Scholarly Works, Virginia-Maryland College of Veterinary Medicine

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Sex-Linked Growth Disorder and Aberrant Pituitary Gene Expression in Nestin-Cre-Mediated Egr1 Conditional Knockout Mice
Swilley, Cody; Lin, Yu; Zheng, Yuze; Xu, Xiguang; Liu, Min; Zimmerman, Kurt; Xie, Hehuang (MDPI, 2023-07-06)
Genes that regulate hormone release are essential for maintaining metabolism and energy balance. Egr1 encodes a transcription factor that regulates hormone production and release, and a decreased in growth hormones has been reported in Egr1 knockout mice. A reduction in growth hormones has also been observed in Nestin-Cre mice, a model frequently used to study the nervous system. Currently, it is unknown how Egr1 loss or the Nestin-Cre driver disrupt pituitary gene expression. Here, we compared the growth curves and pituitary gene expression profiles of Nestin-Cre-mediated Egr1 conditional knockout (Egr1cKO) mice with those of their controls. Reduced body weight was observed in both the Nestin-Cre and Egr1cKO mice, and the loss of Egr1 had a slightly more severe impact on female mice than on male mice. RNA-seq data analyses revealed that the sex-related differences were amplified in the Nestin-Cre-mediated Egr1 conditional knockout mice. Additionally, in the male mice, the influence of Egr1cKO on pituitary gene expression may be overridden by the Nestin-Cre driver. Differentially expressed genes associated with the Nestin-Cre driver were significantly enriched for genes related to growth factor activity and binding. Altogether, our results demonstrate that Nestin-Cre and the loss of Egr1 in the neuronal cell lineage have distinct impacts on pituitary gene expression in a sex-specific manner.
Effects of Dietary Ferulic Acid on Intestinal Health and Ileal Microbiota of Tianfu Broilers Challenged with Lipopolysaccharide
Tang, Ziting; Shu, Gang; Du, Hong; Zheng, Yilei; Fu, Hualin; Zhang, Wei; Lv, Cheng; Xu, Funeng; Li, Haohuan; Ouyang, Ping; Lin, Juchun; Chang, Li-Jen; Amevor, Felix Kwame; Zhao, Xiaoling (MDPI, 2023-02-10)
Lipopolysaccharide (LPS) has been considered the primary agent to establish animal models of inflammation, immunological stress, and organ injury. Previous studies have demonstrated that LPS impaired gastrointestinal development and disrupted intestinal microbial composition and metabolism. Ferulic acid (FA) isolated from multiple plants exhibits multiple biological activities. This study investigated whether FA ameliorated intestinal function and microflora in LPS-challenged Tianfu broilers. The results showed that LPS challenge impaired intestinal function, as evidenced by decreased antioxidant functions (p < 0.05), disrupted morphological structure (p < 0.05), and increased intestinal permeability (p < 0.05); however, these adverse effects were improved by FA supplementation. Additionally, FA supplementation preserved sIgA levels (p < 0.05), increased mRNA expression levels of CLDN and ZO-1 (p < 0.05), and enhanced epithelial proliferation (p < 0.05) in the ileal mucosa in LPS-challenged chickens. Moreover, FA supplementation rectified the ileal microflora disturbances in the LPS-challenged broilers. The results demonstrate that dietary FA supplementation decreased LPS-induced intestinal damage by enhancing antioxidant capacity and maintaining intestinal integrity. Furthermore, FA supplementation protects intestinal tight junctions (TJs), elevates secretory immunoglobulin A (sIgA) levels, and modulates ileal microflora composition in LPS-challenged broilers.
Oral Sampling of Little Brown Bat (Myotis lucifugus) Maternity Colonies for SARS-CoV-2 in the Northeast and Mid-Atlantic, USA
Moran, Megan L.; Boyd, William; De La Cruz, Jesse L.; Bertke, Andrea S.; Ford, W. Mark (MDPI, 2023-02-04)
The potential introduction of SARS-CoV-2, the virus responsible for the COVID-19 pandemic, into North American bat populations is of interest to wildlife managers due to recent disease-mediated declines of several species. Populations of little brown bats (Myotis lucifugus) have collapsed due to white-nose syndrome (WNS), a disease caused by the introduction and spread of the fungal pathogen Pseudogymnoascus destructans (Pd). Throughout much of the United States and southern Canada, large colonies of the species routinely established diurnal roosts in anthropogenic structures, thereby creating the potential for direct human contact and cross-species disease transmission. Given recent declines and the potential for further disease impacts, we collected oral swabs from eight little brown bat maternity colonies to assess the presence and prevalence of SARS-CoV-2 by RT-qPCR analysis. Little brown bat colonies in Maryland (n = 1), New Hampshire (n = 1), New Jersey (n = 2), New York (n = 1), Rhode Island (n = 2), and Virginia (n = 1) were taken during May-August, 2022. From 235 assayed individuals, no bat tested positive for SARS-CoV-2. Our results indicate that little brown bats may not contract SARS-CoV-2 or that the virus persists at undetectable levels in populations of the Mid-Atlantic and Northeast during summer months. Nonetheless, continued monitoring and future work addressing other seasons may still be warranted to conclusively determine infection status.
Herpes Simplex Virus 1 (HSV-1) Infected Cell Protein 0 (ICP0) Targets of Ubiquitination during Productive Infection of Primary Adult Sensory Neurons
Harrell, Telvin L.; Davido, David J.; Bertke, Andrea S. (MDPI, 2023-02-02)
Herpes simplex virus 1 (HSV-1) enters sensory neurons with the potential for productive or latent infection. For either outcome, HSV-1 must curtail the intrinsic immune response, regulate viral gene expression, and remove host proteins that could restrict viral processes. Infected cell protein 0 (ICP0), a virus-encoded E3 ubiquitin ligase, supports these processes by mediating the transfer of ubiquitin to target proteins to change their location, alter their function, or induce their degradation. To identify ubiquitination targets of ICP0 during productive infection in sensory neurons, we immunoprecipitated ubiquitinated proteins from primary adult sensory neurons infected with HSV-1 KOS (wild-type), HSV-1 n212 (expressing truncated, defective ICP0), and uninfected controls using anti-ubiquitin antibody FK2 (recognizing K29, K48, K63 and monoubiquitinated proteins), followed by LC-MS/MS and comparative analyses. We identified 40 unique proteins ubiquitinated by ICP0 and 17 ubiquitinated by both ICP0 and host mechanisms, of which High Mobility Group Protein I/Y (HMG I/Y) and TAR DNA Binding Protein 43 (TDP43) were selected for further analysis. We show that ICP0 ubiquitinates HMG I/Y and TDP43, altering protein expression at specific time points during productive HSV-1 infection, demonstrating that ICP0 manipulates the sensory neuronal environment in a time-dependent manner to regulate infection outcome in neurons.
De Novo Assembly and Annotation of 11 Diverse Shrub Willow (Salix) Genomes Reveals Novel Gene Organization in Sex-Linked Regions
Hyden, Brennan; Feng, Kai; Yates, Timothy B.; Jawdy, Sara; Cereghino, Chelsea; Smart, Lawrence B.; Muchero, Wellington (MDPI, 2023-02-02)
Poplar and willow species in the Salicaceae are dioecious, yet have been shown to use different sex determination systems located on different chromosomes. Willows in the subgenus Vetrix are interesting for comparative studies of sex determination systems, yet genomic resources for these species are still quite limited. Only a few annotated reference genome assemblies are available, despite many species in use in breeding programs. Here we present de novo assemblies and annotations of 11 shrub willow genomes from six species. Copy number variation of candidate sex determination genes within each genome was characterized and revealed remarkable differences in putative master regulator gene duplication and deletion. We also analyzed copy number and expression of candidate genes involved in floral secondary metabolism, and identified substantial variation across genotypes, which can be used for parental selection in breeding programs. Lastly, we report on a genotype that produces only female descendants and identified gene presence/absence variation in the mitochondrial genome that may be responsible for this unusual inheritance.
Characterizing the Ablative Effects of Histotripsy for Osteosarcoma: In Vivo Study in Dogs
Ruger, Lauren N.; Hay, Alayna N.; Vickers, Elliana R.; Coutermarsh-Ott, Sheryl; Gannon, Jessica M.; Covell, Hannah S.; Daniel, Gregory B.; Laeseke, Paul F.; Ziemlewicz, Timothy J.; Kierski, Katharine R.; Ciepluch, Brittany J.; Vlaisavljevich, Eli; Tuohy, Joanne L. (MDPI, 2023-01-25)
Osteosarcoma (OS) is a malignant bone tumor treated by limb amputation or limb salvage surgeries and chemotherapy. Histotripsy is a non-thermal, non-invasive focused ultrasound therapy using controlled acoustic cavitation to mechanically disintegrate tissue. Recent ex vivo and in vivo pilot studies have demonstrated the ability of histotripsy for ablating OS but were limited in scope. This study expands on these initial findings to more fully characterize the effects of histotripsy for bone tumors, particularly in tumors with different compositions. A prototype 500 kHz histotripsy system was used to treat ten dogs with suspected OS at an intermediate treatment dose of 1000 pulses per location. One day after histotripsy, treated tumors were resected via limb amputation, and radiologic and histopathologic analyses were conducted to determine the effects of histotripsy for each patient. The results of this study demonstrated that histotripsy ablation is safe and feasible in canine patients with spontaneous OS, while offering new insights into the characteristics of the achieved ablation zone. More extensive tissue destruction was observed after histotripsy compared to that in previous reports, and radiographic changes in tumor size and contrast uptake following histotripsy were reported for the first time. Overall, this study significantly expands our understanding of histotripsy bone tumor ablation and informs future studies for this application.
Stress Hormones Epinephrine and Corticosterone Selectively Reactivate HSV-1 and HSV-2 in Sympathetic and Sensory Neurons
Goswami, Poorna; Ives, Angela M.; Abbott, Amber R. N.; Bertke, Andrea S. (MDPI, 2022-05-23)
Herpes simplex viruses 1 and 2 (HSV-1 and HSV-2) establish latency in sensory and autonomic neurons, from which they can reactivate to cause recurrent disease throughout the life of the host. Stress is strongly associated with HSV recurrences in humans and animal models. However, the mechanisms through which stress hormones act on the latent virus to cause reactivation are unknown. We show that the stress hormones epinephrine (EPI) and corticosterone (CORT) induce HSV-1 reactivation selectively in sympathetic neurons, but not sensory or parasympathetic neurons. Activation of multiple adrenergic receptors is necessary for EPI-induced HSV-1 reactivation, while CORT requires the glucocorticoid receptor. In contrast, CORT, but not EPI, induces HSV-2 reactivation in both sensory and sympathetic neurons through either glucocorticoid or mineralocorticoid receptors. Reactivation is dependent on different transcription factors for EPI and CORT, and coincides with rapid changes in viral gene expression, although genes differ for HSV-1 and HSV-2, and temporal kinetics differ for EPI and CORT. Thus, stress-induced reactivation mechanisms are neuron-specific, stimulus-specific and virus-specific. These findings have implications for differences in HSV-1 and HSV-2 recurrent disease patterns and frequencies, as well as development of targeted, more effective antivirals that may act on different responses in different types of neurons.
Engrafting Horse Immune Cells into Mouse Hosts for the Study of the Acute Equine Immune Responses
Leeth, Caroline M.; Adkins, Janie; Hay, Alayna N.; Bogers, Sophie Helen; Potter, Ashley; Witonsky, Sharon G.; Zhu, Jing (MDPI, 2021-10-14)
Immunological studies in the horse are frequently hampered by lack of environmental control, complicated study design, and ethical concerns when performing high risk studies. The purpose of the current study was to investigate the utility of a xenograft model for studying acute equine immune responses. Immunocompromised non obese diabetic (NOD). sudden combined immunodeficiency (scid).gamma-/- (NSG) mice were engrafted with either equine peripheral blood lymphocytes (PBLs) or equine bone marrow to determine an optimal protocol for equine lymphocyte engraftment. We found that both PBL and bone marrow grafts populated the host mice successfully. Bone marrow transplants were technically more challenging and required further processing to retard graft versus host disease. Graft vs host disease was apparent at 28 days post-PBL transfer and 56 days post-bone marrow transfer. The results of these studies support the use of mouse hosts to study acute equine immune responses and that different engraftment techniques can be used depending on the experimental design.
Enemy of My Enemy: A Novel Insect-Specific Flavivirus Offers a Promising Platform for a Zika Virus Vaccine
Porier, Danielle L.; Wilson, Sarah N.; Auguste, Dawn I.; Leber, Andrew; Coutermarsh-Ott, Sheryl; Allen, Irving C.; Caswell, Clayton C.; Budnick, James A.; Bassaganya-Riera, Josep; Hontecillas, Raquel; Weger-Lucarelli, James; Weaver, Scott C.; Auguste, Albert J. (MDPI, 2021-10-07)
Vaccination remains critical for viral disease outbreak prevention and control, but conventional vaccine development typically involves trade-offs between safety and immunogenicity. We used a recently discovered insect-specific flavivirus as a vector in order to develop an exceptionally safe, flavivirus vaccine candidate with single-dose efficacy. To evaluate the safety and efficacy of this platform, we created a chimeric Zika virus (ZIKV) vaccine candidate, designated Aripo/Zika virus (ARPV/ZIKV). ZIKV has caused immense economic and public health impacts throughout the Americas and remains a significant public health threat. ARPV/ZIKV vaccination showed exceptional safety due to ARPV/ZIKV’s inherent vertebrate host-restriction. ARPV/ZIKV showed no evidence of replication or translation in vitro and showed no hematological, histological or pathogenic effects in vivo. A single-dose immunization with ARPV/ZIKV induced rapid and robust neutralizing antibody and cellular responses, which offered complete protection against ZIKV-induced morbidity, mortality and in utero transmission in immune-competent and -compromised murine models. Splenocytes derived from vaccinated mice demonstrated significant CD4⁺ and CD8⁺ responses and significant cytokine production post-antigen exposure. Altogether, our results further support that chimeric insect-specific flaviviruses are a promising strategy to restrict flavivirus emergence via vaccine development.
Essential Gene(s) Targeted by Peptide Nucleic Acids Kills Mycobacterium smegmatis in Culture and in Infected Macrophages
Islam, Md. Ariful; Khatun, Mst. Minara; Sriranganathan, Nammalwar; Boyle, Stephen M. (Scientific Research Publishing, 2021-04-19)
Background: Antisense peptide nucleic acids (PNAs) exhibit growth inhibitory effects on bacteria by inhibiting the expression of essential genes and could be promising therapeutic agents for treating bacterial infections. A study was carried out to determine the efficacy of several antisense PNAs in inhibiting extracellular and intracellular growth of Mycobacterium smegmatis. Methods : Six PNAs obtained from a commercial supplier were tested to evaluate the inhibitory effect on bacterial growth by inhibiting the expression of the following essential genes: inhA (a fatty acid elongase), rpsL (ribosomal S12 protein), gyrA (DNA gyrase), pncA (pyrazinamidase), polA (DNA polymerase I) and rpoC (RNA polymerase β subunit) of M. smegmatis . Each PNA was tested at 20 μM, 10 μM, 5 μM and 2.5 μM concentrations to determine whether they caused a dose dependent killing of M. smegmatis cultured in Middlebrook 7H9 broth or in a J774A.1 murine macrophage cell line. Results : In Middlebrook broth, the strong growth inhibitory effect against M. smegmatis was observed by PNAs targeting the inhA and rpsL genes at all four concentrations. The PNAs targeting the pncA, polA and rpoC genes were found to exhibit strong growth inhibition against M. smegmatis but only at 20 μM concentration. No growth inhibition of M. smegmatis was seen in pure culture when treated with PNAs targeting gyrA and a mismatch PNA targeting dnaG (DNA primase). All six PNAs showed killing of M. smegmatis in J774A.1 macrophage cell line that were statistically significant (p < 0.05). Conclusion: It may be concluded from this study that PNAs could be potential therapeutics for mycobacterial infections.
Survival of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and Herpes Simplex Virus 1 (HSV-1) on Foods Stored at Refrigerated Temperature
Dhakal, Janak; Jia, Mo; Joyce, Jonathan D.; Moore, Greyson A.; Ovissipour, Mahmoudreza; Bertke, Andrea S. (MDPI, 2021-05-04)
Outbreaks of coronavirus infectious disease 2019 (COVID-19) in meat processing plants and media reports of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection on foods have raised concerns of a public health risk from contaminated foods. We used herpes simplex virus 1, a non-Biosafety Level 3 (non-BSL3) enveloped virus, as a surrogate to develop and validate methods before assessing the survival of infectious SARS-CoV-2 on foods. Several food types, including chicken, seafood, and produce, were held at 4 °C and assessed for infectious virus survival (herpes simplex virus 1 (HSV-1) and SARS-CoV-2) at 0 h, 1 h, and 24 h post-inoculation (hpi) by plaque assay. At all three time points, recovery of SARS-CoV-2 was similar from chicken, salmon, shrimp, and spinach, ranging from 3.4 to 4.3 log PFU/mL. However, initial (0 h) virus recovery from apples and mushrooms was significantly lower than that from poultry and seafood, and infectious virus decreased over time, with recovery from mushrooms becoming undetectable by 24 hpi. Comparing infectious virus titers with viral genome copies confirmed that PCR-based tests only indicate presence of viral nucleic acid, which does not necessarily correlate with the quantity of infectious virus. The survival and high recovery of SARS-CoV-2 on certain foods highlight the importance of safe food handling practices in mitigating any public health concerns related to potentially contaminated foods.
Escherichia coli-associated granulomatous colitis in a cat
Safian, Alexander M.; Bolton, Timothy (John Wiley & Sons Ltd., 2021)
A 6-year-old male castrated domestic shorthair cat was evaluated for a 6-month history of haematochezia, mucoid diarrhoea, tenesmus and rectal prolapse. Colonic histopathology revealed multifocal mucosal ulceration and lamina propria infiltration with large numbers of periodic acid-Schiff-positivemacrophages. Large clusters of intracellular Escherichia coli were confirmed with fluorescence in situ hybridization testing, similar to that seen in dogs with granulomatous colitis. An 8-week course of marbofloxacin resulted in resolution of clinical signs; however, recurrence occurred 4 weeks later. A 12-week course of marbofloxacin resulted in disease remission for which the cat still remains free of clinical signs (15 months). Escherichia coli-associated granulomatous colitis, although reported with rarity in this species, is an important infectious cause of chronic large intestinal disease in the cat.
On the Fly: Interactions Between Birds, Mosquitoes, and Environment That Have Molded West Nile Virus Genomic Structure Over Two Decades
Duggal, Nisha K.; Langwig, Kate E.; Ebel, Gregory D.; Brault, Aaron C. (Oxford University Pres, 2019-09-24)
West Nile virus (WNV) was first identified in North America almost 20 yr ago. In that time, WNV has crossed the continent and established enzootic transmission cycles, resulting in intermittent outbreaks of human disease that have largely been linked with climatic variables and waning avian seroprevalence. During the transcontinental dissemination of WNV, the original genotype has been displaced by two principal extant genotypes which contain an envelope mutation that has been associated with enhanced vector competence by Culex pipiens L. (Diptera: Culicidae) and Culex tarsalis Coquillett vectors. Analyses of retrospective avian host competence data generated using the founding NY99 genotype strain have demonstrated a steady reduction in viremias of house sparrows over time. Reciprocally, the current genotype strains WN02 and SW03 have demonstrated an inverse correlation between house sparrow viremia magnitude and the time since isolation. These data collectively indicate that WNV has evolved for increased avian viremia while house sparrows have evolved resistance to the virus such that the relative host competence has remained constant. Intrahost analyses of WNV evolution demonstrate that selection pressures are avian species-specific and purifying selection is greater in individual birds compared with individual mosquitoes, suggesting that the avian adaptive and/or innate immune response may impose a selection pressure on WNV. Phylogenomic, experimental evolutionary systems, and models that link viral evolution with climate, host, and vector competence studies will be needed to identify the relative effect of different selective and stochastic mechanisms on viral phenotypes and the capacity of newly evolved WNV genotypes for transmission in continuously changing landscapes.
Duration of seminal Zika viral RNA shedding in immunocompetent mice inoculated with Asian and African genotype viruses
McDonald, Erin M.; Duggal, Nisha K.; Delorey, Mark J.; Oksanish, James; Ritter, Jana M.; Brault, Aaron C. (Elsevier, 2019-06-20)
Prior to the emergence of Asian genotype Zika virus (ZIKV) in the Western hemisphere, sexual transmission in humans was documented. Sexual transmission by African genotype ZIKVs has not been assessed in laboratory animal models, due to rapid and high mortality rates of immunodeficient mice following inoculation. To overcome these limitations, immunocompetent C57Bl/6 mice were used to longitudinally assess Asian and African genotype ZIKV sexual transmission potential. Furthermore, to determine if enhanced pathogenesis of African genotype ZIKVs is due to structural determinants, PRVABC59 prM/E was replaced with African MR766 prM/E (chimeric ZIKV). The African genotype and chimeric ZIKV elicited greater pathogenic effects in the male reproductive tract and generated higher viremias. Yet, the duration, magnitude and efficiency of seminal shedding of infectious virus and viral RNA were similar between chimeric-, African and Asian genotype ZIKVinoculated mice. These data show that increased male reproductive tract pathology does not increase sexual transmission potential.
A 7-week-old male Golden Retriever with extreme leukocytosis: A case report
Bingham, Emily; Conner, Bobbi; Stern, Jere; Vitalo, Amber; Schaer, Michael (Wiley, 2020-09-27)
Although neoplasia should be a top concern for extreme leukocytosis in dogs, infectious causes must also be considered to avoid delays in treatment or undue recommendations for humane euthanasia. Blood film review is of paramount importance.
Incidence of Health and Behavior Problems in Service Dog Candidates Neutered at Various Ages
Zlotnick, Marta G.; Corrigan, Virginia Kiefer; Griffin, Erin; Alayon, Megan; Hungerford, Laura L. (2019-10-08)
Saint Francis Service Dogs (SFSD) trains dogs to aid people with multiple sclerosis, brain injury, and many other conditions. Organizations like SFSD must carefully consider when to neuter dogs to give them the best chance at successfully completing lengthy and expensive training. The objective of this retrospective cohort study was to assess differences in the incidence of health or behavior problems leading to dismissal between dogs neutered at different ages. Data on 245 dogs-including birth date, sex, neuter date, dismissal or successful completion of training, and (where applicable) reason for dismissal-were collected from SFSD records. Age-at-neuter was grouped (<7 months; 7-11 months; >11 months) and compared for dogs who successfully completed training and dogs who were dismissed. Dogs neutered from 7 to 11 months of age were dismissed at a significantly lower overall rate than dogs neutered at an older or younger age. There were no differences between males and females. Labrador and golden retrievers were less likely to be dismissed than other breeds. This pattern was the same for dismissals for behavioral reasons. Dogs neutered at <7 months had more than twice the risk for health-related dismissals as dogs neutered at any older age and this pattern held for orthopedic dismissals. Labradors were at higher risk for orthopedic-related dismissal than golden retrievers and all other breeds. This study suggests that there is a relationship between dogs' age at neuter and the incidence of health and behavioral problems that can lead to dismissal from service dog training.
Endemicity of Yaws and Seroprevalence of Treponema pallidum Antibodies in Nonhuman Primates, Kenya
Zimmerman, Dawn M.; Hardgrove, Emily H.; von Fricken, Michael E.; Kamau, Joseph; Chai, Daniel; Mutura, Samson; Kivali, Velma; Hussein, Fatima; Ambala, Peris; Surmat, Andrea; Maina, Joseph G.; Knauf, Sascha (2019-11)
Human yaws has historically been endemic to Kenya, but current epidemiologic data are lacking. We report seroprevalence for Treponema pallidum antibodies in olive baboons (Papio anubis) and vervet monkeys (Chlorocebus pygerythrus) in Laikipia County, Kenya. Our results suggest endemicity of the yaws bacterium in monkeys, posing a possible zoonotic threat to humans.
Studies Exploring the Interaction of the Organophosphorus Compound Paraoxon with Fullerenes
Magnin, Geraldine; Bissel, Philippe; Council-Troche, Roberto McAlister; Zhou, Zhiguo; Ehrich, Marion F. (2019-11-12)
In vitro experiments previously published demonstrated the ability of fullerenes to decrease the capability of organophosphorus (OP) compounds to inhibit acetylcholinesterase. Experiments described herein demonstrate molecular level affinity interactions between fullerenes and the OP test compound paraoxon with NMR spectroscopy. The calculated binding constant of 19 M-1 indicates that this binding was not covalent.
Pulmonary Exposure to Magnéli Phase Titanium Suboxides Results in Significant Macrophage Abnormalities and Decreased Lung Function
McDaniel, Dylan K.; Ringel-Scaia, Veronica M.; Morrison, Holly A.; Coutermarsh-Ott, Sheryl; Council-Troche, McAlister; Angle, Jonathan W.; Perry, Justin B.; Davis, Grace; Leng, Weinan; Minarchick, Valerie; Yang, Yi; Chen, Bo; Reece, Sky W.; Brown, David A.; Cecere, Thomas E.; Brown, Jared M.; Gowdy, Kymberly M.; Hochella, Michael F. Jr.; Allen, Irving C. (Frontiers, 2019-11-28)
Coal is one of the most abundant and economic sources for global energy production. However, the burning of coal is widely recognized as a significant contributor to atmospheric particulate matter linked to deleterious respiratory impacts. Recently, we have discovered that burning coal generates large quantities of otherwise rare Magnéli phase titanium suboxides from TiO2 minerals naturally present in coal. These nanoscale Magnéli phases are biologically active without photostimulation and toxic to airway epithelial cells in vitro and to zebrafish in vivo. Here, we sought to determine the clinical and physiological impact of pulmonary exposure to Magnéli phases using mice as mammalian model organisms. Mice were exposed to the most frequently found Magnéli phases, Ti6O11, at 100 parts per million (ppm) via intratracheal administration. Local and systemic titanium concentrations, lung pathology, and changes in airway mechanics were assessed. Additional mechanistic studies were conducted with primary bone marrow derived macrophages. Our results indicate that macrophages are the cell type most impacted by exposure to these nanoscale particles. Following phagocytosis, macrophages fail to properly eliminate Magnéli phases, resulting in increased oxidative stress, mitochondrial dysfunction, and ultimately apoptosis. In the lungs, these nanoparticles become concentrated in macrophages, resulting in a feedback loop of reactive oxygen species production, cell death, and the initiation of gene expression profiles consistent with lung injury within 6 weeks of exposure. Chronic exposure and accumulation of Magnéli phases ultimately results in significantly reduced lung function impacting airway resistance, compliance, and elastance. Together, these studies demonstrate that Magnéli phases are toxic in the mammalian airway and are likely a significant nanoscale environmental pollutant, especially in geographic regions where coal combustion is a major contributor to atmospheric particulate matter.
The Komodo dragon (Varanus komodoensis) genome and identification of innate immunity genes and clusters
van Hoek, Monique L.; Prickett, M. D.; Settlage, Robert E.; Kang, Lin; Michalak, Pawel; Vliet, Kent A.; Bishop, Barney M. (2019-08-30)
Background We report the sequencing, assembly and analysis of the genome of the Komodo dragon (Varanus komodoensis), the largest extant lizard, with a focus on antimicrobial host-defense peptides. The Komodo dragon diet includes carrion, and a complex milieu of bacteria, including potentially pathogenic strains, has been detected in the saliva of wild dragons. They appear to be unaffected, suggesting that dragons have robust defenses against infection. While little information is available regarding the molecular biology of reptile immunity, it is believed that innate immunity, which employs antimicrobial host-defense peptides including defensins and cathelicidins, plays a more prominent role in reptile immunity than it does in mammals. . Results High molecular weight genomic DNA was extracted from Komodo dragon blood cells. Subsequent sequencing and assembly of the genome from the collected DNA yielded a genome size of 1.6 Gb with 45x coverage, and the identification of 17,213 predicted genes. Through further analyses of the genome, we identified genes and gene-clusters corresponding to antimicrobial host-defense peptide genes. Multiple β-defensin-related gene clusters were identified, as well as a cluster of potential Komodo dragon ovodefensin genes located in close proximity to a cluster of Komodo dragon β-defensin genes. In addition to these defensins, multiple cathelicidin-like genes were also identified in the genome. Overall, 66 β-defensin genes, six ovodefensin genes and three cathelicidin genes were identified in the Komodo dragon genome. Conclusions Genes with important roles in host-defense and innate immunity were identified in this newly sequenced Komodo dragon genome, suggesting that these organisms have a robust innate immune system. Specifically, multiple Komodo antimicrobial peptide genes were identified. Importantly, many of the antimicrobial peptide genes were found in gene clusters. We found that these innate immunity genes are conserved among reptiles, and the organization is similar to that seen in other avian and reptilian species. Having the genome of this important squamate will allow researchers to learn more about reptilian gene families and will be a valuable resource for researchers studying the evolution and biology of the endangered Komodo dragon.

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