Experience with periprosthetic infection after limb salvage surgery for patients with osteosarcoma.

BACKGROUND: The rate of postoperative infection developing is higher after limb salvage surgery (LSS) following sarcoma resection compared with conventional arthroplasty. The goal of this study is to summarize our experience in management of periprosthetic joint infection (PJI) and the risk factors of early PJI after LSS. METHODS: Between January 2010 and July 2019, 53 patients with osteosarcoma in the lower extremities who encountered periprosthetic infection after segmental tumor endoprosthetic replacement in our center were analyzed. Detailed patient characteristics and therapeutic information were collected from database of our institution or follow-up data and we divided patients according to the interval time between infection and tumor resection (surgery-infection interval) and investigate potential risk factors. RESULTS: A total of 53 (5.08%) patients were suffered postoperative infection. The average interval between surgery and clinical signs of deep infections are 27.5 days. For the drainage culture, positive results were only presented in 11 patients (20.8%). Almost half of this study's (47.2%) patients underwent a traditional two-stage revision, that was, after the removal of the infected prosthesis, we applied antibiotic-loaded bone cements as a spacer. The mean blood loss during initial implantation surgery and operation time both correlated with interval period between PJI and initial implantation significantly (P = 0.028, P = 0.046). For several patients which infection marker was hardly back to normal after spacer implantation, we conservatively introduced an improved combination of bone cement and prosthesis for the second-stage surgery (5.6%). There were six patients needing re-operation, of which three were due to the aseptic loosening of the prosthesis, one developed periprosthetic infection again, and two patients encountered local recurrence and underwent amputation. Two patients were dead from distal metastasis. CONCLUSIONS: A two-stage revision strategy remains effective and standardized methods for PJI patients. Total operation time and blood loss during LSS of osteosarcoma are the main risk factors of early PJI. For the patients without confirmed eradiation of microorganisms, an improved combination of bone cement and prosthesis applied in the second-stage surgery could achieve satisfied functional and oncologic results.

CRISPRi-Based Dynamic Control of Carbon Flow for Efficient N-Acetyl Glucosamine Production and Its Metabolomic Effects in Escherichia coli.

Carbon competition between cell growth and product synthesis is the bottleneck in efficient N-acetyl glucosamine (GlcNAc) production in microbial cell factories. In this study, a xylose-induced T7 RNA polymerase-PT7 promoter system was introduced in Escherichia coli W3110 to control the GlcNAc synthesis. Meanwhile, an arabinose-induced CRISPR interference (CRISPRi) system was applied to adjust cell growth by attenuating the transcription of key growth-related genes. By designing proper sgRNAs, followed by elaborate adjustment of the addition time and concentration of the two inducers, the carbon flux between cell growth and GlcNAc synthesis was precisely redistributed. Comparative metabolomics analysis results confirmed that the repression of pfkA and zwf significantly attenuated the TCA cycle and the synthesis of related amino acids, saving more carbon for the GlcNAc synthesis. Finally, the simultaneous repression of pfkA and zwf in strain GLA-14 increased the GlcNAc titer by 47.6% compared with that in E. coli without the CRISPRi system in a shake flask. GLA-14 could produce 90.9 g/L GlcNAc within 40 h in a 5 L bioreactor, with a high productivity of 2.27 g/L/h. This dynamic strategy for rebalancing cell growth and product synthesis could be applied in the fermentative production of other chemicals derived from precursors synthesized via central carbon metabolism.

Comparative metabolomic analysis reveals different evolutionary mechanisms for branched-chain amino acids production.

Evolution is a powerful tool for the breeding of microorganisms, while the connection between the changes of intracellular metabolism and different evolution directions is still unclear, which once clarified, will greatly expand the application of evolutionary engineering. We aim to clarify the correlation between metabolism changes and evolution directions in two Corynebacterium glutamicum strains for L-valine and L-leucine overproducing originated from the same parental strain by repeated random mutagenesis and selection. GC-MS metabolomics was performed to identify and quantify intracellular metabolites of the evolved and wild-type C. glutamicum strains. Time-series comparison of the fermentation processes was performed. The metabolism differences of three strains mainly exist in central carbon metabolism and the stress-resisting modes. C. glutamicum XV developed an overall "pyruvate-saving" mode for L-valine synthesis, and adopted a trehalose accumulating strategy to resist environmental stresses. C. glutamicum CP depended on an enhanced "pyruvate-producing" mode, together with certain "pyruvate-saving" strategies, for efficient L-leucine synthesis, and accumulated proline, my-inositol, and inositol as the stress-resisting measure. These elaborate regulation strategies could be used in future metabolic engineering, making evolution more informative and applicable.