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The aim of this study was to develop an experimental methodology to measure lipid deposition with contact lenses. Contact lenses were incubated in a lipid solution. The amount and types of adsorbed lipids were assessed using mass spectrometry and confocal microscopy. The recovery of lipids from lenses varied with lipid and lens type. Most non-polar and polar lipids were desorbed from lenses during the first 5 min of extraction. Fluorescently labelled phosphatidylcholine bound within the matrix of Senofilcon A lenses but to the surface of Lotrafilcon B lenses, whereas fluorescently labelled cholesteryl ester was found throughout both lenses. The efficacy of extraction of lipids from contact lenses varies for different lipid classes and different lens materials. Differences in the amount and time of lipid desorption probably resulted from the strength of the bond between lipid and lens polymer and the depth of adsorption of lipid in the polymer.

Isolation of antimicrobial-resistant microbes from ocular infections may be becoming more frequent. Infections caused by these microbes can be difficult to treat and lead to poor outcomes. However, new therapies are being developed which may help improve clinical outcomes. This review examines recent reports on the isolation of antibiotic-resistant microbes from ocular infections. In addition, an overview of the development of some new antibiotic therapies is given. The recent literature regarding antibiotic use and resistance, isolation of antibiotic-resistant microbes from ocular infections and the development of potential new antibiotics that can be used to treat these infections was reviewed. Ocular microbial infections are a global public health issue as they can result in vision loss which compromises quality of life. Approximately 70 per cent of ocular infections are caused by bacteria including Chlamydia trachomatis, Staphylococcus aureus, and Pseudomonas aeruginosa and fungi such as Candida albicans, Aspergillus spp. and Fusarium spp. Resistance to first-line antibiotics such as fluoroquinolones and azoles has increased, with resistance of S. aureus isolates from the USA to fluoroquinolones reaching 32 per cent of isolates and 35 per cent being methicillin-resistant (MRSA). Lower levels of MRSA (seven per cent) were isolated by an Australian study. Antimicrobial peptides, which are broad-spectrum alternatives to antibiotics, have been tested as possible new drugs. Several have shown promise in animal models of keratitis, especially treating P. aeruginosa, S. aureus or C. albicans infections. Reports of increasing resistance of ocular isolates to mainstay antibiotics are a concern, and there is evidence that for ocular surface disease this resistance translates into worse clinical outcomes. New antibiotics are being developed, but not by large pharmaceutical companies and mostly in university research laboratories and smaller biotech companies. Antimicrobial peptides show promise in treating keratitis.

Purpose: To evaluate the effect of age, gender and body mass index (BMI) on the levels of tear film neuromediators and corneal nerve parameters in healthy individuals.Methods: Twenty-six he...

Aim: The ocular surface is continually exposed to bacteria from the environment and traditional culture-based microbiological studies have isolated a low diversity of microorganisms from this region. The use of culture-independent methods to define the ocular microbiome, primarily involving 16S ribosomal RNA gene sequencing studies, have shown that the microbial communities present on the ocular surface have a greater diversity than previously reported. Method: A review of the literature on ocular microbiome research in health and disease. Results: Molecular techniques have been used to investigate the effect of contact lens wear and disease on the microbiota of the ocular surface and eyelids and the immunoregulatory role of the ocular surface microbiota. Studies have shown that compositional changes in the microbiota occur in ocular surface disorders such as blepharitis, trachoma and dry eye and also suggest a role of the ocular and non-ocular microbiome in retinal disease including age-related macular degeneration, glaucoma, uveitis and diabetic retinopathy. However, ocular microbiome studies need to recognise the potential for contamination to impact findings and carefully control each stage of the experimental procedure and to utilise statistical methods to identify contamination signals. Conclusion: The healthy ocular surface is characterised by a relatively stable, comparatively low diversity microbiome with recent findings that the bacteria of the ocular surface appear to have a role in maintaining homeostasis by modulating immune function.

Treatment of Pseudomonas aeruginosa eye infections often becomes a challenge due to the ability of this bacterium to be resistant to antibiotics via intrinsic and acquired mechanisms. Transfer of resistance due to interchangeable genetic elements is an important mechanism for the rapid transfer of antibiotic resistance in this pathogen. As a result, drug-resistant strains are becoming increasingly prevalent worldwide. This review systematically analyses data from recent publications to describe the global prevalence and antibiotic sensitivity of ocular P. aeruginosa. Thirty-seven studies were selected for review from PubMed-based searches using the criteria 'microbial keratitis OR eye infection AND Pseudomonas aeruginosa AND antibiotic resistance' and limiting to papers from 2011 onward, to demonstrate the antibiotic resistance from isolates from around the world. Subsequently, we reviewed the ways in which P. aeruginosa can become resistant to antibiotics. Both the rate of isolation of bacteria in general (79 per cent of cases), and prevalence of P. aeruginosa (68 per cent of all isolates) were highest in contact lens-related microbial keratitis. The average resistance rate to common ocular antibiotics such as ciprofloxacin (9 per cent), gentamicin (22 per cent) and ceftazidime (13 per cent) remained relatively low. However, there were large variations in resistance rates reported in studies from different countries, for example resistance to ciprofloxacin reached up to 33 per cent. We next reviewed the types of mobile genetic elements (MGEs) such as plasmids, integrons and transposons that are frequently associated with drug resistance in P. aeruginosa. MGEs are important for the transmission of resistance to beta-lactams and aminoglycosides and recently have been shown to be potential factors for the transmission of fluoroquinolone resistance. Studies on the molecular mechanisms of resistance transfer in ocular P. aeruginosa have begun to be reported and will provide valuable information on the emergence of new antibiotic resistance and potential to treat resistant strains.

Contact lenses are associated with discomfort during wear. This may be the result of stimulation of the ocular surface and production of pro-inflammatory mediators which are then released into the tears. This study examined changes in the concentration in tears of arachidonic acid metabolites (AAM) prostaglandins, cysteinyl leukotrienes, and resolvin-DTears were collected twice a day (morning and evening) for up to 10 days on two different occasions (with and without contact lens wear) from 30 experienced contact lens wearers for analysis of AAM and a separate group (N = 33) for analysis of histamine. Ocular comfort was rated subjectively on an ordinal scale at each time of tear collection. Tears were analyzed using commercial immunoassay-based kits. Statistical analysis was performed using linear mixed model test.Ocular comfort decreased from morning to evening with and without contact lenses (p = 0.001), and the difference in comfort in the evening was greater with contact lens wear (80.9 ± 16.2 vs. 75.5 ± 16.8; p = 0.008). The total concentration of PGs (10.7 ± 10.8 ng/ml), cysteinyl leukotrienes (8.7 ± 0.38 ng/ml), resolvin-DProstaglandins, cysteinyl leukotrienes, resolvin-D1, or histamine concentrations did not alter in relation to changes in comfort of the eye during the day or during contact lens wear. These results suggest that release of these mediators is not responsible for contact lens discomfort.

The objective of this study was to immobilise and characterise a variety of antimicrobial peptides (AMPs) onto poly-hydroxyethylmethacrylate (pHEMA) surfaces to achieve an antibacterial effect. Four AMPs, viz. LL-37, melimine, lactoferricin and Mel-4 were immobilised on pHEMA by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) which assisted covalent attachment. Increasing concentrations of AMPs were immobilised to determine the effect on the adhesion of Pseudomonas aeruginosa and Staphylococcus aureus. The AMP immobilised pHEMAs were characterised by X-ray photoelectron spectroscopy (XPS) to determine the surface elemental composition and by amino acid analysis to determine the total amount of AMP attached. In vitro cytotoxicity of the immobilised pHEMA samples to mouse L929 cells was investigated. Melimine and Mel-4 when immobilised at the highest concentrations showed 3.1 ± 0.6 log and 1.3 ± 0.2 log inhibition against P. aeruginosa, and 3.9 ± 0.6 log and 2.4 ± 0.5 log inhibition against S. aureus, respectively. Immobilisation of LL-37 resulted in up to 2.6 ± 1.0 log inhibition against only P. aeruginosa, but no activity against S. aureus. LFc attachment showed no antibacterial activity. Upon XPS analysis, immobilised melimine, LL-37, LFc and Mel-4 had 1.57 ± 0.38%, 1.13 ± 1.36%, 0.66 ± 0.47% and 0.73 ± 0.32% amide nitrogen attached to pHEMA compared to 0.12 ± 0.14% in the untreated controls. Amino acid analysis determined that the total amount of AMP attachment to pHEMA was 44.3 ± 7.4 nmol, 3.8 ± 0.2 nmol, 6.5 ± 0.6 nmol and 48.9 ± 2.3 nmol for the same peptides respectively. None of the AMP immobilised pHEMA surfaces showed any toxicity towards mouse L929 cells. The immobilisation of certain AMPs at nanomolar concentration to pHEMA is an effective option to develop a stable antimicrobial surface.

Using an in vitro cell culture model, bovine lactoferrin (BLF) stimulates healing of alkali-induced human corneal epithelial wounds. The present study examined the efficacy of BLF in promoting healing of corneal injury in vivo and explored BLF modulation of interleukin-1 (IL-1) during wound healing.Alkali injury was induced to BALB/c mice by exposure of the mouse cornea to a sodium hydroxide (NaOH)-soaked filter disc for 2 min. The corneal surface was irrigated after the injury with saline. Topical BLF in phosphate buffered saline (PBS) (10 µl, 62.5 μM), bovine serum albumin (BSA) (10 µl, 62.5 μM in PBS) or PBS only (10 µl) were applied three times daily to both the alkali-injured and uninjured eyes for 3 d. Wound healing was assessed using 0.1% fluorescein staining under slit lamp microscope. The corneas at 6 h, 24 h or 3 d post-injury and treatment were excised and examined histologically, homogenized corneal tissue was evaluated for expression of IL-1α and IL-1β.After 6 h post-wounding and treatment no significant reduction of wound area was observed between treatments and infiltrating cells or IL-1 expression were not elevated in any group. By 24 h, BLF-treatment resulted in accelerated wound closure (100%) compared to PBS and BSA treatment (70% and 65%, respectively). BLF treatment reduced infiltrating cells compared to controls and no elevation of IL-1, whereas controls displayed elevated infiltrating cells and increased levels of IL-1. After 3 d, mice treated with BLF exhibited complete wound closure while control corneas still exhibited some minor defects. Resolution of inflammation with minimal remaining infiltrating cells was observed in all corneas by day 3, coincident to normal levels of IL-1α and IL-1β.BLF accelerated healing of corneal alkali injury in BALB/c mice which was associated with suppression of IL-1 and reduced infiltrating cells.

Background Rabbits have a longer inter-blink time (approximately 10 minutes) compared with humans (five to eight seconds), suggesting that rabbits have a much more stable tear film. Using fluorescein, the tear break-up time of rabbits has been reported to be similar to that of humans. This study set out to measure the tear break-up time in rabbits using non-invasive methods and to establish the pattern of tear break-up compared to humans. Methods The tear break-up time was measured and the pattern of tear break-up was observed in six New Zealand White rabbits on two separate occasions using both the Keeler Tearscope-plusTM and a slitlamp biomicroscope. Results The mean rabbit tear break-up time was 29.8 ± 3.4 (SD) minutes. This contrasts with the reports of human tear break-up time of eight to 30 seconds. The tear breaking spread very slowly and was often restricted to the area of the initial break. Conclusion Rabbit tears have a significantly higher tear break-up time than humans and this aligns with previously demonstrated differences in inter-blink time between rabbits and humans. Understanding the underlying mechanisms of tear stability may lead to novel ways of increasing human tear film stability.

This study was designed to use multiple reaction monitoring (MRM) for accurate quantification of contact lens protein deposits. Worn lenses used with a multipurpose disinfecting solution were collected after wear. Individual contact lenses were extracted and then digested with trypsin. MRM in conjunction with stable-isotope-labeled peptide standards was used for protein quantification. The results show that lysozyme was the major protein detected from both lens types. The amount of protein extracted from contact lenses was affected by the lens material. Except for keratin-1 (0.83 ± 0.61 vs 0.77 ± 0.20, p = 0.81) or proline rich protein-4 (0.11 ± 0.04 vs 0.15 ± 0.12, p = 0.97), the amounts of lysozyme, lactoferrin, or lipocalin-1 extracted from balafilcon A lenses (12.9 ± 9.01, 0.84 ± 0.50 or 2.06 ± 1.6, respectively) were significantly higher than that extracted from senofilcon A lenses (0.88 ± 0.13, 0.50 ± 0.10 or 0.27 ± 0.23, respectively) (p 0.05). The amount of protein extracted from contact lenses was dependent on both the individual wearer and the contact lens material. This may have implications for the development of clinical responses during lens wear for different people and with different types of contact lenses. The use of MRM-MS is a powerful analytical tool for the quantification of specific proteins from single contact lenses after wear.

Keratoconus is a degenerating disease of the eye that results in an irregularly-shaped cornea. The etiology of the disease is unknown and the prognosis is difficult due to the variability in outcome. Keratoconus has been associated with eye rubbing, atopy, contact lens wear, as well as genetic conditions, such as Down's syndrome, Ehlers-Danlos syndrome, and Marfan's syndrome. Thinning of the cornea in keratoconus has been well studied and is documented to occur as a result of degradation of corneal collagen. The reason for this tissue degradation is unknown but has been hypothesized to be linked with proteases.This study used a literature search to review the role of proteases and inflammatory molecules in the aetiology of keratoconus.Early studies demonstrated elevated levels of collagenolytic and gelatinolytic activities in laboratory cultures of keratoconic corneas. Matrix metalloproteinases (MMPs) are a family of zinc-dependent proteins that include collagenases and gelatinases. MMPs levels are altered in keratoconus corneas compared to normal corneas and the level of tissue inhibitor of metalloproteinases-1 (TIMP-1) is decreased in keratoconic corneas. Recent studies have demonstrated the involvement of Cathepsin B, G, and K in keratoconus. Although thought to be a non-inflammatory disease, inflammatory molecules, such as interleukins and tumor necrosis factor have been shown to be elevated in keratoconus, and these inflammatory molecules may mediate production and activation of proteases.Proteases may be implicated in keratoconus. An in-depth investigation of these proteases may help in better understanding the course of the disease.

Background: Staphylococcus is the leading cause of microbial keratitis. Staphylococcus aureus isolated from ocular infections with resistance to a wide range of antibiotics, including the commonly prescribed fluoroquinolones, is emerging. The aim of this study was to determine the current antibiotic susceptibilities of ocular S. aureus isolates and also determine whether isolates from different adverse events or those with similar antimicrobial susceptibilities are related. Methods: A collection of 55 S. aureus strains from ocular adverse events were analysed for antibiotic susceptibility using disc diffusion (CDS method) and typed using PCR-ribotyping and pulsed-field gel electrophoresis (PFGE). Results: S. aureus isolated from symptomatic ocular adverse events in the USA exhibited greater resistance to antibiotics than did those isolated from symptomatic ocular adverse events in Australia or India (p

Previously, we reported carboxymethyl cellulose (CMC) binding to human corneal epithelial cells and promoting corneal epithelial wound closure in vitro. Using an animal model, the efficacy of CMC in promoting corneal wound healing was examined.Following corneal epithelial wounding of NZ white rabbits, CMC (0.2% or 1.0%) or control vehicle (PBS) was administered topically (4 times daily for 3 days) to wounded and unwounded eyes with or without contact lens wear. Wound healing in response to the treatments was measured as percentage reduction of fluorescein-stained wound area 0 to 72 hr post-wounding. Corneas were examined histologically and expression of zonula occludens-1 (ZO-1) tight-junction was detected by immunohistochemistry.Percentage wound reduction in CMC-treated groups was significantly greater than controls (p0.05) at 24 and 32 hr. Complete wound closure was observed by 48 hr in 100% of CMC-treated eyes compared to 45% of vehicle-treated eyes. CMC also promoted wound closure dose-dependently. Epithelial cells formed an intact layer following CMC-treatment whereas vehicle-treated cells were less ordered. Strong ZO-1 expression in corneal epithelia of CMC-treated eyes was observed at 72 hr. Contact lens wear appeared to delay wound closure compared to without lens wear during CMC-treatment (p = 0.001).CMC promoted dose-dependent corneal epithelial wound healing. CMC stimulated ZO-1 expression, indicating accelerated corneal epithelial resistance barrier regeneration.

To examine the role of Staphylococcus aureus alpha-toxin in contact lens-induced peripheral ulceration (CLPU).Proteolytic enzyme, hyaluronidase, alpha-toxin, and beta-toxin production by S. aureus 8325-4 and its alpha-toxin-deficient mutant (S. aureus DU1090) were examined. Using a rabbit model of CLPU, animals were fitted with hydrogel contact lenses colonized by either S. aureus 8325-4 or the mutant strain. The clinical presentation, bacterial cultures, and histology of the ulceration were examined.Both strains of S. aureus produced similar levels of caseinase, gelatinase, elastase, hyaluronidase, and beta-toxin. S. aureus DU1090 induced weaker haemolysis of rabbit blood cells than S. aureus 8325-4. Ulceration in the S. aureus DU1090 eye was less frequent and less severe than that caused by S. aureus 8325-4.The enzyme production profile of S. aureus DU1090 was similar to the parent strain. S. aureus strains may produce CLPU-like lesions irrespective of alpha-toxin production, but severe infectious lesions are produced only in the presence of alpha-toxin.

During contact lens wear, bacterial contamination of the lens can lead to the development of microbial keratitis (MK) or contact lens induced acute red eye (CLARE). Inflammatory mediators released by the host may heighten the response. We have examined the ability of human corneal epithelial cells and polymorphonuclear leukocytes (PMNs) to release cytokines upon stimulation with bacteria.Serratia marcescens strains were added to corneal epithelial cells or PMNs. After incubations of up to 20 hours, supernatants were assayed for the presence of inflammatory mediators by ELISA.All strains stimulated PMNs to release IL-8 and LTB(4). Epithelial cells did not release LTB(4). IL-8 was released after exposure to most S. marcescens isolates. A sub-group of strains stimulated the release of TNF-alpha from corneal epithelial cells.S. marcescens can stimulate the release of inflammatory mediators from both PMNs and corneal epithelial cells. This release could be important in the progression and resolution of MK and CLARE.

Background: Ocular infection and inflammation during extended wear with hydrogel lenses are often associated with contamination of the lenses with bacteria. This study examines the colonisation of a high-Dk silicone hydrogel contact lens worn on a 30-night extended wear basis (continuous wear) during asymptomatic lens wear. Methods: Worn lenses were collected aseptically and placed in sterile vials. Microbial growth on various media was enumerated and the number of colony-forming units per whole lens was calculated. The proportions of samples contaminated with the bacteria and the extent of contamination were compared after one month, six months and two years of continuous wear widi monthly replacement. The proportion of lenses that were sterile was calculated. Results: The most frequently isolated and most numerous microbes that colonised the contact lenses during wear were the coagulase-negative staphylococci (in particular Staphylococcus epidermidis), followed by Propionibacterium sp. The frequency of isolation of the majority of bacterial isolates did not increase over time. Generally, lenses were sparsely colonised during asymptomatic wear. Conclusion: Up to two years continuous wear of silicone hydrogel lenses of the type used in this study does not appear to alter the types and number of bacteria that colonise die eye during wear, when lenses are replaced on a monthly schedule and the patients remain asymptomatic.

Tear volume is reduced in symptomatic contact lens wearers, evaporation of the ocular tear film may be a cause. In this study we have focussed on symptomatic or intolerant subjects and compared their tear film lipid-related features to those tolerant to soft contact lens wear.Fourteen tolerant and 10 intolerant to lens wear subjects were recruited for this study. Intolerance to lens wear was defined as experiencing dryness symptoms in the first 6 hours of lens wear and consequently not being regular lens wearers. Lipid layer appearance was graded on a 0-5 scale, meibomian gland obstruction was observed, and the McMonnies questionnaire completed. Tears were collected without reflex stimulation. Degraded lipid (tear aldehyde content), secretory phospholipase A2 enzyme (sPLA2) concentration and activity and lipocalin concentration were analysed using spectrophotometry to quantify colour reactions and enzyme linked immunosorbent assays. Statistical results were calculated using non-parametric tests (median +/- interquartile range) or chi-squared test.Degradation of polyunsaturated fatty acids and related esters leads to the by-products, malondialdehyde and 4-hydroxy-2(E)-nonenal. Intolerant subjects were found to have significantly (p = 0.004) higher concentrations of these by-products in their tears (0.85 +/- 1.0 microM; n = 9) compared to tolerant subjects (0.15 +/- 0.15 microM; n = 10). Intolerant subjects (1.86 +/- 0.05 ng/microl; n = 9) had significantly more (p = 0.047) sPLA2 enzyme in their tears compared with tolerant subjects (1.80 +/- 0.08 ng/microl; n = 12) and significantly more enzyme activity (p = 0.012). Intolerant subjects had significantly higher amounts of lipocalin in their tears (2.40 +/- 1.5 microg/microl; n = 10, p0.001) compared to tolerant subjects (0.45 +/- 0.85 microg/microl; n = 13).Changes to the components of the tear film, however small, can disturb the nature and dynamics of the tear film. Increased lipases, degraded lipids and lipocalins in the aqueous tear film potentiates intolerance to contact lens wear and was associated with increased McMonnies dry eye history scores and symptoms scores.

The tear cationic protein lactoferrin increases the activity of various antimicrobial agents against suspended bacterial cultures including Staphylococcus epidermidis, the predominant causative agent of intraocular lens biofilm infections. We investigated the ability of lactoferrin to enhance the activity of vancomycin and lysozyme against biofilms of a clinical S. epidermidis isolate.Biofilms were prepared on soft contact lenses and cells released from the biofilm surface were collected from the surrounding broth. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against the intact biofilm, released cells and suspended bacteria were evaluated using vancomycin with and without lactoferrin.Lactoferrin induced a two-fold reduction in the MBC of vancomycin for biofilm (p0.01) and biofilm-released cells from 64 microg/ml to 32 microg/ml and similarly decreased the MIC for biofilm-released cells (p0.01) from 32 microg/ml to 16 microg/ml. With or without lactoferrin, the MIC of vancomycin for suspended cells (2 and 4 microg/ml respectively) was less (p0.01) than that of both the biofilm (32 and 32 microg/ml) and biofilm-released cells (16 and 32 microg/ml). Lactoferrin did not reduce the MIC of lysozyme, but, at a lysozyme concentration of 16 mg/ml, did significantly (p0.05) reduce the number of viable biofilm and biofilm-released cells.Lactoferrin displays potential as an adjunctive agent to vancomycin in the treatment of S. epidermidis biofilm infections, such as endophthalmitis, associated with intraocular lenses.

Background: Fibronectin enters the tear film during sleep and on reflex tear stimulation. Also, contact lens wearers with increased levels of fibronectin in their tears are more likely to carry gre...

Purpose. The present investigation sought to define the responses of mouse eyes to challenge with three different strains of P. aeruginosa isolated from human corneas or contact lenses: two different strains produced an ulcerative keratitis, and one strain produced contact lens-induced acute red eye (CLARE).Methods. The corneas of BALB/c mice were inoculated with three different strains of P. aeruginosa. The strains were allowed to interact with the corneas for up to 24 h. In addition, strain Paerl, isolated from CLARE, was subjected to in vitro assays to measure its ability to invade corneal epithelial cells, or to produce cytotoxicity in these cells. Both these assays used cultured rabbit corneal epithelial cells.Results. Both MK isolates were able to infect the corneas of mice, but the CLARE isolate was non-infective. The predominant response to infection with the cytotoxic strain was severe corneal edema and infiltration of the corneal stroma with polymorphonuclear leukocytes (PMNs). The predominant r...

Contact lens wear and trauma are the two most common risk factors associated with microbial keratitis (MK). In this issue, Professor Nathan Efron and Dr Phil Morgan present a new way to classify mi...

This study was designed to determine the effect of overnight eye closure on the rate and composition of protein deposition on high water content ionic matrix soft contact lenses (Group IV SCLs) and to extrapolate from this data information on the probable change in the rate of reflex-type tear secretion associated with eye closure.Group IV SCLs were temporally sampled after equivalent periods of wear under closed eye (C) or open eye (O) conditions. Lenses were rinsed in saline and the majority of the tightly bound protein extracted at 90 degrees C in 40% urea, containing 1% SDS, 1 mM DTT, 100 mM Tris-HCl (pH 8.00). Residual protein was determined by Coomassie staining of the extracted lenses and densitometric analysis. Extracted protein was quantitated and separated by SDS-PAGE. Gels were either stained with Coomassie blue or reversibly stained with imidazole-zinc and blotted. Blots were PAS stained, or lectin and antibody probed for glycoproteins, secretory IgA (sIgA), IgG, lysozyme and complement C3. Laboratory simulated deposition studies were carried out on unworn lenses exposed to HPLC purified lysozyme.The protein in the saline rinse, to a large degree mirrored the composition of tear fluid in which the lens had been residing (O or C). This would suggest that the saline wash consists of residual tear fluid and loosely adherent protein. In contrast, the urea extracts were highly homogeneous consisting primarily of lysozyme and to lesser extent lysozyme dimer. This supports the contention that the Group IV SCL functions in the eye much as cationic exchange resin selectively absorbing lysozyme. C extracts also proved relatively enriched in trace amounts of sIgA, IgG and complement C3 and its breakdown products. High levels of C3 and C3 breakdown products were specifically recovered only in the C worn lens extracts from a subject experiencing unilateral contact lens associated corneal infiltrates from the affected eye. In all subjects, markedly less protein (lysozyme) was recovered in urea extracts of lenses exposed to 7-8 h of closed eye as compared to open eye wear (0.20 +/- .08 versus 0.79 +/- .15 mg/lens (n = 6)). Temporal studies further revealed that deposition was linearly related to duration of wear during the initial phase of conditioning film formation giving rise to rate constants for lysozyme deposition of 2.2 +/- 0.29 (n = 5) and 0.20 +/- 0.06 microgram/min (n = 4) under open and closed eye conditions respectively. With further wear, deposition eventually reached a steady state. Under laboratory conditions, lysozyme was much rapidly and quantitatively removed from solution in a manner following a hyperbolic plot. This suggests that during the initial phase of deposition the rate of deposition is limited by the capacity of the tear fluid to deliver lysozyme to the lens surface under these two extremes of conditions.Eye closure profoundly affects the rate of lysozyme deposition on Group IV hydrogels and the composition of minor biofilm constituents in a manner that could affect biocompatibility. Findings support the contention that eye closure results in a90% reduction in the rate of reflex-type tear secretion.

INTRODUCTION: Secretory immunoglobulin A (sIgA) is the predominant immunoglobulin in tears. The role of sIgA in defending the eye against pathogens has not been established clearly. There have been conflicting reports about the effect of contact lens wear on the concentration of sIgA in tears. This study was conducted to elucidate the role of sIgA in ocular defence and to determine the effect of contact lens wear on sIgA concentration. METHODS: Tears were collected from contact lens wearers and non-wearers using micro-capillary tubes. The concentration of sIgA was evaluated using an in-house ELISA. The specificity of sIgA to a strain of P. aeruginosa was examined using a fluorescent assay and the ability of neutrophils to phagocytose sIgA coated bacteria were assessed by plate counts. RESULTS: Tears contained sIgA that reacted to P. aeruginosa. P. aeruginosa coated with sIgA was phagocytosed by the neutrophils. The level of sIgA and the level of sIgA specific to P. aeruginosa in the tears of contact lens wearers were significantly reduced. CONCLUSIONS: These results indicate that contact lens wear significantly alters the level of sIgA in tears which may lead to changes in the ability of the ocular surface to defend itself against potential pathogens.

Streptococcus sanguis FW213 carries peritrichous fimbriae (216±28 nm long) and 6 mutants derived from it lack fimbriae but carry peritrichous fibrils with a mean length of 77–4 + 3–9 nm. Both wild type strain and mutants have a ruthenium red staining layer (≤ 14.5±2.9 nm thick) external to the cell wall at the base of the fibrils and fimbriae. The thickness of this layer is strain dependent. Ruthenium red also stains extracellular masses of material, probably extracellular polysaccharide, but not the fimbriae. S. sanguis strain FW 213 adheres to saliva‐coated hydroxyapatite and buccal epithelial cells and is not aggregated by saliva. The 6 non‐fimbriate mutants of FW213 adhered poorly to hydroxyapatite coated in heated whole saliva (S‐SHA) but 3/6 mutants adhered to the same extent or higher than the wild type to S‐SHA coated in unheated saliva, indicating that strain FW213 may carry a non‐fimbriate adhesin and that whole saliva contains a heat sensitive adhesin. All the mutants had a significantly thinne...

Pubmed and Medline were searched for articles referring to Pseudomonas keratitis between the years 2007 and 2012 to obtain an overview of the current state of this disease. Keyword searches used the terms “Pseudomonas” + “Keratitis” limit to “2007–2012”, and [“Ulcerative” or “Microbial”] + “Keratitis” + “Contact lenses” limit to “2007–2012”. These articles were then reviewed for information on the percentage of microbial keratitis cases associated with contact lens wear, the frequency of Pseudomonas sp. as a causative agent of microbial keratitis around the world, the most common therapies to treat Pseudomonas keratitis, and the sensitivity of isolates of Pseudomonas to commonly prescribed antibiotics. The percentage of microbial keratitis associated with contact lens wear ranged from 0% in a study from Nepal to 54.5% from Japan. These differences may be due in part to different frequencies of contact lens wear. The frequency of Pseudomonas sp. as a causative agent of keratitis ranged from 1% in Japan to over 50% in studies from India, Malaysia, and Thailand. The most commonly reported agents used to treat Pseudomonas keratitis were either aminoglycoside (usually gentamicin) fortified with a cephalosporin, or monotherapy with a fluoroquinolone (usually ciprofloxacin). In most geographical areas, most strains of Pseudomonas sp. (≥95%) were sensitive to ciprofloxacin, but reports from India, Nigeria, and Thailand reported sensitivity to this antibiotic and similar fluoroquinolones of between 76% and 90%.