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New information is needed about the potential sources and pathways of trans-Atlantic dust plumes. Such knowledge has important implications for the long-distance transport and survivability of microorganisms. Forward trajectories of trans-Atlantic dust plumes were studied over a 14-year period, between 2008 and 2021 (n =>500,000 trajectories). Two major dust transport patterns emerged from these analyses. First, summer trajectories (June – August) that arrive in the southeastern regions of the United States and the Caribbean basin and travel above the marine boundary layer at an average altitude of 1,600 m. Second, winter trajectories (December – February) that arrive in the Amazon basin and travel within the boundary layer at an average altitude of 660 m. Ambient meteorological conditions such as solar radiation and relative humidity along dust trajectories suggest a more suitable condition for the survivability of microorganisms reaching the Amazon during the winter with a lower mean solar radiation flux of 294 W m-2 and mean relative humidity levels at around 61% as compared to averages of 370 W m-2 solar radiation and 45% relative humidity for summer trajectories intruding the Caribbean basin. Nevertheless, 14% of winter trajectories (4,664 out of 32,352) reaching the Amazon basin face intense precipitation of higher than 30 mm and get potentially removed as compared to 8% of trajectories (2,540 out of 31,826) intruding the Caribbean basin during the summer. Collectively, our results have important implications for the survivability of microorganisms in trans-Atlantic dust plumes and their potential for major incursion events at receptor regions.

Little is known about the transport and fate of aerosolized particles associated with harmful algal blooms (HABs). An Airborne DROne Particle-monitoring System (AirDROPS) was developed and used to monitor, collect, and characterize airborne particles over two HABs in Grand Lake St Marys (GLSM) and Lake Erie (LE), Ohio USA in August 2019. The AirDROPS consisted of an impinging device (ID) and an optical particle counter (OPC) mounted on a large commercial quadcopter (DJI Inspire 2). The sensor package was mounted above the airframe to limit the effects of propeller downwash that can corrupt measurements taken below the drone. Nineteen flights were conducted 10 m above water level (AWL) at GLSM, and five flights were conducted 10 m AWL at LE. The sampling height was chosen to minimize the effects of propwash on aerosolization from the lake surface. One intercomparison flight was conducted at GLSM over land adjacent to a sonic anemometer mounted on the top of a flagpole 15 m above ground level (AGL). Particle counts generally decreased from morning to afternoon flights, ranging from >4000 in the morning to 4000 below 4 m s−1 to

New tools and technology are needed to track hazardous agents such as oil and red tides in our oceans. Rhodamine dye (a surrogate hazardous agent) was released into the Atlantic ocean in August 2018, and experiments were conducted to track the movement of the dye near the water surface within three hours following the release. A DrOne Water Sampling SystEm (DOWSE), consisting of a 3D-printed sampling device tethered to a drone, was used to collect 26 water samples at different locations around the dye plume. Rhodamine concentrations were measured from the drone water samples using a fluorometer and ranged from 1 to 93 ppb. Dye images were taken during the drone-sampling of surface water containing dye and at about 10 m above the sampling point. These images were post-processed to estimate dye concentrations across the sampling domain. A comparison of calibrated heat maps showed that the altitude images yielded dye distributions that were qualitatively similar to those from images taken near the ocean surface. Moreover, the association between red ratios and dye concentrations yielded trendlines explaining up to 67% of the variation. Drones may be used to detect, track and assist in mitigating hazardous agents in the future. Published version

Heterogeneous ice nucleation plays an important role in many environmental processes such as ice cloud formation, freezing of water bodies or biological freeze protection in the cryosphere. New information is needed about the seasonal availability, nature, and activity of ice nucleating particles (INPs) in alpine environments. These INPs trigger the phase transition from liquid water to solid ice at elevated subzero temperatures. We collected water samples from a series of alpine rivers and lakes (two valleys and their rivers, an artificial pond, and a natural lake system) in Obergurgl, Austria in June 2016, July 2016, November 2016, and May 2017. Each alpine river and lake was sampled multiple times across different seasons, depending on site access during different times of the year. Water samples were filtered through a 0.22 μm membrane filter to separate microbial INPs from the water, and both fractions were analyzed for ice nucleation activity (INA) by an emulsion freezing method. Microorganisms were cultured from the filters, and the cultures then analyzed for INA. Portions of the filtered samples were concentrated by lyophilization to observe potential enhancement of INA. Two sediment samples were taken as reference points for inorganic INPs. Sub-micron INPs were observed in all of the alpine water sources studied, and a seasonal shift to a higher fraction of microbial ice nucleators cultured on selective media was observed during the winter collections. Particles larger than 0.22 μm showed INA, and microbes were cultured from this fraction. Results from 60 samples gave evidence of a seasonal change in INA, presence of submicrometer INPs, and show the abundance of culturable microorganisms, with late spring and early summer showing the most active biological INPs. With additional future research on this topic ski resorts could make use of such knowledge of geographical and seasonal trends of microbial INPs in freshwater habitats in order to improve the production of artificial snow. Published version

Phytophthora sojae is an oomycete that causes stem and root rot disease in soybean. P. sojae delivers many RxLR effector proteins, including Avr1b, into host cells to promote infection. We show here that Avr1b interacts with the soybean U-box protein, GmPUB1-1, in yeast two-hybrid, pull down, and bimolecular fluorescence complementation (BIFC) assays. GmPUB1-1, and a homeologous copy GmPUB1-2, are induced by infection and encode 403 amino acid proteins with U-Box domains at their N-termini. Non-synonymous mutations in the Avr1b C-terminus that abolish suppression of cell death also abolished the interaction of Avr1b with GmPUB1-1, while deletion of the GmPUB1-1 C-terminus, but not the U box, abolished the interaction. BIFC experiments suggested that the GmPUB1-1-Avr1b complex is targeted to the nucleus. In vitro ubiquitination assays demonstrated that GmPUB1-1 possesses E3 ligase activity. Silencing of the GmPUB1 genes in soybean cotyledons resulted in loss of recognition of Avr1b by gene products encoded by Rps1-b and Rps1-k. The recognition of Avr1k (which did not interact with GmPUB1-1) by Rps1-k plants was not, however, affected following GmPUB1-1 silencing. Furthermore, over-expression of GmPUB1-1 in particle bombardment experiments triggered cell death suggesting that GmPUB1 may be a positive regulator of effector-triggered immunity. In a yeast two-hybrid system, GmPUB1-1 also interacted with a number of other RxLR effectors including Avr1d, while Avr1b and Avr1d interacted with a number of other infection-induced GmPUB proteins, suggesting that the pathogen uses a multiplex of interactions of RxLR effectors with GmPUB proteins to modulate host immunity.

The adverse economic impacts of harmful algal blooms can be mitigated via tailored forecasting methods. Adequate provision of these services requires knowledge of the losses avoided, or, in other words, the economic benefits they generate. The latter can be difficult to measure for broader population segments, especially if forecasting services or features do not yet exist. We illustrate how Stated Preference tools and Choice Experiments are well-suited for this case. Using as example forecasts of respiratory irritation levels associated with airborne toxins caused by Florida red tide, we show that 24-hour predictions of spatially and temporally refined air quality conditions are valued highly by the underlying population. This reflects the numerous channels and magnitude of red tide impacts on locals' life and activities, which are also highlighted by our study. Our approach is broadly applicable to any type of air quality impediment with risk of human exposure. Submitted version

Ocean hazardous spills and search and rescue incidents are becoming more prevalent as maritime activities increase across all sectors of society. However, emergency response time remains a factor due to the lack of information available to accurately forecast the location of small objects. Existing drifting characterization techniques are limited to objects whose drifting properties are not affected by on-board wind and surface current sensors. To address this challenge, we study the application of multirotor small unmanned aerial systems (sUAS), and embedded navigation technology, for on-demand wind velocity and surface flow measurements to characterize drifting properties of small objects. An off-the-shelf quadrotor sUAS was used to measure the speed and direction of the wind field at 10 m above surface level near a drifting object. We also leveraged sUAS-grade attitude and heading reference systems and GPS antennas to build water-proof tracking modules that recorded the position and orientation, as well of translational and rotational velocities, of objects drifting in water. The quadrotor and water-proof tracking modules were deployed during field experiments conducted in Claytor Lake, Virginia and the Atlantic Ocean south of Martha’s Vineyard, Massachusetts to characterize the leeway parameters of manikins that simulated a person in water. Leeway parameters were found to be an order of magnitude within previous estimates that were derived using conventional wind and surface current observations. We also determined that multirotor sUAS and water-proof tracking modules can provide accurate and high-resolution ambient information that is critical to understand how changes in orientation affect the downwind displacement and jibing characteristics of small objects floating in water. These findings support further development and application of multirotor sUAS technology for leeway characterization and understanding the effect of an object’s downwind-relative orientation on its drifting characteristics.

Natural aquatic environments such as oceans, lakes, and rivers are home to a tremendous diversity of microorganisms. Some may cross the air-water interface within droplets and become airborne, with the potential to impact the Earth’s radiation budget, precipitation processes, and spread of disease. Larger droplets are likely to return to the water or adjacent land, but smaller droplets may be suspended in the atmosphere for transport over long distances. Here, we report on a series of controlled laboratory experiments to quantify wind-driven droplet production from a freshwater source for low wind speeds. The rate of droplet production increased quadratically with wind speed above a critical value (10-m equivalent 5.7 m/s) where droplet production initiated. Droplet diameter and ejection speeds were fit by a gamma distribution. The droplet mass flux and momentum flux increased with wind speed. Two mechanisms of droplet production, bubble bursting and fragmentation, yielded different distributions for diameter, speed, and angle. At a wind speed of about 3.5 m/s, aqueous suspensions of the ice-nucleating bacteriumPseudomonas syringaewere collected at rates of 283 cells m−2 s−1at 5 cm above the water surface, and at 14 cells m−2 s−1at 10 cm above the water surface. At a wind speed of about 4.0 m/s, aqueous suspensions ofP. syringaewere collected at rates of 509 cells m−2 s−1at 5 cm above the water surface, and at 81 cells m−2 s−1at 10 cm above the water surface. The potential for microbial flux into the atmosphere from aquatic environments was calculated using known concentrations of bacteria in natural freshwater systems. Up to 3.1 × 104 cells m−2 s−1of water surface were estimated to leave the water in potentially suspended droplets (diameters

Rain and snow collected at ground level have been found to contain biological ice nucleators. These ice nucleators have been proposed to have originated in clouds, where they may have participated in the formation of precipitation via ice phase nucleation. We conducted a series of field experiments to test the hypothesis that at least some of the microbial ice nucleators (prokaryotes and eukaryotes) present in rain may not originate in clouds but instead be scavenged from the lower atmosphere by rainfall. Thirty-three simulated rain events were conducted over four months off the side of the Smart Road Bridge in Blacksburg, VA, USA. In each event, sterile water was dispensed over the side of the bridge and recovered in sterile containers in an open fallow agricultural field below (a distance of ∼55 m). Microbes scavenged from the simulated rain events were cultured and their ice nucleation activity was examined. Putative microbial ice nucleators were cultured from 94% (31/33) of the simulated rain events, and represented 1.5% (121/8331) of the total colonies assayed. Putative ice nucleators were subjected to additional droplet freezing assays, and those confirmed through these repeated assays represented 0.4% (34/8331) of the total. Mean CFUs scavenged by simulated rain ranged from 2 to 267 CFUs/mL. Scavenged ice nucleators belong to a number of taxa including the bacterial genera Pseudomonas , Pantoea , and Xanthomonas , and the fungal genera Fusarium , Humicola , and Mortierella . An ice-nucleating strain of the fungal genus Penicillium was also recovered from a volumetric air sampler at the study site. This work expands our knowledge of the scavenging properties of rainfall, and suggests that at least some ice nucleators in natural precipitation events may have been scrubbed from the atmosphere during rainfall, and thus are not likely to be involved in precipitation.

Soybean suspension culture cells were subjected to mutagenesis with ethyl methane sulfonate and cells resistant to the ATP synthase inhibitor oligomycin were recovered. Suspension cultures showed high levels of resistance over a period of 17 months after mutagenesis. The level of resistance to oligomycin decreased during prolonged growth in the absence of the selection agent. Although mitochondrial inheritance of the resistance phenotype was not tested, this observation is consistent with segregation of resistant and sensitive mitochondria in progeny cells. Cross resistance to venturicidin was also observed which suggested the possible involvement of the mitochondrial ATP synthase subunit 9 gene, however, sequences of ATP synthase subunit 9 cDNA clones from resistant cells were identical to the wild type sequence. In addition, Southern blot analyses of DNA from wild type and resistant cultures did not identify rearrangements. These results indicated that resistance to oligomycin could not be directly attributed to a mutation in the primary gene sequence, an alteration in the RNA editing pattern of transcripts, or gross rearrangements in the region containing and directly adjacent to the ATP synthase subunit 9 gene in the mitochondrial DNA from resistant cultures.

Modern soybean [Glycine max (L.) Merr.] cultivars are derived from a limited number of plant introductions, which restricts germplasm diversity at both the nuclear and cytoplasmic levels. This study was conducted to use molecular techniques to assess the extent of cytoplasmic diversity available to soybean breeders. We have surveyed a collection of old domestic soybean varieties with a cloned 2.3 kilobase (kb) HindIII mitochondrial DNA fragment from the cultivar Williams 82 and tbe mitochondrial ATP synthase subunit 6 (atp6) gene to determine sources of cytoplasmic diversity present within this collection. This strategy revealed unique mitochondrial DNA (mtDNA) polymorphisms and polymorphism combinations. In several cases, the mtDNA polymorphisms provided a different classification than was predicted from recorded pedigrees. Variability was also detected among cultivars assigned the name Peking that were introduced independently into the USA soybean collection. The results of this survey coupled with previous studies can be used to define additional cytoplasmic groups in soybean.

Oomycete plant pathogens such as Phytophthora species and downy mildews cause destructive diseases in an enormous variety of crop plant species as well as forests and native ecosystems. These pathogens are most closely related to algae in the kingdom Stramenopiles, and hence have evolved plant pathogenicity independently of other plant pathogens such as fungi. We have used bioinformatic analysis of genome sequences and EST collections, together with functional genomics to identify plant and pathogen genes that may be key players in the interaction between the soybean pathogen Phytophthora sojae and its host. In P. sojae, we have identified many rapidly diversifying gene families that encode potential pathogenicity factors including protein toxins, and a class of proteins (avirulence or effector proteins) that appear to have the ability to penetrate plant cells. Transcriptomic analysis of quantitative or multigenic resistance against P. sojae in soybean has revealed that there are widespread adjustments in host gene expression in response to infection, and that some responses are unique to particular resistant cultivars. These observations lay the foundation for dissecting the interplay between pathogen and host genes during infection at a whole-genome level.

Gene synthesis attempts to assemble user-defined DNA sequences with base-level precision. Verifying the sequences of construction intermediates and the final product of a gene synthesis project is a critical part of the workflow, yet one that has received the least attention. Sequence validation is equally important for other kinds of curated clone collections. Ensuring that the physical sequence of a clone matches its published sequence is a common quality control step performed at least once over the course of a research project. GenoREAD is a web-based application that breaks the sequence verification process into two steps: the assembly of sequencing reads and the alignment of the resulting contig with a reference sequence. GenoREAD can determine if a clone matches its reference sequence. Its sophisticated reporting features help identify and troubleshoot problems that arise during the sequence verification process. GenoREAD has been experimentally validated on thousands of gene-sized constructs from an ORFeome project, and on longer sequences including whole plasmids and synthetic chromosomes. Comparing GenoREAD results with those from manual analysis of the sequencing data demonstrates that GenoREAD tends to be conservative in its diagnostic. GenoREAD is available at www.genoread.org. National Science Foundation (NSF) [EF-0850100, DBI-1241328]; Agriculture and Food Research Initiative of the National Institute of Food and Agriculture of the USDA [2010-65110-20764]; NSF Award [DBI-1241328]; USDA [2010-65110-20764] National Science Foundation (NSF) [EF-0850100, DBI-1241328 to J.P.]; Agriculture and Food Research Initiative of the National Institute of Food and Agriculture of the USDA [2010-65110-20764 to B. M. T.]. Funding for open access charge: NSF Award [DBI-1241328]; USDA [2010-65110-20764].

Background High throughput methods, such as high density oligonucleotide microarray measurements of mRNA levels, are popular and critical to genome scale analysis and systems biology. However understanding the results of these analyses and in particular understanding the very wide range of levels of transcriptional changes observed is still a significant challenge. Many researchers still use an arbitrary cut off such as two-fold in order to identify changes that may be biologically significant. We have used a very large-scale microarray experiment involving 72 biological replicates to analyze the response of soybean plants to infection by the pathogen Phytophthora sojae and to analyze transcriptional modulation as a result of genotypic variation. Results With the unprecedented level of statistical sensitivity provided by the high degree of replication, we show unambiguously that almost the entire plant genome (97 to 99% of all detectable genes) undergoes transcriptional modulation in response to infection and genetic variation. The majority of the transcriptional differences are less than two-fold in magnitude. We show that low amplitude modulation of gene expression (less than two-fold changes) is highly statistically significant and consistent across biological replicates, even for modulations of less than 20%. Our results are consistent through two different normalization methods and two different statistical analysis procedures. Conclusion Our findings demonstrate that the entire plant genome undergoes transcriptional modulation in response to infection and genetic variation. The pervasive low-magnitude remodeling of the transcriptome may be an integral component of physiological adaptation in soybean, and in all eukaryotes.

SummaryPathogens of plants and animals produce effector proteins that are transferred into the cytoplasm of host cells to suppress host defenses. One type of plant pathogens, oomycetes, produces effector proteins with N-terminal RXLR and dEER motifs that enable entry into host cells. We show here that effectors of another pathogen type, fungi, contain functional variants of the RXLR motif, and that the oomycete and fungal RXLR motifs enable binding to the phospholipid, phosphatidylinositol-3-phosphate (PI3P). We find that PI3P is abundant on the outer surface of plant cell plasma membranes and, furthermore, on some animal cells. All effectors could also enter human cells, suggesting that PI3P-mediated effector entry may be very widespread in plant, animal and human pathogenesis. Entry into both plant and animal cells involves lipid raft-mediated endocytosis. Blocking PI3P binding inhibited effector entry, suggesting new therapeutic avenues.