Iran J Parasitol: Vol. 15, No. 2, Apr-Jun 2020, pp.

272-277

Iran J Parasitol
Tehran University of Medical Open access Journal at Iranian Society of Parasitology
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Case Report

Acanthamoeba Keratitis and Acanthamoeba Conjunctivitis: A

Case Report
*Oktay ALVER 1, Mehmet BAYKARA 2, Merve YÜRÜK 3, Nazmiye ÜLKÜ TÜZEMEN 1

1. Department of Medical Microbiology, Faculty of Medicine, Uludag University, Bursa, Turkey

2. Department of Ophthalmology, Faculty of Medicine, Uludag University, Bursa Turkey
3. Department of Medical Parasitology, Faculty of Medicine, Erciyes University, Kayseri, Turkey

Received 10 Aug 2019 Abstract

Accepted 25 Oct 2019 Acanthamoeba species are vision-threatening agents by causing cornea infections
known as Acanthamoeba keratitis. A 5 year-old kid with the complaints of erythe-
ma, eyelid edema, inflammation, limitation of eye movements in the right eye,
Keywords: and having no history of wearing contact lenses or trauma, was diagnosed of
Acanthamoeba conjunctivi- Acanthamoeba conjunctivitis through laboratory examinations in the Ophthalmol-
tis; ogy clinic. The visual sharpness of the patient improved after the treatment. A 44
Acanthamoeba keratitis; year-old female patient suffering from pain, stinging, irritation, and inability to see
genotype T2 in the left eye with the history of wearing contact lenses or trauma was diagnosed
of Acanthamoeba keratitis through laboratory examinations. The agent was isolated
and identified as "A. castellani" in the Genotype "T2". Examination of the left eye
on the 15th day of treatment indicated that all complaints disappeared except for
*Correspondence the cataract originated visual loss. However, the first diagnosis of Acanthamoeba
Email: keratitis appeared in the literature on a case with no history of wearing contact
[email protected] lenses and trauma it is found to be attention grabbing. We think that Acanthamoe-
ba should not be ignored among microbial agents that cause eye infection with or
without trauma and contact lens usage history.

Introduction

A canthamoeba is a protozoon parasite

exists as trophozoite and cyst living
freely that can be found in soil, dust,
purified and bottled water (1), medical
devices, dialysis, contact lenses and contami-
nated cultures (2,3). Corneal infection such as
Acanthamoeba keratitis (AK) which is caused
some species of Acanthamoeba that recognized

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 Alver et al.: Acanthamoeba Keratitis and Acanthamoeba …

as a worldwide threat with increasing im- Her medical history revealed no contact lens
portance and incidence (4). AK may occur in wear and trauma. Upon identifying conjuncti-
healthy, immunocompromised patients. It has vitis at ocular examination, conjunctival swab
been reported that the most important risk specimen was collected from the conjunctiva
factor in these infections is the wearing of under sterile conditions taking into account
contact lenses in developed countries and ocu- the possibility of bacterial infection and inocu-
lar trauma in developing countries (5). lated onto 5 % sheep blood agar, Eosin Meth-
In this study, we aimed to present a pediatric ylene Blue (EMB) and chocolate agar plates.
case diagnosed as “Acanthamoeba sp.” conjunc- The specimen was inside the plate contain-
tivitis and a female case diagnosed as T2 geno- ing Non-nutritious agar (NNA) with Escherich-
type “A. castellani” keratitis. ia coli for feeding amoeba. No growth was de-
tected during the follow-up of bacteriological
Case Presentation 1 and parasitological inoculations. Identification
A 5 years-old girl had referred to an oph- of Acanthamoeba sp. could not be performed
thalmologist elsewhere with the complaints of since no growth occurred on NNA agar after
gradually increasing swelling, hyperemia and 2 times of inoculation. Some studies in the
discharge and treated with antibiotic eye drops literature (6, 7) as in this study show that there
(gentamycin 0.3% and tobramycin 3%). How- may not always be reproduction in culture.
ever, she had to refer to Department of Oph- This may be related to the very low amoeba
thalmology, School of Medicine at Uludag density due to the fact that our patient re-
University Applied Research Center for ceived antibiotic treatment (gentamycin and
Health since she did not improve with the tobramycin) before admission to the clinic.
treatment after 5 days. Eyelid edema, conjunc- However, the examinations of the direct
tival hyperemia, chemosis and limited eye Giemsa stained specimen under light micro-
movements were noted in her right eye whose scope (X100) revealed Acanthamoeba sp. three
left eye had normal findings and she was hos- cysts and seven trophozoites with pseudopo-
pitalized with the prediagnosis of orbital cellu- dia (Fig. 2).
litis and empirical treatment with systemic
cefotaxime (Sefotaks) 150 mg/kg thrice a day,
systemic clindamycin 50 mg/kg thrice aday,
topical moxifloxacin (Vigamoks) 12 times a
day and topical fluoromethalone (Flarex)
thrice a day was started (Fig. 1).

Fig. 2: Trophozoite of Acanthamoeba sp. in a direct

Fig. 1: a. Chemozis, conjunctival hyperemia and giemsa stained conjunctiva smear (Original pic-
intense secretion (Original picture). ture) (x100)
b. Reduced hyperemia and secretion after Acan-
thamoeba conjunctivitis treatment (Original picture). Therefore, all medications were given up
c. Heal of the right eye after Acanthamoeba con- and topical propamidine isothionate 0.1 %
junctivitis treatment (Original picture) (Brolene), topical neomycin sulphate and
polymxyine sulphate (Cebemyxine) were initi-

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 Iran J Parasitol: Vol. 15, No. 2, Apr-Jun 2020, pp.272-277

ated and continued for 4 months. At 4th day ture plates after 7 days. The patient who re-
after treatment, there was no growth after in- ceived alternating topical fortified vancomycin
oculation of conjunctival swab specimen onto (50 mg/ml), ceftazidime (100 mg/ml), pro-
NNA media and direct Giemsa stained speci- pamidine isothionate 0.1% and chlorhexidine
men of the same specimen was not remarka- at ophthalmology clinic was discharged at 11th
ble. As the patient did not have corneal in- day.
volvement, she recovered completely without In the follow-up, after the inoculation of
sequelae and any visual loss. corneal scraping specimen onto E. coli plated
NNA agar, no growth was identified at 2nd
Case Presentation 2 month visit. The corneal scraping specimen
A 44 years-old female patient pointed out was defined as “T2” genotype “A. castellani”
that burning and itching had started in her left according to PCR and sequence analysis that
eye after sand splash to her eyes at seaside 3 targeted 18S rDNA region performed. DNA
months before she referred to the Department isolation was performed using QIAGEN tis-
of Ophthalmology; School Of Medicine, sue set (QIAGEN, Germany) according to
Uludag University Applied research Center for manufacturers’ instructions. QPCR was pre-
Health. She also told that she had not im- pared using Acanthamoeba specific primer pairs,
proved with the treatment given with the di- SYBR Green (Roche, Germany) and DNA.
agnosis of herpetic infection elsewhere. She QPCR programme was performed by Roche
referred to Department of Ophthalmology at Light Cycler 480 II (Roche, USA) and Cq val-
our center with the complaints of pain, sting- ue of the specimen was detected positive. In
ing, lacrimation and decreased vision in her addition, PCR protocol was carried out by
left eye. She told that she had given up contact amplification of 425 bp DNA fragment of the
lens wear 10 days before her admission to the 18S rDNA gene region with JDP1 and JDP2
clinic. The biomicroscopic examination of the primers. The product of PCR was purified
left eye revealed central keratitis and corneal using electrophoresis (Wizard SV Gel,
scraping specimen was taken and inoculated Promega, USA) and the positive band that was
onto E. coli plated NNA agars and incubated imaged was photographed. Purified PCR
at room temperature (Fig. 3). product was sent for DNA sequence analysis
in order to perform philogenetic analysis. Par-
tial data of Acanthamoeba sp. 18S ribosomal
DNA gene region sequence was obtained.
Phylogenetic tree of the specimen was con-
structed that was subject to sequence analysis
with MEGA 6 and the phylogenetic associa-
tion of the specimen with T2 genotype was
detected by neighbor joining test (Fig. 4). Se-
Fig. 3: a. and b. Corneal- ring shaped infiltrate and quence datum was deposited at GenBank un-
conjunctival hyperemia der the following accession number: (Bank-
It2237548, Seq1 MN099283).
Acanthamoeba sp. cysts were identified in the This study was approved by the Ethics
direct Giemsa stained specimen of corneal Committee of Uludag University Faculty of
scraping. Contact lenses and contact lens solu- Medicine (2016-9/18), and informed consent
tion could not be cultured since they had been was obtained from all patients.
thrown away by the patient. Acanthamoeba sp.
cysts and trophozoites were detected on cul-

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 Alver et al.: Acanthamoeba Keratitis and Acanthamoeba …

Fig. 4: Phylogenetic tree of the specimen named as “Sequence 1”

Discussion thamoeba. However, some PCR based tech-

niques have been currently developed and
Acanthamoeba keratitis maintains its im- usually increase the sensitivity to a significant
portance as is usually misdiagnosed in most of extent (4). The PCR protocol used by
the cases and the lack of consensus on diag- Schroeder et al. (11) is one of the best current
nosis (4). The three most important risk fac- diagnostic PCRs. In addition, genotyping can
tors for Acanthamoeba keratitis development be performed by DNA sequencing of the am-
are contact lens wear, contact with contami- plicon that is obtained by JDP1 and JDP2
nated water and trauma (8). Of patients with primers (9,12). Use of primer sets targeting
Acanthamoeba keratitis, 95% had at least one of 18S rDNA region for classification of Acan-
these mentioned risk factors (9). Our second thamoeba species can define all known subspe-
case also had two of these factors, which are cies and is adequate for detection of genotypes
contact lens wear and trauma. The cases are (11,13).
usually misdiagnosed since the findings of Acanthamoeba spp. were isolated from corne-
herpes simplex keratitis are very similar to the al scraping specimens of 16 patients with kera-
early findings of Acanthamoeba keratitis (10). titis in Hungary and it was reported that with
First case was hospitalized and treated with molecular typing the agent was T4 genotype
the prediagnosis of orbital cellulitis. The Acanthamoeba in all 7 (7/16; 43.75 %) cases
treatment was changed upon diagnosis of con- (14). In Spain Acanthamoeba DNA was positive
junctivitis during ocular examination. The in 87 of 177 healthy contact lens wearers, one
treatment of the second case who had been isolate which was cultured and grew, the geno-
diagnosed as herpetic infection before referral type was identified as T4 (15). The culture re-
our clinic and had no improvement with prior sult of corneal scraping material our second
therapy was changed upon bio microscopic case was positive, the strain was identified as
identification of keratitis findings. T2 genotype “A. castellani” according to mo-
The direct detection of the agent in the cor- lecular evaluation of culture isolate using
neal scraping specimen is the single reliable JDP1 and JDP2 primers. Demirci et al. (16)
diagnostic method. Culturing is the gold reported that they identified Acanthamoeba in
standard in the laboratory diagnosis of Acan- corneal biopsy of a 5 years old child who had
no history of trauma or contact lens wear. Er-

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 Iran J Parasitol: Vol. 15, No. 2, Apr-Jun 2020, pp.272-277

dem et al (17) included 26 patients with con- no effective drug that provides treatment
tact lens associated Acanthamoeba keratitis in alone (22).
their study. In the same study, the positivity
rates of the Acanthamoeba were 15.3% for di- Conclusion
rect microscopy, 46.1% for culturing, 92.3%
for PCR and 100% Q PCR (17). Correct diagnosis and treatment ocular in-
When we review eye literature, these two fection agents taking into account the possibil-
cases are noteworthy such that the first one ity of Acanthamoeba irrespective of contact lens
was the first pediatric case who was diagnosed wear and history of trauma; the protection of
as Acanthamoeba sp. conjunctivitis with no his- individuals from ocular health threatening in-
tory of contact lens and trauma, and the sec- fection by paying attention to hygienic
ond case developed cataract leading to vision measures are crucial.
loss as a result of misdiagnosis of treatment
who had T2 genotype A. castellani keratitis.
Conflict of interest
In our literature review, although Acan-
thamoeba sp. has been found on the nasal mu-
cosa and nasopharynx of healthy individuals The authors declare that there is no conflict
(18,19), and there appears to be solely one of interest.
published report up to date on the presence of
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