Medical Mycology Case Reports 2 (2013) 88–90

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Medical Mycology Case Reports

2211-75http://d

n CorrE-m

journal homepage: www.elsevier.com/locate/mmcr

An environmental Sporothrix as a cause of corneal ulcer

Annie S. Morrison a, Shawn R. Lockhart b, Jennifer G. Bromley c, Joung Y. Kimc, Eileen M. Burd a,n

a Department of Pathology and Laboratory Medicine, Emory University School of Medicine, 1364 Clifton RD NE, Atlanta, GA 30322, USAb Mycotic Diseases Branch, Centers for Disease Control and Prevention, 1600 Clifton RD, Atlanta, GA 30322, USAc Department of Ophthalmology, Emory University School of Medicine, 1365 Clifton RD NE, Atlanta, GA 30322, USA

a r t i c l e i n f o

Article history:Received 22 March 2013Received in revised form29 March 2013Accepted 30 March 2013

Keywords:SporothrixFungal keratitisAntifungal susceptibility

39/$ - see front matter & 2013 International Sx.doi.org/10.1016/j.mmcr.2013.03.002

esponding author. Tel.: +1 404 712 7297.ail address: eburd@emory.edu (E.M. Burd).

a b s t r a c t

In this case we introduce Sporothrix pallida, a non-pathogenic environmental Sporothrix species as acause of infectious keratitis in a corneal transplant recipient. Human infections caused by S. schenckii arewell-known but human infection with Sporothrix pallida has not been previously reported.

& 2013 International Society for Human and Animal Mycology. Published by Elsevier B.V All rightsreserved.

1. Introduction

Fungi are opportunistic agents of infections of the cornea.Although rare, especially in temperate climates, fungal infectiontypically occurs in corneas that are compromised by underlyingdisease or trauma. Fungal keratitis is often a diagnostic andtherapeutic challenge, especially in geographic areas where theseinfections are not commonly seen. The associated stromal inflam-mation is not unique to fungal keratitis and management isrestricted to the availability of topical antifungal agents andeffectiveness depends on susceptibility of the causative agentand the extent to which the antifungal penetrates into cornealtissue. The predominant etiologic agents of fungal keratitis such asFusarium, Aspergillus, Curvularia, Alternaria, Candida etc. are alsoknown to cause localized or systemic infections in other parts ofthe body. Some fungi that are considered nonpathogenic tohumans have been isolated as rare causes of infectious keratitis,usually following corneal injury or the long term use of topicalcorticosteroids [1,2]. Non-pathogenic molds such as Phoma,Dichotomophthoropsis, Rhizoctonia, Cephaliophora, Lasiodiplodia,Colletotrichum, Cladorrhinum, Metarhizium and Phaeoisaria havebeen reported [1,2]. We describe a case of fungal keratitiscaused by Sporothrix pallida, a non-pathogenic environmentalmold closely related to the known pathogen Sporothrix schenckii.S. pallida has not been previously described as causing patho-logic infection in humans. Reporting of cases like this that arecaused by unusual organisms not only adds to the cumulativeliterature but provides some measure of the relative frequency

ociety for Human and Animal Myc

of organisms likely to be encountered. This case also providesthe results of susceptibility testing and their use in the manage-ment of antifungal therapy.

2. Case

The patient is a 75 year old woman who presented to theCornea service in January 2012 complaining of two weeks ofdecreased vision in her right eye. She has a history of bilateralFuchs corneal dystrophy and underwent a combined corneatransplant and cataract extraction in the right eye in 2006.She has worn a rigid gas permeable contact lens in the right eyeto improve visual acuity following the cornea transplant. Shehad an episode of graft rejection in 2008 and has been main-tained on prednisilone acetate eye drops once per day since.Medications also included latanaprost and brimonidine eyedrops for treatment of glaucoma. She denied any history ofrecent trauma. At presentation, her uncorrected visual acuitywas 20/70 in the right eye. Examination of her cornea revealedcentral irregular anterior stromal infiltrates in a branchingpattern with an overlying epithelial defect. The patient's corneawas scraped and specimens were submitted for Gram stain andbacterial, mycobacterial and fungal culture. Gram stain of thecorneal scrapings showed numerous branching septate hyphalelements consistent with fungus. The results of the culture asdetailed below identified Sporothrix pallida as the causativeorganism. The patient was started on topical voriconazole everyhour and the prednisilone acetate was discontinued. Followingsix weeks of treatment with voriconazole, she had a small,residual central scar. Her best corrected visual acuity returned

ology. Published by Elsevier B.V All rights reserved.

A.S. Morrison et al. / Medical Mycology Case Reports 2 (2013) 88–90 89

to 20/30. She resumed her prednisilone acetate and contactlens wear.

Three months later she developed new symptoms of pain,redness and decreased vision. A new infiltrate was noted adjacentto the previous scar. Due to her previous history, repeat cultureswere taken and she was restarted on topical voriconazole.S. pallida was again isolated from her right cornea, and suscept-ibility testing was done, which showed an elevated minimuminhibitory concentration (MIC) to voriconazole. The patient had anincomplete response to treatment and developed a persistentepithelial defect with a central infiltrate. Her best corrected visualacuity was 20/40. She underwent a repeat penetrating kerato-plasty to remove the infected graft.

Laboratory diagnosis was made by inoculating the cornealscrapings onto sheep blood agar, chocolate agar, and Sabourauddextrose agar (SAB) with chloramphenicol and gentamicin (Remel,Lenexa, KS) and submitted to the Microbiology Laboratory forroutine bacterial and fungal culture. After four days of incubationat 30 1C, several flat, glabrous, cream-colored colonies that devel-oped a wrinkled and folded surface with maturation were seen onthe fungal culture. A wet mount demonstrated oval yeast forms. Agerm tube test was negative, an API 20C (bioMerieux, Durham, NC)yeast identification panel resulted in no identification, and a ureaslant was positive for urease production. Culture at 25 1C on cornmeal agar yielded tan colonies consisting of oval yeast, longpseudohyphae, and septate hyphae with terminal obovoid conidiawith short denticles growing sympodially on a conidiogenous cell.The culture did not darken with age.

DNA was prepared from the isolate as previously described [3].Sequence analysis of the internal transcribed spacer (ITS) and largesubunit (LSU) sequence identified 100% matches to isolates ofSporothrix pallida and S. stylites. Additional sequence analysis ofthe β-tubulin gene [4] clearly distinguished the isolate as S. pallidawith 100% sequence identity across ITS, LSU and β-tubulin (1448base pairs total) to S. pallida isolate CBS131.56.

Antifungal susceptibility testing on the second isolate was doneusing the methodology of the Clinical and Laboratory StandardsInstitute, document M38-A2 [5]. Minimum inhibitory concentra-tion (MIC) and minimum effective concentration (MEC) valueswere determined following 48 h of growth at 25 1C. MICs for theazoles were notably elevated and ranged from 4 μg/ml to4256 μg/ml (Table 1). MICs for quality control strains were withinreference range. Slow growth prevented determination of yeastphase susceptibilities with no growth in the drug-free wellfollowing 72 h of incubation at 35 1C.

3. Discussion

The pathogenic dimorphic fungus Sporothrix schenckii hasbeen recovered from many clinical specimens worldwide and iswell-known as the causative agent of sporotrichosis. Infection

Table 1In vitro antifungal susceptibilities of the Sporothrixpallida isolate as determined by the standardizedCLSI M38-A2 methods.

Antifungal agent MIC (mg/ml)

Fluconazole 4256 mg/mlVoriconazole 8 mg/mlItraconazole 416 mg/mlPosaconazole 4 mg/mlFlucytosine 4256 mg/mlCaspofungin 4 mg/ml (MEC)Micafungin 2 mg/ml (MEC)Anidulafungin 1 mg/ml (MEC)

usually involves the skin and subcutaneous tissues but canoccasionally disseminate or occur in other sites, primarily inimmunocompromised patients [6]. Although only one clinicallyrelevant species of Sporothrix was classically identified, recentmolecular and phenotypic data demonstrate that what had beenknown as Sporothrix schenckii is actually a complex of speciesof different genetic lineages that tend to cluster in distinctivegeographic regions [4,7]. Accordingly, Sporothrix schenckii is nownamed the S. schenckii complex and is comprised of a number ofnewly recognized Sporothrix species [7,8].

S. schenckii complex organisms live as saprophytes on wood,decaying vegetation, animal excreta and soil, and some species areconsidered to be associated with human infection. Other S.schenckii complex species that are also found in the environmentare clearly distinct from human isolates and are non-pathogenic inanimal studies [9]. Phylogenetic studies based on 28S rDNA andinternal transcribed spacer ITS sequences have been used to makecomparisons at the genus level and separate the group of clinicalstrains from the environmental strains but are not informativeenough to distinguish relationships within each group [10]. Moreinformation has been gained by sequencing the beta-tubulin andcalmodulin genes. This information, along with morphological andcultural differences have lead to the recognition of distinct taxawithin the environmental strains now described as S. pallida, S.stylites, S. humicola and S. lignivora [4]. The previously describedspecies S. albicans and S. nivea have been found to be synonymouswith S. pallida, which was retained as the accepted name since itwas the first of the three to be described [4]. There are no reportedcases of human infection by S. pallida, but that does not eliminatethe possibility of infections that may have been erroneouslyascribed to S. schenckii.

The source of the infection in the present case is unclear.The patient had recently returned from a vacation in Texas, butdenied any recent trauma. She was on long term topical corticos-teroids due to her previous history of graft rejection and wasat least focally immunocompromised. Despite an initial goodresponse to topical therapy, she ultimately required a repeatkeratoplasty to eradicate the infection.

In vitro susceptibility profiles have been reported for S. schenckiicomplex isolates but not for the non-pathogenic Sporothrix species.Although a standardized method for antifungal susceptibility testingis established, no interpretive breakpoints exist for Sporothrix sp. andcorrelation with clinical outcome remains to be determined [5].Reported studies have shown that Sporothrix susceptibility patternsare somewhat variable and depend on the species tested [11,12].S. brasiliensis had generally low MICs to all of the antifungal agentstested and S. mexicana had high MICs, except against terbinafine.Terbinafine had low geometric meanMICs for all of the species testedand was found to be the most active antifungal agent. Ketoconazolegenerally also had very low geometric mean MICs except againstS. mexicana and S. albicans. Posaconazole was the most active of thesystemic antifungal agents. Fluconazole and micafungin were notactive against any of the species. Although itraconazole has beeneffectively used clinically, in vitro activity was poor for itraconazoleand ravuconazole, except against S. brasiliensis. MICs against eberco-nazole, ravuconazole and amphotericin B were moderately high andamphotericin B has been used clinically with variable results.MIC ranges were wider for voriconazole and flucytosine and highgeometric mean MICs indicate that these agents would generally notbe effective. Itraconazole is currently considered to be the treatmentof choice for cutaneous infections, while amphotericin B is thetreatment of choice for systemic or disseminated infections [13].

For the S. pallida isolate obtained in this case, anidulafunginand micafungin appeared to have the most favorable in vitroactivity. Caspofungin, voriconazole and posaconazole had elevatedMICs above 2 μg/ml. Very high MICs were obtained for fluconazole,

A.S. Morrison et al. / Medical Mycology Case Reports 2 (2013) 88–9090

itraconazole, and flucytosine which were suggestive of a low levelof susceptibility to these drugs.

Conflict of interest statement

There are none.

Acknowledgments

The authors would like to thank Naureen Iqbal for generatingthe MIC values. The findings and conclusions of this article arethose of the authors and do not necessarily represent the views ofthe Centers for Disease Control and Prevention.

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