Biology subjects

Burnham, C.-A. D.

Publications and source records attributed to Burnham, C.-A. D..

4 recordsLinked to original sources

bioRxiv2023

Secretory Leukocyte Protease Inhibitor Protects Against Severe Urinary Tract Infection in Mice

Millions suffer from urinary tract infections (UTIs) worldwide every year with women accounting for the majority of cases. Uropathogenic Escherichia coli (UPEC) causes most of these primary infections and leads to 25% becoming recurrent or chronic. To repel invading pathogens, the urinary tract mounts a vigorous innate immune response that includes the secretion of antimicrobial peptides (AMPs), rapid recruitment of phagocytes and exfoliation of superficial umbrella cells. Here, we investigate secretory leukocyte protease inhibitor (SLPI), an AMP with antiprotease, antimicrobial and immunomodulatory functions, known to play protective roles at other mucosal sites, but not well characterized in UTIs. Using a mouse model of UPEC-caused UTI, we show that urine SLPI increases in infected mice and that SLPI is localized to bladder epithelial cells. UPEC infected SLPI-deficient (Slpi-/-) mice suffer from higher urine bacterial burdens, prolonged bladder inflammation, and elevated urine neutrophil elastase (NE) levels compared to wild-type (Slpi+/+) controls. Combined with bulk bladder RNA sequencing, our data indicate that Slpi-/- mice have a dysregulated immune and tissue repair response following UTI. We also measure SLPI in urine samples from a small group of female subjects 18-49 years old and find that SLPI tends to be higher in the presence of a uropathogen, except in patients with history of recent or recurrent UTI (rUTI), suggesting a dysregulation of SLPI expression in these women. Taken together, our findings show SLPI protects against acute UTI in mice and provides preliminary evidence that SLPI is likewise regulated in response to uropathogen exposure in women.

immunology↗

bioRxiv2020

Antimicrobial prodrug activation by the staphylococcal glyoxalase GloB

With the rising prevalence of multidrug-resistance, there is an urgent need to develop novel antibiotics. Many putative antibiotics demonstrate promising in vitro potency but fail in vivo due to poor drug-like qualities (e.g. serum half-life, oral absorption, solubility, toxicity). These drug-like properties can be modified through the addition of chemical protecting groups, creating "prodrugs" that are activated prior to target inhibition. Lipophilic prodrugging techniques, including the attachment of a pivaloyloxymethyl group, have garnered attention for their ability to increase cellular permeability by masking charged residues and the relative ease of the chemical prodrugging process. Unfortunately, pivaloyloxymethyl prodrugs are rapidly activated by human sera, rendering any membrane permeability qualities absent during clinical treatment. Identification of the bacterial prodrug activation pathway(s) will allow for the development of host-stable and microbe-targeted prodrug therapies. Here, we use two zoonotic staphylococcal species, S. schleiferi and S. pseudintermedius, to establish the mechanism of carboxy ester prodrug activation. Using a forward genetic screen, we identify a conserved locus in both species encoding the enzyme hydroxyacylglutathione hydrolase (GloB), whose loss-of-function confers resistance to carboxy ester prodrugs. We enzymatically characterize GloB and demonstrate that it is a functional glyoxalase II enzyme, which has the capacity to activate carboxy ester prodrugs. As GloB homologs are both widespread and diverse in sequence, our findings suggest that GloB may be a useful mechanism for developing species-or genus-level prodrug targeting strategies.

microbiology↗

bioRxiv2019

Improving characterization of understudied human microbiomes using targeted phylogenetics

Whole genome bacterial sequences are required to better understand microbial functions, niches-pecific bacterial metabolism, and disease states. Although genomic sequences are available for many of the human-associated bacteria from commonly tested body habitats (e.g. stool), as few as 13% of bacterial-derived reads from other sites such as the skin map to known bacterial genomes. To facilitate a better characterization of metagenomic shotgun reads from under-represented body sites, we collected over 10,000 bacterial isolates originating from 14 human body habitats, identified novel taxonomic groups based on full length 16S rRNA sequences, clustered the sequences to ensure that no individual taxonomic group was over-selected for sequencing, prioritized bacteria from under-represented body sites (such as skin, respiratory and urinary tract), and sequenced and assembled genomes for 665 new bacterial strains. Here we show that addition of these genomes improved read mapping rates of HMP metagenomic samples by nearly 30% for the previously under-represented phylum Fusobacteria, and 27.5% of the novel genomes generated here had high representation in at least one of the tested HMP samples, compared to 12.5% of the sequences in the public databases, indicating an enrichment of useful novel genomic sequences resulting from the prioritization procedure. As our understanding of the human microbiome continues to improve and to enter the realm of therapy developments, targeted approaches such as this to improve genomic databases will increase in importance from both an academic and clinical perspective. ImportanceThe human microbiome plays a critically important role in health and disease, but current understanding of the mechanisms underlying the interactions between the varying microbiome and the different host environments is lacking. Having access to a database of fully sequenced bacterial genomes provides invaluable insights into microbial functions, but currently sequenced genomes for the human microbiome have largely come from a limited number of body sites (primarily stool), while other sites such as the skin, respiratory tract and urinary tracts are under-represented, resulting in as little as 13% of bacterial-derived reads mapping to known bacterial genomes. Here, we sequenced and assembled 665 new bacterial genomes, prioritized from a larger database to select under-represented body sites and bacterial taxa in the existing databases. As a result, we substantially improve mapping rates for samples from the Human Microbiome Project and provide an important contribution to human bacterial genomic databases for future studies.

microbiology↗

bioRxiv2019

Potent, specific MEPicides for treatment of zoonotic staphylococci

Coagulase-positive staphylococci, which frequently colonize the mucosal surfaces of animals, also cause a spectrum of opportunistic infections including skin and soft tissue infections, urinary tract infections, pneumonia, and bacteremia. However, recent advances in bacterial identification have revealed that these common veterinary pathogens are in fact, zoonoses that cause serious infections in human patients. The global spread of multidrug-resistant zoonotic staphylococci, in particular the emergence of methicillin-resistant organisms, is now a serious threat to both animal and human welfare. Accordingly, new therapeutic targets that can be exploited to combat staphylococcal infections are urgently needed. Enzymes of the methylerythritol phosphate pathway (MEP) of isoprenoid biosynthesis represent potential targets for treating zoonotic staphylococci. Here we demonstrate that fosmidomycin (FSM) inhibits the first step of the isoprenoid biosynthetic pathway catalyzed by deoxyxylulose phosphate reductoisomerase (DXR) in staphylococci. In addition, we have both enzymatically and structurally determined the mechanism by which FSM elicits its effect. Using a forward genetic screen, the glycerol-3-phosphate transporter GlpT that facilitates FSM uptake was identified in two zoonotic staphylococci, Staphylococcus schleiferi and Staphylococcus pseudintermedius. A series of lipophilic ester prodrugs (termed MEPicides) structurally related to FSM were synthesized, and data indicate that the presence of the prodrug moiety not only substantially increased potency of the inhibitors against staphylococci, but also bypassed the need for GlpT-mediated cellular transport. Collectively, our data indicate that the prodrug MEPicides selectively and robustly inhibit DXR in zoonotic staphylococci, and further, DXR represents a promising, druggable target for future development.\n\nAuthor SummaryThe proliferation of microbial pathogens resistant to the current pool of antibiotics is a major threat to public health. Drug resistance is pervasive in staphylococci, including several species that can cause serious zoonotic infections in humans. Thus, new antimicrobial agents are urgently need to combat these life-threatening, resistant infections. Here we establish the MEP pathway as a promising new target against zoonotic staphylococci. We determine that fosmidomycin (FSM) selectively targets the isoprenoid biosynthesis pathway in zoonotic staphylococci, and use forward genetics to identify the transporter that facilitates phosphonate antibiotic uptake. Employing this knowledge, we synthesized a series of potent antibacterial prodrugs that circumvent the transporter. Together, these novel prodrug inhibitors represent promising leads for further drug development against zoonotic staphylococci.

microbiology↗