Current methods for the detection of Mycobacterium tuberculosis (Mtb) are either time consuming or require expensive instruments and are thus are not suitable for point-of-care diagnosis. The design, synthesis, and evaluation of fluorogenic probes with high specificity for BlaC, a biomarker expressed by Mtb, are described. The fluorogenic probe CDG-3 is based on cephalosporin with substitutions at the 2 and 7 positions and it demonstrates over 120 000-fold selectivity for BlaC over TEM-1 Bla, the most common β-lactamase. CDG-3 can detect 10 colony-forming units of the attenuated Mycobacterium bovis strain BCG in human sputum in the presence of high levels of contaminating β-lactamases expressed by other clinically prevalent bacterial strains. In a trial with 50 clinical samples, CDG-3 detected tuberculosis with 90 % sensitivity and 73 % specificity relative to Mtb culture within one hour, thus demonstrating its potential as a low-cost point-of-care test for use in resource-limited areas.

Mycobacterium tuberculosis (Mtb) has complex and dynamic interactions with the human host, and subpopulations of Mtb that emerge during infection can influence disease outcomes. This study implicates zinc ion (Zn2+) availability as a likely driver of bacterial phenotypic heterogeneity in vivo. Zn2+ sequestration is part of “nutritional immunity”, where the immune system limits micronutrients to control pathogen growth, but this defense mechanism seems to be ineffective in controlling Mtb infection. Nonetheless, Zn2+-limitation is an environmental cue sensed by Mtb, as calprotectin triggers the zinc uptake regulator (Zur) regulon response in vitro and co-localizes with Zn2+-limited Mtb in vivo. Prolonged Zn2+ limitation leads to numerous physiological changes in vitro, including differential expression of certain antigens, alterations in lipid metabolism and distinct cell surface morphology. Furthermore, Mtb enduring limited Zn2+ employ defensive measures to fight oxidative stress, by increasing expression of proteins involved in DNA repair and antioxidant activity, including well described virulence factors KatG and AhpC, along with altered utilization of redox cofactors. Here, we propose a model in which prolonged Zn2+ limitation defines a population of Mtb with anticipatory adaptations against impending immune attack, based on the evidence that Zn2+-limited Mtb are more resistant to oxidative stress and exhibit increased survival and induce more severe pulmonary granulomas in mice. Considering that extracellular Mtb may transit through the Zn2+-limited caseum before infecting naïve immune cells or upon host-to-host transmission, the resulting phenotypic heterogeneity driven by varied Zn2+ availability likely plays a key role during early interactions with host cells.

Silicibacter sp. strain TM1040, a member of the Roseobacter clade, forms a symbiosis with unicellular phytoplankton, which is inextricably linked to the biphasic "swim or stick" lifestyle of the bacteria. Mutations in flaC bias the population toward the motile phase. Renewed examination of the FlaC ؊ strain (HG1016) uncovered that it is composed of two different cells: a pigmented type, PS01, and a nonpigmented cell, PS02, each of which has an identical mutation in flaC. While monocultures of PS01 and PS02 had few motile cells (0.6 and 6%, respectively), coculturing the two strains resulted in a 10-fold increase in the number of motile cells. Cell-free supernatants from coculture or wild-type cells were fully capable of restoring motility to PS01 and PS02, which was due to increased fliC3 (flagellin) transcription, FliC3 protein levels per cell, and flagella synthesis. The motility-inducing compound has an estimated mass of 226 Da, as determined by mass spectrometry, and is referred to as Roseobacter Motility Inducer (RMI). Mutations affecting genes involved in phenyl acetic acid synthesis significantly reduced RMI, while defects in tropodithietic acid (TDA) synthesis had marginal or no effect on RMI. RMI biosynthesis is induced by p-coumaric acid, a product of algal lignin degradation. When added to algal cultures, RMI caused loss of motility, cell enlargement, and vacuolization in the algal cells. RMI is a new member of the roseobacticide family of troponoid compounds whose activities affect roseobacters, by shifting their population toward motility, as well as their phytoplankton hosts, through an algicidal effect.

A single glycan-lectin interaction is often weak and semi-specific. Multiple binding domains in a single lectin can bind with multiple glycan molecules simultaneously, making it difficult for the classic “lock-and-key” model to explain these interactions. We demonstrated that hetero-multivalency, a homo-oligomeric protein simultaneously binding to at least two types of ligands, influences LecA (a Pseudomonas aeruginosa adhesin)-glycolipid recognition. We also observed enhanced binding between P. aeruginosa and mixed glycolipid liposomes. Interestingly, strong ligands could activate weaker binding ligands leading to higher LecA binding capacity. This hetero-multivalency is probably mediated via a simple mechanism, Reduction of Dimensionality (RD). To understand the influence of RD, we also modeled LecA’s two-step binding process with membranes using a kinetic Monte Carlo simulation. The simulation identified the frequency of low-affinity ligand encounters with bound LecA and the bound LecA’s retention of the low-affinity ligand as essential parameters for triggering hetero-multivalent binding, agreeing with experimental observations. The hetero-multivalency can alter lectin binding properties, including avidities, capacities, and kinetics, and therefore, it likely occurs in various multivalent binding systems. Using hetero-multivalency concept, we also offered a new strategy to design high-affinity drug carriers for targeted drug delivery.