Objective To determine enterobacteria DNA load in venous blood of febrile surgical patients using real-time quantitative polymerase chain reaction (RQ-PCR), to study the correlations between DNA load and vital signs/blood cell count, and to compare the difference between different detection methods in terms of positive rates.Methods A total of 72 blood samples were obtained for bacterial cuhure and RQ-PCR.The correlations of enterobacteria DNA load with body temperature, heart rate, while blood cell count,and percentages of leukocyte and lymphocyte were then analyzed.Results The enterobacteria positive rate determined by RQ-PCR (63.89％) was significantly higher than that by bacterial culture (9.72％) (F =4.383, P =0.036).The DNA load was significantly correlated with both body temperature and heart rate (P =0.006, r =0.323;P =0.000, r =0.411), but not with white blood cell count, percentages of leukocyte and lymphocyte, and age (P=0.438, r=0.093;P=0.825, r=0.027;P=0.451, r=-0.090;P =0.096, r =0.198).Conclusions RQ-PCR can quickly determine the enterobacteria DNA load in peripheral blood with high sensitivity.Routine blood cell count may not accurately reflect the enterobacteria DNA load in blood.Body temperature and heart rate may be influenced by various factors.

Objective To establish a multiplex real-time quantitative polymerase chain reaction (MRQPCR) assay for fast and simultaneous detection of Escherichia coli (E.coli) and Candida albicans (C.albicans) genes in human whole blood,in order to facilitate differentiation of the types of microorganism and evaluation of the severity of bacterial or fungi translocation due to impaired gut barrier,hence providing help to select specific antimicrobial agents.Methods The β-D-galactosidase gene of E.coli and ITS2 gene of C.albicans were selected as the target genes for designing primers and probes.E.coli and C.albicans genomes were extracted with QIAamp(R) DNA Blood Mini Kit,and the 25 μl TaqMan MRQ-PCR amplification reaction system was established.18 simulated human whole blood samples and 10 whole blood samples from febrile surgical patients were detected for E.coli and C.albicans genes using MRQ-PCR.Results The specificity of the primers and probes were excellent.The correlation coefficients of the standard curves of E.coli and C.albicans were 0.994-0.999 and 0.994-0.998,respectively;and the efficiency of amplification were 0.894-1.022 and 0.905-1.028,respectively.In the standard samples,the lowest detection limits of E.coli and C.albicans were 13.9 copies/μl and 0.8 cfu/μl,respectively;the sensitivity was 100％ and 99.69％,the specificity was 100％ and 94.73％,respectively;the average recovery rates were (101.89 ± 5.69)％ and (103.74 ± 4.64)％ respectively;the intra-batch coefficients of variance (CV) in detecting the genes were (13.14 ± 10.27)％ and (19.18 ± 8.54)％,respectively,and the inter-batch CV were (14.35 ± 9.34)％ and (18.31 ± 10.25) ％,respectively.In human whole blood,the lowest detection limits of E.coli and C.albicans were 12 455.2 copies/ml and 800.3 cfu/ml,respectively;the average recovery rates were (111.60 ± 11.06) ％ and (99.96 ± 6.16) ％,respectively;the intra-batch CV in detecting the genes were (11.02 ± 5.65) ％ and (8.14 ± 7.29)％,respectively,and the average inter-batch CV were (12.88 ± 7.59)％ and (18.62 ± 9.14)％.Conclusions MRQ-PCR is a rapid,sensitive,specific,accurate,and reproducible method for simultaneous detection of E.coli and C.albicans genes in human whole blood,with sample-,cost-,and time-saving advantages.It is a promising technique for rapid differentiation between fungi and bacteria,which could help targeted administration and evaluation of antimicrobial agents,and help to assess the consequence of gut barrier damage and the efficacy of treatment.

Objective To establish a rapid real-time quantitative polymerase chain reaction (RQ-PCR)assay in quantifying and detecting Staphylococcus aureus DNA from human venous blood samples,so as to quantificationally evaluate the systemic infection caused or deteriorated by intestinal bacteria translocation.Methods Totally 26 clinical blood samples and 15 simulation blood samples were detected.The primers and TaqMan probe were designed targeting the highly conserved house-keeping femA gene of Staphylococcus aureus,and a 20 μl RQ-PCR amplification reaction system was established.The standard curve was built based on the recombinant plasmid DNA containing the amplicon of the target gene,and genomic DNA was extracted using QIAamp DNA Blood Mini Kit.Results The specificity of primers and probe was excellent,the detecting limit was 100 copies/μl (103 CFU/ml),the sensitivity was 99.7％,and the specificity was 94.6％.The correlation coefficient of the standard curve was between 0.9918 and 0.9997.For samples with different Staphylococcus anreus concentrations,the average accuracy of the RQ-PCR assay was (96.25 ± 2.26) ％ ; the intra-and interassay coefficients of variation were (8.06 ±0.07)％ and (10.01 ±4.40)％,respectively.The average recovery rate of Staphylococcus aureus DNA in blood samples was (111.72 ± 20.72) ％.In clinical blood samples,the positive rate of Staphylococcus aureus DNA was 15.4％ (4/26),while the blood culture of these samples all produced negative result for Staphylococcus aureus.Conclusion RQ-PCR assays is a rapid,sensitive,and specific method with good repetitiveness and can be used in the quantitative detection of Staphylococcus aureus in whole blood samples.

Objective To establish a real-time quantitative PCR(RQ-PCR)assay for fast detection of invasive fungi DNA in human whole blood samples with universal fungi primers and probe.Methods The universal fungi primers and the TaqMan-probe were designed on the basis of the multi-copy 5.8S region of the rDNA of the clinically most common invasive fungi.The invasive fungi genomic DNA were extracted with QIAamp?DNA Blood Mini Kit.A 20 μl RQ-PCR amplification system was established,and the simulated blood samples containing various given load of invasive fungi genome and the 71 whole blood samples of the surgical febrile patients were examined.Results The detection limit is 101 copies/μl amplification mixture,namely 105 copies/ml whole blood.The sensitivity and the specificity were 95.5％ and 97.6％,respectively; and the positive predictive value and negative predictive value were 98.7％ and 92.0％,respectively.The correlation coefficient of standard curve was between 0.9931 and 0.9977.The intra-and the inter-assay average coefficients of variation were(10.4 ±4.0)％ and(27.9 ± 2.0)％,respectively.The average relative recovery rate of fungi genomic DNA in blood samples was(91.0 ±7.6)％,and the average coefficients of variation of the relative recovery rate was(14.9 ±4.0)％.No fungi DNA was detected among the 71 blood samples of the surgical febrile patients.Conclusions The RQ-PCR assay for fast quantitative detection of invasive fungal DNA in human whole blood samples with the universal fungi primers and the TaqMan-probe was of high sensitivity,specificity,accuracy and precision,and is able to discriminate fungi from bacteria.The invasive fungi genome was not detected in this group of surgical patients,which may imply the less possibility of fungi translocation in the surgical febrile patients.

ObjectiveTo evaluate the parenteral nutrition formula's character of all-in-one-mixed preparation and pre-mixed preparation applied for hospitalized patients in China.MethodsWe collected parenteral nutrition (PN) prescriptions in hospitalized patients with various diseases at dispensing centers of 6 hospitals in different areas of China. Statistic analysis was made by nutrients application, total liquid, nonprotein calories, total nitrogen, ratio of non-protein calories and total nitrogen, and ratio of glucose and fat. A comparison was made between all-in-one-mixed and pre-mixed preparation as to all the indices.Results The nutrients supply in all-in-one-mixed PN preparation, as well as in the pre-mixed PN preparations of 3 different energy degrees can meet the basic nutrition requirements of most patients. The total nitrogen supply in all-in-one-mixed PN prescriptions meets nutrition requirements in most patients, but the ratio of non-protein calories and total nitrogen[( 180-250) : 1]and the ratio of glucose and fat (0. 56-1.26)were significantly different from what a doctor would recommend[( 100-150) : 1, 1.0]. While the ratios in the pre-mixed PN preparation on 3 different energy degrees are more consistent with recommeded values,which are at 167 and 0. 8 respectively.ConclusionsAll-in-one-mixed and pre-mixed PN preparation have their own individual advantages and limits, and both of them can meet the requirements of most hospitalized patients. More attention should be paid to the nutrients proportion when doctors prescribe all-in-one-mixed PN preparations.

Objective To establish a real-time quantitative polymerase chain reaction (RQ-PCR) assay for fast detection of Aspergillus fumigatus genome in human whole blood samples and explore its clinical application.Methods The primers and the TaqMan-probe were designed on the basis of the multi-copy ITS1-5. 8S region of the rDNA of Aspergillus fumigatus. The Aspergillus fumigatus genomic DNA were extracted with QIAamp(R) DNA Blood Mini Kit.A 20 μl RQ-PCR amplification system was established, and the simulated blood samples containing various given load of Aspergillus fumigatus genome and the 66 whole blood samples of the surgical febrile patients were examined. Results The detection limit of the RQ-PCR instrument is 10-1 genomes/μl DNA sample,namely 78 CFU/ml whole blood. The specificity and the sensitivity were 94. 25% and 99. 04% respectively; and the positive predictive value and negative predictive value were 97. 63% and 97. 62% respectively. The average relative error of the quantitative results was (3. 67 ±13. 19)%, and the intra- and the inter-assay average coefficients of variation were (12.38 ± 1. 53)% and (16. 27 ±2. 72)% , respectively. The average recovery rate of Aspergillus fumigatus genomic DNA in human whole blood samples was (107. 81 ±25. 92)% , and the average coefficient of variation of the average recovery rate was (26. 24 ± 5.62) % . No Aspergillus fumigatus genomic DNA was detected among the 66 blood samples of the surgical febrile patients. Conclusions The RQ-PCR assay for fast quantitative detection of Aspergillus fumigatus genome in human whole blood samples is of high sensitivity, specificity,accuracy and precision. The Aspergillus fumigatus genome was not detected in this group of surgical febrile patients.

Objective To establish the real-time quantitative PCR (RQ-PCR) assay for detecting Candida albicans (C.albicans) in whole blood and its clinical application in the febrile surgical patients who may develop gut barrier damage and gut microorganism translocation.Methods The NAG1 gene,which is a single copy in C.albicans genome,was selected as the target gene for designing the primers and probe.The plasmid was fabricated and produced as standard samples.C.albicans genomes were extracted with QIAamp(R) DNA Blood Mini Kit,and the total 20 μl TaqMan RQ-PCR amplification reaction system was established.The 74 venous blood samples from the surgical febrile patients were detected for C.albicans load.Results The specificities of the primers and probe were excellent,the correlation coefficients of the standard curves were between 0.9918 and 0.9985,and the efficiency of amplification was 0.88-1.027 for the samples above the lowest detection limit (100 copies/μl examine fluid,or nearly 1.1 × 103 cfu/ml whole blood).The average accuracy of the RQ-PCR equipment was (99.64±2.08) %,the sensitivity was 97.46%,the specificity was 100%,and the average coefficients of variation (CV) of the intra-and inter-assay were (14.76±2.64)% and (17.85±3.53)%,respectively.The average recovery rate of C.albicans DNA in whole blood samples was (88.60±5.73) %,and the average CV of recovery rate was (11.70 ±5.36) %.The number of copies of C.albicans genes per unit blood was not significantly different among the same original blood samples stored separately under-20℃ for 3 or 6 months when compared with its freshly collected blood (P = 0.267).In the 74 whole blood samples obtained from the febrile surgical patients,the positive rate of C.albicans genes was 2.7% and the highest load was 4.42×103 cfu/ml.Conclusions RQ-PCR is a rapid,sensitive,highly specific,and reproducible method in detecting C.albicans NAG1 gene.Clinically it can be used to quantitatively evaluate the numbers of C.albicans in the whole blood.A small percentage of the febrile surgical patients may develop blood infection of C.albicans.

Objective To investigate the use of parenteral nutrition in hospitalized patients and to examine the process of care of patients receiving parenteral nutrition (PN) in hospital, in light of European Nutrition Day study, and to preliminarily explore the possible main risk factors for complications.Methods We recruited 314 adult hospitalized patients (≥18 years) requiring PN on a predetermined day (November 25, 2013) in 6 hospitals in Beijing and investigated their disease status, nutrition risk, the use of PN and venous infusion related complications using the European Nutrition Day survey questionnaire.Results Of the 314 patients, 311 completed the survey.In the 311 patients, the proportion of patients who received PN of all-in-one mixed preparation was 76.2％ (237/311), the proportion of patients receiving multibottle system was 23.8％ (74/311);PN was administered via central vein in 56.8％ (171/301) of the patients and via peripheral veins in 43.2％ (130/301) of the patients.The mean duration of infusion was (12.5 ±5.1) hours.The ratio of glucose to fat was 0.84 and the ratio of non-protein to calories 531.1.The total energy provision was less than the recommended intake.38.6％ (120/311) of the patients reported that PN infusion would affect activity, and 33.4％ (104/311) thought PN affected their sleep, and the incidence of infusion-related pain was 19.9％ (62/311).Multivariate analysis result showed that the most important factors of infusion-related pain were intravenous route of PN (x2 =25.911,P =0.000) and total venous infusion volume (x2 =6.053, P =0.014).Conclusions The total energy provision of PN is generally inadequate in hospitalized patients in Beijing.The key factor for reducing transfusion-related pain and enhancing PN tolerance is to establish appropriate infusion route.

Peripherally inserted central venous catheter (PICC)-related upper extremity venous thrombosis (UEVT) is defined as upper extremity venous thrombosis within the veins where PICCs were placed or adjacent to and may result in pulmonary embolism. Malignancies, previous history of venous thrombosis and malposition are common risk factors for PICC-UEVT. Once patients demonstrate clinical manifestations of phlebitis and thrombosis, such as swelling, pain and tenderness at the PICC site, venous duplex ultrasonography is the first choice for diagnosing PICC-UEVT. According to American College of Chest Physicians guidelines, it's not recommended to remove PICCs upon detection of PICC-UEVT. The first-line treatment is to administer systemic anticoagulants while keeping the catheter in place, unless any contraindications. PICCs could continue to be used during anticoagulation therapy, suppose that catheter tip remains well placed and functions as normal. With early diagnosis and standard anticoagulant treatment, a better clinical outcome could be achieved. Prophylactic anticoagulation is not routinely recommended per guidelines. Recommendation for asymptomatic PICC-related thrombosis is still absent and warrants further prospective studies with large sample size.

Elderly inpatients with coronavirus disease 2019 (COVID-19) are often at nutritional risk and at higher risk of critical disease. The standardized nutrition treatment could effectively improve the nutritional status, quality of life, and clinical outcomes of COVID-19 patients, and is an important component of the comprehensive management of COVID-19. The individualized nutrition diagnosis, treatment and monitoring should be conducted in compliance to standard procedures of medical nutrition therapy, with consideration of the clinical characteristics of elderly COVID-19 inpatients. The Department of Clinical Nutrition at Peking Union Medical College Hospital has integrated the latest clinical nutrition guidelines and clinical practice of nutrition support of COVID-19, with the aim to provide evidence-based, concise and practical recommendations on nutritional management for elderly inpatients with COVID-19. The recommendations here are to inform effective and standardized nutrition support practice.