Enigmatic dual symbiosis in the excretory organ of Nautilus macromphalus (Cephalopoda: Nautiloidea)

Symbiosis is an important driving force in metazoan evolution and the study of ancient lineages can provide an insight into the influence of symbiotic associations on morphological and physiological adaptations. In the 'living fossil' Nautilus, bacterial associations are found in the highly specialized pericardial appendage. This organ is responsible for most of the excretory processes (ultrafiltration, reabsorption and secretion) and secretes an acidic ammonia-rich excretory fluid. In this study, we show that Nautilus macromphalus pericardial appendages harbour a high density of a beta-proteobacterium and a coccoid spirochaete using transmission electron microscopy, comparative 16S rRNA sequence analysis and fluorescence in situ hybridization (FISH). These two bacterial phylotypes are phylogenetically distant from any known bacteria, with ammonia-oxidizing bacteria as the closest relatives of the beta-proteobacterium (above or equal to 87.5% sequence similarity) and marine Spirochaeta species as the closest relatives of the spirochaete (above or equal to 89.8% sequence similarity), and appear to be specific to Nautilus. FISH analyses showed that the symbionts occur in the baso-medial region of the pericardial villi where ultrafiltration and reabsorption processes take place, suggesting a symbiotic contribution to the excretory metabolism.     

Habitat suitability models for the conservation of thermophilic grasshoppers and bush crickets—simple or complex?

One goal of conservation biology is the assessment of effects of land use change on species distribution. One approach for identifying the factors, which determine habitat suitability for a species are statistical habitat distribution models. These models are quantitative and can be used for predictions in management scenarios. However, they often have one major shortcoming, which is their complexity. This means that they need several, often costly-to-determine parameters for predictions of species occurrence. We first used habitat suitability models to investigate and determine habitat preferences of three different Orthoptera species. Second, we compared the predictive powers of simple habitat suitability models considering only the ‘habitat type’ as predictor with more complex models taking different habitat factors into account. We found that the habitat type is the most reliable and robust factor, which determines the occurrence of the species studied. Thus, analyses of habitat suitability can easily be carried out on the basis of existing vegetation maps for the conservation of the three species under study. Our results can serve as a basis for the estimation of spatio-temporal distribution and survival probabilities of the species studied and might also be valuable for other species living in dry grasslands.     

Resistance to a CRISPR-based gene drive at an evolutionarily conserved site is revealed by mimicking genotype fixation

CRISPR-based homing gene drives can be designed to disrupt essential genes whilst biasing their own inheritance, leading to suppression of mosquito populations in the laboratory. This class of gene drives relies on CRISPR-Cas9 cleavage of a target sequence and copying (‘homing’) therein of the gene drive element from the homologous chromosome. However, target site mutations that are resistant to cleavage yet maintain the function of the essential gene are expected to be strongly selected for. Targeting functionally constrained regions where mutations are not easily tolerated should lower the probability of resistance. Evolutionary conservation at the sequence level is often a reliable indicator of functional constraint, though the actual level of underlying constraint between one conserved sequence and another can vary widely. Here we generated a novel adult lethal gene drive (ALGD) in the malaria vector Anopheles gambiae, targeting an ultra-conserved target site in a haplosufficient essential gene (AGAP029113) required during mosquito development, which fulfils many of the criteria for the target of a population suppression gene drive. We then designed a selection regime to experimentally assess the likelihood of generation and subsequent selection of gene drive resistant mutations at its target site. We simulated, in a caged population, a scenario where the gene drive was approaching fixation, where selection for resistance is expected to be strongest. Continuous sampling of the target locus revealed that a single, restorative, in-frame nucleotide substitution was selected. Our findings show that ultra-conservation alone need not be predictive of a site that is refractory to target site resistance. Our strategy to evaluate resistance in vivo could help to validate candidate gene drive targets for their resilience to resistance and help to improve predictions of the invasion dynamics of gene drives in field populations.     

Effects of insecticide applications on macroinvertebrate density and biomass in rice-fields in the Rhône-delta,France

The density of 23 macroinvertebrate species and the total macroinvertebrate biomass were compared between rice-fields treated with lindane and diazinon in June and alphamethine in August and untreated controls. The macroinvertebrates could be divided into four groups: (1) Taxa, in which the densities were lower in the insecticide treatment in July and August than in the non-insecticide treatment. (2) The Culicidae which occurred in the insecticide treatment in significantly lower density in July, but in significantly higher density in August. (3) Ischnura elegans (Vander L.) which was found in July after the lindane application in significantly higher numbers in the insecticide treatments, but in significantly lower numbers in the insecticide treatment in August after the application of the pyrethroid. In these three groups, we assumed that direct effects due to the insecticides toxicity were the reason for the differences in density. (4) The fourth group included three taxa in which the densities were significantly higher in the insecticide treatment in July and August than in the control. For this, indirect effects due to reduced biotic interactions may be responsible. The biomass was higher in the insecticide treatments in July, mainly because of a high increase in gastropod density, during the rest of the season it was similar between treatments and controls.     

The Lck inhibitor,AMG-47a,blocks necroptosis and implicates RIPK1 in signalling downstream of MLKL

Necroptosis is a form of caspase-independent programmed cell death that arises from disruption of cell membranes by the mixed lineage kinase domain-like (MLKL) pseudokinase after its activation by the upstream kinases, receptor interacting protein kinase (RIPK)-1 and RIPK3, within a complex known as the necrosome. Dysregulated necroptosis has been implicated in numerous inflammatory pathologies. As such, new small molecule necroptosis inhibitors are of great interest, particularly ones that operate downstream of MLKL activation, where the pathway is less well defined. To better understand the mechanisms involved in necroptosis downstream of MLKL activation, and potentially uncover new targets for inhibition, we screened known kinase inhibitors against an activated mouse MLKL mutant, leading us to identify the lymphocyte-specific protein tyrosine kinase (Lck) inhibitor AMG-47a as an inhibitor of necroptosis. We show that AMG-47a interacts with both RIPK1 and RIPK3, that its ability to protect from cell death is dependent on the strength of the necroptotic stimulus, and that it blocks necroptosis most effectively in human cells. Moreover, in human cell lines, we demonstrate that AMG-47a can protect against cell death caused by forced dimerisation of MLKL truncation mutants in the absence of any upstream signalling, validating that it targets a process downstream of MLKL activation. Surprisingly, however, we also found that the cell death driven by activated MLKL in this model was completely dependent on the presence of RIPK1, and to a lesser extent RIPK3, although it was not affected by known inhibitors of these kinases. Together, these results suggest an additional role for RIPK1, or the necrosome, in mediating human necroptosis after MLKL is phosphorylated by RIPK3 and provide further insight into reported differences in the progression of necroptosis between mouse and human cells.Subject terms: Kinases, Necroptosis     

Modelling complexation and electrostatic attraction in heavy metal biosorption by Sargassum biomass

Biosorption, the passive accumulation of metal ions by biomass, can be used for purifying metal bearing wastewater. Seaweeds represent a readily available source of biosorbent material that possesses a high metal binding capacity. For example, Sargassum can accumulate 2 mequiv of Cd per gram of biomass i.e. 10% of its dry weight. Binding of Cd and Cu by Sargassum is an ion exchange process involving both covalent and ionic bonds. The amount of cations bound covalently or by complexation can be predicted using multi-component sorption isotherms involving 2 types of binding sites, carboxyl and sulphate. A Donnan model was used to account for the effect of ionic strength and electrostatic attraction. The use of a multi-component isotherm that included one term for Na binding was less appropriate than the Donnan model for modelling ionic strength effects. It was possible to predict metal and proton binding as a function of the pH value, metal concentration and ionic strength of the solution. This revised version was published online in June 2006 with corrections to the Cover Date.     

Production of poly(4-hydroxybutyric acid) by fed-batch cultures of recombinant strains of Escherichia coli

A recombinant strain of Escherichia coli was used to produce poly(4-hydroxybutyric acid), P(4HB), homopolyester by fed-batch culture in M9 mineral salts medium containing glucose and 4-hydroxybutyric acid as carbon sources. The final cell dry weight, P(4HB) concentration and P(4HB) content were 12.6 g/l, 4.4 g/l, and 36% of cell dry weight, respectively, in a 27-l stirred and aerated fermenter after 60 h of fed-batch fermentation at constant pH.