Effects of viral infection and microbial diversity on patients with sepsis:A retrospective study based on metagenomic next-generation sequencing

BACKGROUND: The study aims to investigate the performance of a metagenomic next-generationsequencing (NGS)-based diagnostic technique for the identifi cation of potential bacterial and viral infectionsand eff ects of concomitant viral infection on the survival rate of intensive care unit (ICU) sepsis patients.METHODS: A total of 74 ICU patients with sepsis who were admitted to our institution from February1, 2018 to June 30, 2019 were enrolled. Separate blood samples were collected from patients for bloodcultures and metagenomic NGS when the patients’ body temperature was higher than 38 °C. Patients’demographic data, including gender, age, ICU duration, ICU scores, and laboratory results, were recorded.The correlations between pathogen types and sepsis severity and survival rate were evaluated.RESULTS: NGS produced higher positive results (105 of 118;88.98%) than blood cultures(18 of 118;15.25%) over the whole study period. Concomitant viral infection correlated closelywith sepsis severity and had the negative effect on the survival of patients with sepsis. However,correlation analysis indicated that the bacterial variety did not correlate with the severity of sepsis.CONCLUSIONS: Concurrent viral load correlates closely with the severity of sepsis and thesurvival rate of the ICU sepsis patients. This suggests that prophylactic administration of antiviraldrugs combined with antibiotics may be benefi cial to ICU sepsis patients.

Prognostic value of CD4^+CD25^+ Tregs as a valuable biomarker for patients with sepsis in ICU

BACKGROUND: Sepsis is a common complication ofinfections, burns, traumas, surgeries, poisonings, and post-cardiopulmonary resuscitation. The present study aimed to investigate prognostic value of CD4+CD25+ regulatory T cells(Treg) in peripheral blood of patients with sepsis.METHODS: Periphery blood from 28 patients diagnosed with sepsis was collected on day 1 and 7 after hospitalization in the ICU of Shanghai Changzheng Hospital between December 2013 to April 2014. The blood was used for analyses of Treg ratio using flow cytometry and for analyses of blood routine test, C-reactive protein(CRP), bilirubin, procalcitonin(PCT), and coagulation. APACHE II and sequential organ failure assessment(SOFA) scores were also investigated. The results were compared between two outcome groups of survival or death to evaluate prognostic value for sepsis.RESULTS: The patients had an average age of 60.36±15.03 years, APACHE II score 16.68±7.00, and SOFA score 7.18±3.78. Among the 28 patients, 12 had severe trauma(42.9%), 10 had septic shock(35.7%), and 9(32.2%) died. The median ratio of Tregs was 2.10%(0.80%, 3.10%) in the survival group vs. 1.80%(1.15%, 3.65%) in the death group(Z=–0.148, P=0.883) on day 1; however it was signifi cantly changed to 0.90%(0.30%, 2.80%) vs. 5.70%(2.60%, 8.30%)(Z=–2.905, P=0.004).CONCLUSION: With better prospects for clinical application, dynamic monitoring of Tregs ratio in peripheral blood has potential value in predicting prognosis of sepsis.

Protective effects of Toll-like receptor 4 inhibitor on myocardial injury and lung injury in sepsis rat model

Objective:To study the protective effects of Toll-like receptor 4 (TLR4) inhibitor on myocardial injury and lung injury in sepsis rat model.Methods:A total of 18 adult male SD rats were selected as experimental animals and randomly divided into control group, model group and intervention group, with 6 in each group. The sepsis models were established by cecal ligation and given intragastric administration of TLR4 inhibitor C34. Twenty four hours after model establishment, the levels of myocardial injury molecules and lung injury molecules in serum as well as the expression of inflammatory cytokines, the levels of oxidative stress molecules and the expression of apoptosis molecules in myocardial tissue and lung tissue were determined.Results: KL-6, cTnI and CK-MB levels in serum, NF-kB, TNF-α, IL-1β, Bax, Caspase-3 and Caspase-9 mRNA expression as well as MDA and OH- levels in myocardial tissue and lung tissue of model group were significantly higher than those of control group while SOD levels in myocardial tissue and lung tissue were significantly lower than those of control group;KL-6, cTnI and CK-MB levels in serum, NF-kB, TNF-α, IL-1β, Bax, Caspase-3 and Caspase-9 mRNA expression as well as MDA and OH-levels in myocardial tissue and lung tissue of intervention group were significantly lower than those of model group while SOD levels in myocardial tissue and lung tissue were significantly higher than those of model group.Conclusions: Toll-like receptor 4 inhibitor has protective effect on myocardial injury and lung injury in model rats with sepsis.

Use of Platelet Indices for Determining Illness Severity and Predicting Prognosis in Critically III Patients

Background：Decreased platelet （PLT） count is one of the independent risk factors for mortality in intensive care unit （ICU） patients.This study was to investigate the relationship between PLT indices and illness severity and their performances in predicting hospital mortality.Methods：Adult patients who admitted to ICU of Changzheng Hospital from January 2011 to September 2012 and met inclusion criteria were included in this study.Univariate analysis was used to identify potential independent risk factors for mortality.Multiple logistic regression analysis was used to calculate adjusted odds ratio for mortality in patients with normal or abnormal PLT indices.The relationship between PLT indices and illness severity were assessed by the Acute Physiology and Chronic Health Evaluation Ⅱ （APACHE Ⅱ） scores or sequential organ failure assessment （SOFA） scores in patients with normal and abnormal PLT indices.The performances of PLT indices in predicting mortality were assessed by receiver operating curves and diagnostic parameters.The survival curves between patients with normal and abnormal PLT indices were compared using Kaplan-Meier method.Results：From January 2011 to September 2012,261 of 361 patients （204 survivors and 57 nonsurvivors） met the inclusion criteria.After adjustment for clinical variables,PLT count 〈 100 × 10^12/L （P =0.011）,plateletcrit （PCT） 〈0.108 （P =0.002）,mean platelet volume （MPV） 〉11.3 fL （P =0.023） and platelet distribution width （PDW） percentage 〉17% （P =0.009） were identified as independent risk factors for mortality.The APACHE Ⅱ and SOFA scores were 14.0 （9.0-20.0） and 7.0 （5.0-10.5） in the ＂low PLT＂ tertile,13.0 （8.0-16.0） and 7.0 （4.0-11.0） in the ＂low PCT＂ tertile,14.0 （9.3-19.0） and 7.0 （4.0-9.8） in the ＂high MPV＂ tertile,14.0 （10.5-20.0） and 7.0 （5.0-11.0） in the ＂high PDW＂ tertile,all of which were higher than those in patients with normal indices.Patients with decreased PLT and PCT values （all P 〈 0.001）,or increased MPV and PDW values （P =0.007 and 0.003,respectively） had shortened length of survival than those with normal PLT indices.Conclusions：Patients with abnormally low PLT count,high MPV value,and high PDW value were associated with more severe illness and had higher risk of death as compared to patients with normal PLT indices.

Comparative Study of Plasma Endotoxin with Procalcitonin Levels in Diagnosis of Bacteremia in Intensive Care Unit Patients

Background： Both procalcitonin （PCT） and plasma endotoxin levels cannot be solely used for a definite diagnosis ofbacteremia or sepsis, and there has been few study comparing the values of the two biomarkers for the diagnosis of bacteremia.The aim of this study was to identify bacteria causing bacteremia and evaluate the role of the two biomarkers in the diagnosis ofbacteremia in Intensive Care Unit （ICU）.Methods： The medical records of 420 patients in ICU were retrospectively reviewed.Patients （n =241） who met the inclusion criteria were subjected to blood culture （BC） for the analysis of the endotoxin or PCT levels.The exclusion criteria included the presence of infection with human immunodeficiency virus and/or AIDS, neutropenia without sepsis, pregnancy, treatment with immunosuppressive therapies, or blood diseases such as hematological tumors.Patients＆#39; BC episodes were divided into BC negative, Gram-negative （GN） bacteria, Gram-positive bacteria, and fungi groups.The PCT and plasma endotoxin levels were compared in the different groups.Results： A total of 241 patients with 505 episodes of BC were analyzed.The GN bacteria group showed higher levels of PCT and endotoxin than the BC negative, Gram-positive bacteria, and fungi groups.GN bacteremia was more prevalent than Gram-positive bacteremia.The GN bacteremia caused by non-Enterobacteriaceae infection presented higher endotoxin level than that by Enterobacteriaceae, but no significant difference in PCT levels was observed between the two groups.The plasma endotoxin significantly differed among different groups and was bacterial species dependent.Conclusions： Plasma endotoxin was more related to GN than to Gram-positive bacteremia, and that endotoxin level was species dependent, but PCT level remained relatively more stable within the GN bacteria caused bacteremia.Both GN and positive bacteria caused bacteremia in the ICU patients in different regions of China.And PCT is a more valuable biomarker than endotoxin in the diagnosis of bacteremia.

Rhubarb Antagonizes Matrix Metalloproteinase-9-induced Vascular Endothelial Permeability

Background： Intact endothelial structure and function are critical for maintaining microcirculatory homeostasis. Dysfunction of the latter is an underlying cause of various organ pathologies. In a previous study, we showed that rhubarb, a traditional Chinese medicine, protected intestinal mucosal microvascular endothelial cells in rats with metastasizing septicemia. In this study, we investigated the effects and mechanisms of rhubarb on matrix metalloproteinase-9 （MMP9）-induced vascular endothelial （VE） permeability. Methods： Rhubarb monomers were extracted and purified by a series of chromatography approaches. The identity of these monomers was analyzed by hydrogen-1 nuclear magnetic resonance （NMR）, carbon-13 NMR, and distortionless enhancement by polarization transfer magnetic resonance spectroscopy. We established a human umbilical vein endothelial cell （HUVEC） monolayer on a Transwell insert. We measured the HUVEC permeability, proliferation, and the secretion of VE-cadherin into culture medium using fluorescein isothiocyanate-dextran assay, 3-（4,5-dimethyl-2-thiazolyl）-2,5-diphenyl-2-H-tetrazolium bromide assay, and enzyme-linked immunosorbent assay, respectively, in response to treatment with MMP9 and/or rhubarb monomers. Results： A total of 21 rhubarb monomers were extracted and identifed. MMP9 significantly increased the permeability of the HUVEC monolayer, which was significantly reduced by five individual rhubarb monomer （emodin, 3,8-dihydroxy- 1-methyl-anthraquinone-2-carboxylic acid, 1-O-caffeoyl-2-（4-hydroxyl-O-cinnamoyl）-β-D-glucose, daucosterol linoleate, and rhein） or a combination of all five monomers （1μmol/L for each monomer）. Mechanistically, the five-monomer mixture at 1 μmol/L promoted HUVEC proliferation. In addition, MMP9 stimulated the secretion of VE-cadherin into the culture medium, which was significantly inhibited by the five-monomer mixture. Conclusions： The rhubarb mixture ofemodin, 3,8-dihydroxy-l-methyl-anthraquinone-2-carboxylic acid, 1-O-caffeoyl-2-（4-hydroxyl-O -cinnamoyl）-μ-D-glucose, daucosterol linoleate, and rheim at a low concentration, antagonized the MMP9-induced HUVEC monolayer permeability by promoting HUVEC proliferation and reducing extracellular VE-cadherin concentrations.

Rhubarb Monomers Protect Intestinal Mucosal Barrier in Sepsis via Junction Proteins

Background： Leakage of the intestinal mucosal barrier may cause translocation of bacteria, then leading to multiorgan lhilure. This study hypothesized that rhubarb monomers might protect the gut mucosal barrier in sepsis through junction proteins. Methods： Healthy male Sprague-Dawley rats （weighing 230-250 g） tinder anesthesia and sedation were subjected to cecal ligation and perforation （CLP）. After surgical preparation, rats were randornly assigned to eight groups （n = 6 or 8 each group）： sham group （Group A： normal saline gavage）; sepsis group （Group B： nomlal saline gavage）; Group C （intraperitoneally, dexamethasone 0.5 mg/kg） immediately after CLP surgery; and rhubarb monomer （ 100 mg/kg in normal saline）-treated groups （Group D： rhein： Group E： emodin; Group F： 3,8-dihydroxy- l-methyl-anthraquinone-2-carboxylic acid; Group G： 1-O-caffeoyl-2-（4-hydroxy-O-cinnamoyl）-D-glucose; and Group H： daucosterol linoleate）. Animals were sacrificed after 24 h. Intestinal histology, lactulose, mannito[ concentrations were measured, and zonula occludens （ZO）-I, occludin and claudin-5 transcription （polymerase chain reaction）, translation （by Western blot analysis）, and expression （by immunohistochemistry） were also measured. Results： Intestinal histology revealed injury to intestinal mucosal villi induced by sepsis in Group B, compared with Group A. Compared with Group A （0.17 ± 0.41 ）, the pathological scores in Groups B （2.83 ± 0.41, P 〈 0.001）, C （ 1.83 ± 0.41, P 〈 0.001 ）, D （2.00 ± 0.63, P 〈 0.001）, E （ 1.83 ± 0.41, P 〈 0.001 ）, F （ 1.83 ± 0.75, P 〈 0.001 ）, G （2.17 ± 0.41, P 〈 0.001 ）,and H （ 1.83 ± 0.41, P 〈 0.001 ） were significantly increased. Lactulose/mannitol （L/M） ratio in Group B （0.046 ± 0.003） was significantly higher than in Group A （0.013 ± 0.001, P 〈 0.001） while L/M ratios in Groups C （0.028 ± 0.002, P 〈 0.001 ）, D （0.029 ± 0.003, P 〈 0.001 ）, E （0.026 ± 0.003, P 〈 0.001 ）, F （0.027 ± 0.003, P 〈 0.001 ）, G （0.030 ± 0.005, P 〈 0.001 ）, and H （0.026 ± 0.002, P 〈 0.001 ） were significantly lower than that in Group B. ZO- 1, occludin and claudin-5 transcription, translation, and expression in Group B were significantly lower than that in Group A （P 〈 0.001 ）, but they were significantly higher in Groups C, D, E, F, G, and H than those in Group B （P 〈 0.05）. Conclusion： Rhubarb monomer treatment ameliorated rnucosal damage in sepsis via enhanced transcription, translation, and expression of junction proteins.