New PCR Assays for the Identification of Fusarium verticillioides, Fusarium subglutinans, and Other Species of the Gibberella fujikuroi Complex

Fusarium verticillioides and Fusarium subglutinans are important fungal pathogens of maize and other cereals worldwide. In this study, we developed PCR-based protocols for the identification of these pathogens targeting the gaoB gene, which codes for galactose oxidase. The designed primers recognized isolates of F. verticillioides and F. subglutinans that were obtained from maize seeds from several producing regions of Brazil but did not recognize other Fusarium spp. or other fungal genera that were either obtained from fungal collections or isolated from maize seeds. A multiplex PCR protocol was established to simultaneously detect the genomic DNA from F. verticillioides and F. subglutinans. This protocol could detect the DNA from these fungi growing in artificially or naturally infected maize seeds. Another multiplex reaction with a pair of primers developed in this work combined with a pre-existing pair of primers has allowed identifying F. subglutinans, F. konzum, and F. thapsinum. In addition, the identification of F. nygamai was also possible using a combination of two PCR reactions described in this work, and another described in the literature.     

Impact of Leptin on Periodontal Ligament Fibroblasts during Mechanical Strain

Orthodontic treatment to correct dental malocclusions leads to the formation of pressure zones in the periodontal ligament resulting in a sterile inflammatory reaction, which is mediated by periodontal ligament fibroblasts (PDLF). Leptin levels are elevated in obesity and chronic inflammatory responses. In view of the increasing number of orthodontic patients with these conditions, insights into effects on orthodontic treatment are of distinct clinical relevance. A possible influence of leptin on the expression profile of PDLF during simulated orthodontic mechanical strain, however, has not yet been investigated. In this study, PDLF were exposed to mechanical strain with or without different leptin concentrations. The gene and protein expression of proinflammatory and bone-remodelling factors were analysed with RT-qPCR, Western-blot and ELISA. The functional analysis of PDLF-induced osteoclastogenesis was analysed by TRAP (tartrate-resistant acid phosphatase) staining in coculture with human macrophages. Pressure-induced increase of proinflammatory factors was additionally elevated with leptin treatment. PDLF significantly increased RANKL (receptor activator of NF-kB ligand) expression after compression, while osteoprotegerin was downregulated. An additional leptin effect was demonstrated for RANKL as well as for subsequent osteoclastogenesis in coculture after TRAP staining. Our results suggest that increased leptin concentrations, as present in obese patients, may influence orthodontic tooth movement. In particular, the increased expression of proinflammatory factors and RANKL as well as increased osteoclastogenesis can be assumed to accelerate bone resorption and thus the velocity of orthodontic tooth movement in the orthodontic treatment of obese patients.     

Combining fuzzy querying of imprecise data and predictive microbiology using category-based reasoning for prediction of the possible microbial spoilage in foods: application to Listeria monocytogenes

Various predictive models of microbial behavior have been created and extensive data collection has been done by numerous private or public laboratories. However, significant differences between predicted and observed values in foods have been observed and need to be stressed, understood and explained as much as possible. In this paper, we present a software tool (currently at the level of a prototype) able: (i) to store in a database all relevant information expressed on one hand as qualitative or quantitative data and on the other hand as precise or imprecise data; (ii) to retrieve the more relevant information from the database using queries where criteria may be expressed as fuzzy values in order to enhance the flexibility of the search: (iii) to compute, in addition to the nearest data, an estimation of searched values using statistical models. The architecture of this software tool is structured as a category-based reasoning system. Example queries about Listeria monocytogenes (L. monocytogenes) illustrate the functionalities of this tool.     

Fade and quench-resistant emission in calcium phosphate nanoreactors

The fluorescence emission and photodegradation properties of fluorescein dye inside fluid-filled spherical nanoreactors ～ 150 nm in diameter and surrounded by a few nanometres thick layer of calcium phosphate are considered in detail. Steady state, stopped flow, and laser pulsed fluorescence spectroscopies, absorption spectroscopy, dynamic light scattering and electron microscopy were used to characterize the materials as a function of encapsulated dye concentration, particle concentration, illumination time, and pH. Fluorescein tends to form stable J-aggregates inside the nanoreactors. The molecular collision rate constants between the dye aggregates and between the dyes and soluble quenchers are greatly reduced inside the nanoreactors and are responsible for the observed resistance to photodegradation and reduced emission quenching. A model for dye behaviour in nanoreactors is suggested. Nanoreactors can be concentrated to a high suspension concentration, yielding exceptionally strong luminescence affected only by inner filter effects absent particle-particle crosstalk. These and similar nanoreactors can be utilized as building blocks for three-dimensional photo-optical devices, and as versatile and resilient supramolecular chromophores or tracers in complex fluids, cells and microfluidic systems where high resolution visualization is needed.     

Acetyldiaminobutanoic acid,a potential lathyrogenic amino acid in leaves of Acacia angustissima

Acacia angustissima has potential in agroforestry systems for forage production in tropical environments. However, feeding trials indicate that the leaves are potentially toxic. Signs of toxicity in sheep are similar to those presented by sheep fed flatpea (Lathyrus sylvestris) hay which contains a high concentration of the non‐protein amino acid 2,4‐diaminobutanoic acid (DABA). Cation exchange chromatography, high‐performance liquid chromatography (HPLC) and NMR were used to isolate and identify the non‐protein amino acid in A angustissima leaves. A quantitative HPLC method was used to determine the content of the non‐protein amino acid in leaves from 20 accessions. The non‐protein amino acid was identified as 4‐N‐acetyl‐2,4‐diaminobutanoic acid (ADAB), previously reported to occur in seeds. The average and range in ADAB content were 16.8 and 10.6–25.4 mg g^?1 respectively. This content of ADAB is higher than the levels of DABA in flatpea hay that are toxic to sheep. © 2001 Society of Chemical Industry     

Podocyte-Related Mechanisms Underlying Survival Benefit of Long-Term Angiotensin Receptor Blocker

We previously found that short-term treatment (week 8 to 12 after injury) with high-dose angiotensin receptor blocker (ARB) induced the regression of existing glomerulosclerosis in 5/6 nephrectomy rats. We therefore assessed the effects of long-term intervention with ARB vs. nonspecific antihypertensives in this study. Adult rats underwent 5/6 nephrectomy and renal biopsy 8 weeks later. The rats were then divided into three groups with equivalent renal function and glomerular sclerosis and treated with high-dose losartan (ARB), nonspecific antihypertensive triple-therapy (TRX), or left untreated (Control) until week 30. We found that blood pressure, serum creatinine levels, and glomerulosclerosis were lower at sacrifice in ARB and TRX vs. Control. Only ARB reduced proteinuria and maintained the density of WT-1-positive podocytes. Glomerular tufts showed more double-positive cells for CD44, a marker of activated parietal epithelial cells, and synaptopodin after ARB vs. TRX or Control. ARB treatment reduced aldosterone levels. ARB-treated rats had significantly improved survival when compared with TRX or Control. We conclude that both long-term ARB and triple-therapy ameliorate progression, but do not sustain the regression of glomerulosclerosis. ARB resulted in the superior preservation of podocyte integrity and decreased proteinuria and aldosterone, linked to increased survival in the uremic environment.     

Molecular Subtyping of Invasive Breast Cancer Using a PAM50-Based Multigene Expression Test-Comparison with Molecular-Like Subtyping by Tumor Grade/Immunohistochemistry and Influence on Oncologist’s Decision on Systemic Therapy in a Real-World Setting

In intermediate risk hormone receptor (HR) positive, HER2 negative breast cancer (BC), the decision regarding adjuvant chemotherapy might be facilitated by multigene expression tests. In all, 142 intermediate risk BCs were investigated using the PAM50-based multigene expression test Prosigna® in a prospective multicentric study. In 119/142 cases, Prosigna® molecular subtyping was compared with local and two central (C1 and C6) molecular-like subtypes relying on both immunohistochemistry (IHC; HRs, HER2, Ki-67) and IHC + tumor grade (IHC+G) subtyping. According to local IHC, 35.4% were Luminal A-like and 64.6% Luminal B-like subtypes (local IHC+G subtype: 31.9% Luminal A-like; 68.1% Luminal B-like). In contrast to local and C1 subtyping, C6 classified >2/3 of cases as Luminal A-like. Pairwise agreement between Prosigna® subtyping and molecular-like subtypes was fair to moderate depending on molecular-like subtyping method and center. The best agreement was observed between Prosigna® (53.8% Luminal A; 44.5% Luminal B) and C1 surrogate subtyping (Cohen’s kappa = 0.455). Adjuvant chemotherapy was suggested to 44.2% and 88.6% of Prosigna® Luminal A and Luminal B cases, respectively. Out of all Luminal A-like cases (locally IHC/IHC+G subtyping), adjuvant chemotherapy was recommended if Prosigna® testing classified as Prosigna® Luminal A at high / intermediate risk or upgraded to Prosigna® Luminal B.     

Statistical evaluation of dye desorption – mixed two- and three-level design and kinetic modeling

Abstract

Biosorption has been shown in the literature to be effective in the removal of dye from aqueous solution. To be fully useful, a desorption process is necessary, which has a direct impact on the process costs. The aim of this work was to evaluate the desorption of the azo dye Acid Yellow 23 adsorbed in a biosorbent obtained from Bombyx mori cocoons. Temperature and pH media were evaluated through a mixed two- and three-level design, using NaOH as desorbing agent. Kinetic desorption was also obtained at the optimized condition. The statistical evaluation of the experimental design showed that the pH has almost two times more influence on the dye desorption than the temperature, with only the linear parameters being statistically significant. Among four kinetic desorption models adjusted to the experimental data, the proposed n-order model was statistically determined as the best representation of the experimental data.