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  5. FP615 : Spectral Domain Anterior Segment Optical Coherence Tomography (ASOCT) in Fungal Keratits

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Dr. Deepali Singhal, S18862,
Dr. Tushar Agarwal, Dr. Jeewan S Titiyal, Dr.Namrata Sharma

Abstract

Aim: To evaluate the use of Spectral Domain Anterior Segment Optical Coherence Tomography (ASOCT) in Fungal Keratitis.

Settings and Design: This study was conducted in a tertiary care centre, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, AIIMS, New Delhi. This study is a prospective case series study.

Methods: 31 eyes with fungal keratitis (smear or culture proven) were recruited. from the cornea and emergency services.Serial ASOCT scans and slit lamp examinationwas performed on day 0, 7, 14, 21, 28, 42 and 56. Corneal thickness (CT) in the infiltrate area and infiltrate thickness (IT) were measured at each followup.

Statistical analysis:Statistical analysis was performed using the SPSS 20 software program for Windows. Data was presented as mean ± SD/ median (minimum – maximum) and frequency percentage as applicable.

Results: Mean CT and IT at presentation were 654.17±103 μm and 411.17 ± 91 μm respectively. CT and IT reduced significanltly at each follow-up compared to baseline [On day 7, 14, 28 and 42; 630.17±108 μm and 376.51±91 μm (p<0.001), 606.79±105 μm and  355.10 ± 94 μm (p<0.001), 548.51 ± 102 μm and  310.31 ± 83 μm (p<0.001), 525.17 ± 96 μm and  293.56 ± 69 μm (p<0.001) respectively]. These findings corroborated with the healing seen clinically. Mean CT and scar size at complete healing was 514 ± 83 μm and 301.18 ± 88 μm. Corneal perforation occured in 2 cases [on day 14 and 21] which underwent keratoplasty.

Conclusions: ASOCT is useful in quantitative monitoring of fungal keratitis.

Keywords: Fungal keratitis; ASCOT; infiltrate thickness 

Introduction

Microbial keratitis leading to corneal scarring is one of the leading cause of blindness in the world especially in developing nations like Asia, Africa and the Middle East.[1] Fungal keratitis is responsible for 30-40% of corneal ulcers in India.[2-7] Various risk factors associated with fungal keratitis include trauma with vegetative matter (most common), poor ocular surface, long term use of topical steroids, contact lens use, systemic immunosupression and diabetes mellitus.[5] Patients of fungal keratitis present with minimal symptoms as compared to signs which include a dry looking ulcer with raised and feathery margins with stromal infiltrates, satellite lesions (10.05%) with fixed hypopyon in 55.62%.[5] Endothelial plaques and immune ring may also be seen.[7]Titiyal et al reported that trauma with organic matter, central location of ulcer, vision less than FCCF, delay in starting treatment, lack of vascularization and alcoholism were the risk factors for perforation.[8]

Routine clinical examination and monitoring of microbial keratitis is done with the help of slit-lamp biomicroscopy that may be limited by the physical properties of light. With this technique the site along with the size and extent of the corneal ulcer can be measured in the longest dimension and perpendicular to it.[9] However, the depth of the infiltrates and associated corneal edema quantified by thickness are often difficult to assesswith slit-lamp examination.This makes the objective assessment and monitoring of keratitis difficult while, subjective features, such as improvement of patient symptoms, can be used as indicators of resolving infection.

Recently, however, quantitative assessment of microbial keratitis has been reported using serial anterior segment optical coherence tomography (AS-OCT) scans.[10-13]This non contact imaging modality provides cross-sectional imaging of the cornea and quantitative assessment of the depth of stromal infiltration as well as width of the infiltrates along with measuring the corneal thickness; an objective marker of corneal edema.[10] The two most commonly used technologies in ASOCT are time domain (Visante) and spectral domain (Optovue). Spectral domain ASOCT (SD-ASOCT) is an advanced technology which captures the images without a moving mirror, providing a high resolution of 5 μm (axial) and 15 μm (transverse) as compared to time domain OCT having a resolution of  5 μm (axial) and 15 μm (transverse). SDOCT uses a shorter wavelength ligft of 840nm with a lower optical power with a faster scan speed of 25-30,000 A scans/second. The major limitation is that the depth and the length of the scan (2×8 mm) is much less as compared to time domain OCT (6×16 mm). Due to a high resolution of SD-ASCOT, this has been used for imaging the details of stromal infiltrates in microbial keratitis more efficiently as compared to TD OCT.[11]

Hereby, we present a prospective study of patients with fungal keratitis which were monitored with serial ASOCT scans along with the clinical evaluation.

Methods

Patients were recruited from the Cornea Clinic of a tertiary eye care hospital, from August 2015 to August 2016. Written informed consent was obtained from all participants. Institutional ethics committee approval was obtained from the Institutional Review Board/Ethics Committee, AIIMS, New Delhi. The research was conducted adhering to the tenets of the Declaration of Helsinki.Patients had the right to refuse to participate and/or withdraw from the study at any time without being denied their regular, full clinical care.Thirty five patients with smear or culture positive fungal keratitis were included in the study.A detailed clinical history, including age, sex, occupation, symptoms, onset, and progression, and predisposing factors were taken along with slit lamp examination at each visit. The size of the epithelial defect and size the stromal infiltrate was measured in the greatest dimension and along an axis perpendicular to it using a slit lamp. The depth of the infiltrate was noted on slit lamp examination by comparing it with the corneal thickness wherever possible. The presence of hypopyon, cataract, and posterior synechiae, if any, were also recorded. Ultrasonography was performed in patients in whom the posterior segment was not visualized to rule out coexisting endophthalmitis.Baseline clinical photograph with and without fluorescein staining was performed. After clinical examination, corneal scraping was performed in all the patients, and the sample was sent for Gram staining, potassium hydroxide wet-mount preparation, bacterial culture(blood agar and chocolate agar), and fungal culture (Sabouraud dextrose agar).All patients were followed up on days 3, 7, 14, 21, and every week until complete ulcer healing had taken place. The size of the scar was measured using a slit lamp in the greatest dimension and that along an axis perpendicular to it.

Examination and scanning protocol

All included patients underwent serial ASOCT examinations using SD-OCT (RTVue-100; Optovue, Freemont, CA, USA) performed by a single OCT expert. This device is primarily for imaging of the posterior segment and uses a cornea/anterior module-long (CAM-L) to obtain anterior segment images. ASOCT imaging was carried out on day 0, 7, 14, 21, 28 and 56. All patients were administered standard antifungal therapy including topical Natamycin 5% every 1 hour, for the first 48 hours, every 2 hours during waking hours until epithelial healing, and then every 4 hours for 3 weeks. In addition, cycloplegics were prescribed in the form of topical homatropine 2% four times a day. Oral antifungals were prescribed in severe cases (>5 mm in size or involvement of >50% stromal depth).

A standardized scanning protocol was used for all cases. Serial ASOCT scans were taken at the centre of the infiltrate using a 6mm horizontal raster scan at all visits. Stromal infiltrate was seen as the hyper-reflective area often associated with epithelial defect. Corneal thickness (CT) and infiltrate thickness (IT) were measured along an axis perpendicular to the horizontal axis of the scan at the centre of the infiltrate in all cases. However, it was not possible to measure infiltrate width (IW) in all cases due to the limited scan length (upto 8mm) in SD-ASOCT. CT was measured in the center of the infiltration with one caliper arm on the most anterior hyper-reflective corneal surface and the second arm on the hyper-reflective endothe- lium. IT, also in the center of the infiltration, was measured with one caliper arm on the most anterior hyper-reflective corneal surface and the second arm on the posterior border of the hyper-reflective area. Endothelial plaque if present was also measred and monitored. Thus,the CT and IT were measured on days 0, 7, 14, 21, 28 and 56. Mean values were calculated at each visit and compared.

Statistical analysis

Statistical analysis was performed using the SPSS 20 softwareprogram for Windows. Data was presented as mean ± SD/ median (minimum – maximum) and frequency percentage as applicable.Over a period of time the two variables CT and IT were compared from day 0 to day 56 by using repeated measures. The adjustment for multiple post-hoc comparisons was done with Bonferroni correction.

Results

Thirty one patients were included in the study. Out of these, 2 patients developed corneal perforation within 7 days which underwent therapeutic keratoplasty. Thus, 29 patients were analysed at the end of the follow up period. Fungal culture was seen to be positive in 17/29 cases and the organisms isolated were Aspergillus (7), Fusarium (5), Alternaria (1), Curvularia (1), Candida (3) in Sabouraud’s Dextrose Agar. Serial ASOCT imaging was done in all the cases and the changes in the measurement of CT and IT were recorded as illustrated in Figure 3 and 4.

CT was measured in the infiltrate area in most of the cases except in a few (7 cases) where posterior shadowing was seen due to the infiltrates. In these cases, CT was measured just adjacent to the area of infiltrates and was seen to be maximum on day0. Mean CT at day 0 was 654.17±103 μm which reduced to 630.17±108 μm (p<0.001), 606.79±105 μm (p<0.001), 570.93± 105μm (p<0.001), 548.51 ± 102 μm (p<0.001), and 525.17 ± 96 μm (p<0.001), at day 7, 14, 21, 28 and 42 respectively (Figure 3).

IT was measured at the centre of the corneal ulcer in all cases and was seen to be maximum at presentation. Mean IT at day 0 was 411.17 ± 91 μm which reduced to 376.51 ± 91 μm (p<0.001), 355.10 ± 94 μm (p<0.001),325.82 ± 89μm (p<0.001), 310.31 ± 83 μm (p<0.001), and 296.72 ± 81 μm (p<0.001), at day 7, 14, 21, 28 and 42 respectively (Figure 4).The mean CT and the mean scar depth at day 56 was 497.10 ± 97μm and 292.17 ± 79 μm respectively.

A detailed analysis of the SD-ASOCT images showed that infiltrates were seen as hyperreflective areas within the stroma associated with epithelial defect along with stromal edema in active phase. At the time of healing corneal scar was seen as a hyperreflective area with intact epithelium and stromal thinning. Endothelial plaque was seen a hyperreflective area at the endothelium with irregular borders in 10/29 cases (34.4%). Stromal cysts or necrotic areas were seen in 7/29 cases (24.1%).

Discussion

We demonstrated the role of SD-ASOCT in microbial keratitis which allows objective assessment and monitoring of the disease course through serial examinations. Though clinical observation by slit lamp biomicroscopy is the gold standard method for monitoring of corneal infections, the comparison across time by this method may be very subjective. Moreover, the assessment of depth of the infiltrates is often difficult by this method. The measurements done by SD-ASOCT are objective and can also reduce the interobserver variability. Various corneal parameters like CT and IT can be measured using the callipers provided in the software. The corneal epithelium and endothelium are normally seen as hyperreflective layers which are used as reference for the placement of callipers. In this study, we measured the CT using these two layers while the IT was measured using the perpendicular between the epithelial layer and the lowermost border of the hyerreflective area seen within the stroma. It was observed that within the initial stages of keratitis only, significant edema was seen in the form of increased CT. At complete healing,the scar tissue was seen as a hyperreflective area associated with epithelial healing and stromal thinning. In a few cases CT could not be measured in the infiltrate area due to posterior shadowing in cases with dense infiltrates. However, on complete healing it was possible to measure the scar depth as well as the corneal thickness possibly due to decrease in the intensity of hyperreflectivity. Thus, the intensity of the infiltrates is an important limiting factor for measurement in ASOCT.

Konstantopoulos et al. quantified the inflammatory response in bacterial keratitis using the long wavelength TD-ASOCT. They measured CT, IT ans IW on serial scans and showed that there was a rapid decrease in CT and IT in early phase (day 0-3) and in late phase (day 7-14) IT reduced faster than CT. However, the IW did not change during the course of treatment.[12] In our study, we were not able to measure the width of the infiltrate in all cases due to the limited scan size of 8mm in SD-ASOCT. Thus, the major limitation of SD-ASOCT is that only a definite section of the cornea can be visualized and limbus to limbus image is not available. However, the advantages like high resolution and the provision of both horizontal and vertical scans could allow this to be used as an adjunctive method for monitoring of keratitis.

Soliman et al described ten common morphological patterns in microbial keratitis which were hyperreflective stromal lesions, epithelial defect, stromal edema, hyperreflective material over epithelium or attached to corneal endothelium, stromal thinning with defective epithelium, descemetocele, hyperreflective stromal lesion (scar) with intact ith intact epithelium with or without stromal thinning, and diffuse stromal thinning with defective epithelium on top.[11]Two unique patterns for fungal keratitis identified were stromal cystic spaces and necrotic stroma.[11]In our study where cases of fungal keratitis were included, stromal cysts were seen in 24.1% cases. Another common feature identified in fungal keratitis was the endothelial plaque seen in 34.4% cases.

SD-ASOCT has also been used to quantify the decrease in corneal edema as well as the size of keratic precipitates in two patients with disciform keratitis.[13] Many studies have shown OCT to be a precise tool for measurement of central corneal thickness.

To conclude, SD-ASOCT can be used as an adjunctive tool for objective assessment and quantitative monitoring of the patients of keratitis with a greater accuracy. However, it must be emphasized that it’s not an absolutely essential tool for management of fungal keratitis but if available it is an extremely useful adjunctive tool for the treating physician. 

References

  1. Chirambo MC. Causes of blindness among students in blind school institutions in a developing country. Br J Ophthalmol 1976; 60: 665-668.
  2. Srinivasan M, Gonzales CA, George C, Cevallos V, Mascarenhas JM, Asokan B, et al. Epidemiology and aetiological diagnosis of corneal ulceration in Madurai, South India. Br J Ophthalmol 1997;81:965-71.
  3. Gopinathan U, Garg P, Fernandes M, Sharma S, Athmanathan S, Rao GN. The epidemiological features and laboratory results of fungal keratitis: A 10-year review at a referral eye care center in south India. Cornea 2002;21:555-59.
  4. Bharathi MJ, Ramakrishnan R, Vasu S, Meenakshi R, Palaniappan R. Aetiological diagnosis of microbial keratitis in South India. Indian J Med Microbiol 2002;20:19-24.
  5. Bharathi MJ, Ramakrishnan R, Vasu S. Epidemiological characteristics and laboratory diagnosis of fungal keratitis. A three-year study. Indian J Ophthalmol. 2003;51:315–321
  6. Bharathi MJ,Ramakrishnan R, Meenakshi R. Microbial Keratitis in South India: Influence of Risk Factors, Climate, and Geographical Variation. Ophthalmic Epidemiology, 2007;14:61–69
  7. Chowdhary A, Singh K. Spectrum of fungal keratitis in north India. Cornea 2005;24:8–15.
  8. Titiyal JS, Negi S, Anand A, Tandon R, Sharma N. Risk factors for perforation in microbial corneal ulcers in north India Br J Ophthalmol 2006;90:686-689
  9. Efron N, Morgan PB, Hill EA, Raynor MK, Tullo AB. The size, location and clinical severity of corneal infiltrative events associated with contact lens wear. Optom Vis Sci. 2005;82:519–527.
  10. Konstantopoulos A, Kuo J, Anderson DF, Hossain PN. Assessment of the use of anterior segment optical coherence tomography in microbial keratitis. Am J Ophthalmol. 2008;146:534–542.
  11. Soliman W, Fathalla AM, El-Sebaity DM, Al-Hussaini AK. Spectral domain anterior segment optical coherence tomography in microbial keratitis. Graefes Arch Clin Exp Ophthalmol. 2013 Feb;251(2):549-53.
  12. Konstantopoulos A, Yadegarfar G, Fievez M, Anderson DF, Hossain P. In vivo quantification of bacterial keratitis with optical coherence tomography. Invest Ophthalmol Vis Sci. 2011 Feb 25;52(2):1093-7.
  13. Yokogawa H, Kobayashi A, Yamazaki N, Sugiyama K. In vivo imaging of coin-shaped lesions in cytomegalovirus corneal endotheliitis by anterior segment optical coherence tomography. Cornea. 2014 Dec;33(12):1332-5. 

Figure Legends

Figure 1: Clinical photographs and respective Spectral Domain Anterior Segment Optical Coherence Tomography (ASOCT) photographs of a fungal keratitis patient. Figure 1a shows the area of stromal infiltrate as the hyperreflective area with an overlying epithelial defect. Figure 1d shows complete healing with scar formation as a hyperreflective area with healed epithelial defect along with stromal thinning.

Figure 2: Clinical photograph and respective Spectral Domain Anterior Segment Optical Coherence Tomography (ASOCT) photograph of a patient showing the scanning protocol. CT was measured along an axis perpendicular to the horizontal axis in the center of the infiltration with one caliper arm on the most anterior hyper-reflective corneal surface and the second arm on the hyper-reflective endothelium. IT, also in the center of the infiltration, was measured with one caliper arm on the most anterior hyper-reflective corneal surface and the second arm on the posterior border of the hyper-reflective area. Figure 2b also shows stromal necrotic areas with an overlying epithelial defect.

Figure 3: Line diagram showing the trend of changes in corneal thickness with time on Spectral Domain Anterior Segment Optical Coherence Tomography (ASOCT) in fungal keratitis.

Figure 4: Line diagram showing the trend of changes in infiltrate thickness with time on Spectral Domain Anterior Segment Optical Coherence Tomography (ASOCT) in fungal keratitis.

FIGURES?

 

 

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