1. Introduction

Acanthamoeba spp. are free-living amoebas (FLA) widely distributed in the environment, including soil, water, and air [1]. They are capable of causing cerebral (granulomatous amebic encephalitis, GAE) and extracerebral (Acanthamoeba keratitis, AK; Acanthamoeba pneumonia, AP; cutaneous acanthamoebiasis and disseminated acanthamoebiasis) life-threatening infections among humans [2]. Acanthamoeba spp. exist as an active (trophozoite) and dormant (cyst) forms. The cysts can abide and survive in various environments for years and transform into trophozoites under favorable environmental conditions [3].

Cerebral and extracerebral infections caused by Acanthamoeba spp. occur mostly among immunocompromised individuals, including HIV positive, organ transplant recipients, patients with chronic diseases, and those who undergo immunosuppressive therapy. Acanthamoebiasis has also been observed in immunocompetent individuals [4,5]. Despite low occurrence worldwide, the mortality rate of Acanthamoeba spp. infections is very high. The pathogenesis of acanthamoebiasis is not fully understood. Acanthamoeba spp. may invade through different ways, such as the respiratory tract or breaks in the skin, resulting in hematogenous dissemination to the brain [6]. Acanthamoeba spp. infections cases are often underdiagnosed and hence strong clinical suspicion along with laboratory technical expertise is required for early diagnosis and therapeutic intervention [7].

2. Biology of Acanthamoeba spp.

Acanthamoeba spp. exist in two distinct forms: an actively feeding and dividing trophozoite (15–45 μm) and a dormant cyst stage (12–25 μm) [8]. Both stages are infective to humans. Trophozoites of Acanthamoeba spp. actively feed on bacteria, yeast, algae and small organic particles. Cysts are formed after exposure to unfavorable environmental conditions, including changes in humidity, temperature, or environmental pH [4]. Cysts are resistant to many chemical and physical factors, including UV radiation, osmotic pressure, disinfectants, and antiseptics [9,10]. They can survive in the environment for more than 20 years [11].

3. Genome of Acanthamoeba spp.

Acanthamoeba species are identified based on the analysis of the diagnostic fragment 3 (DF3) region of the ribosomal DNA gene, designated Acanthamoeba-specific amplimer S1 (ASA.S1) [12]. DF3 encodes the highly variable stem 29-1 of the nuclear small subunit 18S rRNA gene [13]. Acanthamoeba genotypes are distinguished by a 5% or greater sequence difference between isolates [14,15]. To date, 22 sequence types have been identified, designated T1 through T22 [16]. However, Booton et al. [17], examining the DF3 subregion of ASA.S1 in the samples of T3 and T4 isolates, observed several unique sequences within T3 and T4 isolates. These sequences were designated T3/1–T3/5 and T4/1–T4/10 [17]. Due to the fact that different laboratories applied redundant numerical labels to identify different sequences, Fuerst and Booton [16] suggested new subtypes for the T4 genotype, which have been labeled T4A–T4F and T4Neff. The authors also described different sequences within T3 (labeled T3/01–T3/13), T5 (labeled T5/01–T5/15), T11 (labeled T11/01–T11/14), and T15 (labeled T15/01–T15/11). Types T2 and T6 are the most closely related pair of sequence types within Acanthamoeba, and these two types are considered as a supergroup in which five subtypes are formed: T2, T2/6A, T2/6B, T2/6C and T6 [16].

Among all genotypes, T4 is the most frequently isolated from nature, and it includes many pathogenic strains that have been associated with neurological and pulmonary acanthamoebiasis. It is assumed that T4 is characterized by increased virulence and increased resistance to chemotherapeutic agents [18]. The genotypes T1, T2, T4, T5, T10, T11, and T12 are the factors of granulomatous amebic encephalitis [13,19], whereas T2, T4, T5, T16, and T18 may be the factors of Acanthamoeba pneumonia [20,21,22,23].