Pharmacokinetics, Tissue Distribution, and Efficacy of VIO-001 (Meropenem/Piperacillin/Tazobactam) for Treatment of Methicillin-Resistant Staphylococcus aureus Bacteremia in Immunocompetent Rabbits with Chronic Indwelling Vascular Catheters.

Methicillin-resistant Staphylococcus aureus (MRSA) infections of surgically implanted subcutaneous vascular catheters (SISVCs) cause serious morbidity in patients with chronic illnesses. Previous in vitro and murine models demonstrated the synergistic interaction of equimolar concentrations of meropenem/piperacillin/tazobactam (MPT) (VIO-001) against MRSA infection. We investigated the pharmacokinetics (PK) and efficacy of VIO-001 for the treatment of MRSA bacteremia in immunocompetent rabbits with SISVCs. In PK studies, we determined that optimal dosing to achieve a time above 4× MIC (T>4×MIC) of a duration of 3 to 3.30 h required a 1-h infusion with every-4-h (Q4h) dosing. Study groups in efficacy experiments consisted of MPT combinations of 100/150/100 mg/kg of body weight (MPT100), 200/300/200 mg/kg (MPT200), and 400/600/400 mg/kg (MPT400); vancomycin (VAN) at 15 mg/kg; and untreated controls (UC). The inoculum of MRSA isolate USA300-TCH1516 (1 × 103 organisms) was administered via the SISCV on day 1 and locked for 24 h. The 8-day therapy started at 24 h postinoculation. There was a significant reduction of MRSA in blood cultures from the SISVCs in all treatment groups, with full clearance on day 4, versus UCs (P < 0.05). Consistent with the clearance of SISVC-related infection, full eradication of MRSA was achieved in lungs, heart, liver, spleen, and kidneys at the end of the study versus UC (P < 0.01). These results strongly correlated with time-kill data, where MPT in the range of 4/6/4 µg/ml to 32/48/32 µg/ml demonstrated a significant 6-log decrease in the bacterial burden versus UC (P < 0.01). In summary, VIO-001 demonstrated a favorable PK/pharmacodynamic (PD) profile and activity against SISCV MRSA infection, bacteremia, and disseminated infection. This rabbit model provides a new system for understanding new antimicrobial agents against MRSA SISVC-related infection, and these data provide a basis for future clinical investigation.

Mediator subunit 12 coordinates intrinsic and extrinsic control of epithalamic development.

In the developing brain, the production of neurons from multipotent precursors must be carefully regulated in order to generate the appropriate numbers of various differentiated neuronal types. Inductive signals from extrinsic elements such as growth factors need to be integrated with timely expression of intrinsic elements such as transcription factors that define the competence of the cell. The transcriptional Mediator complex offers a mechanism to coordinate the timing and levels of intrinsic and extrinsic influences by acting as a rapid molecular switch for transcription of poised RNA pol II. The epithalamus is a highly conserved region of the vertebrate brain that differentiates early and rapidly in the zebrafish. It includes the pineal and parapineal organs and the habenular nuclei. Mutation of the Mediator complex subunit Med12 impairs the specification of habenular and parapineal neurons and causes a loss of differentiation in pineal neurons and photoreceptors. Although FGF ligands and transcription factors for parapineal and photoreceptor development are still expressed in the pineal complex of med12 mutants, FGF signaling is impaired and transcription factor expression is reduced and/or delayed. We find that the timely expression of one of these transcription factors, tbx2b, is controlled by Med12 and is vital for parapineal specification. We propose that the Mediator complex is responsible for subtle but significant changes in transcriptional timing and amplitude that are essential for coordinating the development of neurons in the epithalamus.

Microbiota restoration reduces antibiotic-resistant bacteria gut colonization in patients with recurrent Clostridioides difficile infection from the open-label PUNCH CD study.

BACKGROUND: Once antibiotic-resistant bacteria become established within the gut microbiota, they can cause infections in the host and be transmitted to other people and the environment. Currently, there are no effective modalities for decreasing or preventing colonization by antibiotic-resistant bacteria. Intestinal microbiota restoration can prevent Clostridioides difficile infection (CDI) recurrences. Another potential application of microbiota restoration is suppression of non-C. difficile multidrug-resistant bacteria and overall decrease in the abundance of antibiotic resistance genes (the resistome) within the gut microbiota. This study characterizes the effects of RBX2660, a microbiota-based investigational therapeutic, on the composition and abundance of the gut microbiota and resistome, as well as multidrug-resistant organism carriage, after delivery to patients suffering from recurrent CDI. METHODS: An open-label, multi-center clinical trial in 11 centers in the USA for the safety and efficacy of RBX2660 on recurrent CDI was conducted. Fecal specimens from 29 of these subjects with recurrent CDI who received either one (N = 16) or two doses of RBX2660 (N = 13) were analyzed secondarily. Stool samples were collected prior to and at intervals up to 6 months post-therapy and analyzed in three ways: (1) 16S rRNA gene sequencing for microbiota taxonomic composition, (2) whole metagenome shotgun sequencing for functional pathways and antibiotic resistome content, and (3) selective and differential bacterial culturing followed by isolate genome sequencing to longitudinally track multidrug-resistant organisms. RESULTS: Successful prevention of CDI recurrence with RBX2660 correlated with taxonomic convergence of patient microbiota to the donor microbiota as measured by weighted UniFrac distance. RBX2660 dramatically reduced the abundance of antibiotic-resistant Enterobacteriaceae in the 2 months after administration. Fecal antibiotic resistance gene carriage decreased in direct relationship to the degree to which donor microbiota engrafted. CONCLUSIONS: Microbiota-based therapeutics reduce resistance gene abundance and resistant organisms in the recipient gut microbiome. This approach could potentially reduce the risk of infections caused by resistant organisms within the patient and the transfer of resistance genes or pathogens to others. TRIAL REGISTRATION: ClinicalTrials.gov, NCT01925417 ; registered on August 19, 2013.

Impact of Amoxicillin-Clavulanate followed by Autologous Fecal Microbiota Transplantation on Fecal Microbiome Structure and Metabolic Potential.

Strategies to prevent multidrug-resistant organism (MDRO) infections are scarce, but autologous fecal microbiota transplantation (autoFMT) may limit gastrointestinal MDRO expansion. AutoFMT involves banking one's feces during a healthy state for later use in restoring gut microbiota following perturbation. This pilot study evaluated the effect of autoFMT on gastrointestinal microbiome taxonomic composition, resistance gene content, and metabolic capacity after exposure to amoxicillin-clavulanic acid (Amox-Clav). Ten healthy participants were enrolled. All received 5 days of Amox-Clav. Half were randomized to autoFMT, derived from stool collected pre-antimicrobial exposure, by enema, and half to saline enema. Participants submitted stool samples pre- and post-Amox-Clav and enema and during a 90-day follow-up period. Shotgun metagenomic sequencing revealed taxonomic composition, resistance gene content, and metabolic capacity. Amox-Clav significantly altered gut taxonomic composition in all participants (n = 10, P < 0.01); however, only three participants exhibited major changes at the phylum level following exposure. In the cohort as a whole, beta-lactamase genes were enriched following Amox-Clav (P < 0.05), and predicted metabolic capacity was significantly altered (P < 0.01). Species composition, metabolic capacity, and beta-lactamase abundance returned to pre-antimicrobial exposure state 7 days after either autoFMT or saline enema (P > 0.05, compared to enrollment). Alterations to microbial metabolic capacity occurred following antimicrobial exposure even in participants without substantial taxonomic disruption, potentially creating open niches for pathogen colonization. Our findings suggest that metabolic potential is an important consideration for complete assessment of antimicrobial impact on the microbiome. AutoFMT was well tolerated and may have contributed to phylogenetic recovery. (This study has been registered at ClinicalTrials.gov under identifier NCT02046525.)IMPORTANCE The spread of multidrug resistance among pathogenic organisms threatens the efficacy of antimicrobial treatment options. The human gut serves as a reservoir for many drug-resistant organisms and their resistance genes, and perturbation of the gut microbiome by antimicrobial exposure can open metabolic niches to resistant pathogens. Once established in the gut, antimicrobial-resistant bacteria can persist even after antimicrobial exposure ceases. Strategies to prevent multidrug-resistant organism (MDRO) infections are scarce, but autologous fecal microbiota transplantation (autoFMT) may limit gastrointestinal MDRO expansion. AutoFMT involves banking one's feces during a healthy state for later use in restoring gut microbiota following perturbation. This pilot study evaluated the effect of amoxicillin-clavulanic acid (Amox-Clav) exposure and autoFMT on gastrointestinal microbiome taxonomic composition, resistance gene content, and metabolic capacity. Importantly, we found that metabolic capacity was perturbed even in cases where gross phylogeny remained unchanged and that autoFMT was safe and well tolerated.

Mechanism of High-Level Daptomycin Resistance in Corynebacterium striatum.

Daptomycin, a last-line-of-defense antibiotic for treating Gram-positive infections, is experiencing clinical failure against important infectious agents, including Corynebacterium striatum The recent transition of daptomycin to generic status is projected to dramatically increase availability, use, and clinical failure. Here we confirm the genetic mechanism of high-level daptomycin resistance (HLDR; MIC = >256 µg/ml) in C. striatum, which evolved within a patient during daptomycin therapy, a phenotype recapitulated in vitro In all 8 independent cases tested, loss-of-function mutations in phosphatidylglycerol synthase (pgsA2) were necessary and sufficient for high-level daptomycin resistance. Through lipidomic and biochemical analysis, we demonstrate that daptomycin's activity is dependent on the membrane phosphatidylglycerol (PG) concentration. Until now, the verification of PG as the in vivo target of daptomycin has proven difficult since tested cell model systems were not viable without membrane PG. C. striatum becomes daptomycin resistant at a high level by removing PG from the membrane and changing the membrane composition to maintain viability. This work demonstrates that loss-of-function mutation in pgsA2 and the loss of membrane PG are necessary and sufficient to produce high-level resistance to daptomycin in C. striatumIMPORTANCE Antimicrobial resistance threatens the efficacy of antimicrobial treatment options, including last-line-of-defense drugs. Understanding how this resistance develops can help direct antimicrobial stewardship efforts and is critical to designing the next generation of antimicrobial therapies. Here we determine how Corynebacterium striatum, a skin commensal and opportunistic pathogen, evolved high-level resistance to a drug of last resort, daptomycin. Through a single mutation, this pathogen was able to remove the daptomycin's target, phosphatidylglycerol (PG), from the membrane and evade daptomycin's bactericidal activity. We found that additional compensatory changes were not necessary to support the removal of PG and replacement with phosphatidylinositol (PI). The ease with which C. striatum evolved high-level resistance is cause for alarm and highlights the importance of screening new antimicrobials against a wide range of clinical pathogens which may harbor unique capacities for resistance evolution.