Rapid diagnosis of sepsis with TaqMan-Based multiplex real-time PCR.

BACKGROUND: The survival rate of septic patients mainly depends on a rapid and reliable diagnosis. A rapid, broad range, specific and sensitive quantitative diagnostic test is the urgent need. Thus, we developed a TaqMan-Based Multiplex real-time PCR assays to identify bloodstream pathogens within a few hours. METHODS: Primers and TaqMan probes were designed to be complementary to conserved regions in the 16S rDNA gene of different kinds of bacteria. To evaluate accurately, sensitively, and specifically, the known bacteria samples (Standard strains, whole blood samples) are determined by TaqMan-Based Multiplex real-time PCR. In addition, 30 blood samples taken from patients with clinical symptoms of sepsis were tested by TaqMan-Based Multiplex real-time PCR and blood culture. RESULTS: The mean frequency of positive for Multiplex real-time PCR was 96% at a concentration of 100 CFU/mL, and it was 100% at a concentration greater than 1000 CFU/mL. All the known blood samples and Standard strains were detected positively by TaqMan-Based Multiplex PCR, no PCR products were detected when DNAs from other bacterium were used in the multiplex assay. Among the 30 patients with clinical symptoms of sepsis, 18 patients were confirmed positive by Multiplex real-time PCR and seven patients were confirmed positive by blood culture. CONCLUSION: TaqMan-Based Multiplex real-time PCR assay with highly sensitivity, specificity and broad detection range, is a rapid and accurate method in the detection of bacterial pathogens of sepsis and should have a promising usage in the diagnosis of sepsis.

Therapeutic Effects and Mechanism of Chinese Medicine Skin Dialysis for Chronic Renal Failure / 广州中医药大学学报

Objective To observe the influence of Chinese medicine skin dialysis on traditional Chinese medicine (TCM) syndrome scores and microinflammation in patients with chronic renal failure(CRF). Methods A total of 180 patients with CRF were divided into groups A, B and C, 60 cases in each group. All of the 3 groups were given conventional treatment, and additionally, group A was given Chinese medicine skin dialysis, and group B was given pure water dialysis. Meanwhile, 60 healthy volunteers served as normal control group (group D), and had no any medication. The course of treatment lasted for one month. Before and after treatment, we examined the scores of TCM syndromes and the levels of 24-hour urinary protein, serum creatinine (SCr), blood urea nitrogen (BUN), estimated glomerular filtration rate (eGFR), albumin (ALB), immunoglobulin A(IgA), immunoglobulin G (IgG) , immunoglobulin M(IgM), interleukin 6 (IL-6) , interleukin 1(IL-1), hyper-sensitive C reactive protein (hs-CRP), and tumor necrosis factor alpha (TNF-α). Results After treatment, the scores of TCM syndromes and the levels of 24-hour urinary protein, SCr, BUN, eGFR, IL-6, IL-1, hs-CRP, and TNF-α in the 3 groups were decreased (P < 0.05 compared with those before treatment), and were markedly lower in group A than those in group B and group C (P<0.05); serum levels of IgA, IgG, IgM and ALB in the 3 groups were increased(P < 0.05 compared with those before treatment), and the levels in group A were significantly higher than those in group B and group C (P < 0.05). However, the levels of observation indexes did not arrive to the levels of group D(P<0.05). Conclusion Chinese medicine skin dialysis is effective on relieving the symptoms of CRF patients and enhancing the therapeutic effect through decreasing inflammatory mediators, inhibiting micro-inflammation and improving nutritional status.

[Clinical Study of Cytomegalovirus Infection and Preemptive Therapy after Allogenic Hematopoietic Stem Cell Transplantation].

OBJECTIVE: To analyze the clinical characteristics of cytomegalovirus(CMV) infection after allogenic hematopoietic stem cell transplantation(allo-HSCT) and the effect of preemptive therapy. METHODS: A total of 134 patients who underwent allo-HSCT from March 2010 to March 2015 in the Department of Hematology of our hospital were enrolled in this study. The CMV infection rate, the median time of CMV infection occurence, and the risk factors for CMV infection after allo-HSCT, the response rate of preemptive treatment and the median time of CMV-DNA turning negative were analyzed. Five-year overall survival rate was compared between the patients with or without CMV infection. RESULTS: The incidence of CMV viremia was 55.2％(74／134), and the median time for the CMV with CMV-DNA positive for the first time was 34 days(14-283) after allo-HSCT．Both univariate and multivariate analysis showed that the thymoglobulin(ATG) used in conditioning regimen and Ⅱ-Ⅳ grade of aGVHD were the risk factors for CMV viremia. After preemptive treatment the 85.1% of patient with CMV viremia turned negative, and the median time of CMV-DNA turning negative were 15 days(5-82), only 2 patients died of CMV pneumonia. Five-year overall survival rate of the patients with or wihout CMV viremia was 49% and 66.3% respectively, and the difference between the 2 groups was significant(P=0.041）. CONCLUSION: The ATG used in conditioning regimen and Ⅱ-Ⅳ grade of aGVHD may increase the incidence of CMV infection after allo-HSCT, and the preemptive thrapy can effectively prevent the CMV viremia turning to CMV disease.

Cerebral metabonomics study on Parkinson's disease mice treated with extract of Acanthopanax senticosus harms.

Extract of Acanthopanax senticosus harms (EAS) has neuroprotective effect on Parkinson's disease (PD) mice against dopaminergic neuronal damage. However, studies of its anti-PD mechanism are challenging, owing to the complex pathophysiology of PD, and complexity of EAS with multiple constituents acting on different metabolic pathways. Here, we have investigated the metabolic profiles and potential biomarkers in a mice model of MPTP-induced PD after treatment of EAS. Metabonomics based on ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) was used to profile the metabolic fingerprints of mesencephalon obtained from 1-Methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine Hydrochloride (MPTP-HCl)-induced PD mice model with and without EAS treatment. Through partial least squares-discriminate analysis (PLS-DA), it was observed that metabolic perturbations induced by MPTP were restored after treatment with EAS. Metabolites with significant changes induced by MPTP, including L-dopa, 5'-methylthioadenosine, tetradecanoylcarnitine, phytosphingosine-1-P, Cer(d18:0/18:0), LysoPC(20:4(5Z,8Z,11Z,14Z)), L-palmitoyl -carnitine, tetracosanoylglycine, morphiceptin and stearoylcarnitine, were characterized as potential biomarkers involved in the pathogenesis of PD. The derivations of all those biomarkers can be regulated by EAS treatment except Cer(d18:0/18:0), LysoPC(20:4(5Z,8Z,11Z,14Z)), morphiceptin. The therapeutic effect of EAS on PD may involve in regulating the tyrosine metabolism, mitochondrial beta-oxidation of long chain saturated fatty acids, fatty acid metabolism, methionine metabolism, and sphingolipid metabolism. This study indicated that changed metabolites can be certainly recovered by EAS, and the treatment of EAS can be connected with the regulation of related metabolic pathways.

A novel bacteriophage KSL-1 of 2-Keto-gluconic acid producer Pseudomonas fluorescens K1005: isolation, characterization and its remedial action.

BACKGROUND: Bacteriophages have the destructive damage on the industrial bioprocess. 2-Keto-gluconic acid (2KGA) producing bacteria had also been attacked and lysed by bacteriophages which lowered the glucose consumption and 2KGA yield and even stopped the fermentation process. In this study, we presented the characteristics of a novel virulent bacteriophage specifically infecting Pseudomonas fluorescens K1005 and proposed an efficient remedial action for this phage infection to reduce the production loss. RESULTS: The phage KSL-1 of Pseudomonas fluorescens K1005 was isolated from abnormal 2KGA fermentation broth. It belonged to the Siphoviridae family with a hexagonal head diameter of about 99 nm and a non-contractile tail of about 103 nm × 39 nm. The genome size of phage KSL-1 was estimated to be approximately 53 kbp. Its optimal MOI to infect P. fluorescens K1005 was about 0.001. One-step growth curve gave its latent and burst periods of 90 min and 75 min with a burst size of 52 phage particles per infected cell. This phage was stable with a pH range of 7.0-10.0, and sensitive to thermal treatment. Finally, a simple remedial action was proposed by feeding fresh seed culture. Compared with the infected 2KGA fermentation, the remedial experiments restored 2KGA fermentation performance by increasing the produced 2KGA concentration to 159.89 g/L and shortening the total fermentation time of 80 h with the productivity and yield of 2.0 g/L.h and 0.89 g/g. The obtained data proved that this method was effective to combat the phage infections problems during the 2KGA fermentation. CONCLUSION: The phage KSL-1 was a novel bacteriophage specifically infecting Pseudomonas fluorescens K1005. The remedial action of feeding fresh seed culture to the infected broth was an easily-operating and effective method to maintain a high 2KGA yield and avoid the draft of infected broth.

[Effects of Chinese herbal medicine Huanglian Jiedu Decoction on urine metabonomics of healthy people].

OBJECTIVE: To study biomarkers and their diversification in healthy people's urine before and after oral administration of Huanglian Jiedu Decoction, a compound Chinese herbal medicine, and to investigate the influence of Huanglian Jiedu Decoction on urine metabonomics of healthy people. METHODS: Ten healthy volunteers joined in this experiment and signed informed consent. They were given oral administration of Huanglian Jiedu Decoction for 3 days, and their urine was collected every morning. The samples were centrifuged at 13 000×g for 10 min at 4 degrees centigrade, and the supernatant was stored at -75 degrees centigrade. The samples were examined by a core technology called ultra-performance liquid chromatography quadruple time of flight/mass spectrometry and then the data were analyzed by principal components analysis and partial least square-discriminant analysis to investigate trajectory diversification and corresponding relations of the time and amount of urine metabonomics before and after oral administration of Huanglian Jiedu Decoction. RESULTS: Evident variance of urine metabonomics took place after oral administration of Huanglian Jiedu Decoction, which showed that the metabolism of healthy people had been visibly affected. Seven potential biomarkers which play a role in healthy people on behalf of Huanglian Jiedu Decoction were preliminarily found, and one biomarker which had biological significance was identified as 2-(formylamino)-benzoic acid. CONCLUSION: The effects of Huanglian Jiedu Decoction on urine metabonomics in healthy people investigated at the level of metabolism can provide theoretical and technical support for the study of Huanglian Jiedu Decoction's influence on heat syndrome.

[Successful treatment of agammaglobulinemia by HLA-mismatched unrelated cord blood stem cell transplantation--the first case report].

OBJECTIVE: To evaluate cord blood stem cell transplantation (CBT) in the treatment of X-linked agammaglobulinemia, and observe the courses of the hematopoietic and immune reconstitution. METHODS: A 14-year-old male patient with agammaglobulinemia received CBT from a 1/6 HLA-mismatched unrelated cord blood. The conditioning regimen was Bu/Cy/anti-CD3 antibody. CsA was given together with MMF and MTX for prophylaxis of GVHD. The patient received 0.42 x 10(8) nucleated cells/kg, containing 0.35 x 10(6) CD34(+) cells/kg. RESULTS: The recipient showed hematopoietic reconstitution on day 30 post-transplantation when ANC was 0.5 x 10(9)/L and BPC 20 x 10(9)/L. Sex chromosome analysis showed engraftment (donor 46, XX/recipient 46, XY = 4:1) on day 45. The recipient's blood group changed from AB to O, IgG from 1.1 g/L to 3.5 g/L, sex chromosome from 46, XY to full 46, XX, and mature B cells in peripheral blood from 0 to 5% on day 100, indicating immune reconstitution. At the last follow-up of 360 days, the patient without acute or chronic GVHD showed normal hemogram and myelogram, IgG 13.5 g/L and 10% mature B cells in peripheral blood, indicating the hematopoiesis and immune persistent reconstitution. No acute or chronic GVHD was developed. CONCLUSION: This is the first case report of successful treatment of X-linked agammaglobulinemia by HLA-mismatched unrelated CBT.