Objective:To investigate clinicians' compliance with the 2018 Surviving Sepsis Campaign (SSC) update "1-hour sepsis Bundle therapy" (1-hour Bundle) when treating patients with Sepsis 3 in the intensive care unit (ICU), and to analyze its impact on patient outcomes.Methods:A multicenter, prospective observational cohort study was conducted. A total of 153 ICU patients in Ziyang First People's Hospital, Ziyang People's Hospital and Yanjiang District People's Hospital who were diagnosed of sepsis by the definition and diagnostic criteria of Sepsis 3 from January 2019 to December 2020 were selected. Among them, 95 patients who had completed 1-hour Bundle were divided into the Bundle compliance group. 58 patients who did not complete the Bundle within 1 hours were classified as the Bundle non-compliance group. The distribution of pathogenic bacteria and infected sites, 1-hour Bundle compliance and 28-day survival in the 3 hospitals were analyzed. Univariate analysis was used to analyze the risk factors affecting the prognostic between the two groups of sepsis patients. Cox regression model was used to draw a 28-day survival curve to evaluate the survival of the patients in the two groups.Results:Among 153 sepsis patients in 3 hospitals, the detection rate of pathogenic bacteria was 61.44% (94/153), and Gram-negative bacteria accounted for 79.79% (75/94). The top 3 infection sites were respiratory system, gastrointestinal tract and urinary system, accounted for 32.0%, 28.1% and 18.3%, respectively. In the 3 hospitals, 62.09% (95/153) of patients fully implemented the 1-hour Bundle. The poorly implemented indicators in the 1-hour Bundle were 1-hour blood microbial culture [77.78% (119/153)] and 1-hour antimicrobial application [79.74% (122/153)]. There was no significant difference in the baseline indicators between Bundle compliance and non-compliance groups. Univariate analysis showed that the main prognostic indicators: 28-day survival rate in the Bundle compliance group was significantly higher than that in the Bundle non-compliance group [80.00% (76/95) vs. 62.06% (36/58), χ2= 6.447, P = 0.014]. Secondary evaluation indicators: mean arterial pressure (MAP) at 6 hours and 24 hours in the Bundle compliance group were significantly higher than those in the Bundle non-compliance group [mmHg (1 mmHg = 0.133 kPa): 78.22±11.25 vs. 69.86±14.04, 79.78±11.45 vs. 75.35±12.90]. However, the median length of in hospital stay in the Bundle compliance group was significantly longer than that in the Bundle non-compliance group [days: 13 (17) vs. 6 (11)], with statistically significant differences (all P < 0.05). Bivariate Logistic regression analysis showed that 6 hours and 24 hours MAP were risk factors affecting the prognosis of patients with sepsis [odds ratio ( OR), 95% confidence interval (95% CI): 1.064 (0.994-1.102), 1.032 (1.003-1.063), both P < 0.05]. Conclusions:The 1-hour Bundle compliance rate of ICU patients with sepsis in 3 hospitals of Ziyang City was 62.09%, and the compliance is still to be improved, especially for the 2 aspects of empirical antimicrobial use and microbial culture retention before antimicrobial use. The 28-day survival rate in the Bundle compliance group was significantly higher than that in the Bundle non-compliance group, suggesting that the 1-hour Bundle regimen can improve the prognosis of patients with sepsis.

BACKGROUND: Malignant plural effusion (MPE) is one of the most common specimen for liquid biopsy gene detection. This study aims to explore a method for isolating tumor cells from large volume of MPE and evaluate its efficacy and application prospect in gene detection. METHODS: Pleural effusions (>500 mL) from 20 advanced lung cancer patients were obtained by effusion drainage and used to isolate tumor cells with cell separation media Percoll and Ficoll. Cell number and purity were calculated. DNA was extracted from the supernatant (etDNA), total cells and isolated tumor cells of pleural effusion (ETC-DNA) to detect the mutation of tumor-related genes by next-generation sequencing. RESULTS: The median number of cells isolated from malignant pleural effusion was 8.50×10⁴ (interquel range: 9.25×10³-3.75×10⁵), 85.50%±5.80% of the cells were identified as tumor cells. The detection rates of epidermal growth factor receptor (EGFR) gene mutation of etDNA, total cell DNA and ETC-DNA were 70.00%, 50.00% and 70.00%, reseparately, while the median EGFR mutation abundance in 3 components was 16.05% (4.78%-43.06%), 1.09% (0.00%-2.39%), and 33.02% (18.50%-76.70%), respectively. ETC-DNA had good consistency with tissue DNA (P>0.999, kappa=1.000) and etDNA (P>0.999, kappa=1.000). ETC-DNA inclined to have higher EGFR mutation than etDNA, but the result was not statistically significant. CONCLUSIONS Our method can isolate large amount of tumor cells from a large volume of malignant pleural effusion with high purity. Using ETC-DNA as specimen improves the efficacy of gene detection, thus is worth further study.

Objective: To evaluate the external quality assessment (EQA) program for genotyping results of tacrolimus metabolism-related cytochrome P450 family 3 subfamily A member 5 ( CYP3A5 )using plasmid DNA constructed in vitro as quality control samples, discuss the problems in clinical laboratories enrolled in the program and improve the detection quality of CYP3A5 gene. Methods: Recombinant plasmid carrying CYP3A5 *3 (rs776746) AA locus sequence was constructed as wild type sample and plasmid with CYP3A5 *3 GG mutation as mutant type sample. Heterozygous mutant samples were obtained by mixing the two plasmids with equal proportion. Recombinant plasmids DNA were used as the sample panel for EQA scheme. Participating laboratories were asked to test the samples using their routine methods and report the results before deadlines. The scores of each laboratory were calculated based on their results and the overall coincidence of different samples as well as the sensitivity and specificity of different methods. Results: CYP3A5 *3 locus genotypes of the constructed plasmid were verified by Sanger sequencing. The results of 15 and 17 valid laboratories were received respectively in the two EQA programs. The total percentage of 93.33% (14/15) and 100% (17/17) of the laboratories submitted correct results for all the samples. The overall coincidence rates were 96% (72/75) and 100% (85/85) respectively. All the laboratories using digital FISH got full marks in two EQA schemes, while the coincidence rates were 90% (27/30) and 100% (40/40) for Sanger sequencing. Conclusion The recombinant plasmid DNA constructed in this study could effectively detect the performance of reagents with good clinical applicability. The results of EQA programs suggested that the overall accuracy rate of enrolled laboratories was high enough, while the performances in some laboratories still need to be improved. Quality controls in clinical laboratories were essential to assure the accuracy of results.

Objective To evaluate the performance of MTHFR 677 genotyping external quality assessment ( EQA) program using plasmid DNA constructed in vitro as quality control samples and discuss the problems in clinical laboratories enrolled in the program .Methods Recombinant plasmid carrying MTHFR 677C locus sequence was constructed as wild type sample and plasmid with MTHFR 677T mutation was generated with site-directed mutagenesis as mutant type sample .Heterozygous mutant samples were obtained after equal proportion of the two plasmids .EQA scheme were held twice a year in 2016 and 2017, and sample panels contained 5 different samples using recombinant plasmid DNA containing all types of MTHFR 677 locus genotypes.Participating laboratories were asked to test samples using their routine methods and report the results before deadlines .26, 28, 52 and 56 effective reports were received respectively in the four EQA schemes .The scores of each lab were calculated based on their results and the overall compliance of different samples as well as the sensitivity and specificity of different methods were calculated using Microsoft Excel .Results MTHFR 677 locus genotypes of the constructed plasmid were verified by Sanger sequencing and there was no failure of sample detection in the four EQA schemes , which suggest that the plasmid has good clinical applicability .About 96.15％( 25/26 ) ,100％( 28/28 ) ,96.15％ (50/52)and 98.21％(55/56) of the laboratories submitted correct results for all samples in the four EQA schemes.The overall compliance rate were 99.23％ ( 129/130 ) , 100％( 140/140 ) ,96.92％ ( 252/260 ) and 98.93％(277/280) respectively.All laboratories using digital FISH and microarrays got full marks in four EQA schemes.The compliance rates for fluorescent PCR were 97.5％ ( 39/40 ) , 100％ ( 45/45 ) , 94.29％(66/70) and 100％ (95/95) respectively, while the rates were 100％ (20/20), 100％ (15/15), 90％(36/40) and 92.5％(37/40) for Sanger sequencing.Conclusions The recombinant plasmid DNA constructed in this study can effectively detect the performance of reagents with good clinical applicability.The results of EQA programs suggested that the overall accuracy rate of laboratories enrolled was high enough , while some laboratories′performance still needs to be improved .Quality controls in clinical laboratories were essential to assure the accuracy of results .

The current treatments of metastatic malignant melanoma include chemotherapy,targeted therapy,immune therapy and radiation therapy,but the treatment outcome is far from optimism.In order to im-prove the treatment efficiency,it is urgent to improve early diagnosis,and develop more effective treatment drugs and delivery systems.The application of nanotechnology in the diagnosis and therapy of melanoma can re-duce the resistance to the drugs,increase efficacy and reduce side effects.

Objective To investigate the relationship between anti-HCV antibody level and hepatitis C virus genotype in the patients.Methods Total of 603 anti-HCV positive serum samples were collected during 2013 to 2014 by retrospective research method.HCV RNA were detected in anti-HCV positive samples by repeat test and the genotype were detected in HCV RNA positive samples.The distribution of anti-HCV level in different hepatitis C genotype patients was analyzed and the body's response to viral antibodies and viral genotype correlation with anti-HCV concentration interquartile range was explored.Rates among genotype groups were compared using chi-square test.Results Totally 412 of 603 (68.33％) samples were anti-HCV positive by double reagent screening.174(42.3％) samples were detected as HCV RNA positive.The distributions of different anti-HCV level in different genotype patients were 1a(n =8) 1/8,1/8,4/8,2/8;1b(n =112)25.9％ (29/112),17.0％ (19/112),25.9％ (29/112),31.3％ (35/112);2a(n =14)3/14,4/14,5/14,2/14;3a(n =11)3/11,6/11,2/11,0/11;3b(n =16)4/16,11/16,1/16,0/16;6a(n =8)2/8,2/8,1/8,3/8 with anti-HCV concentration interquartile range respectively.The anti-HCV concentration distribution was different in patients with different HCV genotypes.The anti-HCV concentration distribution in patients of 1 b,2a and 6a genotypes were evently,while anti-HCV level was relatively high in 1a (13.65) and relatively low in 3b (8.77).There were differences in different genotypes of antibody concentrations (x2 =35.2,P ＜ 0.05).Conclusions There was correlation between anti-HCV level and HCV genotype.Because there were fewer cases in some genotypes,it was necessary to investigate more samples to corfirm the above results.

Objective To investigate the correlation between fatigue and coping strategies among peritoneal dialysis patients,to improve their health consciousness and reduce the fatigue.Methods A total 107 patients were surveyed with general data questionnaire,Multidimensional Fatigue Symptom Inventory-Short Form( MFSI-SF) ,and Medical Coping Modes Questionnaire,MCMQ.Results The score of fatigue was (83.32 ±1.54) points,which indicated the fatigue remained in high level.Scores from high to low were dialysis≥5years,dialysis<1year and 1year≤dialysis<5years.Patients mainly took the coping style with positive face.Pearson correlation analysis showed that fatigue was negatively correlated withface(r=-0.300,P<0.05),while it was positively correlated with withdrawand yield(r=0.227,0.300,all P<0.05).Conclusion Patiens should be encouraged to actively face the disease,pay more attention to their own state of dialysis and improve the effect of drain,reduce fatigue,then to reply for both mind and body health.

Objective To investigate the effect and mechanism of valproic acid in preventing astrocyte proliferation around the central canal of rats following spinal cord injury.Methods Forty-five Wister rats were divided into normal control group (n =5),injury group (n =20) and treatment group (n =20) according to random number table.Animal models of acute spinal cord injury were produced at T10 using Allen' s method by dropping a 10 g weight from a 15 mm height.Rats in treatment group received intraperitoneal injection of valproic acid (300 mg · kg-1 · d-1 in two divided doses) at 30 minutes postinjury.Instead,rats in injury group were injected with an equal volume of saline in the same way.Hindlimb function was evaluated using BBB scoring system at 1,3,7,and 14 days postinjury.Astrocytes proliferation around central canal and expression of glial fibrous acid protein (GFAP) were examined.Results In normal control group,few astrocytes around spinal central canal and a low expression of GFAP were detected.In injury group,astrocytes began to increase at 24 hours postinjury; fluorescence intensity for GFAP was 24.6 ± 3.6 at 24 hours,reached a peak of 69.2 ± 6.4 at 3 days,maintained a high level of 56.7 ± 5.6 at 7 days,and reduced to 35.4 ± 4.3 at 14 days,a level that remained higher than that in normal control group (11.2 ± 1.6).Whereas in treatment group at 3 and 7 days,astrocyte proliferation around spinal central canal was lower than that in injury group; GFAP expressions (47.8 ± 5.3 and 42.2 ± 6.7) were lower than those in injury group (F =177.6,P ＜ 0.05).At 3,7,and 14 days,BBB scores in treatment group (7.80 ± 0.83,12.00 ± 1.58,and 16.60 ± 1.12 respectively) were significantly higher than those in injury group (4.60 ± 0.54,6.65 ± 0.67,and 9.40 ± 1.14 respectively) (F =1 113.6,P ＜ 0.05).Conclusion After spinal cord injury,valproic acid reduces astrocyte proliferation around central canal via inhibiting GFAP expression to promote functional recovery.

ObjectiveTo investigate the inhibitory effect of dacarbazine and an oncolytic adenovirus carrying interleukin-24 (IL-24) on transplanted melanoma in nude mice.MethodsNude mice were inoculated with human A375 melanoma cells to establish a model of malignant melanoma.Then,the mice were divided into 4 groups to be treated with an oncolytic adenovirus carrying interleukin-24 (ZD55-IL-24),dacarbazine,the combination of ZD55-IL-24 and dacarbazine,and phosphate buffer(PBS),respectively,for 3 days.Seven days after the end of the treatment,some mice were sacrificed followed by the determination of IL-24 and E1A protein levels in tumor tissue by Western blot.The tumor volume was measured on a daily basis for 30 days.ResultsIL-24 and E1A were highly expressed in melanoma cell-bearing nude mice treated with ZD55-IL-24 and dacarbazine.At 30 days after the inoculation,the average volume of transplanted melanoma was (2346.5 ± 576.0) mm3 in the combination group,significantly different from that in the ZD55-IL-24 group((4141.6 ± 1348.2) mm3,P ＜ 0.05),dacarbazine group((5230.1 ± 922.8) mm3,P ＜ 0.05),and the control group ((7135.1 ± 1002.3) mm3,P ＜ 0.05).ConclusionThe ZD55-IL-24 in combination with dacarbazine exhibits a remarkably inhibitory effect on the proliferation of melanoma transplanted into nude mice.

Objective To investigate the production of carbapenemase and 16S rRNA methylase in five isolates of pan-drugs resistant E.cloacae recovered in Ruijin hospital.Methods MICs of the five isolates to 10 antibiotics were determined by E test.Six kinds of 16S rRNA methylase genes and a series of β- lactamase genes were amplified by PCR.Shotgun cloning was performed to detect carbapenem resistance determinant.The conjugal transfer of carbapenemase gene and 16S rRNA methylase gene was performed in broth culture with E.coli J53 as the recipient.Pulsed-field gel electrophoresis (PFGE) was carried out to analyse the genotyping.IEF was performed to detect β-1actamases.Southern blot was performed to determine the location of carbapenem resistance determinant.Results The MICs of 10 antibiotics were ＞32 mg/L.Four β-1actamases with pIs of 5.4 ( TEM-1 ),6.7 ( KPC-2 ),8.2 ( SHV-12 ),8.4 (CTX-M-14) were determined.The insertion sequence in the recombinant plasmid was blaKPC-2 flanked by a transposon.blaKPC-2 was located on a large non-conjugative plasmid whereas armA was located on an other conjugative plasmid.PFGE patterns of 5 isolates were identical.Conclusion KPC-2 was responsible for carbapenem resistance in pandrugs resistant Enterobacter cloacae.There was no relationship between blaKPC-2 and armA.Although pandrug resistant Enterobacteriaceae remain rare,the emergence of this group of organism merits monitoring.