FUNGAL=20 KERATITIS - AN=20 OVERVIEW

Dr.Kalyan=20 Das MS, DNB

Dr.Jnanankar.Medhi MS=20

Consultants,=20 Sri Sankaradeva Nethralaya

Beltola, = Guwahati-28,=20 Assam

=20 Keratomycosis is common in tropical countries like India = specially in=20 dry, humid condition. The Madurai Study, 1993 reported an annual = incidence of=20 corneal ulceration in 113 per 100,000 populations.

=20 The protocol for=20 management of infective keratitis

in the absence of any history or = signs=20 suggestive of a nonbacterial etiology is treatment of the ulcer with a = standard=20 empirical regimen of combined broad-spectrum antibiotics. However, = routine=20 scrapping of suspected bacterial cases for smears, culture, and = sensitivity is=20 advocated. But in our country, where the prevalence of bacterial = and fungal=20 corneal ulceration is almost equal (bacterial keratitis =96 47%, = fungal=20 keratitis - 46.8 %, mixed = infection =96=20 5%, parasitic infection =96 1%) optimal management calls for rapid = clinical and=20 microbiological recognition and timely institution of specific therapy.=20

Fungal=20 keratitis is one of the most difficult forms of microbial keratitis to = diagnose=20 and treat successfully and remains a diagnostic and therapeutic = challenge to the=20 attending ophthalmologist. Difficulties arise in making the correct = diagnosis in=20 time, non-availability of a proper laboratory back up and lack of the = ideal=20 topical antifungal preparations. The presentation is often very late in = many=20 cases leading to limited medical or surgical success.

Diagnosis:

&n= bsp; =20 The proper approach to management of fungal keratitis calls for = timely=20 diagnosis and treatment. Clinical features, which are typical of fungal = corneal=20 ulcer, help in making a provisional diagnosis. The typical clinical features = of a=20 fungal corneal ulcer are

Greyish/Dirty white appearance = (sometimes=20 brown pigmentation)=20
Dry, rough texture surface = (gritty=20 feeling)=20
Elevated, feathery = edges=20
Endothelial = plaque=20
Hyphate / branching=20 ulcers=20
Satellite = lesions=20
Ring infiltrate (Wesseley ring) = ring=20 abscess

=95

However a laboratory confirmation = is=20 essential. Modalities of laboratory diagnosis are direct smear = examination,=20 fungal culture and species identification. Corneal biopsy and blood = culture are=20 done in very rare situations where repeated smear, culture does not = reveal any=20 organism and also does not show any clinical response. Confocal = microscopy and=20 electron microscopy add to the armamentarium of diagnostic modalities. = These=20 helps in direct in- vivo visualization of the fungal elements in the = corneal=20 stroma.

Specimen collection and=20 Processing:

Corneal scrapping is done for both = diagnostic=20 and therapeutic purposes. = It is to=20 be done in a slit lamp after instillation of topical anesthetic drop (4% = lignocaine). Purulent material over the ulcer is removed with a = sterile=20 cotton swab and discarded. Kimura=92s spatula or surgical blade no 15 = are used for=20 scrapping. The material is preferably taken from the margin of the ulcer = so as=20 to include the feathery edges. From the sample, 4-5 direct smears are = made on=20 clean glass slides. It is also inoculated directly into different = culture media=20 in multiple =91C=92 streaks. Besides scrapping, specimen for culture can = also be=20 collected by paracentesis and corneal biopsy.

KOH stain: This is = the most basic=20 test for identifying fungal elements in a direct smear taken from a = corneal=20 scrapping. One drop of = KOH (10-20=20 %) is put on an unfixed smear. =20 After incubating for 5 min at 37 deg C it is observed under 10x = and 40x=20 for fungal filaments. Branching fungal filaments and buds of yeast are = stained=20 blue purple.

Grams Stain: This can = help in=20 identifying bacterial morphology and arrangement. Fungal hyphae are = usually=20 stained gram positive but may show variable results also. Epithelial = cells and=20 polymorphs stain pink

Giemsa Stain: This is = useful for=20 studying host cells in a smear. Also used for fungal hyphae, = acanthaemoeba cysts=20 and inclusions. Fungal hyphae are usually stained blue purple.

GMS: Helpful as it stains both live = and dead=20 fungi. Stains the higher = bacteria=20 like Nocardia, Actinomyces, etc.

Special=20 Stains:

Special staining agents may be used in different = cases. The=20 two types used are Fluorochromes and Immunofluorescence. The = fluorochromes=20 commonly used are,

1. Calcofluor = white =96=20 Used for identification of fungi and acanthaemoeba cysts. Excitation is achieved with = violet light=20 (355-425 l).=20

Acridine=20 Orange - Stains RNA = (orange red)=20 and DNA (yellow green). =20

Culture:

Following culture = media are=20 routinely used for inoculating corneal scrapping materials,

=A7 =20 Blood Agar =20 - 37 deg C (aerobic)

=A7 =20 Chocolate Agar =20 - 37 deg C (5-10 % CO₂)

=A7 =20 Saboroud Dextrosae Agar =96 25 deg C

=A7 =20 BHIB =96 37 deg C (aerobic)- broth

=A7 =20 Thioglycollate =20 - 37 deg C (anaerobic) =96broth

In case of liquid = media (broth)=20 growth of an organism is characterized by turbidity of the media, = appearance of=20 ball of fungal mycelia floating on the fluid surface, fungal hyphae = growing=20 along the wall of the container etc.

Culture: Laboratory = confirmed=20 growth is defined as

=A7 =20 At least=20 semi confluent growth on solid media.

=A7 =20 Any growth on two or more media.

=A7 =20 Growth on one media and is supported by direct = smear=20 findings.

Cultures are = incubated for a=20 period of 14 days before discarding as negative.

Methods for fungal identification and = detention:

Conventional
methods: Organisms are = identified by studying the following,=20
- Growth=20 characteristics at 25 deg C and 37 deg C=20
- Colony=20 morphology=20
- Microscopic=20 morphology =96 wet mount=20
- Slide=20 culture=20
- Biochemical=20 reactions =96 e.g. Urease production

Automated=20 system for biochemical studies=20
Molecular=20 methods =96 usually reserved for slow growers.

Molecular = methods:

PCR: It=20 is a method of detecting very small quantities of DNA or RNA segments in = a=20 sample. Any DNA or RNA that matches the probe DNA or RNA sequence is = amplified=20 in vitro up to 1 million times. PCR appears to be equally or more = sensitive than=20 culture studies.

Nucleic Acid probe testing: = Species-specific rRNA probes tagged with a = chemiluminiscent agent and are measured in a luminometer.=20

Treatment:

Treatment of fungal = keratitis is=20 usually medical, but about 20-30% of cases requires some form of = surgical=20 intervention.

Although many antifungal agents = have been=20 isolated or synthesized, none of the presently available agents has the = features=20 of an ideal drug.

The limitations of anti-fungal = agents include=20 =96

=B7 =20 Poor=20 solubility,

=B7 =20 Low=20 potency,

=B7 =20 Variability=20 between in vitro sensitivity pattern and in vivo = effect,

=B7 =20 Development of=20 resistant organism,

=B7 =20 Development of=20 toxicity at therapeutic concentration.

The medical management=20 includes

=B7 =20 Topical=20 antifungal agents

=B7 =20 Peri-ocular=20 injection-rarely used

=B7 =20 Systemic =96=20 oral

Mechanism of=20 action

Fungal = cell wall is=20 the site of antifungal agents. The ergosterol in the cell membrane is = unique to=20 fungi. The antifungal agents has propensity to bind to ergosterol, get = inserted=20 into the membrane and several molecules together orient themselves in = such a way=20 so as to form a =91micropore.=92This increases cell permeability through = which ions,=20 amino acids and other water soluble substances move out. The cell = becomes=20 dehydrated and dies out.

Antifungal Agents

= ; =20 I. = Polyenes

A. =20 Large = Polyenes

=A7 =20 Nystatin

=A7 =20 Amphotericin B

=A7 =20 Amphotericin B methyl ester

B. =20 Small polyenes

=A7 =20 Natamycin

= II. = ;=20 Azoles=20

A. =20 Imidazoles

=A7 =20 Miconazole

=A7 =20 Ketoconazole

=A7 =20 Clotrimazole

=A7 =20 Econazole

=A7 =20 Thiabendazole

B. =20 Triazoles

=A7 =20 Fluconazole

=A7 =20 Itraconazole

=A7 =20 Saperconazole

= I.

III. = ; =20 Pyrimidine

=A7 =20 Flucytosine

The polyenes are most = commonly=20 used antifungal agents in clinical practice.

Natamycin ( 5%=20 suspension.): It is derived from = Streptomyces=20 natalensis. It is fungicidal in nature with a broad spectrum of = action,=20 acting against both yeast and filamentous fungi. The antifungal action = is dose=20 related and acts as a fungistatic in low doses. The average duration of = therapy is 2 to=20 3 weeks.

Amphotericin B eye = drop=20 (0.15%): Amphotericin B (AMB) is produced by a strain of = Streptomyces nodosus.=20 It can be fungistatic or be fungicidal depending on the dose. AMB is the = drug of=20 choice for yeasts. It is used in filamentary fungus also. The known=20 complications of AMB include renal toxicity, bronchospasm, hypotension = etc. &n= bsp; =20 The AMB eyedrops are not available on a commercial basis. It is = prepared=20 from injection AMB (Fungizone 50 mg dry powder). The powder is divided into 6 = equal parts=20 of 8 mg each. Each part is then reconstituted with 5ml of distilled = water and=20 placed in amber colored bottle/vial. It is to be kept at room = temperature away=20 from direct sunlight. Once prepared, the drops are to be used within 48 = hours.=20

Ketoconazole: =

=20 It is a fungistatic agent available as 200mg tablets. The = dose is=20 200 =96 400 mg daily in divided doses. Duration depends on the response = to=20 therapy. The indications for systemic therapy are - large indolent = ulcers=20 involving the limbus, perforated ulcers with uveal tissue prolapse into = the=20 wound, inadequate therapeutic response with topical therapy alone in a = confirmed=20 case of fungal corneal ulcer etc. C

omplications include = hepatotoxicity,=20 impotence etc.

Fluconazole:=20

It is an alternative = to=20 Ketoconazole and acts as a fungistatic agent. It is available in both = systemic=20 as well as topical form. = The=20 oral dose is 100 to 200mg twice daily. Systemic complications associated = with=20 Fluconazole are found to be lesser than Ketoconazole. It increases the = effects=20 of anticoagulants and so it should be prescribed with=20 caution.

Therapy of Fungal = Keratitis based on=20 culture reports

Organism

Topical

(Alternatives)

Sub = conjunctival

(Alternatives)

Oral

(Alternatives)

Filamentous fungi

Natamycin

(Amphotericin B, Miconazole,=20 Fluconazole)

Miconazole

(Fluconazole)

Fluconazole

(Ketoconazole)

Yeast

Amphotericin = B

(Natamycin, Miconazole,=20 Fluconazole)

Miconazole

(Fluconazole)

Flucytosine

(Fluconazole,=20 Ketoconazole)

After starting = antifungal=20 therapy

, regular monitoring = of the=20 patients is crucial. The compliance level, and ability of the patient to = come=20 for follow-ups is also important for the successful treatment. Any doubt = in this=20 aspect usually calls for admission of the patient for receiving = appropriate=20 care.

Another important = aspect of=20 follow-ups is documentation of the clinical picture of the corneal = ulcer. Digital documentation is best, = however=20 in a developing country like ours manual drawing usually suffices. It is = important to use the proper color code while doing the surface mapping.=20 Dimensions of the ulcer area with depth measurement are noted in each = visit.=20

The following = features are to be=20 noted,

=B7 =20 Progress of the corneal ulcer

=B7 =20 The area and density of infiltration

=B7 =20 Size and depth of ulceration

=B7 =20 Size of epithelial defect

=B7 =20 Amount of stromal edema

=B7 =20 Extent of scleral involvement

=B7 =20 Anterior chamber reaction

The response to treatment is usually slow. = Clinical signs=20 denoting improvements are,

Decrease=20 in infiltrate fluffiness=20
Blunting=20 of the edges=20
Disappearance=20 of satellite lesions=20
Decreased=20 discharge=20
Increase=20 comfort=20
Decreased=20 AC reaction

Late signs=20 of improvement include,

=B7 =20 Decrease in=20 density or size of infiltration

=B7 =20 Re-epithelization

=B7 =20 Vascularisation

Surgical = Treatment:

About = 1/4^th to=20 1/3^rd of fungal corneal ulcers may require therapeutic = Penetrating=20 Keratoplasty (PK). Therapeutic PK is usually reserved for cases where = the ulcer=20 progresses despite aggressive antimicrobial therapy, in cases with = descemetocele=20 or perforation or in patients with severe pain. The goal of the surgery = is to=20 eliminate the infection and restore the integrity of the globe. The size of the graft is = decided on the=20 basis of the size of the ulcer and should include the infected edges. = Topical=20 antifungal drugs are to be continued after the procedure and steroid = drops are=20 usually added after 3-4 days.

Other procedures that = requires to=20 be undertaken in non-healing ulcers,

Glue (cyanoacrylate) and BCL=20 application

=B7 =20 Central or lateral Tarsorrhaphy

=B7 =20 Conjunctival hooding

The management of = fungal=20 keratitis is usually long drawn and involves on an average of 2-3 = months.=20 Patient compliance is a crucial factor. The treatment continues even = after the=20 ulcer has healed in the form of visual rehabilitation.

( e mail: drkdssn@sify.com)<= /P>

&nbs= p; = ; = &= nbsp; =20 XXX &nbs= p; = ; = &= nbsp; =20