Open Access Publications

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Open access publications by University of Delaware faculty, staff, postdocs, and graduate students at the Delaware Biotechnology Institute.

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Bridging the Talent Gap: Connecting Talent to Bioscience Careers
(Delaware Journal of Public Health, 2023-11) Lakofsky, Katherine
There is an urgent need to engage, educate, and train a skilled workforce for Delaware’s growing life science sector. A sizeable number of these jobs can be obtained with a high school diploma or GED, coupled with an industry informed short-term training program. Unfortunately, this is not widely known, and many disadvantaged populations do not have access to the necessary training. Through a partnership between the Delaware Bioscience Association and the Delaware Biotechnology Institute at the University of Delaware, efforts are currently underway to develop a pilot training program, specifically focusing on the skills needed for biomanufacturing and basic laboratory operations. Additionally, the program will devote significant resources to the identification and recruitment of participants with an emphasis on engaging historically underrepresented populations, as well as removing barriers to accessing the training. The goal is to connect talent to available careers in the industry, providing participants with increased economic mobility and financial stability.
Phylogenetic Analysis to Detect COVID Superspreaders
(Microbiology Research Journal International, 2023-10-12) Jungck, John R.; Ko, Hajae
Aims: Detection of superspreading events by phylogenetic analysis of nucleotide sequences from a population of individuals collected from a narrow time interval. Study Design: Retrieve nucleic acid sequences, construct multiple sequence alignments, and build phylogenetic networks to determine sources of infection. Place and Duration of Study: This study was performed at the Delaware Biotechnology Institute of the University of Delaware over the period: June-August, 2022. The data used were from the GIS AID database. Methodology: Sequences for analysis were sampled from the GISAID initiative’s open-access SARS-CoV-2 genome database. We selected high-quality nucleotide sequences submitted by Delaware labs between March 18 and April 14, 2021, an important period of 4 weeks which saw the Alpha variant spread rapidly in the Delaware population. Results: Four sources accounted for 215 of the 401 sequences. In other words, 54% of all cases were rooted in just five sources. Conclusion: Thus, superspreading seems to have a major impact on the proportion of individuals in a population affected with COVID.
Simulated microgravity facilitates stomatal ingression by Salmonella in lettuce and suppresses a biocontrol agent
(Scientific Reports, 2024-01-09) Totsline, Noah; Kniel, Kalmia E.; Sabagyanam, Chandran; Bais, Harsh P.
As human spaceflight increases in duration, cultivation of crops in spaceflight is crucial to protecting human health under microgravity and elevated oxidative stress. Foodborne pathogens (e.g., Salmonella enterica) carried by leafy green vegetables are a significant cause of human disease. Our previous work showed that Salmonella enterica serovar Typhimurium suppresses defensive closure of foliar stomata in lettuce (Lactuca sativa L.) to ingress interior tissues of leaves. While there are no reported occurrences of foodborne disease in spaceflight to date, known foodborne pathogens persist aboard the International Space Station and space-grown lettuce has been colonized by a diverse microbiome including bacterial genera known to contain human pathogens. Interactions between leafy green vegetables and human bacterial pathogens under microgravity conditions present in spaceflight are unknown. Additionally, stomatal dynamics under microgravity conditions need further elucidation. Here, we employ a slow-rotating 2-D clinostat to simulate microgravity upon in-vitro lettuce plants following a foliar inoculation with S. enterica Typhimurium and use confocal microscopy to measure stomatal width in fixed leaf tissue. Our results reveal significant differences in average stomatal aperture width between an unrotated vertical control, plants rotated at 2 revolutions per minute (2 RPM), and 4 RPM, with and without the presence of S. typhimurium. Interestingly, we found stomatal aperture width in the presence of S. typhimurium to be increased under rotation as compared to unrotated inoculated plants. Using confocal Z-stacking, we observed greater average depth of stomatal ingression by S. typhimurium in lettuce under rotation at 4 RPM compared to unrotated and inoculated plants, along with greater in planta populations of S. typhimurium in lettuce rotated at 4 RPM using serial dilution plating of homogenized surface sterilized leaves. Given these findings, we tested the ability of the plant growth-promoting rhizobacteria (PGPR) Bacillus subtilis strain UD1022 to transiently restrict stomatal apertures of lettuce both alone and co-inoculated with S. typhimurium under rotated and unrotated conditions as a means of potentially reducing stomatal ingression by S. typhimurium under simulated microgravity. Surprisingly, rotation at 4 RPM strongly inhibited the ability of UD1022 alone to restrict stomatal apertures and attenuated its efficacy as a biocontrol following co-inoculation with S. typhimurium. Our results highlight potential spaceflight food safety issues unique to production of crops in microgravity conditions and suggest microgravity may dramatically reduce the ability of PGPRs to restrict stomatal apertures.
Matrix Degradability Contributes to the Development of Salivary Gland Progenitor Cells with Secretory Functions
(ACS Applied Materials and Interfaces, 2023-07-12) Metkari, Apoorva S.; Fowler, Eric W.; Witt, Robert L.; Jia, Xinqiao
Synthetic matrices that are cytocompatible, cell adhesive, and cell responsive are needed for the engineering of implantable, secretory salivary gland constructs to treat radiation induced xerostomia or dry mouth. Here, taking advantage of the bioorthogonality of the Michael-type addition reaction, hydrogels with comparable stiffness but varying degrees of degradability (100% degradable, 100DEG; 50% degradable, 50DEG; and nondegradable, 0DEG) by cell-secreted matrix metalloproteases (MMPs) were synthesized using thiolated HA (HA-SH), maleimide (MI)-conjugated integrin-binding peptide (RGD-MI), and MI-functionalized peptide cross-linkers that are protease degradable (GIW-bisMI) or nondegradable (GIQ-bisMI). Organized multicellular structures developed readily in all hydrogels from dispersed primary human salivary gland stem cells (hS/PCs). As the matrix became progressively degradable, cells proliferated more readily, and the multicellular structures became larger, less spherical, and more lobular. Immunocytochemical analysis showed positive staining for stem/progenitor cell markers CD44 and keratin 5 (K5) in all three types of cultures and positive staining for the acinar marker α-amylase under 50DEG and 100DEG conditions. Quantitatively at the mRNA level, the expression levels of key stem/progenitor markers KIT, KRT5, and ETV4/5 were significantly increased in the degradable gels as compared to the nondegradable counterparts. Western blot analyses revealed that imparting matrix degradation led to >3.8-fold increase in KIT expression by day 15. The MMP-degradable hydrogels also promoted the development of a secretary phenotype, as evidenced by the upregulation of acinar markers α-amylase (AMY), aquaporin-5 (AQP5), and sodium-potassium chloride cotransporter 1 (SLC12A2). Collectively, we show that cell-mediated matrix remodeling is necessary for the development of regenerative pro-acinar progenitor cells from hS/PCs.
RNA degradome analysis reveals DNE1 endoribonuclease is required for the turnover of diverse mRNA substrates in Arabidopsis
(The Plant Cell, 2023-04-20) Nagarajan, Vinay K.; Stuart, Catherine J.; DiBattista, Anna T.; Accerbi, Monica; Caplan, Jeffrey L.; Green, Pamela J.
In plants, cytoplasmic mRNA decay is critical for posttranscriptionally controlling gene expression and for maintaining cellular RNA homeostasis. Arabidopsis DCP1-ASSOCIATED NYN ENDORIBONUCLEASE 1 (DNE1) is a cytoplasmic mRNA decay factor that interacts with proteins involved in mRNA decapping and nonsense-mediated mRNA decay (NMD). There is limited information on the functional role of DNE1 in RNA turnover, and the identities of its endogenous targets are unknown. In this study, we utilized RNA degradome approaches to globally investigate DNE1 substrates. Monophosphorylated 5′ ends, produced by DNE1, should accumulate in mutants lacking the cytoplasmic exoribonuclease XRN4, but be absent from DNE1 and XRN4 double mutants. In seedlings, we identified over 200 such transcripts, most of which reflect cleavage within coding regions. While most DNE1 targets were NMD-insensitive, some were upstream ORF (uORF)-containing and NMD-sensitive transcripts, indicating that this endoribonuclease is required for turnover of a diverse set of mRNAs. Transgenic plants expressing DNE1 cDNA with an active-site mutation in the endoribonuclease domain abolished the in planta cleavage of transcripts, demonstrating that DNE1 endoribonuclease activity is required for cleavage. Our work provides key insights into the identity of DNE1 substrates and enhances our understanding of DNE1-mediated mRNA decay.
Limiting Mrs2-dependent mitochondrial Mg2+ uptake induces metabolic programming in prolonged dietary stress
(Cell Reports, 2023-03-28) Madaris, Travis R.; Venkatesan, Manigandan; Maity, Soumya; Stein, Miriam C.; Vishnu, Neelanjan; Venkateswaran, Mridula K.; Davis, James G.; Ramachandran, Karthik; Uthayabalan, Sukanthathulse; Allen, Cristel; Osidele, Ayodeji; Stanley, Kristen; Bigham, Nicholas P.; Bakewell, Terry M.; Narkunan, Melanie; Le, Amy; Karanam, Varsha; Li, Kang; Mhapankar, Aum; Norton, Luke; Ross, Jean; Aslam, M. Imran; Reeves, W. Brian; Singh, Brij B.; Caplan, Jeffrey; Wilson, Justin J.; Stathopulos, Peter B.; Baur, Joseph A.; Madesh, Muniswamy
Highlights: • Mitochondrial Mg2+ channel Mrs2 rheostats MCU Ca2+ signals to maintain bioenergetic circuit • DNL precursor and cellular Mg2+ chelator citrate curbs HIF1α signal and oxidative metabolism • Lowering mMg2+ mitigates prolonged dietary-stress-induced obesity and metabolic syndrome • Mrs2 channel blocker CPACC reduces lipid accumulation and promotes browning and weight loss Summary The most abundant cellular divalent cations, Mg2+ (mM) and Ca2+ (nM-μM), antagonistically regulate divergent metabolic pathways with several orders of magnitude affinity preference, but the physiological significance of this competition remains elusive. In mice consuming a Western diet, genetic ablation of the mitochondrial Mg2+ channel Mrs2 prevents weight gain, enhances mitochondrial activity, decreases fat accumulation in the liver, and causes prominent browning of white adipose. Mrs2 deficiency restrains citrate efflux from the mitochondria, making it unavailable to support de novo lipogenesis. As citrate is an endogenous Mg2+ chelator, this may represent an adaptive response to a perceived deficit of the cation. Transcriptional profiling of liver and white adipose reveals higher expression of genes involved in glycolysis, β-oxidation, thermogenesis, and HIF-1α-targets, in Mrs2−/− mice that are further enhanced under Western-diet-associated metabolic stress. Thus, lowering mMg2+ promotes metabolism and dampens diet-induced obesity and metabolic syndrome. Graphical abstract Available at: https://doi.org/10.1016/j.celrep.2023.112155
Costs of parthenogenesis on growth and longevity in ex situ zebra sharks Stegostoma tigrinum
(Endangered Species Research, 2023-02-16) Adams, Lance; Lyons, Kady; Monday, Janet; Larkin, Elizabeth; Wyffels, Jennifer
The zebra shark Stegostoma tigrinum, a popular aquarium fish, is an endangered species that is known to readily reproduce both sexually and through facultative parthenogenesis while in human care. Artificial insemination trials that took place between 2011 and 2013 resulted in the hatching of 2 sexually produced (herein heterozygotes) and 10 parthenogenetic sharks that allowed for a retrospective comparison of growth, feeding and longevity between offspring produced from 2 distinct reproductive modes. Parthenogenetic offspring were generally smaller at hatch than their heterozygous counterparts and, after the first several months post-hatch, failed to increase in mass and length at the same rate as heterozygotes. Parthenogenetic offspring exhibited non-normal swimming behaviors such as spiraling, spy hopping and head standing, which may have been correlated with a gradual decline in the ability of some sharks to properly suction feed. Median lifespan for the parthenotes was 1.05 yr (range: 0.27-6.64 yr); one of the heterozygotes lived to 2.37 yr of age, and the other was alive at the time of this writing in August 2022 and had reached reproductive maturity. By contrast, the 2 longest surviving parthenotes perished just prior to reaching sexual maturity (~5.5 and ~6.5 yr). Parthenogenesis has been documented among ex situ S. tigrinum maintained in aquariums across the globe, and this study demonstrates substantial negative costs to fitness in parthenogenetic offspring compared with their heterozygous siblings. The reduced fitness of parthenotes has implications for managing populations in human care as well as for in situ conservation efforts.
Transcriptomic Signature of the Simulated Microgravity Response in Caenorhabditis elegans and Comparison to Spaceflight Experiments
(Cells, 2023-01-10) Çelen, İrem; Jayasinghe, Aroshan; Doh, Jung H.; Sabanayagam, Chandran R.
Given the growing interest in human exploration of space, it is crucial to identify the effects of space conditions on biological processes. Here, we analyze the transcriptomic response of Caenorhabditis elegans to simulated microgravity and observe the maintained transcriptomic response after returning to ground conditions for four, eight, and twelve days. We show that 75% of the simulated microgravity-induced changes on gene expression persist after returning to ground conditions for four days while most of these changes are reverted after twelve days. Our results from integrative RNA-seq and mass spectrometry analyses suggest that simulated microgravity affects longevity-regulating insulin/IGF-1 and sphingolipid signaling pathways. Finally, we identified 118 genes that are commonly differentially expressed in simulated microgravity- and space-exposed worms. Overall, this work provides insight into the effect of microgravity on biological systems during and after exposure.
Matrix Adhesiveness Regulates Myofibroblast Differentiation from Vocal Fold Fibroblasts in a Bio-orthogonally Cross-linked Hydrogel
(ACS Applied Materials and Interfaces, 2022-11-23) Song, Jiyeon; Gao, Hanyuan; Zhang, He; George, Olivia J.; Hillman, Ashlyn S.; Fox, Joseph M.; Jia, Xinqiao
Repeated mechanical and chemical insults cause an irreversible alteration of extracellular matrix (ECM) composition and properties, giving rise to vocal fold scarring that is refractory to treatment. Although it is well known that fibroblast activation to myofibroblast is the key to the development of the pathology, the lack of a physiologically relevant in vitro model of vocal folds impedes mechanistic investigations on how ECM cues promote myofibroblast differentiation. Herein, we describe a bio-orthogonally cross-linked hydrogel platform that recapitulates the alteration of matrix adhesiveness due to enhanced fibronectin deposition when vocal fold wound healing is initiated. The synthetic ECM (sECM) was established via the cycloaddition reaction of tetrazine (Tz) with slow (norbornene, Nb)- and fast (trans-cyclooctene, TCO)-reacting dienophiles. The relatively slow Tz–Nb ligation allowed the establishment of the covalent hydrogel network for 3D cell encapsulation, while the rapid and efficient Tz–TCO reaction enabled precise conjugation of the cell-adhesive RGDSP peptide in the hydrogel network. To mimic the dynamic changes of ECM composition during wound healing, RGDSP was conjugated to cell-laden hydrogel constructs via a diffusion-controlled bioorthognal ligation method 3 days post encapsulation. At a low RGDSP concentration (0.2 mM), fibroblasts residing in the hydrogel remained quiescent when maintained in transforming growth factor beta 1 (TGF-β1)-conditioned media. However, at a high concentration (2 mM), RGDSP potentiated TGF-β1-induced myofibroblast differentiation, as evidenced by the formation of an actin cytoskeleton network, including F-actin and alpha-smooth muscle actin. The RGDSP-driven fibroblast activation to myofibroblast was accompanied with an increase in the expression of wound healing-related genes, the secretion of profibrotic cytokines, and matrix contraction required for tissue remodeling. This work represents the first step toward the establishment of a 3D hydrogel-based cellular model for studying myofibroblast differentiation in a defined niche associated with vocal fold scarring.
Maize plants and the brace roots that support them
(New Phytologist, 2022-09-14) Sparks, Erin E.
Brace roots are a unique but poorly understood set of organs found in some large cereal crops such as maize. These roots develop from aerial stem nodes and can remain aerial or grow into the ground. Despite their name, the function of these roots to brace the plant was only recently shown. In this article, I discuss the current understanding of brace root function and development, as well as the multitude of open questions that remain about these fascinating organs.
Maize brace root mechanics vary by whorl, genotype, and reproductive stage
(Annals of Botany, 2022-03-03) Hostetler, Ashley N.; Erndwein, Lindsay; Ganji, Elahe; Reneau, Jonathan W.; Killian, Megan L.; Sparks, Erin E.
Background and Aims: Root lodging is responsible for significant crop losses world-wide. During root lodging, roots fail by breaking, buckling, or pulling out of the ground. In maize, above-ground roots, called brace roots, have been shown to reduce root lodging susceptibility. However, the underlying structural-functional properties of brace roots that prevent root lodging are poorly defined. In this study, we quantified structural mechanical properties, geometry, and bending moduli for brace roots from different whorls, genotypes, and reproductive stages. Methods: Using 3-point bend tests, we show that brace root mechanics are variable by whorl, genotype, and reproductive stage. Key Results: Generally, we find that within each genotype and reproductive stage, the brace roots from the first whorl (closest to the ground) had higher structural mechanical properties and a lower bending modulus than brace roots from the second whorl. There was additional variation between genotypes and reproductive stages. Specifically, genotypes with higher structural mechanical properties also had a higher bending modulus, and senesced brace roots had lower structural mechanical properties than hydrated brace roots. Conclusions: Collectively these results highlight the importance of considering whorl-of-origin, genotype, and reproductive stage for quantification of brace root mechanics, which is important for mitigating crop loss due to root mechanical failure.
Erythroid differentiation in mouse erythroleukemia cells depends on Tmod3-mediated regulation of actin filament assembly into the erythroblast membrane skeleton
(FASEB Journal, 2022-02-23) Ghosh, Arit; Coffin, Megan; West, Richard; Fowler, Velia M.
Erythroid differentiation (ED) is a complex cellular process entailing morphologically distinct maturation stages of erythroblasts during terminal differentiation. Studies of actin filament (F-actin) assembly and organization during terminal ED have revealed essential roles for the F-actin pointed-end capping proteins, tropomodulins (Tmod1 and Tmod3). Tmods bind tropomyosins (Tpms), which enhance Tmod capping and F-actin stabilization. Tmods can also nucleate F-actin assembly, independent of Tpms. Tmod1 is present in the red blood cell (RBC) membrane skeleton, and deletion of Tmod1 in mice leads to a mild compensated anemia due to mis-regulated F-actin lengths and membrane instability. Tmod3 is not present in RBCs, and global deletion of Tmod3 leads to embryonic lethality in mice with impaired ED. To further decipher Tmod3’s function during ED, we generated a Tmod3 knockout in a mouse erythroleukemia cell line (Mel ds19). Tmod3 knockout cells appeared normal prior to ED, but showed defects during progression of ED, characterized by a marked failure to reduce cell and nuclear size, reduced viability, and increased apoptosis. Tmod3 does not assemble with Tmod1 and Tpms into the Triton X-100 insoluble membrane skeleton during ED, and loss of Tmod3 had no effect on α1,β1-spectrin and protein 4.1R assembly into the membrane skeleton. However, F-actin, Tmod1 and Tpms failed to assemble into the membrane skeleton during ED in absence of Tmod3. We propose that Tmod3 nucleation of F-actin assembly promotes incorporation of Tmod1 and Tpms into membrane skeleton F-actin, and that this is integral to morphological maturation and cell survival during erythroid terminal differentiation.
Nanocrystalline protein domains via salting-out
(Acta Crystallographica Section F: Structural Biology Communications, 2021-11-02) Greene, D. G.; Modla, S.; Sandler, S. I.; Wagner, N. J.; Lenhoff, A. M.
Protein salting-out is a well established phenomenon that in many cases leads to amorphous structures and protein gels, which are usually not considered to be useful for protein structure determination. Here, microstructural measurements of several different salted-out protein dense phases are reported, including of lysozyme, ribonuclease A and an IgG1, showing that salted-out protein gels unexpectedly contain highly ordered protein nanostructures that assemble hierarchically to create the gel. The nanocrystalline domains are approximately 10–100 nm in size, are shown to have structures commensurate with those of bulk crystals and grow on time scales in the order of an hour to a day. Beyond revealing the rich, hierarchical nanoscale to mesoscale structure of protein gels, the nanocrystals that these phases contain are candidates for structural biology on next-generation X-ray free-electron lasers, which may enable the study of biological macromolecules that are difficult or impossible to crystallize in bulk.
Mechanisms of extracellular S0 globule production and degradation in Chlorobaculum tepidum via dynamic cell–globule interactions
(Microbiology Society, 2016-01-07) Marnocha, C. L.; Levy, A. T.; Powell, D. H.; Hanson, T. E.; Chan, C. S.; ; Marnocha, C. L.; Levy, A. T.; Powell, D. H.; Hanson, T. E.; Chan, C. S.
The Chlorobiales are anoxygenic phototrophs that produce solid, extracellular elemental sulfur globules as an intermediate step in the oxidation of sulfide to sulfate. These organisms must export sulfur while preventing cell encrustation during S0 globule formation; during globule degradation they must find and mobilize the sulfur for intracellular oxidation to sulfate. To understand how the Chlorobiales address these challenges, we characterized the spatial relationships and physical dynamics of Chlorobaculum tepidum cells and S0 globules by light and electron microscopy. Cba. tepidum commonly formed globules at a distance from cells. Soluble polysulfides detected during globule production may allow for remote nucleation of globules. Polysulfides were also detected during globule degradation, probably produced as an intermediate of sulfur oxidation by attached cells. Polysulfides could feed unattached cells, which made up over 80% of the population and had comparable growth rates to attached cells. Given that S0 is formed remotely from cells, there is a question as to how cells are able to move toward S0 in order to attach. Time-lapse microscopy shows that Cba. tepidum is in fact capable of twitching motility, a finding supported by the presence of genes encoding type IV pili. Our results show how Cba. tepidum is able to avoid mineral encrustation and benefit from globule degradation even when not attached. In the environment, Cba. tepidum may also benefit from soluble sulfur species produced by other sulfur-oxidizing or sulfur-reducing bacteria as these organisms interact with its biogenic S0 globules.
Genome assembly with in vitro proximity ligation data and whole-genome triplication in lettuce
(Nature Publishing Group, 2017-04-12) Reyes-Chin-Wo, Sebastian; Wang, Zhiwen; Yang, Xinhua; Kozik, Alexander; Arikit, Siwaret; Song, Chi; Xia, Liangfeng; Froenicke, Lutz; Lavelle, Dean O.; Truco, Marı´a-Jose´; Xia, Rui; Zhu, Shilin; Xu, Chunyan; Xu, Huaqin; Xu, Xun; Cox, Kyle; Korf, Ian; Meyers, Blake C.; Michelmore, Richard W.; Sebastian Reyes-Chin-Wo, Zhiwen Wang, Xinhua Yang, Alexander Kozik, Siwaret Arikit, Chi Song, Liangfeng Xia, Lutz Froenicke, Dean O. Lavelle, Marı´a-Jose´ Truco, Rui Xia, Shilin Zhu, Chunyan Xu, Huaqin Xu, Xun Xu, Kyle Cox, Ian Korf, Blake C. Meyers & Richard W. Michelmore; Arikit, Siwaret; Meyers, Blake C
Lettuce (Lactuca sativa) is a major crop and a member of the large, highly successful Compositae family of flowering plants. Here we present a reference assembly for the species and family. This was generated using whole-genome shotgun Illumina reads plus in vitro proximity ligation data to create large superscaffolds; it was validated genetically and superscaffolds were oriented in genetic bins ordered along nine chromosomal pseudomolecules. We identify several genomic features that may have contributed to the success of the family, including genes encoding Cycloidea-like transcription factors, kinases, enzymes involved in rubber biosynthesis and disease resistance proteins that are expanded in the genome. We characterize 21 novel microRNAs, one of which may trigger phasiRNAs from numerous kinase transcripts. We provide evidence for a whole-genome triplication event specific but basal to the Compositae. We detect 26% of the genome in triplicated regions containing 30% of all genes that are enriched for regulatory sequences and depleted for genes involved in defence.
Presence of pathogenic Escherichia coli is correlated with bacterial community diversity and composition on pre-harvest cattle hides
(Biomed Central Ltd, 3/22/16) Chopyk,Jessica; Moore,Ryan M.; DiSpirito,Zachary; Stromberg,Zachary R.; Lewis,Gentry L.; Renter,David G.; Cernicchiaro,Natalia; Moxley,Rodney A.; Wommack,K. Eric; Jessica Chopyk, Ryan M. Moore, Zachary DiSpirito, Zachary R. Stromberg, Gentry L. Lewis, David G. Renter, Natalia Cernicchiaro, Rodney A. Moxley and K. Eric Wommack; Wommack, K Eric
Background: Since 1982, specific serotypes of Shiga toxin-producing Escherichia coli (STEC) have been recognized as significant foodborne pathogens acquired from contaminated beef and, more recently, other food products. Cattle are the major reservoir hosts of these organisms, and while there have been advancements in food safety practices and industry standards, STEC still remains prevalent within beef cattle operations with cattle hides implicated as major sources of carcass contamination. To investigate whether the composition of hide-specific microbial communities are associated with STEC prevalence, 16S ribosomal RNA (rRNA) bacterial community profiles were obtained from hide and fecal samples collected from a large commercial feedlot over a 3-month period. These community data were examined amidst an extensive collection of prevalence data on a subgroup of STEC that cause illness in humans, referred to as enterohemorrhagic E. coli (EHEC). Fecal 16S rRNA gene OTUs (operational taxonomic units) were subtracted from the OTUs found within each hide 16S rRNA amplicon library to identify hide-specific bacterial populations. Results: Comparative analysis of alpha diversity revealed a significant correlation between low bacterial diversity and samples positive for the presence of E. coli O157:H7 and/or the non-O157 groups: O26, O111, O103, O121, O45, and O145. This trend occurred regardless of diversity metric or fecal OTU presence. The number of EHEC serogroups present in the samples had a compounding effect on the inverse relationship between pathogen presence and bacterial diversity. Beta diversity data showed differences in bacterial community composition between samples containing O157 and non-O157 populations, with certain OTUs demonstrating significant changes in relative abundance. Conclusions: The cumulative prevalence of the targeted EHEC serogroups was correlated with low bacterial community diversity on pre-harvest cattle hides. Understanding the relationship between indigenous hide bacterial communities and populations may provide strategies to limit EHEC in cattle and provide biomarkers for EHEC risk assessment.
MCUR1 Is a Scaffold Factor for the MCU Complex Function and Promotes Mitochondrial Bioenergetics
(Cell Press, 5/24/16) Tomar,Dhanendra; Dong,Zhiwei; Shanmughapriya,Santhanam; Koch,Diana A.; Thomas,Toby; Hoffman,Nicholas E.; Timbalia,Shrishiv A.; Goldman,Samuel J.; Breves,Sarah L.; Corbally,Daniel P.; Nemani,Neeharika; Fairweather,Joseph P.; Cutri,Allison R.; Zhang,Xueqian; Song,Jianliang; Jana,Fabian; Huang,Jianhe; Barrero,Carlos; Rabinowitz,Joseph E.; Luongo,Timothy S.; Schumacher,Sarah M.; Rockman,Michael E.; Dietrich,Alexander; Merali,Salim; Caplan,Jeffrey; Stathopulos,Peter; Ahima,Rexford S.; Cheung,Joseph Y.; Houser,Steven R.; Koch,Walter J.; Patel,Vickas; Gohil,Vishal M.; Elrod,John W.; Rajan,Sudarsan; Madesh,Muniswamy; Dhanendra Tomar, Zhiwei Dong,Santhanam Shanmughapriya, Diana A. Koch, Toby Thomas, Nicholas E. Hoffman, Shrishiv A. Timbalia, Samuel J. Goldman, Sarah L. Breves, Daniel P. Corbally, Neeharika Nemani, Joseph P. Fairweather, Allison R. Cutri, Xueqian Zhang, Jianliang Song, Fabian Jana, Jianhe Huang, Carlos Barrero, Joseph E. Rabinowitz,Timothy S. Luongo, Sarah M. Schumacher, Michael E. Rockman, Alexander Dietrich, Salim Merali, Jeffrey Caplan, Peter Stathopulos, Rexford S. Ahima, Joseph Y. Cheung, Steven R. Houser, Walter J. Koch, Vickas Patel, Vishal M. Gohil, John W. Elrod, Sudarsan Rajan and Muniswamy Madesh; Caplan, Jeffrey Lewis
Mitochondrial Ca2+ Uniporter (MCU)-dependent mitochondrial Ca2+ uptake is the primary mechanism for increasing matrix Ca2+ in most cell types. However, a limited understanding of the MCU complex assembly impedes the comprehension of the precise mechanisms underlying MCU activity. Here, we report that mouse cardiomyocytes and endothelial cells lacking MCU regulator 1 (MCUR1) have severely impaired [Ca2+](m) uptake and I-MCU current. MCUR1 binds to MCU and EMRE and function as a scaffold factor. Our protein binding analyses identified the minimal, highly conserved regions of coiled-coil domain of both MCU and MCUR1 that are necessary for heterooligomeric complex formation. Loss of MCUR1 perturbed MCU heterooligomeric complex and functions as a scaffold factor for the assembly of MCU complex. Vascular endothelial deletion of MCU and MCUR1 impaired mitochondrial bioenergetics, cell proliferation, and migration but elicited autophagy. These studies establish the existence of a MCU complex that assembles at the mitochondrial integral membrane and regulates Ca2+-dependent mitochondrial metabolism.
D1FHS, the Type Strain of the Ammonia-Oxidizing Bacterium Nitrosococcus wardiae spec. nov.: Enrichment, Isolation, Phylogenetic, and Growth Physiological Characterization
(Frontiers Media S.A., 4/14/16) Wang,Lin; Lim,Chee Kent; Dang,Hongyue; Hanson,Thomas E.; Klotz,Martin G.; Lin Wang, Chee Kent Lim, Hongyue Dang, Thomas E. Hanson and Martin G. Klotz; Hanson, Thomas E
An ammonia-oxidizing bacterium, strain D1FHS, was enriched into pure culture from a sediment sample retrieved in Jiaozhou Bay, a hyper-eutrophic semi-closed water body hosting the metropolitan area of Qingdao, China. Based on initial 16S rRNA gene sequence analysis, strain D1FHS was classified in the genus Nitrosococcus, family Chromatiaceae, order Chromatiales, class Gammaproteobacteria; the 16S rRNA gene sequence with highest level of identity to that of D1FHS was obtained from Nitrosococcus halophilus Nc4T. The average nucleotide identity between the genomes of strain D1FHS and N. halophilus strain Nc4 is 89.5%. Known species in the genus Nitrosococcus are obligate aerobic chemolithotrophic ammonia-oxidizing bacteria adapted to and restricted to marine environments. The optimum growth (maximum nitrite production) conditions for D1FHS in a minimal salts medium are: 50 mM ammonium and 700 mM NaCl at pH of 7.5 to 8.0 and at 37 degrees C in dark. Because pertinent conditions for other studied Nitrosococcus spp. are 100200 mM ammonium and <700 mM NaCl at pH of 7.5 to 8.0 and at 2832 degrees C, D1FHS is physiologically distinct from other Nitrosococcus spp. in terms of substrate, salt, and thermal tolerance.
CO2 fixation by anaerobic non-photosynthetic mixotrophy for improved carbon conversion
(Nature Publishing Group, 9/30/16) Jones,Shawn W.; Fast,Alan G.; Carlson,Ellinor D.; Wiedel,Carrissa A.; Au,Jennifer; Antoniewicz,Maciek R.; Papoutsakis,Eleftherios T.; Tracy,Bryan P.; Shawn W. Jones, Alan G. Fast, Ellinor D. Carlson, Carrissa A. Wiedel, Jennifer Au, Maciek R. Antoniewicz, Eleftherios T. Papoutsakis & Bryan P. Tracy; Antoniewicz, Maciej Robert; Papoutsakis, Eleftherios
Maximizing the conversion of biogenic carbon feedstocks into chemicals and fuels is essential for fermentation processes as feedstock costs and processing is commonly the greatest operating expense. Unfortunately, for most fermentations, over one-third of sugar carbon is lost to CO2 due to the decarboxylation of pyruvate to acetyl-CoA and limitations in the reducing power of the bio-feedstock. Here we show that anaerobic, non-photosynthetic mixotrophy, defined as the concurrent utilization of organic (for example, sugars) and inorganic (for example, CO2) substrates in a single organism, can overcome these constraints to increase product yields and reduce overall CO2 emissions. As a proof-of-concept, Clostridium ljungdahlii was engineered to produce acetone and achieved a mass yield 138% of the previous theoretical maximum using a high cell density continuous fermentation process. In addition, when enough reductant (that is, H-2) is provided, the fermentation emits no CO2. Finally, we show that mixotrophy is a general trait among acetogens.
Mechanisms of extracellular S0 globule production and degradation in Chlorobaculum tepidum via dynamic cell–globule interactions
(Microbiology Society, 2016-01-07) Marnocha, C. L.; Levy, A. T.; Powell, D. H.; Hanson, T. E.; Chan, C. S.; C. L. Marnocha, A. T. Levy, D. H. Powell, T. E. Hanson and C. S. Chan; Marnocha, C. L.; Levy, A. T.; Powell, D. H.; Hanson, T. E.; Chan, C. S.
The Chlorobiales are anoxygenic phototrophs that produce solid, extracellular elemental sulfur globules as an intermediate step in the oxidation of sulfide to sulfate. These organisms must export sulfur while preventing cell encrustation during S0 globule formation; during globule degradation they must find and mobilize the sulfur for intracellular oxidation to sulfate. To understand how the Chlorobiales address these challenges, we characterized the spatial relationships and physical dynamics of Chlorobaculum tepidum cells and S0 globules by light and electron microscopy. Cba. tepidum commonly formed globules at a distance from cells. Soluble polysulfides detected during globule production may allow for remote nucleation of globules. Polysulfides were also detected during globule degradation, probably produced as an intermediate of sulfur oxidation by attached cells. Polysulfides could feed unattached cells, which made up over 80% of the population and had comparable growth rates to attached cells. Given that S0 is formed remotely from cells, there is a question as to how cells are able to move toward S0 in order to attach. Time-lapse microscopy shows that Cba. tepidum is in fact capable of twitching motility, a finding supported by the presence of genes encoding type IV pili. Our results show how Cba. tepidum is able to avoid mineral encrustation and benefit from globule degradation even when not attached. In the environment, Cba. tepidum may also benefit from soluble sulfur species produced by other sulfur-oxidizing or sulfur-reducing bacteria as these organisms interact with its biogenic S0 globules.

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