The evolution of spectrum in antibiotics and bacteriocins.

A key property of many antibiotics is that they will kill or inhibit a diverse range of microbial species. This broad-spectrum of activity has its evolutionary roots in ecological competition, whereby bacteria and other microbes use antibiotics to suppress other strains and species. However, many bacteria also use narrow-spectrum toxins, such as bacteriocins, that principally target conspecifics. Why has such a diversity in spectrum evolved? Here, we develop an evolutionary model to understand antimicrobial spectrum. Our first model recapitulates the intuition that broad-spectrum is best, because it enables a microbe to kill a wider diversity of competitors. However, this model neglects an important property of antimicrobials: They are commonly bound, sequestered, or degraded by the cells they target. Incorporating this toxin loss reveals a major advantage to narrow-spectrum toxins: They target the strongest ecological competitor and avoid being used up on less important species. Why then would broad-spectrum toxins ever evolve? Our model predicts that broad-spectrum toxins will be favored by natural selection if a strain is highly abundant and can overpower both its key competitor and other species. We test this prediction by compiling and analyzing a database of the regulation and spectrum of toxins used in inter-bacterial competition. This analysis reveals a strong association between broad-spectrum toxins and density-dependent regulation, indicating that they are indeed used when strains are abundant. Our work provides a rationale for why bacteria commonly evolve narrow-spectrum toxins such as bacteriocins and suggests that the evolution of antibiotics proper is a signature of ecological dominance.

Microcin MccI47 selectively inhibits enteric bacteria and reduces carbapenem-resistant Klebsiella pneumoniae colonization in vivo when administered via an engineered live biotherapeutic.

The gastrointestinal (GI) tract is the reservoir for multidrug resistant (MDR) pathogens, specifically carbapenem-resistant (CR) Klebsiella pneumoniae and other Enterobacteriaceae, which often lead to the spread of antimicrobial resistance genes, severe extraintestinal infections, and lethal outcomes. Selective GI decolonization has been proposed as a new strategy for preventing transmission to other body sites and minimizing spreading to susceptible individuals. Here, we purify the to-date uncharacterized class IIb microcin I47 (MccI47) and demonstrate potent inhibition of numerous Enterobacteriaceae, including multidrug-resistant clinical isolates, in vitro at concentrations resembling those of commonly prescribed antibiotics. We then genetically modify the probiotic bacterium Escherichia coli Nissle 1917 (EcN) to produce MccI47 from a stable multicopy plasmid by using MccI47 toxin production in a counterselection mechanism to engineer one of the native EcN plasmids, which renders provisions for inducible expression and plasmid selection unnecessary. We then test the clinical relevance of the MccI47-producing engineered EcN in a murine CR K. pneumoniae colonization model and demonstrate significant MccI47-dependent reduction of CR K. pneumoniae abundance after seven days of daily oral live biotherapeutic administration without disruption of the resident microbiota. This study provides the first demonstration of MccI47 as a potent antimicrobial against certain Enterobacteriaceae, and its ability to significantly reduce the abundance of CR K. pneumoniae in a preclinical animal model, when delivered from an engineered live biotherapeutic product. This study serves as the foundational step toward the use of engineered live biotherapeutic products aimed at the selective removal of MDR pathogens from the GI tract.

Bacterial species rarely work together.

Competition is prevalent and could be harnessed as an alternative to antibiotics.

An overview of Salmonella enterica metal homeostasis pathways during infection.

Nutritional immunity is a powerful strategy at the core of the battlefield between host survival and pathogen proliferation. A host can prevent pathogens from accessing biological metals such as Mg, Fe, Zn, Mn, Cu, Co or Ni, or actively intoxicate them with metal overload. While the importance of metal homeostasis for the enteric pathogen Salmonella enterica Typhimurium was demonstrated many decades ago, inconsistent results across various mouse models, diverse Salmonella genotypes, and differing infection routes challenge aspects of our understanding of this phenomenon. With expanding access to CRISPR-Cas9 for host genome manipulation, it is now pertinent to re-visit past results in the context of specific mouse models, identify gaps and incongruities in current knowledge landscape of Salmonella homeostasis, and recommend a straight path forward towards a more universal understanding of this historic host-microbe relationship.

A Bioinspired Molybdenum Catalyst for Aqueous Perchlorate Reduction.

Perchlorate (ClO4-) is a pervasive, harmful, and inert anion on both Earth and Mars. Current technologies for ClO4- reduction entail either harsh conditions or multicomponent enzymatic processes. Herein, we report a heterogeneous (L)Mo-Pd/C catalyst directly prepared from Na2MoO4, a bidentate nitrogen ligand (L), and Pd/C to reduce aqueous ClO4- into Cl- with 1 atm of H2 at room temperature. A suite of instrument characterizations and probing reactions suggest that the MoVI precursor and L at the optimal 1:1 ratio are transformed in situ into oligomeric MoIV active sites at the carbon-water interface. For each Mo site, the initial turnover frequency (TOF0) for oxygen atom transfer from ClOx- substrates reached 165 h-1. The turnover number (TON) reached 3840 after a single batch reduction of 100 mM ClO4-. This study provides a water-compatible, efficient, and robust catalyst to degrade and utilize ClO4- for water purification and space exploration.

SATB2 and MDM2 Immunoexpression and Diagnostic Role in Primary Osteosarcomas of the Jaw.

Primary osteosarcomas of the jaw (OSJ) are rare, accounting for 6% of all osteosarcomas. This study aims to determine the value of SATB2 and MDM2 immunohistochemistry (IHC) in differentiating OSJ from other jawbone mimickers, such as benign fibro-osseous lesions (BFOLs) of the jaw or Ewing sarcoma of the jaw. Certain subsets of osteosarcoma harbor a supernumerary ring and/or giant marker chromosomes with amplification of the 12q13-15 region, including the murine double-minute type 2 (MDM2) and cyclin-dependent kinase 4 (CDK4) genes. Special AT-rich sequence-binding protein 2 (SATB2) is an immunophenotypic marker for osteoblastic differentiation. Cases of OSJ, BFOLs (ossifying fibroma and fibrous dysplasia) of the jaw, and Ewing sarcoma of the jaw were retrieved from the Departments of Oral Pathology and Oral Medicine, Faculty of Dentistry, Obafemi Awolowo University and Lagos State University College of Medicine, Nigeria. All OSJ retrieved showed histologic features of high-grade osteosarcoma. IHC for SATB2 (clone EP281) and MDM2 (clone IF2), as well as fluorescence in situ hybridization (FISH) for MDM2 amplification, were performed on all cases. SATB2 was expressed in a strong intensity and diffuse staining pattern in all cases (11 OSJ, including a small-cell variant, 7 ossifying fibromas, and 5 fibrous dysplasias) except in Ewing sarcoma, where it was negative in neoplastic cells. MDM2 was expressed in a weak to moderate intensity and scattered focal to limited diffuse staining pattern in 27% (3/11) of cases of OSJ and negative in all BFOLs and the Ewing sarcoma. MDM2 amplification was negative by FISH in interpretable cases. In conclusion, the three cases of high-grade OSJs that expressed MDM2 may have undergone transformation from a low-grade osteosarcoma (LGOS). SATB2 is not a dependable diagnostic marker to differentiate OSJ from BFOLs of the jaw; however, it could serve as a valuable diagnostic marker in differentiating the small-cell variant of OSJ from Ewing sarcoma of the jaw, while MDM2 may be a useful diagnostic marker in differentiating OSJ from BFOLs of the jaw, especially in the case of an LGOS or high-grade transformed osteosarcoma.

Microcin H47: A Class IIb Microcin with Potent Activity Against Multidrug Resistant Enterobacteriaceae.

Microcin H47 (MccH47) is an antimicrobial peptide produced by some strains of Escherichia coli that has demonstrated inhibitory activity against enteric pathogens in vivo and has been heterologously overexpressed in proof-of-concept engineered probiotic applications. While most studies clearly demonstrate inhibitory activity against E. coli isolates, there are conflicting results on the qualitative capacity for MccH47 to inhibit strains of Salmonella. Here, we rectify these inconsistencies via the overexpression and purification of a form of MccH47, termed MccH47-monoglycosylated enterobactin (MccH47-MGE). We then use purified MccH47 to estimate minimum inhibitory concentrations (MICs) against a number of medically relevant Enterobacteriaceae, including Salmonella and numerous multidrug resistant (MDR) strains. While previous reports suggested that the spectrum of activity for MccH47 is quite narrow and restricted to activity against E. coli, our data demonstrate that MccH47 has broad and potent activity within the Enterobacteriaceae family, suggesting it as a candidate for further development toward treating MDR enteric infections.

Engineered Probiotic for the Inhibition of Salmonella via Tetrathionate-Induced Production of Microcin H47.

Complications arising from antibiotic-resistant bacteria are becoming one of the key issues in modern medicine. Members of drug-resistant Enterobacteriaceae spp. include opportunistic pathogens (e.g., Salmonella spp.) that are among the leading causes of morbidity and mortality worldwide. Overgrowth of these bacteria is considered a hallmark of intestinal dysbiosis. Microcins (small antimicrobial peptides) produced by some gut commensals can potentially cure these conditions by inhibiting these pathogens and have been proposed as a viable alternative to antibiotic treatment. In this proof-of-concept work, we leverage this idea to develop a genetically engineered prototype probiotic to inhibit Salmonella spp. upon exposure to tetrathionate, a molecule produced in the inflamed gut during the course of Salmonella infection. We developed a plasmid-based system capable of conferring the ability to detect and utilize tetrathionate, while at the same time producing microcin H47. We transferred this plasmid-based system to Escherichia coli and demonstrated the ability of the engineered strain to inhibit growth of Salmonella in anaerobic conditions while in the presence of tetrathionate, with no detectable inhibition in the absence of tetrathionate. In direct competition assays between the engineered E. coli and Salmonella, the engineered E. coli had a considerable increase in fitness advantage in the presence of 1 mM tetrathionate as compared to the absence of tetrathionate. In this work, we have demonstrated the ability to engineer a strain of E. coli capable of using an environmental signal indicative of intestinal inflammation as an inducing molecule, resulting in production of a microcin capable of inhibiting the organism responsible for the inflammation.

Remote beam output audits: a global assessment of results out of tolerance.

BACKGROUND AND PURPOSE: Remote beam output audits, which independently measure an institution's machine calibration, are a common component of independent radiotherapy peer review. This work reviews the results and trends of these audit results across several organisations and geographical regions. MATERIALS AND METHODS: Beam output audit results from the Australian Clinical Dosimetry Services, International Atomic Energy Agency, Imaging and Radiation Oncology Core, and Radiation Dosimetry Services were evaluated from 2010 to the present. The rate of audit results outside a +/-5% tolerance was evaluated for photon and electron beams as a function of the year of irradiation and nominal beam energy. Additionally, examples of confirmed calibration errors were examined to provide guidance to clinical physicists and auditing bodies. RESULTS: Of the 210,167 audit results, 1323 (0.63%) were outside of tolerance. There was a clear trend of improved audit performance for more recent dates, and while all photon energies generally showed uniform rates of results out of tolerance, low (6 MeV) and high (≥18 MeV) energy electron beams showed significantly elevated rates. Twenty nine confirmed calibration errors were explored and attributed to a range of issues, such as equipment failures, errors in setup, and errors in performing the clinical reference calibration. Forty-two percent of these confirmed errors were detected during ongoing periodic monitoring, and not at the time of the first audit of the machine. CONCLUSIONS: Remote beam output audits have identified, and continue to identify, numerous and often substantial beam calibration errors.

Motion of the esophagus due to cardiac motion.

When imaging studies (e.g. CT) are used to quantify morphological changes in an anatomical structure, it is necessary to understand the extent and source of motion which can give imaging artifacts (e.g. blurring or local distortion). The objective of this study was to assess the magnitude of esophageal motion due to cardiac motion. We used retrospective electrocardiogram-gated contrast-enhanced computed tomography angiography images for this study. The anatomic region from the carina to the bottom of the heart was taken at deep-inspiration breath hold with the patients' arms raised above their shoulders, in a position similar to that used for radiation therapy. The esophagus was delineated on the diastolic phase of cardiac motion, and deformable registration was used to sequentially deform the images in nearest-neighbor phases among the 10 cardiac phases, starting from the diastolic phase. Using the 10 deformation fields generated from the deformable registration, the magnitude of the extreme displacements was then calculated for each voxel, and the mean and maximum displacement was calculated for each computed tomography slice for each patient. The average maximum esophageal displacement due to cardiac motion for all patients was 5.8 mm (standard deviation: 1.6 mm, maximum: 10.0 mm) in the transverse direction. For 21 of 26 patients, the largest esophageal motion was found in the inferior region of the heart; for the other patients, esophageal motion was approximately independent of superior-inferior position. The esophagus motion was larger at cardiac phases where the electrocardiogram R-wave occurs. In conclusion, the magnitude of esophageal motion near the heart due to cardiac motion is similar to that due to other sources of motion, including respiratory motion and intra-fraction motion. A larger cardiac motion will result into larger esophagus motion in a cardiac cycle.

Low bone mineral density in highly trained male master cyclists.

The purpose of this study was to determine total and regional bone mineral density (BMD) in highly competitive young adult and master male cyclists. Three groups of men were studied: older cyclists (51.2+/-5.3 years, n=27); young adult cyclists (31.7+/-3.5 years, n=16); and 24 non-athletes matched by age (+/-2 years) and body weight (+/-2 kg) to the master cyclists. All of the master cyclists had been training and racing for a minimum of 10 years (mean 20.2+/-8.4 years) and engaging in little to no weight-bearing exercise. The younger cyclists also engaged in little weight-bearing exercise and had been training and racing for 10.9+/-3.2 years. Age-matched controls were normally active. The History of Leisure Activity Questionnaire was used to determine the influence on BMD of self-reported total and weight-bearing exercise during three periods of life: 12-18 years, 19-34 years, and 35-49 years. BMD (measured by DXA) of the spine (L2-L4) and total hip was significantly (P<0.033) lower in the master cyclists compared to both age-matched controls and young adult cyclists. Total body BMD was lower in the master cyclists compared to the young-adults (P<0.033). Furthermore, four (15%) of the master cyclists, but none of the men in the other groups, had T-scores (spine and/or hip) lower than -2.5. Weight-bearing exercise performed during teen and young adult years did not appear to influence BMD, as there were no differences at any site between those within the upper and lower 50th percentiles for weight-bearing exercise during the 12-18, 19-34, or 35-49 year time periods. These data indicate that master cyclists with a long history of training exclusively in cycling have low BMD compared to their age-matched peers. Although highly trained and physically fit, these athletes may be at high risk for developing osteoporosis with advancing age.