The Neisseria gonorrhoeae type IV pilus promotes resistance to hydrogen peroxide- and LL-37-mediated killing by modulating the availability of intracellular,labile iron

The Neisseria gonorrhoeae Type IV pilus is a multifunctional, dynamic fiber involved in host cell attachment, DNA transformation, and twitching motility. We previously reported that the N. gonorrhoeae pilus is also required for resistance against hydrogen peroxide-, antimicrobial peptide LL-37-, and non-oxidative, neutrophil-mediated killing. We tested whether the hydrogen peroxide, LL-37, and neutrophil hypersensitivity phenotypes in non-piliated N. gonorrhoeae could be due to elevated iron levels. Iron chelation in the growth medium rescued a nonpiliated pilE mutant from both hydrogen peroxide- and antimicrobial peptide LL-37-mediated killing, suggesting these phenotypes are related to iron availability. We used the antibiotic streptonigrin, which depends on free cytoplasmic iron and oxidation to kill bacteria, to determine whether piliation affected intracellular iron levels. Several non-piliated, loss-of-function mutants were more sensitive to streptonigrin killing than the piliated parental strain. Consistent with the idea that higher available iron levels in the under- and non-piliated strains were responsible for the higher streptonigrin sensitivity, iron limitation by desferal chelation restored resistance to streptonigrin in these strains and the addition of iron restored the sensitivity to streptonigrin killing. The antioxidants tiron and dimethylthiourea rescued the pilE mutant from streptonigrin-mediated killing, suggesting that the elevated labile iron pool in non-piliated bacteria leads to streptonigrin-dependent reactive oxygen species production. These antioxidants did not affect LL-37-mediated killing. We confirmed that the pilE mutant is not more sensitive to other antibiotics showing that the streptonigrin phenotypes are not due to general bacterial envelope disruption. The total iron content of the cell was unaltered by piliation when measured using ICP-MS suggesting that only the labile iron pool is affected by piliation. These results support the hypothesis that piliation state affects N. gonorrhoeae iron homeostasis and influences sensitivity to various host-derived antimicrobial agents.     

Microbial light-activatable proton pumps as neuronal inhibitors to functionally dissect neuronal networks in C. elegans

Essentially any behavior in simple and complex animals depends on neuronal network function. Currently, the best-defined system to study neuronal circuits is the nematode Caenorhabditis elegans, as the connectivity of its 302 neurons is exactly known. Individual neurons can be activated by photostimulation of Channelrhodopsin-2 (ChR2) using blue light, allowing to directly probe the importance of a particular neuron for the respective behavioral output of the network under study. In analogy, other excitable cells can be inhibited by expressing Halorhodopsin from Natronomonas pharaonis (NpHR) and subsequent illumination with yellow light. However, inhibiting C. elegans neurons using NpHR is difficult. Recently, proton pumps from various sources were established as valuable alternative hyperpolarizers. Here we show that archaerhodopsin-3 (Arch) from Halorubrum sodomense and a proton pump from the fungus Leptosphaeria maculans (Mac) can be utilized to effectively inhibit excitable cells in C. elegans. Arch is the most powerful hyperpolarizer when illuminated with yellow or green light while the action spectrum of Mac is more blue-shifted, as analyzed by light-evoked behaviors and electrophysiology. This allows these tools to be combined in various ways with ChR2 to analyze different subsets of neurons within a circuit. We exemplify this by means of the polymodal aversive sensory ASH neurons, and the downstream command interneurons to which ASH neurons signal to trigger a reversal followed by a directional turn. Photostimulating ASH and subsequently inhibiting command interneurons using two-color illumination of different body segments, allows investigating temporal aspects of signaling downstream of ASH.     

Wetland conservation and sustainable use under global change: a tropical Australian case study using magpie geese

Imminent shifts in environmental parameters due to climatic change might have profound ramifications for wetlands listed under the Ramsar convention. Although the exact mechanisms by which global change will affect these systems are not known, models that simulate component drivers, particularly at a broad spatial scale, can nevertheless allow for more informed conservation decision making. Such general inference is particularly needed for wetlands across the tropics, where less knowledge and fewer resources are available to mitigate the impacts on important conservation sites. Here we develop a case study of wetland loss to sea level rise across tropical north Australia (including Ramsar‐listed sites), and link these to a metapopulation model for a keystone endemic waterbird, the magpie goose Anseranas semipalmata. We projected published models on sea level rise through to the year 2400, and found a non‐linear trajectory of inundation up to 20 m above present levels. Digital elevation models were used to simulate sea level rise and the spatially differentiated loss of wetland habitat used by geese. Range retraction was linked to decline in ecological carrying capacity, and we coupled wetland‐specific habitat loss projections to a spatially explicit demographic metapopulation model. Additionally, we included alternate harvest strategies based on present‐day estimates of indigenous and non‐indigenous offtake of geese, and examined the synergy between wetland loss and hunting on extinction risk. Our results suggest that Australia's once‐abundant and widespread magpie goose will be reduced to a fragmented population of just a few thousand individuals within the next 200–300 yr. Harvest could continue for some time, up to a “tipping point” at around 5% loss of current wetland habitat, after which the decline of geese is rapid. Given the inexorable nature of sea level rise, short‐ to medium‐term conservation of waterbirds across Ramsar wetlands must prepare for adaptive wetland management, such as through buffer‐placement, and ongoing monitoring of harvest.     

Starch mobilization in leaves

Starch mobilization is well understood in cereal endosperms, but both the pathway and the regulation of the process are poorly characterized in other types of plant organs. Arabidopsis leaves offer the opportunity for rapid progress in this area, because of the genomic resources available in this species and the ease with which starch synthesis and degradation can be monitored and manipulated. Progress in understanding three aspects of starch degradation is described: the role of disproportionating enzyme, the importance of phosphorolytic degradation, and new evidence about the involvement of a starch-phosphorylating enzyme in the degradative process. Major areas requiring further research are outlined.     

Late stage antibacterial drugs in the clinical pipeline

Bacterial resistance to antimicrobial agents is a growing problem worldwide. Not only is issue compounded by the fact that there are fewer pharmaceutical companies conducting research to discover novel antimicrobials than in the past but development time lines have stretched so that a dozen years from discovery to the market is now the standard. Eleven antibacterial drugs in late stage clinical development are discussed. Whereas many of these may successfully deal with resistant strains of Gram-positive pathogens, there is very little in development to address the gorwing unmet medical need of multi-drug resistant Gram-negative infections.     

Achieving Molecular Selectivity in Imaging Using Multiphoton Raman Spectroscopy Techniques

In the case of most optical imaging methods, contrast is generated either by physical properties of the sample (Differential Image Contrast, Phase Contrast), or by fluorescent labels that are localized to a particular protein or organelle. Standard Raman and infrared methods for obtaining images are based upon the intrinsic vibrational properties of molecules, and thus obviate the need for attached fluorophores. Unfortunately, they have significant limitations for live-cell imaging. However, an active Raman method, called Coherent Anti-Stokes Raman Scattering (CARS), is well suited for microscopy, and provides a new means for imaging specific molecules. Vibrational imaging techniques, such as CARS, avoid problems associated with photobleaching and photo-induced toxicity often associated with the use of fluorescent labels with live cells. Because the laser configuration needed to implement CARS technology is similar to that used in other multiphoton microscopy methods, such as two-photon fluorescence and harmonic generation, it is possible to combine imaging modalities, thus generating simultaneous CARS and fluorescence images. A particularly powerful aspect of CARS microscopy is its ability to selectively image deuterated compounds, thus allowing the visualization of molecules, such as lipids, that are chemically indistinguishable from the native species.     

The Influences of Environment, Mating Habitat, and Predation on Evolution of Pinniped Lactation Strategies

Seals have adapted their social systems and lactation strategies to marine environments that include open and ice-covered oceans, high and low productivity, extremes in seasonality, and ocean- and terrestrial-type predators. Different explanations for the variation in pinniped lactation systems have been proposed but tests of alternative hypotheses have not sufficiently accounted for phylogeny and body size. After controlling for this variation, I predicted that environment, mating habitat, and predation would yield a fuller explanation. Lactation traits, duration, pup growth rate, and fat content were significantly influenced by both body size and phylogeny, which together explained 20–69% of the variation. After controlling for this variation, initial results did not support the environment hypothesis, as no differences in lactation traits were found between species living in polar (≥60°N) versus equatorial (<60°N) environments. In contrast, seals that nurse in areas of Arctic sea ice contending with ice-hunting predators, such as polar bears, had relatively short lactation compared to species living in the Antarctic and more equatorial regions. Also, the availability of predator-free islands for terrestrial mating and parturition was related to a harem mating system, increased sexual size dimorphism (SSD), and slow juvenile growth rates, less fat in milk, and longer lactation. Using structural equation modeling, latitude and size of harems provided independent explanations for all three lactation traits. Thus, use of islands in ice-free waters, predation in Arctic ice-covered waters, and more milk fat in high-latitude seals together provided adequate explanations for the evolution of lactation diversity among pinnipeds.     

Why is big Pharma getting out of antibacterial drug discovery?

Since the advent of the antibiotic era in the late 1940s drug discovery and development has evolved into an expensive, time consuming, cumbersome and bureaucratic process involving multiple interest groups such as pharmaceutical manufacturers, governmental regulatory authorities, patent officers, academic and clinical researchers and trial lawyers. It would seem that the least involved among the interest groups are the consumers of health care themselves. Politicians and the public alike complain loudly about drug prices although fewer and fewer new therapies are being developed. The cost and complexities of drug discovery and development have shifted the investment equation away from the development of drugs targeting short course therapies for acute diseases and towards long-term treatment of chronic conditions. Coupled with the failure of large investments into target-based approaches to produce novel antibacterial agents, companies large and small have exited from this field despite a growing clinical need.