Bioavailable levels of trace metals, such as iron and zinc, for bacterial growth in nature are sufficiently low that most microbes have evolved high-affinity binding and transport systems. The microbe Campylobacter jejuni lives in the gastrointestinal tract of chickens, the principal source of human infection. A high-affinity ABC transporter for zinc uptake is required for Campylobacter survival in chicken intestines in the presence of a normal microbiota but not when chickens are raised with a limited microbiota. Mass spectrometric analysis of cecal contents revealed the presence of numerous zinc-binding proteins in conventional chicks compared to the number in limited-microbiota chicks. The presence of a microbiota results in the production of host zinc-binding enzymes, causing a growth restriction for bacteria that lack the high-affinity zinc transporter. Such transporters in a wide range of pathogenic bacteria make them good targets for the development of broad-spectrum antimicrobials.Importance Zinc is an essential trace element for the growth of most organisms. Quantities of zinc inside cells are highly regulated, as too little zinc does not support growth, while too much zinc is toxic. Numerous bacterial cells require zinc uptake systems for growth and virulence. The work presented here demonstrates that the microbiota in the gastrointestinal tract reduces the quantity of zinc. Without a high-affinity zinc transporter, Campylobacter jejuni, a commensal organism of chickens, is unable to replicate or colonize the gastrointestinal tract. This is the first demonstration of zinc competition between microbiota in the gastrointestinal tract of a host. These results could have profound implications in the field of microbial pathogenesis and in our understanding of host metabolism and the microbiota.
SUMMARY Pneumonic plague is a deadly respiratory disease caused by Yersinia pestis. The bacterial protease Pla contributes to disease progression and manipulation of host immunity, but the mechanisms by which this occurs are largely unknown. Here we show that Pla degrades the apoptotic signaling molecule Fas ligand (FasL) to prevent host cell apoptosis and inflammation. Wild-type Y. pestis, but not a Pla mutant (Δpla), degrades FasL, which results in decreased downstream caspase-3/7 activation and reduced apoptosis. Similarly, lungs of mice challenged with wild-type Y. pestis show reduced levels of FasL and activated caspase-3/7 compared to Δpla infection. Consistent with a role for FasL in regulating immune responses, Δpla infection results in aberrant pro-inflammatory cytokine levels. The loss of FasL or inhibition of caspase activity alters host inflammatory responses and enables enhanced Y. pestis outgrowth in the lungs. Thus, by degrading FasL, Y. pestis manipulates host cell death pathways to facilitate infection.
Many Gram-negative bacteria produce outer membrane vesicles (OMVs) during cell growth and division, and some bacterial pathogens deliver virulence factors to the host via the release of OMVs during infection. Here we show that Yersinia pestis, the causative agent of the disease plague, produces and releases native OMVs under physiological conditions. These OMVs, approximately 100 nm in diameter, contain multiple virulence-associated outer membrane proteins including the adhesin Ail, the F1 outer fimbrial antigen, and the protease Pla. We found that OMVs released by Y. pestis contain catalytically active Pla that is competent for plasminogen activation and α2-antiplasmin degradation. The abundance of OMV-associated proteins released by Y. pestis is significantly elevated at 37°C compared to 26°C and is increased in response to membrane stress and mutations in RseA, Hfq, and the major Braun lipoprotein (Lpp). In addition, we show that Y. pestis OMVs are able to bind to components of the extracellular matrix such as fibronectin and laminin. These data suggest that Y. pestis may produce OMVs during mammalian infection and we propose that dispersal of Pla via OMV release may influence the outcome of infection through interactions with Pla substrates such as plasminogen and Fas ligand.
Direct transmission of bacteria to subsequent generations highlights the beneficial nature of host-bacteria relationships. In insects, this process is often mediated by the production of microbe-containing secretions. The objective of this study was to determine if the burying beetle, Nicrophorus defodiens, utilizes anal secretions to transmit adult digestive tract bacteria onto a small vertebrate carcass; thus creating the potential to aid in carcass preservation or pass digestive tract bacteria to their larval offspring. Using high-throughput Illumina sequencing of the 16S rRNA gene, we characterized bacterial communities of adult beetle digestive tracts, their anal secretions, and prepared mouse carcasses. We also examined unprepared carcass bacterial communities as a means to interpret community shifts that take place during carcass preservation. We found a vast reduction in diversity on prepared carcasses after anal secretion application. Overall, there was little similarity in bacterial communities among adult digestive tracts, anal secretions, and prepared carcasses, suggesting bacterial communities found in adult digestive tracts do not successfully colonize and achieve dominance on prepared carcasses by way of beetle anal secretions. We concluded that N. defodiens does not transmit their digestive tract bacterial communities to prepared carcasses in a wholesale manner, but may transmit key microbes, including core microbiome members, to preserved carcasses that may ultimately act to sustain larvae and serve as inocula for larval digestive tracts.
ObjectiveThe expansion of molecular techniques in medical diagnosis, forensics, and education requires the development of improved techniques of DNA extraction from fixed tissues. Cadaver tissues are not commonly used for genetic analysis due to DNA degradation resulting from the embalming fixation. Modification of existing techniques of tissue disruption combined with phenol–chloroform treatment was done to produce an efficient method of extracting amplifiable DNA of high quality and quantity from non-paraffin embedded embalmed cadaver tissue.ResultsTissues (cerebellum, cerebral cortex, heart, and bone) from four cadavers were used to develop a procedure for DNA isolation, which includes a high heat treatment. The location and age of the tissue had a significant effect on the quantity of DNA recovered. Targeted PCR amplification of the Apolipoprotein gene was used to assess the efficacy of genotypic analysis from the recovered DNA. We report the development of a simple, reliable, and low-cost method of DNA isolation utilizing brain tissue from embalmed tissues that could be used for PCR amplification and genetic analysis.Electronic supplementary materialThe online version of this article (10.1186/s13104-017-3066-y) contains supplementary material, which is available to authorized users.
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