Original article / research
Year : 2016 Month : April Volume : 5 Issue : 2 Page : 30 - 33 Full Version

Mycotic Keratitis in Solapur (A Two Years Study)
Karan Indra Ostwal, Rebecca Lalngaihzuali, Nasi ra khalid Shaik h, Kis hore ingole, Rajaram powar
1. Assistant Professor, Department of Microbiology, Government Medical College, Akola, India. 2. Junior Resident, Department of Microbiology, Dr. VMGMC Solapur, India. 3. Associate Professor, Department of Microbiology, Dr. VMGMC Solapur, India. 4. Professor and Head, Department of Microbiology, Dr. VMGMC Solapur, India. 5. Dean, Department of Microbiology, Dr. VMGMC, Solapur, India.
 
Correspondence Address :
Karan Indra Ostwal, Rebecca Lalngaihzuali, Nasi ra khalid Shaik h, Kis hore ingole, Rajaram powar,
Dr. Karan Indra Ostwal,
Assistant Professor, Department of Microbiology,
Government Medical College Akola,
Maharashtra-444001, India.
E-mail: Karanost55@gmail.com
 
ABSTRACT
: Introduction: Corneal ulcer is the second most common cause of visual disability and blindness in developing countries after cataract. Corneal scarring due to keratitis can be prevented. This therefore necessitates the knowledge of its etiology for early intervention.

Aim: To study the occurrence, etiology and predisposing factors of mycotic keratitis at a tertiary care centre in Solapur, Maharashtra.

Materials and Methods: The retrospective study was carried out for duration of 2 years (April 2013 to March 2015). All aseptically collected corneal scrapings from clinically suspected cases of fungal keratitis were included. Conventional methods i.e. KOH mount, Gram’s stain and fungal cultures on a pair of Saubaraud Dextrose Agar were used for diagnosis. Slide cultures on Corn Meal Agar with tween 80 were used for confirmation of moulds and germ tube tests done for yeasts.

Results: Hundred corneal scrapings were studied out of which fungal hyphae were identified in 20% of the KOH mounts and in 17% of Gram’s stained smears. Fungal culture was positive in 24 samples. Predominant isolate was Fusarium spp (29%. 7/24) followed by Alternaria spp (21%, 5/24) and Aspergillus spp (17%, 4/24) and 8% (2/24) each of Cladosporium spp, Penicillium spp, Bipolaris spp and Candida albicans. Ocular injury was the most commonly encountered predisposing factor.

Conclusion: Fungal aetiology constitutes a significant proportion of corneal ulcers, out of which Fusarium spp is the most common isolate in Solapur, Maharashtra followed by Alternaria spp unlike majority of studies in the country where Aspergillus spp are the most common isolates.
Keywords : Corneal scrapings, Fungal culture, Fungal isolates, Microscopy
DOI and Others : 10.7860/NJLM/2016/17664:2108
 
INTRODUCTION
Keratitis is a condition in which the cornea becomes inflamed by infectious organisms like virus, bacteria, fungus and parasites or non-infectious agents (1). In developed countries viral infections are common whereas bacteria, fungi and Acanthamoeba are more common causes in developing countries (2). The condition is marked by moderate to intense pain, impaired eyesight, photophobia, red eye and a gritty sensation and often results in corneal scarring and opacification ultimately leading to blindness (3).

Retinal diseases (40–54%) are important cause of blindness in developed nations while it is cataract (44–60%) and corneal diseases (8–25%) for developing countries (4). Certain conditions like injury to the eye and therapy with antibiotics and corticosteroids render the eye susceptible to infection with various fungi especially in tropical parts of the world (5).

The incidence of fungal keratitis is higher in tropical regions of the world especially India (6), Aspergillus flavus is the most common isolate in Northern India (22.5%) (2), Eastern India (15.85%) (5), Western India (45.29%) (1) and Mumbai 14.7 % (6), Aspergillus fumigatus in Chennai 40% (7), Aspergillus spp in Chandigarh (41.18%) (8), and Fusarium spp in South India (42.%) (9). Yeast isolation from corneal ulcers is generally low in India (10),(11).

It is expected that individuals suffering from unilateral corneal blindness in India will be 10.6 million by 2020 (10). Corneal scaring due to keratitis can be prevented (12). Hence, this study aims to find out the occurrence of fungal keratitis, its aetiology and predisposing factors with respect to the city of Solapur.
 
 
Material and Methods
The retrospective study was carried out for duration of 2 years (April 2013 to March 2015), in the Department of Microbiology Dr V.M Government Medical college Solapur, Maharashtra, India. Based on inclusion and exclusion criteria sample was collected from 100 corneal ulcers scrapings and were studied.

Inclusion criteria

All corneal scrapings from clinically suspected cases of fungal keratitis received in the Microbiology Department Dr VMGMC, Solapur were included in the study.

Exclusion criteria

Clinically suspected bacterial and viral keratitis was excluded from the study.

Procedure

The corneal scrapings were collected aseptically by Ophthalmologists and sent to Microbiology Laboratory. They were then inoculated on to one Chocolate Agar and two Sabouraund’s Dextrose Agar (SDA) by making “C” or “S” streak to ensure fungal growth in the form of streak was from inoculum and not a laboratory contaminant (13). Due to paucity of samples and for better yield minimum media was used for inoculation. The remaining were taken on to the centre of clean glass slide to be used for direct microscopic examination.

The Chocolate Agar and one SDA were incubated at 370C and the other SDA at 250C and were observed daily for growth. Confirmation of moulds was done by putting slide cultures on Corn Meal Agar (CMA) with tween 80 and germ tube tests done for yeasts (14). The glass slides were used for performing KOH mount and Gram’s staining for the demonstration of fungal hyphae. Detailed relevant history including age, sex, occupation, address, history of ocular trauma/injury, duration of symptoms, previous treatment, predisposing ocular conditions, use of topical medicines / steroids and contact lens was recorded (Table/Fig 1),(Table/Fig 2).

Reading of Results

Fungus identification was done macroscopically based on the texture, growth rate, pigmentation, colony morphology on SDA slants, reverse and obverse surface colorof SDA slant and microscopically based on features such as mycelium and conidia types on Lactophenol cotton blue (LPCB) mounts of culture positive fungi.
 
 
Results
100 corneal scrapings were studied out of which fungal hyphae were identified in 20% of the KOH mounts and in 17% of Gram’s stained smears. Ocular injury was found to be the most commonly encountered predisposing factor. Predominant isolates were Fusarium solani (6) and Fusarium chlamydosporum (1) followed by Alternaria alternata (5) as shown in (Table/Fig 3). Pictures of F. solani and Alternaria alternata grown on SDA are shown in (Table/Fig 4),(Table/Fig 5) respectively.

Statistical analysis

KOH mount showed evidence of fungal hyphae in 20% of corneal scrapings examined. With culture taken as standard test,the sensitivity and specificity of KOH mount and Gram’s stain were calculated as shown in (Table/Fig 6),(Table/Fig 7) respectively. Kappa factor was found out to have a value of 0.901 in the study (Table/Fig 8). This reveals that the degree of agreement is quite high for the two tests. The above tables revealed that KOH mount preparation and Gram stained smears are simple and sensitive methods for early diagnosis.
 
 
Discussion
In Solapur, young adults (around 35 years) were most commonly affected and males showed higher positivity rates as compared to females. This could be due to more outdoor activity of males in their early adulthood. 77% of positive cases hailed from the rural areas where the occupation was mostly agricultural work in the fields. This again could be due to more exposure to environment in daily activity from which the fungal sources were contracted. These findings are similar with majority of studies on the topic all over India, though variation in percentage do exits. Winter months are associated with more cases as during this time as sugarcane harvesting starts and farmers are exposed to husks produced. Summer time arrived in Solapur with winds which also play a role in spreading the infectious fungi as well as in exposure to the population.

The predominant isolate in our study was F.solani in contrast to most of the studies and what the textbooks mention as Aspergillus Spp. Being the most common isolate, this suggest that there can be variation in isolation of fungal isolates depending on geographical aspect. Moulds were the commonly isolated fungi in the various studies as mentioned in (Table/Fig 9) which comprises of 95-98% which a small proportion of yeast were also found to be the causative factor in our study.

Limitation

Some of the genus like Cladosporium and Penicillium were not identified to genus level. Antifungal susceptibility testing was not performed in our study.
 
 
Conclusion
A significant proportion of corneal ulcer in Solapur is of fungal origin (24%). This proves that many ocular complications including corneal opacification and blindness can be avoided by prompt management of mycotic keratiis. Fusarium spp. being the most frequent isolates reveals that the geographical distribution of fungus varies even within the country. The most frequently encountered predisposing factor is ocular trauma. Hence, history of ocular trauma should raise a high suspicion of mycotic keratitis.
 
REFERENCES
1.
Jadhav SV, Gandham NR, Misra RN, Ujagare MT, Sharma M, Sardar RM. Prevalence of fungal keratitis from tertiary care hospital from western part of India. International Journal of Microbiology Research. 2012;4(4): 211-14.
2.
Gupta A, Capoor MR, Gupta S, Kochhar S, Tomer A, Gupta V. Clinico demograghical profile of keratomycosis in Delhi,North India. Indian Journal of Medical Microbiology. 2014; 32(3):310-14.
3.
Garg P, Krishna P, Stratis A, Gopinathan U. The value of corneal transplantation in reducing blindness. Eye. 2005;19(10):1106-14.
4.
Sharma S. Ocular infections: Research in India. Indian Journal of Medical Microbiology. 2010;28(2):91.
5.
Rautaraya B, Sharma S, Kar S, Das S, Sahu S. Diagnosis and treatment outcome of mycotic keratitis at a tertiary eye care center in eastern india. BMC Ophthalmology. 2011;11(1):39.   [Google Scholar]
6.
Baradkar VP, De A, Mathur M, Lanjewar M, Kumar S. Mycotic keratitis from Mumbai. Bombay Hospital Journal. 2008; 50(2):200-05.
7.
Kindo AJ, Suresh K, Anita S, Kalyani J. Fungus as an etiology in keratitis-our experience in SRMC. Sri Ramachandra Journal of Medicine. 2009;2(2):14-18.
8.
Chander J, Singla N, Agnihotri N, Arya S, Deep A. Keratomycosis in and around Chandigarh: A five-year study from a north Indian tertiary care hospital. Indian Journal of Pathology and Microbiology. 2008;51(2):304.
9.
Bharathi M, Ramakrishnan R, Meenakshi R, Padmavathy S, Shivakumar C, Srinivasan M. Microbial keratitis in South India: Influence of risk factors, climate, and geographical variation. Ophthalmic Epidemiology. 2007;14(2):61-69.
10.
Dandona R. Corneal blindness in a southern Indian population: need for health promotion strategies. British Journal of Ophthalmology. 2003;87(2): 133-41.
11.
Chowdhary A, Singh K. Spectrum of fungal keratitis in North India. Cornea. 2005;24(1):8-15.
12.
Basak S, Basak S, Mohanta A, Bhowmick A. Epidemiological and microbiological diagnosis of suppurative keratitis in Gangetic West Bengal, Eastern India. Indian J Ophthalmol. 2005;53(1):17.
13.
Chander J. Textbook of Medical Mycology. 3rd ed. New Delhi: Mehta Publishers;1995.
14.
Bharathi M, Ramakrishnan R, Meenakshi R, Shivakumar C, Raj D. Analysis of the risk factors predisposing to fungal, bacterial and Acanthamoeba keratitis in south India. Indian J Med Res. 2009;130:749-57.
15.
Jastaneiah S, Ali A, Al-Rajhi, Abbott A. Ocular mycosis at a referral center in Saudi Arabia, a 20-year study. Saudi Journal of Ophthalmology. 2011;25:231-38.
16.
Sengupta J, Saha S, Banerjee D, Khetan A. Epidemiological profile of fungal keratitis in urban population of West Bengal, India. Oman Journal of Ophthalmology. 2009;2(3):114.
17.
Punia S, Kind R, Chander J, Arya S, Handa U, Mohan H. Spectrum of fungal keratitis, clinicopathologic study of 44 cases. Int J Ophthalmol. 2014;7(1):114-17.
18.
Nath et al. Mycotic corneal ulcers in upper Assam. Indian J Ophthalmol. 2011;59(5):367–71.  [Google Scholar]
 
TABLES AND FIGURES