Optical Density Index Research Articles

Immunomodulatory activity of aqueous extract of Nyctanthes arbor-tristis flowers with particular reference to splenocytes proliferation and cytokines induction.

Objectives:To investigate the immunomodulatory activity of aqueous extract of Nyctanthes arbor-tristis flowers (NAFE) with particular reference to splenocytes proliferation and induction of cytokines.Materials and Methods:Antibody titer was determined by tube agglutination and indirect ELISA assay in four groups of mice-control, antigen alone, and NAFE-treated (400 and 800 mg/kg for 21 days) after immunization with Salmonella antigen while cellular immunity was studied in three groups of rats (control and NAFE-treated - 400 and 800 mg/kg) following DNCB application. Splenocytes from untreated and NAFE-treated rats were stimulated using concanavalin-A (Con-A) and optical density (OD) and stimulation index were determined. Splenocytes from control rats were also treated in vitro with NAFE (50–1600 μg/ml) and Con-A to determine the effect on splenocytes proliferation. Interleukin-2 (IL-2) and IL-6 levels in splenocytes supernatant from control and NAFE-treated rats and following in vitro treatment of splenocytes with NAFE (50–1600 μg/ml) were determined using ELISA kits.Results:Marked to a significant increase in antibody titer by both the methods in NAFE-treated mice and a significant increase in skin thickness in rats after challenge with DNCB, respectively suggested humoral and cell-mediated immunostimulant potential of NAFE. Significant increase in OD and stimulation index following e x vivo and in vitro exposure of splenocytes and sensitization with Con-A and significant elevation in IL-2 and IL-6 levels in splenocytes supernantant was also observed after their ex vivo and in vitro exposure to NAFE.Conclusion:Humoral and cell-mediated immunostimulant activity of NAFE seems to be mediated through splenocytes proliferation and increased production of cytokines, especially IL-2 and IL-6.

Galactomannan detection for invasive aspergillosis in immunocompromised patients.

Invasive aspergillosis is the most common life-threatening opportunistic invasive mycosis in immunocompromised patients. A test for invasive aspergillosis should neither be too invasive nor too great a burden for the already weakened patient. The serum galactomannan enzyme-linked immunosorbent assay (ELISA) seems to have the potential to meet both requirements. To obtain summary estimates of the diagnostic accuracy of galactomannan detection in serum for the diagnosis of invasive aspergillosis. We searched MEDLINE, EMBASE and Web of Science with both MeSH terms and text words for both aspergillosis and the sandwich ELISA. We checked the reference lists of included studies and review articles for additional studies. We conducted the searches in February 2014. We included cross-sectional studies, case-control designs and consecutive series of patients assessing the diagnostic accuracy of galactomannan detection for the diagnosis of invasive aspergillosis in patients with neutropenia or patients whose neutrophils are functionally compromised. The reference standard was composed of the criteria given by the European Organization for Research and Treatment of Cancer (EORTC) and the Mycoses Study Group (MSG). Two review authors independently assessed quality and extracted data. We carried out meta-analysis using the bivariate method. We investigated sources of heterogeneity by adding potential sources of heterogeneity to the model as covariates. We included 54 studies in the review (50 in the meta-analyses), containing 5660 patients, of whom 586 had proven or probable invasive aspergillosis. When using an optical density index (ODI) of 0.5 as a cut-off value, the sensitivity of the test was 82% (73% to 90%) and the specificity was 81% (72% to 90%). At a cut-off value of 1.0 ODI, the sensitivity was 72% (65% to 80%) and the specificity was 88% (84% to 92%). At a cut-off value of 1.5 ODI, the sensitivity was 61% (47% to 75%) and the specificity was 93% (89% to 97%). None of the potential sources of heterogeneity had a statistically significant effect on either sensitivity or specificity. If we used the test at a cut-off value of 0.5 ODI in a population of 100 patients with a disease prevalence of 9% (overall median prevalence), two patients who have invasive aspergillosis would be missed (sensitivity 82%, 18% false negatives), and 17 patients would be treated unnecessarily or referred unnecessarily for further testing (specificity 81%, 19% false negatives). If we used the test at a cut-off value of 1.5 in the same population, that would mean that four invasive aspergillosis patients would be missed (sensitivity 61%, 39% false negatives), and six patients would be treated or referred for further testing unnecessarily (specificity 93%, 7% false negatives). These numbers should, however, be interpreted with caution because the results were very heterogeneous.

Plasmalyte: No Longer a Culprit in Causing False-Positive Galactomannan Test Results.

False-positive galactomannan (GM) results have been reported in patients treated with gluconate-containing solutions, such as Plasmalyte. The GM optical density index was tested on 33 distinct batches of Plasmalyte and was found to be negative in all of the batches, confirming that Plasmalyte is no longer a cause of false-positive GM results.

Diagnosis of Invasive Aspergillosis: Use of the Galactomannan Assay

Invasive aspergillosis is the cause of severe morbidity and mortality in immunocompromised patients. Given the challenges of fungal cultures, non-culture surrogates are crucial to the timely diagnosis of invasive aspergillosis (IA) to initiate expedited treatment. The Platelia™ Aspergillus EIA (Bio-Rad, Hercules, California) is a double-sandwich ELISA that detects the galactomannan (GM) of the fungal cell wall and was cleared by the FDA for use in serum and bronchoalveolar fluid (BAL) in 2003 and 2011, respectively. The population in which GM has been studied the most and has shown the greatest accuracy is that of hematologic malignancy. The optimal optical density index (ODI) cutoff to define test positivity in the serum is still a matter of debate because this value influences test performance. Using a lower ODI threshold (≥0.5 vs. ≥1 vs. ≥1.5) optimizes sensitivity at the expense of specificity and vice versa using a higher ODI threshold. One must be alert to the potential for false-positive results, particularly if a ODI cutoff of 0.5 is used in the serum. False positives can occur due to medications, i.e., piperacillin/tazobactam, though there is increasing evidence that newer formulations are less cross-reactive with GM, and false-positive results occur in the presence of other molds that cross-react with GM. As the use of mold-active antifungal prophylaxis increases, one must be aware that GM may not perform as well due to lower pre-test probability of IA and lower test sensitivity. Emerging evidence indicates that use of GM in combination with other tests, e.g., Aspergillus PCR or lateral flow device (LFD), may enhance diagnostic accuracy beyond GM alone; however, further validation of these diagnostics in combination are required before routine implementation can be recommended. BAL GM performs better than serum as it is significantly more sensitive, though the optimal ODI cutoff is also debated (≥0.5 vs. ≥1).False-positive results can be due to use of medications, as with serum GM. False negatives can occur with the use of certain agents which decrease the viscosity of the BAL fluid, and use of such agents need to be considered by the clinician when evaluating a test result. Again, BAL GM, in conjunction with other tests, e.g., PCR and LFD, are promising, but further studies are needed. GM in other fluids, i.e., CSF, urine, and tissue, may be useful, but the studies are very limited. In summary, when employed in the right clinical context and interpreted appropriately, serum and BAL GM can facilitate the diagnosis and early treatment of IA. While there are significant limitations and the landscape is evolving, the test has an important role in clinical practice today.

Correlation between Circulating Fungal Biomarkers and Clinical Outcome in Invasive Aspergillosis.

Objective means are needed to predict and assess clinical response in patients treated for invasive aspergillosis (IA). We examined whether early changes in serum galactomannan (GM) and/or β-D-glucan (BDG) can predict clinical outcomes. Patients with proven or probable IA were prospectively enrolled, and serial GM and BDG levels and GM optical density indices (GMI) were calculated twice weekly for 6 weeks following initiation of standard-of-care antifungal therapy. Changes in these biomarkers during the first 2 and 6 weeks of treatment were analyzed for associations with clinical response and survival at weeks 6 and 12. Among 47 patients with IA, 53.2% (25/47) and 65.9% (27/41) had clinical response by weeks 6 and 12, respectively. Changes in biomarkers during the first 2 weeks were associated with clinical response at 6 weeks (GMI, P = 0.03) and 12 weeks (GM+BDG composite, P = 0.05; GM, P = 0.04; GMI, P = 0.02). Changes in biomarkers during the first 6 weeks were also associated with clinical response at 6 weeks (GM, P = 0.05; GMI, P = 0.03) and 12 weeks (BDG+GM, P = 0.02; GM, P = 0.02; GMI, P = 0.01). Overall survival rates at 6 weeks and 12 weeks were 87.2% (41/47) and 79.1% (34/43), respectively. Decreasing biomarkers in the first 2 weeks were associated with survival at 6 weeks (BDG+GM, P = 0.03; BDG, P = 0.01; GM, P = 0.03) and at 12 weeks (BDG+GM, P = 0.01; BDG, P = 0.03; GM, P = 0.01; GMI, P = 0.007). Similar correlations occurred for biomarkers measured over 6 weeks. Patients with negative baseline GMI and/or persistently negative GMI during the first 2 weeks were more likely to have CR and survival. These results suggest that changes of biomarkers may be informative to predict and/or assess response to therapy and survival in patients treated for IA.

Generic piperacillin/tazobactam is not associated with galactomannan false-positivity in adult patients with cancer: a case-control study.

Recent products of piperacillin/tazobactam (PTZ) from the original manufacturer, previously considered a major cause of galactomannan (GM) false-positivity, are reported not to be related to it. However, data regarding generic PTZ are limited and controversial. To evaluate the effect of generic PTZ on GM false-positivity in Korea, we performed a case-control study in adult patients with cancer. A case-control study was designed. Electronic medical records of cancer patients who were admitted and tested for serum GM between March and June 2014 at a tertiary care university hospital were reviewed. During the study period, a single generic PTZ (C manufacturer, Korea) was used. Patients who received PTZ within 24h prior to serum GM testing were enrolled. Age- and GM test date-matched non-PTZ patients were selected as controls. A total of 110 patients received PTZ within 24h prior to serum GM testing during the study period. The GM optical density index (ODI) of the PTZ group did not vary significantly from that of the control group (p = 0.251). The percentage of false-positive patients in the PTZ group was also similar to that of the control group (p = 0.538). There was no statistical relationship between GM ODI titer and time interval from PTZ administration (p = 0.095) or cumulative PTZ dose (p = 0.416). In a case-control study that evaluated 220 patients, a generic PTZ in Korea was not related to GM false-positivity.

Improved Standardization of the Bio-Rad Platelia Aspergillus Galactomannan Antigen Sandwich Enzyme Immunoassay Using the DS2 (Dynex) Enzyme-Linked Immunosorbent Assay (ELISA) Processing System

The galactomannan enzyme immunoassay (GM-EIA) is widely utilized for the diagnosis of invasive aspergillosis (IA). There is inconsistent reproducibility of results between centers when the assay is processed manually. Automation of EIAs can reduce variation. This study investigated the semiautomation of the GM-EIA on the DS2 (Dynex) platform in the following three stages: (i) DS2 GM-EIA method validation with experimental samples, (ii) DS2 retesting of case-defined clinical samples, and (iii) a 12-month audit of DS2 GM-EIA performance. In stage i, Bland-Altman analysis demonstrated a reduced variance between optical density index (ODI) values for samples processed on two DS2 platforms (mean difference, -0.02; limits of agreement [LOA], -0.19 to 0.14) compared with the variance between samples processed manually and on a DS2 platform (mean difference, 0.02; LOA, -0.25 to 0.3). In stage ii, 100% (14/14 samples) qualitative agreement was observed for serum samples from patients with IA, with no significant change in the ODI values when samples were processed on the DS2 platform. A significant decrease in ODI values was observed for control serum samples on the DS2 platform (difference, 0.01; P = 0.042). In stage iii, a significant reduction in the frequency of equivocal results, from 5.56% (136/2,443 samples) to 1.56% (15/961 samples), was observed after DS2 automation (difference, 4.0%; 95% confidence interval [CI], 2.7 to 5.2%; P < 0.01), with an equivalent increase in negative results. This study demonstrates that GM-EIA automation may reduce intersite variability. Automation does not have an impact on the repeatability of truly positive results but contributes to a reduction in false-positive (equivocal) GM-EIA results, reducing the need to retest a significant proportion of samples.

Some newer marker phytoconstituents in methanolic extract of Moringa oleifera leaves and evaluation of its immunomodulatory and splenocytes proliferation potential in rats.

Objectives:The present study was undertaken to unravel the newer marker phytoconstituents in methanolic extract of Moringa oleifera leaves (MOLE) and evaluation of its immunomodulatory and splenocytes proliferation potential in rats.Materials and Methods:Hot methanolic extract of MOLE was subjected to gas chromatography-mass spectrometry (GC-MS) analysis. Immunomodulatory potential was studied in four groups of rats following administration of MOLE at 62.5 and 125 mg/kg for 21 days, followed by immunization with Salmonella typhimurium “O” antigen and antibody titer determined using indirect enzyme-linked immunosorbent assay kit. Total lymphocytes and T- and B-lymphocytes count were determined in control and after MOLE administration (62.5 and 125 mg/kg) to rats for 42 days. Splenocytes (2 × 106 spleen cells/ml) from MOLE treated rats were harvested and stimulated using concanavalin A and optical density (OD) and stimulation index were determined. Splenocytes from healthy control rats were also collected and treated in vitro with different concentrations of MOLE (5, 10, 25, 50, and 100 µg/ml) and concanavalin A to determine effect of MOLE on OD and stimulation index.Results:GC-MS analysis revealed presence of 9,12,15-octadecatrienoic acid ethyl ester, 6-octadecenoic acid, cis-vaccenic acid and 2-octyl-cyclopropaneoctanal in MOLE. MOLE at 125 mg/kg increased the antibody titer by 50%. Although there was slight decline in lymphocytes count (total, B- and T-lymphocytes) in MOLE treated rats, percentage of T-lymphocytes was increased nonsignificantly. Ex vivo and in vitro studies revealed marked increase in OD and stimulation index indicating MOLE-induced splenocytes proliferation.Conclusion:GC-MS study revealed four new compounds in MOLE apart from promising its immunomodulatory potential based on humoral immune response, percentage increase in T-lymphocytes count, and induction of splenocytes proliferation.

Serum and urine galactomannan testing for screening in patients with hematological malignancies

Testing for serum galactomannan (GM) has been established as an important method for diagnosing invasive aspergillosis (IA); however, limited data exist regarding the application of urine GM testing. The objective of this study was to evaluate the performance of GM screening of urine specimens and to compare results with serum GM. The study was performed between July 2012 and March 2013 in adult patients with underlying hematological malignancies who were hospitalized at the Medical University of Graz, Austria. Serum and urine screening samples were collected and tested twice weekly (always on the same day). In total, 242 serum samples and a similar number of urine samples were collected from 75 patients. A total of 21/242 (8.7%) serum samples from 13 patients were GM positive. Sensitivity, specificity, positive predictive value, and negative predictive value using a 0.1 optical density index cutoff for urine samples (compared with same-day serum results) were as follows: 47.6%, 86%, 24.4%, and 94.5%, respectively. In 8/10 patients with probable IA, at least one positive GM result was found with this cutoff. After calculating clinical performance of the urine GM test, we found that sensitivity increased to 71.4% and specificity to 88.2%. Spearman-Rho correlation analysis revealed a significant positive correlation between serum and urine samples (P<0.001; ρ = 0.252). In conclusion, GM detection in urine might be a promising method for IA screening. However, further studies are needed.

Performance of Galactomannan, Beta- d -Glucan, Aspergillus Lateral-Flow Device, Conventional Culture, and PCR Tests with Bronchoalveolar Lavage Fluid for Diagnosis of Invasive Pulmonary Aspergillosis

Galactomannan detection in bronchoalveolar lavage (BAL) fluid samples (GM test) is currently considered the gold standard test for diagnosing invasive pulmonary aspergillosis (IPA). The limitations, however, are the various turnaround times and availability of testing. We compared the performance of GM testing with that of conventional culture, an Aspergillus lateral-flow-device (LFD) test, a beta-d-glucan (BDG) test, and an Aspergillus PCR assay by using BAL fluid samples from immunocompromised patients. A total of 78 BAL fluid samples from 78 patients at risk for IPA (74 samples from Graz and 4 from Mannheim) collected between December 2012 and May 2013 at two university hospitals in Austria and Germany were included. Three patients had proven IPA, 14 probable IPA, and 17 possible IPA, and 44 patients had no IPA. The diagnostic accuracies of the different methods for probable/proven IPA were evaluated. The diagnostic odds ratios were the highest for the GM, PCR, and LFD tests. The sensitivities for the four methods (except culture) were between 70 and 88%. The combination of the GM (cutoff optical density index [ODI], >1.0) and LFD tests increased the sensitivity to 94%, while the combination of the GM test (>1.0) and PCR resulted in 100% sensitivity (specificity for probable/proven IPA, 95 to 98%). The performance of conventional culture was limited by low sensitivity, while that of the BDG test was limited by low specificity. We evaluated established and novel diagnostic methods for IPA and found that the Aspergillus PCR, LFD, and GM tests were the most useful methods for diagnosing the disease by using BAL fluid samples. In particular, the combination of the GM test and PCR or, if PCR is not available, the LFD test, allows for sensitive and specific diagnosis of IPA.

The effects of erythropoietin on STAT1 and STAT3 levels following cerebral ischemia-reperfusion in rats

Objective To explore the effects of erythropoietin （EPO） on the expression of signal transducer and activator of transcription （STAT） 1, phosphorylated STAT1 （ P-STAT1 ）, STAT3, P-STAT3 and cell apoptosis in rat models of focal cerebral ischemia-reperfusion. Methods Eighty male Sprague- Dawley rats were randomly and evenly divided into four groups by completely random design method： shamoperation （group A ）, cerebral isehemia-reperfusion （group B ）, cerebral ischemia-reperfusion ＋ saline （group C） and cerebral ischemia-reperfusion ＋ EPO （group D）. The model of focal cerebral ischemiareperfusion injury was established by blocking the left middle cerebral artery. All rats underwent MRI for the detection of the changes of infarct area between 2 h post ischemia and 24 h of reperfusion. Western blot was used to observe the expression of STAT1, P-STAT1, STAT3, P-STAT3. Terminal oxynucleotidyl transferase mediated dUTP biotin nick end labeling （TUNEL） was used to evaluate the cell apoptosis including the relative area （ ROI area/whole brain area of the same layer x 100% ） of abnormal signal region, relative optical density （rOD） and apoptotic index. One-way analysis of variance and q test were used to analyze the data. Results On T2WI imaging, rats in group B and group C presented large hyperintense areas in the cortex and subcortex of left hemispheric （ （28.00±4.60） % and （29.70±4.80） % respectively）. Group D presented less hyperintense areas in the cortex and subcortex of left hemispheric compared with group B and group C （（21.10±2. 40）%; F= 11.285, P〈0.01 ）. The expression of STAT1 and STAT3 proteins was not significandy affected by ischemia-reperfusion and EPO intervention compared with normal brain tissue （ F= 0.806, 1.558, both P〉0.05）. However, the level of P-STAT1 was low in group A （rOD = 0.75±0.13） but increased after cerebral ischemia-reperfusion. Compared with group B and group C, P-STAT1 expression was lower in group D （B-D： 2. 08±0.15, 2.05±0.16, 1.92±0.05; F= 3.274, P〉0.05）. The level of P-STAT3 was also low in group A （rOD= 1.02＋0.09）. Compared with group B and group C, P-STAT3 expression in group D was significantly higher （ B- D ： 2.22 ± 0.13, 2.04 ± 0.14, 4.21± 0.21 ; F = 40.719, P 〈 0.01 ）. The apoptotic index of group B and group C was （42.00±1.30）% and （41.20±2.50）%, respectively, which was significantly higher than that of group D （ （20. 90±1.46）%; F= 378.704, P〈0.01 ）. Conclusion Intraperitoneal injection of EPO can reduce the cerebral ischemic area and the number of apoptotic cells in the ischemic penumbra in rat ischemia-reperfusion models through increasing P-STAT3 and decreasing P-STAT1 levels. Key words: Cerebral ischemia; Reperfusion; STAT transcription factors; Erythropoietin; Magnetic resonance imaging; Rats

Comparative evaluation of galactomannan optical density indices and culture results in bronchoscopic specimens obtained from neutropenic and non‐neutropenic patients

Aspergillus infections are major causes of morbidity and mortality among immunocompromised patients. This study was designed to investigate the galactomannan assay optical density (OD) indices relative to the culture results in bronchoscopic samples obtained from neutropenic and non-neutropenic patients. Galactomannan OD indices from 1427 samples from 2005 to 2012, which were sent from 839 patients and were composed of bronchial lavage (BL=727) and bronchoalveolar lavage fluids (BAL=700), were retrospectively analysed. The recovery rates of Aspergillus species from these specimens were 9.4% from the combined patient group and 13.3% from the neutropenic group. Aspergillus fumigatus complex was the most frequently isolated species. The mean and median OD indices of the positive and negative culture samples are approximately 5 and 1, respectively, and 91% of all culture-positive samples have ≥1 OD index value. The receiver-operating characteristics curve analysis demonstrated that the feasibility of the Aspergillus galactomannan assay and Aspergillus galactomannan test has superior accuracy in BAL compared to BL fluids, and the test is not affected by the immune status of the patient. We suggest that the Aspergillus galactomannan test, which uses bronchoscopic material, leads to an earlier diagnosis and if the OD index is found ≥1, fungal growth can be expected.

Comparative study on hepatic toxicity of Gardeniae Fructus and Huanglian Jiedu decoction

To observe the effect of ingredients in Huanglian Jiedu decoction (HLJDT) combined with Gardeniae Fructus on the hepatic toxicity of Gardeniae Fructus and its mechanism. Rats were given Gardeniae Fructus and HLJDT decoction at the dose of 10 times of clinical dosage for 3 days. Their ALT AST, ALP, TBA were detected, and their liver weight index was calculated. SOD activity, MDA content, GSH-PX activity, TNF-alpha content in hepatic tissues were determined. The cell apoptosis in liver tissue was determined by TUNEL, and the expressions of apoptosis related proteins Bax, Bcl-2 were measured by immunohistochemical method. Compared with the normal control group, the Gardeniae Fructus group showed significant increase in the liver weight index, ALT, AST, TBA and ALP, notable decrease in SOD, SOD/MDA and GSH-PX, and remarkable rise in MDA, TNF-a concentration, accumulated optical density, apoptosis index, Bax and Bax/Bcl-2. Compare with that in the Gardeniae Fructus group, the liver index, ALT, AST, TBA, ALP reduced obviously; SOD, SOD/MDA and GSH-PX markedly increased; MDA and TNF-alpha significantly reduced; the accumulated optical density and apoptosis index significantly reduced; and Bax/Bcl-2 was much lower in HLJDT group. The hepatic toxicity caused by Gardeniae Fructus may be related to inflammation, oxidative stress-induced hepatocyte necrosis and apoptosis. Other ingredients in HLJDT, apart from Gardeniae Fructus, can decrease the hepatic toxicity caused by Gardeniae Fructus by increasing the enzyme activity eliminating radicals and inhibiting hepatocyte injury caused by inflammatory reaction against Gardeniae Fructus.

The retrospective study of serum aspergillus galactomannan (GM) antigen assay in invasive aspergillosis on hematological diseases

To explore the relationship between the optical density index of serum aspergillus galactomannan (GM) assay and invasive aspergillosis (IA). From Jan 2008 to Dec 2011, 825 hematological diseases patients with neutrophil count <0.5×10⁹/L⁹ by continuous blood count tests were admitted into our hospital. The optical density index of GM assay was ≥0.5 at least once. Of 825 patients, 247 cases were manifested as fever during hospitalization. The optical density index of GM antigen was detected by enzyme-linked immunosorbent assay, and the sensitivity and specificity of optical density ranged in 0.5-1.5. In this study, the sensitivity and specificity of GM assay with continuous twice samples (73% and 93%, respectively) were higher than single sample (66% and 80%, respectively) when optical density index ≥1.0. 69 cases were diagnosed as proven IA with the incidence rate of 8.36%. The cut-off level for serum GM antigen assay should be decided as optical density index in two continuous samples of ≥1.0.

Serum galactomannan assay for diagnosis of probable invasive Aspergillosis in acute leukemia and hematopoietic stem cell transplantation

Background:Invasive aspergillosis (IA) is a leading cause of mortality in acute leukemia and hematopoietic stem cell transplantation (HSCT).Aims:To determine the yield of galactomannan (GM) assay for the diagnosis of probable IA, its temporal relationship with the computed tomography (CT) scans and correlation with mortality in AL and HSCT.Patients and Methods:Consecutive neutropenic episodes (n=150) among inpatients aged ≥15 years with AL or recipients of HSCT were prospectively evaluated over 1½ years. All patients underwent weekly serum GM assay and optical density index >0.5 for ≥2 samples was defined as positive. IA was diagnosed according to EORTC 2008 guidelines.Results:Of the 150 episodes enrolled, 43 (28.7%) were diagnosed with IA: possible 25 (16.7%), probable 17 (11.3%) and proven 1 (0.7%). The yield of GM assay in diagnosing probable IA was 17/42 (40.5%). In 88.2% of probable IA episodes, GM was positive before high-resolution CT at a median of 10 days (range 1-16). In the episodes with ≥2 samples tested, fatality was higher in those ≥2 values positive for GM, compared to the rest (31% vs. 13.2%, odd ratio 2.96, 95% CI 1.09-8.00; P=0.04).Conclusions:In AL and HSCT, GM assay could identify patients with probable IA earlier than CT chest and also predicted a higher risk of death.

Macrophage migration inhibitory factor in head and neck squamous cell carcinoma: clinical and experimental studies

The present in vivo/in vitro study was undertaken in order to evaluate the importance of macrophage migration inhibitory factor (MIF) in the progression of head and neck squamous cell carcinoma (HNSCC). Tumor tissue expression (MIF immunostaining) and plasma levels (ELISA) of MIF were determined in HNSCC patients and correlated with tumor recurrence and metastasis, and overall survival. Furthermore, the impact of MIF expression on cell proliferation and anticancer drug sensitivity was examined in murine squamous carcinoma cell line SCCVII after MIF knockdown (MIF-KD). As revealed by quantitative analysis of MIF immunostaining, tumor progression was accompanied by an increase in mean optical density (MOD) and labeling index (LI). Likewise, an elevation of MIF serum levels was noted in HNSCC patients (n = 66) versus healthy individuals (n = 16). Interestingly, comparison of laryngeal carcinoma patients on the basis of MIF tissue expression (high expression, LI ≥ 47, versus low expression, LI < 47) disclosed a significant difference between disease-free survival curves for local and nodal recurrence, and overall survival curve. In vitro, MIF knockdown in murine SCCVII cells resulted in reduced cell proliferation and a decrease in cell motility. In mice inoculated with SCCVII cells, MIF-KD tumors grew more slowly and also appeared more sensitive to chemotherapy. Both clinical observations and experimental data suggest that MIF plays a pivotal role in the progression of HNSCC.

Evaluation of serum galactomannan detection for diagnosis of feline upper respiratory tract aspergillosis

Measurement of serum galactomannan (GM), a polysaccharide fungal cell-wall component, is a non-invasive test for early diagnosis of invasive aspergillosis in humans. Feline upper respiratory tract (URT) aspergillosis is an emerging infectious disease in cats. Diagnosis requires biopsy for procurement of tissue specimens for cytological or histological detection of fungal hyphae and for fungal culture. The aim of this study was to evaluate serum GM measurement as a non-invasive diagnostic test for URT aspergillosis in cats. A one-stage, immunoenzymatic sandwich ELISA was used to detect serum GM in 4 groups of cats; Group 1 (URT aspergillosis) – confirmed URT aspergillosis (n=13, sinonasal aspergillosis (SNA) n=6 and sino-orbital aspergillosis (SOA) n=7), Group 2 (URT other) – other URT diseases (n=15), Group 3 (β-lactam) – cats treated with β-lactam antibiotics for non-respiratory tract disease (n=14), Group 4a – healthy young cats (≤1y of age, n=28), Group 4b – healthy adult cats (>1y of age, n=16). One cat with SNA and two cats with SOA caused by an Aspergillus fumigatus-mimetic species, tested positive for serum GM. For a cut-off optical density index of 1.5, the overall sensitivity and specificity of the assay was 23% and 78% respectively. False positive results occurred in 29% of cats in Group 3 and 32% of cats in Group 4a. Specificity increased to 90% when Groups 3 and 4a were excluded from the analysis. Overall, serum GM measurement has a poor sensitivity but is a moderately specific, non-invasive screening test to rule out infection in patients with suspected feline upper respiratory tract aspergillosis.

Diagnostic Accuracy of PCR Alone Compared to Galactomannan in Bronchoalveolar Lavage Fluid for Diagnosis of Invasive Pulmonary Aspergillosis: a Systematic Review

PCR in bronchoalveolar lavage (BAL) fluid has not been accepted as a diagnostic criterion for invasive pulmonary aspergillosis (IPA). We conducted a systematic review assessing the diagnostic accuracy of PCR in BAL fluid with a direct comparison versus galactomannan (GM) in BAL fluid. We included prospective and retrospective cohort and case-control studies. Studies were included if they used the EORTC/MSG consensus definition criteria of IPA and assessed ≥80% of patients at risk for IPA. Two reviewers abstracted data independently. Risk of bias was assessed using QUADAS-2. Summary sensitivity and specificity values were estimated using a bivariate model and reported with a 95% confidence interval (CI). Nineteen studies published between 1993 and 2012 were included. The summary sensitivity and specificity values (CIs) for diagnosis of proven or probable IPA were 90.2% (77.2 to 96.1%) and 96.4% (93.3 to 98.1%), respectively. In nine cohort studies strictly adherent to the 2002 or 2008 EORTC/MSG criteria for reference standard definitions, the summary sensitivity and specificity values (CIs) were 77.2% (62 to 87.6%) and 93.5% (90.6 to 95.6%), respectively. Antifungal treatment before bronchoscopy significantly reduced sensitivity. The diagnostic performance of PCR was similar to that of GM in BAL fluid using an optical density index cutoff of 0.5. If either PCR or GM in BAL fluid defined a positive result, the pooled sensitivity was higher than that of GM alone, with similar specificity. We conclude that the diagnostic performance of PCR in BAL fluid is good and comparable to that of GM in BAL fluid. Performing both tests results in optimal sensitivity with no loss of specificity. Results are dependent on the reference standard definitions.

Galactomannan Antigen Testing for Diagnosis of Invasive Aspergillosis in Pediatric Hematology Patients

Invasive aspergillosis (IA) can cause significant morbidity and mortality in immunocompromised children. The galactomannan (GM) enzyme immunoassay (EIA) has been shown in adult studies to be a useful adjunct in diagnosing IA. Data on this assay in children are limited by small sample sizes and conflicting results; false-positive assays were a concern in historical studies. We sought to evaluate the GM EIA in a large cohort of children who received intensive chemotherapy and/or hematopoietic stem cell transplant. A focus was placed on evaluating the assay specificity, and the potential of measuring GM antigen in urine. A multicenter prospective observational study in children with anticipated prolonged neutropenia was performed. Serum specimens were collected twice weekly, and urine was collected once weekly during neutropenic periods. Operating characteristics were calculated using the GM EIA optical density index cutoffs of 0.5 and 1.0 for both serum and urine specimens. At least one serum or urine specimen was tested from 198 patients. Ten patients had one or more repeatedly positive serum specimens, while 37 patients had one or more repeatedly positive urine specimens. The specificity of serum and urine testing was 95% and 80%, respectively. Although the urine test resulted in a higher false positivity rate, it successfully identified the only case of probable IA. Data suggest that the serum GM EIA does not provide frequent false-positive results as previously reported. Screening for galactomannan, or a related antigen in urine, needs to be further evaluated as it may be amenable to development of surveillance strategies.

Computerized Scheimpflug densitometry as a measure of corneal optical density after excimer laser refractive surgery in myopic eyes

To evaluate changes in anterior corneal optical density and the refractive index after photorefractive keratectomy (PRK) using a rotating Scheimpflug system. Department of Ophthalmology, University Federico II, Naples, Italy. Comparative case series. Anterior corneal optical density was evaluated with a rotating Scheimpflug system at baseline and 3 months and 12 months after PRK in eyes with a refractive error between -6.00 diopters (D) and -12.00 D (study group). A control group of unoperated eyes with the same refraction range was used to calculate corneal optical density and the Gladstone-Dale constant in unoperated eyes using the Gladstone-Dale formula. In the study group, changes in the anterior corneal optical density were evaluated over time and variations in the anterior corneal refractive index were obtained using the Gladstone-Dale constant. The study group comprised 37 eyes and the control group, 200 eyes. In the study group, the mean anterior corneal optical density and refractive index, respectively, were 27.71 ± 4.39 and 1.360 ± 0.05 at baseline, 37.812 ± 12.31 and 1.491 ± 0.16 after 3 months (P<.001 compared with baseline), and 26.29 ± 4.93 and 1.341 ± 0.06 after 12 months (P=.03 compared with baseline). The mean corneal optical density in the control group was 27.71 ± 4.31 (SD), and the resultant Gladstone-Dale constant was 0.013. An early increase and a subsequent reduction in anterior corneal optical density and the refractive index were present in myopic eyes during 1 year after PRK.