Extraction, Purification, and Next-Generation Sequencing (NGS) Analysis of DNA and RNA from Formalin-Fixed and Paraffin-Embedded (FFPE) Tissue.

Formalin fixed paraffin embedded (FFPE) tissues have long been used for immunohistological analyses. FFPE tissues can be stored at room temperature for several years enabling analyses to be performed later. Ease of storage and transport makes these tissues an attractive source of biological material.

Use of high throughput DNA analysis to characterize the nodule-associated bacterial community from four ages of trees in a Costa Rican cloud forest.

Leguminous tree root nodule nitrogen-fixing bacteria are critical for recuperation of soil C and N cycle processes after disturbance in tropical forests, while other nodule-associated bacteria (NAB) may enhance nodule development and activity, and plant growth. However, little is known of these root nodule microbiomes. Through DNA analysis, we evaluated the bacterial taxa associated with the root nodules of the 1-year-old, 2-year-old, 13-year-old, and old growth trees in a cloud forest.

Time-Course Progression of Whole Transcriptome Expression Changes of Trigeminal Ganglia Compared to Dorsal Root Ganglia in Rats Exposed to Nerve Injury.

Mechanisms underlying neuropathic pain (NP) are complex with multiple genes, their interactions, environmental and epigenetic factors being implicated. Transcriptional changes in the trigeminal (TG) and dorsal root (DRG) ganglia have been implicated in the development and maintenance of NP. Despite efforts to unravel molecular mechanisms of NP, many remain unknown.

Cutting Edge: Neutrophils License the Maturation of Monocytes into Effective Antifungal Effectors.

Neutrophils are critical for the direct eradication of conidia, but whether they mediate antifungal defense beyond their role as effectors is unclear. In this study, we demonstrate that neutrophil depletion impairs the activation of protective antifungal CCR2 inflammatory monocytes. In the absence of neutrophils, monocytes displayed limited differentiation into monocyte-derived dendritic cells, reduced formation of reactive oxygen species, and diminished conidiacidal activity.

Comprehensive Analysis of Disease Pathology in Immunocompetent and Immunocompromised Hosts following Pulmonary SARS-CoV-2 Infection.

The Coronavirus disease 2019 (COVID-19) pandemic disproportionately affects immunocompetent and immunocompromised individuals, with the latter group being more vulnerable to severe disease and death. However, the differential pathogenesis of SARS-CoV-2 in the context of a specific immunological niche remains unknown. Similarly, systematic analysis of disease pathology in various extrapulmonary organs in immunocompetent and immunocompromised hosts during SARS-CoV-2 infection is not fully understood.

Biomechanical Forces Regulate Gene Transcription During Stretch-Mediated Growth of Mammalian Neurons.

At birth, there are 100 billion neurons in the human brain, with functional neural circuits extending through the spine to the epidermis of the feet and toes. Following birth, limbs and vertebrae continue to grow by several orders of magnitude, forcing established axons to grow by up to 200 cm in length without motile growth cones. The leading regulatory paradigm suggests that biomechanical expansion of mitotic tissue exerts tensile force on integrated nervous tissue, which synchronizes ongoing growth of spanning axons.

Analysis of 1,25-Dihydroxyvitamin D Genomic Action Reveals Calcium-Regulating and Calcium-Independent Effects in Mouse Intestine and Human Enteroids.

Although vitamin D is critical for the function of the intestine, most studies have focused on the duodenum. We show that transgenic expression of the vitamin D receptor (VDR) only in the distal intestine of VDR null mice (KO/TG mice) results in the normalization of serum calcium and rescue of rickets. Although it had been suggested that calcium transport in the distal intestine involves a paracellular process, we found that the 1,25-dihydroxyvitamin D [1,25(OH)D]-activated genes in the proximal intestine associated with active calcium transport (, , and ) are also induced by 1,25(OH)D in the distal intestine of KO/TG mice.

Mycobacterium tuberculosis progresses through two phases of latent infection in humans.

Little is known about the physiology of latent Mycobacterium tuberculosis infection. We studied the mutational rates of 24 index tuberculosis (TB) cases and their latently infected household contacts who developed active TB up to 5.25 years later, as an indication of bacterial physiological state and possible generation times during latent TB infection in humans.

The Role of Immunological Synapse in Predicting the Efficacy of Chimeric Antigen Receptor (CAR) Immunotherapy.

Chimeric Antigen Receptor (CAR) immunotherapy utilizes genetically-engineered immune cells that express a unique cell surface receptor that combines tumor antigen specificity with immune cell activation. In recent clinical trials, the adoptive transfer of CAR-modified immune cells (including CAR-T and CAR-NK cells) into patients has been remarkably successful in treating multiple refractory blood cancers. To improve safety and efficacy, and expand potential applicability to other cancer types, CARs with different target specificities and sequence modifications are being developed and tested by many laboratories.

Rapidly Correcting Frameshift Mutations in the Mycobacterium tuberculosis Gene Produce Reversible Ethambutol Resistance and Small-Colony-Variant Morphology.

We have identified a previously unknown mechanism of reversible high-level ethambutol (EMB) resistance in that is caused by a reversible frameshift mutation in the gene. A frameshift mutation in produces the small-colony-variant (SCV) phenotype, but this mutation does not change the MICs of any drug for wild-type However, the same mutation in a low-level EMB-resistant double mutant (MIC = 8 μg/ml) produces an SCV with an EMB MIC of 32 μg/ml. Reversible resistance is indistinguishable from a drug-persistent phenotype, because further culture of these SCV mutants results in rapid reversion of the frameshifts, reestablishing the correct open reading frame, returning the culture to normal colony size, and reversing the EMB MIC back to that (8 μg/ml) of the parental strain.

Trichinella spiralis-induced mastocytosis and erythropoiesis are simultaneously supported by a bipotent mast cell/erythrocyte precursor cell.

Anti-helminth responses require robust type 2 cytokine production that simultaneously promotes worm expulsion and initiates the resolution of helminth-induced wounds and hemorrhaging. However, how infection-induced changes in hematopoiesis contribute to these seemingly distinct processes remains unknown. Recent studies have suggested the existence of a hematopoietic progenitor with dual mast cell-erythrocyte potential.

Differential gene expression changes in the dorsal root versus trigeminal ganglia following peripheral nerve injury in rats.

Background: The dorsal root (DRG) and trigeminal (TG) ganglia contain cell bodies of sensory neurons of spinal and trigeminal systems, respectively. They are homologs of each other; however, differences in how the two systems respond to injury exist. Trigeminal nerve injuries rarely result in chronic neuropathic pain (NP).

A Mechanosensitive Channel Governs Lipid Flippase-Mediated Echinocandin Resistance in Cryptococcus neoformans.

Echinocandins show fungicidal activity against common invasive mycoses but are ineffective against cryptococcosis. The underlying mechanism for echinocandin resistance in remains poorly understood but has been shown to involve Cdc50, the regulatory subunit of lipid flippase. In a forward genetic screen for Δ suppressor mutations that are caspofungin resistant, we identified Crm1 (aspofungin esistant utation ), a homolog of mechanosensitive channel proteins, and showed that Δ restored caspofungin resistance in Δ cells.

Antitubercular Triazines: Optimization and Intrabacterial Metabolism.

The triazine antitubercular JSF-2019 was of interest due to its in vitro efficacy and the nitro group shared with the clinically relevant delamanid and pretomanid. JSF-2019 undergoes activation requiring FH and one or more nitroreductases in addition to Ddn. An intrabacterial drug metabolism (IBDM) platform was leveraged to demonstrate the system kinetics, evidencing formation of NO and a des-nitro metabolite.

Phase variation in produces transiently heritable drug tolerance.

The length and complexity of tuberculosis (TB) therapy, as well as the propensity of to develop drug resistance, are major barriers to global TB control efforts. is known to have the ability to enter into a drug-tolerant state, which may explain many of these impediments to TB treatment. We have identified a mechanism of genetically encoded but rapidly reversible drug tolerance in caused by transient frameshift mutations in a homopolymeric tract (HT) of 7 cytosines (7C) in the gene.

Entry of the bat influenza H17N10 virus into mammalian cells is enabled by the MHC class II HLA-DR receptor.

Haemagglutinin and neuraminidase surface glycoproteins of the bat influenza H17N10 virus neither bind to nor cleave sialic acid receptors, indicating that this virus employs cell entry mechanisms distinct from those of classical influenza A viruses. We observed that certain human haematopoietic cancer cell lines and canine MDCK II cells are susceptible to H17-pseudotyped viruses. We identified the human HLA-DR receptor as an entry mediator for H17 pseudotypes, suggesting that H17N10 possesses zoonotic potential.

Publisher Correction: Toxoplasma Modulates Signature Pathways of Human Epilepsy, Neurodegeneration & Cancer.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Paraspinous muscle gene expression profiling following simulated staged endovascular repair of thoracoabdominal aortic aneurysm: exploring potential therapeutic pathways.

Objectives: Thoracic endovascular techniques for aneurysm repair offer less invasive alternatives to open strategies. Both approaches, however, are associated with the risk for neurological complications. Despite adjuncts to maintain spinal cord perfusion, ischaemia and paraplegia continue to occur during thoracoabdominal aortic aneurysm (TAAA) repair.

Synergistic Lethality of a Binary Inhibitor of Mycobacterium tuberculosis KasA.

We report GSK3011724A (DG167) as a binary inhibitor of β-ketoacyl-ACP synthase (KasA) in Genetic and biochemical studies established KasA as the primary target. The X-ray crystal structure of the KasA-DG167 complex refined to 2.0-Å resolution revealed two interacting DG167 molecules occupying nonidentical sites in the substrate-binding channel of KasA.

Differential gene expression in trigeminal ganglia of male and female rats following chronic constriction of the infraorbital nerve.

Background: The mechanisms underlying sex-based differences in pain and analgesia are poorly understood. In this study, we investigated gene expression changes in trigeminal ganglia (TG) of male and female rats exposed to infraorbital nerve chronic constriction injury (IoN-CCI).

Methods: Somatosensory assessments were performed prior to IoN-CCI and at selected time points postsurgery.

The human GCOM1 complex gene interacts with the NMDA receptor and internexin-alpha.

The known functions of the human GCOM1 complex hub gene include transcription elongation and the intercalated disk of cardiac myocytes. However, in all likelihood, the gene's most interesting, and thus far least understood, roles will be found in the central nervous system. To investigate the functions of the GCOM1 gene in the CNS, we have cloned human and rat brain cDNAs encoding novel, 105 kDa GCOM1 combined (Gcom) proteins, designated Gcom15, and identified a new group of GCOM1 interacting genes, termed Gints, from yeast two-hybrid (Y2H) screens.

Toxoplasma Modulates Signature Pathways of Human Epilepsy, Neurodegeneration & Cancer.

One third of humans are infected lifelong with the brain-dwelling, protozoan parasite, Toxoplasma gondii. Approximately fifteen million of these have congenital toxoplasmosis. Although neurobehavioral disease is associated with seropositivity, causality is unproven.

A Novel Small-Molecule Inhibitor of the Demethylmenaquinone Methyltransferase MenG Is Bactericidal to Both Growing and Nutritionally Deprived Persister Cells.

Active tuberculosis (TB) and latent infection both require lengthy treatments to achieve durable cures. This problem has partly been attributable to the existence of nonreplicating "persisters" that are difficult to kill using conventional anti-TB treatments. Compounds that target the respiratory pathway have the potential to kill both replicating and persistent and shorten TB treatment, as this pathway is essential in both metabolic states.