BCG has been one of the vehicles for multi-recombinant vaccine. However, low transformation efficiency of BCG with plasmid DNA hampered studies involving expression of foreign antigens in BCG. In an effort to determine the optimal conditions, this study was initiated to investigate factors involved in the transformation of BCG with a Mycobacterium-Escherichia coli shuttle vector, pYUB18, by electroporation. Mycobacterium bovis BCG (strain 1173P2) was grown in Middlebrook (M) 7H9 broth containing albumin-dextrose-catalase and 0.05% tween 80, and transformed BCG was grown in M7H10 agar containing kanamycin for counting viable cells. Pretreatment of BCG with 10 mM CaCl2 improved the transformation efficiency, but overnight incubation of BCG with 1% glycine did not. The transformation efficiency in BCG also varied depending on voltage, resistance, and DNA concentration. The maximum transformation efficiency was obtained when the infinity resistance, 12.5 Kv/cm, and 100 ng of DNA were used, and reached 1.4 x 10(5) CFU/microgram of plasmid DNA, which is about 3-100 times greater than those from previous reports. The transformation conditions described in this study, therefore, will give us a better position for employing BCG as a vehicle for developing multi-recombinant vaccines.
Calcium Chloride/pharmacology ; Comparative Study ; DNA/metabolism ; Electrophysiology ; Electroporation* ; Escherichia coli/genetics* ; Genetic Vectors* ; Glycine/pharmacology ; Mycobacterium/genetics* ; Mycobacterium bovis/genetics* ; Osmolar Concentration ; Transformation, Bacterial/physiology* ; Transformation, Bacterial/drug effects

Objective: : Psoas and masseter muscles are known markers of sarcopenia. However, the relative superiority of either muscle as a marker is unclear. Therefore, this study analyzed the two muscles in patients with a prognosis of traumatic brain injury (TBI). Methods: : Patients with TBI visiting a regional trauma center between January 2017 and December 2018 were selected, and their medical records were reviewed. TBI patients with an abbreviated injury score (AIS) of 4 or 5 were selected. Patients with an AIS of 4 or 5 at the chest, abdomen, and extremity were excluded. Patients with a hospital stay of 1 to 2 days were excluded. Both muscle areas were measured based on the initial computed tomography. The psoas muscle index (PMI) and the masseter muscle index (MMI) were calculated by dividing both muscle areas by height in meters squared (cm2/m2). These muscle parameters along with other medical information were used to analyze mortality and the Glasgow outcome scale (GOS). Results: : A total of 179 patients, including 147 males (82.1%), were analyzed statistically. The mean patient age was 58.0 years. The mortality rate was 16.8% (30 patients). The mean GOS score was 3.7. Analysis was performed to identify the parameters associated with mortality, which was a qualitative study outcome. The psoas muscle area (16.9 vs. 14.4 cm2, p=0.028) and PMI (5.9 vs. 5.1 cm2/m2, p=0.004) showed statistical differences between the groups. The PMI was also statistically significant as a risk factor for mortality in logistic regression analysis (p=0.023; odds ratio, 0.715; 95% confidence interval, 0.535–0.954). Quantitative analyses were performed with the GOS scores. Bivariate correlation analysis showed a statistically significant correlation between PMI and GOS scores (correlation coefficient, 0.168; p=0.003). PMI (p=0.004, variation inflation factor 1.001) was significant in multiple regression analysis. The masseter muscle area and MMI did not show significance in the study. Conclusion : Larger PMI was associated with statistically significant improved survival and GOS scores, indicating its performance as a superior prognostic marker. Further analyses involving a larger number of patients, additional parameters, and more precise settings would yield a better understanding of sarcopenia and TBI.