Improving Adenovirus Based Gene Transfer: Strategies to Accomplish Immune Evasion

Adenovirus (Ad) based gene transfer vectors continue to be the platform of choice for an increasing number of clinical trials worldwide. In fact, within the last five years, the number of clinical trials that utilize Ad based vectors has doubled, indicating growing enthusiasm for the numerous positive characteristics of this gene transfer platform. For example, Ad vectors can be easily and relatively inexpensively produced to high titers in a cGMP compliant manner, can be stably stored and transported, and have a broad applicability for a wide range of clinical conditions, including both gene therapy and vaccine applications. Ad vector based gene transfer will become more useful as strategies to counteract innate and/or pre-existing adaptive immune responses to Ads are developed and confirmed to be efficacious. The approaches attempting to overcome these limitations can be divided into two broad categories: pre-emptive immune modulation of the host, and selective modification of the Ad vector itself. The first category of methods includes the use of immunosuppressive drugs or specific compounds to block important immune pathways, which are known to be induced by Ads. The second category comprises several innovative strategies inclusive of: (1) Ad-capsid-display of specific inhibitors or ligands; (2) covalent modifications of the entire Ad vector capsid moiety; (3) the use of tissue specific promoters and local administration routes; (4) the use of genome modified Ads; and (5) the development of chimeric or alternative serotype Ads. This review article will focus on both the promise and the limitations of each of these immune evasion strategies, and in the process delineate future directions in developing safer and more efficacious Ad-based gene transfer strategies

Immune Recognition of Gene Transfer Vectors: Focus on Adenovirus as a Paradigm

Recombinant Adenovirus (Ad) based vectors have been utilized extensively as a gene transfer platform in multiple pre-clinical and clinical applications. These applications are numerous, and inclusive of both gene therapy and vaccine based approaches to human or animal diseases. The widespread utilization of these vectors in both animal models, as well as numerous human clinical trials (Ad-based vectors surpass all other gene transfer vectors relative to numbers of patients treated, as well as number of clinical trials overall), has shed light on how this virus vector interacts with both the innate and adaptive immune systems. The ability to generate and administer large amounts of this vector likely contributes not only to their ability to allow for highly efficient gene transfer, but also their elicitation of host immune responses to the vector and/or the transgene the vector expresses in vivo. These facts, coupled with utilization of several models that allow for full detection of these responses has predicted several observations made in human trials, an important point as lack of similar capabilities by other vector systems may prevent detection of such responses until only after human trials are initiated. Finally, induction of innate or adaptive immune responses by Ad vectors may be detrimental in one setting (i.e., gene therapy) and be entirely beneficial in another (i.e., prophylactic or therapeutic vaccine based applications). Herein, we review the current understanding of innate and adaptive immune responses to Ad vectors, as well some recent advances that attempt to capitalize on this understanding so as to further broaden the safe and efficient use of Ad-based gene transfer therapies in general

Stream hydrological fragmentation drives bacterioplankton community composition

In Mediterranean intermittent streams, the hydrological fragmentation in summer and the successive water flow reconvergence in autumn allow exploring how local processes shape the microbial community within the same habitat. The objectives of this study were to determine how bacterial community composition responded to hydrological fragmentation in summer, and to evaluate whether the seasonal shifts in community composition predominate over the effects of episodic habitat fragmentation. The bacterial community was assessed along the intermittent stream Fuirosos (Spain), at different levels of phylogenetic resolution by in situ hybridization, fingerprinting, and 16S rRNA gene sequencing. The hydrological fragmentation of the stream network strongly altered the biogeochemical conditions with the depletion of oxidized solutes and caused changes in dissolved organic carbon characteristics. In the isolated ponds, beta-Proteobacteria and Actinobacteria increased their abundance with a gradual reduction of the alpha-diversity as pond isolation time increased. Moreover, fingerprinting analysis clearly showed a shift in community composition between summer and autumn. In the context of a seasonal shift, the temporary stream fragmentation simultaneously reduced the microbial dispersion and affected local environmental conditions (shift in redox regime and quality of the dissolved organic matter) tightly shaping the bacterioplankton community composition

Alternatives for testing of context-aware software systems in non-academic settings:results from a <i>Rapid Review</i>

Context: Context-awareness challenges the engineering of contemporary software systems and jeopardizes their testing. The variation of context represents a relevant behavior that deepens the limitations of available software testing practices and technologies. However, such software systems are mainstream. Therefore, researchers in non-academic settings also face challenges when developing and testing contemporary software systems. Objective: To understand how researchers deal with the variation of context when testing context-aware software systems developed in non-academic settings. Method: To undertake a secondary study (Rapid Review) to uncover the necessary evidence from primary sources describing the testing of context-aware software systems outside academia. Results: The current testing initiatives in non-academic settings aim to generate or improve test suites that can deal with the context variation and the sheer volume of test input possibilities. They mostly rely on modeling the systems’ dynamic behavior and increasing computing resources to generate test inputs to achieve this. We found no evidence of test results aiming at managing context variation through the testing lifecycle process. Conclusions: So far, the identified testing initiatives and strategies are not ready for mainstream adoption. They are all domain-specific, and while the ideas and approaches can be reproduced in distinct settings, the technologies are to be re-engineered and tailored to the context-awareness of contemporary software systems in different problem domains. Further and joint investigations in academia and experiences in non-academic set- tings can evolve the body of knowledge regarding the testing of contemporary software systems in the field

Zooplankton as refuge for invading bacteria

Nonetheless, studies on the impact of non-parasitic bacteria are still scarce. In this study we analysed the consequences of this colonisation on the distribution of potentially pathogenic and antibiotic resistant bacteria in freshwater systems. This is based on the hypothesis that potential pathogens have a selective advantage when growing in biofilms, as has been shown in clinical settings. Thus we postulate that attaching to animals might allow potential pathogens, e.g. deriving from the efflux of a waste water treatment plant (WWTP), to survive in freshwaters. Additionally the movement of the animal transports the bacteria away from the point-contamination to more pristine waters. We experimentally show that, if added to a natural bacterial community, Escherichia coli attaches to daphnids and derives particularly well in the gut of the animals, where they naturally do not form part of their microflora, whilst decreasing in abundance in the surrounding water. Moreover, the presence of daphnids decreased the abundance of a tetracycline resistance gene (tetA) in the surrounding water, whilst a higher number of tetA gene copies were detected in the animals. In addition we tested whether zooplankton species form a surface for the horizontal gene transfer, of e.g. antibiotic resistant genes, from one E. coli strain to another. In order to expand our hypothesis to natural environment we sampled Lake Maggiore at three points with increasing distance to an inflow of a WWTP and compared the microbial community on various zooplankton species as well as on stones and in the sediment. Whereas similar relationships have been described in marine systems, e.g. Vibrio cholera and Copepods, this is the first study that confronts the potential role of zooplankton in the spread of potential pathogens in freshwater systems

Quality and reactivity of dissolved organic matter in a Mediterranean river across hydrological and spatial gradients.

Understanding DOM transport and reactivity in rivers is essential to having a complete picture of the global carbon cycle. In this study, we explore the effects of hydrological variability and downstream transport on dissolved organic matter (DOM) dynamics in a Mediterranean river. We sampled the main stem of the river Tordera from the source to the sea, over a range of fifteen hydrological conditions including extreme events (flood and drought). By exploring spatial and temporal gradients of DOM fluorescence properties, river hydrology was found to be a significant predictor of DOM spatial heterogeneity. An additional space-resolved mass balance analysis performed on four contrasting hydrological conditions revealed that this was due to a shift in the biogeochemical function of the river. Flood conditions caused a conservative transport of DOM, generating a homogeneous, humic-like spatial profile of DOM quality. Lower flows induced a non-conservative, reactive transport of DOM, which enhanced the spatial heterogeneity of DOM properties. Moreover, the downstream evolution of DOM chemostatic behaviour revealed that the role of hydrology in regulating DOM properties increased gradually downstream, indicating an organised inter-dependency between the spatial and the temporal dimensions. Overall, our findings reveal that riverine DOM dynamics is in constant change owing to varying hydrological conditions, and emphasize that in order to fully understand the role of rivers in the global carbon cycle, it is necessary to take into account the full range of hydrological variability, from floods to droughts

EVIDENCE OF OPPORTUNISTIC FEEDING BETWEEN ICHTHYOSAURS AND THE OLDEST OCCURRENCE OF THE HEXANCHID SHARK NOTIDANODON FROM THE UPPER JURASSIC OF NORTHERN ITALY

In 2016, two fossil marine reptiles were re-discovered in the collections of the Museo Civico di Storia Naturale di Verona. Originally recovered near Asiago, Vicenza province (northern Italy) from an outcrop of the Rosso Ammonitico Veronese Fm. (Middle-Upper Jurassic), they were never described. Morphological analysis carried out under UV-light allowed enhancing contrast with the surrounding matrix and better identifying some anatomical details. Both specimens consist of partially articulated postcranial elements from two distinct ichthyosaurs, including vertebrae, ribs, and some fragmentary elements of the appendicular skeleton. The first specimen V7101 is here tentatively assigned to Ophthalmosauridae based on a combination of features shared with other taxa in this family, such as the regionalization of the vertebral column. Taphonomical analysis suggests a long exposure of the carcass on the sea floor before burial; two teeth of the hexanchiform shark Notidanodon found near the ribcage could indicate scavenging. An ichthyosaur tooth most probably not belonging to the same specimen was found stuck on a rib and can also be attributed to scavenging – the first ever record of this interaction between two ichthyosaurs. The second specimen V7102 is represented by a poorly preserved partial vertebral column and is here referred to Ichthyosauria indet. due to the absence of taxonomically significant characters. Calcareous nannofossil data and microfacies analyses allow us to assign both specimens to the basal Kimmeridgian. This makes the two Notidanodon teeth associated with V7101 the oldest recorded occurrence of this genus

Dissolved organic matter in continental hydro-geothermal systems: insights from two hot springs of the East African Rift valley

Little is known about the quantity and quality of dissolved organic matter (DOM) in waters from continental geothermal systems, with only a few reports available from the Yellowstone US National Park. In this study, we explored the chemodiversity of DOM in water samples collected from two geothermal hot springs from the Kenyan East African Rift Valley, a region extremely rich in fumaroles, geysers, and spouting springs, located in close proximity to volcanic lakes. The DOM characterization included in-depth assessments performed by negative electrospray ionization Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR-MS). Reduced, saturated and little aromatic DOM compounds were dominant in the hot spring waters collected from either the Ol Njorowa gorge (ON) or the south shore of the soda-saline Lake Elementaita (ELM). Oxygen-poor and sulfur-bearing DOM molecules prevailed in ON, probably reflecting abiotic sulfurization from sulfide-rich geofluids. Nitrogen-bearing aliphatic and protein-like molecules were abundant in ELM, possibly perfusing through the organic-rich sediments of the adjacent Lake Elementaita. Notably, the heat-altered DOM of ancient autochthonous derivation could represent an overlooked source of aliphatic organic carbon for connected lentic environments, with a potential direct impact on nutrient cycling in lakes that receive geothermal water inputs

Microbial colonization patterns and biodegradation of petrochemical and biodegradable plastics in lake waters: insights from a field experiment

IntroductionOnce dispersed in water, plastic materials become promptly colonized by biofilm-forming microorganisms, commonly known as plastisphere.MethodsBy combining DNA sequencing and Confocal Laser Scanning Microscopy (CLSM), we investigated the plastisphere colonization patterns following exposure to natural lake waters (up to 77 days) of either petrochemical or biodegradable plastic materials (low density polyethylene - LDPE, polyethylene terephthalate - PET, polylactic acid - PLA, and the starch-based MaterBi® - Mb) in comparison to planktonic community composition. Chemical composition, water wettability, and morphology of plastic surfaces were evaluated, through Transform Infrared Spectroscopy (ATR-FTIR), Scanning Electron Microscopy (SEM), and static contact angle analysis, to assess the possible effects of microbial colonization and biodegradation activity.Results and DiscussionThe phylogenetic composition of plastisphere and planktonic communities was notably different. Pioneering microbial colonisers, likely selected from lake waters, were found associated with all plastic materials, along with a core of more than 30 abundant bacterial families associated with all polymers. The different plastic materials, either derived from petrochemical hydrocarbons (i.e., LDPE and PET) or biodegradable (PLA and Mb), were used by opportunistic aquatic microorganisms as adhesion surfaces rather than carbon sources. The Mb-associated microorganisms (i.e. mostly members of the family Burkholderiaceae) were likely able to degrade the starch residues on the polymer surfaces, although the Mb matrix maintained its original chemical structure and morphology. Overall, our findings provide insights into the complex interactions between aquatic microorganisms and plastic materials found in lake waters, highlighting the importance of understanding the plastisphere dynamics to better manage the fate of plastic debris in the environment

Compound heterozygosity for loss‐of‐function FARSB variants in a patient with classic features of recessive aminoacyl‐tRNA synthetase‐related disease

Aminoacyl‐tRNA synthetases (ARSs) are ubiquitously expressed enzymes that ligate amino acids onto tRNA molecules. Genes encoding ARSs have been implicated in phenotypically diverse dominant and recessive human diseases. The charging of tRNAPHE with phenylalanine is performed by a tetrameric enzyme that contains two alpha (FARSA) and two beta (FARSB) subunits. To date, mutations in the genes encoding these subunits (FARSA and FARSB) have not been implicated in any human disease. Here, we describe a patient with a severe, lethal, multisystem, developmental phenotype who was compound heterozygous for FARSB variants: p.Thr256Met and p.His496Lysfs*14. Expression studies using fibroblasts isolated from the proband revealed a severe depletion of both FARSB and FARSA protein levels. These data indicate that the FARSB variants destabilize total phenylalanyl‐tRNA synthetase levels, thus causing a loss‐of‐function effect. Importantly, our patient shows strong phenotypic overlap with patients that have recessive diseases associated with other ARS loci; these observations strongly support the pathogenicity of the identified FARSB variants and are consistent with the essential function of phenylalanyl‐tRNA synthetase in human cells. In sum, our clinical, genetic, and functional analyses revealed the first FARSB variants associated with a human disease phenotype and expand the locus heterogeneity of ARS‐related human disease.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/144241/1/humu23424_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/144241/2/humu23424.pd