Haemosporidians are a diverse group of vector-borne parasitic protozoa that includes the agents of human malaria; however, most of the described species are found in birds and reptiles. Although our understanding of these parasites’ diversity has expanded by analyses of their mitochondrial genes, there is limited information on these genes’ evolutionary rates. Here, 114 mitochondrial genomes (mtDNA) were studied from species belonging to four genera: Leucocytozoon, Haemoproteus, Hepatocystis, and Plasmodium. Contrary to previous assertions, the mtDNA is phylogenetically informative. The inferred phylogeny showed that, like the genus Plasmodium, the Leucocytozoon and Haemoproteus genera are not monophyletic groups. Although sensitive to the assumptions of the molecular dating method used, the estimated times indicate that the diversification of the avian haemosporidian subgenera/genera took place after the Cretaceous–Paleogene boundary following the radiation of modern birds. Furthermore, parasite clade differences in mtDNA substitution rates and strength of negative selection were detected. These differences may affect the biological interpretation of mtDNA gene lineages used as a proxy to species in ecological and parasitological investigations. Given that the mitochondria are critically important in the parasite life cycle stages that take place in the vector and that the transmission of parasites belonging to particular clades has been linked to specific insect families/subfamilies, this study suggests that differences in vectors have affected the mode of evolution of haemosporidian mtDNA genes. The observed patterns also suggest that the radiation of haemosporidian parasites may be the result of community-level evolutionary processes between their vertebrate and invertebrate hosts.

We present a simple, automated method for high-throughput formation of droplet interface bilayers (DIBs) in a microfluidic device. We can form complex DIB networks that are able to fill predefined three dimensional architectures. Moreover, we demonstrate the flexibility of the system by using a variety of lipids including 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC) and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC).

Soil is the habitat of countless organisms and encompasses an enormous variety of dynamic environmental conditions. While it is evident that a thorough understanding of how organisms interact with the soil environment may have substantial ecological and economical impact, current laboratory-based methods depend on reductionist approaches that are incapable of simulating natural diversity. The application of Lab-on-a-Chip or microfluidic technologies to organismal studies is an emerging field, where the unique benefits afforded by system miniaturisation offer new opportunities for the experimentalist. Indeed, precise spatiotemporal control over the microenvironments of soil organisms in combination with high-resolution imaging has the potential to provide an unprecedented view of biological events at the single-organism or single-cell level, which in turn opens up new avenues for environmental and organismal studies. Herein we review some of the most recent and interesting developments in microfluidic technologies for the study of soil organisms and their interactions with the environment. We discuss how so-called "Soil-on-a-Chip" technology has already contributed significantly to the study of bacteria, nematodes, fungi and plants, as well as inter-organismal interactions, by advancing experimental access and environmental control. Most crucially, we highlight where distinct advantages over traditional approaches exist and where novel biological insights will ensue.

Interactions between fungi and prokaryotes are abundant in many ecological systems. A wide variety of biomolecules regulate such interactions and many of them have found medicinal or biotechnological applications. However, studying a fungal-bacterial system at a cellular level is technically challenging. New microfluidic devices provided a platform for microscopic studies and for long-term, time-lapse experiments. Application of these novel tools revealed insights into the dynamic interactions between the basidiomycete Coprinopsis cinerea and the bacterium Bacillus subtilis. Direct contact was mediated by polar attachment of bacteria to only a subset of fungal hyphae suggesting a differential competence of fungal hyphae and thus differentiation of hyphae within a mycelium. The fungicidal activity of B. subtilis was monitored at a cellular level and showed a novel mode of action on fungal hyphae.

A chiral tris(urea) organogelator gels dmso-water and methanol-water mixtures at low weight percent. The formation of the helical gel fibres is partially inhibited by addition of chloride, which is bound by the gelator, resulting in fully crystalline material characterised by X-ray crystallography.

35Root hairs are tubular protrusions of the root epidermis that significantly enlarge the exploitable soil 36 volume in the rhizosphere. Trichoblasts, the cell type responsible for root hair formation, switch 37 from cell elongation to tip growth through polarization of the growth machinery to a pre-defined root 38 hair initiation domain (RHID) at the plasma membrane. The emergence of this polar domain 39 resembles the establishment of cell polarity in other eukaryotic systems [1][2][3]. Rho-type GTPases 40 of plants (ROPs) are among the first molecular determinants of the RHID [4, 5] and later play a 41 central role in polar growth [6]. Numerous studies have elucidated mechanisms that position the 42 RHID in the cell [7][8][9] or regulate ROP activity [10][11][12][13][14][15][16][17][18]. The molecular players that target ROPs to 43 the RHID and initiate outgrowth, however, have not been identified. We dissected the timing of the 44 growth machinery assembly in polarizing hair cells and found that positioning of molecular players 45 and outgrowth are temporally separate processes that are each controlled by specific ROP guanine 46 nucleotide exchange factor (GEFs). A functional analysis of trichoblast-specific GEFs revealed 47 GEF3 to be required for normal ROP polarization and thus efficient root hair emergence, while 48 GEF4 predominantly regulates subsequent tip growth. Ectopic expression of GEF3 induced the 49 formation of spatially confined, ROP-recruiting domains in other cell types, demonstrating the role 50 of GEF3 to serve as a membrane landmark during cell polarization. 51 3 RESULTS AND DISCUSSION 52 53 Temporal analysis of hair cell polarization reveals phased deployment of the tip-growth 54 machinery 55To dissect the process of breaking cellular symmetry and initiating polar growth in plants, we 56 analyzed the gradual assembly of the tip growth machinery in trichoblasts using specific markers 57 for cytoskeletal rearrangement, plasma membrane specialization, cell wall modification, vesicle 58 trafficking, and Rho-GTPase signaling. Using stable transgenic Arabidopsis lines expressing 59 fluorescently labeled versions of these protein markers, we determined the timing of their 60 polarization at the RHID during trichoblast differentiation. As the Arabidopsis root represents a time-61 axis of development from stem cells to mature cells, we numbered the developmental stages (-7 62 to +3) within a cell file, with the last cell before bulging labeled -1 and the first cell after the onset 63 of bulging labeled +1 ( Figure 1A). We used the integral plasma membrane marker GFP-LTI6B [19] 64 as a reference and calculated the polarity index of fluorescently labeled marker proteins to quantify 65 protein accumulation at the RHID (Figure 1B, C). Our survey unveiled a two-phase assembly of 66 the tip growth machinery: an initiation phase, during which the RHID is positioned and predefined, 67 followed by the tip growth phase. Consistent with previous reports [4, 5], mCitrine-labeled (mCit) 68GTPase ROP2 associ...

Roots grow in highly dynamic and heterogeneous environments. Biological activity as well as uneven nutrient availability or localized stress factors result in diverse microenvironments. Plants adapt their root morphology in response to changing environmental conditions, yet it remains largely unknown to what extent developmental adaptations are based on systemic or cell-autonomous responses. We present the dual-flow-RootChip, a microfluidic platform for asymmetric perfusion of Arabidopsis roots to investigate root-environment interactions under simulated environmental heterogeneity. Applications range from investigating physiology, root hair development and calcium signalling upon selective exposure to environmental stresses to tracing molecular uptake, performing selective drug treatments and localized inoculations with microbes. Using the dual-flow-RootChip, we revealed cell-autonomous adaption of root hair development under asymmetric phosphate (Pi) perfusion, with unexpected repression in root hair growth on the side exposed to low Pi and rapid tip-growth upregulation when Pi concentrations increased. The asymmetric root environment further resulted in an asymmetric gene expression of RSL4, a key transcriptional regulator of root hair growth. Our findings demonstrate that roots possess the capability to locally adapt to heterogeneous conditions in their environment at the physiological and transcriptional levels. Being able to generate asymmetric microenvironments for roots will help further elucidate decision-making processes in root-environment interactions.

Highlights d Polarization and outgrowth of root hairs are temporally separate processes d Distinct RopGEFs are involved in ROP GTPase recruitment and growth regulation d GEF3 is required for ROP2 targeting to the emerging root hair initiation domain d GEF3 expression is sufficient to induce polar domain formation in epidermal cells