In situ NIR spectroscopy monitoring of plasmid production processes effect of producing strain, medium composition and the cultivation strategy

BACKGROUNDWhile the pharmaceutical industry keeps an eye on plasmid DNA production for new generation gene therapies, real-time monitoring techniques for plasmid bioproduction are as yet unavailable. This work shows the possibility of in situ monitoring of plasmid production in Escherichia coli cultures using a near infrared (NIR) fiber optic probe. RESULTSPartial least squares (PLS) regression models based on the NIR spectra were developed for predicting bioprocess critical variables such as the concentrations of biomass, plasmid, carbon sources (glucose and glycerol) and acetate. In order to achieve robust models able to predict the performance of plasmid production processes, independently of the composition of the cultivation medium, cultivation strategy (batch versus fed-batch) and E. coli strain used, three strategies were adopted, using: (i) E. coliDH5 cultures conducted under different media compositions and culture strategies (batch and fed-batch); (ii) engineered E. coli strains, MG1655endArecApgi and MG1655endArecA, grown on the same medium and culture strategy; (iii) diverse E. coli strains, over batch and fed-batch cultivations and using different media compositions. PLS models showed high accuracy for predicting all variables in the three groups of cultures. CONCLUSIONNIR spectroscopy combined with PLS modeling provides a fast, inexpensive and contamination-free technique to accurately monitoring plasmid bioprocesses in real time, independently of the medium composition, cultivation strategy and the E. coli strain used.

Resting energy expenditure, macronutrient utilization, and body composition in term infants after corrective surgery of major congenital anomalies: a case-study

Introduction - Knowledge on the metabolic changes and nutritional needs during the postsurgical anabolic phase in infants is scarce. This analysis explores the associations of resting energy expenditure (REE) and macronutrient utilization with body composition of full-term infants, during catch-up growth after corrective surgery of major congenital anomalies. Methods - A cohort of full-term appropriate for-gestational-age neonates subjected to corrective surgery of major congenital anomalies were recruited after gaining weight for at least one week. REE and macronutrient utilization, measured by respiratory quotient (RQ), were assessed by indirect calorimetry using the Deltatrac II Metabolic Monitor ®. Body composition, expressed as fat-free mass (FFM), fat mass (FM) and adiposity defined as percentage of FM (% FM), was measured by air displacement plethysmography using the Pea Pod ®. Results - Four infants were included at 3 to 5 postnatal weeks. Recommended energy and macronutrient intakes for healthy term infants were provided. Through the study, the median (min-max) REE (Kcal/Kg FFM/d) was 70.8 (60.6-96.1) and RQ was 0.99 (0.72-1.20). Steady increases in both body weight and FFM were associated with initial decrease in FM and adiposity followed by their increase. Low RQ preceded decrease in adiposity. Conclusion - The marked adiposity depletion, not expected during steady weight gain in the postsurgical period, prompts us to report this finding. The subsequent adiposity catch-up was associated with relatively high REE and RQ, suggesting preferential oxidation of carbohydrates and preservation of lipids for fat storage.