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The importance of Antarctic sea ice and Southern Ocean warming has come into the focus of polar research during the last couple of decades. Especially around West Antarctica, where warm water masses approach the continent and where sea ice has declined, the distribution and evolution of sea ice play a critical role in the stability of nearby ice shelves. Organic geochemical analyses of marine seafloor surface sediments from the Antarctic continental margin allow an evaluation of the applicability of biomarker-based sea-ice and ocean temperature reconstructions in these climate-sensitive areas. We analysed highly branched isoprenoids (HBIs), such as the sea-ice proxy IPSO25 and phytoplankton-derived HBI-trienes, as well as phytosterols and isoprenoidal glycerol dialkyl glycerol tetraethers (GDGTs), which are established tools for the assessment of primary productivity and ocean temperatures respectively. The combination of IPSO25 with a phytoplankton marker (i.e. the PIPSO25 index) permits semi-quantitative sea-ice reconstructions and avoids misleading over- or underestimations of sea-ice cover. Comparisons of the PIPSO25-based sea-ice distribution patterns and TEX86L- and RI-OH′-derived ocean temperatures with (1) sea-ice concentrations obtained from satellite observations and (2) instrument measurements of sea surface and subsurface temperatures corroborate the general capability of these proxies to determine oceanic key variables properly. This is further supported by model data. We also highlight specific aspects and limitations that need to be taken into account for the interpretation of such biomarker data and discuss the potential of IPSO25 as an indicator for the former occurrence of platelet ice and/or the export of ice-shelf water.

Science-based forest management requires quantitative estimation of forest attributes traditionally collected via sampled field plots in a forest inventory program. Three-dimensional (3D) remotely sensed data such as Light Detection and Ranging (lidar), are increasingly utilized to supplement and even replace field-based forest inventories. However, lidar remains cost prohibitive for smaller areas and repeat measurements, often limiting its use to single acquisitions of large contiguous areas. Recent advancements in unpiloted aerial systems (UAS), digital aerial photogrammetry (DAP) and high precision global positioning systems (HPGPS) have the potential to provide low-cost time and place flexible 3D data to support forest inventory and monitoring. The primary objective of this study was to assess the ability of low-cost commercial off the shelf UAS DAP and HPGPS to create accurate 3D data and predictions of key forest attributes, as compared to both lidar and field observations, in a wide range of forest conditions in California, USA. A secondary objective was to assess the accuracy of nadir vs. off-nadir UAS DAP, to determine if oblique imagery provides more accurate 3D data and forest attribute predictions. UAS DAP digital terrain models (DTMs) were comparable to lidar DTMS across most sites and nadir vs. off-nadir imagery collection (R2 = 0.74–0.99), although model accuracy using off-nadir imagery was very low in mature Douglas-fir forest (R2 = 0.17) due to high canopy density occluding the ground from the image sensor. Surface and canopy height models were shown to have less agreement to lidar (R2 = 0.17–0.69), with off-nadir imagery surface models at high canopy density sites having the lowest agreement with lidar. UAS DAP models predicted key forest metrics with varying accuracy compared to field data (R2 = 0.53–0.85), and were comparable to predictions made using lidar. Although lidar provided more accurate estimates of forest attributes across a range of forest conditions, this study shows that UAS DAP models, when combined with low-cost HPGPS, can accurately predict key forest attributes across a range of forest types, canopies densities, and structural conditions.

Fusarium keratoplasticum is the Fusarium species most commonly associated with human infections (fusariosis). Antifungal treatment of fusariosis is often hampered by limited treatment options due to resistance towards azole antifungals. The mechanisms of antifungal resistance and sterol biosynthesis in fusaria are poorly understood. Therefore, in this study we assessed the transcriptional response of F. keratoplasticum when exposed to voriconazole. Our results revealed a group of dramatically upregulated ergosterol biosynthesis gene duplicates, most notably erg6A (912-fold), cyp51A (52-fold) and ebp1 (20-fold), which are likely part of an alternative ergosterol biosynthesis salvage pathway. The presence of human cholesterol biosynthesis gene homologs in F. keratoplasticum (ebp1, dhcr7 and dhcr24_1, dhcr24_2 and dhcr24_3) suggests that additional sterol biosynthesis pathways may be induced in fusaria under other growth conditions or during host invasion. Voriconazole also induced the expression of a number of ABC efflux pumps. Further investigations suggested that the highly conserved master regulator of ergosterol biosynthesis, FkSR, and the pleiotropic drug resistance network that induces zinc-cluster transcription factor FkAtrR coordinate the response of FSSC species to azole antifungal exposure. In-depth genome mining also helped clarify the ergosterol biosynthesis pathways of moulds and provided a better understanding of antifungal drug resistance mechanisms in fusaria.

Fusarium infections in humans (fusariosis) and in economically important plants involve species of several Fusarium species complexes. Species of the Fusarium solani species complex (FSSC) are the most frequent cause of human fusariosis. The FSSC comprises more than 60 closely related species that can be separated into three major clades by multi-locus sequence typing (MLST) using translation elongation factor 1-alpha (TEF1-α) and RNA polymerase II (RPB2) DNA sequences. The MLST nomenclature for clade 3 of the FSSC assigns numbers to species types (e.g., FSSC 2) and lowercase letters to identify unique haplotypes. The aim of this study was to analyse the genotypic and phenotypic characteristics of 15 environmental and 15 clinical FSSC isolates from Malaysia. MLST was used for the genotypic characterisation of FSSC isolates from various locations within Malaysia, which was complemented by their morphological characterisation on potato dextrose and carnation leaf agar. MLST identified eight different FSSC species: thirteen Fusarium keratoplasticum (i.e., FSSC 2), six Fusarium suttonianum (FSSC 20), five Fusarium falciforme (FSSC 3+4), two Fusarium cyanescens (FSSC 27), and one each of Fusarium petroliphilum (FSSC 1), Fusarium waltergamsii (FSSC 7), Fusarium sp. (FSSC 12), and Fusarium striatum (FSSC 21). Consistent with previous reports from Malaysia, most (11 of 15) clinical FSSC isolates were F. keratoplasticum and the majority (9 of 15) of environmental isolates were F. suttonianum (5) or F. falciforme (4) strains. The taxonomic relationships of the isolates were resolved phylogenetically. The eight Fusarium species also showed distinct morphological characteristics, but these were less clearly defined and reached across species boundaries. Although TEF1-α and RPB2 sequences were sufficient for the species identification of most FSSC isolates, a more precise MLST scheme needs to be established to reliably assign individual isolates of the species-rich FSSC to their geographically-, epidemiologically-, and host-associated sub-lineages.

The successful biochemical and biophysical characterization of ABC transporters depends heavily on the choice of the heterologous expression system. Over the past two decades, we have developed a yeast membrane protein expression platform that has been used to study many important fungal membrane proteins. The expression host Saccharomyces cerevisiae ADΔΔ is deleted in seven major endogenous ABC transporters and it contains the transcription factor Pdr1-3 with a gain-of-function mutation that enables the constitutive overexpression of heterologous membrane protein genes stably integrated as single copies at the genomic PDR5 locus. The creation of versatile plasmid vectors and the optimization of one-step cloning strategies enables the rapid and accurate cloning, mutagenesis, and expression of heterologous ABC transporters. Here, we describe the development and use of a novel protease-cleavable mGFPHis double tag (i.e., the monomeric yeast enhanced green fluorescent protein yEGFP3 fused to a six-histidine affinity purification tag) that was designed to avoid possible interference of the tag with the protein of interest and to increase the binding efficiency of the His tag to nickel-affinity resins. The fusion of mGFPHis to the membrane protein ORF (open reading frame) enables easy quantification of the protein by inspection of polyacrylamide gels and detection of degradation products retaining the mGFPHis tag. We demonstrate how this feature facilitates detergent screening for membrane protein solubilization. A protocol for the efficient, fast, and reliable isolation of the small-scale plasma membrane preparations of the C-terminally tagged Candida albicans multidrug efflux transporter Cdr1 overexpressed in S. cerevisiae ADΔΔ, is presented. This small-scale plasma membrane isolation protocol generates high-quality plasma membranes within a single working day. The plasma membrane preparations can be used to determine the enzyme activities of Cdr1 and Cdr1 mutant variants.

The SkyVision system was developed by the State of Ohio and the Air Force Research Laboratory (AFRL) to promote the research and development of Unmanned Aerial Systems (UAS) technologies. With the goal and vision to deliver a ground-based detect and avoid (GBDAA) capability, SkyVision leverages Air Traffic Control (ATC) level radar data through the Federal Aviation Administration's (FAA) Standard Terminal Automation Replacement System (STARS). This paper details the research and development of a set of subsystems that integrate into SkyVision to provide the system with UAS flight information, seeking to enhance the system, enabling safe beyond visual line of sight (BVLOS) UAS operations throughout the nation.

The successful biochemical and biophysical characterization of ABC transporters depends heavily on the choice of the heterologous expression system. Over the past two decades, we have developed a yeast membrane protein expression platform that has been used to study many important fungal membrane proteins. The expression host Saccharomyces cerevisiae ADΔΔ is deleted in seven major endogenous ABC transporters and it contains the transcription factor Pdr1-3 with a gain-of-function mutation that enables the constitutive overexpression of heterologous membrane protein genes stably integrated as single copies at the genomic PDR5 locus. The creation of versatile plasmid vectors and the optimization of one-step cloning strategies enables the rapid and accurate cloning, mutagenesis, and expression of heterologous ABC transporters. Here, we describe the development and use of a novel protease-cleavable mGFPHis double tag (i.e., the monomeric yeast enhanced green fluorescent protein yEGFP3 fused to a six-histidine affinity purification tag) that was designed to avoid possible interference of the tag with the protein of interest and to increase the binding efficiency of the His tag to nickel-affinity resins. The fusion of mGFPHis to the membrane protein ORF (open reading frame) enables easy quantification of the protein by inspection of polyacrylamide gels and detection of degradation products retaining the mGFPHis tag. We demonstrate how this feature facilitates detergent screening for membrane protein solubilization. A protocol for the efficient, fast, and reliable isolation of the small-scale plasma membrane preparations of the C-terminally tagged Candida albicans multidrug efflux transporter Cdr1 overexpressed in S. cerevisiae ADΔΔ, is presented. This small-scale plasma membrane isolation protocol generates high-quality plasma membranes within a single working day. The plasma membrane preparations can be used to determine the enzyme activities of Cdr1 and Cdr1 mutant variants.

Fusarium keratoplasticum is arguably the most common Fusarium solani species complex (FSSC) species associated with human infections. Invasive fusariosis is a life-threatening fungal infection that is difficult to treat with conventional azole antifungals. Azole drug resistance is often caused by the increased expression of pleiotropic drug resistance (PDR) ATP-binding cassette (ABC) transporters of the ABCG sub-family. Most investigations of Fusarium ABC transporters associated with azole antifungal drug resistance are limited to plant pathogens. Through the manual curation of the entire ABCG protein family of four FSSC species including the fully annotated genome of the plant pathogen Nectria haematococca we identified PDR transporters ABC1 and ABC2 as the efflux pump candidates most likely to be associated with the innate azole resistance phenotype of Fusarium keratoplasticum. An initial investigation of the transcriptional response of logarithmic phase F. keratoplasticum cells to 16 mg/L voriconazole confirmed strong upregulation (372-fold) of ABC1 while ABC2 mRNA levels were unaffected by voriconazole exposure over a 4 h time-period. Overexpression of F. keratoplasticum ABC1 and ABC2 in the genetically modified Saccharomyces cerevisiae host ADΔΔ caused up to ∼1,024-fold increased resistance to a number of xenobiotics, including azole antifungals. Although ABC1 and ABC2 were only moderately (20% and 10%, respectively) expressed compared to the Candida albicans multidrug efflux pump CDR1, overexpression of F. keratoplasticum ABC1 caused even higher resistance levels to certain xenobiotics (e.g., rhodamine 6G and nigericin) than CDR1. Our investigations suggest an important role for ABC1 orthologues in the innate azole resistance phenotype of FSSC species.