STUDIES ON AVIAN MALARIA IN JAPAN : III. TISSUE CULTURE OF THE EXOERYTHROCYTIC FROM OF THE PARASITE

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  • 日本に見られる鶏マラリアについて : III. 赤血球外発育型の培養
  • ニホン ニ ミラレル ケイ マラリア ニ ツイテ 3 セッケッキュウ ガイ ハツイクガタ ノ バイヨウ

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Abstract

Nlaray studies laave been made on the cultivation of tlae exoerythrocytic form ofmntl;trial parasites. In most of them, however, cultivation was achieved merely by theint xitro transfer of tissues infected with this form by the plasma clot method or thetrypsizt digestion method. Only Mlll, YE, R22), utilizing the plasma clot method, has suc-ceecled in cultivating this form in cultured cells whiclt had been inoculated with bloodparasitized by Plasmodium gallinaceum.An attempt was made to cultivate the exoerythrocytic form in a monolayer cultureof chick embryo cells by inoculation with blood parasitized by P. juxtanucleare. Itwas reported previously that this parasite was easily transferred to an exoerythrocyticform from the erythrocytic form in vivo18). If a method was established for the in vitrocultivation of the exoerythrocytic form, it would be possible to make an attempt toclarify the mechanism of transfer of both forms in vivo.Parasitized blood was obtained from experimental chickens by heart puncture andwas anticoagulated by the addition of 5% citrate solution at the rate of 10:1. It wascentrifuged at 2, 500 rpm for 10 minutes. The sediment was resuspended in Eaglesmedium and recentrifuged at 2, 500 rpm for 10 minutes. The sedimented blood wasresuspended in Eagles medium at the rate of 0.25 per cent. Then 1.0 ml aliquots oftJae resulting suspension were inoculated into cell cultures and incubated in stationaryracks at 37C or 40C for 24 to 72 hours.1VIonolayer cultures were prepared in tubes (containing coverslips approximately6332 rnm) frorn chick embryo cells by the trypsin digestion method. The cell growthmedium employed was Eagles medium (with 10% bovine serum added).After inoculation 2 or more coverslips were removed daily, fixed with methanol, and stained with diluted Giemsas solution for 20 hours. The entire field of each cover-slip was examined for parasites of exoerythrocytic form. The following results wereobtained.l. Alany different types of the exoerythrocytic form were developed in cultures 4. The most suitable conditions for the in vitro cultivation of the exoerythrocyticform consisted of incubation at 40C, contact maintained between the cell culture andthe parasitized blood for 2 -3 days, and a blood concentration of O.25%.5. The addition of bovine or chicken serum did not affect the development of theparasite.6. There was no relationship between the number of inoculated parasites andgrowth of the exoerythrocytic form in cultures.7. When parasitized blood harvested from infected chickens had no detectable?exoerythrocytic form, it was impossible for this form to grow in a culture inoculatedwith it.

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