Generation of a mutator parasite to drive resistome discovery in Plasmodium falciparum

In vitro evolution of drug resistance is a powerful approach for identifying antimalarial targets, however, key obstacles to eliciting resistance are the parasite inoculum size and mutation rate. Here we sought to increase parasite genetic diversity to potentiate resistance selections by editing catalytic residues of Plasmodium falciparum DNA polymerase δ. Mutation accumulation assays reveal a ~5–8 fold elevation in the mutation rate, with an increase of 13–28 fold in drug-pressured lines. Upon challenge with the spiroindolone PfATP4-inhibitor KAE609, high-level resistance is obtained more rapidly and at lower inocula than wild-type parasites. Selections also yield mutants with resistance to an “irresistible” compound, MMV665794 that failed to yield resistance with other strains. We validate mutations in a previously uncharacterised gene, PF3D7_1359900, which we term quinoxaline resistance protein (QRP1), as causal for resistance to MMV665794 and a panel of quinoxaline analogues. The increased genetic repertoire available to this “mutator” parasite can be leveraged to drive P. falciparum resistome discovery

Mitigating the risk of antimalarial resistance via covalent dual-subunit inhibition of the Plasmodium proteasome

The Plasmodium falciparum proteasome constitutes a promising antimalarial target, with multiple chemotypes potently and selectively inhibiting parasite proliferation and synergizing with the first-line artemisinin drugs, including against artemisinin-resistant parasites. We compared resistance profiles of vinyl sulfone, epoxyketone, macrocyclic peptide, and asparagine ethylenediamine inhibitors and report that the vinyl sulfones were potent even against mutant parasites resistant to other proteasome inhibitors and did not readily select for resistance, particularly WLL that displays covalent and irreversible binding to the catalytic β2 and β5 proteasome subunits. We also observed instances of collateral hypersensitivity, whereby resistance to one inhibitor could sensitize parasites to distinct chemotypes. Proteasome selectivity was confirmed using CRISPR/Cas9-edited mutant and conditional knockdown parasites. Molecular modeling of proteasome mutations suggested spatial contraction of the β5 P1 binding pocket, compromising compound binding. Dual targeting of P. falciparum proteasome subunits using covalent inhibitors provides a potential strategy for restoring artemisinin activity and combating the spread of drug-resistant malaria

Oropharynx microbial diversity of phyla in the asthma, chronic obstructive pulmonary disease (COPD), and healthy control groups.

<p>Oropharynx microbial diversity of phyla in the asthma, chronic obstructive pulmonary disease (COPD), and healthy control groups.</p

Characteristics of 47 subjects in the asthma, COPD, and healthy control groups.

<p>FEV1, forced expiratory volume in 1 s; FVC, functional vital capacity.</p><p>Date for FEV1% and FVC % are expressed as mean 6 SD.</p><p>Characteristics of 47 subjects in the asthma, COPD, and healthy control groups.</p

Microbial Communities in the Upper Respiratory Tract of Patients with Asthma and Chronic Obstructive Pulmonary Disease

<div><p>Respiratory infections are well-known triggers of chronic respiratory diseases. Recently, culture-independent tools have indicated that lower airway microbiota may contribute to pathophysiologic processes associated with asthma and chronic obstructive pulmonary disease (COPD). However, the relationship between upper airway microbiota and chronic respiratory diseases remains unclear. This study was undertaken to define differences of microbiota in the oropharynx of asthma and COPD patients relative to those in healthy individuals. To account for the qualitative and quantitative diversity of the 16S rRNA gene in the oropharynx, the microbiomes of 18 asthma patients, 17 COPD patients, and 12 normal individuals were assessed using a high-throughput next-generation sequencing analysis. In the 259,572 total sequence reads, α and β diversity measurements and a generalized linear model revealed that the oropharynx microbiota are diverse, but no significant differences were observed between asthma and COPD patients. <i>Pseudomonas</i> spp. of Proteobacteria and <i>Lactobacillus</i> spp. of Firmicutes were highly abundant in asthma and COPD. By contrast, <i>Streptococcus</i>, <i>Veillonella</i>, <i>Prevotella</i>, and <i>Neisseria</i> of Bacteroidetes dominated in the healthy oropharynx. These findings are consistent with previous studies conducted in the lower airways and suggest that oropharyngeal airway microbiota are important for understanding the relationships between the various parts of the respiratory tract with regard to bacterial colonization and comprehensive assessment of asthma and COPD.</p></div

Abundance table of normal, asthma and COPD.

<p>Abundance table of normal, asthma and COPD.</p

Statistical analysis of the healthy control, asthma, and chronic obstructive pulmonary disease (COPD) groups.

<p>Statistical analysis of the healthy control, asthma, and chronic obstructive pulmonary disease (COPD) groups.</p

Roles of the gut virome and mycobiome in faecal microbiota transplantation

Faecal microbiota transplantation (FMT) is an innovative approach to treat diseases that are associated with gut dysbiosis, by transferring a healthy stool microbiota to a recipient with disease. Beyond the bacteriome, the human gut also harbours diverse communities of viruses and fungi, collectively known as the virome and the mycobiome. The effect of the virome and the mycobiome on the success of FMT therapy has not been appreciated until recently. In this Review, we summarise the current literature on the effects of the gut virome and mycobiome on the treatment of various diseases with FMT. We discuss the beneficial effects and health concerns of viral and fungal transfer during FMT, and highlight the roles of bacteriophages and Candida species in FMT efficacy. We also summarise the intricate relationships between the gut virome, mycobiome, bacteriome, and host immunity underlying FMT effectiveness. Future efforts should be devoted to understanding the versatile roles and the therapeutic mechanisms of viral and fungal lineages, and their combinations, in different diseases. Harnessing the gut virome, mycobiome, and bacteriome in combination is a promising prospect for the future of FMT and microbiota-based therapies

Timing of Tributyrin Supplementation Differentially Modulates Gastrointestinal Inflammation and Gut Microbial Recolonization Following Murine Ileocecal Resection

Background: Gastrointestinal surgery imparts dramatic and lasting imbalances, or dysbiosis, to the composition of finely tuned microbial ecosystems. The aim of the present study was to use a mouse ileocecal resection (ICR) model to determine if tributyrin (TBT) supplementation could prevent the onset of microbial dysbiosis or alternatively enhance the recovery of the gut microbiota and reduce gastrointestinal inflammation. Methods: Male wild-type (129 s1/SvlmJ) mice aged 8–15 weeks were separated into single cages and randomized 1:1:1:1 to each of the four experimental groups: control (CTR), preoperative TBT supplementation (PRE), postoperative TBT supplementation (POS), and combined pre- and postoperative supplementation (TOT). ICR was performed one week from baseline assessment with mice assessed at 1, 2, 3, and 4 weeks postoperatively. Primary outcomes included evaluating changes to gut microbial communities occurring from ICR to 4 weeks. Results: A total of 34 mice that underwent ICR (CTR n = 9; PRE n = 10; POS n = 9; TOT n = 6) and reached the primary endpoint were included in the analysis. Postoperative TBT supplementation was associated with an increased recolonization and abundance of anaerobic taxa including Bacteroides thetaiotomicorn, Bacteroides caecimuris, Parabacteroides distasonis, and Clostridia. The microbial recolonization of PRE mice was characterized by a bloom of aerotolerant organisms including Staphylococcus, Lactobacillus, Enteroccaceae, and Peptostreptococcacea. PRE mice had a trend towards decreased ileal inflammation as evidenced by decreased levels of IL-1β (p = 0.09), IL-6 (p = 0.03), and TNF-α (p &lt; 0.05) compared with mice receiving TBT postoperatively. In contrast, POS mice had trends towards reduced colonic inflammation demonstrated by decreased levels of IL-6 (p = 0.07) and TNF-α (p = 0.07). These changes occurred in the absence of changes to fecal short-chain fatty acid concentrations or histologic injury scoring. Conclusions: Taken together, the results of our work demonstrate that the timing of tributyrin supplementation differentially modulates gastrointestinal inflammation and gut microbial recolonization following murine ICR