Bioconversion of Phytosterols to 9-Hydroxy-3-Oxo-4,17-Pregadiene-20-Carboxylic Acid Methyl Ester by Enoyl-CoA Deficiency and Modifying Multiple Genes in Mycolicibacterium neoaurum.

Steroid drug precursors, including C19 and C22 steroids, are crucial to steroid drug synthesis and development. However, C22 steroids are less developed due to the intricacy of the steroid metabolic pathway. In this study, a C22 steroid drug precursor, 9-hydroxy-3-oxo-4,17-pregadiene-20-carboxylic acid methyl ester (9-OH-PDCE), was successfully obtained from Mycolicibacterium neoaurum by 3-ketosteroid-Δ1-dehydrogenase and enoyl-CoA hydratase ChsH deficiency. The production of 9-OH-PDCE was improved by the overexpression of 17ß-hydroxysteroid dehydrogenase Hsd4A and acyl-CoA dehydrogenase ChsE1-ChsE2 to reduce the accumulation of by-products. The purity of 9-OH-PDCE in fermentation broth was improved from 71.7% to 89.7%. Hence, the molar yield of 9-OH-PDCE was improved from 66.7% to 86.7%, with a yield of 0.78 g/L. Furthermore, enoyl-CoA hydratase ChsH1-ChsH2 was identified to form an indispensable complex in Mycolicibacterium neoaurum DSM 44704. IMPORTANCE C22 steroids are valuable precursors for steroid drug synthesis, but the development of C22 steroids remains unsatisfactory. This study presented a strategy for the one-step bioconversion of phytosterols to a C22 steroid drug precursor, 9-hydroxy-3-oxo-4,17-pregadiene-20-carboxylic acid methyl ester (9-OH-PDCE), by 3-ketosteroid-Δ1-dehydrogenase and enoyl-CoA hydratase deficiency with overexpression of 17ß-hydroxysteroid dehydrogenase acyl-CoA dehydrogenase in Mycolicibacterium. The function of the enoyl-CoA hydratase ChsH in vivo was revealed. Construction of the novel C22 steroid drug precursor producer provided more potential for steroid drug synthesis, and the characterization of the function of ChsH and the transformation of steroids further revealed the steroid metabolic pathway.

Routine Use of Biomarkers to Rationalize Antibiotic Use During Febrile Episodes in Pediatric Bone Marrow Transplantation Units.

BACKGROUND: Children frequently develop fever after hematopoietic stem cell transplant (HSCT). Although the etiology of many febrile episodes (FEs) is not an infection, patients often receive broad-spectrum antibiotics in response. METHODS: To improve the judicious use of antibiotics in pediatric HSCT patients, we performed a prospective cohort study of children receiving an HSCT in Clínica Imbanaco (Cali, Colombia) between September 2016 and December 2019. We assessed all FEs occurring during 3 periods (infusion, neutropenic and engraftment). We measured procalcitonin and C-reactive protein (CRP) sequentially during each FE and compared levels among patients with fever due to significant infection (FSI) versus fever not attributable to infection (FNI) in each transplant period. RESULTS: There were 166 FEs in 95 patients. FSI accounted for 12%, 42% and 42% of FE during infusion, neutropenic and engraftment periods, respectively. CRP had better discriminatory capacity for FSI versus FNI in the infusion period [area under the curve (AUC) 0.80 (95% confidence interval [CI], 0.62-0.96) for a CRP level of 50 mg/L]. Neither biomarker performed well in the neutropenic period. During the engraftment period, a CRP of 65 mg/L had an AUC of 0.81 (95% CI, 0.65-0.96), while a procalcitonin level of 0.25 ng/mL had an AUC of 0.83 (95% CI, 0.63-1.0). In contrast to procalcitonin, the CRP's pattern of change throughout the first 3 days of fever in each transplant period was different in FSI compared with FNI. CONCLUSION: Sequential measurement of biomarkers, especially CRP, may allow clinicians to more appropriately manage antibiotic use in pediatric HSCT units.