nature : (1) either it is a parasite quite distinct from those already known to occur in Cottus, or (2) it may represent a phase in the life-history of one or other of the blood parasites already recorded in Cottus, i.e., of Trypanosoma cotti or of ~ ~ ~ ~ o ~ r e g a r i n a catti. One was left, therefore, in a dilemma with regard to the parasite until an unlooked-for solution presented itself during the investigation of Hwtnogregarina simondi (Henry, 1913 ”. It has been found during this investigation that Namao- gregarincx, simondi exhibits a granule-shedding phase, and that along with this phenomenon there are to be found in the red corpuscles of the vertebrate host, Solea vulgaris, cell inclusions which pass through a certain cycle of development. One of the phases in this cycle I have described as the “ peripheral cliromatin phase,” and the forms of this phase met with in Solea are comparable to those which occur in the present case. I believe, therefore, that the organism described herein is none other than the “ peripheral chromatin phase ” of Hmmogregarina eotti.

REFERENCES.

1. ERUMPT ET LEBAILLY . . “Description de quelques nouvelles espbces de Trypanosomes et d’HQmogr6garines de Tklkostdens marins,” Conapt. rend. Acatl. d. Sc., Paris, 1904, tome cxxxix. p. 613,

JOURNAL OF PATHOLOGY.-VOL. S V I I I PLATE XVI. I

INTRA CORPUSCULAR PARASITES IN FISH. 227

2. HENRY . . . . . . . “On the Hzemoprotozoa of British Sea-fish (A Preliminary Note),” Journ. Path. and Bacterial., Cambridge, 1910, vol. xiv. p. 463.

3. ,, . . . . . . . ‘<A Summary of the Blood Parasites of British Sea-fish,’’ Ibid. , 1913, vol. xviii. p. 218. ’

4. ,, . . . . . . . “The {Granule Shedding of Hmmogregarina sinion&,” Ibid., 1913, vol. xviii. p. 240.

DESCRIPTION OF PLATE XVI.

FIGS. l-l5.-Single infections. FIGS. 16-23.-Double infections. FIGS. 24-30. -Treble infections.

The centre of the body of the parasite is paler than the surrounding corpuscle protoplasm. The periphery of the parasite is differentiated into a pale blue portion and a chromatinic portion, the latter being frequently broken up into irregular granules.

All the corpuscles are taken from preparations stained with Leishnlan’s stain.