Comparison of an immortalized human corneal epithelial cell line with Vero cells in the isolation of Herpes simplex virus-1 for the laboratory diagnosis of Herpes simplex keratitis

BACKGROUND: Herpes simplex keratitis (HSK) is a sight threatening ocular infection often requiring a specific and prompt laboratory diagnosis. Isolation of Herpes simplex virus (HSV-1) in culture provides the most reliable and specific method and is considered as the "Gold Standard" in the laboratory diagnosis of HSK in spite of its low sensitivity. Using "cell lines of corneal origin" for virus isolation may be beneficial under such circumstances, since these cells have been shown to be excellent substrates for the growth of HSV-1 isolated from the cornea. We report a comparative study of a novel human corneal epithelial cell line (HCE) and the Vero cell line in the isolation of HSV-1 from corneal scrapings employing a shell vial assay. METHODS: Corneal scrapings were obtained from 17 patients with a clinical diagnosis of HSK. All the cases were confirmed by virological investigations (PCR and viral antigen detection positive, n = 15, PCR positive, n = 1, Viral antigen positive, n = 1). Scrapings obtained from 10 patients with infectious keratitis of non-viral origin were included as controls. All the scrapings were simultaneously inoculated into shell vials of HCE and Vero cells. Cultures were terminated at 24 h post-infection. Isolation of HSV-1 was confirmed using an indirect immunofluorescence/ immunoperoxidase assay. RESULTS: Virus could be isolated using both or either of the cell lines in 10/17 (58.82%) patients with HSK. HSV-1 was isolated from 10/ 17 (58.82%) and 4/17(23.52%) specimens in HCE and Vero cells, respectively (P = 0.036). None of the controls yielded HSV-1. While all the 10 (100%) strains were isolated in HCE, Vero yielded only 4/10 (40%) strains in the shell vial culture (P = 0.014). CONCLUSIONS: HCE showed a statistically significant difference in the virus isolation rate with respect to Vero cells. HCE may be an excellent alternative cell line for the isolation of HSV-1, especially from corneal scrapings, for the laboratory diagnosis of HSK

Serratamolide is a hemolytic factor produced by Serratia marcescens

Serratia marcescens is a common contaminant of contact lens cases and lenses. Hemolytic factors of S. marcescens contribute to the virulence of this opportunistic bacterial pathogen. We took advantage of an observed hyper-hemolytic phenotype of crp mutants to investigate mechanisms of hemolysis. A genetic screen revealed that swrW is necessary for the hyper-hemolysis phenotype of crp mutants. The swrW gene is required for biosynthesis of the biosurfactant serratamolide, previously shown to be a broad-spectrum antibiotic and to contribute to swarming motility. Multicopy expression of swrW or mutation of the hexS transcription factor gene, a known inhibitor of swrW expression, led to an increase in hemolysis. Surfactant zones and expression from an swrW-transcriptional reporter were elevated in a crp mutant compared to the wild type. Purified serratamolide was hemolytic to sheep and murine red blood cells and cytotoxic to human airway and corneal limbal epithelial cells in vitro. The swrW gene was found in the majority of contact lens isolates tested. Genetic and biochemical analysis implicate the biosurfactant serratamolide as a hemolysin. This novel hemolysin may contribute to irritation and infections associated with contact lens use. © 2012 Shanks et al

Validity and reliability of a modified english version of the physical activity questionnaire for adolescents

BACKGROUND: Adaptation of physical activity self-report questionnaires is sometimes required to reflect the activity behaviours of diverse populations. The processes used to modify self-report questionnaires though are typically underreported. This two-phased study used a formative approach to investigate the validity and reliability of the Physical Activity Questionnaire for Adolescents (PAQ-A) in English youth. Phase one examined test content and response process validity and subsequently informed a modified version of the PAQ-A. Phase two assessed the validity and reliability of the modified PAQ-A. METHODS: In phase one, focus groups (n = 5) were conducted with adolescents (n = 20) to investigate test content and response processes of the original PAQ-A. Based on evidence gathered in phase one, a modified version of the questionnaire was administered to participants (n = 169, 14.5 ± 1.7 years) in phase two. Internal consistency and test-retest reliability were assessed using Cronbach’s alpha and intra-class correlations, respectively. Spearman correlations were used to assess associations between modified PAQ-A scores and accelerometer-derived physical activity, self-reported fitness and physical activity self-efficacy. RESULTS: Phase one revealed that the original PAQ-A was unrepresentative for English youth and that item comprehension varied. Contextual and population/cultural-specific modifications were made to the PAQ-A for use in the subsequent phase. In phase two, modified PAQ-A scores had acceptable internal consistency (α = 0.72) and test-retest reliability (ICC = 0.78). Modified PAQ-A scores were significantly associated with objectively assessed moderate-to-vigorous physical activity (r = 0.39), total physical activity (r = 0.42), self-reported fitness (r = 0.35), and physical activity self-efficacy (r = 0.32) (p ≤ 0.01). CONCLUSIONS: The modified PAQ-A had acceptable internal consistency and test-retest reliability. Modified PAQ-A scores displayed weak-to-moderate correlations with objectively measured physical activity, self-reported fitness, and self-efficacy providing evidence of satisfactory criterion and construct validity, respectively. Further testing with more diverse English samples is recommended to provide a more complete assessment of the tool. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13690-016-0115-2) contains supplementary material, which is available to authorized users

Comparison of the sensitivity of a 24 h-shell vial assay, and conventional tube culture, in the isolation of Herpes simplex virus – 1 from corneal scrapings

BACKGROUND: Herpes simplex keratitis is a sight threatening ocular infection. A rapid and specific diagnosis is essential for the institution of specific antiviral therapy and to avoid complications that can arise from misdiagnosis and inappropriate treatment. Though a variety of techniques are available, isolation of Herpes simplex virus 1 (HSV-1) in culture provides the most reliable and specific method, and is considered as the gold standard in laboratory diagnosis of herpes simplex keratitis. We report a comparative study of the sensitivity of a 24 h-shell vial assay and conventional tube culture in the isolation of HSV-1 from corneal scrapings. METHODS: A total of 74 corneal scrapings obtained from 74 patients with a clinical suspicion of herpes simplex keratitis submitted for the isolation of HSV-1, were simultaneously inoculated into shell vial and tube cultures employing the vero cell line. Shell vial and tube cultures were terminated at 24 h and fifth day respectively. Isolation of HSV-1 was confirmed employing an indirect immunofluorescence assay. RESULTS: HSV-1 was isolated from 24/74 (32.4%) specimens employing both the methods. Sensitivity of both the techniques were found to be similar (20/24, 83.3%) (P = 1.0). CONCLUSION: A 24 h-shell vial assay is a rapid alternative technique in comparison to the time consuming conventional tube cultures for the isolation of HSV-1, especially from corneal scrapings for the laboratory diagnosis of herpes simplex keratitis

Oral cancer treatment costs in Greece and the effect of advanced disease

BACKGROUND: The main purpose of the study was to quantify the direct costs of oral cancer treatment to the healthcare system of Greece. Another aim was to identify factors that affect costs and potential cost reduction items. More specifically, we examined the relationship between stage of disease, modality of treatment and total direct costs. METHODS: The medical records and clinic files of the Oral and Maxillofacial Clinic of the Athens General Hospital "Genimatas" were abstracted to investigate clinical treatment characteristics, including length of hospitalization, modes of treatment, stage of disease etc. Records of 95 patients with oral squamous cell carcinoma (OSSC), with at least six months of follow-up, were examined. The clinical data was then used to calculate actual direct costs, based on 2001 market values. RESULTS: The mean total direct costs for OSSC treatment estimated at euro 8,450 or approximately US$ 7,450. Costs depended on the stage of the disease, with significant increases in stages III and IV, as compared with stages I and II (p < 0.05). Multi-modality treatment applied mainly to patients in stages III and IV was the factor that affected the cost. Disease stage was also associated with the total duration of hospitalization (p < 0.05). CONCLUSIONS: The clinical management of advanced oral cancer is strongly associated with higher costs. Although the ideal would be to prevent cancer, the combination of high-risk screening, early diagnosis and early treatment seems the most efficient way to reduce costs, and most importantly, prolong life

Microparticle-mediated transfer of the viral receptors CAR and CD46, and the CFTR channel in a CHO cell model confers new functions to target cells

Cell microparticles (MPs) released in the extracellular milieu can embark plasma membrane and intracellular components which are specific of their cellular origin, and transfer them to target cells. The MP-mediated, cell-to-cell transfer of three human membrane glycoproteins of different degrees of complexity was investigated in the present study, using a CHO cell model system. We first tested the delivery of CAR and CD46, two monospanins which act as adenovirus receptors, to target CHO cells. CHO cells lack CAR and CD46, high affinity receptors for human adenovirus serotype 5 (HAdV5), and serotype 35 (HAdV35), respectively. We found that MPs derived from CHO cells (MP-donor cells) constitutively expressing CAR (MP-CAR) or CD46 (MP-CD46) were able to transfer CAR and CD46 to target CHO cells, and conferred selective permissiveness to HAdV5 and HAdV35. In addition, target CHO cells incubated with MP-CD46 acquired the CD46-associated function in complement regulation. We also explored the MP-mediated delivery of a dodecaspanin membrane glycoprotein, the CFTR to target CHO cells. CFTR functions as a chloride channel in human cells and is implicated in the genetic disease cystic fibrosis. Target CHO cells incubated with MPs produced by CHO cells constitutively expressing GFP-tagged CFTR (MP-GFP-CFTR) were found to gain a new cellular function, the chloride channel activity associated to CFTR. Time-course analysis of the appearance of GFP-CFTR in target cells suggested that MPs could achieve the delivery of CFTR to target cells via two mechanisms: the transfer of mature, membrane-inserted CFTR glycoprotein, and the transfer of CFTR-encoding mRNA. These results confirmed that cell-derived MPs represent a new class of promising therapeutic vehicles for the delivery of bioactive macromolecules, proteins or mRNAs, the latter exerting the desired therapeutic effect in target cells via de novo synthesis of their encoded proteins

Small Polarons in Transition Metal Oxides

The formation of polarons is a pervasive phenomenon in transition metal oxide compounds, with a strong impact on the physical properties and functionalities of the hosting materials. In its original formulation the polaron problem considers a single charge carrier in a polar crystal interacting with its surrounding lattice. Depending on the spatial extension of the polaron quasiparticle, originating from the coupling between the excess charge and the phonon field, one speaks of small or large polarons. This chapter discusses the modeling of small polarons in real materials, with a particular focus on the archetypal polaron material TiO2. After an introductory part, surveying the fundamental theoretical and experimental aspects of the physics of polarons, the chapter examines how to model small polarons using first principles schemes in order to predict, understand and interpret a variety of polaron properties in bulk phases and surfaces. Following the spirit of this handbook, different types of computational procedures and prescriptions are presented with specific instructions on the setup required to model polaron effects.Comment: 36 pages, 12 figure

External Ocular Surface Bacterial Isolates and their Antimicrobial Susceptibility Patterns among Pre-operative Cataract Patients at Mulago National Hospital in Kampala, Uganda.

Endophthalmitis is a severe complication of cataract surgery which leads to high ocular morbidity and visual loss even with antibiotic treatment. Bacterial ocular floras are the implicated causative agents. This study was undertaken to evaluate the external ocular surface bacterial isolates and their antimicrobial susceptibility patterns among pre-operative cataract patients at Mulago National Hospital. This cross sectional study enrolled consecutively 131 patients scheduled for routine cataract surgery in the Department of Ophthalmology at Mulago National Hospital in Kampala, Uganda. Eyelid margin and conjunctival swabs were collected and processed using standard microbiological procedures to identify bacterial isolates and their respective antimicrobial susceptibility patterns. Of 131 patients involved (mean age 63.3 ± 14.5 years), 54.2% (71/131) were females. The eyelid margin and conjunctival samples were culture positive in 59.5% (78/138) and 45.8% (60/138) respectively. The most common organisms identified were Coagulase-negative Staphylococci (CoNS) [65.9% (91/138)] and Staphylococcus aureus [21.0% (29/138)]. CoNS showed the highest resistance to tetracycline (58.2%, 53/91) and erythromycin (38.5%, 35/91), whereas in S. aureus the resistance to tetracycline and erythromycin were 55.2% (16/29) and 31.0% (9/29) respectively. Methicillin resistant CoNS (MRS) and Methicillin resistance S. aureus (MRSA) were 31.9% (29/91) and 27.6% (8/29) respectively. There were low resistance rates for CoNS, S. aureus and other bacterial isolates to ciprofloxacin (11.1%-24.2%), gentamicin (5.6-31.0%), tobramycin (17.2% -25.3%) and vancomycin (0.0%). CoNS and S. aureus are the most common bacterial isolates found on the external ocular surface of the pre-operative cataract patients. Ciprofloxacin, gentamicin, tobramycin and vancomycin showed the lowest resistance rates to all bacterial isolates, therefore may be used to reduce bacteria load in the conjunctiva sac among cataract patients prior to surgery