Impact of phage predation on bacterial transcriptome under simulated human airway conditions

Bacteriophages have been proven to be efficient in the combat of bacterial multidrug-resistant infections, including those caused by Pseudomonas aeruginosa. Nevertheless, the interactions of phages with bacteria in the human body remains unexplained and its disclosure could lead to advance research and development in phage-based therapies. In this work, RNA-sequencing of phage-infected P. aeruginosa PAO1 adhered to a human epithelial cell monolayer (Nuli-1 ATCC® CRL-4011) was performed to assess bacterial transcriptional processes occurring in phagebacteriahuman cells, i.e., mimicking phage predation under more realistic settings. To achieve that, adhered bacteria were infected with phage LUZ19, and total RNA was extracted from the complex cell mixture. Thereafter, bacterial rRNA/human RNA was depleted and cDNA libraries were prepared to sequence. The differentially expressed genes (DEGs) were quantified using uninfected bacteria as control. In human airway-simulated conditions, there were 21, 39, and 129 bacterial DEGs after 5, 10, and 15 min-post infection, respectively. From DEGs, some genes were identified as part of LUZ19 typical induced responses (prophage, glycerol metabolism, and spermidine synthesis genes). However, unique responses were also captured including upregulation of pyochelin syntheses, LPS modification, sulfate starvation, exopolysaccharide-related genes, and downregulation of bacterial global regulators. These changes are associated with starvation-like conditions (iron and sulfate) and bacteria adaptation to the host, but its role in phage infection progression is still unknown. The study of its impact on bacterial virulence or phage efficient infectivity under human physiology is of most importance. This comprehensive study allows the comparison of bacterial and phage transcripts in the presence of host cells, contributing to a better understanding of phage-bacteria-host interactions, which are relevant in a phage therapy context.info:eu-repo/semantics/publishedVersio

Explicit kinetic heterogeneity: mechanistic models for interpretation of labeling data of heterogeneous cell populations

Estimation of division and death rates of lymphocytes in different conditions is vital for quantitative understanding of the immune system. Deuterium, in the form of deuterated glucose or heavy water, can be used to measure rates of proliferation and death of lymphocytes in vivo. Inferring these rates from labeling and delabeling curves has been subject to considerable debate with different groups suggesting different mathematical models for that purpose. We show that the three models that are most commonly used are in fact mathematically identical and differ only in their interpretation of the estimated parameters. By extending these previous models, we here propose a more mechanistic approach for the analysis of data from deuterium labeling experiments. We construct a model of "kinetic heterogeneity" in which the total cell population consists of many sub-populations with different rates of cell turnover. In this model, for a given distribution of the rates of turnover, the predicted fraction of labeled DNA accumulated and lost can be calculated. Our model reproduces several previously made experimental observations, such as a negative correlation between the length of the labeling period and the rate at which labeled DNA is lost after label cessation. We demonstrate the reliability of the new explicit kinetic heterogeneity model by applying it to artificially generated datasets, and illustrate its usefulness by fitting experimental data. In contrast to previous models, the explicit kinetic heterogeneity model 1) provides a mechanistic way of interpreting labeling data; 2) allows for a non-exponential loss of labeled cells during delabeling, and 3) can be used to describe data with variable labeling length

API480: features towards therapy in honeybee hives

American foulbrood disease (AFB) is a devastating bacterial disease affecting honeybees. It is caused by Paenibacillus larvae, a worldwide-distributed spore forming Gram-positive bacterium which spread easily across apiaries producing highly resistant spores. When AFB symptoms are found the burning of contaminated hives is mandatory causing serious economic losses [1]. Bacteriophages (phages) are being considered valuable solutions to the control of this infection [2-5]. So far, 48 Siphoviridae P. larvae phages sequences are known and most encode integration genes suggesting a temperate lifestyle. All of these 48 phages seem to have a common evolutionary ancestor showing an overall common structure. Their genomes were grouped into four clusters (with Fern, Harrison, Vegas and Halcyone as representative phages) and one singleton (phage Lily) [6].Project APILYSE,PTDC/CVT-EPI/4008/2014-POCI-01-0145-FEDER-016598,-funded by FEDER through COMPETE2020-Programa Operacional Competitividade e Internacionalização(POCI) and the Portuguese Foundation for Science and Technology(FCT) under the scope of the strategic funding of UID/BIO/04469/2019 unit and BioTec Norte operation(NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020-Programa Operacional Regional do Norte.The work was also supported by CEB-UM that provided the laboratorial facilities to perform this research. HR was supported by FCT through the grant SFRH/BD/128859/2017. MB and RL were supported by KULeuven through a GOAgrant[3E140356]info:eu-repo/semantics/publishedVersio

Enterococcus spp. from chicken meat collected 20 years apart overcome multiple stresses occurring in the poultry production chain : Antibiotics, copper and acids

Poultry meat has been a vehicle of antibiotic resistant bacteria and genes. Yet, the diversity of selective pressures associated with their maintenance in the poultry-production chain remains poorly explored. We evaluated the susceptibility of Enterococcus spp. from chicken meat collected 20 years apart to antibiotics, metals, acidic pH and peracetic acid-PAA. Contemporary chicken-meat samples (n = 53 batches, each including a pool of neck skin from 10 single carcasses) were collected in a slaughterhouse facility using PAA as disinfectant (March-August 2018, North of Portugal). Broilers were raised in intensive farms (n = 29) using CuSO4 and organic acids as feed additives. Data were compared with that of 67 samples recovered in the same region during 1999-2001. All 2018 samples had multidrug resistant-MDR isolates, with >45 % carrying Enterococcus faecalis, Enterococcus faecium or Enterococcus gallinarum resistant to tetracycline, erythromycin, ampicillin, quinupristin-dalfopristin, ciprofloxa-cin, chloramphenicol or aminoglycosides. Resistance rates were similar (P > 0.05) to those of 1999-2001 samples for all but five antibiotics. The decrease of samples carrying vancomycin-resistant isolates from 46 % to % between 1999-2001 and 2018 was the most striking difference. Isolates from both periods were similarly susceptible to acid pH [minimum-growth pH (4.5-5.0), minimum-survival pH (3.0-4.0)] and to PAA (MIC90 = 100-120 mg/L/MBC90 = 140-160 mg/L; below concentrations used in slaughterhouse). Copper tolerance genes (tcrB and/or cueO) were respectively detected in 21 % and 4 % of 2018 and 1999-2001 samples. The tcrB gene was only detected in E. faecalis (MICCuSO4 > 12 mM), and their genomes were compared with other international ones of chicken origin (PATRIC database), revealing a polyclonal population and a plasmid or chromosomal location for tcrB. The tcrB plasmids shared diverse genetic modules, including multiple antimicrobial resistance genes (e.g. to tetracyclines, chloramphenicol, macrolide-lincosamide-streptogramin B-MLSB, aminoglycosides, bacitracin, coccidiostats). When in chromosome, the tcrB gene was co-located closely to merA (mercury) genes. Chicken meat remains an important vehicle of MDR Enterococcus spp. able to survive under diverse stresses (e.g. copper, acid) potentially contributing to these bacteria maintenance and flux among animal-environment -humans.Peer reviewe

Histological and ultrastructural analysis of cryopreserved sheep preantral follicles

The aim of this study was to verify the histological and ultrastructural characteristics of sheep preantral follicles after exposure of ovarian tissue to cryopreservation in glycerol (GLY), ethylene glycol (EG), propanediol (PROH) or dimethyl sulfoxide (DMSO) in order to determine the optimum method to store sheep ovarian tissue for later experimental or clinical use. Each ovarian pair from five mixed-breed ewes was divided into 17 fragments. One (control) fragment was immediately fixed for routine histological and ultrastructural studies and the remaining (test) fragments were randomly distributed in cryotubes, equilibrated at 20 °C/20 min in 1.8 mL of minimal essential medium (MEM) containing 1.5 or 3 M GLY, EG, PROH or DMSO and then either fixed for morphological studies to determine their possible toxic effect or frozen/thawed and then fixed to test the effect of cryopreservation on preantral follicles. Histological analysis showed that, compared to control fragments, all cryoprotectants at both concentrations significantly reduced the percentage of normal preantral follicles in ovarian fragments prior to or after cryopreservation. PROH 3.0 M appeared to exert a more toxic effect (P < 0.05) than the other cryoprotectants in noncryopreserved tissues. After freezing/thawing, the highest (P < 0.05) percentages of lightmicroscopical normal preantral follicles were observed in ovarian fragments cryopreserved in EG (1.5 and 3 M) or DMSO (1.5 M). However, transmission electronic microscopical (TEM) examination showed that only the DMSO-cryopreserved preantral follicles had normal ultrastructure. The data suggest that sheep preantral follicles should be cryopreserved with 1.5 M DMSO for later clinical or experimental application

Modular coherent photonic-aided payload receiver for communications satellites

Ubiquitous satellite communications are in a leading position for bridging the digital divide. Fulfilling such a mission will require satellite services on par with fibre services, both in bandwidth and cost. Achieving such a performance requires a new generation of communications payloads powered by large-scale processors, enabling a dynamic allocation of hundreds of beams with a total capacity beyond 1 Tbit s−1. The fact that the scale of the processor is proportional to the wavelength of its signals has made photonics a key technology for its implementation. However, one last challenge hinders the introduction of photonics: while large-scale processors demand a modular implementation, coherency among signals must be preserved using simple methods. Here, we demonstrate a coherent photonic-aided receiver meeting such demands. This work shows that a modular and coherent photonic-aided payload is feasible, making way to an extensive introduction of photonics in next generation communications satellites

Structure of the Vesicular Stomatitis Virus N0-P Complex

Replication of non-segmented negative-strand RNA viruses requires the continuous supply of the nucleoprotein (N) in the form of a complex with the phosphoprotein (P). Here, we present the structural characterization of a soluble, heterodimeric complex between a variant of vesicular stomatitis virus N lacking its 21 N-terminal residues (NΔ21) and a peptide of 60 amino acids (P60) encompassing the molecular recognition element (MoRE) of P that binds RNA-free N (N0). The complex crystallized in a decameric circular form, which was solved at 3.0 Å resolution, reveals how the MoRE folds upon binding to N and competes with RNA binding and N polymerization. Small-angle X-ray scattering experiment and NMR spectroscopy on the soluble complex confirms the binding of the MoRE and indicates that its flanking regions remain flexible in the complex. The structure of this complex also suggests a mechanism for the initiation of viral RNA synthesis

Developing multiscale and integrative nature–people scenarios using the Nature Futures Framework

1. Scientists have repeatedly argued that transformative, multiscale global scenarios are needed as tools in the quest to halt the decline of biodiversity and achieve sustainability goals. 2. As a first step towards achieving this, the researchers who participated in the scenarios and models expert group of the Intergovernmental Science‐Policy Platform on Biodiversity and Ecosystem Services (IPBES) entered into an iterative, participatory process that led to the development of the Nature Futures Framework (NFF). 3. The NFF is a heuristic tool that captures diverse, positive relationships of humans with nature in the form of a triangle. It can be used both as a boundary object for continuously opening up more plural perspectives in the creation of desirable nature scenarios and as an actionable framework for developing consistent nature scenarios across multiple scales. 4. Here we describe the methods employed to develop the NFF and how it fits into a longer term process to create transformative, multiscale scenarios for nature. We argue that the contribution of the NFF is twofold: (a) its ability to hold a plurality of perspectives on what is desirable, which enables the development of joint goals and visions and recognizes the possible convergence and synergies of measures to achieve these visions and (b), its multiscale functionality for elaborating scenarios and models that can inform decision‐making at relevant levels, making it applicable across specific places and perspectives on nature. 5. If humanity is to achieve its goal of a more sustainable and prosperous future rooted in a flourishing nature, it is critical to open up a space for more plural perspectives of human–nature relationships. As the global community sets out to develop new goals for biodiversity, the NFF can be used as a navigation tool helping to make diverse, desirable futures possible

Developing multiscale and integrative nature–people scenarios using the Nature Futures Framework

1. Scientists have repeatedly argued that transformative, multiscale global scenarios are needed as tools in the quest to halt the decline of biodiversity and achieve sustainability goals. 2. As a first step towards achieving this, the researchers who participated in the scenarios and models expert group of the Intergovernmental Science-Policy Platform on Biodiversity and Ecosystem Services (IPBES) entered into an iterative, participatory process that led to the development of the Nature Futures Framework (NFF). 3. The NFF is a heuristic tool that captures diverse, positive relationships of humans with nature in the form of a triangle. It can be used both as a boundary object for continuously opening up more plural perspectives in the creation of desirable nature scenarios and as an actionable framework for developing consistent nature scenarios across multiple scales. 4. Here we describe the methods employed to develop the NFF and how it fits into a longer term process to create transformative, multiscale scenarios for nature. We argue that the contribution of the NFF is twofold: (a) its ability to hold a plurality of perspectives on what is desirable, which enables the development of joint goals and visions and recognizes the possible convergence and synergies of measures to achieve these visions and (b), its multiscale functionality for elaborating scenarios and models that can inform decision-making at relevant levels, making it applicable across specific places and perspectives on nature. 5. If humanity is to achieve its goal of a more sustainable and prosperous future rooted in a flourishing nature, it is critical to open up a space for more plural per- spectives of human–nature relationships. As the global community sets out to de- velop new goals for biodiversity, the NFF can be used as a navigation tool helping to make diverse, desirable futures possible

Accelerated Immunodeficiency by Anti-CCR5 Treatment in HIV Infection

In 50% of progressing HIV-1 patients, CXCR4-tropic (X4) virus emerges late in infection, often overtaking CCR5-tropic (R5) virus as the dominant viral strain. This “phenotypic switch” is strongly associated with rapidly declining CD4+ T cell counts and AIDS onset, yet its causes remain unknown. Here, we analyze a mathematical model for the mechanism of X4 emergence in late-stage HIV infection and use this analysis to evaluate the utility of a promising new class of antiretroviral drugs—CCR5 inhibitors—in dual R5, X4 infection. The model shows that the R5-to-X4 switch occurs as CD4+ T cell activation levels increase above a threshold and as CD4+ T cell counts decrease below a threshold during late-stage HIV infection. Importantly, the model also shows that highly active antiretroviral therapy (HAART) can inhibit X4 emergence but that monotherapy with CCR5 blockers can accelerate X4 onset and immunodeficiency if X4 infection of memory CD4+ T cells occurs at a high rate. Fortunately, when CXCR4 blockers or HAART are used in conjunction with CCR5 blockers, this risk of accelerated immunodeficiency is eliminated. The results suggest that CCR5 blockers will be more effective when used in combination with CXCR4 blockers and caution against CCR5 blockers in the absence of an effective HAART regimen or during HAART failure