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Application of a Human Blood Brain Barrier Organ-on-a-Chip Model to Evaluate Small Molecule Effectiveness against Venezuelan Equine Encephalitis Virus
(Viruses-basel, 2022-12-15) Boghdeh, Niloufar A.; Risner, Kenneth H.; Barrera, Michael D.; Britt, Clayton M.; Schaffer, David K.; Alem, Farhang; Brown, Jacquelyn A.; Wikswo, John P.; Narayanan, Aarthi
The blood brain barrier (BBB) is a multicellular microenvironment that plays an important role in regulating bidirectional transport to and from the central nervous system (CNS). Infections by many acutely infectious viruses such as alphaviruses and flaviviruses are known to impact the integrity of the endothelial lining of the BBB. Infection by Venezuelan Equine Encephalitis Virus (VEEV) through the aerosol route causes significant damage to the integrity of the BBB, which contributes to long-term neurological sequelae. An effective therapeutic intervention strategy should ideally not only control viral load in the host, but also prevent and/or reverse deleterious events at the BBB. Two dimensional monocultures, including trans-well models that use endothelial cells, do not recapitulate the intricate multicellular environment of the BBB. Complex in vitro organ-on-a-chip models (OOC) provide a great opportunity to introduce human-like experimental models to understand the mechanistic underpinnings of the disease state and evaluate the effectiveness of therapeutic candidates in a highly relevant manner. Here we demonstrate the utility of a neurovascular unit (NVU) in analyzing the dynamics of infection and proinflammatory response following VEEV infection and therapeutic effectiveness of omaveloxolone to preserve BBB integrity and decrease viral and inflammatory load.
The Astonishing Behavior of Electric Eels
(Frontiers in Integrative Neuroscience, 2019-07-16) Catania, Kenneth C.
The remarkable physiology of the electric eel (Electrophorus electricus) made it one of the first model species in science. It was pivotal for understanding animal electricity in the 1700s, was investigated by Humboldt and Faraday in the 1800s, was leveraged to isolate the acetylcholine receptor in the 20th century, and has inspired the design of new power sources and provided insights to electric organ evolution in the 21st century. And yet few studies have investigated the electric eel's behavior. This review focuses on a series of recently discovered behaviors that evolved alongside the eel's extreme physiology. Eels use their high-voltage electric discharge to remotely control prey by transcutaneously activating motor neurons. Hunting eels use this behavior in two different ways. When prey have been detected, eels use high-voltage to cause immobility by inducing sustained, involuntary muscle contractions. On the other hand, when prey are hidden, eels often use brief pulses to induce prey twitch, which causes a water movement detected by the eel's mechanoreceptors. Once grasped in the eel's jaws, difficult prey are often subdued by sandwiching them between the two poles (head and tail) of the eel's powerful electric organ. The resulting concentration of the high-voltage discharge, delivered at high-rates, causes involuntary fatigue in prey muscles. This novel strategy for inactivating muscles is functionally analogous to poisoning the neuromuscular junction with venom. For self-defense, electric eels leap from the water to directly electrify threats, efficiently activating nociceptors to deter their target. The latter behavior supports a legendary account by Alexander von Humboldt who described a battle between electric eels and horses in 1800. Finally, electric eels use high-voltage not only as a weapon, but also to efficiently track fast-moving prey with active electroreception. In conclusion, remarkable behaviors go hand in hand with remarkable physiology.
Chemical Control of Mosquitoes and the Pesticide Treadmill: A Case for Photosensitive Insecticides as Larvicides
(Insects, 2022-11-28) Meier, Cole J.; Rouhier, Matthew F.; Hillyer, Julian F.
Simple Summary Mosquitoes transmit disease, and over the past century, mosquito control has mostly relied on chemical insecticides that target the adult life stage. We review methods of mosquito control and argue that photoactive molecules that target larvae-called photosensitive insecticides or PSIs-are an environmentally friendly addition to our mosquitocidal arsenal. Insecticides reduce the spread of mosquito-borne disease. Over the past century, mosquito control has mostly relied on neurotoxic chemicals-such as pyrethroids, neonicotinoids, chlorinated hydrocarbons, carbamates and organophosphates-that target adults. However, their persistent use has selected for insecticide resistance. This has led to the application of progressively higher amounts of insecticides-known as the pesticide treadmill-and negative consequences for ecosystems. Comparatively less attention has been paid to larvae, even though larval death eliminates a mosquito's potential to transmit disease and reproduce. Larvae have been targeted by source reduction, biological control, growth regulators and neurotoxins, but hurdles remain. Here, we review methods of mosquito control and argue that photoactive molecules that target larvae-called photosensitive insecticides or PSIs-are an environmentally friendly addition to our mosquitocidal arsenal. PSIs are ingested by larvae and produce reactive oxygen species (ROS) when activated by light. ROS then damage macromolecules resulting in larval death. PSIs are degraded by light, eliminating environmental accumulation. Moreover, PSIs only harm small translucent organisms, and their broad mechanism of action that relies on oxidative damage means that resistance is less likely to evolve. Therefore, PSIs are a promising alternative for controlling mosquitoes in an environmentally sustainable manner.
Clinical characteristics and electrophysiologic properties of SCN5A variants in fever-induced Brugada syndrome
(Ebiomedicine, 2022-12) Li, Bian
Background Brugada syndrome (BrS) is a severe inherited arrhythmia syndrome that can be unmasked by fever.Methods A multicentre clinical analysis was performed in 261 patients diagnosed with fever-induced BrS, including 198 (75.9%) and 27 (10.3%) patients who received next-generation genetic sequencing and epicardial arrhythmogenic substrate (AS) mapping, respectively.Findings In fever-induced BrS patients, pathogenic or likely pathogenic (P/LP) SCN5A variant carriers developed fever-induced BrS at a younger age, and more often in females and those of Caucasian descent. They exhibited significant electrophysical abnormalities, including a larger epicardial AS area, and more prolonged abnormal epicardial electrograms. During a median follow-up of 50.5 months (quartiles 32.5-81.5 months) after the diagnosis, major cardiac events (MCE) occurred in 27 (14.4%) patients. Patients with P/LP SCN5A variants had a higher ratio of MCE compared with the rest. Additionally, history of syncope, QRS duration, and Tpe interval could also predict an increased risk for future MCE according to univariate analysis. Multivariate analysis indicated that only P/LP SCN5A variants were independent significant predictors of MCE. Computational structural modelling showed that most variants are destabilizing, suggesting that Nav1.5 structure destabilization caused by SCN5A missense variants may contribute to fever-induced BrS.Interpretation In our cohort, P/LP SCN5A variant carriers with fever-induced BrS are more prevalent among patients of Caucasian descent, females, and younger patients. These patients exhibit aggressive electrophysiological abnormalities and worse outcome, which warrants closer monitoring and more urgent management of fever.
Diverse environmental perturbations reveal the evolution and context-dependency of genetic effects on gene expression levels
(Genome Research, 2022-10) Lea, Amanda J.; Peng, Julie; Ayroles, Julien F.
There is increasing appreciation that, in addition to being shaped by an individual's genotype and environment, most complex traits are also determined by poorly understood interactions between these two factors. So-called "genotype x environment" (GxE) interactions remain difficult to map at the organismal level but can be uncovered using molecular phenotypes. To do so at large scale, we used TM3'seq to profile transcriptomes across 12 cellular environments in 544 immortalized B cell lines from the 1000 Genomes Project. We mapped the genetic basis of gene expression levels across environments and revealed a context-dependent genetic architecture: The average heritability of gene expression levels increased in treatment relative to control conditions, and on average, each treatment revealed new expression quantitative trait loci (eQTLs) at 11% of genes. Across our experiments, 22% of all identified eQTLs were context-dependent, and this group was enriched for trait- and disease-associated loci. Further, evolutionary analyses suggested that positive selection has shaped GxE loci involved in responding to immune challenges and hormones but not to man-made chemicals. We hypothesize that this reflects a reduced opportunity for selection to act on responses to molecules recently introduced into human environments. Together, our work highlights the importance of considering an exposure's evolutionary history when studying and interpreting GxE interactions, and provides new insight into the evolutionary mechanisms that maintain GxE loci in human populations.
Efficacy of virtual and asynchronous teaching of computer-assisted diagnosis of genetic diseases seen in clinics
(Wiley, 2021-12-11) Hash, Mary Grace; Walker, Phillip; Laferriere, Heather; Melton, Leeanna; Heller, Lauren; Phillips, John
We studied if clinicians could gain sufficient working knowledge of a computer-assisted diagnostic decision support system (DDSS) (SimulConsult), to make differential diagnoses (DDx) of genetic disorders. We hypothesized that virtual training could be convenient, asynchronous, and effective in teaching clinicians how to use a DDSS. We determined the efficacy of virtual, asynchronous teaching for clinicians to gain working knowledge to make computer-assisted DDx. Our study consisted of three surveys (Baseline, Training, and After Use) and a series of case problems sent to clinicians at Vanderbilt University Medical Center. All participants were able to generate computer-assisted DDx that achieved passing scores of the case problems. Between 75% and 92% agreed/completely agreed the DDSS was useful to their work and for clinical decision support and was easy to use. Participants' use of the DDSS resulted in statistically significant time savings in key tasks and in total time spent on clinical tasks. Our results indicate that virtual, asynchronous teaching can be an effective format to gain a working knowledge of a DDSS, and its clinical use could result in significant time savings across multiple tasks as well as facilitate synergistic interaction between clinicians and lab specialists. This approach is especially pertinent and offers value amid the COVID-19 pandemic.
Immune repertoire fingerprinting by principal component analysis reveals shared features in subject groups with common exposures
(BMC Bioinformatics, 2019-12-04) Sevy, Alexander M.; Soto, Cinque; Bombardi, Robin G.; Meiler, Jens; Crowe, James E., Jr.
Background: Advances in next-generation sequencing (NGS) of antibody repertoires have led to an explosion in B cell receptor sequence data from donors with many different disease states. These data have the potential to detect patterns of immune response across populations. However, to this point it has been difficult to interpret such patterns of immune response between disease states in the absence of functional data. There is a need for a robust method that can be used to distinguish general patterns of immune responses at the antibody repertoire level. Results: We developed a method for reducing the complexity of antibody repertoire datasets using principal component analysis (PCA) and refer to our method as "repertoire fingerprinting." We reduce the high dimensional space of an antibody repertoire to just two principal components that explain the majority of variation in those repertoires. We show that repertoires from individuals with a common experience or disease state can be clustered by their repertoire fingerprints to identify common antibody responses. Conclusions: Our repertoire fingerprinting method for distinguishing immune repertoires has implications for characterizing an individual disease state. Methods to distinguish disease states based on pattern recognition in the adaptive immune response could be used to develop biomarkers with diagnostic or prognostic utility in patient care. Extending our analysis to larger cohorts of patients in the future should permit us to define more precisely those characteristics of the immune response that result from natural infection or autoimmunity.
Large protein organelles form a new iron sequestration system with high storage capacity
(eLife, 2019-07-08) Giessen, Tobias W.; Orlando, Benjamin J.; Verdegaal, Andrew A.; Chambers, Melissa G.; Gardener, Jules; Bell, David C.; Birrane, Gabriel; Liao, Maofu; Silver, Pamela A.
Iron storage proteins are essential for cellular iron homeostasis and redox balance. Ferritin proteins are the major storage units for bioavailable forms of iron. Some organisms lack ferritins, and it is not known how they store iron. Encapsulins, a class of protein-based organelles, have recently been implicated in microbial iron and redox metabolism. Here, we report the structural and mechanistic characterization of a 42 nm two-component encapsulin-based iron storage compartment from Quasibacillus thermotolerans. Using cryo-electron microscopy and x-ray crystallography, we reveal the assembly principles of a thermostable T = 4 shell topology and its catalytic ferroxidase cargo and show interactions underlying cargo-shell co-assembly. This compartment has an exceptionally large iron storage capacity storing over 23,000 iron atoms. Our results reveal a new approach for survival in diverse habitats with limited or fluctuating iron availability via an iron storage system able to store 10 to 20 times more iron than ferritin.
Mitofusins Mfn1 and Mfn2 Are Required to Preserve Glucose- but Not Incretin-Stimulated beta-Cell Connectivity and Insulin Secretion
(Diabetes, 2022-07) Xu, Yanwen; Gu, Guoqiang
Mitochondrial glucose metabolism is essential for stimulated insulin release from pancreatic beta-cells. Whether mitofusin gene expression, and hence, mitochondrial network integrity, is important for glucose or incretin signaling has not previously been explored. Here, we generated mice with beta-cell-selective, adult-restricted deletion knock-out (dKO) of the mitofusin genes Mfn1 and Mfn2 (beta Mfn1/2 dKO). beta Mfn1/2-dKO mice displayed elevated fed and fasted glycemia and a more than fivefold decrease in plasma insulin. Mitochondrial length, glucose-induced polarization, ATP synthesis, and cytosolic and mitochondrial Ca2+ increases were all reduced in dKO islets. In contrast, oral glucose tolerance was more modestly affected in beta Mfn1/2-dKO mice, and glucagon-like peptide 1 or glucose-dependent insulinotropic peptide receptor agonists largely corrected defective glucose-stimulated insulin secretion through enhanced EPAC-dependent signaling. Correspondingly, cAMP increases in the cytosol, as measured with an Epac-camps-based sensor, were exaggerated in dKO mice. Mitochondrial fusion and fission cycles are thus essential in the beta-cell to maintain normal glucose, but not incretin, sensing. These findings broaden our understanding of the roles of mitofusins in beta-cells, the potential contributions of altered mitochondrial dynamics to diabetes development, and the impact of incretins on this process.
Nutritional Heterogeneity Among Aspergillus fumigatus Strains Has Consequences for Virulence in a Strain- and Host-Dependent Manner
(FRONTIERS IN MICROBIOLOGY, 2019-04-24) Steenwyk, Jacob L.
Acquisition and subsequent metabolism of different carbon and nitrogen sources have been shown to play an important role in virulence attributes of the fungal pathogen Aspergillus fumigatus, such as the secretion of host tissue-damaging proteases and fungal cell wall integrity. We examined the relationship between the metabolic processes of carbon catabolite repression (CCR), nitrogen catabolite repression (NCR) and virulence in a variety of A. fumigatus clinical isolates. A considerable amount of heterogeneity with respect to the degree of CCR and NCR was observed and a positive correlation between NCR and virulence in a neutropenic mouse model of pulmonary aspergillosis (PA) was found. Isolate Afs35 was selected for further analysis and compared to the reference strain A1163, with both strains presenting the same degree of virulence in a neutropenic mouse model of PA. Afs35 metabolome analysis in physiological-relevant carbon sources indicated an accumulation of intracellular sugars that also serve as cell wall polysaccharide precursors. Genome analysis showed an accumulation of missense substitutions in the regulator of protease secretion and in genes encoding enzymes required for cell wall sugar metabolism. Based on these results, the virulence of strains Afs35 and A1163 was assessed in a triamcinolone murine model of PA and found to be significantly different, confirming the known importance of using different mouse models to assess strain-specific pathogenicity. These results highlight the importance of nitrogen metabolism for virulence and provide a detailed example of the heterogeneity that exists between A. fumigatus isolates with consequences for virulence in a strain-specific and host-dependent manner.
Species-level Repertoire Size Predicts a Correlation Between Individual Song Elaboration and Reproductive Success
(ECOLOGY AND EVOLUTION, 2019-07) Robinson, Cristina M.; Creanza, Nicole
Birdsong has long been considered a sexually selected trait that relays honest information about male quality, and laboratory studies generally suggest that female songbirds prefer larger repertoires. However, analysis of field studies across species surprisingly revealed a weak correlation between song elaboration and reproductive success, and it remains unknown why only certain species show this correlation in nature. Taken together, these studies suggest that females in numerous species can detect and prefer larger repertoires in a laboratory setting, but larger individual repertoires correlate with reproductive success only in a subset of these species. This prompts the question: Do the species that show a stronger correlation between reproductive success and larger individual repertoires in nature have anything in common? In this study, we test whether between-species differences in two song-related variables-species average syllable repertoire size and adult song stability over time-can be used to predict the importance of individual song elaboration in reproductive success within a species. Our cross-species meta-analysis of field studies revealed that species with larger average syllable repertoire sizes exhibited a stronger correlation between individual elaboration and reproductive success than species with smaller syllable repertoires. Song stability versus plasticity in adulthood provided little predictive power on its own, suggesting that the putative correlation between repertoire size and age in open-ended learners does not explain the association between song elaboration and reproductive success.
Synechocystis: A model system for expanding the study of cyanobacterial circadian rhythms
(2023-01-04) Zhao, Chi; Xu, Yao; Wang, Bo; Johnson, Carl Hirschie
The study of circadian rhythms in bacteria was transformed by studies of the cyanobacterium Synechococcus elongatus. However, in a number of respects S. elongatus is atypical, and while those unusual characteristics were helpful for rapid progress in the past, another commonly used cyanobacterial species, Synechocystis sp. PCC 6803, may be more representative and therefore more productive for future insights into bacterial clock mechanisms. In the past, circadian studies of Synechocystis have suffered from not having an excellent reporter of circadian gene expression, but we introduce here a new luminescence reporter that rivals the reporters that have been used so successfully in S. elongatus. Using this new system, we generate for the first time in Synechocystis circadian period mutants resulting from point mutations. The temperature compensation and dark-pulse resetting that mediates entrainment to the environment is characterized. Moreover, we analyse the complex organization of clock genes in Synechocystis and identify which genes are essential for circadian rhythmicity and adaptive fitness for entrainment and optimal phase alignment to environmental cycles (and which genes are not). These developments will provide impetus for new approaches towards understanding daily timekeeping mechanisms in bacteria.
The Wolbachia Mobilome in Culex Pipiens Includes a Putative Plasmid (vol 10, 1050, 2019)
(NATURE COMMUNICATIONS, 2019-07-12) Reveillaud, Julie; Bordenstein, Sarah R.; Cruaud, Corinne; Shaiber, Alon; Esen, Ozcan C.; Weill, Mylene; Makoundou, Patrick; Lolans, Karen; Watson, Andrea; Rakotoarivony, Ignace; Bordenstein, Seth R.; Eren, A. Murat
Wolbachia is a genus of obligate intracellular bacteria found in nematodes and arthropods worldwide, including insect vectors that transmit dengue, West Nile, and Zika viruses. Wolbachia's unique ability to alter host reproductive behavior through its temperate bacteriophage WO has enabled the development of new vector control strategies. However, our understanding of Wolbachia's mobilome beyond its bacteriophages is incomplete. Here, we reconstruct near-complete Wolbachia genomes from individual ovary metagenomes of four wild Culex pipiens mosquitoes captured in France. In addition to viral genes missing from the Wolbachia reference genome, we identify a putative plasmid (pWCP), consisting of a 9.23-kbp circular element with 14 genes. We validate its presence in additional Culex pipiens mosquitoes using PCR, long-read sequencing, and screening of existing metagenomes. The discovery of this previously unrecognized extrachromosomal element opens additional possibilities for genetic manipulation of Wolbachia.

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