Pythium insidiosum and Acanthamoeba keratitis in a contact lens user

Abstract

The oomycete Pythium and the protozoan Acanthamoeba can cause fulminant and recalcitrant keratitis, respectively. These infections are not only sight-threatening but can also threaten the structural integrity of the eye. A high index of suspicion is required to identify Pythium keratitis given its uncommon occurrence. Acanthamoeba keratitis is most commonly associated with contact lens wear. However, its coexistence with Pythium has not been reported. We present the successful management of a case of contact lens-related keratitis, coinfected with Pythium and Acanthamoeba.

Background

The use of contact lenses is associated with a higher risk of keratitis; bacterial infections being the most common.¹ However, fungal and Acanthamoeba keratitis are also associated with contact lens use and current literature shows an increasing trend in both types of infections.² In temperate climates, a common risk factor for infectious keratitis is contact lens use; whereas in the tropics it is usually related to trauma with organic material.^{1 2}

Pythium insidiosum can cause a fulminant keratitis which is very often unresponsive to medical and surgical therapy; its relentless progression very often necessitates evisceration. The first reported cases of this devastating infection were associated with prolonged contact with water, in farmers in Thailand.³ Increasing familiarity with the pathogen has led to its identification in keratitis from all over the world. Pythium keratitis has also been reported in contact lens users.^4–8

Acanthamoeba keratitis is a relatively uncommon but important cause of keratitis; delays in diagnosis, poor response to therapy and the long duration of treatment can cause significant visual loss. The most common risk factor is the use of contact lenses, followed by trauma.⁹

The management of Pythium or Acanthamoeba keratitis poses considerable diagnostic and therapeutic challenges to the clinician, worldwide. Our first report on the successful management of a coinfection with these two pathogens in a contact lens user is therefore significant and would be of particular interest to clinicians and optometrists, with a focus on contact lenses and anterior segment infections

Case presentation

A 21-year-old healthy man, a known soft contact lens user, presented at our clinic with a history of dust particles falling into his right eye 10 days earlier. He had removed his contact lenses immediately after the injury, but subsequently developed redness, irritation and mild pain in the right eye. At the time of his consultation at our clinic, he was already on 5% natamycin and 0.5% moxifloxacin eye drops on an hourly basis for 7 days.

Investigations

The unaided visual acuity was 6/36 in both eyes; improving with correction to 6/6 in the left eye. Slit lamp evaluation of the right eye was significant for mild lid oedema and 1+ conjunctival congestion, and a 4×5 mm patchy stromal infiltrate in the temporal cornea with involvement up to the midstromal level. Stromal oedema adjacent to the infiltrate was noted (figure 1). The anterior chamber had 1+ cells and flare, the lens was clear. Slit lamp examination of the left eye was normal in all respects. The intraocular pressure was digitally normal in both eyes. Fundus evaluation of the right eye was deferred, but was normal in all respects in the left eye.

Figure 1.

Slit lamp examination, showing stromal infiltrate.

Corneal scrapings were obtained for both smears and cultures. Gram staining showed poorly staining filaments with sparsely septate hyphae (figure 2); while the 10% potassium hydroxide mount was negative for hyphae. The scrapings were plated on blood agar, Sabouraud dextrose agar, non-nutrient agar (NNA) and into brain heart infusion broth. Samples for PCR testing were also taken. Since the patient did not have his contact lenses, neither the lenses nor the solution could be used for microbiological work-up. Based on the smear reports, a diagnosis of fungal keratitis was made and therapy was initiated with topical 5% natamycin and 1% voriconazole hourly, 2% homatropine eye drops three times a day and oral ketoconazole 200 mg twice daily.

Figure 2.

Gram stain, corneal scraping smear, showing aseptate poorly staining hyphae; 100× oil immersion.

Flat greyish-white colonies with few aerial mycelia and possessing filiform margins grew on blood agar within 2 days, which was suggestive of Pythium insidiosum (figure 3). Pythium also grew on Sabouraud dextrose agar over the next few days as a slightly yellowish-cream flat colony. A lactophenol cotton blue mount of the culture showed sparsely septate non-sporulating filaments. However, the NNA plates were negative. Since the index of suspicion was strong for polymicrobial keratitis, confocal microscopy was done on the third day. Both hyphae and Acanthamoeba cysts were present (figure 4).

Figure 3.

Flat greyish-white Pythium colony on blood agar with filiform margins.

Figure 4.

Confocal biomicroscopy: yellow arrows show cysts, and blue arrowheads show hyphae; Heidelberg HRT-III.

The identification of the mycelial colony as P. insidiosum was confirmed via zoospore production on rice husk induction medium, and DNA sequencing of the ITS region of P. insidiosum.

Differential diagnosis

A tentative diagnosis of fungal versus polymicrobial keratitis was made. However, based on Gram stain, the culture characteristics, as well as the confocal evaluation, a clinical diagnosis of coexistent Pythium and Acanthamoeba keratitis was made

Treatment

The topical therapy was therefore supplemented with 0.04% polyhexamethylene biguanide (PHMB) hourly and fortified 0.2% linezolid eye drops hourly. Tablet linezolid 300 mg twice daily was also added.

Three days after initiating changes in the therapy, there was no change in the clinical picture, except for the presence of a minimal hypopyon. Intrastromal 1% preservative-free voriconazole was administered. Azithromycin 1% eye ointment was added four times a day. Acanthamoeba trophozoites were seen on the sixth day on the NNA plate overlaid with Escherichia coli.

Ten days after presentation, signs of healing in the periphery were noted; the epithelial defect had reduced in size and oedema had also reduced. A prominent surface plaque was present. Intrastromal voriconazole injections were repeated on days 12 and 24 with tapering of topical medications. The infiltrate continued to reduce in size with scarring at the edge of the lesion but the surface plaque continued to persist (figure 5). By the 25th day the uncorrected visual acuity had also improved to 6/18.

Figure 5.

Slit lamp examination, showing scarring with surface plaque.

On day 32, a new lesion adjacent to the prior one was noted (figure 6). Scrapings were repeated. Gram staining and 10% potassium hydroxide revealed few wide aseptate hyphae, suggestive of Pythium, while cultures were once again positive for Acanthamoeba. On the same day an intracameral injection of preservative-free 1% voriconazole injection was given and oral azithromycin 500 mg once daily was prescribed for 3 days, along with the prior topical medications.

Figure 6.

Slit lamp examination, showing new lesion adjacent to the original infiltrate.

Outcome and follow-up

Due to the obdurate nature of the lesion, a therapeutic keratoplasty was performed on the 38th day. Intrastromal 1% voriconazole was also administered at the graft–host junction. The excised corneal button was cultured but was negative for both Pythium and Acanthamoeba. However, the Gomori methenamine silver stain showed diffuse stromal infiltration with hyphae (figure 7).

Figure 7.

Gomori methenamine silver stain of histopathology section of keratoplasty button, showing fungal hyphae and elements, 40×.

Postoperatively, the graft was clear. The topical medications were continued on an hourly basis. Oral prednisolone 40 mg was started on postoperative day 1 and topical fluorometholone 0.1% twice daily, on postoperative day 6. A gradual taper of topical medications was instituted 20 days after surgery and by the 28th day the oral steroids were discontinued—the fluorometholone eye drops replaced with prednisolone acetate eye drops. At the last review 4 months postoperatively, the graft was clear with best-corrected visual acuity of 6/9, N8 (figure 8). Given the nature of the offending pathogens, the patient continues to be maintained on PHMB and azithromycin eye drops twice daily, along with prednisolone acetate eye drops.

Figure 8.

Slit lamp examination, showing clear graft 4 months postoperatively.

Discussion

P. insidiosum, an oomycete which exhibits mycelial growth on agar media, is very often misdiagnosed or treated as a fungal infection.¹⁰ It is more closely related to algae than fungi, and was first reported as a cause of systemic infection; cutaneous granulomas and arteritis being the most common. The infection is associated with significant mortality and morbidity.¹¹ It was subsequently identified as a cause of a fulminant keratitis in rice workers in Thailand.³ Increasing familiarity with the organism and its identification by characteristic appearances on smears, colony morphology, zoospore formation and nucleic acid sequencing¹⁰ have led to earlier diagnosis, but clear-cut and effective strategies for combating infection still elude us. Failure of medical therapy and recurrences after therapeutic keratoplasty necessitate evisceration in many patients.^{3 6} Contact with water seems to be the most common risk factor for contracting the infection,³ with a few cases being related to contact lens wear.^{5 6} Lekhanont in 2009 described the first case of contact lens-related Pythium keratitis, which in spite of multiple keratoplasties still required enucleation.⁵ Tanhehco also described a contact lens user who failed antifungal therapy and two keratoplasties, necessitating enucleation.⁶ However Badenoch⁴ and Lelievre⁸ reported the successful treatment of Pythium keratitis with a combination of antifungal therapy and therapeutic keratoplasty. Barequet et al reported long-term follow-up of 6 years after keratoplasty with retention of good vision in one patient, treated with topical and systemic voriconazole and therapeutic keratoplasty.⁷

The appropriate medical management of Pythium keratitis is still undefined. Since P. insidiosum lacks ergosterol in its cytoplasmic membranes, antifungal therapy usually fails.¹² Caspofungin, which inhibits glucan synthesis, a substance found in Pythium membranes, exhibits only limited fungistatic activity in vitro and in vivo.¹³

A review of the various therapies used in treating Pythium keratitis is not encouraging. Lekhamont described failed therapy to natamycin, amphotericin B and ketoconazole eye drops, as well as oral itraconazole and terbinafine.⁵ Lelievre described successful therapy of contact lens-associated Pythium keratitis with topical and intravenous caspofungin, as well as systemic, topical and intrastromal voriconazole, followed by a large therapeutic keratoplasty⁸; while Barequet reported long-term retention of useful vision with a combination of topical amphotericin B and voriconazole, systemic voriconazole and keratoplasty.⁷ To the best of our knowledge, all reported cases of Pythium keratitis required keratoplasty for successful eradication of the infection, the only exception to this is the single case of presumptive P. insidiosum keratitis successfully treated with linezolid and azithromycin eye drops by Ramappa and coworkers.¹⁴

Our rationale for choice of drugs was based on information available in the literature. A single case report by Barequet et al described the successful eradication of the infection with the use of topical and intravenous voriconazole, and therapeutic keratoplasty.⁷ One report which evaluated the efficacy of seven antifungals against eight strains of P. insidiosum obtained from vascular and ocular pythiosis noted that the highest Minimum Inhibitory Concentration (MIC) was detected for amphotericin B, while voriconazole, fluconazole, anidulafungin, caspofungin, itraconazole and terbinafine were found to have the lowest MICs.¹⁵ Behrens-Bohmann et al have described excellent efficacy of PHMB in vitro against hyaline fungi such as Fusarium and Rhizopus,¹⁶ and PHMB has well-known efficacy against Acanthamoeba. Linezolid and azithromycin eye drops were also used based on the successful medical therapy of a single case of presumed Pythium keratitis.¹⁴ Tigecycline, minocycline, clarithromycin and azithromycin were found to be effective in subcutaneous infections in the rabbit model. Additionally, azithromycin, with or without minocycline, was noted to significantly reduce the P. insidiosum load.¹⁷

Apart from the lack of clear-cut strategies for medical therapy, delays in diagnosis are common since smear evaluations may be negative or the organism fails to grow in culture. Given its mycelial-like growth on culture media and the lack of sporulation, it is very often classified as an unidentified hyaline fungus. Thus, the window period for a successful keratoplasty which could surgically debulk the infection is very often lost. Even with keratoplasty, recurrences have been common; early diagnosis is therefore the key to successful outcomes after keratoplasty.

Acanthamoeba keratitis is also recalcitrant to therapy, and a number of authors consider that keratoplasty is best delayed until the organism is eradicated.^18–20 However, our patient fared well with keratoplasty.

Learning points.

• Pythium insidiosum can coexist with Acanthamoeba keratitis.

• Delays in diagnosis are common since smear evaluations may be negative, the organism fails to grow in culture or may be erroneously treated as fungal keratitis.

• A high index of suspicion is required to identify this particular combination of pathogens.

• Appropriate medical management of Pythium keratitis still lacks clarity.

• Early diagnosis is therefore the key to successful outcomes after keratoplasty.