College of Agriculture and Life Sciences (CALS)

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Caging the blob: using a slime mold to teach concepts about barriers that constrain the movement of organisms
Bohland, Cynthia E.; Schmale, David G. III; Ross, Shane D. (University of California Press, 2011-11-01)
Few laboratory exercises are designed to teach biology students about barriers that may constrain the movement of organisms. We describe a unique inquiry-based exercise involving Lego mazes (the barrier) and the plasmodial slime mold, Physarum polycephalum (the organism). During guided inquiry, students construct mazes using Lego brand building blocks and the slime mold is allowed to "navigate" through the maze and "respond" to the barrier. Students then generate and test hypotheses about the movement of the slime mold in response to different barriers in the open-inquiry phase of the investigation.
The cardiolipin-binding peptide elamipretide mitigates fragmentation of cristae networks following cardiac ischemia reperfusion in rats
Allen, Mitchell E.; Pennington, Edward Ross; Perry, Justin B.; Dadoo, Sahil; Makrecka-Kuka, Marina; Dambrova, Maija; Moukdar, Fatiha; Patel, Hetal D.; Han, Xianlin; Kidd, Grahame K.; Benson, Emily K.; Raisch, Tristan B.; Poelzing, Steven; Brown, David A.; Shaikh, Saame Raza (2020-07-17)
Allen and Pennington et al. show that the cardiolipin-binding peptide elamipretide mitigates disease-induced fragmentation of cristae networks following cardiac ischemia reperfusion in rats. This study suggests that elamipretide targets mitochondrial membranes to sustain cristae networks, improving their bioenergetic function. Mitochondrial dysfunction contributes to cardiac pathologies. Barriers to new therapies include an incomplete understanding of underlying molecular culprits and a lack of effective mitochondria-targeted medicines. Here, we test the hypothesis that the cardiolipin-binding peptide elamipretide, a clinical-stage compound under investigation for diseases of mitochondrial dysfunction, mitigates impairments in mitochondrial structure-function observed after rat cardiac ischemia-reperfusion. Respirometry with permeabilized ventricular fibers indicates that ischemia-reperfusion induced decrements in the activity of complexes I, II, and IV are alleviated with elamipretide. Serial block face scanning electron microscopy used to create 3D reconstructions of cristae ultrastructure reveals that disease-induced fragmentation of cristae networks are improved with elamipretide. Mass spectrometry shows elamipretide did not protect against the reduction of cardiolipin concentration after ischemia-reperfusion. Finally, elamipretide improves biophysical properties of biomimetic membranes by aggregating cardiolipin. The data suggest mitochondrial structure-function are interdependent and demonstrate elamipretide targets mitochondrial membranes to sustain cristae networks and improve bioenergetic function.
Characterization of sequentially-staged cancer cells using electrorotation
Trainito, Claudia I.; Sweeney, Daniel C.; Čemazăr, Jaka; Schmelz, Eva M.; Français; Le Pioufle, Bruno; Davalos, Rafael V. (PLOS, 2019-09-19)
The identification and separation of cells from heterogeneous populations is critical to the diagnosis of diseases. Label-free methodologies in particular have been developed to manipulate individual cells using properties such as density and morphology. The electrical properties of malignant cells, including the membrane capacitance and cytoplasmic conductivity, have been demonstrated to be altered compared to non-malignant cells of similar origin. Here, we exploit these changes to characterize individual cells in a sequentially-staged in vitro cancer model using electrorotation (EROT)—the rotation of a cell induced by a rotating electric field. Using a microfabricated device, a dielectrophoretic force to suspend cells while measuring their angular velocity resulting from an EROT force applied at frequencies between 3 kHz to 10 MHz. We experimentally determine the EROT response for cells at three stages of malignancy and analyze the resultant spectra by considering models that include the effect of the cell membrane alone (single-shell model) and the combined effect of the cell membrane and nucleus (double-shell model). We find that the cell membrane is largely responsible for a given cell’s EROT response between 3 kHz and 10 MHz. Our results also indicate that membrane capacitance, membrane conductance, and cytoplasmic conductivity increase with an increasingly malignant phenotype. Our results demonstrate the potential of using electrorotation as a means making of non-invasive measurements to characterize the dielectric properties of cancer cells.
Classical and adaptive control of ex vivo skeletal muscle contractions using Functional Electrical Stimulation (FES)
Cienfuegos, Paola Jaramillo; Shoemaker, Adam; Grange, Robert W.; Abaid, Nicole; Leonessa, Alexander (PLOS, 2017-03-08)
Functional Electrical Stimulation is a promising approach to treat patients by stimulating the peripheral nerves and their corresponding motor neurons using electrical current. This technique helps maintain muscle mass and promote blood flow in the absence of a functioning nervous system. The goal of this work is to control muscle contractions from FES via three different algorithms and assess the most appropriate controller providing effective stimulation of the muscle. An open-loop system and a closed-loop system with three types of model-free feedback controllers were assessed for tracking control of skeletal muscle contractions: a Proportional-Integral (PI) controller, a Model Reference Adaptive Control algorithm, and an Adaptive Augmented PI system. Furthermore, a mathematical model of a muscle-mass-spring system was implemented in simulation to test the open-loop case and closed-loop controllers. These simulations were carried out and then validated through experiments ex vivo. The experiments included muscle contractions following four distinct trajectories: a step, sine, ramp, and square wave. Overall, the closed-loop controllers followed the stimulation trajectories set for all the simulated and tested muscles. When comparing the experimental outcomes of each controller, we concluded that the Adaptive Augmented PI algorithm provided the best closed-loop performance for speed of convergence and disturbance rejection.
Dielectrophoretic differentiation of mouse ovarian surface epithelial cells, macrophages, and fibroblasts using contactless dielectrophoresis
Salmanzadeh, Alireza; Kittur, Harsha; Sano, Michael B.; Roberts, Paul C.; Schmelz, Eva M.; Davalos, Rafael V. (American Institute of Physics, 2012-06-01)
Ovarian cancer is the leading cause of death from gynecological malignancies in women. The primary challenge is the detection of the cancer at an early stage, since this drastically increases the survival rate. In this study we investigated the dielectrophoretic responses of progressive stages of mouse ovarian surface epithelial (MOSE) cells, as well as mouse fibroblast and macrophage cell lines, utilizing contactless dielectrophoresis (cDEP). cDEP is a relatively new cell manipulation technique that has addressed some of the challenges of conventional dielectrophoretic methods. To evaluate our microfluidic device performance, we computationally studied the effects of altering various geometrical parameters, such as the size and arrangement of insulating structures, on dielectrophoretic and drag forces. We found that the trapping voltage of MOSE cells increases as the cells progress from a non-tumorigenic, benign cell to a tumorigenic, malignant phenotype. Additionally, all MOSE cells display unique behavior compared to fibroblasts and macrophages, representing normal and inflammatory cells found in the peritoneal fluid. Based on these findings, we predict that cDEP can be utilized for isolation of ovarian cancer cells from peritoneal fluid as an early cancer detection tool. (C) 2012 American Institute of Physics. [http://dx.doi.org/10.1063/1.3699973] Actual pdf downloaded from NCBI.
Elevated perfusate [Na+] increases contractile dysfunction during ischemia and reperfusion
King, D. Ryan; Padget, Rachel L.; Perry, Justin B.; Hoeker, Gregory S.; Smyth, James W.; Brown, David A.; Poelzing, Steven (2020-10-14)
Recent studies revealed that relatively small changes in perfusate sodium ([Na+](o)) composition significantly affect cardiac electrical conduction and stability in contraction arrested ex vivo Langendorff heart preparations before and during simulated ischemia. Additionally, [Na+](o) modulates cardiomyocyte contractility via a sodium-calcium exchanger (NCX) mediated pathway. It remains unknown, however, whether modest changes to [Na+](o) that promote electrophysiologic stability similarly improve mechanical function during baseline and ischemia-reperfusion conditions. The purpose of this study was to quantify cardiac mechanical function during ischemia-reperfusion with perfusates containing 145 or 155 mM Na+ in Langendorff perfused isolated rat heart preparations. Relative to 145 mM Na+, perfusion with 155 mM [Na+](o) decreased the amplitude of left-ventricular developed pressure (LVDP) at baseline and accelerated the onset of ischemic contracture. Inhibiting NCX with SEA0400 abolished LVDP depression caused by increasing [Na+](o) at baseline and reduced the time to peak ischemic contracture. Ischemia-reperfusion decreased LVDP in all hearts with return of intrinsic activity, and reperfusion with 155 mM [Na+](o) further depressed mechanical function. In summary, elevating [Na+](o) by as little as 10 mM can significantly modulate mechanical function under baseline conditions, as well as during ischemia and reperfusion. Importantly, clinical use of Normal Saline, which contains 155 mM [Na+](o), with cardiac ischemia may require further investigation.
Establishing an immunocompromised porcine model of human cancer for novel therapy development with pancreatic adenocarcinoma and irreversible electroporation
Hendricks-Wenger, Alissa; Aycock, Kenneth N.; Nagai-Singer, Margaret A.; Coutermarsh-Ott, Sheryl; Lorenzo, Melvin F.; Gannon, Jessica; Uh, Kyungjun; Farrell, Kayla; Beitel-White, Natalie; Brock, Rebecca M.; Simon, Alexander; Morrison, Holly A.; Tuohy, Joanne L.; Clark-Deener, Sherrie; Vlaisavljevich, Eli; Davalos, Rafael V.; Lee, Kiho; Allen, Irving C. (Nature Research, 2021-04-07)
New therapies to treat pancreatic cancer are direly needed. However, efficacious interventions lack a strong preclinical model that can recapitulate patients’ anatomy and physiology. Likewise, the availability of human primary malignant tissue for ex vivo studies is limited. These are significant limitations in the biomedical device field. We have developed RAG2/IL2RG deficient pigs using CRISPR/Cas9 as a large animal model with the novel application of cancer xenograft studies of human pancreatic adenocarcinoma. In this proof-of-concept study, these pigs were successfully generated using on-demand genetic modifications in embryos, circumventing the need for breeding and husbandry. Human Panc01 cells injected subcutaneously into the ears of RAG2/IL2RG deficient pigs demonstrated 100% engraftment with growth rates similar to those typically observed in mouse models. Histopathology revealed no immune cell infiltration and tumor morphology was highly consistent with the mouse models. The electrical properties and response to irreversible electroporation of the tumor tissue were found to be similar to excised human pancreatic cancer tumors. The ample tumor tissue produced enabled improved accuracy and modeling of the electrical properties of tumor tissue. Together, this suggests that this model will be useful and capable of bridging the gap of translating therapies from the bench to clinical application.
Exploring Host-Microbiome Interactions using an in Silico Model of Biomimetic Robots and Engineered Living Cells
Heyde, Keith C.; Ruder, Warren C. (Springer Nature, 2015-07-16)
The microbiome's underlying dynamics play an important role in regulating the behavior and health of its host. In order to explore the details of these interactions, we created an in silico model of a living microbiome, engineered with synthetic biology, that interfaces with a biomimetic, robotic host. By analytically modeling and computationally simulating engineered gene networks in these commensal communities, we reproduced complex behaviors in the host. We observed that robot movements depended upon programmed biochemical network dynamics within the microbiome. These results illustrate the model's potential utility as a tool for exploring inter-kingdom ecological relationships. These systems could impact fields ranging from synthetic biology and ecology to biophysics and medicine.
Intercomparison of Small Unmanned Aircraft System (sUAS) Measurements for Atmospheric Science during the LAPSE-RATE Campaign
Barbieri, Lindsay; Kral, Stephan T.; Bailey, Sean C. C.; Frazier, Amy E.; Jacob, Jamey D.; Reuder, Joachim; Brus, David; Chilson, Phillip B.; Crick, Christopher; Detweiler, Carrick; Doddi, Abhiram; Elston, Jack; Foroutan, Hosein; González-Rocha, Javier; Greene, Brian R.; Guzman, Marcelo I.; Houston, Adam L.; Islam, Ashraful; Kemppinen, Osku; Lawrence, Dale; Pillar-Little, Elizabeth A.; Ross, Shane D.; Sama, Michael P.; Schmale, David G. III; Schuyler, Travis J.; Shankar, Ajay; Smith, Suzanne W.; Waugh, Sean; Dixon, Cory; Borenstein, Steve; de Boer, Gijs (MDPI, 2019-05-10)
Small unmanned aircraft systems (sUAS) are rapidly transforming atmospheric research. With the advancement of the development and application of these systems, improving knowledge of best practices for accurate measurement is critical for achieving scientific goals. We present results from an intercomparison of atmospheric measurement data from the Lower Atmospheric Process Studies at Elevation—a Remotely piloted Aircraft Team Experiment (LAPSE-RATE) field campaign. We evaluate a total of 38 individual sUAS with 23 unique sensor and platform configurations using a meteorological tower for reference measurements. We assess precision, bias, and time response of sUAS measurements of temperature, humidity, pressure, wind speed, and wind direction. Most sUAS measurements show broad agreement with the reference, particularly temperature and wind speed, with mean value differences of 1.6 ± 2.6 ∘ C and 0.22 ± 0.59 m/s for all sUAS, respectively. sUAS platform and sensor configurations were found to contribute significantly to measurement accuracy. Sensor configurations, which included proper aspiration and radiation shielding of sensors, were found to provide the most accurate thermodynamic measurements (temperature and relative humidity), whereas sonic anemometers on multirotor platforms provided the most accurate wind measurements (horizontal speed and direction). We contribute both a characterization and assessment of sUAS for measuring atmospheric parameters, and identify important challenges and opportunities for improving scientific measurements with sUAS.
Investigating dielectric properties of different stages of syngeneic murine ovarian cancer cells
Salmanzadeh, Alireza; Sano, Michael B.; Gallo-Villanueva, R. C.; Roberts, Paul C.; Schmelz, Eva M.; Davalos, Rafael V. (American Institute of Physics, 2013-01-01)
In this study, the electrical properties of four different stages of mouse ovarian surface epithelial (MOSE) cells were investigated using contactless dielectrophoresis (cDEP). This study expands the work from our previous report describing for the first time the crossover frequency and cell specific membrane capacitance of different stages of cancer cells that are derived from the same cell line. The specific membrane capacitance increased as the stage of malignancy advanced from 15.39 +/- 1.54 mF m(-2) for a non-malignant benign stage to 26.42 +/- 1.22 mF m(-2) for the most aggressive stage. These differences could be the result of morphological variations due to changes in the cytoskeleton structure, specifically the decrease of the level of actin filaments in the cytoskeleton structure of the transformed MOSE cells. Studying the electrical properties of MOSE cells provides important information as a first step to develop cancer-treatment techniques which could partially reverse the cytoskeleton disorganization of malignant cells to a morphology more similar to that of benign cells. (C) 2013 American Institute of Physics. [http://dx.doi.org/10.1063/1.4788921] Actual pdf downloaded from NCBI.
Lagrangian coherent structures are associated with fluctuations in airborne microbial populations
Tallapragada, Phanindra; Ross, Shane D.; Schmale, David G. III (American Institute of Physics, 2011-09-01)
Many microorganisms are advected in the lower atmosphere from one habitat to another with scales of motion being hundreds to thousands of kilometers. The concentration of these microbes in the lower atmosphere at a single geographic location can show rapid temporal changes. We used autonomous unmanned aerial vehicles equipped with microbe-sampling devices to collect fungi in the genus Fusarium 100 m above ground level at a single sampling location in Blacksburg, Virginia, USA. Some Fusarium species are important plant and animal pathogens, others saprophytes, and still others are producers of dangerous toxins. We correlated punctuated changes in the concentration of Fusarium to the movement of atmospheric transport barriers identified as finite-time Lyapunov exponent-based Lagrangian coherent structures (LCSs). An analysis of the finite-time Lyapunov exponent field for periods surrounding 73 individual flight collections of Fusarium showed a relationship between punctuated changes in concentrations of Fusarium and the passage times of LCSs, particularly repelling LCSs. This work has implications for understanding the atmospheric transport of invasive microbial species into previously unexposed regions and may contribute to information systems for pest management and disease control in the future.
N-(3-oxododecanoyl)-L-homoserine lactone interactions in the breast tumor microenvironment: Implications for breast cancer viability and proliferation in vitro
Balhouse, Brittany N.; Patterson, Logan; Schmelz, Eva M.; Slade, Daniel J.; Verbridge, Scott S. (PLOS, 2017-07-10)
It is well documented that the tumor microenvironment profoundly impacts the etiology and progression of breast cancer, yet the contribution of the resident microbiome within breast tissue remains poorly understood. Tumor microenvironmental conditions, such as hypoxia and dense tumor stroma, predispose progressive phenotypes and therapy resistance, however the role of bacteria in this interplay remains uncharacterized. We hypothesized that the effect of individual bacterial secreted molecules on breast cancer viability and proliferation would be modulated by these tumor-relevant stressors differentially for cells at varying stages of progression. To test this, we incubated human breast adenocarcinoma cells (MDA-MB-231, MCF-DCIS.com) and non-malignant breast epithelial cells (MCF-10A) with N-(3-oxododecanoyl)-L-homoserine lactone (OdDHL), a quorum-sensing molecule from Pseudomonas aeruginosa that regulates bacterial stress responses. This molecule was selected because Pseudomonas was recently characterized as a significant fraction of the breast tissue microbiome and OdDHL is documented to impact mammalian cell viability. After OdDHL treatment, we demonstrated the greatest decrease in viability with the more malignant MDA-MB-231 cells and an intermediate MCF-DCIS.com (ductal carcinoma in situ) response. The responses were also culture condition (i.e. microenvironment) dependent. These results contrast the MCF-10A response, which demonstrated no change in viability in any culture condition. We further determined that the observed trends in breast cancer viability were due to modulation of proliferation for both cell types, as well as the induction of necrosis for MDA-MB-231 cells in all conditions. Our results provide evidence that bacterial quorum-sensing molecules interact with the host tissue environment to modulate breast cancer viability and proliferation, and that the effect of OdDHL is dependent on both cell type as well as microenvironment. Understanding the interactions between bacterial signaling molecules and the host tissue environment will allow for future studies that determine the contribution of bacteria to the onset, progression, and therapy response of breast cancer.
Quantification of zearalenone and α-zearalenol in swine liver and reproductive tissues using GC-MS
Pack, Erica; Stewart, Jacob; Rhoads, Michelle; Knight, James W.; De Vita, Raffaella; Clark-Deener, Sherrie; Schmale, David G. III (Elsevier, 2020-12-01)
The mycotoxin zearalenone (ZEN) is a common contaminant of swine feed which has been related to a wide range of reproductive anomalies in swine, such as pelvic organ prolapse, anestrous, and pseudopregnancy. New information is needed to understand how ZEN and related metabolites accumulate in swine reproductive tissues. We conducted a feeding study to track ZEN and the metabolite α-zearalenol (α-ZEL) in swine liver and reproductive tissues. Thirty pubertal gilts were randomly assigned one of three treatments, with ten pigs in each treatment group: (1) base feed with solvent for 21 days, (2) ZEN-spiked feed for seven days followed by base feed with solvent for 14 days, and (3) ZEN-spiked feed for 21 days. At the end of the trial, liver, anterior vagina, posterior vagina, cervix, uterus, ovaries, and broad ligament were collected from pigs. ZEN was found in the anterior vagina, posterior vagina, cervix, and ovaries, with significantly higher concentrations in the cervix relative to other reproductive tissues. ZEN and α-ZEL were found in liver tissue from pigs in each treatment group. Our results show that ZEN accumulates more in the cervix than other reproductive tissues. The presence of ZEN in reproductive tissues may be indicative of ZEN-related reproductive symptoms. Future work could examine how ZEN concentrations vary in reproductive tissues as a factor of the pigs age, weight, sex, or parity, to establish parameters that make pig more sensitive to ZEN.
Raman chemometric urinalysis (Rametrix) as a screen for bladder cancer
Huttanus, Herbert M.; Vu, Tommy; Guruli, Georgi; Tracey, Andrew; Carswell, William; Said, Neveen; Du, Pang; Parkinson, Bing G.; Orlando, Giuseppe; Robertson, John L.; Senger, Ryan S. (2020-08-21)
Bladder cancer (BCA) is relatively common and potentially recurrent/progressive disease. It is also costly to detect, treat, and control. Definitive diagnosis is made by examination of urine sediment, imaging, direct visualization (cystoscopy), and invasive biopsy of suspect bladder lesions. There are currently no widely-used BCA-specific biomarker urine screening tests for early BCA or for following patients during/after therapy. Urine metabolomic screening for biomarkers is costly and generally unavailable for clinical use. In response, we developed Raman spectroscopy-based chemometric urinalysis (Rametrix (TM)) as a direct liquid urine screening method for detecting complex molecular signatures in urine associated with BCA and other genitourinary tract pathologies. In particular, the Rametrix(TM)screen used principal components (PCs) of urine Raman spectra to build discriminant analysis models that indicate the presence/absence of disease. The number of PCs included was varied, and all models were cross-validated by leave-one-out analysis. In Study 1 reported here, we tested the Rametrix (TM) screen using urine specimens from 56 consented patients from a urology clinic. This proof-of-concept study contained 17 urine specimens with active BCA (BCA-positive), 32 urine specimens from patients with other genitourinary tract pathologies, seven specimens from healthy patients, and the urinalysis control Surine(TM). Using a model built with 22 PCs, BCA was detected with 80.4% accuracy, 82.4% sensitivity, 79.5% specificity, 63.6% positive predictive value (PPV), and 91.2% negative predictive value (NPV). Based on the number of PCs included, we found the Rametrix(TM)screen could be fine-tuned for either high sensitivity or specificity. In other studies reported here, Rametrix(TM)was also able to differentiate between urine specimens from patients with BCA and other genitourinary pathologies and those obtained from patients with end-stage kidney disease (ESKD). While larger studies are needed to improve Rametrix(TM)models and demonstrate clinical relevance, this study demonstrates the ability of the Rametrix(TM)screen to differentiate urine of BCA-positive patients. Molecular signature variances in the urine metabolome of BCA patients included changes in: phosphatidylinositol, nucleic acids, protein (particularly collagen), aromatic amino acids, and carotenoids.
The Rametrix (TM) PRO Toolbox v1.0 for MATLAB (R)
Senger, Ryan S.; Robertson, John L. (2020-01-06)
Background. Existing tools for chemometric analysis of vibrational spectroscopy data have enabled characterization of materials and biologicals by their broad molecular composition. The Rametrix (TM) LITE Toolbox v1.0 for MATLAB (R) is one such tool available publicly. It applies discriminant analysis of principal components (DAPC) to spectral data to classify spectra into user-defined groups. However, additional functionality is needed to better evaluate the predictive capabilities of these models when "unknown" samples are introduced. Here, the Rametrix (TM) PRO Toolbox v1.0 is introduced to provide this capability. Methods. The Rametrix (TM) PRO Toolbox v1.0 was constructed for MATLAB (R) and works with the Rametrix (TM) LITE Toolbox v1.0. It performs leave-one-out analysis of chemometric DAPC models and reports predictive capabilities in terms of accuracy, sensitivity (true-positives), and specificity (true-negatives). Rametrix (TM) PRO is available publicly through GitHub under license agreement at: https://github.com/SengerLab/RametrixPROToolbox. Rametrix (TM) PRO was used to validate Rametrix (TM) LITE models used to detect chronic kidney disease (CKD) in spectra of urine obtained by Raman spectroscopy. The dataset included Raman spectra of urine from 20 healthy individuals and 31 patients undergoing peritoneal dialysis treatment for CKD. Results. The number of spectral principal components (PCs) used in building the DAPC model impacted the model accuracy, sensitivity, and specificity in leave-one-out analyses. For the dataset in this study, using 35 PCs in the DAPC model resulted in 100% accuracy, sensitivity, and specificity in classifying an unknown Raman spectrum of urine as belonging to a CKD patient or a healthy volunteer. Models built with fewer or greater number of PCs showed inferior performance, which demonstrated the value of Rametrix (TM) PRO in evaluating chemometric models constructed with Rametrix (TM) LITE.
Spectral characteristics of urine specimens from healthy human volunteers analyzed using Raman chemometric urinalysis (Rametrix)
Senger, Ryan S.; Kavuru, Varun; Sullivan, Meaghan; Gouldin, Austin; Lundgren, Stephanie; Merrifield, Kristen; Steen, Caitlin; Baker, Emily; Vu, Tommy; Agnor, Ben; Martinez, Gabrielle; Coogan, Hannah; Carswell, William; Karageorge, Lampros; Dev, Devasmita; Du, Pang; Sklar, Allan; Orlando, Giuseppe; Pirkle, James, Jr.; Robertson, John L. (PLOS, 2019-09-27)
Raman chemometric urinalysis (Rametrix™) was used to analyze 235 urine specimens from healthy individuals. The purpose of this study was to establish the “range of normal” for Raman spectra of urine specimens from healthy individuals. Ultimately, spectra falling outside of this range will be correlated with kidney and urinary tract disease. Rametrix™ analysis includes direct comparisons of Raman spectra but also principal component analysis (PCA), discriminant analysis of principal components (DAPC) models, multivariate statistics, and it is available through GitHub as the Rametrix™ LITE Toolbox for MATLAB®. Results showed consistently overlapping Raman spectra of urine specimens with significantly larger variances in Raman shifts, found by PCA, corresponding to urea, creatinine, and glucose concentrations. A 2-way ANOVA test found that age of the urine specimen donor was statistically significant (p < 0.001) and donor sex (female or male identification) was less so (p = 0.0526). With DAPC models and blind leave-one-out build/test routines using the Rametrix™ PRO Toolbox (also available through GitHub), an accuracy of 71% (sensitivity = 72%; specificity = 70%) was obtained when predicting whether a urine specimen from a healthy unknown individual was from a female or male donor. Finally, from female and male donors (n = 4) who contributed first morning void urine specimens each day for 30 days, the co-occurrence of menstruation was found statistically insignificant to Rametrix™ results (p = 0.695). In addition, Rametrix™ PRO was able to link urine specimens with the individual donor with an average of 78% accuracy. Taken together, this study established the range of Raman spectra that could be expected when obtaining urine specimens from healthy individuals and analyzed by Rametrix™ and provides the methodology for linking results with donor characteristics.
Vortex-induced dispersal of a plant pathogen by raindrop impact
Kim, Seungho; Park, Hyunggon; Gruszewski, Hope A.; Schmale, David G. III; Jung, Sunghwan (NAS, 2019)
Raindrop impact on infected plants can disperse micron-sized propagules of plant pathogens (e.g., spores of fungi). Little is known about the mechanism of how plant pathogens are liberated and transported due to raindrop impact.We used high-speed photography to observe thousands of dry-dispersed spores of the rust fungus Puccinia triticina being liberated from infected wheat plants following the impact of a single raindrop.We revealed that an air vortex ring was formed during the raindrop impact and carried the dry-dispersed spores away from the surface of the host plant. The maximum height and travel distance of the airborne spores increased with the aid of the air vortex. This unique mechanism of vortex-induced dispersal dynamics was characterized to predict trajectories of spores. Finally, we found that the spores transported by the air vortex can reach beyond the laminar boundary layer of leaves, which would enable the long-distance transport of plant pathogens through the atmosphere.
Wind-driven spume droplet production and the transport of Pseudomonas syringae from aquatic environments
Pietsch, Renee B.; Grothe, Hinrich; Hanlon, Regina; Powers, Craig W.; Jung, Sunghwan; Ross, Shane D.; Schmale, David G. III (PeerJ, 2018-09-26)
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 bacterium Pseudomonas syringae were collected at rates of 283 cells m−2 s−1 at 5 cm above the water surface, and at 14 cells m−2 s−1 at 10 cm above the water surface. At a wind speed of about 4.0 m/s, aqueous suspensions of P. syringae were collected at rates of 509 cells m−2 s−1 at 5 cm above the water surface, and at 81 cells m−2 s−1 at 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−1 of water surface were estimated to leave the water in potentially suspended droplets (diameters <100 µm). Understanding the sources and mechanisms for bacteria to aerosolize from freshwater aquatic sources may aid in designing management strategies for pathogenic bacteria, and could shed light on how bacteria are involved in mesoscale atmospheric processes.

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