Cross-reactivity and immunotherapeutic potential of BamA recombinant protein from Acinetobacter baumannii

Acinetobacter baumannii is an important nosocomial pathogen. BamA is a protein that belongs to a complex responsible for organizing the proteins on the bacterial outer membrane. In this work, we aimed to evaluate murine immune responses to BamA recombinant protein (rAbBamA) from A. baumannii in an animal model of infection, and to assess cross-reactivity of this target for the development of anti-A. baumannii vaccines or diagnostics. Immunization of mice with rAbBamA elicited high antibody titers and antibody recognition of native A. baumannii BamA. Immunofluorescence also detected binding to the bacterial surface. After challenge, immunized mice demonstrated a 40% survival increase and better bacterial clearance in kidneys. Immunoblot of anti-rAbBamA against other medically relevant bacteria showed binding to proteins of approximately 35 kDa in Klebsiella pneumoniae and Escherichia coli lysates, primarily identified as OmpA and OmpC, respectively. Altogether, our data show that anti-rAbBamA antibodies provide a protective response against A. baumannii infection in mice. However, the response elicited by immunization with rAbBamA is not completely specific to A. baumannii. Although a broad-spectrum vaccine that protects against various pathogens is an appealing strategy, antibody reactivity against the human microbiota is undesired. In fact, immunization with rAbBamA produced noticeable effects on the gut microbiota. However, the changes elicited were small and non-specific, given that no significant changes in the abundance of Proteobacteria were observed. Overall, rAbBamA is a promising target, but specificity must be considered in the development of immunological tools against A. baumannii.     

Arp2/3 promotes junction formation and maintenance in the Caenorhabditis elegans intestine by regulating membrane association of apical proteins

It has been proposed that Arp2/3, which promotes nucleation of branched actin, is needed for epithelial junction initiation but is less important as junctions mature. We focus here on how Arp2/3 contributes to the Caenorhabditis elegans intestinal epithelium and find important roles for Arp2/3 in the maturation and maintenance of junctions in embryos and adults. Electron microscope studies show that embryos depleted of Arp2/3 form apical actin-rich microvilli and electron-dense apical junctions. However, whereas apical/basal polarity initiates, apical maturation is defective, including decreased apical F-actin enrichment, aberrant lumen morphology, and reduced accumulation of some apical junctional proteins, including DLG-1. Depletion of Arp2/3 in adult animals leads to similar intestinal defects. The DLG-1/AJM-1 apical junction proteins, and the ezrin-radixin-moesin homologue ERM-1, a protein that connects F-actin to membranes, are required along with Arp2/3 for apical F-actin enrichment in embryos, whereas cadherin junction proteins are not. Arp2/3 affects the subcellular distribution of DLG-1 and ERM-1. Loss of Arp2/3 shifts both ERM-1 and DLG-1 from pellet fractions to supernatant fractions, suggesting a role for Arp2/3 in the distribution of membrane-associated proteins. Thus, Arp2/3 is required as junctions mature to maintain apical proteins associated with the correct membranes.     

Successful aging and sustained good health in the naked mole rat: a long-lived mammalian model for biogerontology and biomedical research

Naked mole rats (NMRs; Heterocephalus glaber) are the longest-living rodents known, with a maximum lifespan of 30 years--5 times longer than expected on the basis of body size. These highly social mouse-sized rodents, naturally found in subterranean burrows in the arid and semiarid regions of the horn of Africa, are commonly used in behavioral, neurological, and ecophysiological research. Very old NMRs (>28 years), like humans, show signs of age-associated pathologies (e.g., muscle loss) as well as the accumulation of lipofuscin pigments, but no signs of tumorigenesis. Indeed, for at least 80% of their lives NMRs maintain normal activity, body composition, and reproductive and physiological functions with no obvious age-related increases in morbidity or mortality rate. Their long lifespan is attributed to sustained good health and pronounced cancer resistance. Clearly physiological and biochemical processes in this species have evolved to dramatically extend both their good health- and lifespan. We and others have tested various current theories using this species as an exceptionally long-lived animal model of successful abrogated aging. Surprisingly, NMRs have high levels of oxidative stress and relatively short telomeres, yet they are extremely resilient when subjected to cellular stressors and appear capable of sustaining both their genomic and protein integrity under hostile conditions. The challenge is to understand how these animals are able to do this. Elucidating these mechanisms will provide useful information for enhancing human life- and healthspan, making the naked mole rat a true "supermodel" for aging research and resistance to chronic age-associated diseases.     

Endocytosis and toxicity of clostridial binary toxins depend on a clathrin‐independent pathway regulated by Rho‐GDI

Clostridial binary toxins, such as Clostridium perfringens Iota and Clostridium botulinum C2, are composed of a binding protein (Ib and C2II respectively) that recognizes distinct membrane receptors and mediates internalization of a catalytic protein (Ia and C2‐I respectively) with ADP‐ribosyltransferase activity that disrupts the actin cytoskeleton. We show here that the endocytic pathway followed by these toxins is independent of clathrin but requires the activity of dynamin and is regulated by Rho‐GDI. This endocytic pathway is similar to a recently characterized clathrin‐independent pathway followed by the interleukin‐2 (IL2) receptor. We found indeed that Ib and C2II colocalized intracellularly with the IL2 receptor but not the transferrin receptor after different times of endocytosis. Accordingly, the intracellular effects of Iota and C2 on the cytoskeleton were inhibited by inactivation of dynamin or by Rho‐GDI whereas inhibitors of clathrin‐dependent endocytosis had no protective effect.     

Informed consent and genomic incidental findings: IRB chair perspectives

It is unclear how genomic incidental finding (GIF) prospects should be addressed in informed consent processes. An exploratory study on this topic was conducted with 34 purposively sampled Chairs of institutional review boards (IRBs) at centers conducting genome-wide association studies. Most Chairs (96%) reported no knowledge of local IRB requirements regarding GIFs and informed consent. Chairs suggested consent processes should address the prospect of, and study disclosure policy on, GIFs; GIF management and follow-up; potential clinical significance of GIFs; potential risks of GIF disclosure; an opportunity for participants to opt out of GIF disclosure; and duration of the researcher's duty to disclose GIFs. Chairs were concerned about participant disclosure preferences changing over time; inherent limitations in determining the scope and accuracy of claims about GIFs; and making consent processes longer and more complex. IRB Chair and other stakeholder perspectives can help advance informed consent efforts to accommodate GIF prospects.     

Spatial dynamics of human-origin H1 influenza A virus in North American swine

The emergence and rapid global spread of the swine-origin H1N1/09 pandemic influenza A virus in humans underscores the importance of swine populations as reservoirs for genetically diverse influenza viruses with the potential to infect humans. However, despite their significance for animal and human health, relatively little is known about the phylogeography of swine influenza viruses in the United States. This study utilizes an expansive data set of hemagglutinin (HA1) sequences (n = 1516) from swine influenza viruses collected in North America during the period 2003-2010. With these data we investigate the spatial dissemination of a novel influenza virus of the H1 subtype that was introduced into the North American swine population via two separate human-to-swine transmission events around 2003. Bayesian phylogeographic analysis reveals that the spatial dissemination of this influenza virus in the US swine population follows long-distance swine movements from the Southern US to the Midwest, a corn-rich commercial center that imports millions of swine annually. Hence, multiple genetically diverse influenza viruses are introduced and co-circulate in the Midwest, providing the opportunity for genomic reassortment. Overall, the Midwest serves primarily as an ecological sink for swine influenza in the US, with sources of virus genetic diversity instead located in the Southeast (mainly North Carolina) and South-central (mainly Oklahoma) regions. Understanding the importance of long-distance pig transportation in the evolution and spatial dissemination of the influenza virus in swine may inform future strategies for the surveillance and control of influenza, and perhaps other swine pathogens.     

Root tips moving through soil: An intrinsic vulnerability

Root elongation occurs by the generation of new cells from meristematic tissue within the apical 1–2 mm region of root tips. Therefore penetration of the soil environment is carried out by newly synthesized plant tissue, whose cells are inherently vulnerable to invasion by pathogens. This conundrum, on its face, would seem to reflect an intolerable risk to the successful establishment of root systems needed for plant life. Yet root tip regions housing the meristematic tissues repeatedly have been found to be free of microbial infection and colonization. Even when spore germination, chemotaxis, and/or growth of pathogens are stimulated by signals from the root tip, the underlying root tissue can escape invasion. Recent insights into the functions of root border cells, and the regulation of their production by transient exposure to external signals, may shed light on long-standing observations.Key words: border cells, chemotaxis, zoospores, neutrophil extracellular traps (NETs)…The evidence suggests that there has evolved within plants, mechanisms for extremely rapid adjustment to changes in the soil environment. The logical conclusion is that plants can and do selectively manipulate the ecological balances within the rhizosphere to their own advantage.¹“Sloughed root cap cells” that detach from the root tip were long presumed to be moribund tissue serving to lubricate passage of the elongating root.² The discovery nearly a century ago that these cells from Zea mays L. and Pisum sativum L. can remain 100% viable for weeks after detachment into hydroponic culture did not alter this perception.³ In recent decades, studies have shown that the cells from root caps of most species are metabolically active and can survive even after detachment into the soil.⁴ Moreover, the cell populations express distinct patterns of gene expression reflecting tissue specialization and were therefore given the name root ‘border’ cells.⁵ Like ‘border towns’ that exist at the boundary of disparate countries and cultures, border cells are part of the plant and part of the soil, yet distinct from both.The soil is a dynamic environment whose pH, surface charge, water availability, texture and composition can range markedly on a large and small scale.^1,6,7 The concept of a ‘microniche’ emphasizes that the biological requirements for a particular soil microorganism may be met within one site but not another site only a micron away.⁸ Thus, the rhizosphere—the region adjacent to root surfaces—can support much higher levels of microorganisms than bulk soil a few millimeters distant.⁹ This phenomenon is recognized to be driven by an increased availability of nutrients released from plants into the external environment.¹⁰ Less well recognized is the dynamic variation that occurs along the root surface, and its significance in patterns of disease development. As roots emerge and the new tissue differentiates progressively through stages from root cap, root apical meristem, elongation zone, and finally mature roots with lignified cell walls, the material released into the environment also changes.^11–13 More than 90% of bulk carbon released from young roots of legumes is delivered by the root cap, a 1 mm zone at the apex.¹⁴ Some pathogens are attracted specifically to the root tip region, presumably in response to such exudates.^15,16 For example, instantaneous swarming occurs when a cotton root is placed into a suspension of Pythium dissotocum zoospores (Sup. Fig. 1). This host-specific attraction is specific to the root tip region where border cells are present (Sup. Fig. 2). Border cells remain attractive to zoospores when removed from the root (Sup. Fig. 3). The nature of the attractant is not known, but its impact is localized and transient (Sup. Fig. 4).Newly generated tissue is highly susceptible to infection by pathogens, in general, so elongating root tips would be predicted to be vulnerable to invasion. And yet, root apices repeatedly have been found to escape infection and colonization.^17–19 Recent discoveries about parallels between mammalian white blood cells and root border cells may provide new insight into this apparent conundrum.²⁰ Neutrophils, a type of white blood cell, are produced in response to infection. Neutrophil extracellular traps (NETs) then attract and kill the invader through a process that requires extracellular DNA (exDNA) and an array of extracellular proteins.^21,22 Border cell production, like that of neutrophils, also is induced in response to signals from pathogens and root tip resistance to infection requires exDNA and an array of extracellular proteins.^20,23 Root tip specific chemotaxis, like that seen with Pythium zoospores, has been presumed to involve steps in a process of pathogen invasion.^15,16 It may, instead, involve a process of extracellular trapping and killing by cells designed to protect root meristems from invasion, in a manner analogous to that which occurs in mammalian defense. If tests confirm this model, the mystery of how root tips escape infection by soilborne pathogens they attract could be resolved.     

Curcumin inhibits growth of Saccharomyces cerevisiae through iron chelation

Curcumin, a polyphenol derived from turmeric, is an ancient therapeutic used in India for centuries to treat a wide array of ailments. Interest in curcumin has increased recently, with ongoing clinical trials exploring curcumin as an anticancer therapy and as a protectant against neurodegenerative diseases. In vitro, curcumin chelates metal ions. However, although diverse physiological effects have been documented for this compound, curcumin's mechanism of action on mammalian cells remains unclear. This study uses yeast as a model eukaryotic system to dissect the biological activity of curcumin. We found that yeast mutants lacking genes required for iron and copper homeostasis are hypersensitive to curcumin and that iron supplementation rescues this sensitivity. Curcumin penetrates yeast cells, concentrates in the endoplasmic reticulum (ER) membranes, and reduces the intracellular iron pool. Curcumin-treated, iron-starved cultures are enriched in unbudded cells, suggesting that the G(1) phase of the cell cycle is lengthened. A delay in cell cycle progression could, in part, explain the antitumorigenic properties associated with curcumin. We also demonstrate that curcumin causes a growth lag in cultured human cells that is remediated by the addition of exogenous iron. These findings suggest that curcumin-induced iron starvation is conserved from yeast to humans and underlies curcumin's medicinal properties.     

Prevalence and patterns of nitrosatable drug use among U.S. women during early pregnancy

BACKROUND

Experimental evidence indicates that certain drugs, that are secondary or tertiary amines or amides, form N‐nitroso compounds in the presence of nitrite in an acidic environment. Nitrosatable drugs have been associated with birth defects in a few epidemiologic studies. This study describes the prevalence and patterns of nitrosatable drug use among U.S. women during early pregnancy and examines maternal factors associated with such use.

METHODS

Data were analyzed from the National Birth Defects Prevention Study and included 6807 mothers who gave birth to babies without major congenital malformations during 1997 to 2005. Information was collected by telephone interview about medication use, demographic factors, and maternal health. Drugs taken during the first trimester were classified according to nitrosatability, amine and amide functional groups, and primary indication of use.

RESULTS

Approximately 24% of the women took one or more nitrosatable drugs during the first trimester, including 12.4%, 12.2%, and 7.6% who respectively took secondary amines, tertiary amines, or amides. Five of the ten most commonly taken drugs were available over the counter. Women who were non‐Hispanic white (29.5%), with 1 year or more college education (27.3%) or 40 years or older (28.8%) had the highest prevalence of use. Supplemental vitamin C, an inhibitor of nitrosation, was not taken by 41.6% and 19.3% of nitrosatable drug users during the first and second months of pregnancy, respectively.

CONCLUSIONS

In this U.S. population, ingestion of drugs classified as nitrosatable was common during the first trimester of pregnancy, especially among non‐Hispanic white, more educated, and older mothers. Birth Defects Research (Part A) 2011. © 2011 Wiley‐Liss, Inc.