Weighted-norm preconditioners for a multilayer tide model

We derive a linearized rotating shallow water system modeling tides, which can be discretized by mixed finite elements. Unlike previous models, this model allows for multiple layers stratified by density. Like the single-layer case [R. C. Kirby and T. Kernell, Comput. Math. Appl., 82 (2021), pp. 212–227], a weighted-norm preconditioner gives a (nearly) parameter-robust method for solving the resulting linear system at each time step, but the all-to-all coupling between the layers in the model poses a significant challenge to efficiency. Neglecting the inter-layer coupling gives a preconditioner that degrades rapidly as the number of layers increases. By a careful analysis of the matrix that couples the layers, we derive a robust method that requires solving a reformulated system that only involves coupling between adjacent layers. Numerical results obtained using Firedrake [F. Rathgeber et al., ACM Trans. Math. Software, 43 (2016), 24] confirm the theory

Influenza Virus Infection of the Murine Uterus: A New Model for Antiviral Immunity in the Female Reproductive Tract

Secretory IgA (S-IgA) mediates local immunity to influenza virus in the murine upper respiratory tract and may play an important role in local immunity to various microorganisms in the female reproductive tract as well. Although the presence of IgA in cervicovaginal or uterine secretions has been correlated with immunity to a number of pathogens, there has been no direct demonstration of the mediation of uterine antiviral immunity by S-IgA. Influenza virus, although not a normal pathogen of the reproductive tract, was used to develop a model for the investigation of mucosal immunity in the uterus. PR8 (H1N1) influenza virus injected into the ovarian bursa of BALB/c mice grew well, with peak titers between days 3 and 5. Intravenous injection of polymeric IgA anti-influenza virus monoclonal antibody before or 30 min after viral challenge protected mice against viral infection. We believe this work to be the first direct demonstration of S-IgA-mediated antiviral uterine immunity. It provides a model for further investigation of immunity in the female reproductive tract.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63226/1/vim.2006.19.613.pd

Rapid Immunomagnetic Negative Enrichment of Neutrophil Granulocytes from Murine Bone Marrow for Functional Studies In Vitro and In Vivo

Polymorphonuclear neutrophils (PMN) mediate early immunity to infection but can also cause host damage if their effector functions are not controlled. Their lack or dysfunction is associated with severe health problems and thus the analysis of PMN physiology is a central issue. One prerequisite for PMN analysis is the availability of purified cells from primary organs. While human PMN are easily isolated from peripheral blood, this approach is less suitable for mice due to limited availability of blood. Instead, bone marrow (BM) is an easily available reservoir of murine PMN, but methods to obtain pure cells from BM are limited. We have developed a novel protocol allowing the isolation of highly pure untouched PMN from murine BM by negative immunomagnetic isolation using a complex antibody cocktail. The protocol is simple and fast (∼1 h), has a high yield (5–10*106 PMN per animal) and provides a purity of cells equivalent to positive selection (>80%). Most importantly, cells obtained by this method are non-activated and remain fully functional in vitro or after adoptive transfer into recipient animals. This method should thus greatly facilitate the study of primary murine PMN in vitro and in vivo

Bordetella pertussis Autotransporter Vag8 Binds Human C1 Esterase Inhibitor and Confers Serum Resistance

Bordetella pertussis employs numerous strategies to evade the immune system, including the ability to resist killing via complement. Previously we have shown that B. pertussis binds a complement regulatory protein, C1 esterase inhibitor (C1inh) to its surface in a Bvg-regulated manner (i.e. during its virulence phase), but the B. pertussis factor was not identified. Here we set out to identify the B. pertussis C1inh-binding factor. Using a serum overlay assay, we found that this factor migrates at approximately 100 kDa on an SDS-PAGE gel. To identify this factor, we isolated proteins of approximately 100 kDa from wild type strain BP338 and from BP347, an isogenic Bvg mutant that does not bind C1inh. Using mass spectrometry and bioinformatics, we identified the autotransporter protein Vag8 as the putative C1inh binding protein. To prove that Vag8 binds C1inh, vag8 was disrupted in two different B. pertussis strains, namely BP338 and 18–323, and the mutants were tested for their ability to bind C1inh in a surface-binding assay. Neither mutant strain was capable of binding C1inh, whereas a complemented strain successfully bound C1inh. In addition, the passenger domain of Vag8 was expressed and purified as a histidine-tagged fusion protein and tested for C1inh-binding in an ELISA assay. Whereas the purified Vag8 passenger bound C1inh, the passenger domain of BrkA (a related autotransporter protein) failed to do so. Finally, serum assays were conducted to compare wild type and vag8 mutants. We determined that vag8 mutants from both strains were more susceptible to killing compared to their isogenic wild type counterparts. In conclusion, we have discovered a novel role for the previously uncharacterized protein Vag8 in the immune evasion of B. pertussis. Vag8 binds C1inh to the surface of the bacterium and confers serum resistance

Predation on Multiple Trophic Levels Shapes the Evolution of Pathogen Virulence

The pathogen virulence is traditionally thought to co-evolve as a result of reciprocal selection with its host organism. In natural communities, pathogens and hosts are typically embedded within a web of interactions with other species, which could affect indirectly the pathogen virulence and host immunity through trade-offs. Here we show that selection by predation can affect both pathogen virulence and host immune defence. Exposing opportunistic bacterial pathogen Serratia marcescens to predation by protozoan Tetrahymena thermophila decreased its virulence when measured as host moth Parasemia plantaginis survival. This was probably because the bacterial anti-predatory traits were traded off with bacterial virulence factors, such as motility or resource use efficiency. However, the host survival depended also on its allocation to warning signal that is used against avian predation. When infected with most virulent ancestral bacterial strain, host larvae with a small warning signal survived better than those with an effective large signal. This suggests that larval immune defence could be traded off with effective defence against bird predators. However, the signal size had no effect on larval survival when less virulent control or evolved strains were used for infection suggesting that anti-predatory defence against avian predators, might be less constrained when the invading pathogen is rather low in virulence. Our results demonstrate that predation can be important indirect driver of the evolution of both pathogen virulence and host immunity in communities with multiple species interactions. Thus, the pathogen virulence should be viewed as a result of both past evolutionary history, and current ecological interactions

Genome Mining for Radical SAM Protein Determinants Reveals Multiple Sactibiotic-Like Gene Clusters

Thuricin CD is a two-component bacteriocin produced by Bacillus thuringiensis that kills a wide range of clinically significant Clostridium difficile. This bacteriocin has recently been characterized and consists of two distinct peptides, Trnβ and Trnα, which both possess 3 intrapeptide sulphur to α-carbon bridges and act synergistically. Indeed, thuricin CD and subtilosin A are the only antimicrobials known to possess these unusual structures and are known as the sactibiotics (sulplur to alpha carbon-containing antibiotics). Analysis of the thuricin CD-associated gene cluster revealed the presence of genes encoding two highly unusual SAM proteins (TrnC and TrnD) which are proposed to be responsible for these unusual post-translational modifications. On the basis of the frequently high conservation among enzymes responsible for the post-translational modification of specific antimicrobials, we performed an in silico screen for novel thuricin CD–like gene clusters using the TrnC and TrnD radical SAM proteins as driver sequences to perform an initial homology search against the complete non-redundant database. Fifteen novel thuricin CD–like gene clusters were identified, based on the presence of TrnC and TrnD homologues in the context of neighbouring genes encoding potential bacteriocin structural peptides. Moreover, metagenomic analysis revealed that TrnC or TrnD homologs are present in a variety of metagenomic environments, suggesting a widespread distribution of thuricin-like operons in a variety of environments. In-silico analysis of radical SAM proteins is sufficient to identify novel putative sactibiotic clusters

Essential versus accessory aspects of cell death: recommendations of the NCCD 2015

Cells exposed to extreme physicochemical or mechanical stimuli die in an uncontrollable manner, as a result of their immediate structural breakdown. Such an unavoidable variant of cellular demise is generally referred to as ‘accidental cell death’ (ACD). In most settings, however, cell death is initiated by a genetically encoded apparatus, correlating with the fact that its course can be altered by pharmacologic or genetic interventions. ‘Regulated cell death’ (RCD) can occur as part of physiologic programs or can be activated once adaptive responses to perturbations of the extracellular or intracellular microenvironment fail. The biochemical phenomena that accompany RCD may be harnessed to classify it into a few subtypes, which often (but not always) exhibit stereotyped morphologic features. Nonetheless, efficiently inhibiting the processes that are commonly thought to cause RCD, such as the activation of executioner caspases in the course of apoptosis, does not exert true cytoprotective effects in the mammalian system, but simply alters the kinetics of cellular demise as it shifts its morphologic and biochemical correlates. Conversely, bona fide cytoprotection can be achieved by inhibiting the transduction of lethal signals in the early phases of the process, when adaptive responses are still operational. Thus, the mechanisms that truly execute RCD may be less understood, less inhibitable and perhaps more homogeneous than previously thought. Here, the Nomenclature Committee on Cell Death formulates a set of recommendations to help scientists and researchers to discriminate between essential and accessory aspects of cell death

Cooling athletes with a spinal cord injury

Cooling strategies that help prevent a reduction in exercise capacity whilst exercising in the heat have received considerable research interest over the past 3 decades, especially in the lead up to a relatively hot Olympic and Paralympic Games. Progressing into the next Olympic/Paralympic cycle, the host, Rio de Janeiro, could again present an environmental challenge for competing athletes. Despite the interest and vast array of research into cooling strategies for the able-bodied athlete, less is known regarding the application of these cooling strategies in the thermoregulatory impaired spinal cord injured (SCI) athletic population. Individuals with a spinal cord injury (SCI) have a reduced afferent input to the thermoregulatory centre and a loss of both sweating capacity and vasomotor control below the level of the spinal cord lesion. The magnitude of this thermoregulatory impairment is proportional to the level of the lesion. For instance, individuals with high-level lesions (tetraplegia) are at a greater risk of heat illness than individuals with lower-level lesions (paraplegia) at a given exercise intensity. Therefore, cooling strategies may be highly beneficial in this population group, even in moderate ambient conditions (~21 °C). This review was undertaken to examine the scientific literature that addresses the application of cooling strategies in individuals with an SCI. Each method is discussed in regards to the practical issues associated with the method and the potential underlying mechanism. For instance, site-specific cooling would be more suitable for an athlete with an SCI than whole body water immersion, due to the practical difficulties of administering this method in this population group. From the studies reviewed, wearing an ice vest during intermittent sprint exercise has been shown to decrease thermal strain and improve performance. These garments have also been shown to be effective during exercise in the able-bodied. Drawing on additional findings from the able-bodied literature, the combination of methods used prior to and during exercise and/or during rest periods/half-time may increase the effectiveness of a strategy. However, due to the paucity of research involving athletes with an SCI, it is difficult to establish an optimal cooling strategy. Future studies are needed to ensure that research outcomes can be translated into meaningful performance enhancements by investigating cooling strategies under the constraints of actual competition. Cooling strategies that meet the demands of intermittent wheelchair sports need to be identified, with particular attention to the logistics of the sport

Activity of Bdellovibrio Hit Locus Proteins, Bd0108 and Bd0109, Links Type IVa Pilus Extrusion/Retraction Status to Prey-Independent Growth Signalling

Bdellovibrio bacteriovorus are facultatively predatory bacteria that grow within gram-negative prey, using pili to invade their periplasmic niche. They also grow prey-independently on organic nutrients after undergoing a reversible switch. The nature of the growth switching mechanism has been elusive, but several independent reports suggested mutations in the hit (host-interaction) locus on the Bdellovibrio genome were associated with the transition to preyindependent growth. Pili are essential for prey entry by Bdellovibrio and sequence analysis of the hit locus predicted that it was part of a cluster of Type IVb pilus-associated genes, containing bd0108 and bd0109. In this study we have deleted the whole bd0108 gene, which is unique to Bdellovibrio, and compared its phenotype to strains containing spontaneous mutations in bd0108 and the common natural 42 bp deletion variant of bd0108. We find that deletion of the whole bd0108 gene greatly reduced the extrusion of pili, whereas the 42 bp deletion caused greater pilus extrusion than wild-type. The pili isolated from these strains were comprised of the Type IVa pilin protein; PilA. Attempts to similarly delete gene bd0109, which like bd0108 encodes a periplasmic/secreted protein, were not successful, suggesting that it is likely to be essential for Bdellovibrio viability in any growth mode. Bd0109 has a sugar binding YD- repeat motif and an N-terminus with a putative pilin-like fold and was found to interact directly with Bd0108. These results lead us to propose that the Bd0109/Bd0108 interaction regulates pilus production in Bdellovibrio (possibly by interaction with the pilus fibre at the cell wall), and that the presence (and possibly retraction state) of the pilus feeds back to alter the growth state of the Bdellovibrio cell. We further identify a novel small RNA encoded by the hit locus, the transcription of which is altered in different bd0108 mutation background