Challenges and opportunities for quantifying roots and rhizosphere interactions through imaging and image analysis

The morphology of roots and root systems influences the efficiency by which plants acquire nutrients and water, anchor themselves and provide stability to the surrounding soil. Plant genotype and the biotic and abiotic environment significantly influence root morphology, growth and ultimately crop yield. The challenge for researchers interested in phenotyping root systems is, therefore, not just to measure roots and link their phenotype to the plant genotype, but also to understand how the growth of roots is influenced by their environment. This review discusses progress in quantifying root system parameters (e.g. in terms of size, shape and dynamics) using imaging and image analysis technologies and also discusses their potential for providing a better understanding of root:soil interactions. Significant progress has been made in image acquisition techniques, however trade-offs exist between sample throughput, sample size, image resolution and information gained. All of these factors impact on downstream image analysis processes. While there have been significant advances in computation power, limitations still exist in statistical processes involved in image analysis. Utilizing and combining different imaging systems, integrating measurements and image analysis where possible, and amalgamating data will allow researchers to gain a better understanding of root:soil interactions

Changes in the allocation of endogenous strigolactone improve plant biomass production on phosphate-poor soils.

Strigolactones (SLs) are carotenoid-derived phytohormones shaping plant architecture and inducing the symbiosis with endomycorrhizal fungi. In Petunia hybrida, SL transport within the plant and towards the rhizosphere is driven by the ABCG-class protein PDR1. PDR1 expression is regulated by phytohormones and by the soil phosphate abundance, and thus SL transport integrates plant development with nutrient conditions. We overexpressed PDR1 (PDR1 OE) to investigate whether increased endogenous SL transport is sufficient to improve plant nutrition and productivity. Phosphorus quantification and nondestructive X-ray computed tomography were applied. Morphological and gene expression changes were quantified at cellular and whole tissue levels via time-lapse microscopy and quantitative PCR. PDR1 OE significantly enhanced phosphate uptake and plant biomass production on phosphate-poor soils. PDR1 OE plants showed increased lateral root formation, extended root hair elongation, faster mycorrhization and reduced leaf senescence. PDR1 overexpression allowed considerable SL biosynthesis by releasing SL biosynthetic genes from an SL-dependent negative feedback. The increased endogenous SL transport/biosynthesis in PDR1 OE plants is a powerful tool to improve plant growth on phosphate-poor soils. We propose PDR1 as an as yet unexplored trait to be investigated for crop production. The overexpression of PDR1 is a valuable strategy to investigate SL functions and transport routes

Derivation of a Triple Mosaic Adenovirus for Cancer Gene Therapy

A safe and efficacious cancer medicine is necessary due to the increasing population of cancer patients whose particular diseases cannot be cured by the currently available treatment. Adenoviral (Ad) vectors represent a promising therapeutic medicine for human cancer therapy. However, several improvements are needed in order for Ad vectors to be effective cancer therapeutics, which include, but are not limited to, improvement of cellular uptake, enhanced cancer cell killing activity, and the capability of vector visualization and tracking once injected into the patients. To this end, we attempted to develop an Ad as a multifunctional platform incorporating targeting, imaging, and therapeutic motifs. In this study, we explored the utility of this proposed platform by generating an Ad vector containing the poly-lysine (pK), the herpes simplex virus type 1 (HSV-1) thymidine kinase (TK), and the monomeric red fluorescent protein (mRFP1) as targeting, tumor cell killing, and imaging motifs, respectively. Our study herein demonstrates the generation of the triple mosaic Ad vector with pK, HSV-1 TK, and mRFP1 at the carboxyl termini of Ad minor capsid protein IX (pIX). In addition, the functionalities of pK, HSV-1 TK, and mRFP1 proteins on the Ad vector were retained as confirmed by corresponding functional assays, indicating the potential multifunctional application of this new Ad vector for cancer gene therapy. The validation of the triple mosaic Ad vectors also argues for the ability of pIX modification as a base for the development of multifunctional Ad vectors

Immunogenic Comparison of Chimeric Adenovirus 5/35 Vector Carrying Optimized Human Immunodeficiency Virus Clade C Genes and Various Promoters

Adenovirus vector-based vaccine is a promising approach to protect HIV infection. However, a recent phase IIb clinical trial using the vector did not show its protective efficacy against HIV infection. To improve the vaccine, we explored the transgene protein expression and its immunogenicity using optimized codon usage, promoters and adaptors. We compared protein expression and immunogenicity of adenovirus vector vaccines carrying native or codon usage-optimized HIV-1 clade C gag and env genes expression cassettes driven by different promoters (CMV, CMVi, and CA promoters) and adapters (IRES and F2A). The adenovirus vector vaccine containing optimized gag gene produced higher Gag protein expression and induced higher immune responses than the vector containing native gag gene in mice. Furthermore, CA promoter generated higher transgene expression and elicited higher immune responses than other two popularly used promoters (CMV and CMVi). The second gene expression using F2A adaptor resulted in higher protein expression and immunity than that of using IRES and direct fusion protein. Taken together, the adenovirus vector containing the expression cassette with CA promoter, optimized HIV-1 clade C gene and an F2A adaptor produced the best protein expression and elicited the highest transgene-specific immune responses. This finding would be promising for vaccine design and gene therapy

Hemodynamic parameters to guide fluid therapy

The clinical determination of the intravascular volume can be extremely difficult in critically ill and injured patients as well as those undergoing major surgery. This is problematic because fluid loading is considered the first step in the resuscitation of hemodynamically unstable patients. Yet, multiple studies have demonstrated that only approximately 50% of hemodynamically unstable patients in the intensive care unit and operating room respond to a fluid challenge. Whereas under-resuscitation results in inadequate organ perfusion, accumulating data suggest that over-resuscitation increases the morbidity and mortality of critically ill patients. Cardiac filling pressures, including the central venous pressure and pulmonary artery occlusion pressure, have been traditionally used to guide fluid management. However, studies performed during the past 30 years have demonstrated that cardiac filling pressures are unable to predict fluid responsiveness. During the past decade, a number of dynamic tests of volume responsiveness have been reported. These tests dynamically monitor the change in stroke volume after a maneuver that increases or decreases venous return (preload) and challenges the patients' Frank-Starling curve. These dynamic tests use the change in stroke volume during mechanical ventilation or after a passive leg raising maneuver to assess fluid responsiveness. The stroke volume is measured continuously and in real-time by minimally invasive or noninvasive technologies, including Doppler methods, pulse contour analysis, and bioreactance

Genetic Incorporation of Human Metallothionein into the Adenovirus Protein IX for Non-Invasive SPECT Imaging

As the limits of existing treatments for cancer are recognized, clearly novel therapies must be considered for successful treatment; cancer therapy using adenovirus vectors is a promising strategy. However tracking the biodistribution of adenovirus vectors in vivo is limited to invasive procedures such as biopsies, which are error prone, non-quantitative, and do not give a full representation of the pharmacokinetics involved. Current non-invasive imaging strategies using reporter gene expression have been applied to analyze adenoviral vectors. The major drawback to approaches that tag viruses with reporter genes is that these systems require initial viral infection and subsequent cellular expression of a reporter gene to allow non-invasive imaging. As an alternative to conventional vector detection techniques, we developed a specific genetic labeling system whereby an adenoviral vector incorporates a fusion between capsid protein IX and human metallothionein. Our study herein clearly demonstrates our ability to rescue viable adenoviral particles that display functional metallothionein (MT) as a component of their capsid surface. We demonstrate the feasibility of 99mTc binding in vitro to the pIX-MT fusion on the capsid of adenovirus virions using a simple transchelation reaction. SPECT imaging of a mouse after administration of a 99mTc-radiolabeled virus showed clear localization of radioactivity to the liver. This result strongly supports imaging using pIX-MT, visualizing the normal biodistribution of Ad primarily to the liver upon injection into mice. The ability we have developed to view real-time biodistribution in their physiological milieu represents a significant tool to study adenovirus biology in vivo

Derivation of a Myeloid Cell-Binding Adenovirus for Gene Therapy of Inflammation

The gene therapy field is currently limited by the lack of vehicles that permit efficient gene delivery to specific cell or tissue subsets. Native viral vector tropisms offer a powerful platform for transgene delivery but remain nonspecific, requiring elevated viral doses to achieve efficacy. In order to improve upon these strategies, our group has focused on genetically engineering targeting domains into viral capsid proteins, particularly those based on adenovirus serotype 5 (Ad5). Our primary strategy is based on deletion of the fiber knob domain, to eliminate broad tissue specificity through the human coxsackie-and-adenovirus receptor (hCAR), with seamless incorporation of ligands to re-direct Ad tropism to cell types that express the cognate receptors. Previously, our group and others have demonstrated successful implementation of this strategy in order to specifically target Ad to a number of surface molecules expressed on immortalized cell lines. Here, we utilized phage biopanning to identify a myeloid cell-binding peptide (MBP), with the sequence WTLDRGY, and demonstrated that MBP can be successfully incorporated into a knob-deleted Ad5. The resulting virus, Ad.MBP, results in specific binding to primary myeloid cell types, as well as significantly higher transduction of these target populations ex vivo, compared to unmodified Ad5. These data are the first step in demonstrating Ad targeting to cell types associated with inflammatory disease

Increased 30-Day Mortality in Very Old ICU Patients with COVID-19 Compared to Patients with Respiratory Failure without COVID-19

Purpose: The number of patients ≥ 80 years admitted into critical care is increasing. Coronavirus disease 2019 (COVID-19) added another challenge for clinical decisions for both admission and limitation of life-sustaining treatments (LLST). We aimed to compare the characteristics and mortality of very old critically ill patients with or without COVID-19 with a focus on LLST. Methods: Patients 80 years or older with acute respiratory failure were recruited from the VIP2 and COVIP studies. Baseline patient characteristics, interventions in intensive care unit (ICU) and outcomes (30-day survival) were recorded. COVID patients were matched to non-COVID patients based on the following factors: age (± 2 years), Sequential Organ Failure Assessment (SOFA) score (± 2 points), clinical frailty scale (± 1 point), gender and region on a 1:2 ratio. Specific ICU procedures and LLST were compared between the cohorts by means of cumulative incidence curves taking into account the competing risk of discharge and death. Results: 693 COVID patients were compared to 1393 non-COVID patients. COVID patients were younger, less frail, less severely ill with lower SOFA score, but were treated more often with invasive mechanical ventilation (MV) and had a lower 30-day survival. 404 COVID patients could be matched to 666 non-COVID patients. For COVID patients, withholding and withdrawing of LST were more frequent than for non-COVID and the 30-day survival was almost half compared to non-COVID patients. Conclusion: Very old COVID patients have a different trajectory than non-COVID patients. Whether this finding is due to a decision policy with more active treatment limitation or to an inherent higher risk of death due to COVID-19 is unclear.info:eu-repo/semantics/publishedVersio

Erratum to: 36th International Symposium on Intensive Care and Emergency Medicine

[This corrects the article DOI: 10.1186/s13054-016-1208-6.]