Journal: ACS infectious diseases
The parasitic disease onchocerciasis is the second leading cause of preventable blindness, afflicting more than 18 million people worldwide. Despite an available treatment, ivermectin, and control efforts by the World Health Organization, onchocerciasis remains a burden in many regions. With an estimated 120 million people living in areas at risk of infection, efforts are now shifting from prevention to surveillance and elimination. The lack of a robust, point-of-care diagnostic for an active Onchocerca infection has been a limiting factor in these efforts. Previously, we reported the discovery of the biomarker N-acetyl-tyramine-O-glucuronide (NATOG) in human urine samples and its ability to track treatment progression between medicated patients relative to placebo; we also established its capability to monitor disease burden in a jird model. NATOG is a human-produced metabolite of tyramine which, itself is produced as a nematode neurotransmitter. The ability of NATOG to distinguish between active and past infection overcomes the limitations of antibody biomarkers and PCR methodologies. Lateral flow immunoassay (LFIA) diagnostics offer the versatility and simplicity to be employed in the field and are inexpensive enough to be utilized in largescale screening efforts. Herein, we report the development and assessment of a NATOG-based urine LFIA for onchocerciasis, which accurately identified 85% of analyzed patient samples (N = 27).
The recent outbreak of coronavirus disease 2019 (COVID-19) highlights an urgent need for therapeutics. Through a series of drug repurposing screening campaigns, niclosamide, an FDA-approved anthelminthic drug, was found to be effective against various viral infections with nanomolar to micromolar potency such as SARS-CoV, MERS-CoV, ZIKV, HCV and human adenovirus, indicating its potential as an antiviral agent. In this brief article, we summarize the broad antiviral activities of niclosamide and highlight its potential clinical use for treatment of COVID-19.
Staphylococcus aureus produces a cocktail of metallophores (staphylopine, staphyloferrin A, and staphyloferrin B) to scavenge transition metals during infection of a host. In addition, S. aureus displays the extracellular surface lipoproteins FhuD1 and FhuD2 along with the ABC transporter complex FhuCBG to facilitate the use of hydroxamate xenosiderophores such as desferrioxamine B (DFOB) for iron acquisition. DFOB is used as a chelation therapy to treat human iron overload diseases and has been linked to an increased risk of S. aureus infections. We used a panel of synthetic DFOB analogs and a FhuD2-selective trihydroxamate sideromycin to probe xenosiderophore utilization in S. aureus and establish structure-activity relationships for Fe(III) binding, FhuD2 binding, S. aureus growth promotion, and competition for S. aureus cell entry. Fe(III) binding assays and FhuD2 intrinsic fluorescence quenching experiments revealed that diverse chemical modifications of the terminal ends of linear ferrioxamine siderophores influences Fe(III) affinity, but not FhuD2 binding. Siderophore-sideromycin competition assays and xenosiderophore growth promotion assays revealed that S. aureus SG511 and ATCC 11632 can distinguish between competing siderophores based exclusively on net charge of the siderophore-Fe(III) complex. Our work provides a roadmap for tuning hydroxamate xenosiderophore scaffolds to suppress (net negative charge) or enhance (net positive or neutral charge) uptake by S. aureus for applications in metal chelation therapy and siderophore-mediated antibiotic delivery, respectively.
Streptococcus agalactiae (Group B Streptococcus, GBS) is a Gram-positive bacterial pathogen that causes invasive infections of both children and adults. During pregnancy, GBS is a significant cause of infection of the fetal membranes (chorioamnionitis), which can lead to intra-amniotic infection, preterm birth, stillbirth, and neonatal sepsis. Recently, breastfeeding has been thought to represent a potential mode of GBS transmission from mother to newborn, which might increase the risk for late-onset sepsis. Little is known, however, about the molecular components of breast milk that may support or prevent GBS colonization. In this study, we examine how human milk oligosaccharides (HMOs) affect the pathogenesis of GBS. HMOs from discrete donor samples were isolated and profiled by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS). Growth and biofilm assays show that HMOs from mothers of specific milk groups can modulate the growth and biofilm formation of GBS. High-resolution field-emission gun scanning electron microscopy (SEM) and confocal laser scanning microscopy confirmed the quantitative biofilm assays and demonstrated cell arrangement perturbations in bacterial cultures treated with specific oligosaccharides. These findings demonstrate that HMOs affect the growth and cell biology of GBS. Finally, this study provides the first example of HMOs functioning as anti-biofilm agents against GBS.
The spread of antimicrobial resistance continues to be a priority health concern worldwide, necessitating exploration of alternative therapies. Cannabis sativa has long been known to contain antibacterial cannabinoids, but their potential to address antibiotic resistance has only been superficially investigated. Here, we show that cannabinoids exhibit antibacterial activity against MRSA, inhibit its ability to form biofilms and eradicate pre-formed biofilms and stationary phase cells persistent to antibiotics. We show that the mechanism of action of cannabigerol is through targeting the cytoplasmic membrane of Gram-positive bacteria and demonstrate in vivo efficacy of cannabigerol in a murine systemic infection model caused by MRSA. We also show that cannabinoids are effective against Gram-negative organisms whose outer membrane is permeabilized, where cannabigerol acts on the inner membrane. Finally, we demonstrate that cannabinoids work in combination with polymyxin B against multi-drug resistant Gram-negative pathogens, revealing the broad-spectrum therapeutic potential for cannabinoids.
Zika virus has emerged as a global concern because neither a vaccine nor antiviral compounds targeting it exist. A structure for the positive-sense RNA genome has not been established, leading us to look for potential G-quadruplex sequences (PQS) in the genome. The analysis identified >60 PQSs in the Zika genome. To minimize the PQS population, conserved sequences in the Flaviviridae family were found by sequence alignment, identifying seven PQSs in the prM, E, NS1, NS3, and NS5 genes. Next, alignment of 78 Zika strain genomes identified a unique PQS near the end of the 3'-UTR. Structural studies on the G-quadruplex sequences found four of the conserved Zika virus sequences to adopt stable, parallel-stranded folds that bind a G-quadruplex-specific compound, and one that was studied caused polymerase stalling when folded to a G-quadruplex. Targeting these PQSs with G-quadruplex binding molecules validated in previous clinical trials may represent a new approach for inhibiting viral replication.
Monoclonal antibody (mAb) therapeutics targeting cancer, autoimmune diseases, inflammatory diseases, and infectious diseases are growing exponentially. Although numerous panels of mAbs targeting infectious disease agents have been developed, their progression into clinically useful mAbs is often hindered by the lack of sequence information and/or loss of hybridoma cells that produce them. Here we combine the power of crystallography and mass spectrometry to determine the amino acid sequence and glycosylation modification of the Fab fragment of a potent human astrovirus-neutralizing mAb. We used this information to engineer a recombinant antibody single-chain variable fragment that has the same specificity as the parent monoclonal antibody to bind to the astrovirus capsid protein. This antibody can now potentially be developed as a therapeutic and diagnostic agent.
Enterohemorrhagic Escherichia coli O157:H7 presents a serious threat to human health and sanitation and is a leading cause in many food- and waterborne ailments. While conventional bacterial detection methods such as PCR, fluorescent immunoassays and ELISA exhibit high sensitivity and specificity, they are relatively laborious and require sophisticated instruments. In addition, these methods often demand extensive sample preparation and have lengthy readout times. We propose a simpler and more sensitive diagnostic technique featuring multiparametric magneto-fluorescent nanosensors (MFnS). Through a combination of magnetic relaxation and fluorescence measurements, our nanosensors are able to detect bacterial contamination with concentrations as little as 1 colony-forming unit (CFU). The magnetic relaxation property of our MFnS allow for sensitive screening at low target CFU, which is complemented by fluorescence measurements of higher CFU samples. Together, these qualities allow for the detection and quantification of broad-spectrum contaminations in samples ranging from aquatic reservoirs to commercially produced food.
Growing evidence suggests the importance of lipid metabolism in pathogenesis of tuberculosis. Neutral lipids form the majority of lipids in a caseous granuloma, pathology characteristic of tuberculosis. Cytosolic lipid droplets (LDs) of macrophages form the store house of these lipids, and have been demonstrated to contribute to the inflammatory response to infection. The proteome of lipid droplets reflects the mechanisms of lipid metabolism active under a condition. However, infection induced changes in the proteome of these dynamic organelles remains elusive. Here we employed quantitative proteomics to identify alterations induced upon infection with live Mycobacterium tuberculosis (Mtb) in comparison with heat killed bacilli or uninfected macrophages. We found increased abundance of proteins coupled with lipid metabolism, protein synthesis and vesicular transport function in LDs upon infection with live Mtb. Using biochemical methods and microscopy, we validated ARL8B to be increased on the lipid droplet surface of live Mtb infected macrophages and that ARL8B is a bonafide LD protein. This study provides the first proteomic evidence that the dynamic responses to infection also encompass changes at the level of LDs. This information will be important in understanding how Mtb manipulates lipid metabolism and defence mechanisms of the host macrophage.
There has been a very limited number of high throughput screening campaigns carried out with Leishmania drug targets. In part, this is due to the small number of suitable target genes that have been shown by genetic or chemical methods to be essential for the parasite. In this perspective, we discuss the state of genetic target validation in the field of Leishmania research and review the 200 Leishmania genes and 36 Trypanosoma cruzi genes for which gene deletion attempts have been made since the first published case in 1990. We define a quality score for the different genetic deletion techniques that can be used to identify potential drug targets. We also discuss how the advances in genome-scale gene disruption techniques have been used to assist target based and phenotypic based drug development in other parasitic protozoa and why Leishmania has lacked a similar approach so far. The prospects for this scale of work are considered in the context of the application of CRISPR/Cas9 gene editing as a useful tool in Leishmania.