The resurgence of tuberculosis has been characterized by the emergence of significant numbers of drug-resistant strains. Furthermore, microorganisms of the Mycobacterium avium complex, opportunistic pathogens common in AIDS patients, are inherently resistant to many traditional antimycobacterial agents (20,23). Hence, the development of novel drugs for the treatment of atypical infections by M. avium, Mycobacterium intracellulare, and multiple-drug-resistant Mycobacterium tuberculosis is urgently needed.The mycobacterial cell wall is an effective barrier that contributes to drug resistance (45). Inhibitors of cell wall biosynthesis not only are potential antimycobacterial agents but also increase mycobacterial susceptibility to other antimicrobial agents ( (26,31,33). Moreover, the biosynthesis of mycolyl-arabinogalactan-peptidoglycan complex is inhibited by DCS in M. tuberculosis (10), and biochemical studies indicated that D-alanine ligase is one of the targets in mycobacteria (11).DCS is an effective antimycobacterial agent but is rarely prescribed and used only in combined therapies due to its adverse effects (21,22,54). These side effects are due to binding of DCS to neuronal N-methyl aspartate receptors (44) and inhibition of enzymes that metabolize and synthesize the neurotransmitter ␥-aminobutyric acid (53). Nevertheless, DCS is an excellent candidate for the development of a new generation of antibiotics. Two important considerations predict that rationally designed derivatives of DCS may be more efficacious antimicrobial agents. First, DCS targets participate in essential steps of cell wall synthesis. Second, DCS resistance has not yet become an important clinical problem. Therefore, the identification of DCS targets and the elucidation of the mechanisms leading to DCS resistance may contribute to the development of new therapeutics with fewer side effects and mechanisms of action which do not favor the emergence of resistance.Few studies on the mode of action and mechanisms of DCS resistance in mycobacteria have been conducted. David (9) isolated and characterized step-wise DCS-resistant (DCS r ) mutants of M. tuberculosis and discovered mutants that showed either normal or reduced cellular permeability to DCS. It was hypothesized that mutants with normal uptake carried mutations in the D-alanine ligase gene, but no biochemical or molecular evidence in support of this hypothesis was provided.Here we describe the first molecular genetic analysis of DCS resistance in mycobacteria, which led to the identification of one of the DCS targets and resistance mechanisms in Mycobacterium smegmatis. A spontaneous DCS r mutant strain of M. smegmatis exhibited a promoter-up mutation in the D-alanine racemase gene (alrA) which increased the levels of expression

The data plead for combining phenotypic and genotypic approaches in the diagnosis of efflux-mediated resistance in P. aeruginosa.

The BCR-ABL-negative myeloproliferative neoplasms (MPNs), polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF), entered the spotlight in 2005 when the unique somatic acquired JAK2 V617F mutation was described in 495% of PV and in 50% of ET and PMF patients. For the very rare PV patients who do not harbor the JAK2 V617F mutation, exon 12 JAK2 mutants were discovered also to result in activated forms of JAK2. A minority of ET and PMF patients harbor mutations that constitutively activate the thrombopoietin receptor (TpoR). In bone marrow reconstitution models based on retroviral transduction, the phenotype induced by JAK2 V617F is less severe and different from the rapid fatal myelofibrosis induced by TpoR W515L. The reasons for these differences are unknown. Exactly by which mechanism(s) one acquired somatic mutation, JAK2 V617F, can promote three different diseases remains a mystery, although gene dosage and host genetic variation might have important functions. We review the recent progress made in deciphering signaling anomalies in PV, ET and PMF, with an emphasis on the relationship between JAK2 V617F and cytokine receptor signaling and on cross-talk with several other signaling pathways.