Identification of Glaucoma in Diabetics Using the Laguna-ONhE Colourimetric Method and OCT Spectralis.

BACKGROUND: Previous retrospective results are evaluated prospectively and blinded. METHODS: A total of 221 eyes previously classified as normal (G1), 279 as moderate risk of glaucoma (G2) and 217 as high risk (G3) according to the Globin Discriminant Function (GDF) Laguna-ONhE index were examined with OCT Spectralis- Results: In G1, the Bruch's Membrane Opening Minimum Rim Width (BMO-MRW) was 332 ± 55 microns; in G2, it was 252 ± 47 (p < 0.0001); and in G3, 231 ± 44 (p < 0.0001). In G1, the 1% and 5% percentiles were 233 and 248, respectively; in G2, they were lower in 28.80% and 42.29% of cases, respectively; and in G3, in 50.23% and 63.59% of cases, respectively. Most of the cases were normal-tension glaucomas. Laguna-ONhE indices showed a curvilinear correlation with BMO-MRW results. The Retinal Nerve Fibre Layer (RNFL) showed a poor relationship with BMO. Assuming G1 to be truly normal, BMO-MRW would have a Receiver operating characteristic (ROC) curve area of 0.901 for G2 and G3 and 0.651 for RNFL. A significant reduction in pixels corresponding to vessels was found in G2 and G3 vs. G1 (p < 0.0001). CONCLUSIONS: In some cases, these defects appear to be mainly glaucomatous, and in others, they are associated with diabetic microangiopathy. In normal tension glaucoma, RNFL defects may be less severe than those inside the nerve.

Glaucoma Incidence and Progression in Diabetics: The Canary Islands Study Using the Laguna ONhE Application.

Background: Laguna ONhE provides a globin distribution function (GDF), in which a glaucoma discriminator based on deep learning plays an important role, and there is also an optimized globin individual pointer (GIP) for progression analysis. Methods: Signs of optic nerve glaucoma were identified in 1,124,885 fundus images from 203,115 diabetics obtained over 15 years and 117,813 control images. Results: A total of 743,696 images from 313,040 eyes of 173,661 diabetics were analysed. Some exclusions occurred due to excessive illumination, poor quality, or the absence of optic discs. Suspicion of glaucoma was reported in 6.70%, for an intended specificity of 99% (GDF < −15). More signs of glaucoma occur in diabetics as their years of disease increase, and after age 60, compared to controls. The GIP detected progression (p < 0.01) in 2.59% of cases with 4 controls and in 42.6% with 14 controls was higher in cases with lower GDF values. The GDF was corrected for the disc area and proved to be independent of it (r = 0.001925; p = 0.2814). Conclusions: The GDF index suggests a higher and increasing glaucoma probability in diabetics over time. Doubling the number of check-ups from four to eight increases the ability to detect GIP index progression by a factor of 5.

Optic disc area frequency distribution in a large sample of retinographic images.

OBJECTIVE: To describe a new method to estimate the frequency distribution of optic nerve disc area, using digital retinographic images. METHODS AND ANALYSIS: We analysed 492 023 fundus images obtained with seven fundus cameras, mainly in Caucasian subjects. They were grouped by resolution and zoom. They were automatically segmented by identifying the inner edge of the Elschnig scleral ring. For this purpose, a neural network trained by deep learning previously described was used. The number of pixels contained within the segmentation and their frequency distribution were calculated. The results of each camera, using different number of images, were compared with the global results using the Kolmogorov-Smirnov test to confront frequency distributions. RESULTS: The frequency distribution was non-Gaussian, more limited in small sizes than in large ones. If the median is assigned a theoretical value of 1.95 mm2, the 1th, 5th, 25th, 50th, 75th, 95th and 99th percentiles would correspond to 1.29, 1.46, 1.73, 1.95, 2.20, 2.64 and 3.03 mm2 in all the dataset. The overall differences were significant for the smaller series, but for each percentile their mean value was only 0.01 mm2 and the maximum 0.10 mm2, so they can be considered similar for practical purposes in all cameras. CONCLUSION: By automatically segmenting the edges of the optic nerve and observing the frequency distribution of the number of pixels it delimits, it is possible to estimate the frequency distribution of the disc area in the population as a whole and that of each individual case.

Comparison of age-related vascular changes in the optic disc of patients with diabetes, with glaucomatous and non-glaucomatous features.

OBJECTIVE: To identify age-related vascular changes in the optic discs of patients with diabetes with and without signs of glaucoma. METHODS AND ANALYSIS: A total of 2153 eyes of 1797 patients with diabetes without significant retinopathy were monitored with 10 Topcon-NW400 images obtained over 10.27±1.58 years. 571 non-diabetics eyes were selected as controls. Laguna ONhE uses convolutional neural networks to identify optic disc edges, vessels, cup and rim, and provides a glaucoma assessment index-Globin Distribution Function (GDF). RESULTS: In the first image, vessel pixels accounted for 33.88% of the disc area (SD=3.72) in non-glaucoma (DN) and 31.35% (SD=4.05; p<0.0001) in glaucoma cases (DG). This number of pixels was reduced by -0.55% each year (SD=0.77) in the DN and -0.76% (SD=0.86; p=0.0014) in the DG. In the first image, 76.55% of the disc pixels (SD=11.13) belonged to the rim in the DN and 62.05% (SD=11.00; p=0.0014) in the DG, decreasing annually by -0.33% (SD=0.99) in the DN and -0.68% (SD=1.08; p<0.00001) in the DG groups. All rim sectors were reduced over time in the DG group, particularly superotemporal (41°-80°) and inferotemporal (271°-310°). The reduction was smaller in DN, presenting as progressive thickening of the temporal sector (311°-40°). No changes in age were observed in healthy controls. CONCLUSION: Patients with diabetes show progressive reduction of vessels and neuroretinal rim at the optic disc, which is more intense in association with glaucoma. In the absence of glaucoma, the temporal sector of the diabetic rim was not reduced but thickened, displacing the cup nasally.

Fully Automated Colorimetric Analysis of the Optic Nerve Aided by Deep Learning and Its Association with Perimetry and OCT for the Study of Glaucoma.

BACKGROUND: Laguna-ONhE is an application for the colorimetric analysis of optic nerve images, which topographically assesses the cup and the presence of haemoglobin. Its latest version has been fully automated with five deep learning models. In this paper, perimetry in combination with Laguna-ONhE or Cirrus-OCT was evaluated. METHODS: The morphology and perfusion estimated by Laguna ONhE were compiled into a "Globin Distribution Function" (GDF). Visual field irregularity was measured with the usual pattern standard deviation (PSD) and the threshold coefficient of variation (TCV), which analyses its harmony without taking into account age-corrected values. In total, 477 normal eyes, 235 confirmed, and 98 suspected glaucoma cases were examined with Cirrus-OCT and different fundus cameras and perimeters. RESULTS: The best Receiver Operating Characteristic (ROC) analysis results for confirmed and suspected glaucoma were obtained with the combination of GDF and TCV (AUC: 0.995 and 0.935, respectively. Sensitivities: 94.5% and 45.9%, respectively, for 99% specificity). The best combination of OCT and perimetry was obtained with the vertical cup/disc ratio and PSD (AUC: 0.988 and 0.847, respectively. Sensitivities: 84.7% and 18.4%, respectively, for 99% specificity). CONCLUSION: Using Laguna ONhE, morphology, perfusion, and function can be mutually enhanced with the methods described for the purpose of glaucoma assessment, providing early sensitivity.

New visual field indices of disharmony for early diagnosis of glaucoma, alone or associated with conventional parameters.

PURPOSE: To evaluate the specificity of new perimetric indices based on harmony, alone and in combination with structural data, for glaucoma detection. METHODS: In this prospective observational cross-sectional study, one eye of 105 healthy subjects and 97 early and suspect glaucomas were sequentially included and examined with Cirrus optical coherence tomography, twice with OCULUS Smartfield perimeter (SPARK strategy) and twice with Humphrey Analyzer (24-2 SITA-Fast) at the Ophthalmology Department from the University Hospital La Candelaria. Disharmony in the visual field was evaluated including vertical threshold symmetry, threshold rank), and homogeneity (threshold standard deviation from its maximum) using the patient himself/herself as a reference. We also evaluated disharmony in combination with the mean deviation and the pattern standard deviation in a single index (mismatch) and various combinations of morphological and functional indices. Combinations used a new score based on values above certain critical cut-off levels of each index. RESULTS: For 95% specificity, the highest sensitivities were as follows: vertical cup/disc ratio: 28.9%; SPARK threshold rank: 29.9%; and SITA-Fast threshold standard deviation: 28.9%. For the combined indices and 100% specificity, they were 5 SPARK indices mismatch: 10.3%; 5 SITA-Fast indices mismatch: 11.3%; 8 optical coherence tomography indices: 21.9%; 13 SPARK and optical coherence tomography indices: 27.8%; and 13 SITA-Fast and optical coherence tomography indices: 32.0%. CONCLUSION: Disharmony combined with normative value-based indices and/or optical coherence tomography indices is useful for very specific early diagnosis of glaucoma.

Relationship between Retinal Nerve Fiber Layer Thickness and Hemoglobin Present in the Optic Nerve Head in Glaucoma.

PURPOSE: To observe the relationship between topographic hemoglobin levels in the optic nerve head (ONH), the rim thickness (BMO-MRW), and retinal nerve fiber layer (RNFL) thickness. METHODS: 96 normal eyes and 82 glaucomas were examined using TOP strategy (Octopus 300 perimeter), SPECTRALIS OCT, and Laguna ONhE program which estimates hemoglobin from conventional color photographs (Horus Scope DEC 200 fundus camera). RESULTS: The correlation between Laguna ONhE glaucoma discriminant function (GDF) and SPECTRALIS BMO-MRW was R = 0.81 (P < 0.0001), similar to that between the BMO-MRW and BMO-RNFL thicknesses (R = 0.85, P < 0.0001) (P = 0.227 between both R values). GDF correlated well with RNFL thicknesses in the 360 degrees around the nerve, similar to mean perimetric sensitivity (MS) and BMO-MRW. The amount of hemoglobin in the nasal and temporal sectors showed low correlation with superior and inferior RNFL thicknesses. The superotemporal and inferotemporal sectors located on the vertical diameter of the disk showed good intercorrelation but without a clear RNFL topographic relationship. CONCLUSION: GDF showed high correlation with RNFL thickness. Except in the nasal and temporal sectors, ONH hemoglobin correlated well with RNFL thickness.

Estimating the Amount of Hemoglobin in the Neuroretinal Rim Using Color Images and OCT.

PURPOSE: To calculate the amount of hemoglobin (Hb) in the optic nerve head (ONH), using superimposed color fundus images with disc, rim and cup boundaries obtained by OCT-Cirrus. MATERIAL AND METHODS: We examined 100 healthy and 121 glaucomatous eyes using Oculus-Spark perimetry, Cirrus-OCT and Visucam (Zeiss) ONH color images. The Laguna ONhE program was then used to calculate the amount of Hb in the cup and six sectors of the rim. Receiver operating characteristic (ROC) analysis was performed and correlations between parameters were calculated. RESULTS: In suspected and confirmed glaucoma, Hb was significantly lower than controls in all rim sectors, especially the inferior and superonasal (p < 0.0001). Mean deviation (MD) of visual field regions showed greater correlation with the amount of Hb in the superior and inferior sectors of the rim than with rim area (p = 0.02) or nerve fiber layer thickness (p < 0.0001). On ROC analysis, the best diagnostic indicators were OCT rim area, vertical cup/disc ratio (C/D) and Glaucoma Discriminant Function (GDF) of Laguna ONhE, without significant differences. CONCLUSIONS: The amount of Hb in the ONH seems to have an important relationship with glaucomatous visual field sensitivity. The remaining rim has insufficient perfusion in many cases of glaucoma.

Evaluation of Visual Field Progression in Glaucoma: Quasar Regression Program and Event Analysis.

PURPOSE: To determine the sensitivity, specificity and agreement between the Quasar program, glaucoma progression analysis (GPA II) event analysis and expert opinion in the detection of glaucomatous progression. MATERIALS AND METHODS: The Quasar program is based on linear regression analysis of both mean defect (MD) and pattern standard deviation (PSD). Each series of visual fields was evaluated by three methods; Quasar, GPA II and four experts. The sensitivity, specificity and agreement (kappa) for each method was calculated, using expert opinion as the reference standard. RESULTS: The study included 439 SITA Standard visual fields of 56 eyes of 42 patients, with a mean of 7.8 ± 0.8 visual fields per eye. When suspected cases of progression were considered stable, sensitivity and specificity of Quasar, GPA II and the experts were 86.6% and 70.7%, 26.6% and 95.1%, and 86.6% and 92.6% respectively. When suspected cases of progression were considered as progressing, sensitivity and specificity of Quasar, GPA II and the experts were 79.1% and 81.2%, 45.8% and 90.6%, and 85.4% and 90.6% respectively. The agreement between Quasar and GPA II when suspected cases were considered stable or progressing was 0.03 and 0.28 respectively. The degree of agreement between Quasar and the experts when suspected cases were considered stable or progressing was 0.472 and 0.507. The degree of agreement between GPA II and the experts when suspected cases were considered stable or progressing was 0.262 and 0.342. CONCLUSIONS: The combination of MD and PSD regression analysis in the Quasar program showed better agreement with the experts and higher sensitivity than GPA II.

Human Endogenous Retrovirus Type K (HERV-K) Particles Package and Transmit HERV-K-Related Sequences.

UNLABELLED: Human endogenous retroviruses (HERV) make up 8% of the human genome. While the youngest of these retroviruses, HERV-K(HML-2), termed HK2, is able to code for all viral proteins and produce virus-like particles, it is not known if these virus particles package and transmit HK2-related sequences. Here, we analyzed the capacity of HK2 for packaging and transmitting HK2 sequences. We created an HK2 probe, termed Bogota, which can be packaged into HK2 viruses, and transfected it into cells that make HK2 particles. Supernatants of the transfected cells, which contained HK2 viral particles, then were added to target cells, and the transmissibility of the HK2 Bogota reporter was tracked by G418 resistance. Our studies revealed that contemporary HK2 virions produced by some teratocarcinoma and breast cancer cell lines, as well as by peripheral blood lymphocytes from lymphoma patients, can package HK2 Bogota probes, and these viruses transmitted these probes to other cells. After transmission, HK2 Bogota transcripts undergo reverse transcription, a step impaired by antiretroviral agents or by introduction of mutations into the probe sequences required for reverse transcription. HK2 viruses were more efficiently transmitted in the presence of HK2 Rec or HIV-1 Tat and Vif. Transmitted Bogota probes formed episomes but did not integrate into the cellular genome. Resistance to integration might explain the relatively low number of HK2 insertions that were acquired during the last 25 million years of evolution. Whether transient transmission of modern HK2 sequences, which encode two putative oncoproteins, can lead to disease remains to be studied. IMPORTANCE: Retroviruses invaded the genome of human ancestors over the course of millions of years, yet these viruses generally have been inactivated during evolution, with only remnants of these infectious sequences remaining in the human genome. One of these viruses, termed HK2, still is capable of producing virus particles, although these particles have been regarded as being noninfectious. Using a genetic probe derived from HK2, we have discovered that HK2 viruses produced in modern humans can package HK2 sequences and transmit them to various other cells. Furthermore, the genetic sequences packaged in HK2 undergo reverse transcription. The transmitted probe circularized in the cell and failed to integrate into the cellular genome. These findings suggest that modern HK2 viruses can package viral RNA and transmit it to other cells. Contrary to previous views, we provide evidence of an extracellular viral phase of modern HK2 viruses. We have no evidence of sustained, spreading infection.

Glaucoma morphologic damage estimated from functional tests.

PURPOSE: To compare direct morphologic measurements with the ones deduced from functional tests. METHODS: A total of 111 normal eyes (examined twice) and 112 consecutive eyes with suspected or confirmed glaucoma were analyzed using Spark strategy (Oculus Easyfield Perimeter) and Cirrus optical coherence tomography (OCT). The threshold values were used to deduce the thickness of the retinal nerve fiber layer (RNFL) in 25 sectors. They were also used to derive a normalized value of the neuroretinal rim area, correcting the influence of the optic disc area size, and expressing the result as a percentage of their normal average relation. RESULTS: The correlation between the deduced and the measured RNFL thicknesses were r = 0.77 (p<0.001, standard error [SE] = 15.4 µm in normal and 15.1 µm in glaucomatous eyes) in the sector to sector comparison, and r = 0.80 (p<0.000001, SE = 10.0 µm in normal and 10.4 µm in glaucomatous eyes) for the average thickness. In normal subjects, the SE of deducing a second OCT result from the previous one was 13.2 µm in the sector by sector analysis and 4.6 µm for average thickness. The correlation between the deduced and measured rim area was r = 0.87 (p<0.000001, SE = 6.7% in normal and 15.9% in glaucomatous eyes). Receiver operating characteristic analysis yielded the following areas, sensitivities, and specificities: measured thickness: 0.89, 75.5%, 91.5%; deduced thickness: 0.91, 86.5%, 93.4%; measured rim area: 0.96, 89.2%, 95.3%; deduced rim area: 0.94, 82.9%, 96.2%. Additionally, the results in a new series of 71 normal and 68 glaucomatous eyes are verified. CONCLUSIONS: Spark perimetry allowed deducing glaucomatous morphologic alterations with significant accuracy.

Estimation of the relative amount of hemoglobin in the cup and neuroretinal rim using stereoscopic color fundus images.

PURPOSE: To calculate the relative amount of hemoglobin (Hb) in sectors of the optic nerve head (ONH) from stereoscopic color fundus images using the Laguna ONhE method and compare the results with the visual field evaluation and optical coherence tomography (OCT). METHODS: Healthy eyes (n = 87) and glaucoma eyes (n = 71) underwent reliable Oculus Spark perimetry and Cirrus OCT. Optical nerve head color images were acquired with a nonmydriatic stereoscopic Wx Kowa fundus camera. Laguna ONhE program was applied to these images to calculate the relative Hb amount in the cup and six sectors of the rim. Receiver operating characteristic (ROC) analysis and correlations between parameters were calculated. RESULTS: We did not observe any variations in the relative amount of Hb in relation to age in healthy subjects (R(2) = 0.033, P > 0.05). Maximum ROC area confidence intervals were observed for a combination between perimetric indices and the Laguna ONhE Glaucoma discriminant function (0.970-0.899) followed by rim area (0.960-0.883), and mean deviation (MD; 0.944-0.857). In glaucoma cases, relative Hb amount presented significant reduction in all rim sectors, especially 231° to 270° and 81° to 120° (P < 0.001), except in the temporal 311° to 40° (P = 0.11). Perimetry mean sensitivity by sectors was better correlated with respective Hb levels than with rim areas or the corresponding nerve fiber thickness, especially the superior and inferior sectors (P < 0.05). CONCLUSIONS: Visual field sensitivity was better correlated with Hb levels than with rim sector areas or the corresponding nerve fiber thickness. In many cases the remaining rim show low perfusion, especially in the superior and inferior sectors.

Small molecule deubiquitinase inhibitors promote macrophage anti-infective capacity.

The global spread of anti-microbial resistance requires urgent attention, and diverse alternative strategies have been suggested to address this public health concern. Host-directed immunomodulatory therapies represent one approach that could reduce selection for resistant bacterial strains. Recently, the small molecule deubiquitinase inhibitor WP1130 was reported as a potential anti-infective drug against important human food-borne pathogens, notably Listeria monocytogenes and noroviruses. Utilization of WP1130 itself is limited due to poor solubility, but given the potential of this new compound, we initiated an iterative rational design approach to synthesize new derivatives with increased solubility that retained anti-infective activity. Here, we test a small library of novel synthetic molecules based on the structure of the parent compound, WP1130, for anti-infective activity in vitro. Our studies identify a promising candidate, compound 9, which reduced intracellular growth of L. monocytogenes at concentrations that caused minimal cellular toxicity. Compound 9 itself had no bactericidal activity and only modestly slowed Listeria growth rate in liquid broth culture, suggesting that this drug acts as an anti-infective compound by modulating host-cell function. Moreover, this new compound also showed anti-infective activity against murine norovirus (MNV-1) and human norovirus, using the Norwalk virus replicon system. This small molecule inhibitor may provide a chemical platform for further development of therapeutic deubiquitinase inhibitors with broad-spectrum anti-infective activity.

Genomic flexibility of human endogenous retrovirus type K.

UNLABELLED: Human endogenous retrovirus type K (HERV-K) proviruses are scattered throughout the human genome, but as no infectious HERV-K virus has been detected to date, the mechanism by which these viruses replicated and populated the genome remains unresolved. Here, we provide evidence that, in addition to the RNA genomes that canonical retroviruses package, modern HERV-K viruses can contain reverse-transcribed DNA (RT-DNA) genomes. Indeed, reverse transcription of genomic HERV-K RNA into the DNA form is able to occur in three distinct times and locations: (i) in the virus-producing cell prior to viral release, yielding a DNA-containing extracellular virus particle similar to the spumaviruses; (ii) within the extracellular virus particle itself, transitioning from an RNA-containing particle to a DNA-containing particle; and (iii) after entry of the RNA-containing virus into the target cell, similar to canonical retroviruses, such as murine leukemia virus and HIV. Moreover, using a resuscitated HERV-K virus construct, we show that both viruses with RNA genomes and viruses with DNA genomes are capable of infecting target cells. This high level of genomic flexibility historically could have permitted these viruses to replicate in various host cell environments, potentially assisting in their many integration events and resulting in their high prevalence in the human genome. Moreover, the ability of modern HERV-K viruses to proceed through reverse transcription and package RT-DNA genomes suggests a higher level of replication competency than was previously understood, and it may be relevant in HERV-K-associated human diseases. IMPORTANCE: Retroviral elements comprise at least 8% of the human genome. Of all the endogenous retroviruses, HERV-K viruses are the most intact and biologically active. While a modern infectious HERV-K has yet to be found, HERV-K activation has been associated with cancers, autoimmune diseases, and HIV-1 infection. Thus, determining how this virus family became such a prevalent member of our genome and what it is capable of in its current form are of the utmost importance. Here, we provide evidence that HERV-K viruses currently found in the human genome are able to proceed through reverse transcription and historically utilized a life cycle with a surprising degree of genomic flexibility in which both RNA- and DNA-containing viruses were capable of mediating infection.

Murine norovirus: propagation, quantification, and genetic manipulation.

Murine norovirus (MNV) is a positive-sense, plus-stranded RNA virus in the Caliciviridae family. It is the most common pathogen in biomedical research colonies. MNV is also related to the human noroviruses, which cause the majority of nonbacterial gastroenteritis worldwide. Like the human noroviruses, MNV is an enteric virus that replicates in the intestine and is transmitted by the fecal-oral route. MNV replicates in murine macrophages and dendritic cells in cells in culture and in the murine host. This virus is often used to study mechanisms in norovirus biology, because human noroviruses are refractory to growth in cell culture. MNV combines the availability of a cell culture and reverse genetics system with the ability to study infection in the native host. Herein, we describe a panel of techniques that are commonly used to study MNV biology.

Regulation of the human endogenous retrovirus K (HML-2) transcriptome by the HIV-1 Tat protein.

UNLABELLED: Approximately 8% of the human genome is made up of endogenous retroviral sequences. As the HIV-1 Tat protein activates the overall expression of the human endogenous retrovirus type K (HERV-K) (HML-2), we used next-generation sequencing to determine which of the 91 currently annotated HERV-K (HML-2) proviruses are regulated by Tat. Transcriptome sequencing of total RNA isolated from Tat- and vehicle-treated peripheral blood lymphocytes from a healthy donor showed that Tat significantly activates expression of 26 unique HERV-K (HML-2) proviruses, silences 12, and does not significantly alter the expression of the remaining proviruses. Quantitative reverse transcription-PCR validation of the sequencing data was performed on Tat-treated PBLs of seven donors using provirus-specific primers and corroborated the results with a substantial degree of quantitative similarity. IMPORTANCE: The expression of HERV-K (HML-2) is tightly regulated but becomes markedly increased following infection with HIV-1, in part due to the HIV-1 Tat protein. The findings reported here demonstrate the complexity of the genome-wide regulation of HERV-K (HML-2) expression by Tat. This work also demonstrates that although HERV-K (HML-2) proviruses in the human genome are highly similar in terms of DNA sequence, modulation of the expression of specific proviruses in a given biological situation can be ascertained using next-generation sequencing and bioinformatics analysis.

Chemical derivatives of a small molecule deubiquitinase inhibitor have antiviral activity against several RNA viruses.

Most antiviral treatment options target the invading pathogen and unavoidably encounter loss of efficacy as the pathogen mutates to overcome replication restrictions. A good strategy for circumventing drug resistance, or for pathogens without treatment options, is to target host cell proteins that are utilized by viruses during infection. The small molecule WP1130 is a selective deubiquitinase inhibitor shown previously to successfully reduce replication of noroviruses and some other RNA viruses. In this study, we screened a library of 31 small molecule derivatives of WP1130 to identify compounds that retained the broad-spectrum antiviral activity of the parent compound in vitro but exhibited improved drug-like properties, particularly increased aqueous solubility. Seventeen compounds significantly reduced murine norovirus infection in murine macrophage RAW 264.7 cells, with four causing decreases in viral titers that were similar or slightly better than WP1130 (1.9 to 2.6 log scale). Antiviral activity was observed following pre-treatment and up to 1 hour postinfection in RAW 264.7 cells as well as in primary bone marrow-derived macrophages. Treatment of the human norovirus replicon system cell line with the same four compounds also decreased levels of Norwalk virus RNA. No significant cytotoxicity was observed at the working concentration of 5 µM for all compounds tested. In addition, the WP1130 derivatives maintained their broad-spectrum antiviral activity against other RNA viruses, Sindbis virus, LaCrosse virus, encephalomyocarditis virus, and Tulane virus. Thus, altering structural characteristics of WP1130 can maintain effective broad-spectrum antiviral activity while increasing aqueous solubility.

Measuring hemoglobin levels in the optic disc of Parkinson's disease patients using new colorimetric analysis software.

Objective. To evaluate a new method of measuring hemoglobin (Hb) levels and quantifying the color changes in the optic nerve head of Parkinson's disease (PD) patients. We also compared differences in retinal nerve fiber layer (RNFL) thicknesses obtained using spectral domain optical coherence tomography (OCT) device between PD group and healthy group. Methods. One hundred and fifty-five PD patients and 91 sex- and age-matched healthy subjects were included in this cross-sectional study. OCT examinations and one photograph of the optic disc were performed. The Laguna ONhE ("optic nerve hemoglobin"; Insoft SL, Tenerife, Spain) software was used to analyze the Hb level on the acquired optic disc photographs. Results. PD patients exhibited significantly reduced mean optic disc Hb percentages (57.56% in PD, 67.63% in healthy subjects; P = 0.001) as well as reduced Hb in almost all analyzed sectors, with the largest differences detected in the inferior and nasal sectors. RNFL parameters were significantly reduced in PD patients compared with healthy subjects, especially in the inferior quadrant. Conclusions. Measurements of optic disc Hb levels obtained with the Laguna ONhE software had good ability to detect optic nerve color changes (more papillary paleness and consequently this could suggest optic atrophy and axonal loss) in PD patients.

HIV infection reveals widespread expansion of novel centromeric human endogenous retroviruses.

Human endogenous retroviruses (HERVs) make up 8% of the human genome. The HERV-K (HML-2) family is the most recent group of these viruses to have inserted into the genome, and we have detected the activation of HERV-K (HML-2) proviruses in the blood of patients with HIV-1 infection. We report that HIV-1 infection activates expression of a novel HERV-K (HML-2) provirus, termed K111, present in multiple copies in the centromeres of chromosomes throughout the human genome yet not annotated in the most recent human genome assembly. Infection with HIV-1 or stimulation with the HIV-1 Tat protein leads to the activation of K111 proviruses. K111 is present as a single copy in the genome of the chimpanzee, yet K111 is not found in the genomes of other primates. Remarkably, K111 proviruses appear in the genomes of the extinct Neanderthal and Denisovan, while modern humans have at least 100 K111 proviruses spread across the centromeres of 15 chromosomes. Our studies suggest that the progenitor K111 integrated before the Homo-Pan divergence and expanded in copy number during the evolution of hominins, perhaps by recombination. The expansion of K111 provides sequence evidence suggesting that recombination between the centromeres of various chromosomes took place during the evolution of humans. K111 proviruses show significant sequence variations in each individual centromere, which may serve as markers in future efforts to annotate human centromere sequences. Further, this work is an example of the potential to discover previously unknown genomic sequences through the analysis of nucleic acids found in the blood of patients.