Purine Nucleosides Interfere with c-di-AMP Levels and Act as Adjuvants To Re-Sensitize MRSA To ß-Lactam Antibiotics.

The purine-derived signaling molecules c-di-AMP and (p)ppGpp control mecA/PBP2a-mediated ß-lactam resistance in methicillin-resistant Staphylococcus aureus (MRSA) raise the possibility that purine availability can control antibiotic susceptibility. Consistent with this, exogenous guanosine and xanthosine, which are fluxed through the GTP branch of purine biosynthesis, were shown to significantly reduce MRSA ß-lactam resistance. In contrast, adenosine (fluxed to ATP) significantly increased oxacillin resistance, whereas inosine (which can be fluxed to ATP and GTP via hypoxanthine) only marginally increased oxacillin susceptibility. Furthermore, mutations that interfere with de novo purine synthesis (pur operon), transport (NupG, PbuG, PbuX) and the salvage pathway (DeoD2, Hpt) increased ß-lactam resistance in MRSA strain JE2. Increased resistance of a nupG mutant was not significantly reversed by guanosine, indicating that NupG is required for guanosine transport, which is required to reduce ß-lactam resistance. Suppressor mutants resistant to oxacillin/guanosine combinations contained several purine salvage pathway mutations, including nupG and hpt. Guanosine significantly increased cell size and reduced levels of c-di-AMP, while inactivation of GdpP, the c-di-AMP phosphodiesterase negated the impact of guanosine on ß-lactam susceptibility. PBP2a expression was unaffected in nupG or deoD2 mutants, suggesting that guanosine-induced ß-lactam susceptibility may result from dysfunctional c-di-AMP-dependent osmoregulation. These data reveal the therapeutic potential of purine nucleosides, as ß-lactam adjuvants that interfere with the normal activation of c-di-AMP are required for high-level ß-lactam resistance in MRSA. IMPORTANCE The clinical burden of infections caused by antimicrobial resistant (AMR) pathogens is a leading threat to public health. Maintaining the effectiveness of existing antimicrobial drugs or finding ways to reintroduce drugs to which resistance is widespread is an important part of efforts to address the AMR crisis. Predominantly, the safest and most effective class of antibiotics are the ß-lactams, which are no longer effective against methicillin-resistant Staphylococcus aureus (MRSA). Here, we report that the purine nucleosides guanosine and xanthosine have potent activity as adjuvants that can resensitize MRSA to oxacillin and other ß-lactam antibiotics. Mechanistically, exposure of MRSA to these nucleosides significantly reduced the levels of the cyclic dinucleotide c-di-AMP, which is required for ß-lactam resistance. Drugs derived from nucleotides are widely used in the treatment of cancer and viral infections highlighting the clinical potential of using purine nucleosides to restore or enhance the therapeutic effectiveness of ß-lactams against MRSA and potentially other AMR pathogens.

Inborn Errors of Metabolism in Pediatric Epilepsy.

Patients with refractory epilepsy from inborn errors of metabolism typically present as neonates. Direct supplementation with the deficient vitamin or cofactor is recommended, and case series report both efficacy and safety data of these agents. Some conditions may also occur together, necessitating multiple treatments. Despite effective and early treatment, patients are at heightened risk for neurological sequela. The literature on seizures related to metabolic deficiencies for pediatric patients is limited but has some guidance on appropriate dosing and monitoring for agents to target specific deficiencies, which may help with narrowing antiepileptic therapies, reducing side effects, and improving neurodevelopmental outcomes and quality of life. The focus of this review is to discuss the pharmacotherapy, including the most updated published efficacy and safety data, involved in treating refractory epilepsy as a result of metabolic errors.

Medication adherence, molecular monitoring, and clinical outcomes in patients with chronic myelogenous leukemia in a large HMO.

OBJECTIVE: Our objective was to examine the association between adherence to tyrosine kinase inhibitors (TKIs) and molecular monitoring and the risk of disease progression or mortality among patients with chronic phase chronic myeloid leukemia (CML). DESIGN: We assembled a retrospective cohort of patients with CML (chronic phase, no prior cancer history, and confirmed to be Philadelphia chromosome positive) using data from electronic health records and chart reviews. Medication possession ratio (MPR) was used to measure drug adherence. SETTING: A large, community-based, integrated health plan in Southern California. PARTICIPANTS: The cohort consisted of 245 adult patients (≥18 years old) with Philadelphia positive chronic phase CML diagnosed from 2001 to 2012 and followed through 2013. MAIN OUTCOME MEASURES: In survival analyses, we examined the association of TKI adherence (MPR) and polymerase chain reaction (PCR) monitoring test frequency with the composite clinical outcome, progression to accelerated phase disease-blast crisis or mortality (progression-free survival). The cohort was followed for a maximum of 13 years (median 4.6 years). RESULTS: Over 90% of the cohort initiated TKI therapy within 3 months of diagnosis, and the mean MPR was 88% (SD 18%). Virtually all patients (96%) started on imatinib. The rates of progression to accelerated phase-blast crisis and mortality were lower in patients with greater TKI adherence (20.4/1000 person-years) versus lower adherence (27.0/1000 person-years). Patients who underwent PCR monitoring had a significantly reduced risk of progression or mortality, which was seen in patients with high and low TKI adherence status from both the groups (hazard ratio [HR] 0.07 [95% CI 0.03-0.19 if MPR >90%] and HR 0.70 [95% CI 0.02-0.21 if MPR<90%]). CONCLUSION: Our results suggest that close clinical monitoring, which includes PCR monitoring in patients with high and low TKI drug adherence, is associated with a lower risk of progression or mortality.

Effects of apomixis and polyploidy on diversification and geographic distribution in Amelanchier (Rosaceae).

UNLABELLED: • PREMISE OF THE STUDY: Amelanchier polyploid apomicts differ from sexual diploids in their more complex diversification, greater species problems, and geographic distribution. To understand these differences, we investigated the occurrence of polyploidy and frequency of apomixis. This research helps clarify species delimitation in an evolutionarily complex genus.• METHODS: We used flow cytometry to estimate genome size of 1355 plants. We estimated the frequency of apomixis from flow-cytometrically determined ploidy levels of embryo and endosperm and from a progeny study using RAPD markers. We explored relationships of triploids to other ploidy levels and of ploidy levels to latitude plus elevation.• KEY RESULTS: Diploids (32% of sample) and tetraploids (62%) were widespread. Triploids (6%) mostly occurred in small numbers with diploids from two or more species or with diploids and tetraploids. Seeds from diploids were 2% apomictic, the first report of apomixis in Amelanchier diploids. Seeds from triploids were 75% apomictic. We documented potential triploid bridge and triploid block from unbalanced endosperm and low pollen viability. Seeds from tetraploids were 97% apomictic, and tetraploids often formed microspecies. We did not find strong evidence for geographical parthenogenesis in North American Amelanchier. Most currently recognized species contained multiple ploidy levels that were morphologically semicryptic.• CONCLUSIONS: Documentation of numerous transitions from diploidy to polyploidy helps clarify diversification, geographic distribution, and the species problem in Amelanchier. Despite the infrequent occurrence of triploids, their retention of 25% sexuality and capacity for triploid bridge may be important steps between sexual diploids and predominantly apomictic tetraploids.