A Nomogram Model for Individualized Prediction of the Risk of Respiratory Tract Infection within Six Months after Diagnosis in Patients with Primary Immune Thrombocytopenia.

The risk factors of upper respiratory tract infection (URI) within 6 months after diagnosis in patients with idiopathic thrombocytopenic purpura (ITP) were analyzed, and the nomogram model was established and verified, with 242 and 50 ITP patients as the training and validation set, respectively. The patients were followed up for six months after the diagnosis of ITP. The clinical data of patients were collected, and the risk factors of URI in ITP patients within six months after diagnosis were analyzed using univariable, followed by multivariable logistic regression. Among the 242 ITP patients in the training set, 52 cases (21.49%) had URI, including 24 cases of viral infection, 11 cases of Mycoplasma pneumoniae infection, and 17 cases of bacterial infection. Logistic regression analysis showed that advanced age, use of glucocorticoid, smoking history, platelet count, serum CRP level, and lymphocyte subsets CD4 + and CD8 + were all risk factors for ITP patients to develop symptoms within six months after diagnosis (P < 0.05). Using the above five indicators, a nomogram prediction model was built for URI occurrence in patients with ITP within half a year after diagnosis, and the results showed an AUC, a sensitivity, and a specificity of 0.936 (95% CI: 0.878-0.983), 0.942, and 0.865, respectively. The nomogram model was internally verified by the bootstrap method for 500 self-sampling times, and the prediction of the calibration curve was in high consistency with the real results. External validation of the nomogram model resulted in an AUC, a sensitivity, and a specificity of 0.890 (95% CI: 0.757-0.975), 0.949, and 0.727, respectively. The nomogram model of URI in ITP patients within half a year after diagnosis based on logistic regression analysis has good discrimination and prediction accuracy. This provides important guidance value for individualized prediction of URI in ITP patients.

Branched-chain 2-keto acid decarboxylases derived from Psychrobacter.

The conversion of branched-chain amino acids to branched-chain acids or alcohols is an important aspect of flavor in the food industry and is dependent on the Ehrlich pathway found in certain lactic acid bacteria. A key enzyme in the pathway, the 2-keto acid decarboxylase (KDC), is also of interest in biotechnology applications to produce small branched-chain alcohols that might serve as improved biofuels or other commodity feedstocks. This enzyme has been extensively studied in the model bacterium Lactococcus lactis, but is also found in other bacteria and higher organisms. In this report, distinct homologs of the L. lactis KDC originally annotated as pyruvate decarboxylases from Psychrobacter cryohalolentis K5 and P. arcticus 273-4 were cloned and characterized, confirming a related activity toward specific branched-chain 2-keto acids derived from branched-chain amino acids. Further, KDC activity was confirmed in intact cells and cell-free extracts of P. cryohalolentis K5 grown on both rich and defined media, indicating that the Ehrlich pathway may also be utilized in some psychrotrophs and psychrophiles. A comparison of the similarities and differences in the P. cryohalolentis K5 and P. arcticus 273-4 KDC activities to other bacterial KDCs is presented.

Construction of a genetically engineered microorganism for phenanthrene biodegradation.

The bacterium Pseudomonas sp. CGMCC2953, isolated from oil-polluted soil, was used as a recipient for a biodegradative gene encoding catechol 2,3-dioxygenase (C23O), which was successfully cloned into the plasmid pK4 derived from pRK415 with a broad host range. The apparent phenanthrene biodegradation parameters of the recombinant microorganism (Pseudomonas sp. CGMCC2953-pK) were determined and compared with those of the wild type. As the key enzyme of phenanthrene degradation, C23O, could be expressed constitutively in the recombinant strain, Pseudomonas sp. CGMCC2953-pK showed an increased ability to degrade phenanthrene. The excessive production of C23O in Pseudomonas sp. CGMCC2953-pK could serve as an effective approach to construct genetically engineered microorganisms for the bioremediation of environmental contaminations.

Differences in substrate specificities of five bacterial wax ester synthases.

Wax esters are produced in certain bacteria as a potential carbon and energy storage compound. The final enzyme in the biosynthetic pathway responsible for wax ester production is the bifunctional wax ester synthase/acyl-coenzyme A (acyl-CoA):diacylglycerol acyltransferase (WS/DGAT), which utilizes a range of fatty alcohols and fatty acyl-CoAs to synthesize the corresponding wax ester. We report here the isolation and substrate range characterization for five WS/DGAT enzymes from four different bacteria: Marinobacter aquaeolei VT8, Acinetobacter baylyi, Rhodococcus jostii RHA1, and Psychrobacter cryohalolentis K5. The results from kinetic studies of isolated enzymes reveal a differential activity based on the order of substrate addition and reveal subtle differences between the substrate selectivity of the different enzymes. These in vitro results are compared to the wax ester and triacylglyceride product profiles obtained from each organism grown under neutral lipid accumulating conditions, providing potential insights into the role that the WS/DGAT enzyme plays in determining the final wax ester products that are produced under conditions of nutrient stress in each of these bacteria. Further, the analysis revealed that one enzyme in particular from M. aquaeolei VT8 showed the greatest potential for future study based on rapid purification and significantly higher activity than was found for the other isolated WS/DGAT enzymes. The results provide a framework to test prospective differences between these enzymes for potential biotechnological applications such as high-value petrochemicals and biofuel production.

Rational design of catechol-2, 3-dioxygenase for improving the enzyme characteristics.

Catechol-2, 3-dioxygenase (C23O) from Pseudomonas sp. CGMCC2953 identified in our laboratory, which is one of the key enzymes responsible for phenanthrene biodegradation, was expected to get better characteristics tolerant to environment for its further application. With the aim of improving the enzyme properties by introducing intermolecular disulfide bonds, X-ray structure of a C23O from Pseudomonas putida MT-2, a highly conserved homologous with the C23O from Pseudomonas sp. CGMCC2953, was directly used to find the potential sites for forming disulfide bonds between two monomers of the target C23O. Two sites, Ala229 and His294, were identified and mutated to cysteine, respectively, by using site mutagenesis. The expected disulfide bond between these two CYS residues was confirmed with both molecular modeling and experimental results. The optimum temperature of the mutated enzyme was widened from 40 to 40 approximately 50 degrees C. The mutated C23O became more alkalescency stable compared with the wild-type enzyme, e.g., 75% of the maximal enzyme activity retained even under pH 9.5 while 50% residue for the wild-type one. Improvement of thermostability of the mutated C230 with the redesigned disulfide was also confirmed.