Full-field electroretinography, visual acuity and visual fields in Usher syndrome: a multicentre European study
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Usher syndrome (USH) is a multisensory deficiency involving vision, hearing and the vestibular system. The purpose of this study is to report on the functional data (i.e. electroretinography, visual fields, visual acuity) of patients with retinitis pigmentosa (RP) due to Usher syndrome that were collected in a multicentre European study (TREATRUSH).
A total of 268 genetically confirmed USH patients underwent electrophysiological examinations in the context of multimodal ophthalmological examination in the study (75 USH1, 189 USH2 and four USH3). Full-field electroretinography (ERG) was performed according to ISCEV standards, visual field determination was carried out with either the Octopus or Goldmann perimeters and visual acuity was examined with either ETDRS or Snellen charts. The data were compared between USH subtypes (USH1/USH2/USH3) and correlated with age.
Visual acuity decreases significantly with age for both USH1 and USH2 (p < 0.001), without a difference between the two cohorts. When corrected for age, the preserved kinetic visual field was significantly larger in USH2 than in USH1 (p = 0.04). Furthermore, the preserved kinetic visual field area showed a significant decrease with age (based on an exponential fit) in both USH1 and USH2 (p < 0.001). In USH1 patients, however, the visual field was already vastly reduced at an early age. The ERG results were abnormal in all patients. Detectable data for scotopic ERG were obtained from nine patients, and data of photopic ERG were obtained from 24 patients, without a difference between USH1 and USH2 subtypes.
There are differences in the phenotypes of RP in USH subtypes, most visible in the progression of visual fields between USH1 and USH2. The perimetric reduction occurs earlier in USH1 than in USH2. In both subtypes, visual acuity decreases significantly with age and the ERG is not detectable already at early ages.
KeywordsERG Usher syndrome Visual field Visual acuity Functional diagnostics
Usher syndrome (USH) is an autosomal recessive sensory disease combining deafness, visual impairment (retinitis pigmentosa, RP) and in some cases vestibular and olfactory deficits [1, 2]. There are three different clinical subtypes (USH1, USH2 and USH3). USH type 1 (USH1) patients are normally born completely deaf with delayed sitting and walking due to vestibular deficits, and night blindness diagnosed at about 10 years of age. In USH2 patients, on the other hand, there are no balance defect and usually a hearing impairment but no deafness is present at birth, which does not progress over time in most of the cases. Night blindness also normally occurs later than in USH1 . In USH3, which accounts for only about 2% of patients, the hearing impairment is progressive and a balance defect may be present. This phenotype is found mostly in Finland and Ashkenazi Jews [4, 5]. At least 11 causative genes have been found to be associated with USH: six for USH1 (MYO7A/USH1B, CDH23/USH1D, USH1C/DFNB18, USH1G/SANS, PCDH15/USH1F, CIB2/USH1J), three for USH2 (i.e. USH2A, ADGRV1/USH2C (formerly called GPR98 or VLGR1), DFNB31/USH2D) and two for USH3 (CLRN1/USH3A, HARS) [2, 6].
USH3 is a very rare subtype with a later, post-lingual and progressive onset of auditory symptoms . USH1 and USH2 have distinct auditory and vestibular phenotypes, with visual symptoms exhibiting differences with a large heterogeneity. Some observations show that the retinal degeneration in USH is similar to non-syndromic RP, but there are differences in the three subtypes and some inconsistency in the published results. Research shows that especially for the USH2 subtype the natural course of the disease exhibits the same phenotypic heterogeneity as the non-syndromic retinal disease and once the disease becomes symptomatic, visual fields deteriorate following stereotyped kinetics, even in patients with late-onset retinal disease . Other longitudinal data indicate that the prognosis of the USH2A phenotype is more severe than that of non-syndromic RP [8, 9]. In the USH1B phenotype (MYO7A), central vision typically remains preserved until the third decade of life . Based on ERG results, the retinal degeneration in Usher syndrome has been previously described as an equal reduction or loss of rod and cone function, in contrary to the classical non-syndromic RP where rod function deteriorates first followed by cone function loss . However, the majority of published reports confirm that the retinal degeneration follows a rod–cone dystrophy pattern, corresponding to the non-syndromic RP, even in the early childhood [12, 13, 14].
Although published reports are not always consistent, there are differences in severity between USH1 and USH2 retinal phenotypes: the age of symptom onset is earlier in USH1 and this subgroup seems to have a more severe visual impairment—especially regarding the progression of visual functions [3, 15, 16, 17]. Morphologically, both USH1 and USH2 phenotypes show similar pattern of disease progression, as shown by fundus autofluorescence or optical coherence tomography, as well as cataract development [3, 18]. However, studies have failed to show a difference in the rate of progression or age at reaching legal blindness between the subtypes [3, 18].
Here, we examine the ERG, visual acuity and visual field data gathered from a large cohort of 268 USH patients in a multicentre European study TREATRUSH (https://cordis.europa.eu/project/rcn/95259_en.html), which was carried out to advance our understanding of the disease through phenotypic characterization and genetic analysis.
A total of 268 genetically confirmed Usher syndrome patients underwent electrophysiological testing in the context of multimodal ophthalmological examination in the study (75 USH1, 189 USH2 and four USH3). Following genotypes were included in USH1: 52 USH1B (MYO7A), five USH1C (USH1C), eight USH1D (CDH23), eight USH1F (USH1F) and two USH1G (USH1G) patients. In the USH2 cohort, there were 172 USH2A (USH2A) patients and 17 USH2C (ADGRV1) patients. The four USH3 patients were classified as USH3A.
The patients were examined at four different centres: Centre for Ophthalmology, University of Tuebingen, Germany; CHNO Quinze-Vingts, Paris, France; University Eye Hospital Ljubljana, Slovenia and Academic Hospital University of Campania “Luigi Vanvitelli”, Naples, Italy. Data were entered into a central database . Informed consent was obtained from all individual participants included in the study.
Visual acuity: Best-corrected visual acuity was measured with either the ETDRS or Snellen charts, at a distance of 4 m. It was converted to logMAR units.
Visual field: The kinetic visual field was examined using the III4e target with either the Octopus (Haag-Streit International, Germany) or Goldmann perimeters. In centres using Octopus perimeter (Haag-Streit International, Germany), the visual field area was quantified by the automated device software. In centres using Goldman perimeter, the visual fields were scanned and measured the visual field area using the “Area tool” in Adobe Acrobat Pro. The grid markings on the Goldmann visual field were used to calibrate the tool. The preserved area (deg2) was taken for further analysis.
Full-field: ERG recordings were performed according to ISCEV standards [11, 12], with either the Espion (Diagnosys Ltd, UK) or the EREV 2000 (LACE Elettronica srl, Italy) devices. The protocol included scotopic and photopic flash ERG recordings and 30 Hz flicker.
The right eye was taken for analysis. The data were evaluated using Microsoft Excel (2010) and JMP® 13.0.0. Because USH1 and USH2 groups do not have the same age range, differences between subtypes were tested using an ANCOVA, with age as a covariate, after standard least squares modelling. Significance was set to p < 0.05.
The patients were aged on average 42.48 years ± 13.36. The average age for USH1 was 37.64 years ± 13.21, USH2 44.55 years ± 12.88 and USH3 33.96 years ± 11.68. There was a significant difference in age between USH1 and USH2 subtypes (p = 0.0001). Two of the USH1 patients (2.7%) and ten of the USH2 (5.3%) had cataract. Among the patients were 125 females and 143 males.
From the total of 268 patients, from 19 patients (seven USH1 (9.3%), 11 USH2 (5.9%) and one USH3 (25%)) no data for visual acuity were documented, usually due to poor vision.
The average age of USH1 and USH2 patients whose ERG was detectable is not significantly different.
There is no correlation between the ERG results and the visual field area for either USH1 or USH2, and there is no significant relationship between visual acuity and visual field. There was no correlation between the ERG results and the visual acuity for either USH1 or USH2.
Our analysis presents functional visual results from a large multicentre European study TREATRUSH (https://cordis.europa.eu/project/rcn/95259_en.html), carried out to examine the genotype–phenotype correlation in the so far largest published cohort of 268 patients suffering from Usher syndrome. We examined full-field ERG, visual fields and visual acuity in a transversal manner and correlated the findings with age. Additionally, we searched for differences between the subtypes of Usher syndrome (types 1, 2 and 3). Due to a very low number of USH3 cases, only differences between USH1 and USH2 could be analysed statistically.
We found that full-field ERG responses were not detectable in the majority of the patients: from the 221 available recordings, 192 were below the measurable threshold (87%) for any standard response (scotopic and photopic); however, the patients with still detectable ERG signals were statistically similarly represented in USH1 and USH2, none of them presenting with a normal ERG result. In the USH3 cohort, none of the patients had a detectable ERG response. Although the retinal degeneration associated with Usher syndrome is classically considered a rod–cone dystrophy, descriptions of a rather equal loss of rod and cone function over the time course have also been published [7, 11, 12]. Our data indicate that the loss of full-field ERG responses occurs very early in most of the USH patients; no detectable scotopic ERG responses after the age of 40 years, but still some detectable photopic signals around the age of 60 in single patients indicate rather a rod–cone type of degeneration. Also, preserved visual acuity at the time of non-detectable ERG suggests preservation of central cones.
Although the difference between ERG responses of USH1 and USH2 was not statistically significant, our data show that the oldest patients for whom scotopic and photopic ERG responses were detectable belonged to the USH2 group. This supports the more severe course of retinal disease in the USH1 cohort, visible also in our data of the visual field area in USH1 vs. USH2. An earlier retrospective study on 22 USH patients with average age of 27 years showed non-detectable dark-adapted responses in all patients and non-detectable cone responses in 95% of patients. Average 30 Hz amplitude was approximately 3 µV in USH1 and 1 µV in USH2; implicit times were not significantly different between the two groups . Zein et al.  published recently that a microvolt level ERG recording of the cone flicker might represent a reliable response evaluation of USH patients for whom the standard recordings do not show measurable responses. Herewith, it could be shown that USH1 and USH2 have similar progression rates even after standard ERG is not detectable. This analysis requires a cycle-by-cycle recording as published by Sieving et al. earlier  that was not included in the study protocol of the presented multicentre study. A large study on 105 USH2A patients (average age of 32 years) reported average 30 Hz cone amplitude of 6 µV (0.1–59.0 µV) which decreased at rate of 13% per year. No difference between syndromic and non-syndromic RP was found .
Furthermore, a correlation between the maximal ERG b-wave amplitude and visual field size has been shown ; however, due to the non-detectable responses in most of the patients this relationship could not be analysed statistically in this study.
The full-field ERG is an important diagnostic test in combination with an ophthalmological examination to screen for Usher syndrome in children with hearing loss. However, clinical experience and earlier reports show that the full-field ERG responses decrease to non-detectable values in the early stages of retinal degeneration and thus are not a sensible tool to assess remaining retinal function. A study on 14 children with Usher syndrome showed early alterations in full-field ERG, corresponding to a rod–cone dystrophy in all. Remaining rod function could be verified in the majority of the children up to 4 years of age. After 4 years of age, there was a further deterioration of the rod function. In all children, the cone function was moderately reduced, in a few cases almost normal . Another study included five children with USH1 (aged 6 months to 5 years at the time of ERG recordings) in which all had abnormal ERG recordings with mostly non-detectable responses. Only two children had detectable rod responses (ages 6 months and 4 years) and only two others had detectable cone responses .
Unlike full-field ERG, the visual field represents an established, even though subjective, clinical progression parameter over years or decades. It is the preserved area of a kinetic visual field, where the difference between USH1 and USH2 becomes obvious: in our cohort of USH1, the visual field is vastly reduced mostly around 20 years of age, showing some further, statistically significant, exponential reduction with age. In USH2, the heterogeneity of the visual field findings is high, even up to the age of 50–60 years and there is an exponential significant decrease in the visual field area with age for the USH2 group, too. Although there are severely affected individuals in USH2, having massive visual field constriction already at an early age, similar to the USH1 phenotype, the large variability of the preserved visual field area in the USH2 cohort makes it phenotypically less severe. This may explain why some reports describe the functional progression in both USH types as similar , whereas most publications report a more rapid progression for USH1 compared to USH2 [15, 16, 17]. Due to the small sample size of USH1C, USH1D and USH1F in comparison with USH1B, as well as the small sample size of USH2C in comparison with USH2A, no valid group statistics could be performed to compare the genotype-specific differences in visual field progression. However, the descriptively plotted values in USH1 do not indicate any obvious difference in progression among USH1 genotypes, except the one outlier of USH1F. Almost all USH1 patients had a visual field area corresponding to a concentric reduction to approximately 10 degrees or less. In the USH2 cohort, the visual field was better preserved for some USH2A patients under 60 years of age; in USH2C, all but one had a similar visual field constriction as USH1 patients.
For the visual acuity, we found in both USH1 and USH2 a significant decrease with age without a difference between the two subtypes. In contrary to ERG and to some extent visual field areas, visual acuity remains preserved in RP usually until late stages of the disease. However, Edwards and co-workers  reported that the maximum difference in visual acuity between the two groups occurred during the third and fourth decades of life, with the USH1 patients being more impaired. A post hoc analysis in our cohort of patients in the third and fourth decades shows only a tendency for a difference in the visual acuity (p = 0.052) between USH1 and USH2. Again, due to the small sample size of USH1C, USH1D and USH1F in comparison with USH1B, as well as the small sample size of USH2C in comparison with USH2A, we did not consider group statistics to be a valid indicator of the visual acuity differences between the genotypes. The plotted values of genotypic subgroups neither for USH1 nor for USH2 indicate any obvious difference in visual acuity progressions.
Our data did not show any correlation between the functional tests: there was no correlation between the ERG results and the visual field area for either USH1 or USH2 and there was no significant relationship between visual acuity and visual field. There was no correlation between the ERG results and the visual acuity for either USH1 or USH2.
Although descriptions of the USH1 and USH2 phenotype are not consistent throughout the literature, our results support the fact that the retinal degeneration of Usher syndrome is of the classical rod–cone type and that the USH1 retinal phenotype has a more severe course than the USH2 phenotype. The visual field area analysis represents the most obvious difference between the severity and the progression speed of USH1 and USH2. This supports the clinical value of kinetic visual field measurement with the III4e stimulus for follow-up and consulting of patients with Usher syndromes.
In contrary to the auditory phenotype, this difference in severity is not inborn, but reflects a different progression of the retinal degeneration, which happens earlier and faster in USH1 [15, 16, 17]. The difference between USH types reflects the different retinal pathologies underlying the photoreceptor degeneration in the two types of syndrome. The USH1 protein complex is necessary for the growth of both cone and rod photoreceptor cells [24, 25] and is required for coupling between the outer segment of the photoreceptor and the inner segment and the calyceal processes, as well as between calyceal processes. On the other hand, USH2 proteins are localized to an inner segment membrane that surrounds the photoreceptor connecting cilium, the periciliary membrane complex [26, 27, 28].
Although we report here on the largest cohort of clinical functional data with USH so far, our data have some limitations. In this multicentre study, examinations were performed by several different investigators with different technical set-ups for each test, which may have led to larger variances in the data. Also, compliance to the study protocol adherence was different with many missing values in the database. Additionally, the visual acuity was also measured by two different methods, ETDRS and Snellen; they are not completely equivalent: Snellen is on average 0.13 logMAR units higher than ETDRS . Nevertheless, a re-analysis of the data with this as a correction factor did not significantly alter the conclusions of the study. Despite these weak points of the cohort report, the results represent the largest published cohort of patients with Usher syndrome so far.
This work was supported by the European Union Seventh Framework Programme under the Grant Agreement HEALTH-F2-2010-242013 (TREATRUSH) and the Tistou and Charlotte Kerstan Foundation.
Compliance with ethical standards
Conflict of interest
All authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest (such as honoraria; educational grants; participation in speakers’ bureaus; membership, employment, consultancies, stock ownership or other equity interest; and expert testimony or patent licensing arrangements) or non-financial interest (such as personal or professional relationships, affiliations, knowledge or beliefs) in the subject matter or materials discussed in this manuscript.
Statement of human rights
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.
Statement on the welfare of animals
This article does not contain any studies with animals performed by any of the authors.
Informed consent was obtained from all individual participants included in the study.
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