Protein kinases PknA and PknB independently and coordinately regulate essential Mycobacterium tuberculosis physiologies and antimicrobial susceptibility.

The Mycobacterium tuberculosis Ser/Thr protein kinases PknA and PknB are essential for growth and have been proposed as possible drug targets. We used a titratable conditional depletion system to investigate the functions of these kinases. Depletion of PknA or PknB or both kinases resulted in growth arrest, shortening of cells, and time-dependent loss of acid-fast staining with a concomitant decrease in mycolate synthesis and accumulation of trehalose monomycolate.

Licensed Bacille Calmette-Guérin (BCG) formulations differ markedly in bacterial viability, RNA content and innate immune activation.

Background: Bacille Calmette-Guérin (BCG), the live attenuated tuberculosis vaccine, is manufactured under different conditions across the globe generating formulations that may differ in clinical efficacy. Innate immune recognition of live BCG contributes to immunogenicity suggesting that differences in BCG viability may contribute to divergent activity of licensed formulations.

Methods: We compared BCG-Denmark (DEN), -Japan (JPN), -India (IND), -Bulgaria (BUL) and -USA in vitro with respect to a) viability as measured by colony-forming units (CFU), mycobacterial membrane integrity, and RNA content, and b) cytokine/chemokine production in newborn cord and adult peripheral blood.

Multisystem Analysis of Reveals Kinase-Dependent Remodeling of the Pathogen-Environment Interface.

Tuberculosis is the leading killer among infectious diseases worldwide. Increasing multidrug resistance has prompted new approaches for tuberculosis drug development, including targeted inhibition of virulence determinants and of signaling cascades that control many downstream pathways. We used a multisystem approach to determine the effects of a potent small-molecule inhibitor of the essential Ser/Thr protein kinases PknA and PknB.

Review of Experimental Compounds Demonstrating Anti-Toxoplasma Activity.

Toxoplasma gondii is a ubiquitous apicomplexan parasite capable of infecting humans and other animals. Current treatment options for T. gondii infection are limited and most have drawbacks, including high toxicity and low tolerability.

Investigating essential gene function in Mycobacterium tuberculosis using an efficient CRISPR interference system.

Despite many methodological advances that have facilitated investigation of Mycobacterium tuberculosis pathogenesis, analysis of essential gene function in this slow-growing pathogen remains difficult. Here, we describe an optimized CRISPR-based method to inhibit expression of essential genes based on the inducible expression of an enzymatically inactive Cas9 protein together with gene-specific guide RNAs (CRISPR interference). Using this system to target several essential genes of M.

Comprehensive Definition of the SigH Regulon of Mycobacterium tuberculosis Reveals Transcriptional Control of Diverse Stress Responses.

Expression of SigH, one of 12 Mycobacterium tuberculosis alternative sigma factors, is induced by heat, oxidative and nitric oxide stresses. SigH activation has been shown to increase expression of several genes, including genes involved in maintaining redox equilibrium and in protein degradation. However, few of these are known to be directly regulated by SigH.

ESTIMATING BACTERIAL DIVERSITY IN (THYSANOPTERA: THRIPIDAE) VIA NEXT GENERATION SEQUENCING.

The last 2 decades have produced a better understanding of insect-microbial associations and yielded some important opportunities for insect control. However, most of our knowledge comes from model systems. Thrips (Thysanoptera: Thripidae) have been understudied despite their global importance as invasive species, plant pests and disease vectors.

De novo Assembly and Analysis of the Northern Leopard Frog Rana pipiens Transcriptome.

The northern leopard frog Rana (Lithobates) pipiens is an important animal model, being used extensively in cancer, neurology, physiology, and biomechanical studies. R. pipiens is a native North American frog whose range extends from northern Canada to southwest United States, but over the past few decades its populations have declined significantly and is now considered uncommon in large portions of the United States and Canada.

Host-directed antimicrobial drugs with broad-spectrum efficacy against intracellular bacterial pathogens.

We sought a new approach to treating infections by intracellular bacteria, namely, by altering host cell functions that support their growth. We screened a library of 640 Food and Drug Administration (FDA)-approved compounds for agents that render THP-1 cells resistant to infection by four intracellular pathogens. We identified numerous drugs that are not antibiotics but were highly effective in inhibiting intracellular bacterial growth with limited toxicity to host cells.

ZipA is required for FtsZ-dependent preseptal peptidoglycan synthesis prior to invagination during cell division.

Rod-shaped bacteria grow by a repetitive cycle of elongation followed by division, and the mechanisms responsible for these two processes have been studied for decades. However, little is known about what happens during the transition between the two activities. At least one event occurs after elongation ends and before division commences, that being the insertion of new cell wall peptidoglycan into a narrowly circumscribed ribbon around midcell where septation is destined to take place.

Escherichia coli low-molecular-weight penicillin-binding proteins help orient septal FtsZ, and their absence leads to asymmetric cell division and branching.

Escherichia coli cells lacking low-molecular-weight penicillin-binding proteins (LMW PBPs) exhibit morphological alterations that also appear when the septal protein FtsZ is mislocalized, suggesting that peptidoglycan modification and division may work together to produce cell shape. We found that in strains lacking PBP5 and other LMW PBPs, higher FtsZ concentrations increased the frequency of branched cells and incorrectly oriented Z rings by 10- to 15-fold. Invagination of these rings produced improperly oriented septa, which in turn gave rise to asymmetric cell poles that eventually elongated into branches.