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Pembuatan Kultur In Vitro Stadium Eritrosit Plasmodium Knowlesi A1-H.1

1Department of Epidemiology and Tropical Diseases, Faculty of Public Health, Universitas Diponegoro, Indonesia

2Department of Environmental Biology and Tropical Medicine, College of Medicine, Kangwon National University, Indonesia

Received: 13 Sep 2023; Revised: 30 Oct 2023; Accepted: 5 Nov 2023; Published: 1 Dec 2023.
Open Access Copyright (c) 2023 MEDIA KESEHATAN MASYARAKAT INDONESIA under http://creativecommons.org/licenses/by-sa/4.0/.

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Abstract

Latar belakang: Plasmodium knowlesi adalah spesies Plasmodium yang terkenal mampu menyebabkan malaria zoonosis pada manusia. Infeksi P knowlesi dapat menyebabkan penyakit yang bersifat parah hingga kematian. Parasit malaria ini dapat menginfeksi darah manusia dan juga kera. Kultur in vitro parasit stadium deritrosit (blood stage) jangka panjang sangat memungkinkan untuk P falciparum dan P knowlesi. P knowlesi saat ini digunakan sebagai model eksperimental untuk penelitian in vivo, ex vivo dan in vitro. Penelitian ini bertujuan untuk  mengkultur P knowlesi yang dilakukan secara kontinyu dalam kondisi laboratorium.

Metode: Penelitian ini merupakan penelitian yang menggunakan rancangan eksperimental dengan pendekatan observasional. Kami menggunakan strain P knowlesi A1-H.1 (PkA1-H.1) dengan beberapa kondisi laboratorium, seperti optimasi serum dan konsentrasi glokosa yang digunakan. P knowlesi di kultur selama 10 hari dan morfologi parasit dikonfirmasi dengan pewarnaan giemsa 10%.

Hasil: Strain PkA1-H.1 mampu mempertahankan kemampuannya untuk menginfeksi 100% sel darah merah manusia dalam kondisi laboratorium. Serum AB manusia dan serum kuda (10%, v/v) merupakan kondisi optimal untuk pertumbuhan PkA1-H.1. Total 2-4 gram dekstrosa anhidrat merupakan glukosa optimal yang dibutuhkan parasit untuk tumbuh. Parasit tumbuh perlahan pada 5 hari pertama setelah proses kultur dilakukan (thawing), namun pertumbuhan bisa meningkat pesat setelah hari ke 6 kultur. Parasit nampak sehat dari hari pertama hingga hari kesepuluh, yang dibuktikan dengan penampakan morfologi ring, tropozoit dan schizont.

Simpulan: Hasil ini menunjukkan pentingnya faktor pertumbuhan yang optimal seperti konsentrasi serum dan dekstrosa untuk mendukung pertumbuhan PkA1-H.1. Keberhasilan kultur in vitro bisa membantu penelitian tentang biologi invasi, vaksin dan skrining obat baru.

Kata kunci: malaria; zoonosis; P knowlesi, kultur in-vitro

 

Title: Re-setting-up the Continous in-vitro Culture of Plasmodium knowlesi Blood Stages

Background: Plasmodium knowlesi is well-known Plasmodium species causing zoonotic malaria in humans. The infection of P knowlesi cause severe illness to death. This simian malaria parasite infects both macaque and human bloods. Long-term in vitro cultures of blood-stage parasites are feasible for Plasmodium falciparum and P knowlesi. However, P knowlesi is recently used as experimental model for in vivo, ex vivo and in vitro studies. This study aimed to culture continously the P knowlesi in laboratory condition with human red blood cells.

Method: This study used an experimental study design with observasional approach. We used P knowlesi A1-H.1 strain (PkA1-H.1) under several laboratory conditions, such as serum and glocuse concentration. The culture was conducted for 10 days. The morphology of parasites was confirmed using 10% Giemsa staining.

Result: PkA1-H.1 human-adapted strain was successfully maintained ability to grow under the 100% human red blood cells. Pooled-human AB serum and horse serum (10%, v/v) was optimal condition for PkA1-H.1 to grow. Total 2-4 gram of dextrose anhydrous was optimal glucose required for parasite to grow. The parasites grew slowly in the first 5 days after thawing, but the growth increased rapidly after day 6 of culture. The morphology of parasites was normally observed as seen healthy rings, trophozoites and schizonts.

Conclusion: This result showed the importance of optimal growth factors (serum and dextrose) to support the PkA1-H.1 under the laboratory condition. Its successful of continous culture highlighted the use of culture for understanding of the invasion biology, vaccine or drug discoveries.

Keywords: malaria; zoonosis; P knowlesi, in vitro culture

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Keywords: malaria; zoonosis; P knowlesi, kultur in-vitro

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