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Case Report

Fungal Keratitis Caused by Podospora austroamericana: A First

Case Report
Gunasekaran Rameshkumar, Mariappan Ponlakshmi, Appavu Selvapandiyan, Ravindran Ramsudharsan1, Raja Subramanian Krishnan2, Prajna Lalitha
Departments of Ocular Microbiology, 1Intraocular Lens and Cataract Services and 2Cornea and Refractive Surgery Services, Aravind Eye Hospital and Post Graduate
Institute of Ophthalmology, Madurai, Tamil Nadu, India

Abstract
We report a case of keratitis caused by a rare fungus Podospora austroamericana. Clinical and microbiological evaluation of the corneal ulcer
was done and the treatment outcome was studied. The fungus was grown from the corneal scraping, and it was identified as P. austroamericana
based on DNA sequence and analysis of the internal transcribed spacer region. The patient was treated with topical azithromycin, natamycin and
voriconazole. Despite maximum medical therapy, the ulcer progressed very rapidly and the patient developed panophthalmitis and evisceration
of the eye had to be done. This is the first reported case of keratitis caused by P. austroamericana.

Keywords: DNA sequencing, keratitis, panophthalmitis, polymerase chain reaction

Introduction stromal oedema [Figure 1a]. As per our clinical protocol, the

Fungal keratitis is an important cause of visual loss globally.
The fungi implicated appear to vary depending on the
geographical locations. Fusarium spp. and Aspergillus spp. are
the most prevalent fungus causing mycotic keratitis in South
India.[1,2] However, various rare fungi causing mycotic keratitis
have been reported from different parts of India.[3] Podospora
austroamericana is a plant pathogen. Podospora is a ubiquitous
genus in the family Lasiosphaeriaceae (Ascomycota and fungi)
and now includes approximately 78 species.[4] The Podospora
species is reported mainly from soil[5] and cow dung.[6] We
report a case of keratitis caused by P. austroamericana and
discuss the presentation, identification of the fungus and the
management.
Case Report
A 65‑year‑old male presented with redness and pain in his
right eye 4  days after undergoing cataract surgery. He did
not recall any history of ocular trauma or fall of foreign
body. He did not have any other systemic complaints. On
examination, his visual acuity in the right eye was only
perception of light. The conjunctiva was congested and lid
edema was seen. The cornea had a circumcorneal congestion,
multiple stromal infiltrates along the limbus with diffuse
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ulcer was scraped using a Kimura spatula and subjected to
microbiological evaluation which included smear and culture
methods. The smears employed were Gram staining and 10%
potassium hydroxide (KOH) mount, while the culture methods
included blood agar and potato dextrose agar. The Gram stain
and 10% KOH mount showed the presence of filamentous
fungi  [Figure  2a]. Colonies were pale white in colour and
velvety in nature in front  [Figure  2b]. Spore morphology
was identified using lactophenol cotton blue mounting, but
the fungus could not identified using conventional techniques
due to lack of sporulation even after 2–3 weeks of incubation.
After 4 weeks of incubation, the ascospore morphology was
identified and presented in Figure 2c‑e. DNA extraction from
the fungus was carried out using the phenol–chloroform
method. Polymerase chain reaction  (PCR) targeting the
internal transcribed spacer (ITS) region 1 and 4 was carried
out in 50 μl reactions, containing 1 μl of 200 mM dNTPs,
Address for correspondence: Dr. Prajna Lalitha,
Department of Ocular Microbiology, Aravind Eye Hospital and Post Graduate
Institute of Ophthalmology, No. 1, Anna Nagar,
Madurai ‑ 625 020, Tamil Nadu, India.
E‑mail: lalitha@aravind.org
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How to cite this article: Rameshkumar G, Ponlakshmi M,

DOI: Selvapandiyan A, Ramsudharsan R, Krishnan RS, Lalitha P. Fungal keratitis
10.4103/ijmm.IJMM_19_1 caused by Podospora austroamericana: A first case report. Indian J Med
Microbiol 2018;36:594-6.

594 © 2019 Indian Journal of Medical Microbiology | Published by Wolters Kluwer - Medknow
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Rameshkumar, et al.: Keratitis caused by P. austroamericana

a
a b

b
Figure 1: Clinical findings of patients with Podospora austroamericana
keratitis. (a) Slit‑lamp picture of the cornea showing multifocal anterior
stromal infiltrate. (b) Total limbus‑to‑limbus full‑thickness infiltrate
c d e
Figure 2: (a) Gram‑stain image showing septate fungal hyphae seen
under ×100. (b) Pale white colour fungus grown in potato dextrose
agar plate after 3 days of incubation at 27°C. (c) Ascoma of Podospora
austroamericana. (d) Asci with ascospores. (e) Ascospores

5 μl of reaction buffer  (Tris with Mgcl2), 10  pM forward

primer  (ITS 1:  5’ TCCGTAGGTGAACCTGCGG 3’) and Table 1: Antifungal susceptibilities of the clinical isolate
reverse primer (ITS4:5’ TCCTCCGCTTATTGATATG 3’),[7] of Podospora austroamericana
1.2 U/μl of Taq polymerase and 5 μl of genomic DNA, and Antifungal drugs Range (µg/ml) MIC (µg/ml)
amplification was carried out in Agilent SureCycler 8800 Amphotericin B 16‑0.03 2
(Agilent Technologies, USA). DNA sequencing was done Voriconazole 16‑0.03 0.5
using an ABI 3100 genetic analyzer (PE Applied Biosystems, Ketoconazole 16‑0.03 1
Foster City, CA, USA). The sequence was analysed and Posaconazole 16‑0.03 0.5
identified using MegaBlast search programme of GenBank Natamycin 32‑0.06 4
database. The query coverage and identity were 100%. The Caspofungin 8‑0.01 0.25
MIC – Minimum inhibitory concentration
sequence was submitted in NCBI  (GenBank Accession
number: MH973201). In vitro antifungal susceptibility
testing was performed by broth microdilution method species re‑introduced the genus Schizothecium corda and
(CLSI guidelines M38A2) against following drugs: transferring 31 species of Podospora to it. He re‑established
amphotericin B, voriconazole, ketoconazole, posaconazole, the relationship between these two species based on the spore
natamycin and caspofungin.[8] The organism was susceptible morphology.[9] Podospora species are mostly coprophilic in
to all the antifungal drugs  [Table  1]. The patient was habit and also reported as plant pathogen.[5,6] In our case report,
started with topical azithromycin  (1%), natamycin  (5%) the patient did not recall any history of ocular trauma. However,
and voriconazole (1%) hourly with atropine eye drops and the possibility of a trivial ocular trauma cannot be ruled out.
The case described in this report deserves attention because
painkillers. He was reviewed daily. The size of stromal
of the unique characteristics of the organism, difficulties
infiltrate increased despite active treatment. Hence,
in identification and non‑responsive to treatment. There is
itraconozole eye drops and oral ketoconazole 200  mg bd
no clinical information in the medical literature concerning
were also added. Azithromycin (1%) was added as the clinical
Podospora species because it has not been identified in clinical
picture was thought to resemble Pythium keratitis. However,
samples in the previous reports and is unreported in ocular
the eye became phthisical within 4 weeks and nothing further
infections.
could be done to restore the vision [Figure 1b].
Declaration of patient consent
Discussion The authors certify that they have obtained all appropriate
patient consent forms. In the form the patient(s) has/have
Fungal keratitis is a major public health problem in India and
given his/her/their consent for his/her/their images and other
other developing countries. Fusarium and Aspergillus species
clinical information to be reported in the journal. The patients
are the commonly reported major pathogens in most of the
understand that their names and initials will not be published
geographical areas.[1‑3] The Podospora species is a large and
and due efforts will be made to conceal their identity, but
polymorphic genus. There is notable variation in the structure,
anonymity cannot be guaranteed.
especially ascomatal wall and ascospore appendages. Mirza
and Cain revised the genus of these fungi and listed 64 species[5] Financial support and sponsorship
and Lundqvist (1972) who reviewed the Nordic coprophilous Nil.

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Rameshkumar, et al.: Keratitis caused by P. austroamericana

Conflicts of interest
There are no conflicts of interest.
References
1. Lalitha  P, Prajna  NV, Manoharan  G, Srinivasan  M, Mascarenhas  J,
Das  M, et al. Trends in bacterial and fungal keratitis in South India,
2002‑2012. Br J Ophthalmol 2015;99:192‑4.
2. Sengupta  S, Rajan  S, Reddy  PR, Thiruvengadakrishnan  K,
Ravindran RD, Lalitha P, et al. Comparative study on the incidence and
outcomes of pigmented versus non pigmented keratomycosis. Indian J
Ophthalmol 2011;59:291‑6.
3. Saha  S, Banerjee  D, Khetan  A, Sengupta  J. Epidemiological profile
of fungal keratitis in urban population of West Bengal, India. Oman J
Ophthalmol 2009;2:114‑8.
4. Cai  L, Jeewon  R, Hyde  KD. Phylogenetic evaluation and taxonomic
revision of Schizothecium based on ribosomal DNA and protein coding
genes. Fungal Divers 2005;19:1‑17.
5. Stchigel AM, Calduch M, Guarro J, Zaror L. A new species of Podospora
from soil in Chile. Mycologia 2002;94:554‑8.
6. Mirza  JH, Cain  RF. Revision of the genus Podospora. Can J Bot
1969;47:1999‑2048.
7. Jaeger  EE, Carroll  NM, Choudhury  S, Dunlop  AA, Towler  HM,
Matheson  MM, et al. Rapid detection and identification of Candida,
Aspergillus and Fusarium species in ocular samples using nested PCR.
J Clin Microbiol 2000;38:2902‑8.
8. Clinical and Laboratory Standards Institute. Reference Method for
Broth Dilution Antifungal Susceptibility Testing of Filamentous Fungi;
Approved Standard. CLSI Document M38‑A2. 2nd  ed. Wayne, PA:
Clinical and Laboratory Standards Institute; 2008.
9. Lundqvist N. Nordic Sordariaceae s. lat. Symb Bot Ups 1972;20:1‑374.

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