Purpose: The purpose of this study was to assess changes in the neural retina in eyes with different stages of diabetic retinopathy (DR) in comparison to age-matched healthy subjects. Methods: Retrospective analysis of spectral-domain optical coherence tomography (SD-OCT) scans of 76 naïve eyes of 62 subjects with diabetes was performed. Key exclusion criteria included presence of diabetic macular edema, any other retinal disease, history of any treatment for DR, or incorrect segmentation of the retinal layers on SD-OCT scans. Eyes from diabetic patients were divided into three groups, including no DR, nonproliferative DR (NPDR), and proliferative DR (PDR). A control group of 67 eyes of 66 age-matched healthy volunteers was included for comparison. Average, minimum, and sectoral thicknesses for the ganglion cell-inner plexiform layer (GCIPL) and retinal nerve fiber layer (RNFL) were collected from both groups and compared using an ANOVA test. Results: Among the 76 included eyes, 43 had NPDR, 13 had PDR, and 20 had no signs of DR. Average and minimum GCIPL showed significant thinning in diabetic subjects compared with controls in all stages of DR (P < 0.05), especially involving the papillo-macula bundle. However, GCIPL thickness was similar between diabetic groups. There was no significant difference in average or sectoral RNFL thicknesses among groups; however, the minimum RNFL thickness was lower in diabetics compared with controls (P < 0.05). No relationship between GCIPL and RNFL thicknesses and duration of diabetes was present. Conclusions: Early thinning on the inner retina happens in type 2 diabetes, even before visible vascular signs of DR. This supports the presence of a neurodegenerative process in eyes of patients with diabetes and warrants neuroprotective intervention to prevent chronic neurodegeneration. The SD-OCT may represent an indispensable tool for identifying early signs of neurodegeneration in diabetic patients.

Acanthamoeba, an opportunistic pathogen is known to cause an infection of the cornea, central nervous system, and skin. Acanthamoeba feeds different microorganisms, including potentially pathogenic prokaryotes; some of microbes have developed ways of surviving intracellularly and this may mean that Acanthamoeba acts as incubator of important pathogens. A systematic review of the literature was performed in order to capture a comprehensive picture of the variety of microbial species identified within Acanthamoeba following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. Forty-three studies met the inclusion criteria, 26 studies (60.5%) examined environmental samples, eight (18.6%) studies examined clinical specimens, and another nine (20.9%) studies analysed both types of samples. Polymerase chain reaction (PCR) followed by gene sequencing was the most common technique used to identify the intracellular microorganisms. Important pathogenic bacteria, such as E. coli, Mycobacterium spp. and P. aeruginosa, were observed in clinical isolates of Acanthamoeba, whereas Legionella, adenovirus, mimivirus, and unidentified bacteria (Candidatus) were often identified in environmental Acanthamoeba. Increasing resistance of Acanthamoeba associated intracellular pathogens to antimicrobials is an increased risk to public health. Molecular-based future studies are needed in order to assess the microbiome residing in Acanthamoeba, as a research on the hypotheses that intracellular microbes can affect the pathogenicity of Acanthamoeba infections.

To determine the frequency of different types of spectral domain optical coherence tomography (SD-OCT) scan artifacts and errors in ganglion cell algorithm (GCA) in healthy eyes. Infrared image, color-coded map and each of the 128 horizontal b-scans acquired in the macular ganglion cell-inner plexiform layer scans using the Cirrus HD-OCT (Carl Zeiss Meditec, Dublin, CA) macular cube 512 × 128 protocol in 30 healthy normal eyes were evaluated. The frequency and pattern of each artifact was determined. Deviation of the segmentation line was classified into mild (less than 10 microns), moderate (10–50 microns) and severe (more than 50 microns). Each deviation, if present, was noted as upward or downward deviation. Each artifact was further described as per location on the scan and zones in the total scan area. A total of 1029 (26.8%) out of total 3840 scans had scan errors. The most common scan error was segmentation error (100%), followed by degraded images (6.70%), blink artifacts (0.09%) and out of register artifacts (3.3%). Misidentification of the inner retinal layers was most frequent (62%). Upward Deviation of the segmentation line (47.91%) and severe deviation (40.3%) were more often noted. Artifacts were mostly located in the central scan area (16.8%). The average number of scans with artifacts per eye was 34.3% and was not related to signal strength on Spearman correlation (p = 0.36). This study reveals that image artifacts and scan errors in SD-OCT GCA analysis are common and frequently involve segmentation errors. These errors may affect inner retinal thickness measurements in a clinically significant manner. Careful review of scans for artifacts is important when using this feature of SD-OCT device.

Background: Acanthamoeba is an environmental host for various microorganisms. Acanthamoeba is also becoming an increasingly important pathogen as a cause of keratitis. In Acanthamoeba keratitis (AK), coinfections involving pathogenic bacteria have been reported, potentially attributed to the carriage of microbes by Acanthamoeba. This study assessed the presence of intracellular bacteria in Acanthamoeba species recovered from domestic tap water and corneas of two different AK patients and examined the impact of naturally occurring intracellular bacteria within Acanthamoeba on the severity of corneal infections in rats. Methodology/Principal findings: Household water and corneal swabs were collected from AK patients. Acanthamoeba strains and genotypes were confirmed by sequencing. Acanthamoeba isolates were assessed for the presence of intracellular bacteria using sequencing, fluorescence in situ hybridization (FISH), and electron microscopy. The viability of the bacteria in Acanthamoeba was assessed by labelling with alkyne–functionalized D–alanine (alkDala). Primary human macrophages were used to compare the intracellular survival and replication of the endosymbiotic Pseudomonas aeruginosa and a wild type strain. Eyes of rats were challenged intrastromally with Acanthamoeba containing or devoid of P. aeruginosa and evaluated for the clinical response. Domestic water and corneal swabs were positive for Acanthamoeba. Both strains belonged to genotype T4F. One of the Acanthamoeba isolates harboured P. aeruginosa which was seen throughout the Acanthamoeba’s cytoplasm. It was metabolically active and could be seen undergoing binary fission. This motile strain was able to replicate in macrophage to a greater degree than strain PAO1 (p<0.05). Inoculation of Acanthamoeba containing the intracellular P. aeruginosa in rats eyes resulted in a severe keratitis with increased neutrophil response. Acanthamoeba alone induced milder keratitis. Conclusions/Significance: Our findings indicate the presence of live intracellular bacteria in Acanthamoeba can increase the severity of acute keratitis in vivo. As P. aeruginosa is a common cause of keratitis, this may indicate the potential for these intracellular bacteria in Acanthamoeba to lead to severe polymicrobial keratitis.

Hard exudates (HEs) are the classical sign of diabetic retinopathy (DR) which is one of the leading causes of blindness, especially in developing countries. Accordingly, disease screening involves examining HEs qualitatively using fundus camera. However, for monitoring the treatment response, quantification of HEs becomes crucial and hence clinicians now seek to measure the area of HEs in the digital colour fundus (CF) photographs. Against this backdrop, we proposed an algorithm to quantify HEs using CF images and compare with previously reported technique using ImageJ. CF photographs of 30 eyes (20 patients) with diabetic macular edema were obtained. A robust semi-automated algorithm was developed to quantify area covered by HEs. In particular, the proposed algorithm, a two pronged methodology, involved performing top-hat filtering, second order statistical filtering, and thresholding of the colour fundus images. Subsequently, two masked observers performed HEs measurements using previously reported ImageJ-based protocol and compared with those obtained through proposed method. Intra and inter-observer grading was performed for determining percentage area of HEs identified by the individual algorithm. Of the 30 subjects, 21 were males and 9 were females with a mean age of the 50.25 ± 7.80 years (range 33–66 years). The correlation between the two measurements of semi-automated and ImageJ were 0.99 and 0.99 respectively. Previously reported method detected only 0–30% of the HEs area in 9 images, 30–60% in 12 images and 60–90% in remaining images, and more than 90% in none. In contrast, proposed method, detected 60–90% of the HEs area in 13 images and 90–100% in remaining 17 images. Proposed method semi-automated algorithm achieved acceptable accuracy, qualitatively and quantitatively, on a heterogeneous dataset. Further, quantitative analysis performed based on intra- and inter-observer grading showed that proposed methodology detects HEs more accurately than previously reported ImageJ-based technique. In particular, we proposed algorithm detect faint HEs also as opposed to the earlier method.

Aim: To compare the outcomes of subthreshold microsecond (STM) and continuous-wave laser (CWL) panretinal photocoagulation (PRP). Methods: In this randomized, prospective, pilot study, 20 eyes of 10 subjects with symmetric severe non-proliferative (NPDR) or low-risk proliferative diabetic retinopathy (PDR) were included. Each eye of the subject was randomized into either CWL or STM PRP group. Patients were evaluated at baseline and at months 3, 6, and 9 with color fundus photographs and visual field tests at each visit; however, electroretinography (ERG) was conducted at baseline and at month 9. The primary outcome measure was the difference in disease progression between the groups. Secondary outcome measures included change in visual acuity, contrast visual acuity, retinal sensitivity on visual field test, and change in ERG parameters. Results: During the 9-month follow-up, one eye of the STM group progressed to vitreous hemorrhage at the month 6 follow-up and required rescue conventional laser. The CWL group showed a drop in low-contrast visual acuity, visual field index, and scotopic b/a ratio in comparison to the STM group, although the difference was statistically insignificant (p>0.05). Conclusion: This prospective pilot study proposes microsecond PRP is non-inferior to CWL PRP and could be an alternative to CWL PRP to avoid associated complications in cases of severe NPDR and early PDR.