Evaluation of In2Care mosquito stations for suppression of the Australian backyard mosquito, Aedes notoscriptus (Diptera: Culicidae).

Aedes notoscriptus (Skuse) is a container-inhabiting mosquito endemic to Australia that vectors arboviruses and is suspected to transmit Mycobacterium ulcerans, the cause of Buruli ulcer. We evaluated the effectiveness of the In2Care station, which suppresses mosquito populations via the entomopathogenic fungus, Beauveria bassiana, and the insect growth regulator pyriproxyfen, the latter of which is autodisseminated among larval habitats by contaminated mosquitoes. A field trial was conducted using 110 In2Care stations in a 50,000 m2 area and results were compared to 4 control areas that did not receive the treatment. Efficacy was evaluated by comparing egg counts and measuring larvicidal impact in surrounding breeding sites. Laboratory experiments validated the effect of B. bassiana on adult survival. Results of this field trial indicate that, 6 wk after the In2Care stations were deployed, treatment site ovitraps contained 43% fewer eggs than control site ovitraps, and 33% fewer eggs after 10 wk, suggesting that the In2Care station was able to reduce the egg density of Ae. notoscriptus. Population reduction remained evident for up to 3 wk after In2Care stations were removed. Treatment site ovitraps had significantly fewer Ae. notoscriptus eclosing than control site ovitraps, confirming the pyriproxyfen autodissemination feature of the stations. An average reduction of 50% in adult eclosion was achieved. Exposure to B. bassiana resulted in four-times higher mortality among adult mosquitoes. Additionally, using fresh In2Care nettings led to an 88% decrease in average survival compared to 4-wk-old nettings. The use of In2Care stations has potential for suppressing Ae. notoscriptus egg density.

A diversity of endosymbionts across Australian aphids and their persistence in aphid cultures.

There is increasing interest in the use of endosymbionts in pest control, which will benefit from the identification of endosymbionts from potential donor species for transfer to pest species. Here, we screened for endosymbionts in 123 Australian aphid samples across 32 species using 16S DNA metabarcoding. We then developed a qPCR method to validate the metabarcoding data set and to monitor endosymbiont persistence in aphid cultures. Pea aphids (Acyrthosiphon pisum) were frequently coinfected with Rickettsiella and Serratia, and glasshouse potato aphids (Aulacorthum solani) were coinfected with Regiella and Spiroplasma; other secondary endosymbionts detected in samples occurred by themselves. Hamiltonella, Rickettsia and Wolbachia were restricted to a single aphid species, whereas Regiella was found in multiple species. Rickettsiella, Hamiltonella and Serratia were stably maintained in laboratory cultures, although others were lost rapidly. The overall incidence of secondary endosymbionts in Australian samples tended to be lower than recorded from aphids overseas. These results indicate that aphid endosymbionts probably exhibit different levels of infectivity and vertical transmission efficiency across hosts, which may contribute to natural infection patterns. The rapid loss of some endosymbionts in cultures raises questions about factors that maintain them under field conditions, while endosymbionts that persisted in laboratory culture provide candidates for interspecific transfers.

Single-molecule force spectroscopy reveals binding and bridging dynamics of PARP1 and PARP2 at DNA double-strand breaks.

Poly(ADP-ribose) polymerases (PARPs) play key roles in DNA damage repair pathways in eukaryotic cells. Human PARPs 1 and 2 are catalytically activated by damage in the form of both double-strand and single-strand DNA breaks. Recent structural work indicates that PARP2 can also bridge two DNA double-strand breaks (DSBs), revealing a potential role in stabilizing broken DNA ends. In this paper, we have developed a magnetic tweezers-based assay in order to measure the mechanical stability and interaction kinetics of proteins bridging across the two ends of a DNA DSB. We find that PARP2 forms a remarkably stable mechanical link (rupture force ~85 pN) across blunt-end 5'-phosphorylated DSBs and restores torsional continuity allowing DNA supercoiling. We characterize the rupture force for different overhang types and show that PARP2 switches between bridging and end-binding modes depending on whether the break is blunt-ended or has a short 5' or 3' overhang. In contrast, PARP1 was not observed to form a bridging interaction across blunt or short overhang DSBs and competed away PARP2 bridge formation, indicating that it binds stably but without linking together the two broken DNA ends. Our work gives insights into the fundamental mechanisms of PARP1 and PARP2 interactions at double-strand DNA breaks and presents a unique experimental approach to studying DNA DSB repair pathways.

Direct Costs of Second Aqueous Shunt Implant Versus Transscleral Cyclophotocoagulation (The Assists Trial).

PRCIS: The cost of cyclophotocoagulation is less than the cost of a second glaucoma drainage device. PURPOSE: To compare the total direct costs of implantation of a second glaucoma drainage device (SGDD) with transscleral cyclophotocoagulation (CPC) for patients with inadequately controlled intraocular pressure (IOP) reduction, despite the presence of a preexisting glaucoma drainage device in the ASSISTS clinical trial. METHODS: We compared the total direct cost per patient, including the initial study procedure, medications, additional procedures, and clinic visits during the study period. The relative costs for each procedure during the 90-day global period and the entire study period were compared. The cost of the procedure, including facility fees and anesthesia costs, were determined using the 2021 Medicare fee schedule. Average wholesale prices for self-administered medications were obtained from AmerisourceBergen.com. The Wilcoxon rank sum test was used to compare costs between procedures. RESULTS: Forty-two eyes of 42 participants were randomized to SGDD (n=22) or CPC (n=20). One CPC eye was lost to follow-up after initial treatment and was excluded. The mean (±SD, median) duration of follow-up was 17.1 (±12.8, 11.7) months and 20.3 (±11.4, 15.1) months for SGDD and CPC, respectively ( P =0.42, 2 sample t test). The mean total direct costs (±SD, median) per patient during the study period were $8790 (±$3421, $6805 for the SGDD group) and $4090 (±$1424, $3566) for the CPC group ( P <0.001). Similarly, the global period cost was higher in the SGDD group than in the CPC group [$6173 (±$830, $5861) vs. $2569 (±$652, $2628); P <0.001]. The monthly cost after the 90-day global period was $215 (±$314, $100) for SGDD and $103 (±$74, $86) for CPC ( P =0.31). The cost of IOP-lowering medications was not significantly different between groups during the global period ( P =0.19) or after the global period ( P =0.23). CONCLUSION: The total direct cost in the SGDD group was more than double that in the CPC group, driven largely by the cost of the study procedure. The costs of IOP-lowering medications were not significantly different between groups. When considering treatment options for patients with a failed primary GDD, clinicians should be aware of differences in costs between these treatment strategies.

Use of urgent, emergency and acute care by mental health service users: A record-level cohort study.

BACKGROUND: People with serious mental illness experience worse physical health and greater mortality than the general population. Crude rates of A&E attendance and acute hospital admission are higher in people with serious mental illness than other hospital users. We aimed to further these findings by undertaking a standardised comparison of urgent and emergency care pathway use among users of mental health services and the general population. METHODS: Retrospective cohort analysis using routine data from 2013-2016 from the CUREd dataset for urgent and emergency care contacts (NHS 111, ambulance, A&E and acute admissions) and linked mental health trust data for Sheffield, England. We compared annual age- and sex-standardised usage rates for each urgent and emergency care service between users of mental health services and those without a recent history of mental health service use. RESULTS: We found marked differences in usage rates for all four urgent and emergency care services between the general population and users of mental health services. Usage rates and the proportion of users were 5-6 times and 3-4 times higher in users of mental health services, respectively, for all urgent and emergency care services. Users of mental health services were often more likely to experience the highest or lowest acuity usage characteristics. CONCLUSIONS: Current users of mental health services were heavily over-represented among urgent and emergency care users, and they made more contacts per-person. Higher service use among users of mental health services could be addressed by improved community care, more integrated physical and mental health support, and more proactive primary care. A complex pattern of service use among users of mental health services suggests this will need careful targeting to reduce avoidable contacts and optimise patient outcomes.

Outcomes of the Second Aqueous Shunt Implant Versus Transscleral Cyclophotocoagulation Treatment Study: A Randomized Comparative Trial.

PRCIS: Short-term overall success rates were high with either SGDD or CPC. However, SGDD was associated with more clinic visits and an increased risk of additional glaucoma surgery. Both treatments were reasonable options for eyes with inadequately controlled IOP after a single GDD. PURPOSE: The purpose of this study is to compare the implantation of a second glaucoma drainage device (SGDD) and transscleral cyclophotocoagulation (CPC) in eyes with inadequately controlled intraocular pressure (IOP), despite the presence of a preexisting glaucoma drainage device. METHODS: Patients with inadequately controlled IOP, despite the medical therapy and a preexisting glaucoma drainage device, were enrolled at 14 clinical centers and randomly assigned to treatment with a SGDD or CPC. MAIN OUTCOME MEASURES: Surgical failure was defined as: (1) IOP ≤5 mm Hg or >18 mm Hg or <20% reduction below baseline on maximum tolerated topical ocular hypotensive therapy, (2) reoperation for glaucoma, or (3) loss of light perception. The primary outcome measure was overall success with or without adjunctive medical therapy. RESULTS: Forty-two eyes of 42 participants were randomized to SGDD (n=22) or CPC (n=20). Mean duration of follow-up was 18.6 (±12.1; range: 1.1-38.6) months. The cumulative success rate was 79% for SGDD and 88% for CPC at 1 year ( P =0.63). Although the study was underpowered, no significant differences in IOP, postoperative number of IOP-lowering medications, or adverse events were observed. The number of additional glaucoma surgeries ( P =0.003), office visits during the first 3 months ( P <0.001), and office visits per month after month 3 ( P <0.001) were greater in the SGDD group. CONCLUSIONS: Short-term overall success rates were high with either SGDD or CPC. However, SGDD was associated with more clinic visits and an increased risk of additional glaucoma surgery.

Efficient golden gate assembly of DNA constructs for single molecule force spectroscopy and imaging.

Single-molecule techniques such as optical tweezers and fluorescence imaging are powerful tools for probing the biophysics of DNA and DNA-protein interactions. The application of these methods requires efficient approaches for creating designed DNA structures with labels for binding to a surface or microscopic beads. In this paper, we develop a simple and fast technique for making a diverse range of such DNA constructs by combining PCR amplicons and synthetic oligonucleotides using golden gate assembly rules. We demonstrate high yield fabrication of torsionally-constrained duplex DNA up to 10 kbp in length and a variety of DNA hairpin structures. We also show how tethering to a cross-linked antibody substrate significantly enhances measurement lifetime under high force. This rapid and adaptable fabrication method streamlines the assembly of DNA constructs for single molecule biophysics.

Preventive hoof trimming in dairy cattle: Determining current practices and identifying future research areas.

BACKGROUND: The extent to which preventive hoof trimming is implemented on Great Britain's (GB) dairy farms is unknown. The aims of this study were to determine common practices and capture producers' input on key areas that require further research. METHODS: An online survey was conducted over a 4-week period from March to April 2017. RESULTS: A total of 338 valid survey responses were received. The majority of farmers undertook preventive hoof trimming (82.4%), and this was significantly positively associated with increased herd yield (p < 0.001). Drying off was the most common time trimming was undertaken, with 72.2% of farmers who implemented preventive trimming doing so at this point in the management cycle. Of those undertaking preventive trimming, 46.4% solely used an external hoof trimmer, 31.7% solely used farm staff and the remainder (21.9%) used a combination of operators. Four over-arching themes were identified within the key research questions; when to trim, why we should trim, how to trim and who to trim. The most frequent questions related to optimal trimming timing, frequency and method. CONCLUSION: This study highlighted that preventive hoof trimming is a widespread practice on GB dairy farms, undertaken by both external hoof trimmers and farm staff. Despite this, farmers still want to know when they should undertake preventive hoof trimming and which technique they should use.

Associations of 12-year sleep behaviour trajectories from childhood to adolescence with myopia and ocular biometry during young adulthood.

PURPOSE: Cross-sectional studies have variably reported that poor sleep quality may be associated with myopia in children. Longitudinal data, collected over the ages when myopia develops and progresses, could provide new insights into the sleep-myopia paradigm. This study tested the hypothesis that 12-year trajectories of sleep behaviour from childhood to adolescence is associated with myopia during young adulthood. METHODS: At the 5-, 8-, 10-, 14- and 17-year follow-ups of the longitudinal Raine Study, which has been following a cohort since their birth in 1989-1992, participants' parents/guardians completed the Child Behaviour Checklist questionnaire (CBCL), which collected information on their child's sleep behaviour and quality. The CBCL includes six questions measuring sleep behaviour, which parents rated as 0 = not true, 1 = somewhat/sometimes true, or 2 = very/often true. Scores were summed at each follow-up to form a composite "sleep behaviour score". Latent Class Growth Analysis (LCGA) was used to classify participants according to their 12-year trajectory of sleep behaviour. At the 20-year follow-up, an eye examination was performed which included cycloplegic autorefraction and axial length measurement. RESULTS: The LCGA identified three clusters of participants based on their trajectory of sleep behaviour: those with minimal' (43.6% of the total Raine Study sample), 'declining' (48.9%), or 'persistent' (7.5%) sleep problems. A total of 1194 participants had ophthalmic data and longitudinal sleep data available for analysis (47.2% female, 85.6% Caucasian). No significant differences were observed in regards to age, sex, ethnicity or ocular parameters between trajectory groups. Unadjusted and fully adjusted analyses demonstrated that sleep problem behaviour was not significantly associated with changes in refractive error, axial length or corneal radius. CONCLUSIONS: Our findings do not support the hypothesis that there is an association between sleep behaviour and myopia. Future longitudinal studies should explore sleep trajectory data pre- and post-myopia diagnosis to confirm our results.

Using a learning health system to understand the mismatch between medicines supply and actual medicines use among adults with cystic fibrosis.

BACKGROUND: Studies in separate cohorts suggest possible discrepancies between inhaled medicines supplied (median 50-60%) and medicines used (median 30-40%). We performed the first study that directly compares CF medicine supply against use to identify the cost of excess medicines supply. METHODS: This cross-sectional study included participants from 12 UK adult centres with ≥1 year of continuous adherence data from data-logging nebulisers. Medicine supply was measured as medication possession ratio (MPR) for a 1-year period from the first suitable supply date. Medicine use was measured as electronic data capture (EDC) adherence over the same period. The cost of excess medicines was calculated as whole excess box(es) supplied after accounting for the discrepancy between EDC adherence and MPR with 20% contingency. RESULTS: Among 275 participants, 133 (48.4%) were females and mean age was 30 years (95% CI 29-31 years). Median EDC adherence was 57% (IQR 23-86%), median MPR was 74% (IQR 46-96%) and the discrepancy between measures was median 14% (IQR 2-29%). Even with 20% contingency, mean potential cost of excess medicines was £1,124 (95% CI £855-1,394), ranging from £183 (95% CI £29-338) for EDC adherence ≥80% to £2,017 (95% CI £1,507-2,526) for EDC adherence <50%. CONCLUSIONS: This study provides a conservative estimate of excess inhaled medicines supply cost among adults with CF in the UK. The excess supply cost was highest among those with lowest EDC adherence, highlighting the importance of adherence support and supplying medicine according to actual use. MPR provides information about medicine supply but over-estimates actual medicine use.

Single-molecule measurements reveal that PARP1 condenses DNA by loop stabilization.

Poly(ADP-ribose) polymerase 1 (PARP1) is an abundant nuclear enzyme that plays important roles in DNA repair, chromatin organization and transcription regulation. Although binding and activation of PARP1 by DNA damage sites has been extensively studied, little is known about how PARP1 binds to long stretches of undamaged DNA and how it could shape chromatin architecture. Here, using single-molecule techniques, we show that PARP1 binds and condenses undamaged, kilobase-length DNA subject to sub-piconewton mechanical forces. Stepwise decondensation at high force and DNA braiding experiments show that the condensation activity is due to the stabilization of DNA loops by PARP1. PARP inhibitors do not affect the level of condensation of undamaged DNA but act to block condensation reversal for damaged DNA in the presence of NAD+ Our findings suggest a mechanism for PARP1 in the organization of chromatin structure.

Microfluidic flow-cell with passive flow control for microscopy applications.

We present a fast, inexpensive and robust technique for constructing thin, optically transparent flow-cells with pump-free flow control. Using layers of glass, patterned adhesive tape and polydimethylsiloxane (PDMS) connections, we demonstrate the fabrication of planar devices with chamber height as low as 25 µm and with millimetre-scale (x,y) dimensions for wide-field microscope observation. The method relies on simple benchtop equipment and does not require microfabrication facilities, glass drilling or other workshop infrastructure. We also describe a gravity perfusion system that exploits the strong capillary action in the flow chamber as a passive limit-valve. Our approach allows simple sequential sample exchange with controlled flow rates, sub-5 µL sample chamber size and zero dead volume. We demonstrate the system in a single-molecule force spectroscopy experiment using magnetic tweezers.

Using Anterior Segment Optical Coherence Tomography (ASOCT) Parameters to Determine Pupillary Block Versus Plateau Iris Configuration.

PRéCIS:: The potential parameters for differentiating pupillary block (PB) from plateau iris configuration (PIC) on anterior segment optical coherence tomography (ASOCT) are lens/pupil size parameters and angles. Further study is needed to determine a landmark peripheral to the centroid of the iris. PURPOSE: Investigate anterior segment parameters to distinguish between 2 mechanisms of angle closure, PB and PIC, using swept-source Fourier domain ASOCT. PATIENTS AND METHODS: Retrospective ASOCT images from narrow angle eyes were reviewed. PIC was defined either by ultrasound biomicroscopy and/or clinically when an iridoplasty was performed. Images were read by a masked reader using Anterior Chamber Analysis and Interpretation software to identify scleral spur landmarks and calculate anterior chamber, peripheral angle, iris size, iris shape, and lens/pupil size parameters. ASOCT parameters were summarized and compared using the 2-sample t test. Thresholds and area under receiver operating characteristic curve were calculated using logistic regression analysis. RESULTS: One hundred eyes (66 PB and 34 PIC) of 100 participants were reviewed. Of all ASOCT parameters, iris length in each quadrant, pupil arc, lens/pupil parameters (pupil arc, lens vault, and pupil diameter), all pupillary margin-center point-scleral spur landmark (PM-C-SSL) parameters, and all except superior central iris vault parameters were significantly different between PB and PIC. On threshold evaluation, lens/pupil parameters had the greatest area under receiver operating characteristic curve values (0.77 to 0.80), followed by PM-C-SSL angles (0.71 to 0.75). CONCLUSIONS: We propose that the pupil size parameters and PM-C-SSL angle are the most reliable novel ASOCT parameters to distinguish between PB and PIC eyes. These parameters do not rely on the visibility of the posterior iris surface, which is difficult to identify with ASOCT, but may be ambient lighting dependent.

Origin of resistance to pyrethroids in the redlegged earth mite (Halotydeus destructor) in Australia: repeated local evolution and migration.

BACKGROUND: Halotydeus destructor is a major pest of crops and pastures across southern parts of Australia. This invasive mite has been chemically controlled for over 50 years, but resistance to synthetic pyrethroids and organophosphates is developing. Understanding processes behind the emerging resistance is important for effective management efforts. We undertook a ddRAD pool-sequencing approach to analyse genome-wide single nucleotide polymorphism variation in H. destructor population samples at two scales: local resistance across a set of fields, and regional resistance across their Australian range, along with toxicology bioassays to screen for pyrethroid resistance. RESULTS: Spatial patterns of genomic variation and resistance at a local scale indicated that genetic similarity among samples was more closely correlated with distance along roads and fence-lines than with straight-line geographic distance. This pattern was particularly strong in resistant samples, which were also more related than susceptible samples, suggesting local spread of resistance within an area after it emerged. By contrast, regional data suggest resistance has emerged repeatedly within parts of Australia. Our de novo annotation of the H. destructor draft genome sequence and Bayesian analysis identified several candidate loci strongly associated with population-level resistance to pyrethroids, located in genomic regions that code for transmembrane transport and signalling proteins that have previously been linked to insecticide resistance in other arthropods. CONCLUSION: Our findings highlight multiple independent evolutionary events leading to resistance in H. destructor, and demonstrate the utility and cost-effectiveness of a cross-population, genome-wide association study to reveal processes underlying adaptive evolution in a non-model invasive species. © 2019 Society of Chemical Industry.

A LAMP assay for the rapid and robust assessment of Wolbachia infection in Aedes aegypti under field and laboratory conditions.

With Wolbachia-based arbovirus control programs being scaled and operationalised around the world, cost effective and reliable detection of Wolbachia in field samples and laboratory stocks is essential for quality control. Here we validate a modified loop-mediated isothermal amplification (LAMP) assay for routine scoring of Wolbachia in mosquitoes from laboratory cultures and the field, applicable to any setting. We show that this assay is a rapid and robust method for highly sensitive and specific detection of wAlbB Wolbachia infection within Aedes aegypti under a variety of conditions. We test the quantitative nature of the assay by evaluating pooled mixtures of Wolbachia-infected and uninfected mosquitoes and show that it is capable of estimating infection frequencies, potentially circumventing the need to perform large-scale individual analysis for wAlbB infection status in the course of field monitoring. These results indicate that LAMP assays are useful for routine screening particularly under field conditions away from laboratory facilities.

PEGylated surfaces for the study of DNA-protein interactions by atomic force microscopy.

DNA-protein interactions are vital to cellular function, with key roles in the regulation of gene expression and genome maintenance. Atomic force microscopy (AFM) offers the ability to visualize DNA-protein interactions at nanometre resolution in near-physiological buffers, but it requires that the DNA be adhered to the surface of a solid substrate. This presents a problem when working in biologically relevant protein concentrations, where proteins may be present in large excess in solution; much of the biophysically relevant information can therefore be occluded by non-specific protein binding to the underlying substrate. Here we explore the use of PLLx-b-PEGy block copolymers to achieve selective adsorption of DNA on a mica surface for AFM studies. Through varying both the number of lysine and ethylene glycol residues in the block copolymers, we show selective adsorption of DNA on mica that is functionalized with a PLL10-b-PEG113/PLL1000-2000 mixture as viewed by AFM imaging in a solution containing high concentrations of streptavidin. We show - through the use of biotinylated DNA and streptavidin - that this selective adsorption extends to DNA-protein complexes and that DNA-bound streptavidin can be unambiguously distinguished in spite of an excess of unbound streptavidin in solution. Finally, we apply this to the nuclear enzyme PARP1, resolving the binding of individual PARP1 molecules to DNA by in-liquid AFM.

Latent Sensitization in a Mouse Model of Ocular Neuropathic Pain.

PURPOSE: Chronic ocular pain is poorly understood and difficult to manage. We developed a murine model of corneal surface injury (CSI)-induced chronic ocular neuropathic pain. The study focuses on changes in corneal nerve morphology and associated short- and long-term pain-like behavior after CSI. METHODS: CSI was induced in mice by local application of an alkali solution (0.75 N NaOH). Corneal nerve architecture, morphology, density, and length were studied. Eye-wiping was evaluated before and after CSI in response to hypertonic saline (2 M NaCl). Naltrexone (NTX) or Naloxone-methiodide (NLX-me), opioid receptor antagonists, were given subcutaneously (s.c., 3 mg/kg) or topically (eye drop, 100 µM), and then an eye-wiping test was performed. RESULTS: CSI caused partial corneal deinnervation followed by gradual reinnervation. Regenerated nerves displayed increased tortuosity, beading, and branching. CSI enhanced hypertonic saline-induced eye-wiping behavior compared to baseline or sham-injury (P < 0.01). This hypersensitivity peaked at 10 days and subsided 14 days after CSI. Administration of NTX, or NLX-me, a selective peripheral opioid antagonist, reinstated eye-wiping behavior in the injury group, but not in the sham groups (P < 0.05). CONCLUSIONS: This study introduces a model of chronic ocular pain and corneal neuropathy following CSI. CSI induces central and peripheral opioid receptor-dependent latent sensitization (LS) that is unmasked by systemic or topical administration of opioid antagonists. TRANSLATIONAL RELEVANCE: This model of chronic ocular pain establishes LS as a new inhibitory mechanism in the oculotrigeminal system and may be used for potential diagnostic and therapeutic interventions for ocular neuropathy.

The African Descent and Glaucoma Evaluation Study (ADAGES) III: Contribution of Genotype to Glaucoma Phenotype in African Americans: Study Design and Baseline Data.

PURPOSE: To describe the study protocol and baseline characteristics of the African Descent and Glaucoma Evaluation Study (ADAGES) III. DESIGN: Cross-sectional, case-control study. PARTICIPANTS: Three thousand two hundred sixty-six glaucoma patients and control participants without glaucoma of African or European descent were recruited from 5 study centers in different regions of the United States. METHODS: Individuals of African descent (AD) and European descent (ED) with primary open-angle glaucoma (POAG) and control participants completed a detailed demographic and medical history interview. Standardized height, weight, and blood pressure measurements were obtained. Saliva and blood samples to provide serum, plasma, DNA, and RNA were collected for standardized processing. Visual fields, stereoscopic disc photographs, and details of the ophthalmic examination were obtained and transferred to the University of California, San Diego, Data Coordinating Center for standardized processing and quality review. MAIN OUTCOME MEASURES: Participant gender, age, race, body mass index, blood pressure, history of smoking and alcohol use in POAG patients and control participants were described. Ophthalmic measures included intraocular pressure, visual field mean deviation, central corneal thickness, glaucoma medication use, or past glaucoma surgery. Ocular conditions, including diabetic retinopathy, age-related macular degeneration, and past cataract surgery, were recorded. RESULTS: The 3266 ADAGES III study participants in this report include 2146 AD POAG patients, 695 ED POAG patients, 198 AD control participants, and 227 ED control participants. The AD POAG patients and control participants were significantly younger (both, 67.4 years) than ED POAG patients and control participants (73.4 and 70.2 years, respectively). After adjusting for age, AD POAG patients had different phenotypic characteristics compared with ED POAG patients, including higher intraocular pressure, worse visual acuity and visual field mean deviation, and thinner corneas (all P < 0.001). Family history of glaucoma did not differ between AD and ED POAG patients. CONCLUSIONS: With its large sample size, extensive specimen collection, and deep phenotyping of AD and ED glaucoma patients and control participants from different regions in the United States, the ADAGES III genomics study will address gaps in our knowledge of the genetics of POAG in this high-risk population.

Genetic Architecture of Primary Open-Angle Glaucoma in Individuals of African Descent: The African Descent and Glaucoma Evaluation Study III.

PURPOSE: To find genetic contributions to glaucoma in African Americans. DESIGN: Cross-sectional, case-control study. PARTICIPANTS: One thousand eight hundred seventy-five primary open-angle glaucoma (POAG) patients and 1709 controls, self-identified as being of African descent (AD), from the African Descent and Glaucoma Evaluation Study (ADAGES) III and Wake Forest School of Medicine. METHODS: MegaChip genotypes were imputed to Thousand Genomes data. Association of single nucleotide polymorphisms (SNPs) with POAG and advanced POAG was tested by linear mixed model correcting for relatedness and population stratification. Genetic risk scores were tested by receiver operator characteristic curves (ROC-AUCs). MAIN OUTCOME MEASURES: Primary open-angle glaucoma defined by visual field loss without other nonocular conditions (n = 1875). Advanced POAG was defined by age-based mean deviation of visual field (n = 946). RESULTS: Eighteen million two hundred eighty-one thousand nine hundred twenty SNPs met imputation quality of r2 > 0.7 and minor allele frequency > 0.005. Association of a novel locus, EN04, was observed for advanced POAG (rs185815146 ß, 0.36; standard error, 0.065; P < 3×10-8). For POAG, an AD signal was observed at the 9p21 European descent (ED) POAG signal (rs79721419; P < 6.5×10-5) independent of the previously observed 9p21 ED signal (rs2383204; P < 2.3×10-5) by conditional analyses. An association with POAG in FNDC3B (rs111698934; P < 3.9×10-5) was observed, not in linkage disequilibrium (LD) with the previously reported ED SNP. Additional previously identified loci associated with POAG in persons of AD were: 8q22, AFAP1, and TMC01. An AUC of 0.62 was observed with an unweighted genetic risk score comprising 11 SNPs in candidate genes. Two additional risk scores were studied by using a penalized matrix decomposition with cross-validation; risk scores of 50 and 400 SNPs were identified with ROC of AUC = 0.74 and AUC = 0.94, respectively. CONCLUSIONS: A novel association with advanced POAG in the EN04 locus was identified putatively in persons of AD. In addition to this finding, this genome-wide association study in POAG patients of AD contributes to POAG genetics by identification of novel signals in prior loci (9p21), as well as advancing the fine mapping of regions because of shorter average LD (FNDC3B). Although not useful without confirmation and clinical trials, the use of genetic risk scores demonstrated that considerable AD-specific genetic information remains in these data.