Inactivation of DltA Modulates Virulence Factor Expression in Streptococcus pyogenes

D-alanylated lipoteichoic acid is a virtually ubiquitous component of gram-positive cell walls. Mutations in the dltABCD operon of numerous species exhibit pleiotropic effects, including reduced virulence, which has been attributed to increased binding of cationic antimicrobial peptides to the more negatively charged cell surface. In this study, we have further investigated the effects that mutating dltA has on virulence factor expression in Streptococcus pyogenes.Isogenic Delta dltA mutants had previously been created in two distinct M1T1 isolates of S. pyogenes. Immunoblots, flow cytometry, and immunofluorescence were used to quantitate M protein levels in these strains, as well as to assess their ability to bind complement. Bacteria were tested for their ability to interact with human PMN and to grow in whole human blood. Message levels for emm, sic, and various regulatory elements were assessed by quantitative RT-PCR. Cell walls of Delta dltA mutants contained much less M protein than cell walls of parent strains and this correlated with reduced levels of emm transcripts, increased deposition of complement, increased association of bacteria with polymorphonuclear leukocytes, and reduced bacterial growth in whole human blood. Transcription of at least one other gene of the mga regulon, sic, which encodes a protein that inactivates antimicrobial peptides, was also dramatically reduced in Delta dltA mutants. Concomitantly, ccpA and rofA were unaffected, while rgg and arcA were up-regulated.This study has identified a novel mechanism for the reduced virulence of dltA mutants of Streptococcus pyogenes in which gene regulatory networks somehow sense and respond to the loss of DltA and lack of D-alanine esterification of lipoteichoic acid. The mechanism remains to be determined, but the data indicate that the status of D-alanine-lipoteichoic acid can significantly influence the expression of at least some streptococcal virulence factors and provide further impetus to targeting the dlt operon of gram-positive pathogens in the search for novel antimicrobial compounds

Gene therapy for monogenic liver diseases: clinical successes, current challenges and future prospects

Over the last decade, pioneering liver-directed gene therapy trials for haemophilia B have achieved sustained clinical improvement after a single systemic injection of adeno-associated virus (AAV) derived vectors encoding the human factor IX cDNA. These trials demonstrate the potential of AAV technology to provide long-lasting clinical benefit in the treatment of monogenic liver disorders. Indeed, with more than ten ongoing or planned clinical trials for haemophilia A and B and dozens of trials planned for other inherited genetic/metabolic liver diseases, clinical translation is expanding rapidly. Gene therapy is likely to become an option for routine care of a subset of severe inherited genetic/metabolic liver diseases in the relatively near term. In this review, we aim to summarise the milestones in the development of gene therapy, present the different vector tools and their clinical applications for liver-directed gene therapy. AAV-derived vectors are emerging as the leading candidates for clinical translation of gene delivery to the liver. Therefore, we focus on clinical applications of AAV vectors in providing the most recent update on clinical outcomes of completed and ongoing gene therapy trials and comment on the current challenges that the field is facing for large-scale clinical translation. There is clearly an urgent need for more efficient therapies in many severe monogenic liver disorders, which will require careful risk-benefit analysis for each indication, especially in paediatrics

Clinical and in vitro

Treatment of infections caused by Burkholderia cepacia complex (Bcc) in cystic fibrosis (CF) patients poses a complex problem. Bcc is multidrug-resistant due to innate and acquired mechanisms of resistance. As CF patients receive multiple courses of antibiotics, susceptibility patterns of strains from CF patients may differ from those noted in strains from non-CF patients. Thus, there was a need for assessing in vitro and clinical data to guide antimicrobial therapy in these patients. A systematic search of literature, followed by extraction and analysis of available information from human and in vitro studies was done. The results of the analysis are used to address various aspects like use of antimicrobials for pulmonary and non-pulmonary infections, use of combination versus monotherapy, early eradication, duration of therapy, route of administration, management of biofilms, development of resistance during therapy, pharmacokinetics–pharmacodynamics correlations, therapy in post-transplant patients and newer drugs in Bcc-infected CF patients