Investigating the presence of aspergillus fumigatus and A. Flavus using galactomannan enzyme assay and taqman real-time PCR technique

Background: Invasive aspergillosis (IA), a serious and fatal disease, is caused by numerous opportunistic fungi including Aspergillus species. Lack of early diagnosis and delay in treatment lead to the rapid spread of the infection and relapse, increase treatment costs, and ultimately cause death. Objectives: This study aimed at investigating the presence of Aspergillus species by Galactomannan EIA (GM) and TaqMan q PCR methods. Methods: Fluid samples of bronchoalveolar lavage (BAL) were collected from 89 patients, who were at risk for IA and underwent a bronchoscopy at Shariati hospital in Tehran, Iran. The specimens were examined using direct examination, culture methods, and GM and TaqMan real-time PCR. Results: A total of 23 samples were found to be positive in direct examination, 7 were identified as Aspergillus fumigatus, and 11 were positive as A. flavus in culture assays, 27 were GM positive, and 29 were positive with PAN Aspergillus probe. Moreover, 7 samples were positive with A. fumigatus probe and 11 were positive with A. flavus probe. Negative predictive value (NPV), sensitivity, positive predictive value (PPV), and specificity for a � 1.0 BAL GM level were 98.4, 94.4, 62.9, and 85.9, respectively, and were 100 for TaqMan Real time PCR. Conclusions: Our findings revealed that TaqManreal-time PCR has a great value for detection of Aspergillus species in BAL specimen. Since early diagnosis has been highly considered to prevent the occurrence of drug resistance and mortality rate, it is necessary to use this method directly to detect Aspergillus species in BAL or blood specimens without culture. Onthe other hand, Galactomannan assay with a 98.4 NPVcould be usedto screen the patients with suspected invasive aspergillosis. Additionally, dueto the lack of need for specialized devices, an appropriate diagnostic technique should be used in laboratories that do not have access to specialized procedures. © 2018, Jundishapur Journal of Microbiology

Investigating the presence of aspergillus fumigatus and A. Flavus using galactomannan enzyme assay and taqman real-time PCR technique

Background: Invasive aspergillosis (IA), a serious and fatal disease, is caused by numerous opportunistic fungi including Aspergillus species. Lack of early diagnosis and delay in treatment lead to the rapid spread of the infection and relapse, increase treatment costs, and ultimately cause death. Objectives: This study aimed at investigating the presence of Aspergillus species by Galactomannan EIA (GM) and TaqMan q PCR methods. Methods: Fluid samples of bronchoalveolar lavage (BAL) were collected from 89 patients, who were at risk for IA and underwent a bronchoscopy at Shariati hospital in Tehran, Iran. The specimens were examined using direct examination, culture methods, and GM and TaqMan real-time PCR. Results: A total of 23 samples were found to be positive in direct examination, 7 were identified as Aspergillus fumigatus, and 11 were positive as A. flavus in culture assays, 27 were GM positive, and 29 were positive with PAN Aspergillus probe. Moreover, 7 samples were positive with A. fumigatus probe and 11 were positive with A. flavus probe. Negative predictive value (NPV), sensitivity, positive predictive value (PPV), and specificity for a � 1.0 BAL GM level were 98.4, 94.4, 62.9, and 85.9, respectively, and were 100 for TaqMan Real time PCR. Conclusions: Our findings revealed that TaqManreal-time PCR has a great value for detection of Aspergillus species in BAL specimen. Since early diagnosis has been highly considered to prevent the occurrence of drug resistance and mortality rate, it is necessary to use this method directly to detect Aspergillus species in BAL or blood specimens without culture. Onthe other hand, Galactomannan assay with a 98.4 NPVcould be usedto screen the patients with suspected invasive aspergillosis. Additionally, dueto the lack of need for specialized devices, an appropriate diagnostic technique should be used in laboratories that do not have access to specialized procedures. © 2018, Jundishapur Journal of Microbiology

Decolorization of Reactive Black-5 High Concentration by Vermicompost Microflora and Detoxification of By-Products by UV-C/H2O2 Post-Treatment

The presence of synthetic dyes in textile wastewater is a problematic issue for environmentalist. Nowadays, dye removal is practiced via different methods. Among all these methods, biodecolorization is an ideal technique. The present research apples vermicompost microflora to remove reactive black-C, pH = 7, and under anaerobic condition. At 36h, removal efficiencies of 94.79%, 94.06%, and 93.6% are obtained for concentrations of 800, 850, and 950 mg/L, respectively. It has also been observed that when the initial concentration rises to 1400 mg/L, the efficiency drops to 51.57% at 36h. Also, methyl red, methyl orange, eriochrome black-t, and acid blue-113 could be decolorized by the isolated bacterial strain with an efficiency of 94.29%, 92.10%, 90.83%, and 88.95%, respectively. Phytotoxicity Test shows that the parent form of reactive black-5 has not been toxic for the seeds (100% germination for Triticum aestivum and 90% for Maize). When reactive black-5 is treated with isolated bacterial strain under anaerobic condition, none of the seeds remain germinated which might be due to the possible formation of toxic aromatic amines intermediates. Therefore, ultraviolet C + 100 mM H2O2 has been used as the post-treatment process for detoxifying of by-products. After the integrated treatment of synthetic wastewater, containing RB-5, complete germination (100%) of Triticum aestivum and Maize is observed. In the post-treatment process, due to the generation and activation of hydroxyl radicals, the toxic aromatic amines compounds convert to the less toxic compounds. Keywords:azo dyes; anaerobic removal; integrated treatment; phytotoxicity; hydroxyl radical

Designer Exosomes: A New Platform for Biotechnology Therapeutics

Abstract: Desirable features of exosomes have made them a suitable manipulative platform for biomedical applications, including targeted drug delivery, gene therapy, cancer diagnosis and therapy, development of vaccines, and tissue regeneration. Although natural exosomes have various potentials, their clinical application is associated with some inherent limitations. Recently, these limitations inspired various attempts to engineer exosomes and develop designer exosomes. Mostly, designer exosomes are being developed to overcome the natural limitations of exosomes for targeted delivery of drugs and functional molecules to wounds, neurons, and the cardiovascular system for healing of damage. In this review, we summarize the possible improvements of natural exosomes by means of two main approaches: parental cell-based or pre-isolation exosome engineering and direct or post-isolation exosome engineering. Parental cell-based engineering methods use genetic engineering for loading of therapeutic molecules into the lumen or displaying them on the surface of exosomes. On the other hand, the post-isolation exosome engineering approach uses several chemical and mechanical methods including click chemistry, cloaking, bio-conjugation, sonication, extrusion, and electroporation. This review focuses on the latest research, mostly aimed at the development of designer exosomes using parental cell-based engineering and their application in cancer treatment and regenerative medicine. Graphic Abstract: Figure not available: see fulltext. © 2020, Springer Nature Switzerland AG

Mechanism Underlying Defective Interferon Gamma-Induced IDO Expression in Non-obese Diabetic Mouse Fibroblasts

Indoleamine 2,3-dioxygenase (IDO) can locally suppress T cell-mediated immune responses. It has been shown that defective self-tolerance in early prediabetic female non-obese diabetic (NOD) mice can be attributed to the impaired interferon-gamma (IFN-γ)- induced IDO expression in dendritic cells of these animals. As IFN-γ can induce IDO in both dendritic cells and fibroblasts, we asked the question of whether there exists a similar defect in IFN-γ-induced IDO expression in NOD mice dermal fibroblasts. To this end, we examined the effect of IFN-γ on expression of IDO and its enzymatic activity in NOD dermal fibroblasts. The results showed that fibroblasts from either prediabetic (8 wks of age) female or male, and diabetic female or male (12 and 24 wks of age respectively) NOD mice failed to express IDO in response to IFN-γ treatment. To find underlying mechanisms, we scrutinized the IFN- γ signaling pathway and investigated expression of other IFN-γ-modulated factors including major histocompatibility complex class I (MHC-I) and type I collagen (COL-I). The findings revealed a defect of signal transducer and activator of transcription 1 (STAT1) phosphorylation in NOD cells relative to that of controls. Furthermore, we found an increase in MHC-I and suppression of COL-I expression in fibroblasts from both NOD and control mice following IFN-γ treatment; indicating that the impaired response to IFN-γ in NOD fibroblasts is specific to IDO gene. Finally, we showed that an IFN-γ-independent IDO expression pathway i.e. lipopolysaccharide (LPS)-mediated-c-Jun kinase is operative in NOD mice fibroblast. In conclusion, the findings of this study for the first time indicate that IFN-γ fails to induce IDO expression in NOD dermal fibroblasts; this may partially be due to defective STAT1 phosphorylation in IFN-γ-induced-IDO signaling pathway

An Observational Cohort Study of the Kynurenine to Tryptophan Ratio in Sepsis: Association with Impaired Immune and Microvascular Function

Both endothelial and immune dysfunction contribute to the high mortality rate in human sepsis, but the underlying mechanisms are unclear. In response to infection, interferon-γ activates indoleamine 2,3-dioxygenase (IDO) which metabolizes the essential amino acid tryptophan to the toxic metabolite kynurenine. IDO can be expressed in endothelial cells, hepatocytes and mononuclear leukocytes, all of which contribute to sepsis pathophysiology. Increased IDO activity (measured by the kynurenine to tryptophan [KT] ratio in plasma) causes T-cell apoptosis, vasodilation and nitric oxide synthase inhibition. We hypothesized that IDO activity in sepsis would be related to plasma interferon-γ, interleukin-10, T cell lymphopenia and impairment of microvascular reactivity, a measure of endothelial nitric oxide bioavailability. In an observational cohort study of 80 sepsis patients (50 severe and 30 non-severe) and 40 hospital controls, we determined the relationship between IDO activity (plasma KT ratio) and selected plasma cytokines, sepsis severity, nitric oxide-dependent microvascular reactivity and lymphocyte subsets in sepsis. Plasma amino acids were measured by high performance liquid chromatography and microvascular reactivity by peripheral arterial tonometry. The plasma KT ratio was increased in sepsis (median 141 [IQR 64–235]) compared to controls (36 [28–52]); p<0.0001), and correlated with plasma interferon-γ and interleukin-10, and inversely with total lymphocyte count, CD8+ and CD4+ T-lymphocytes, systolic blood pressure and microvascular reactivity. In response to treatment of severe sepsis, the median KT ratio decreased from 162 [IQR 100–286] on day 0 to 89 [65–139] by day 7; p = 0.0006) and this decrease in KT ratio correlated with a decrease in the Sequential Organ Failure Assessment score (p<0.0001). IDO-mediated tryptophan catabolism is associated with dysregulated immune responses and impaired microvascular reactivity in sepsis and may link these two fundamental processes in sepsis pathophysiology

Fibroblast cell-based therapy prevents induction of alopecia areata in an experimental model

YesAlopecia areata (AA) is an autoimmune hair loss disease with infiltration of proinflammatory cells into hair follicles. Current therapeutic regimens are unsatisfactory mainly because of the potential for side effects and/or limited efficacy. Here we report that cultured, transduced fibroblasts, which express the immunomodulatory molecule indoleamine 2,3-dioxygenase (IDO), can be applied to prevent hair loss in an experimental AA model. A single intraperitoneal (IP) injection of IDO-expressing primary dermal fibroblasts was given to C3H/HeJ mice at the time of AA induction. While 60–70% of mice that received either control fibroblasts or vehicle injections developed extensive AA, none of the IDO-expressing fibroblast-treated mice showed new hair loss up to 20 weeks post injection. IDO cell therapy significantly reduced infiltration of CD4+ and CD8+ T cells into hair follicles and resulted in decreased expression of TNF-α, IFN-γ and IL-17 in the skin. Skin draining lymph nodes of IDO fibroblast-treated mice were significantly smaller, with more CD4+ CD25+ FoxP3+ regulatory T cells and fewer Th17 cells than those of control fibroblast and vehicle-injected mice. These findings indicate that IP injected IDO-expressing dermal fibroblasts can control inflammation and thereby prevent AA hair loss.Canadian Institutes of Health Researches (Funding Reference Number: 134214 and 136945)