Page 8 - Read Online

Saleh et al. J Transl Genet Genom 2021;5:250-64 https://dx.doi.org/10.20517/jtgg.2021.23 Page 252

consistent with ABCA4-IRD [Table 1]. ABCA4 variants were classified based on the standards and
guidelines recommended by the American College of Medical Genetics and Genomics and the Association
[35]
for Molecular Pathology . Molecular genetic testing was performed in 9 patients by ABCR genotyping
[36]
[4]
microarray (ABCR400 chip) , 2 patients by a custom designed resequencing array (RetChip) , and 41
patients by Next Generation Sequencing using targeted gene panels including ABCA4 and a variable
[4]
number of other genes that have been associated with macular dystrophy . Additional causal sequence
changes in other genes than ABCA4 have not been identified in this group of patients.

All patients underwent clinical ophthalmological examination as well as detailed retinal imaging following
informed consent after detailed explanation about the background of the study. The study was performed in
adherence to the tenets of the Declaration of Helsinki. All applicable institutional and governmental
regulations concerning the ethical use of human volunteers were followed during this research.

Retinal imaging
Retinal imaging including multicolor spectral reflectance imaging, FAF, NIA, and OCT were performed as
[5]
described previously . All images were obtained after medical dilatation of the pupil (phenylephrine 2.5%
and tropicamide 1% achieving a minimal diameter of 5 mm) by trained retinal imaging specialists. FAF and
NIA were obtained with a confocal scanning laser ophthalmoscope (Heidelberg Retina Angiograph 2,
Heidelberg Engineering, Heidelberg, Germany) using 30° (M-FAF, M-NIA) and 55° lenses (W-FAF, W-
NIA). Multicolor spectral reflectance images and OCT were obtained with a spectral domain OCT
(Spectralis OCT, Heidelberg Engineering, Heidelberg, Germany). A standard volume scan macular OCT
(M-OCT) was recorded using 49 B-scans with a distance between B-scans of 129 µm in a 20 × 20 degree
field (6.2 mm × 6.2 mm) using ART mode with 16 images averaged. A wide-field (W-OCT) was recorded
using 31 B-scans with a distance between B-scans of 245 µm in a 55 × 25 degree field (16.1 mm × 7.3 mm)
using ART mode with 16 images averaged. All images were evaluated by two observers (Saleh M and
Kellner U), and W-OCT images were compared to M-OCT as well as M-/W-FAF and M-/W-NIA findings.
Lesions detected on retinal imaging were similar on both eyes in nearly all patients; therefore, the results are
presented combined for both eyes. In two patients (#38 and #52), the results of the more severely affected
eye were selected. In addition, unilateral focal choroidal excavation was identified in three patients (#12,
#44, and #48).

Statistical analysis
Correlation of the ABCA4 genotype with lesions detected by W-OCT was determined by applying a Fisher’s
exact test for count data implemented in the statistical programming software R . Differences in age
[37]
distributions were evaluated by a two-sided t test for independent samples by the t test function
implemented in R. Statistical significance was defined by a P-value below 0.05.

RESULTS
Fifty-two unrelated ABCA4-IRD patients were included in this study (32 females, 20 males; Table 2). The
age at first examination ranged 7-66 years, and it ranged 8-68 years at last examination. W-OCTs could be
recorded in 50/52 patients using 31 B-scans. Due to problems with fixation, in one patient the scan pattern
was reduced to 11 B-scans and in one patient only single B-scans were possible. Clinical findings as well as
retinal imaging results identified similar characteristics in both eyes of all patients except when specifically
mentioned.

W-OCT vs. M-OCT
Macular alterations on M-OCT and W-OCT presented as photoreceptor and RPE atrophy (n = 34), foveal
or perifoveal subretinal material (SRM; n = 8), a preserved foveal island with perifoveal atrophy (n = 4), a