Bacterial variability in the mammalian gut captured by a single-cell synthetic oscillator

Funder: Harvard Medical School; doi: https://doi.org/10.13039/100006691Funder: Wyss Institute for Biologically Inspired EngineeringAbstract: Synthetic gene oscillators have the potential to control timed functions and periodic gene expression in engineered cells. Such oscillators have been refined in bacteria in vitro, however, these systems have lacked the robustness and precision necessary for applications in complex in vivo environments, such as the mammalian gut. Here, we demonstrate the implementation of a synthetic oscillator capable of keeping robust time in the mouse gut over periods of days. The oscillations provide a marker of bacterial growth at a single-cell level enabling quantification of bacterial dynamics in response to inflammation and underlying variations in the gut microbiota. Our work directly detects increased bacterial growth heterogeneity during disease and differences between spatial niches in the gut, demonstrating the deployment of a precise engineered genetic oscillator in real-life settings

Author Correction: Bacterial variability in the mammalian gut captured by a single-cell synthetic oscillator (Nature Communications, (2019), 10, 1, (4665), 10.1038/s41467-019-12638-z)

In the original version of the Supplementary Information provided with this Article, Supplementary Table 1 incorrectly provided the details for strain LPT239 as “E. coli MC4100 + pLPT234 + pLPT145” and for strain LPT320 as “E. coli MC4100 + pLPT234 + pLPT41”. The details for strain LPT239 should be “E. coli MC4100 + pLPT234 + pLPT41” and for strain LPT320 should be “E. coli MC4100 + pLPT234 + pLPT145”. This has been corrected in the HMTL version of the article and the correct table appended below: Original (Table presented.)

High‐precision stable isotope analysis of <5 μg CaCO 3

Rationale Oxygen (δ18O) and carbon (δ13C) isotope analysis of foraminifera and other CaCO3 samples has been a key technique for paleoceanographical and paleoclimatological research for more than 60 years. There is ongoing demand for the analysis of ever smaller CaCO3 samples, driven, for example, by the desire to analyse single specimen planktic foraminifera, or small samples of tooth enamel. Methods We present a continuous‐flow mass spectrometric technique that uses cryo‐focusing of sample CO2 to analyse CaCO3 samples in a weight range between 10 and 3 μg. These are considerably lower sample weights than achievable on most currently available standard instrumentation. The technique is automated, so that sample throughput lies at >60 samples per day. The method involves an on‐line vial‐flushing routine designed to remove machine drift due to blank CO2 build‐up in the sample vials. Results In a series of experiments the effect of blank CO2 build‐up is quantified, and outgassing from the chlorobutyl septa identified as the source. An improved flushing routine together with the use of a cryo‐focusing step in the analysis is demonstrated to provide the analytical stability and sensitivity to analyse CaCO3 samples in a weight range between 10 and 3 μg at ≤0.1‰ precision (1σ) for both δ18O and δ13C values. The technique yields similarly precise results for the analysis of the structural carbonate fraction of small tooth enamel samples. Conclusions This study demonstrates that high‐precision oxygen and carbon isotope analysis is possible on CaCO3 samples smaller than 5 μg by use of a continuous‐flow isotope technique. Of key importance are (1) the application of a cold trap that drastically reduces sample gas loss, and (2) a modified flushing regime that eliminates increasing background CO2 build‐up in sample vials during longer automated sample runs

The Biology and Management of Cartilaginous Tumors: A Role For Targeting Isocitrate Dehydrogenase

Chondrosarcomas are rare mesenchymal neoplasms defined by the production of abnormal cartilaginous matrix. Conventional chondrosarcoma is the most common histology. The management of primary conventional chondrosarcoma generally is surgical with the possible addition of radiation therapy. Treatment of conventional chondrosarcoma is problematic in unresectable or metastatic disease because the tumors tend to be resistant to standard sarcoma chemotherapy regimens. Previous attempts at targeted therapy, including inhibitors of Hedgehog signaling, the mTOR pathway, and platelet-derived growth factor receptor (PDGFR) have been largely disappointing. However, heterozygous mutations in isocitrate dehydrogenase (IDH) enzymes recently have been identified in chondrogenic neoplasms, with mutations reported in approximately 87% of benign enchondromas, 70% of conventional chondrosarcomas, and 54% of dedifferentiated chondrosarcomas. The normal IDH protein continues to produce alpha-ketoglutarate (alpha-KG) whereas the mutant IDH protein converts KG to the oncometabolite 2-hydroxyglutarate (2-HG). Clinical trials of novel IDH inhibitors are ongoing, with evidence of early activity in IDH-mutant leukemias. IDH inhibitors show antitumor effects against IDH-mutant chondrosarcoma cell lines, supporting the inclusion of patients with chondrosarcoma with IDH mutations on IDH inhibitor clinical trials for solid tumors. Targeting IDH mutations may offer hope of a novel antineoplastic strategy not only for patients with chondrosarcomas, but also for other solid tumors with aberrant IDH activity