Streptolysin O and NAD-Glycohydrolase Prevent Phagolysosome Acidification and Promote Group A Streptococcus Survival in Macrophages

ABSTRACT Group A Streptococcus (GAS, Streptococcus pyogenes) is an ongoing threat to human health as the agent of streptococcal pharyngitis, skin and soft tissue infections, and life-threatening conditions such as necrotizing fasciitis and streptococcal toxic shock syndrome. In animal models of infection, macrophages have been shown to contribute to host defense against GAS infection. However, as GAS can resist killing by macrophages in vitro and induce macrophage cell death, it has been suggested that GAS intracellular survival in macrophages may enable persistent infection. Using isogenic mutants, we now show that the GAS pore-forming toxin streptolysin O (SLO) and its cotoxin NAD-glycohydrolase (NADase) mediate GAS intracellular survival and cytotoxicity for macrophages. Unexpectedly, the two toxins did not inhibit fusion of GAS-containing phagosomes with lysosomes but rather prevented phagolysosome acidification. SLO served two essential functions, poration of the phagolysosomal membrane and translocation of NADase into the macrophage cytosol, both of which were necessary for maximal GAS intracellular survival. Whereas NADase delivery to epithelial cells is mediated by SLO secreted from GAS bound to the cell surface, in macrophages, the source of SLO and NADase is GAS contained within phagolysosomes. We found that transfer of NADase from the phagolysosome to the macrophage cytosol occurs not by simple diffusion through SLO pores but rather by a specific translocation mechanism that requires the N-terminal translocation domain of NADase. These results illuminate the mechanisms through which SLO and NADase enable GAS to defeat macrophage-mediated killing and provide new insight into the virulence of a major human pathogen

Interventional Psychiatry In Pediatric Depression: Community Practices And Preliminary Ketamine Data

Pediatric depression is a significant public health problem, and adolescents whose depression does not respond to first-line treatments have few evidence-based options. Interventional psychiatric services (IPS), including ketamine, esketamine, electroconvulsive therapy (ECT), and repetitive transcranial magnetic stimulation (rTMS) are promising approaches for adult depression, but data on efficacy and safety in the pediatric population are limited. The purpose of this thesis is two-fold: first, to better understand the current attitudes, level of access, and community practices surrounding the use of IPS in pediatric populations. Secondly, to build the IPS pediatric evidence base by assessing the intermediate-term safety and efficacy of intravenous (IV) ketamine in adolescent treatment-resistant depression (TRD). The first aim was accomplished through an online, anonymous survey of psychiatric healthcare providers who treat pediatric patients and providers of any discipline who advertise IPS. 898 healthcare providers were contacted, 330 of whom were sent the survey, with a response rate of 12.7%. Out of a total of 42 respondents, 4 providers treat pediatric patients with ketamine, 2 with rTMS, and 1 with ECT targeting diagnoses that include TRD, obsessive-compulsive disorder, post-traumatic stress disorder, Tourette’s syndrome, and schizophrenia/schizoaffective disorder. The second aim was accomplished through an interim analysis of an ongoing double-blind, midazolam-controlled clinical trial of a multiple-dose paradigm of IV ketamine to treat adolescent TRD. Of the three study participants, the one ketamine-randomized participant had a significant clinical response during the blinded phase, whereas the two midazolam-randomized participants did not have a response to midazolam. The midazolam-randomized subjects also did not respond to subsequent open ketamine treatment but did improve in the open phase of the trial from other treatments. Regarding safety, all infusion-related side effects were transient and resolved within 1.5 hours of infusion completion, and our intermediate-term safety outcome of cognitive function showed no significant decreases on composite scores of the Cogstate battery. In conclusion, here we demonstrate in a regional sample that IPS are being used off-label in the community to treat pediatric mental illness, despite having little data available for safety or efficacy in this population. We show preliminary data, however, that adolescent patients with TRD tolerate IV ketamine treatments, with variable efficacy in this small sample size, highlighting the need for continued study