M2 Polarization of Human Macrophages Favors Survival of the Intracellular Pathogen Chlamydia pneumoniae

Intracellular pathogens have developed various strategies to escape immunity to enable their survival in host cells, and many bacterial pathogens preferentially reside inside macrophages, using diverse mechanisms to penetrate their defenses and to exploit their high degree of metabolic diversity and plasticity. Here, we characterized the interactions of the intracellular pathogen Chlamydia pneumoniae with polarized human macrophages. Primary human monocytes were pre-differentiated with granulocyte macrophage colony-stimulating factor or macrophage colony-stimulating factor for 7 days to yield M1-like and M2-like macrophages, which were further treated with interferon-γ and lipopolysaccharide or with interleukin-4 for 48 h to obtain fully polarized M1 and M2 macrophages. M1 and M2 cells exhibited distinct morphology with round or spindle-shaped appearance for M1 and M2, respectively, distinct surface marker profiles, as well as different cytokine and chemokine secretion. Macrophage polarization did not influence uptake of C. pneumoniae, since comparable copy numbers of chlamydial DNA were detected in M1 and M2 at 6 h post infection, but an increase in chlamydial DNA over time indicating proliferation was only observed in M2. Accordingly, 72±5% of M2 vs. 48±7% of M1 stained positive for chlamydial lipopolysaccharide, with large perinuclear inclusions in M2 and less clearly bordered inclusions for M1. Viable C. pneumoniae was present in lysates from M2, but not from M1 macrophages. The ability of M1 to restrict chlamydial replication was not observed in M1-like macrophages, since chlamydial load showed an equal increase over time for M1-like and M2-like macrophages. Our findings support the importance of macrophage polarization for the control of intracellular infection, and show that M2 are the preferred survival niche for C. pneumoniae. M1 did not allow for chlamydial proliferation, but failed to completely eliminate chlamydial infection, giving further evidence for the ability of C. pneumoniae to evade cellular defense and to persist in human macrophages.     

Rivers,not refugia,drove diversification in arboreal,sub‐Saharan African snakes

The relative roles of rivers versus refugia in shaping the high levels of species diversity in tropical rainforests have been widely debated for decades. Only recently has it become possible to take an integrative approach to test predictions derived from these hypotheses using genomic sequencing and paleo‐species distribution modeling. Herein, we tested the predictions of the classic river, refuge, and river‐refuge hypotheses on diversification in the arboreal sub‐Saharan African snake genus Toxicodryas. We used dated phylogeographic inferences, population clustering analyses, demographic model selection, and paleo‐distribution modeling to conduct a phylogenomic and historical demographic analysis of this genus. Our results revealed significant population genetic structure within both Toxicodryas species, corresponding geographically to river barriers and divergence times from the mid‐Miocene to Pliocene. Our demographic analyses supported the interpretation that rivers are indications of strong barriers to gene flow among populations since their divergence. Additionally, we found no support for a major contraction of suitable habitat during the last glacial maximum, allowing us to reject both the refuge and river‐refuge hypotheses in favor of the river‐barrier hypothesis. Based on conservative interpretations of our species delimitation analyses with the Sanger and ddRAD data sets, two new cryptic species are identified from east‐central Africa. This study highlights the complexity of diversification dynamics in the African tropics and the advantages of integrative approaches to studying speciation in tropical regions.     

Management intensity and vegetation complexity affect web-building spiders and their prey

Agricultural management and vegetation complexity affect arthropod diversity and may alter trophic interactions between predators and their prey. Web-building spiders are abundant generalist predators and important natural enemies of pests. We analyzed how management intensity (tillage, cutting of the vegetation, grazing by cattle, and synthetic and organic inputs) and vegetation complexity (plant species richness, vegetation height, coverage, and density) affect rarefied richness and composition of web-building spiders and their prey with respect to prey availability and aphid predation in 12 habitats, ranging from an uncut fallow to a conventionally managed maize field. Spiders and prey from webs were collected manually and the potential prey were quantified using sticky traps. The species richness of web-building spiders and the order richness of prey increased with plant diversity and vegetation coverage. Prey order richness was lower at tilled compared to no-till sites. Hemipterans (primarily aphids) were overrepresented, while dipterans, hymenopterans, and thysanopterans were underrepresented in webs compared to sticky traps. The per spider capture efficiency for aphids was higher at tilled than at no-till sites and decreased with vegetation complexity. After accounting for local densities, 1.8 times more aphids were captured at uncut compared to cut sites. Our results emphasize the functional role of web-building spiders in aphid predation, but suggest negative effects of cutting or harvesting. We conclude that reduced management intensity and increased vegetation complexity help to conserve local invertebrate diversity, and that web-building spiders at sites under low management intensity (e.g., semi-natural habitats) contribute to aphid suppression at the landscape scale.     

A novel feeding behaviour in wild redfronted lemurs (Eulemur rufifrons): depletion of spider nests

Reports on behavioural innovations in wild primate populations as well as on their transmission are rare. Here, we report observations suggesting that redfronted lemurs (Eulemur rufifrons) invent new behaviours in the wild. We observed a novel feeding behaviour in redfronted lemurs in Kirindy Forest, western Madagascar. The feeding behaviour consisted of depletion of nests of a social spider species (Stegodyphus sp.). The behaviour was observed in only one out of four study groups, although spider nests were present in the home ranges of all four groups. The behaviour was exhibited in three different years (2009, 2011, 2012) and appears to be re-invented from time to time. Interestingly, in 2011 this behaviour was shown by four individuals and probably spread within the group. This feeding behaviour has only been observed between the middle of June and early August (i.e. the early dry season), and nests were found to be empty later on, suggesting that these nests are available as a food source only seasonally. Our observation contributes a rare case of behavioural innovations in a wild primate population.     

A limited survey of aflatoxin B1 contamination in Indonesian palm kernel cake and copra meal sampled from batches

Samples from large (100–200 tons) batches of palm kernel cake (PKC, n?=?20) and copra meal (CM, n?=?13) were collected at production facilities of four Indonesian feed mill manufacturers and analysed for aflatoxin B₁ (AFB₁) by ELISA. Recoveries using spiked samples ranged from 86 to 113 %, with relative standard deviations of <9 % (PKC) and <6 % (CM). All batches were positive for AFB₁: in PKC, at levels of 5.8–93.1 μg/kg (mean 49 μg/kg), and in CM, at levels of 1.1–147 μg/kg (mean 38.1 μg/kg). AFB₁ levels were, in most batches, below the maximum level (100 μg/kg) recommended by the National Standardisation Agency, Republic of Indonesia. However, about half of the batches exceeded both the European Union and USA regulations for AFB₁ in animal feed. In conclusion, serious efforts are necessary to control production, storage and shipment of palm kernel cake and copra meal for feed purposes, and clearly not only for products intended for export but also to reduce AFB₁ levels in domestic Indonesian feed.     

Extreme genetic depauperation and differentiation of both populations and species in Eurasian feather grasses (Stipa)

A highly selfing breeding system affects gene flow, which may have consequences for patterns of genetic variation and differentiation on both the population and species level. Feather grasses (Stipa spp.) are dominant elements of Eurasian steppes that persist in Central Europe in scattered isolated populations that are of great conservation interest. Cleistogamy is common in the Stipa pennata group, the phylogeny of which is largely unresolved. Intraspecific patterns of genetic variation can be characterised by lack of gene flow due to selfing, but also by large-scale historical migrations and long-term isolation. We analysed both 5 species within the S. pennata group and 33 populations of Stipa pulcherrima sampled across a large part of its range. Using AFLP markers we assessed phylogenetic relationships of the S. pennata group and patterns of genetic variation within and among populations. The S. pennata group formed a consistent clade separated from S. capillata. Stipa pulcherrima was sister to S. eriocaulis, but the relationships among S. pennata s. str., S. borysthenica., and S. tirsa remained unresolved. Within-population genetic variation was extremely low in all species of the S. pennata group (H _e = 0.0–0.013). In S. pulcherrima, genetic variation was consistently relatively high in the east (Romania, Russia) and declined toward western populations, with many populations at the western range edge lacking genetic variation entirely. Populations were strongly differentiated (F _ST = 0.762), and this differentiation did not follow a classical pattern of isolation by distance. Bayesian cluster analysis revealed nine gene pools in S. pulcherrima, which were mostly geographically clustered. Overall the results suggest that S. pulcherrima and species of the S. pennata group are characterised by a cleistogamous breeding system leading to extremely low levels of genetic variation and high levels of population differentiation at both the population and species level. Postglacial colonisation, current population isolation, and population bottlenecks at the western range periphery have further reduced genetic variation and obviated gene exchange. Thus, genetic variation can only be preserved by the conservation of multiple populations.     

Protective effects of dietary EPA and DHA on ischemia–reperfusion-induced intestinal stress

The immunoregulatory effects of dietary omega-3 fatty acids are still not fully characterized. The aim of this study was to determine whether dietary eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) intake limits intestinal ischemia–reperfusion (IR) injury. To test this, rats were fed either control or EPA/DHA supplemented diet for 3 weeks following which they underwent either a sham or an IR surgical protocol. A significant reduction in mucosal damage was observed after EPA/DHA supplemented diet as reflected by maintenance of total protein content. To address the underlying mechanisms of protection, we measured parameters of oxidative stress, intestinal and serological cytokines and intestinal eicosanoids. Interestingly, EPA/DHA fed animals displayed a higher activity of oxidative stress enzyme machinery, i.e., superoxide dismutase and catalase in addition to a reduction in total nitrate/nitrite content. While no changes in cytokines were observed, eicosanoid analyses of intestinal tissue revealed an increase in metabolites of the 12-lipoxygenase pathway following IR. Further, IR in EPA/DHA fed animals was accompanied by a significant increase of 17,18-epoxyeicosatetraenoic acid, 8-Iso prostaglandin F_3α and thromboxane B₃, by more than 12-, 6-, 3-fold, respectively. Thus, the data indicate that EPA/DHA supplementation may be able to reduce early intestinal IR injury by anti-oxidative and anti-inflammatory mechanisms.     

Similar performance in central and range‐edge populations of a Eurasian steppe grass under different climate and soil pH regimes

The abundant centre hypothesis predicts that changing environmental conditions are detrimental to a species’ abundance and performance towards the periphery of its range. We tested these predictions for the perennial grass Stipa capillata, a species that is commonly found in steppes of Asia but is rare at its north‐western range edge, in central Europe. We compared 21 populations in dry grassland fragments in central Europe and 20 populations in steppe habitats of Kazakhstan. We studied 15 plant performance traits both in situ and under laboratory and common greenhouse conditions, including local density, plant size and biomass production, seed size, weight and viability. To assess environmental conditions, we assembled data on topography, soil properties and climatic parameters. Using variance components analysis and multivariate methods we analyzed whether plant performance and environmental attributes differed more, as predicted, between the core and peripheral regions or whether they differed more among their subregions or populations. Additionally, we tested whether performance was affected by the same set of environmental predictors in each region. Contrary to our expectations, plant performance traits were virtually unchanged between the range periphery and centre. As expected, macroclimatic conditions showed a significant difference between the two regions (annual mean temperature, annual precipitation). The only other measured environmental variable that differed significantly between the two regions was soil pH, which was lower in core‐range populations. Our findings thus do not support the notion of reduced performance at the range edge. Instead, our data lend support to earlier theories of relative habitat constancy, suggesting that peripheral populations can shift to other habitats through plasticity or adaptation.     

CK2 phosphorylation of an acidic Ser/Thr di-isoleucine motif in the Na+/H+ exchanger NHE5 isoform promotes association with beta-arrestin2 and endocytosis

Internalization of the Na(+)/H(+) exchanger NHE5 into recycling endosomes is enhanced by the endocytic adaptor proteins β-arrestin1 and -2, best known for their preferential recognition of ligand-activated G protein-coupled receptors (GPCRs). However, the mechanism underlying their atypical association with non-GPCRs, such as NHE5, is unknown. In this study, we identified a highly acidic, serine/threonine-rich, di-isoleucine motif (amino acids 697-723) in the cytoplasmic C terminus of NHE5 that is recognized by β-arrestin2. Gross deletions of this site decreased the state of phosphorylation of NHE5 as well as its binding and responsiveness to β-arrestin2 in intact cells. More refined in vitro analyses showed that this site was robustly phosphorylated by the acidotropic protein kinase CK2, whereas other kinases, such as CK1 or the GPCR kinase GRK2, were considerably less potent. Simultaneous mutation of five Ser/Thr residues within 702-714 to Ala ((702)ST/AA(714)) abolished phosphorylation and binding of β-arrestin2. In transfected cells, the CK2 catalytic α subunit formed a complex with NHE5 and decreased wild-type but not (702)ST/AA(714) NHE5 activity, further supporting a regulatory role for this kinase. The rate of internalization of (702)ST/AA(714) was also diminished and relatively insensitive to overexpression of β-arrestin2. However, unlike in vitro, this mutant retained its ability to form a complex with β-arrestin2 despite its lack of responsiveness. Additional mutations of two di-isoleucine-based motifs (I697A/L698A and I722A/I723A) that immediately flank the acidic cluster, either separately or together, were required to disrupt their association. These data demonstrate that discrete elements of an elaborate sorting signal in NHE5 contribute to β-arrestin2 binding and trafficking along the recycling endosomal pathway.