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Purpose: To demonstrate that high-seed, ultra-high-resolution spectral-domain optical coherence tomography (SD-OCT) technology can image in vivo fine morphological features in the healthy and pathological human limbus., Methods: A compact, fiberoptic SD-OCT system was developed for imaging the human limbus. It combines ∼1.5-µm isotropic spatial resolution in ocular tissue and an acquisition rate of 250,000 A-scans per second. The imaging probe was outfitted with two microscope objectives to provide flexibility in the choice of wide field of view and extended depth of focus versus high lateral resolution. The clinical potential of the system was evaluated by imaging subjects with limbal stem cell dysfunction (LSCD; n = 4) and healthy controls (n = 6)., Results: Limbus images acquired from the healthy controls showed normal cellular structure of the limbal crypts, palisades of Vogt (POVs), and vasculature of the underlying scleral tissue. Images acquired from the LSCD subjects showed distortions or absence of POVs, invasion of highly scattering conjunctival tissue over the limbal and peripheral corneal epithelium, scarring and thinning of the limbal epithelium, and neovascularization., Conclusions: The combination of high OCT spatial resolution and rapid image acquisition rate allows for in vivo, contactless, volumetric visualization of fine morphological details that could be beneficial for the precise diagnosis and grading of LSCD, planning of treatment, and evaluation of the effectiveness of the treatment approaches., Translational Relevance: The OCT technology described here could improve the clinical diagnostics and grading of LSCD, preoperative planning, and postoperative evaluation of LSCD subjects, in addition to monitoring the effectiveness of various LSCD treatments.

Mycolicibacterium hassiacum (homotypic synonym: Mycobacterium hassiacum ) represents an ungrouped thermotolerant rapidly growing mycobacteria (RGM) species occasionally associated with infections and disease in humans. In this report, we describe a case of pyogranulomatous dermatitis and panniculitis due to M. hassiacum in an immunocompetent adult cat. To the best of our knowledge, this represents the first report of M. hassiacum infection in animals. We also report the results of the in-depth genome characterization of the isolate using a combined short- and long-read whole-genome sequencing (WGS) approach. We observed the lack of acquired-resistance genes and no evidence of mutations in housekeeping genes associated with resistance to rifampicin and isoniazid. We detected some virulence factors in our isolate, such as some associated with the interaction of mycobacteria with host cells, and the presence of multiple copies of heavy metal resistance genes ( ars B, ars R, and ars L/ cad L). In conclusion, M. hassiacum should be included among the RGM species associated with feline subcutaneous atypical mycobacteriosis (SAM). A reliable and fast RGM laboratory identification and characterization is important not only for an accurate etiological diagnosis but also for a correct approach to SAM treatment options.

Tuberculosis (TB) affects humans and other animals, and it is caused by bacteria within the Mycobacterium tuberculosis complex (MTBC). In this study, we report the characterisation of Mycobacterium pinnipedii that caused a TB case in a sea lion ( Otaria flavescens ) kept in an Italian zoo. The animal died due to severe, progressive disorders involving the respiratory and gastro-enteric systems and the skin. At necropsy, typical gross lesions referable to a TB generalised form were found. In particular, nodular granulomatous lesions were detected in the lungs and several lymph nodes, and colonies referable to Mycobacterium spp. were isolated from lung, mesenteric, and mediastinal lymph nodes. The isolate was identified by PCR as a MTBC, had a spoligotype SB 1480 ("seal lineage"), and was characterised and characterised by whole-genome sequencing analysis confirming that the MTBC involved was M. pinnipedii . The analysis of the resistome and virulome indicated the presence of macrolide and aminoglycoside resistance genes intrinsic in M. tuberculosis [ erm-37 and aac (2')-Ic] and confirmed the presence of the region of difference 1 (RD1), harbouring the esx A and esx B virulence genes, differently from its closest taxon, M. microti . As for other MTCB members, M. pinnipedii infection can spill over into non-pinniped mammalian species; therefore, zoological gardens, veterinary practitioners, and public health officers should be aware of the hazard posed by tuberculosis from marine mammals. Since the isolate under study, as well as all available genomes of M. pinnipedii investigated in this study retains almost all the M. tuberculosis virulence genes, it could indeed cause infection, lesions, and disease in other animal species, including humans., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2023 Alba, Caprioli, Cocumelli, Eleni, Diaconu, Donati, Ianzano, Sorbara, Stravino, Cerini, Boniotti, Zanoni, Franco and Battisti.)

Purpose: To investigate changes in the corneal volume, corneal densitometry and pachymetry of young myopes wearing over-night orthokeratology (OK) contact lenses., Methods: The medical records of 28 right eyes of young myopes wearing OK between 2013 and 2018 were reviewed retrospectively. The baseline refractive error, best corrected visual acuities at baseline and uncorrected visual acuity at the most recent visit were recorded. Corneal volume of the central 10 mm cornea; densitometry at central, nasal and temporal cornea; and pachymetry along the horizontal and vertical meridians were collected from the Pentacam® HR at baseline, after one night of lens wear, and at the latest visit., Results: The mean age of subjects was 12.03 ± 3.80 years at the time of OK lens fitting and wore OK overnight for a mean duration of 666 days (range 206-1736 days). The baseline spherical equivalent refractive error was -3.03 ± 1.56 D (range -1.00 to -6.00 D). The corneal volume increased significantly after OK wear (p = 0.001). Corneal densitometry increased after OK wear, but the change did not reach statistical significance (p = 0.113). Pachymetry in the central cornea did not change significantly across all visits (p > 0.05) but increased significantly in the mid-peripheral regions of the cornea. Baseline refractive error was not found to be correlated with the changes in corneal volume, corneal densitometry, or pachymetry., Conclusion: The increase in corneal volume and densitometry and no significant change in the central corneal thickness may indicate the presence of corneal oedema from long-term OK wear. The baseline refraction was not correlated with the changes in corneal volume, densitometry or pachymetry., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Ltd.)

The increasing prevalence of pESI(like)-positive, multidrug-resistant (MDR) S. Infantis in Europe is a cause of major concern. As previously demonstrated, the pESI(like) megaplasmid is not only a carrier of antimicrobial resistant (AMR) genes (at least tet, dfr, and sul genes), but also harbours several virulence and fitness genes, and toxin/antitoxin systems that enhance its persistence in the S. Infantis host. In this study, five prototype pESI(like) plasmids, of either CTX-M-1 or CTX-M-65 ESBL-producing strains, were long-read sequenced using Oxford Nanopore Technology (ONT), and their complete sequences were resolved. Comparison of the structure and gene content of the five sequenced plasmids, and further comparison with previously published pESI(like) sequences, indicated that although the sequence of such pESI(like) 'mosaic' plasmids remains almost identical, their structures appear different and composed of regions inserted or transposed after different events. The results obtained in this study are essential to better understand the plasticity and the evolution of the pESI(like) megaplasmid, and therefore to better address risk management options and policy decisions to fight against AMR and MDR in Salmonella and other food-borne pathogens. Graphical representation of the pESI-like plasmid complete sequence (ID 12037823/11). Block colours indicate the function of the genes: red: repB gene; pink: class I integrons (IntI); yellow; mobile elements; blue: resistance genes; green: toxin/anti-toxin systems; grey: mer operon; light green: genes involve in conjugation., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

Carbapenemase-producing Enterobacterales (CPE) are considered a major public health issue. In the frame of the EU Harmonized AMR Monitoring program conducted in Italy in 2021, 21 epidemiological units of fattening pigs (6.98%; 95% CI 4.37-10.47%; 21/301) and four epidemiological units of bovines <12 months (1.29%; 95% CI 0.35-3.27%, 4/310) resulted positive to OXA-48-like-producing E. coli ( n  = 24 OXA-181, n  = 1 OXA-48). Whole Genome Sequencing (WGS) for in-depth characterization, genomics and cluster analysis of OXA-181-(and one OXA-48) producing E. coli isolated, was performed. Tracing-back activities at: (a) the fattening holding of origin of one positive slaughter batch, (b) the breeding holding, and (c) one epidemiologically related dairy cattle holding, allowed detection of OXA-48-like-producing E. coli in different units and comparison of further human isolates from fecal samples of farm workers. The OXA-181-producing isolates were multidrug resistant (MDR), belonged to different Sequence Types (STs), harbored the IncX and IncF plasmid replicons and multiple virulence genes. Bioinformatics analysis of combined Oxford Nanopore Technologies (ONT) long reads and Illumina short reads identified bla OXA-181 as part of a transposon in IncX1, IncX3, and IncFII fully resolved plasmids from 16 selected E. coli, mostly belonging to ST5229, isolated during the survey at slaughter and tracing-back activities. Although human source could be the most likely cause for the introduction of the bla OXA-181 -carrying IncX1 plasmid in the breeding holding, concerns arise from carbapenemase OXA-48-like-producing E. coli spreading in 2021 in Italian fattening pigs and, to a lesser extent, in veal calf holdings., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Carfora, Diaconu, Ianzano, Di Matteo, Amoruso, Dell'Aira, Sorbara, Bottoni, Guarneri, Campana, Franco, Alba and Battisti.)

Purpose: To describe the development and report psychometric properties of the Contact Lens Risk Survey (CLRS) to identify patients at risk for soft contact lens-related complications., Methods: Psychometric properties of the CLRS, a web-based survey with branching logic, were determined using data from 5 multi-site fieldings (n = 1059), including re-fielding to sub groups. Responses from participants with and without an active red eye were used to identify risk factors of a red eye event and calculate an overall risk score. A 6th fielding of the CLRS (n = 171) was used to assess discriminate validity., Results: Participants needed 11-12 min to complete the survey with a negligible difference by age. Internal consistency was excellent (Cronbach's α ≥ 0.70) for 3 of the 5 constructs identified by factor analysis. Twelve of the 17 survey items exhibited excellent within-subject repeatability (Kappa ≥ 0.61). Between-subject agreement, assessed in 18-25 year olds, was high for the majority of items, suggesting good generalizability across different populations of SCL wearers. The ability of the model using individual items of the CLRS to discriminate Controls and participants with a red eye was good with an area under the curve of 0.779., Conclusion: The CLRS tool is a repeatable and valid instrument to standardize documentation of demographic, behavior, and exposure factors which appear to drive the greatest risk of a contact lens related red eye event., (Copyright © 2020 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.)

Purpose: To evaluate the agreement of central corneal thickness (CCT) measurement between Scheimpflug imaging and ultrasound (U/S) pachymetry in keratoconic eyes, and investigate factors that affect the agreement., Methods: This post hoc analysis within a prospective, observational non-randomised study preformed at the Kensington Eye Institute, Toronto, Ontario, Canada, included crosslinking candidates with progressive keratoconus (KC). Main outcome measures were the agreement of CCT measurement between Scheimpflug imaging (Pentacam Oculus, Wetzlar, Germany) and U/S pachymetry (PachPen 24-5100, Accutome Inc., Malvern, Pennsylvania, USA), and factors that affect the agreement., Results: A total of 794 keratoconic eyes of 456 subjects with a mean age of 27.6±8.0 years (69.7% males and 49.6% right eyes) were included. Agreement between devices was moderate (intraclass correlation coefficient: 74.9%, Bland-Altman limits of agreement: -48.5 μm to +62.5 μm). In a multivariable analysis, cone decentration (p<0.001, coefficient +10.13 [+6.73 to +13.53 95% CIs]) and Kmax (p<0.001, coefficient +0.68 [+0.46 to +0.90 95% CIs]) were significantly associated (both clinically and statistically) with the level of agreement between the devices; the discrepancy in CCT between the devices increased on average by 10.13 μm for every mm of cone decentration, and by 6.8 μm for every 10D of Kmax. Age, corneal astigmatism and spherical equivalent were statistically but not clinically significant factors affecting agreement., Conclusion: The agreement of CCT measurement between Scheimpflug imaging and U/S pachymetry in KC was moderate. To ensure the safety of crosslinking in keratoconic corneas, Scheimpflug and U/S CCT measurements should not be used interchangeably, especially in steep corneas and corneas with decentred cones., Competing Interests: Competing interests: MM is a paid consultant for Eye Yon Medical and Lapidot Medical. CC has received support from Alcon Laboratories Inc, Allergan, Bausch & Lomb, Santen, Shire, Tearlab, Labtician Thea, and Johnson & Johnson. AS has been a paid consultant for Abbott Medical Optics, AbbVie, Alcon Laboratories Inc, Allergan, Bausch & Lomb, Santen Inc, Shire, Acqueous, and Labtician Thea. DR has received support from Alcon Laboratories Inc and Johnson & Johnson. TR has been a paid consultant for Alcon Laboratories Inc, Johnson & Johnson, and AbbVie., (© Author(s) (or their employer(s)) 2021. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: To test the Contact Lens Assessment in Youth Contact Lens Risk Survey (CLRS) scoring algorithm in soft contact lens (SCL) wearers presenting with SCL-related adverse events and healthy matched controls., Methods: This prospective case-control study compared CLRS responses in SCL wearers presenting with symptomatic red eyes (Cases) with age-, sex- and site-matched healthy SCL wearers (Controls) at six locations across North America. Responses to individual questions from Cases and Controls were analyzed using either the Pearson, Mantel-Haenszel chi-square, or Fisher's exact test. Differences in the CLRS scores were compared using t-tests., Results: A total of 171 SCL wearers were enrolled (n = 57 Cases, 114 Controls). Cases were adjudicated to consensus and classified as contact lens-related serious and significant (S&S) or non-serious events. S&S Cases scored significantly higher on the CLRS (56.1 ± 11.1) than either their matched Controls (44.3 ± 11.1) (P <  0.001) or the non-Serious Cases (44.8± 12.8 P =  0.002). Scores of non-serious Cases were not significantly different than scores of their Controls (43.4 ± 10.7, P = 0.33) CONCLUSIONS: The CLRS scoring algorithm discriminated between SCL wearers presenting with S&S CL-related adverse events from healthy SCL wearers and those with non-serious events. This survey and scoring system could help practitioners identify patients at greatest risk for CL-related adverse events and support targeted interventions aimed at reducing risk behaviors., (Copyright © 2020 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.)

Significance: Cytokine and protease analysis revealed relative changes in the post-lens tear film of scleral lenses with low and high limbal clearances. Results from this study indicate that midperipheral lens fit is an important fitting feature that can impact the inflammatory response of a keratoconic eye., Purpose: The purpose of this study was to investigate changes in levels of inflammatory mediators in the post-lens tear film of keratoconic scleral lens wearers with varying limbal clearance designs., Methods: Twenty-two keratoconic eyes were fitted with two sets of scleral lenses that were consistent in lens diameter and central sagittal depth but varied in limbal clearance by approximately 50 μm. Lenses were worn in a randomly assigned order for a 2-week period each. At each follow-up visit, immediately after lens removal, tear samples were collected with a microcapillary tube (10 μL, 0.5 mm in diameter) from the bowl of the inverted scleral lens. Tear cytokine and protease analysis was performed using a multiplex electrochemiluminescent array (Meso Scale Discovery, Rockville, MD) instrument. Levels of interleukins 1, 6, and 8; tumor necrosis factor α; and matrix metalloproteinases 1 and 9 were compared and analyzed., Results: Levels of interleukin 1β, tumor necrosis factor α, and matrix metalloproteinase 1 increased with high limbal clearance (P = .01, .006, and .02, respectively). No change in interleukins 6 and 8 levels was found (P > .05). A decrease in matrix metalloproteinase 9 was noted in post-lens tear film of scleral lenses with high limbal clearance (P = .10)., Discussion: Relative changes in the cytokine and protease levels were found when comparing low and high limbal clearance, indicating that the midperipheral lens fit is an important feature that can impact the inflammatory response of the keratoconic eye., Competing Interests: Conflict of Interest Disclosure: None of the authors have reported a financial conflict of interest., (Copyright © 2021 American Academy of Optometry.)

Purpose: The purpose of this study was to characterize the central epithelial thickness (CET) of penetrating keratoplasty corneal specimens obtained from patients with keratoconus (KC) and correlate the histological patterns with their clinical history., Methods: Ex vivo histological imaging was performed to measure CET and total corneal thickness (TCT) in 56 patients with KC. Microscopic slides from penetrating keratoplasty corneal specimens, stained with hematoxylin and eosin were evaluated using bright field microscopy. CET and TCT were measured, and morphological features were studied. Clinical history regarding duration of KC prior to surgery and length of and tolerance to contact lens wear were compared and analyzed., Results: The microscopic slides of all patients available for follow up (n=48) were analyzed and CET and TCT were measured. The histological evaluation revealed 3 distinctive epithelial patterns. Pattern 1 with central hypertrophic and hydropic changes (n=19) measured 70.89±25.88μm in CET and 308.63±100.74μm in TCT; Pattern 2 (n=14) had not changed, similar to normal epithelium CET and TCT measuring 36.5±7.02μm and 260.14±87.93μm respectively. Pattern 3 (n=15) demonstrated thinner central epithelium characterized by atrophy and focal hydropic changes measuring 19.93±4.60μm and 268.00±79.39μm in CET and TCT respectively (all p<0.0001). The presence of Pattern 2 characterized by similar to normal CET was correlated with the duration of the condition (R=0.600, p=0.030). There was a significant difference in the length of CL wear comparing those with patterns 1 and 2 versus 3 (least no. of CL years) (p=0.05 and p=0.33 respectivelly)., Conclusions: Patients with advanced disease have various central corneal epithelial changes detected with histology. Although each central epithelial pattern type was distinctive comparing the 3 patterns, there was no correlation with years of CL wear but only with the duration of the condition., (Copyright © 2020 Spanish General Council of Optometry. Published by Elsevier España, S.L.U. All rights reserved.)

Objectives: To the best of our knowledge, we describe the first evidence in Europe of an MDR, blaNDM-4-positive Escherichia coli isolated from a food-producing animal, harboured by a novel IncFII plasmid of which we report the complete sequence., Methods: One blaNDM-4-positive E. coli isolated in 2019 from the caecal contents of a fattening pig in Italy was in-depth characterized by combined bioinformatic analysis of Oxford Nanopore long reads and Illumina short reads, for in silico typing, determination of the blaNDM-4 genetic context and full reconstruction of the blaNDM-4-carrying plasmid., Results: The isolate belonged to ST641 and to the genoserotype O108:H23 and tested positive for different virulence genes and plasmid replicons. The MDR phenotype of resistance to all β-lactams, carbapenems, sulfamethoxazole and trimethoprim was mediated by blaTEM-1B, blaNDM-4, sul1/sul3 and dfrA12, respectively. The blaNDM-4 gene was harboured by a novel 53 043 bp IncFII plasmid (pMOL412&#95;FII) composed of four main genetic regions, including an MDR region (MRR-NDM-4) of 16 kb carrying blaNDM-4 and several antimicrobial resistance genes located in a class 1 integron. pMOL412&#95;FII was closely related to another ∼90.3 kb plasmid (pM109&#95;FII) harbouring blaNDM-4 in an E. coli isolated from a human patient in Myanmar., Conclusions: To the best of our knowledge, we have identified for the first time in Europe an NDM-producing Enterobacterales in livestock and resolved the complete sequence of the novel pMOL412&#95;FII plasmid harbouring blaNDM-4 in an MRR. A global One Health approach, comparing genomic data from different sources and geographical areas, may help to trace back and control possible plasmid-borne carbapenemase gene transmission between animals and humans and along the food chain at international level., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)

Significance: This study evaluates how limbal clearance (LC) may impact the clinical performance of scleral lenses based on subjective response on comfort and ocular responses. Limbal zone designs in scleral lenses might affect the integrity of the limbal epithelial tissue as a result of a combined hypoxic and compression-related etiology., Purpose: This study aimed to investigate the clinical performance and ocular and subjective responses to the wear of scleral lenses having varying LC., Methods: Lenses with varying LC with a difference of 50 μm were fitted on and compared among keratoconic participants. The lenses were worn during a 2-week period. Visual analog scales concerning subjective comfort were completed. Ocular surface findings including hyperemia, limbal staining, and corneal swelling were compared., Results: Participants reported greater comfort achieved with high LC. No difference in limbal and bulbar hyperemia was found (P > .05). Corneal response to scleral lens wear with both low and high LCs seems to result in similar perilimbal staining and negative corneal staining. An increase in corneal pachymetric values was noted at the central and peripheral cornea, with no difference between low- and high-LC lenses (all, P < .05). Quadrant-specific analysis indicated that all meridians at both 6 and 8 mm were increased for the low-LC lenses (all, P < .05) and only the temporal region for the high-LC lenses (P = .02)., Conclusions: Limbal clearance may play an important role in subjective performance in scleral lenses but does not impact the degree of hyperemia in either the limbal or bulbar region. Although low LC might result in more compression-related changes to ocular surface, high LC is associated with greater comfort and greater edematous changes. Limbal zone designs in scleral lenses might affect the integrity of the limbal epithelial tissue as a result of a combined hypoxic and compression-related etiology.

We conducted a multilaboratory assessment to determine the suitability of a new commercially available reference material with 40 cancer variants in a background of wild-type DNA at four different variant allele frequencies (VAFs): 2%, 0.50%, 0.125%, and 0%. The variants include single nucleotides, insertions, deletions, and two structural variations selected for their clinical importance and to challenge the performance of next-generation sequencing (NGS) methods. Fragmented DNA was formulated to simulate the size distribution of circulating wild-type and tumor DNA in a synthetic plasma matrix. DNA was extracted from these samples and characterized with different methods and multiple laboratories. The various extraction methods had differences in yield, perhaps because of differences in chemistry. Digital PCR assays were used to measure VAFs to compare results from different NGS methods. Comparable VAFs were observed across the different NGS methods. This multilaboratory assessment demonstrates that the new reference material is an appropriate tool to determine the analytical parameters of different measurement methods and to ensure their quality assurance., (Copyright © 2019 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

We present the first spectral domain optical coherence tomography (SD-OCT) system that combines an isotropic imaging resolution of ~1.5 µm in biological tissue with a 250 kHz image acquisition rate, for in vivo non-contact, volumetric imaging of the cellular structure of the human cornea. OCT images of the healthy human cornea acquired with this system reveal the cellular structure of the corneal epithelium, cellular debris and mucin clusters in the tear film, the shape, size and spatial distribution of the sub-basal corneal nerves and keratocytes in the corneal stroma, as well as reflections from endothelial nuclei. The corneal images presented here demonstrate the potential clinical value of the new high speed, high resolution OCT system for non-invasive diagnostics and monitoring the treatment of corneal diseases., Competing Interests: The authors declare that there are no conflicts of interest related to this article.

Purpose: To assess how varying degrees of corneal clearance of scleral contact lenses (ScCL) impact visual acuity (VA) and comfort in patients with corneal ectasia., Method: Three ScCL were fitted to 20 subjects with previous diagnosis of either keratoconus (n=18) or pellucid marginal degeneration (n=2). Fitting of ScCL was based on corneal sagittal height (CSH) measured with Visante OCT at a 15-mm chord on the horizontal meridian. To select the ScCL from the diagnostic lens set, values of 325, 375, and 425 μm were randomly added in sequence to CSH. Subjects wore ScCL for 1 hr. Central corneal clearance (CCC) and topographic corneal clearance (TCC) along the vertical meridian were assessed using an ultralong optical coherence tomographer. High-contrast VA (HCVA) and low-contrast VA (LCVA) were measured using a logarithm of the minimum angle of resolution VA chart, and comfort ratings were obtained with a standard comfort scale (0-100)., Results: Mean CSH in the horizontal meridian was 3.78±0.53 (range: 3.33-4.17) mm at a 15-mm chord. Mean CCC was 190±100 μm (TCC: 160±94 μm at +3 mm and 180±94 μm at -3 mm), 360±120 μm (TCC: 260±100 μm at +3 mm and 330±110 μm at -3 mm), and 450±170 μm (TCC: 320±120 μm at +3 mm and 400±120 μm at -3 mm) for each lens (P=0.001). Mean HCVA for lenses 1, 2, and 3 were 0.05±0.12, 0.07±0.11, and 0.11±0.08 respectively, which were significantly different (P=0.02). Tukey post hoc analysis showed that this difference was only significant between lenses 1 and 3 (P=0.01). Similar findings were found for LCVA. Comfort ratings for lenses 1, 2, and 3 were 74.9±9.2, 79.7±11.6, and 78.6±10.8, respectively (P=0.24)., Conclusion: The CSH is an effective method of determining the appropriate lens/cornea relationship. Lens 2 (+375 μm) gave the best combination of acuity and comfort ratings. Evaluation of the fluorescein pattern must be balanced with VA and comfort ratings for successful fitting in a clinical setting.

Purpose: First, to examine how wearing high and low modulus lenses with two different base curves affected lens fit, and the corneal tissue and bulbar conjunctival vascular tissue (bulbar redness and blood velocity). Secondly, to quantify the associations between these baseline and outcome variables and the third purpose was to correlate these variables with end of day comfort., Methods: Thirty participants wore higher (PureVision (PV) 8.3, 8.6) and lower (Acuvue Advance (AA) 8.3, 8.7) modulus silicone hydrogel lenses for two weeks on a daily wear basis. Lens fitting characteristics were examined. Corneal epithelial thickness was measured and the cornea and conjunctiva were assessed. RBC velocity was estimated from high magnification bulbar conjunctival images. Subjective comfort/dryness was reported by participants using visual analogue scales., Results: AA lenses were rated the most comfortable (ANOVA, p=0.041). The least movement was while using the AA 8.3 base curve lens (Tukey p=0.028). Steep AA and PV lenses showed significantly higher conjunctival staining at the 2 week visit (ANOVA, p=0.029). There was a significant decrease in RBC velocity with both steeper AA lenses vs PV lenses (Tukey, p=0.001). Comparing baseline and 2 week visits, there was a significant negative correlation for the PV 8.3 between comfort and superior bulbar staining (r=-0.53). For both the PV 8.3 and AA 8.3 reduced RBC velocity was correlated with dryness (r=0.61 and r=0.91, respectively)., Conclusions: Physical differences in contact lenses affect structural and vascular functional aspects of the ocular surface and these may be associated with symptoms of dryness., (Copyright © 2017 British Contact Lens Association. All rights reserved.)

Background: We demonstrate the feasibility of creating a pair of reference samples to be used as surrogates for clinical samples measured in either a research or clinical laboratory setting. The reference sample paradigm presented and evaluated here is designed to assess the capability of a measurement process to detect true differences between two biological samples. Cell-based reference samples can be created with a biomarker signature pattern designed in silico. Clinical laboratories working in regulated applications are required to participate in proficiency testing programs; research laboratories doing discovery typically do not. These reference samples can be used in proficiency tests or as process controls that allow a laboratory to evaluate and optimize its measurement systems, monitor performance over time (process drift), assess changes in protocols, reagents, and/or personnel, maintain standard operating procedures, and most importantly, provide evidence for quality results., Results: The biomarkers of interest in this study are microRNAs (miRNAs), small non-coding RNAs involved in the regulation of gene expression. Multiple lung cancer associated cell lines were determined by reverse transcription (RT)-PCR to have sufficiently different miRNA profiles to serve as components in mixture designs as reference samples. In silico models based on the component profiles were used to predict miRNA abundance ratios between two different cell line mixtures, providing target values for profiles obtained from in vitro mixtures. Two reference sample types were tested: total RNA mixed after extraction from cell lines, and intact cells mixed prior to RNA extraction. MicroRNA profiling of a pair of samples composed of extracted RNA derived from these cell lines successfully replicated the target values. Mixtures of intact cells from these lines also approximated the target values, demonstrating potential utility as mimics for clinical specimens. Both designs demonstrated their utility as reference samples for inter- or intra-laboratory testing., Conclusions: Cell-based reference samples can be created for performance assessment of a measurement process from biomolecule extraction through quantitation. Although this study focused on miRNA profiling with RT-PCR using cell lines associated with lung cancer, the paradigm demonstrated here should be extendable to genome-scale platforms and other biomolecular endpoints.

Background: The potential utility of microRNA as biomarkers for early detection of cancer and other diseases is being investigated with genome-scale profiling of differentially expressed microRNA. Processes for measurement assurance are critical components of genome-scale measurements. Here, we evaluated the utility of a set of total RNA samples, designed with between-sample differences in the relative abundance of miRNAs, as process controls., Results: Three pure total human RNA samples (brain, liver, and placenta) and two different mixtures of these components were evaluated as measurement assurance control samples on multiple measurement systems at multiple sites and over multiple rounds. In silico modeling of mixtures provided benchmark values for comparison with physical mixtures. Biomarker development laboratories using next-generation sequencing (NGS) or genome-scale hybridization assays participated in the study and returned data from the samples using their routine workflows. Multiplexed and single assay reverse-transcription PCR (RT-PCR) was used to confirm in silico predicted sample differences. Data visualizations and summary metrics for genome-scale miRNA profiling assessment were developed using this dataset, and a range of performance was observed. These metrics have been incorporated into an online data analysis pipeline and provide a convenient dashboard view of results from experiments following the described design. The website also serves as a repository for the accumulation of performance values providing new participants in the project an opportunity to learn what may be achievable with similar measurement processes., Conclusions: The set of reference samples used in this study provides benchmark values suitable for assessing genome-scale miRNA profiling processes. Incorporation of these metrics into an online resource allows laboratories to periodically evaluate their performance and assess any changes introduced into their measurement process.

Purpose: To test the ability of responses to the Contact Lens Assessment in Youth (CLAY) Contact Lens Risk Survey (CLRS) to differentiate behaviors among participants with serious and significant (S&S) contact lens-related corneal inflammatory events, those with other events (non-S&S), and healthy controls matched for age, gender, and soft contact lens (SCL) wear frequency., Methods: The CLRS was self-administered electronically to SCL wearers presenting for acute clinical care at 11 clinical sites. Each participant completed the CLRS before their examination. The clinician, masked to CLRS responses, submitted a diagnosis for each participant that was used to classify the event as S&S or non-S&S. Multivariate logistic regression analyses were used to compare responses., Results: Comparison of responses from 96 participants with S&S, 68 with non-S&S, and 207 controls showed that patients with S&S were more likely (always or fairly often) to report overnight wear versus patients with non-S&S (adjusted odds ratio [aOR], 5.2; 95% confidence interval [CI], 1.4-18.7) and versus controls (aOR, 5.8; CI, 2.2-15.2). Patients with S&S were more likely to purchase SCLs on the internet versus non-S&S (aOR, 4.9; CI, 1.6-15.1) and versus controls (aOR, 2.8; CI, 1.4-5.9). The use of two-week replacement lenses compared with daily disposables was significantly higher among patients with S&S than those with non-S&S (aOR, 4.3; CI, 1.5-12.0). Patients with S&S were less likely to regularly discard leftover solution compared with controls (aOR, 2.5; CI, 1.1-5.6)., Conclusions: The CLRS is a clinical survey tool that can be used to identify risky behaviors and exposures directly associated with an increased risk of S&S events.

Significance: It is important to be able to accurately estimate the central corneal clearance when fitting scleral contact lenses. Tools available have intrinsic biases due to the angle of viewing, and therefore an idea of the amount of error in estimation will benefit the fitter., Purpose: To compare the accuracy of observers' ability to estimate scleral contact lens central corneal clearance (CCC) with biomicroscopy to measurements using slit-lamp imaging and anterior segment optical coherence tomography (AS-OCT)., Methods: In a Web-based survey with images of four scleral lens fits obtained with a slit-lamp video imaging system, participants were asked to estimate the CCC. Responses were compared with known values of CCC of these images determined with an image-processing program (digital CCC) and using the AS-OCT (AS-OCT CCC). Bland-Altman plots and concordance correlation coefficients were used to assess the agreement of CCC measured by the various methods., Results: Sixty-six participants were categorized for analysis based on the amount of experience with scleral lens fitting into novice, intermediate, or advanced fitters. Comparing the estimated CCC to the digital CCC, all three groups overestimated by an average of +27.3 ± 67.3 μm. The estimated CCC was highly correlated to the digital CCC (0.79, 0.92, and 0.94 for each group, respectively). Compared with the CCC measurements using AS-OCT, the three groups of participants overestimated by +103.3 μm and had high correlations (0.79, 0.93, and 0.94 for each group)., Discussion: Results from this study validate the ability of contact lens practitioners to observe and estimate the CCC in scleral lens fittings through the use of biomicroscopic viewing. Increasing experience with scleral lens fitting does not improve the correlation with measured CCC from digital or the AS-OCT. However, the intermediate and advanced groups display significantly less inter-observer variability compared with the novice group.

Significance: With the increase in the use of scleral contact lenses among practitioners, questions regarding lens settling are gradually gaining attention. This is because current studies support the notion that scleral lenses settle back over time. More research is needed to understand the exact cause and the factors that underpin such phenomenon., Purpose: The present study aims to assess the effect of time on topographic corneal clearance of three scleral contact lenses of varying sagittal depths., Methods: Three scleral contact lenses were fitted to 20 subjects with previous diagnosis of keratoconus (n = 18) or pellucid marginal degeneration (n = 2). The fit was based on corneal sagittal height measured with the Visante optical coherence tomographer (OCT) at 15 mm along the horizontal meridian. To select an appropriate lens from the diagnostic lens set, values of 325 μm (lens 1), 375 μm (lens 2), and 425 μm (lens 3) were randomly added in sequence to the corneal sagittal height. Subjects wore each lens for 1 hour. Corneal clearance was measured at 10-minute intervals for 1 hour using a custom ultra-long OCT. To assess change in clearance, central point and two mid-peripheral points (+3 mm and -3 mm) along an 8-mm chord were measured by taking differences at each time point up to 1 hour. Measurements were repeated for the two other lenses., Results: Mean central corneal clearance loss for all three lenses was 33.83 ± 48.40 μm. This was 26 ± 27 μm (13 ± 14 μm, +3 mm; 34 ± 37 μm, -3 mm), lens 1; 35 ± 59 μm (38 ± 61 μm, +3 mm; 52 ± 69 μm, -3 mm), lens 2; and 41 ± 54 μm (33 ± 26 μm, +3 mm; 52 ± 48 μm, -3 mm), lens 3, respectively. There was no significant difference (P = 0.06) at central and other locations for lens 1 (location and over time). There were significant differences for both lenses 2 and 3 (P < .001, P = .01, respectively) for all three locations and over time., Conclusions: There is a likelihood of clearance loss after 1 hour of lens wear. This varies between subjects, initial lens-fit relationship, and over time.

A research-grade OCT system was used to image in-vivo and without contact with the tissue, the cellular structure and microvasculature of the healthy human corneo-scleral limbus. The OCT system provided 0.95 µm axial and 4 µm (2 µm) lateral resolution in biological tissue depending on the magnification of the imaging objective. Cross-sectional OCT images acquired tangentially from the inferior limbus showed reflective, loop-like features that correspond to the fibrous folds of the palisades of Vogt (POV). The high OCT resolution allowed for visualization of individual cells inside the limbal crypts, capillaries extending from the inside of the POV's fibrous folds and connecting to a lateral grid of micro-vessels located in the connective tissue directly below the POV, as well as reflections from individual red blood cells inside the capillaries. Difference in the reflective properties of the POV was observed among subjects of various pigmentation levels of the POV. Morphological features observed in the high resolution OCT images correlated well with histology. The ability to visualize the limbal morphology and microvasculature in-vivo at cellular level can aid the diagnostics and treatment of limbal stem cell dysfunction and dystrophies.

Purpose: To assess the repeatability and reproducibility of Visante™ OCT and Oculus Pentacam HR® in measuring central (CCT) and topographic corneal thickness (TCT) along the principle meridians in keratoconus (KC) corneae., Method: Twenty participants diagnosed with KC were recruited. There were two study visits. On the first study visit, two repeated corneal thickness measurements were obtained with each instrument. Measurements were repeated at least 48hrs later in the same order. TCT were recorded in the 90, 180, 45 and 135 meridians at 1mm intervals across 8mm chord., Results: Mean CCT for Visante™ OCT was 484.97±43.14μm (range: 484.84-486.09) and Oculus Pentacam HR® was 478.86±45.31μm (range: 477.20-480.53). No significant difference in TCT between the two visits (p=0.54) and measurements (p=0.63) for Visante™ OCT. For Oculus Pentacam HR®, no significant difference was found for each visit (p=0.18) but differences existed in the measurements outside of the central region (p=0.001). Tukey post-hoc analysis shows the differences (p˂0.05) were found in the +1 and +4 (supero-temporal) locations in the 135 meridian. Significant differences were found comparing the two instruments, (p<0.05). Bland Altman plots were used to demonstrate the differences between the two instruments and indicate their limits of agreement., Conclusion: Both instruments gave repeatable measurements as no significant differences were found in most locations in all meridians. Comparing the two instruments, they were not reproducible in all locations., (Copyright © 2017 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.)

Purpose: To understand soft contact lens (SCL) and gas-permeable (GP) lens wearers' behaviors and knowledge regarding exposure of lenses to water., Methods: The Contact Lens Risk Survey (CLRS) and health behavior questions were completed online by a convenience sample of 1056 SCL and 85 GP lens wearers aged 20 to 76 years. Participants were asked about exposing their lenses to water and their understanding of risks associated with these behaviors. Chi-square analyses examined relationships between patient behaviors and perceptions., Results: GP lens wearers were more likely than SCL wearers to ever rinse or store lenses in water (rinsing: 91% GP, 31% SCL, P < 0.001; storing: 33% GP, 15% SCL P < 0.001). Among SCL wearers, men were more likely to store (24% vs. 13%, P = 0.003) or rinse (41% vs. 29%, P = 0.012) their lenses in water. Showering while wearing lenses was more common in SCL wearers (86%) than GP lens wearers (67%) (P < 0.0001). Swimming while wearing lenses was reported by 62% of SCL wearers and 48% of GP lens wearers (P = 0.027). Wearers who rinsed (SCL; P < 0.0001, GP; P = 0.11) or stored lenses in water (SCL; P < 0.0001, GP P = 0.007) reported that this behavior had little or no effect on their infection risk, compared with those who did not. Both SCL (P < 0.0001) and GP lens wearers (P < 0.0001) perceived that distilled water was safer than tap water for storing or rinsing lenses., Conclusions: Despite previously published evidence of Acanthamoeba keratitis' association with water exposure, most SCL, and nearly all GP lens wearers, regularly expose their lenses to water, with many unaware of the risk.

Purpose: The purpose of this study was to determine the change in over-refraction, if any, after a scleral lens settled on the eye for 6-8 h., Methods: Sixteen patients of varying refractive errors and normal corneal curvatures (measured with Pentacam ™ Oculus) were fitted with trial Mini-Scleral Design (MSD) scleral lenses (15.8 mm diameter) in one eye. The sagittal depths of the scleral lenses were selected by adding 350 μm to the corneal sagittal heights measured at a chord length of 15 mm with the Visante ™ optical coherence tomographer (OCT) anterior segment scans and picking the closest available trial lens in the set. Initial measurements were taken 30 min after lens insertion and included an auto-refraction, subjective refraction, and best sphere refraction over the contact lens. Visual acuities and Visante ™ OCT anterior segment scans were also taken. These measurements were repeated after 6-8 h of lens wear., Results: Over the trial wearing period, the average change in the spherical component of the over-refraction was +0.06 D (S.D. 0.17) (p = 0.16). The average change in cylinder was +0.04 D (S.D. 0.19) (p = 0.33). The average absolute change in axis was 1.06° (S.D. 12.11) (p = 0.74). The average change in best sphere was +0.13 ± 0.30 D (p = 0.12). There was no significant change in visual acuity with the best sphere over-refraction over the 6-8 h wearing period. There was a significant change in central corneal clearance over the wearing period of 83 μm (S.D. 22) (p < 0.0001)., Conclusion: Despite a significant change in the central corneal clearance due to thinning of the fluid reservoir as the scleral lens settled (an average decrease of 83 μm after wearing the lenses for 6-8 h), there was not a statistically significant change in the subjective over-refraction (sphere, cylinder, and axis) or best sphere or visual acuity. This study has confirmed that there is no link between reduction in central corneal clearance and change in over-refraction for average corneas., (© 2017 The Authors Ophthalmic & Physiological Optics © 2017 The College of Optometrists.)

Corneal degenerative conditions such as keratoconus (KC) cause progressive damage to the anterior corneal tissue and eventually severely compromise visual acuity. The ability to visualize corneal tissue damage in-vivo at cellular or sub-cellular level at different stages of development of KC and other corneal diseases, can aid the early diagnostics as well as the development of more effective treatment approaches for various corneal pathologies, including keratoconus. Here, we present the optical design of an optical coherence tomography system that can achieve 0.95 µm axial resolution in biological tissue and provide test results for the system's spatial resolution and sensitivity. Corneal images acquired in-vivo with this system from healthy and keratoconic human subjects reveal the cellular and sub-cellular structure of the corneal epithelium, as well as the normal and abnormal structure of the Bowman's membrane and the anterior corneal stroma.

Purpose: To compare the habits of United States (US) soft contact lens (SCL) wearers who bought SCLs from their eye care practitioner (ECP), on the internet/telephone, or at retail (not where they were examined) to test the effect of proximity to the prescriber on SCL wear and care practices., Methods: Adult SCL wearers completed an adapted Contact Lens Risk Survey (CLRS) online that queried items related to risk factors for SCL-related complications. Responses from subjects who purchased at the ECP, via the internet/telephone, or at a retail store were compared (Chi-Square)., Results: Purchase sources were: ECP 646 (67%, 44±12 yrs, 17% male), Retail 104 (11%, 45±13 yrs, 28% male), and Internet/telephone 218 (23%, 45±12 yrs, 18% male); age (p=0.51), gender (p=0.021). Internet purchasers had fewer annual eye exams (79% ECP, 83% retail, 66% internet/telephone, p=0.007), purchased more hydrogel SCLs (34% ECP, 29% retail, 45% internet/telephone, p=0.0034), and paid for SCLs with insurance less often (39% ECP, 29% retail, 19% internet/telephone, p<0.0001). Other behaviors were similar across groups (p>0.05)., Conclusions: In this sample, the purchase location of SCL wearers had limited impact on known risk factors for SCL-related complications. Internet purchasers reported less frequent eye exams and were more likely to be wearing hydrogel SCLs. Closer access to the ECP through in-office SCL purchase did not improve SCL habits or reduce the prevalence of risk behaviors., (Copyright © 2016 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.)

Photobacterium damselae subsp. damselae (PDD) is a known pathogen of fish, humans and marine mammals. In this study, a Multilocus Sequence Typing (MLST) scheme based on six housekeeping genes (g lp, gyrB, metG, pnt, pyrC , and toxR ) was developed to better understand the PDD population structure and used to type 73 PDD isolates from cetaceans, mainly striped dolphins ( Stenella coeruleoalba ) involved in mortality episodes, and from a few marine chelonians. Five reference ATCC strains were also included in the study. Typing allowed the discrimination of groups of PDD strains isolated from different host species, at different times and from different geographic areas, suggesting that a clonal PDD group may have spread in the Tyrrhenian sea at the time of an Unusual Mortality Event (UME) among cetaceans, mainly striped dolphins, occurred in early 2013 along the Italian western coasts.

Hodgkin/Reed-Sternberg (HRS) cells in the setting of chronic lymphocytic leukemia (CLL) exist in 2 forms: type I with isolated HRS cells in a CLL background (Hodgkin-like lesion) and type II with typical classic Hodgkin lymphoma, a variant of Richter transformation (CHL-RT). The clinical significance of the 2 morphological patterns is unclear, and their biological features have not been compared. We retrospectively reviewed 77 cases: 26 of type I and 51 of type II CHL-RT; 3 cases progressed from type I to type II. We examined clinical features, Epstein-Barr virus (EBV) status, and clonal relatedness after microdissection. Median age for type I was 62 years versus 73 years for type II (P=.01); 27% (type I) versus 73% (type II) had a history of CLL. HRS cells were positive for EBV in 71% (55/77), similar in types I and II. Clonality analysis was performed in 33 cases (type I and type II combined): HRS cells were clonally related to the underlying CLL in 14 and unrelated in 19. ZAP-70 expression of the CLL cells but not EBV status or morphological pattern was correlated with clonal relatedness: all 14 clonally related cases were ZAP-70 negative, whereas 74% (14/19) of clonally unrelated cases were ZAP-70 positive. Overall median survival (types I and II) after diagnosis was 44 months. Advanced age was an adverse risk factor for survival, but not histologic pattern, type I versus type II. HRS-like cells in a background of CLL carries a similar clinical risk to that of CHL-RT and may progress to classic Hodgkin lymphoma in some cases., (Published by Elsevier Inc.)

A Methicillin-resistantStaphylococcus aureus(MRSA) was isolated in Italy from a pathological sample of a mare presenting chronic purulent sinusitis and that had undergone frontal-sinus surgery three months before. Humans, horses, dogs and environmental samples were subsequently collected at the mare's stable and at the Veterinary Hospital, where the mare was operated/hospitalized, and screened for the presence of MRSA that was detected from other horses and from the environment at both sites. All the MRSA isolates belonged to clonal complex (CC)8, ST8-t11469-SCCmec-IVa, and showed similar phenotypic and genetic multidrug resistance patterns and macrorestriction-pulsed-field gel electrophoresis profiles. The only MRSA detected from humans was a CC1, ST1-t127-SCCmec-IVa. This paper represents the first report of a clinical MRSA infection in a horse in Italy. This study also supports the opinion that improper use of antibiotics and hospitalization/surgery can represent risk factors for MRSA colonization/infection in horses, and that the environment is among important sources for exposure., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Staphylococcus aureus is involved in a wide variety of diseases in humans and animals, and it is considered one of the most significant etiological agents of intramammary infection in dairy ruminants, causing both clinical and subclinical infections. In this study, the intra-farm prevalence and circulation of methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) were investigated on an Italian dairy sheep farm previously identified as MRSA-positive by testing bulk tank milk (first isolation in 2012). Human samples (nasal swabs, hand skin samples, and oropharyngeal swabs) from 3 persons working in close contact with the animals were also collected, and the genetic characteristics and relatedness of the MRSA isolates from human and animal sources within the farm were investigated. After 2yr from the first isolation, we confirmed the presence of the same multidrug-resistant strain of MRSA sequence type (ST)1, clonal complex (CC)1, spa type t127, staphylococcal cassette chromosome mec (SCCmec) type IVa, showing identical pulsed field gel electrophoresis (PFGE) and resistance profiles at the farm level in bulk tank milk. Methicillin-resistant S. aureus isolates were detected in 2 out of 556 (0.34%) individual milk samples, whereas MSSA isolates were detected in 10 samples (1.8%). The MRSA were further isolated from udder skin samples from the 2 animals that were MRSA-positive in milk and in 2 of the 3 examined farm personnel. All MRSA isolates from both ovine and human samples belonged to ST(CC)1, spa type t127, SCCmec type IVa, with some isolates from animals harboring genes considered markers of human adaptation. In contrast, all MSSA isolates belonged to ruminant-associated CC130, ST700, spa type t528. Analysis by PFGE performed on selected MRSA isolates of human and animal origin identified 2 closely related (96.3% similarity) pulsotypes, displaying only minimal differences in gene profiles (e.g., presence of the immune evasion cluster genes). Although we observed low MRSA intra-farm prevalence, our findings highlight the importance of considering the possible zoonotic potential of CC1 livestock-associated MRSA, in view of the ability to persist over years at the farm level. Biosecurity measures and good hygiene practices could be useful to prevent MRSA spread at the farm level and to minimize exposure in the community and in categories related to farm animal industry (e.g., veterinarians, farmers, and farm workers)., (Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

The rate of fragile elderly patients affected by chronic kidney disease stage 5-5D is rapidly increasing. The decision making process regarding the start and the withdrawal of dialysis is often difficult for all those involved: patients, relatives, nephrologists and renal nurses. Therefore nephrologists and renal nurses are called to rapidly improve their theoretical and practical competence about the end-of-life care. The quality of clinical intervention and management requires a sound expertise in the ethical, legal, organizational and therapeutic aspects, not trivial nor even deductible from purely private and individual opinions nor from traditional medical practice. The present paper discusses the ethical and legal implications related to the start rather than to withdrawn from dialysis, preferring a non-dialysis medical treatment and / or palliative care. Operational aspects regarding the regional network of palliative care, the path of shared decision making process and a systematic approach to optimize medical and nursing interventions through the Liverpool Care Pathway program are discussed thereafter.

Purpose: To visualize in vivo and quantify the thickness of the posterior corneal layers: the acellular pre-Descemet's layer (PDL), Descemet's membrane (DM), and endothelium (END) in healthy subjects, using ultrahigh-resolution optical coherence tomography (UHR-OCT)., Methods: A research-grade, 800-nm UHR-OCT system with 0.95-μm axial resolution in corneal tissue was used to image in vivo the posterior cornea in healthy subjects. The system offers approximately 98 dB sensitivity for 680 μW optical power incident on the cornea and 34,000 A-scans/s image acquisition rate. This study comprised 20 healthy subjects, aged 20 to 60 years. The thickness of the PDL, DM, and END layers was measured both with a custom, automatic segmentation algorithm and manually., Results: The boundaries and structure of the posterior corneal layers were clearly visible in the UHR-OCT images. The average thickness was measured to be 6.6 ± 1.4 μm (PDL), 10.4 ± 2.9 μm (DM), and 4.8 ± 0.4 μm (END), which agrees well with published data from ex vivo studies. Both the END and DM thickness showed minor spatial variations, whereas the PDL showed up to 2× thickness change for different locations on the same cross-sectional corneal image or over the entire imaged region of the cornea., Conclusions: Our data indicate that all three layers of the posterior cornea can be clearly visualized in vivo and their thicknesses measured precisely with UHR-OCT. Although the PDL thickness showed large spatial variations, the thickness of the DM and END layers was consistent over the entire imaged region of the cornea.

Purpose: The purpose of this study was to assess risk factors associated with soft contact lens (SCL)-related corneal infiltrative events (CIEs)., Methods: This was a single-visit, case-control study conducted at five academic centers in North America. Cases were defined as current SCL wearers with a symptomatic CIE. For each case, three age- and sex-matched controls were enrolled. Subjects completed the Contact Lens Risk Survey (CLRS), a standardized scripted medical interview, supplied a recent health history, and underwent an ocular examination. Microbial culturing of the ocular surface, SCL, and lens storage case was conducted for all cases and one of the three matched controls. Univariate and multivariate logistic regression modeling were used to assess the risk of developing a CIE., Results: Thirty cases and 90 controls 13 to 31 years of age completed the study. Corneal infiltrative event diagnosis included contact lens-associated red eye, infiltrative keratitis, and contact lens peripheral ulcer. Subjects with symptomatic CIEs were more likely to harbor substantial levels of gram-negative bioburden on the ocular surface and contact lens. Significant risk factors for developing a CIE were overnight wear of SCLs, use of multipurpose solution, rinsing SCLs with water, lens storage case older than 6 months, previous "red eye" event, use of ocular drops in the past week, and illness during the past week., Conclusions: This pilot study demonstrated feasibility of enrolling a representative pool of SCL wearers with an untreated, symptomatic CIE and assessing CIE risk factors by using standardized methods. A larger sample size is needed to determine relationships between patient-reported behaviors and exposures, microbial bioburden, and CIE development.

Contact lenses provide safe and effective vision correction for many Americans. However, contact lens wearers risk infection if they fail to wear, clean, disinfect, and store their contact lenses as directed. Over the past decade, CDC has investigated several multistate outbreaks of serious eye infections among contact lens wearers, including Acanthamoeba keratitis. Each investigation identified frequent contact lens hygiene-related risk behaviors among patients. To guide prevention efforts, a population-based survey was used to estimate the number of contact lens wearers aged ≥18 years in the United States. A separate online survey of contact lens wearers assessed the prevalence of contact lens hygiene-related risk behaviors. Approximately 99% of wearers reported at least one contact lens hygiene risk behavior. Nearly one third of contact lens wearers reported having experienced a previous contact lens-related red or painful eye requiring a doctor's visit. An estimated 40.9 million U.S. adults wear contact lenses, and many could be at risk for serious eye infections because of poor contact lens wear and care behaviors. These findings have informed the creation of targeted prevention messages aimed at contact lens wearers such as keeping all water away from contact lenses, discarding used disinfecting solution from the case and cleaning with fresh solution each day, and replacing their contact lens case every 3 months.

Background: Understanding the factors associated with myopic progression is critical to properly recruit subjects into clinical trials for control of myopia. The purpose of this study is to describe the rate of change in soft contact lens (SCL) power and the associated predictive factors in a young clinical population from the Contact Lens Assessment in Youth study., Methods: Data from a retrospective chart review of myopic SCL wearers aged eight to 22 years were analysed for rate of progression of myopia and associated characteristics using multivariate methods., Results: Myopic subjects (n = 912) with at least six months of follow-up were observed (4,341 visits, mean follow-up 25 months, 37 per cent hydrogel and 63 per cent silicone hydrogel SCLs). During observation, 36 per cent of subjects experienced a change in soft contact lens power of -0.50 D or more. Significant predictors of future increase in minus lens power were: ages eight to 13 years, shorter time to the first increase in minus power and hydrogel soft contact lens material. The mean annualised increase in minus decreased with age (-0.31D per year for eight to 13 year olds to -0.10 D per year for 20 to 22 year olds, p < 0.0001). Increases in minus were less common among users of silicone hydrogel materials than hydrogel daily disposable lenses after controlling for age (p = 0.039)., Conclusion: In this retrospective chart review of young soft contact lens wearers, the mean annualised rate of increase in minus soft contact lens power decreased with age, longer time to first increase in power and was greater with hydrogel soft contact lenses. The rates observed were similar to progression rates in prospective myopia clinical trials that employed cylcoplegic autorefraction., (© 2015 The Authors. Clinical and Experimental Optometry © 2015 Optometry Australia.)

Purpose: Conjunctival compression observed in ultrahigh resolution optical coherence tomography (UHR-OCT) images of contact lens edges could be actual tissue alteration, may be an optical artefact arising from the difference between the refractive indexes of the lens material and the conjunctival tissue, or could be a combination of the two. The purpose of this study is to image the artefact with contact lenses on a non-biological (non-indentable) medium and to determine the origins of the observed conjunctival compression., Methods: Two-dimensional cross-sectional images of the edges of a selection of marketed silicone hydrogel and hydrogel lenses (refractive index ranging from 1.40 to 1.43) were acquired with a research grade UHR-OCT system. The lenses were placed on three continuous surfaces, a glass sphere (refractive index n = 1.52), a rigid contact lens (n = 1.376) and the cornea of a healthy human subject (average n = 1.376). The displacement observed was analysed using ImageJ., Results: The observed optical displacement ranged between 5.39(0.06) μm with Acuvue Advance and 11.99(0.18) μm with Air Optix Night & Day when the lens was imaged on the glass reference sphere. Similarly, on a rigid contact lens displacement ranged between 5.51(0.03) and 9.72(0.12) μm. Displacement was also observed when the lenses were imaged on the human conjunctiva and ranged from 6.49(0.80) μm for the 1-day Acuvue Moist to 17.4(0.22) μm for the Pure Vision contact lens., Conclusions: An optical displacement artefact was observed when imaging a contact lens on two rigid continuous surfaces with UHR-OCT where compression or indentation of the surface could not have been a factor. Contact lenses imaged in situ also exhibited displacement at the intersection of the contact lens edge and the conjunctiva, likely a manifestation of both the artefact and compression of the conjunctiva., (© 2015 The Authors Ophthalmic & Physiological Optics © 2015 The College of Optometrists.)

Purpose: To compare live and photographic (still) grades of corneal staining of the same eyes and the repeatability of grading between two investigators., Methods: Thirty patients were recruited to participate in a contact lens study, and their level of corneal staining was graded by two investigators in situ (live images), using slit lamp biomicroscopy. Digital still images of the corneal staining were also captured during the study visits. An independent observer selected 105 of the still images graded by investigator 1 and another 105 images graded by investigator 2 and presented them to the original investigator in a random order, on three separate occasions. Grading was performed at the time of the live grading and the three still image sessions, using the Centre for Contact Lens Research corneal staining scale that combines grades of both extent and type to provide an overall "global staining score" from 0 to 10,000 for corneal staining. A comparison was made between live and still grades as well as the intrainvestigator repeatability for the multiple grading of the still images., Results: The mean (±SD) of corneal staining grades recorded for the same eyes examined live and then later on three occasions was 1795 (±1083) and 714 (±974), respectively, for participants examined by investigator 1 (p < 0.001) and 1854 (±1075) and 461 (±411) for those examined by investigator 2 (p < 0.001). There was a significant difference over the three repeated still grading sessions for each investigator (p < 0.001), although there was a high degree of consistency among the three still grading sessions for each of the investigators: the intraclass correlation for investigator 1 was 0.91 (confidence interval, 0.87 to 0.93) and that for investigator 2 was 0.82 (confidence interval, 0.77 to 0.87)., Discussion: Digital still image grading of corneal staining significantly underrepresented the amount of corneal staining observed through a slit lamp. Clinical investigators graded corneal staining with a high degree of consistency.

Tear exchange beneath a contact lens facilitates ongoing fluid replenishment between the ocular surface and the lens. This exchange is considerably lower during the wear of soft lenses compared with rigid lenses. As a result, the accumulation of tear film debris and metabolic by-products between the cornea and a soft contact lens increases, potentially leading to complications. Lens design innovations have been proposed, but no substantial improvement in soft lens tear exchange has been reported. Researchers have determined post-lens tear exchange using several methods, notably fluorophotometry. However, due to technological limitations, little remains known about tear hydrodynamics around the lens and, to-date, true tear exchange with contact lenses has not been shown. Further knowledge regarding tear exchange could be vital in aiding better contact lens design, with the prospect of alleviating certain adverse ocular responses. This article reviews the literature to-date on the significance, implications and measurement of tear exchange with contact lenses., (Copyright © 2014 Spanish General Council of Optometry. Published by Elsevier Espana. All rights reserved.)

Background: A 3.4kb deletion (3.4kbΔ ) in mitochondrial DNA (mtDNA) found in histologically normal prostate biopsy specimens has been reported to be a biomarker for the increased probability of prostate cancer. Increased mtDNA copy number is also reported as associated with cancer., Objective: Independent evaluation of these two potential prostate cancer biomarkers using formalin-fixed paraffin-embedded (FFPE) prostate tissue and matched urine and serum from a high risk cohort of men with and without prostate cancer., Methods: Biomarker levels were detected via qPCR., Results: Both 3.4kbΔ and mtDNA levels were significantly higher in cancer patient FFPE cores (p= 0.045 and p= 0.070 respectively at > 90% confidence). Urine from cancer patients contained significantly higher levels of mtDNA (p= 0.006, 64.3% sensitivity, 86.7% specificity). Combining the 3.4kbΔ and mtDNA gave better performance of detecting prostate cancer than either biomarker alone (FFPE 73.7% sensitivity, 65% specificity; urine 64.3% sensitivity, 100% specificity). In serum, there was no difference for any of the biomarkers., Conclusions: This is the first report on detecting the 3.4kbΔ in urine and evaluating mtDNA levels as a prostate cancer biomarker. A confirmation study with increased sample size and possibly with additional biomarkers would need to be conducted to corroborate and extend these observations.

Purpose: Previous studies have reported that the risk of corneal infectious and inflammatory events (CIEs) with soft contact lens (SCL) wear is highest in late adolescence and early adulthood. This study assesses the associations between patient age and other factors that may contribute to CIEs in young SCL wearers., Methods: After ethics approvals and informed consent, a nonclinical population of young SCL wearers was surveyed in five US cities. Data from 542 SCL wearers aged 12-33 years were collected electronically. Responses were analyzed by age bins (12-14, 15-17, 18-21, 22-25, 26-29, and 30-33 years) using chi-square test., Results: The cohort was 34% male and balanced across age bins. There were several significant associations between survey response and age (in bins). Wearers aged 18-21 years reported more recent nights with less than 6 hours of sleep (p < 0.001), more colds/flu (p = 0.049), and higher stress levels (p < 0.001). Wearers 18-21 and those 22-25 years were more likely to wear SCLs when showering (p < 0.001) and also reported more frequent naps with SCLs (p < 0.001). They reported sleeping in SCLs after alcohol use (p = 0.031), when traveling (p = 0.001), and when away from home (p = 0.024). Lower rates of regular hand washing before lens application (p = 0.054) was also associated with these groups. In addition, the relationship between reactive replacement and recommended replacement was dependent on age (p < 0.0001)., Conclusions: Patient age influences lens wearing behaviors, environmental exposures, and other determinants of health that may contribute to increased CIEs in younger wearers. Targeted, age-specific education should be considered for both new and established SCL wearers.

Objectives: This was a secondary analysis of an existing dataset of soft contact lens wearers age 8-33 years, who received eye care outside of a clinical trial. The aim was to identify geographical and temporal factors associated with interruptions to contact lens wear., Methods: Data from six academic centers in North America captured 522 events in 3549 patients. Cases were analyzed overall and in subcategories of allergic, and serious and significant adverse events. General estimating equations were used to model the effect of geographic (Northwest, West, Central, Northeast, Southeast) and temporal (season, month, day of the week) factors, along with previously identified risk factors associated with interruptions in lens wear (patient age, contact lens material, overnight wear, lens care system, replacement schedule, and years of contact lens wear)., Results: After controlling for established risk factors, both region and temporal factors disrupted the patients' ability to maintain contact lens wear. About 4% of all visits had complications that led to an interruption in wear. Allergic events were highest in the Central region. Serious and significant adverse events peaked in the Southeast during the Autumn and Winter months (September, October, December). Day of the week was not significant in any model., Conclusions: This study provides evidence of seasonal and regional challenges to contact lens wear. As the use of soft contact lenses expands for both cosmetic and medical reasons, practitioners must examine ways to maintain continuous, safe, and healthy use of contact lenses across all patients., (Published by Elsevier Ltd.)

Aim: To describe the meibomian gland (MG) appearance in a case of eyelid chalazia, using a novel meibography method., Methods: A 29-year-old female of South Asian origin presented with mild swelling in the lower lid of the left eye. The patient also presented with a history of a recurrent chalazion in the upper lid of the same eye, which later progressed to an active chalazion. A chalazion also developed in the upper lid of the right eye. Meibography was performed using a novel Keratograph (Keratograph 4, OCULUS, Wetzlar, Germany) to evaluate the structure of the MG in the area affected by the chalazia., Results: The area of the recurrent chalazion in the upper left lid showed partial and/or complete MG loss. The active chalazia in the lower left lid and the upper right lid showed inflammation and MG drop out at the affected site. The inflammation was found to be reduced during the follow-up visits, however disappearance of MG very specific to the region of the chalazion was observed., Conclusion: The Keratograph 4 was able to image the MG structures clearly, allowing the clinician to monitor the progression of chalazia and the MG loss in the affected areas., (Copyright © 2013 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.)

Purpose: The primary purpose of this pilot study was to compare epithelial barrier function (EBF) and staining in a small group of participants using a silicone hydrogel (SH) lens worn on a daily basis with two different care regimens. Secondarily, the aim was to see if there was any correlation between corneal staining and EBF., Methods: The corneal EBF of 10 non-lens wearers (control) and 15 age-matched asymptomatic SH contact lens wearers (test) were assessed using fluorophotometry. Biomicroscopy was performed to assess corneal staining after the EBF was measured. The lens wearers wore PureVision (FDA group V) SH lenses for two consecutive 1-month periods while using either Alcon Opti-Free Express or Renu Fresh using a randomized, investigator-masked, crossover design. Control subjects were assessed on one occasion, and lens wearers were examined before fitting with lenses and after 7, 14, and 28 days of lens wear, with each combination., Results: Compared with the control group, both study groups had an increase in epithelial permeability at baseline (p ≤ 0.04). There were no changes in EBF during the treatment period for either solution (p = 0.87). A significant difference in EBF was found between the test groups during the treatment period (p = 0.02), with greater permeability in the Renu Fresh-disinfected lenses. There was poor correlation between corneal staining and EBF (r = 0.35, p > 0.05) because of large individual variations., Conclusions: Daily wear of highly oxygen-permeable SH lenses increases corneal epithelial permeability to fluorescein probably because of increased mechanical effects. In addition, certain lens-solution interactions can add to this effect, as seen in this study. Despite having a low amount of central corneal staining in the Renu Fresh group, staining and EBF did not prove to be well correlated. The presence of central corneal staining is a confounding factor when measuring EBF.

Recent discoveries in cancer biology have greatly increased the understanding of cancer at the molecular level, but translating this knowledge into clinically useful diagnostic tests has proved challenging. More efficient transfer of new molecular tests into patient care requires better standardization of laboratory practices, measurement methods and data management. The workshop assembled experts from National Cancer Institute, US FDA, National Institute of Standards and Technology, academia and industry, to address the most efficient approaches to biomarker standardization and validation. The workshop participants described the current state of research in molecular diagnostics standardization and addressed three questions: what has worked? What has not?And what needs improving?

Background: The mission of the National Cancer Institute's Early Detection Research Network (EDRN) is to identify and validate cancer biomarkers for clinical use. Since its inception, EDRN investigators have learned a great deal about the process of validating biomarkers for clinical use. Translational research requires a broad spectrum of research expertise, and coordinating collaborative activities can be challenging. The EDRN has developed a robust triage and validation system that serves the roles of both "facilitator" and "brake.", Content: The system consists of (a) establishing a reference set of specimens collected under PRoBE (Prospective Specimen Collection Retrospective Blinded Evaluation) design criteria; (b) using the reference set to prevalidate candidate biomarkers before committing to full-scale validation; (c) performing full-scale validation for those markers that pass prevalidation testing; and (d) ensuring that the reference set is sufficiently large in numbers and volumes of sample that it can also be used to study future candidate biomarkers. This system provides rigorous and efficient evaluation of candidate biomarkers and biomarker panels. Reference sets should also be constructed to enable high-quality biomarker-discovery research., Summary: We describe the process of establishing our system in the hope that it will serve as an example of how to validate biomarkers for clinical application. We also hope that this description of the biospecimen reference sets available from the EDRN will encourage the biomarker research community--from academia or industry--to use this resource to advance biomarkers into clinical use., (© 2012 American Association for Clinical Chemistry)

We have developed a system to isolate rare cells from whole blood using commercially available components and simple microfluidics. We characterized the capture of MCF-7 cells spiked into whole human blood using this system to demonstrate that enrichment and enumeration studies give results similar to in situ surface-modified devices while reducing fabrication and operation complexity.