Antimicrobial, Anti-Inflammatory, Antiparasitic, and Cytotoxic Activities of Laennecia confusa

The current paper investigated the potential benefit of the traditional Mexican medicinal plant Laennecia confusa (Cronquist) G. L. Nesom (Asteraceae). Fractions from the hexane, chloroform, methanol, and aqueous extracts were analyzed for antibacterial, antifungal, anti-inflammatory, and antiparasitic activities. The antimicrobial activity of the extracts and fractions was assessed on bacterial and fungal strains, in addition to the protozoa Leishmania donovani, using a microdilution assay. The propensity of the plant's compounds to produce adverse effects on human health was also evaluated using propidium iodine to identify damage to human macrophages. The anti-inflammatory activity of the extracts and fractions was investigated by measuring the secretion of interleukin-6. Chemical analyses demonstrated the presence of flavonoids, cyanogenic and cardiotonic glycosides, saponins, sesquiterpene lactones, and triterpenes in the chloroform extract. A number of extracts and fractions show antibacterial activity. Of particular interest is antibacterial activity against Staphylococcus aureus and its relative methicillin-resistant strain, MRSA. Hexanic and chloroformic fractions also exhibit antifungal activity and two extracts and the fraction CE 2 antiparasitic activity against Leishmania donovani. All bioactive extracts and fractions assayed were also found to be cytotoxic to macrophages. In addition, the hexane and methane extracts show anti-inflammatory activity by suppressing the secretion of interleukine-6

Defining the burden of febrile illness in rural South and Southeast Asia: an open letter to announce the launch of the Rural Febrile Illness project.

In rural areas of South and Southeast Asia malaria is declining but febrile illnesses still account for substantial morbidity and mortality. Village health workers (VHWs) are often the first point of contact with the formal health system, and for patients with febrile illnesses they can provide early diagnosis and treatment of malaria. However, for the majority of febrile patients, VHWs lack the training, support and resources to provide further care. Consequently, treatable bacterial illnesses are missed, antibiotics are overused and poorly targeted, and patient attendance wanes along with declining malaria. This announces the start of a new initiative, the Rural Febrile Illness (RFI) project, the first in a series of projects to be implemented as part of the South and Southeast Asian Community-based Trials Network (SEACTN) research programme. This multi-country, multi-site project will begin in Bangladesh, Cambodia, Lao PDR, and Myanmar and will define the epidemiological baseline of febrile illness in five remote and underserved areas of Asia where malaria endemicity is declining and access to health services is limited. The RFI project aims to determine the incidence, causes and outcomes of febrile illness; understand the opportunities, barriers and appetite for adjustment of the role of VHWs to include management of non-malarial febrile illnesses; and establish a network of community healthcare providers and facilities capable of implementing interventions designed to triage, diagnose and treat patients presenting with febrile illnesses within these communities in the future. [Abstract copyright: Copyright: © 2021 Chandna A et al.

The role of ergothioneine in the physiology and pathogenesis of Mycobacterium tuberculosis.

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), is capable of synthesizing the small sulfur compound ergothioneine (EGT). The physiological role and regulation of EGT biosynthesis in Mtb is unknown. However, mammalian cells deficient in EGT demonstrate augmented oxidative stress and cell death, suggesting that EGT is an antioxidant. Through this work, we identified the Mtb EGT biosynthesis genes and characterized rv3701c (egtD) to encode for a histidine methyltransferase that forms the EGT biosynthetic pathway intermediate, hercynine (N-α-trimethylhistidine). Construction of a knockout (ΔegtD) demonstrated the methyltransferase to be essential for EGT biosynthesis in Mtb. We investigated the role of EGT in protecting Mtb during macrophage infection and observed a small but significant difference in the ex vivo growth and survival of the ΔegtD strain relative to H37Rv wildtype and the complement strains 120 h post-infection of murine macrophages. However, there was no difference in survival of the ΔegtD mutant in human THP-1 cells over this time period suggesting an alternative role for EGT in Mtb physiology and pathogenesis. Upon further analysis we found EgtD to be phosphorylated by and interact with the Mtb Serine/Threonine Protein Kinase PknD. Phosphorylation of EgtD both in vitro and inside Escherichia coli identified phosphorylation at site Thr-213, permitting us to generate an EgtD phosphomimetic and phosphoablative mutant strains with our ΔegtD mutant. In vitro analysis, demonstrated phosphorylated EgtD to produce significantly lower quantities of methylated histidine relative to the non-phosphorylated form. Further quantification of intracellular EGT levels in the Mtb EgtD phosphomutants and Mtb PknD transposon mutant identified PknD to negatively regulate EGT biosynthesis. From these results, we identified that EGT biosynthesis is up-regulated in response to nutrient starvation. Under these conditions, the Mtb ΔegtD mutant was unable to maintain viability compared to its parental wildtype and complement strains. As starvation induces a non-replicative state in Mtb, these findings indicate that EGT plays a role in mediating persistent infection or disease latency. Further metabolic analysis identified Mtb intracellular EGT levels to be directly correlated with carbon source type and availability, suggesting a role for EGT in long-term energy storage.Medicine, Faculty ofExperimental Medicine, Division ofMedicine, Department ofGraduat

Integrated fever management: disease severity markers to triage children with malaria and non-malarial febrile illness

Abstract Febrile symptoms in children are a leading cause of health-care seeking behaviour worldwide. The majority of febrile illnesses are uncomplicated and self-limited, without the need for referral or hospital admission. However, current diagnostic tools are unable to identify which febrile children have self-limited infection and which children are at risk of progressing to life-threatening infections, such as severe malaria. This paper describes the need for a simple community-based tool that can improve the early recognition and triage of febrile children, with either malarial or non-malarial illness, at risk of critical illness. The integration of a disease severity marker into existing malaria rapid diagnostic tests (RDT) could enable detection of children at risk of severe infection in the hospital and community, irrespective of aetiology. Incorporation of a disease severity marker could inform individualized management and early triage of children at risk of life-threatening infection. A child positive for both malaria and a disease severity marker could be prioritized for urgent referral/admission and parenteral therapy. A child positive for malaria and negative for a disease severity marker could be managed conservatively, as an out-patient, with oral anti-malarial therapy. An RDT with a disease severity marker could facilitate an integrated community-based approach to fever syndromes and improve early recognition, risk stratification, and prompt treatment of severe malaria and other life-threatening infections

Endothelial Activation: The Ang/Tie Axis in Sepsis

Sepsis, a dysregulated host response to infection that causes life-threatening organ dysfunction, is a highly heterogeneous syndrome with no specific treatment. Although sepsis can be caused by a wide variety of pathogenic organisms, endothelial dysfunction leading to vascular leak is a common mechanism of injury that contributes to the morbidity and mortality associated with the syndrome. Perturbations to the angiopoietin (Ang)/Tie2 axis cause endothelial cell activation and contribute to the pathogenesis of sepsis. In this review, we summarize how the Ang/Tie2 pathway is implicated in sepsis and describe its prognostic as well as therapeutic utility in life-threatening infections

Practical Methods to Permit the Analysis of Host Biomarkers in Resource-Limited Settings.

Host biomarker testing can be used as an adjunct to the clinical assessment of patients with infections and might be particularly impactful in resource-constrained settings. Research on the merits of this approach at peripheral levels of low- and middle-income country health systems is limited. In part, this is due to resource-intense requirements for sample collection, processing, and storage. We evaluated the stability of 16 endothelial and immune activation biomarkers implicated in the host response to infection stored in venous plasma and dried blood spot specimens at different temperatures for 6 months. We found that -80°C storage offered no clear advantage over -20°C for plasma aliquots, and most biomarkers studied could safely be stored as dried blood spots at refrigeration temperatures (4°C) for up to 3 months. These results identify more practical methods for host biomarker testing in resource-limited environments, which could help facilitate research in rural and remote environments

Protein Kinase G Induces an Immune Response in Cows Exposed to Mycobacterium avium Subsp. paratuberculosis

To establish infection, pathogens secrete virulence factors, such as protein kinases and phosphatases, to modulate the signal transduction pathways used by host cells to initiate immune response. The protein MAP3893c is annotated in the genome sequence of Mycobacterium avium subspecies paratuberculosis (MAP), the causative agent of Johne’s disease, as the serine/threonine protein kinase G (PknG). In this work, we report that PknG is a functional kinase that is secreted within macrophages at early stages of infection. The antigen is able to induce an immune response from cattle exposed to MAP in the form of interferon gamma production after stimulation of whole blood with PknG. These findings suggest that PknG may contribute to the pathogenesis of MAP by phosphorylating macrophage signalling and/or adaptor molecules as observed with other pathogenic mycobacterial species

Pulmonary Nontuberculous Mycobacteria, Ontario, Canada, 2020

We measured annual prevalence of microbiologically defined nontuberculous mycobacterial lung disease in Ontario, Canada. Mycobacterium avium prevalence was 13 cases/100,000 persons in 2020, a 2.5-fold increase from 2010, indicating a large increase in true M. avium lung disease. During the same period, M. xenopi decreased nearly 50%, to 0.84 cases/100,000 persons

Soluble urokinase-type plasminogen activator receptor as a prognostic marker of Ugandan children at risk of severe and fatal malaria

Background: Current malaria diagnostic tests do not reliably identify children at risk of severe and fatal infection. Host immune and endothelial activation contribute to malaria pathogenesis. Soluble urokinase-type plasminogen activator receptor (suPAR) is a marker of these pathways. We hypothesized that measuring suPAR at presentation could risk-stratify children with malaria. Methods: Plasma suPAR levels were determined in consecutive febrile children with malaria at presentation to hospital in Jinja, Uganda. We evaluated the accuracy of suPAR in predicting in-hospital mortality, and whether suPAR could improve a validated clinical scoring system (Lambaréné Organ Dysfunction Score [LODS]). Results: Of the 1226 children with malaria, 39 (3.2%) died. suPAR concentrations at presentation were significantly higher in children who went on to die than in those who survived (P \u3c .0001). suPAR levels were associated with disease severity (LODS: 0 vs 1, P = .001; 1 vs 2, P \u3c .001; 2 vs 3, 0 vs 2, 1 vs 3, and 0 vs 3, P \u3c .0001). suPAR concentrations were excellent predictors of in-hospital mortality (area under the receiver operating characteristic curve [AUROC], 0.92 [95% confidence interval {CI}, .91–.94]). The prognostic accuracy of LODS (AUROC, 0.93 [95% CI, .91–.94]) was improved when suPAR was added (AUROC, 0.97 [95% CI, .96–.98]; P \u3c .0001). Conclusions Conclusions: Measuring suPAR at presentation can identify children at risk of severe and fatal malaria. Adding suPAR to clinical scores could improve the recognition and triage of children at risk of death. suPAR can be detected with a point-of-care test and can now be evaluated in prospective trials

Biophysical Characterization of the Tandem FHA Domain Regulatory Module from the Mycobacterium tuberculosis ABC Transporter Rv1747.

The Mycobacterium tuberculosis ATP-binding cassette transporter Rv1747 is a putative exporter of cell wall biosynthesis intermediates. Rv1747 has a cytoplasmic regulatory module consisting of two pThr-interacting Forkhead-associated (FHA) domains connected by a conformationally disordered linker with two phospho-acceptor threonines (pThr). The structures of FHA-1 and FHA-2 were determined by X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy, respectively. Relative to the canonical 11-strand β-sandwich FHA domain fold of FHA-1, FHA-2 is circularly permuted and lacking one β-strand. Nevertheless, the two share a conserved pThr-binding cleft. FHA-2 is less stable and more dynamic than FHA-1, yet binds model pThr peptides with moderately higher affinity (∼50 μM versus 500 μM equilibrium dissociation constants). Based on NMR relaxation and chemical shift perturbation measurements, when joined within a polypeptide chain, either FHA domain can bind either linker pThr to form intra- and intermolecular complexes. We hypothesize that this enables tunable phosphorylation-dependent multimerization to regulate Rv1747 transporter activity