Extended Abstract
1. Introduction
Acanthamoeba is a free-living amoeba that plenty of them live around us [1]. The high prevalence of this parasite increases the risk of exposure to humans. Because the parasite is opportunistic, under appropriate conditions, human health at risk and caused acanthamoebiasis disease. The neurological disorders caused by this parasite often lead to death and the treatment of visual disorders has not been effective so far [2] because the parasite in vitro and in vivo environments become highly resistant cysts and hard to be treated and controlled [3]. 
Although a wide range of chemical drugs can be used to treat the disease, they may have side effects [5, 6]. Therefore, the searches for an alternative natural and traditional medicine rather than chemical drugs continue [2].
 Recently the effect of some herbal compounds in the treatment of this disease has been studied and confirmed. Since human beings are exposed to high-frequency waves through natural and artificial sources, the effects of these waves on the control or treatments of microorganisms including parasites have also been addressed in some studies [8, 9]. In this study, we investigated the effect of microwave radiation on Acanthamoeba cysts in vitro.
2. Materials and Methods
An experimental study was performed on Acanthamoeba cysts in vitro. The parasitic cysts were cultured and a parasitic suspension containing 19000 to 20,000 cysts per ml was prepared. Sixteen test tubes containing 100 µl of parasitic suspension were prepared and divided into two groups including 8 tubes [16]. These tubes were irradiated by microwaves oven with a power of 1550 watts and a frequency of 2450 MHz. The contents of 8 tubes were irradiated with continuous microwave in 0-120 s time interval (first group) and the next 8 tubes were irradiated with repetitive microwave in 0-360 s time interval (second group). In both groups, the first tube with zero radiation time was considered as the control. The mortality of cysts at the end of irradiation was recorded and compared with control. The temperature of the parasitic suspension was recorded before and after the irradiation and the difference between the initial and final temperature of the experiment was recorded as ΔT. Each experiment was repeated 3 times. 
3. Results
The results of this study showed that continuous and repetitive microwave irradiation resulted in 100% death of the parasite cysts in 120 and 360 s, respectively (Table 1 and 2). ANOVA analysis between the rate of the parasite after continuous microwave irradiation indicates that the parasitic mortality rate in the control group was significantly different from other irradiation times (except 10 s) (P<0.0001). Statistical analysis also showed that parasite mortality rates were significantly different between all continuous irradiation times (except for 30 and 90 s) (P<0.0001).

ANOVA analysis between the rate of the parasite after repetitive microwave irradiation indicates that the parasitic mortality rate in the control group was significantly different from other irradiation times (except 10, 30, and 60 s) (P<0.0001). 
4. Discussion
Acanthamoeba is a pathogenic organism that produces resistant cysts. These cysts lead to the ineffectiveness of conventional drugs against this parasite. Cysts of this parasite are resistant to chemical drugs or require overdosage of drugs to eliminate cysts. So finding and using drugs of natural origin or other treatments that destroy the cyst wall or wall synthesis obstacle have been the focus of researchers [17]. 
Also, the use of therapeutic properties of mechanical factors such as sound waves has been considered in recent years. So that some researchers use the waves (mechanical or electromagnetic) to eliminate pathogenic organisms and treat diseases. One of the first medical applications of electromagnetic waves has been to destroy cancer cells [20]. In recent years the effects of High-intensity Focused Ultrasound (HIFU) and microwave on some parasitic agents have been proven [8, 9, 15]. 
In this study, the effect of microwave as a high-frequency non-ionizing radiation with thermal effects on highly resistant Acanthamoeba cysts was investigated. The results showed that the effect of microwave irradiation on these cysts depends on the duration of irradiation. On the other hand, the mortality rate of Acanthamoeba parasites in continuous and repetitive microwave irradiation did not exceed 6% during 105 and 270 s, respectively, but when the irradiation time increased to 120 and 360 s respectively, the mortality rate rapidly increased to 100%.
The results of the study showed that during continuous irradiation of the microwave for 105 s, the mortality rate of Acanthamoeba cysts was only 5.7%, and ΔT about 83°C. However, during the repetitive microwave irradiation for 270 s, the ΔT was about 42° C, and the mortality was almost similar to the continuous irradiation mode. According to these results, the change in the mode of microwave irradiation, from continuous to repetitive, resulted in a reduction of ΔT but did not affect the mortality rate of Acanthamoeba cysts. Therefore, it seems that the effect of microwaves is not only caused by the increase in ambient heat but also another property of this type of wave is involved in the death of Acanthamoeba.

Ethical Considerations
Compliance with ethical guidelines
The project was approved by the Ethical Committee of Arak University of Medical Sciences (IR.ARAKMU.REC.1394.192).
Funding
This study extracted from a research proposal approved by the Arak University of Medical Sciences. We received financial support from the Deputy for Research and Technology of this University.
Authors' contributions
Conceptualization and Review & editing: Zahra Eslamirad and Homa Soleimani; Investigation: Reza Hajihossein, Zahra Eslamirad, and Homa soleimani; Validation: Zahra Eslamirad; Supervision: Homa soleimani.
Conflicts of interest
The authors declared no conflict of interest.
Acknowledgements
The authors would like to thank the Research Deputy of Arak University of Medical Sciences for their financial support of this research (Number: 2348).