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Korean J Parasitol > Volume 36(1):1998 > Article

Original Article
Korean J Parasitol. 1998 Mar;36(1):7-13. English.
Published online Mar 20, 1998.  http://dx.doi.org/10.3347/kjp.1998.36.1.7
Copyright © 1998 by The Korean Society for Parasitology
Ultrastructural changes of Acanthamoeba cyst of clinical isolates after treatment with minimal cysticidal concentration of polyhexamethylene biguanide
Hyun-Hee Kong and Dong-Il Chung*
Department of Parasitology, Kyungpook National University School of Medicine, Taegu, Korea.
Received November 06, 1997; Accepted February 04, 1998.

Abstract

In order to understand the action mechanism of polyhexamethylene biguanide (PHMB) to the cyst of Acanthamoeba on the morphological basis, the cysts of four corneal isolates of Acanthamoeba were treated with minimal cysticidal concentration (MCC) of PHMB and their ultrastructural changes were examined by transmission electron microscopy. The most striking change of cysts treated with PHMB compared with normal cysts was the shrinkage of intracystic amoebae, which resulted in the separation of the plasma membrane of intracystic amoeba from endocystic wall. Subplasmalemmal lipid droplets became irregularly shaped. In severely damaged cysts, cytoplasm was aggregated and organelles were severely deformed. Cytoplasmic materials were leaked out through the damaged plasma membrane. Most cysts showed aggregation of nuclear chromatin material. Number of mitochondrial cristae was also reduced. Ecto- and endo-cystic walls were relatively well tolerated. Findings in the present study revealed that PHMB affected mainly on plasma membrane, but lesser on organellar membrane of intracystic amoeba. It seemed likely that PHMB might kill cystic forms of Acanthamoeba by similar mechanism in which this environmental biocide can damage the cell wall of Escherichia coli by binding with acidic phospholipids.

Figures


Fig. 1
Agarose gel electrophoretic restriction fragment patterns for mitochondria DNA of 4 Korean corneal isolates of Acanthamaoeba. M: Hind III digested λ phage DNA as DNA molecular size standard.


Figs. 2-10
Fig. 2 & 3. Normal cyst of Acanthamaoeba KA/E2 isolate. The plasma membrance (arrows) is lined along with endocystic wall. The morphology of nucleus. mitochondria (arrow heads) and other intracellular organelles are normal. The mitochondria had 4-10 cristae. Fig. 2, ×5,500; Fig. 3, ×18,500. Bars = 1µm. Fig. 4. The nucleus with typical central karyosome of control cyst of KA/E3 isolate. ×16,500. Bar = 1µm. Fig. 5. Severely damaged cyst of KA/E1 strain treated with MCC8. ×7,600. Bar = 1µm. Fig. 6. Separation of plasma membrane from endocyst of KA/E2 cyst treated with MCC8. ×23,700. Bar = 1µm. Fig. 7. Irregular shaped subplasmalemmal lipid droplet of KA/E4 cyst treated with MCC48. ×75,000. Bar = 0.1µm. Fig. 8. Severely deformed intracellular organelles and aggregation of intracytoplasmic materials of KA/E3 cyst treated with MCC48. ×9,600. Bar = 1µm. Fig. 9. The number of mitochondrial cristae was decreased, and autophagic vacuole was observed in the cyst of KA/E4 treated with MCC8. ×38,300. Bar = 1µm. Fig. 10. Leakage of intracellular materials (arrows) and lipid droplets through plasma membrane (arrow heads) in KA/E3 cyst treated with MCC8. ×14,000. Bar = 1µm.


Figs. 11-13
Fig. 11. Intranuclear aggregation of chromatin materials of KA/E1 cyst treated with MCC48. ×11,300. Bar = 1µm. Figs. 12 & 13. In the cyst treated with MCC8 for 2 hr, invagination of plasma membrance was observed, and mitochondria and intracellular organelles were not deformed. Fig. 12, ×27,300; Fig. 13, ×16,900. Bar = 1µm. APV, autophagic vacuole; CW, cyst wall; EC, ectocyst; EN, endocyst; K, karyosome; L, lipid droplet; M, mitochondria; N, nucleus; O, ostyole; P, peroxisome; PM, plasma membrane.

Tables


Table 1
Average MCC (µm/ml) of PHMB against four Acanthamaoeba isolates

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