Distribution and antibiotic resistance of 532 bacterial panthogens in blood culture in a hematology de-partment / 中华微生物学和免疫学杂志

Objective To analyze the distribution and antibiotic resistance of bacterial pathogens in blood culture in the Department of Hematology of the First Affiliated Hospital of Nanjing Medical Universi-ty from 2013 to 2017. Methods BACTEC FX automatic blood culture instrument was used for blood cul-ture. VITEK-2 Compact automatic microbial identification and drug susceptibility system was used for bacte-rial identification, drug sensitivity testing and fungal identification. K-B and ATB FUNGUS 3 methods were used for Streptococcus and fungi susceptibility testing, respectively. WHONET5. 6 software was used for sta-tistical analysis. Results The positive results of blood culture accounted for 7. 6% during 2013 to 2017 and the top five isolated pathogenic bacteria were Escherichia coli (20. 9% ), Klebsiella pneumoniae (18. 0% ), coagulase negative Staphylococcus (12. 4% ), Pseudomonas aeruginosa (11. 5% ) and Enterobacter cloacae (3. 6% ). Carbapenems (94. 4% ) were the most active antibiotics against Enterobacteriaceae. Pseudo-monas aeruginosa strains were sensitive to all antibacterial dugs except aztreonam. Acinetobacter baumannii strains had a lower sensitivity to almost all antibacterial dugs. Vancomycin, linezolid and teicoplanin were active antimicrobial agents against gram-positive cocci. Conclusion Patients with hematological diseases are more susceptible to nosocomial infection, which should be paid more attention to in clinical and laborato-ry practice. Rational antimicrobial therapy is an effective way for control of antimicrobial resistance.

Effects of betaine on formation and dispersion of Pseudomonas aeruginosa biofilm and its drug-resistance / 临床检验杂志

Objective To investigate the effects of betaine on the formation and dispersion of biofilm of Pseudomonas aeruginosa and its drug-resistance.Methods A total of 20 strains of Pseudomonas aeruginosa were obtained from clinical inpatients.The biofilm formation abilities of the Pseudomonas aeruginosa were evaluated by violet staining,and the effects of betaine on the formation and dispersion of biofilm were studied.The minimum inhibitory concentration (MIC) values of Pseudomonas aeruginosa on ciprofloxacin were compared with the controls when biofilm was formed and inhibited.Results Biofilm was formed in all the 20 strains of Pseudomonas aeruginosa in 24 hours with absorbance (A590 nm) (1.90 ± 0.66).Betaine significantly inhibited biofilm formation of Pseudomonasaeruginosa in 24 hours compared with control group(t =4.36,P ＜ 0.01) and the maximum inhibition reached in 48 hours with absorbance(A590 nm) (1.12 ±0.60).The maximum dispersion of betaine on mature biofilm of Pseudomonas aeruginosa reached in 24 hours.The MIC range of ciprofloxacin to the 20 strains of Pseudomonas aeruginosa was 0.03 to 4 μg/mL with 0.25 μg/mL of MIC50 and 2 μg/mL of MIC90.After the biofilm was inhibited by belaine,the MIC of ciprofloxacin to Pseudomonas aeruginosa did not changed.The MIC of ciprofloxacin to biofilm-formed Pseudomonas aeruginosa was more than 16 μg/mL.Conclusion Betaine could effectively inhibit the formation of biofilm and disperse the mature biofilm of Pseudomonas aeruginosa,which may provide more choices for the treatment of clinical infection.The germicidal efficacy of ciprofloxacin has no changed on the biofilm-formed bacteria when inhibition of betaine was involved.

Inhibitory and dispersive effects of betaine on formation of Staphylococcus aureus biofilm / 临床检验杂志

Objective To observe the formation of Staphylococcus aureus biofilm and the inhibitory and dispersive effects of betaine on the biofilm.Methods The inhibitory and dispersive effects of 0.1％ betaine on the biofilm from 20 strains of Staphylococcus aureus were examined by crystal violet assay.Results All the 20 strains of Staphylococcus aureus formed biofilm.The biofilm of methicillinsensitive Staphylococcus aureus (MSSA) was formed in 24 hours with peak value of absorbance (A590 nm) (1.99 ± 0.53).The biofilm of methicillin-resistant Staphylococcus atureus(MRSA) was formed in 48 hours with peak value of absorbance(A590 nm) (1.13 ±0.47).After adding betaine,the absorbance(A590 nm) of MSSA biofilm fell down to(1.74 ± 0.61) in 24 hours,while the absorbance(A590 nm) of MRSA biofilm fell down to(0.40 ± 0.12) in 48 hours,which was significantly reduced compared with the controls (t =2.43,5.84,P ＜ 0.05 respectively).When adding betaine after the biofilm formed,the absorbancies (A590 nm) of both MSSA and MRSA showed no significant difference compared with the controls (P ＞ 0.05).Conclusion Betaine could inhibit biofilm formation of Staphylococcus aureus at concentration of 0.1％,but it could not disperse the mature biofilm of Staphylococcus aureus.

Pathogen distribution and susceptibility profile of fungal isolates from bloodstream infections during the period from 2013 through 2015 / 中国感染与化疗杂志

Objective To investigate the pathogen distribution and susceptibility profile of fungal isolates from bloodstream infections,and valuate the clinical utility of G test in diagnosis of fungal infections for the purpose to improve antifungal therapy.Methods A retrospective analysis was carried out to analyze the fungal pathogens isolated from bloodstream infections in the First Affiliated Hospital of Nanjing Medical University during the period from January 2013 through December 2015 and their antimicrobial susceptibility.Results A total of 114 fungal strains were isolated from bloodstream infections during the 3-year period,most of which were Candida (99/114,86.8％),especially Candida albicans (30.7％).About 41.2％ (47/114) of the fungal strains were isolated from Department of Thoracic Surgery (10,5 and 4 strains in 2013,2014 and 2015),Hematology (11 strains in 2014),and ICU (7 strains in 2014).Antimicrobial susceptibility testing showed that all the fungal strains (100％) were susceptible to amphotericin B,but 83.5％ susceptible to itraconazole (the lowest).G test was positive before the result of blood culture in 13 of the 54 patients who received G test.Conclusions Candida was the most common fungus in fungal bloodstream infection.Amphotericin B is the most active antifungal agent in vitro.Blood culture combined with serological test can provide clinicians an earlier and reliable diagnosis.

Development and Evaluation of a Duplex Real-Time PCR Assay With a Novel Internal Standard for Precise Quantification of Plasma DNA

BACKGROUND: Circulating levels of cell-free DNA increase in many pathologic conditions. However, notable discrepancies in the quantitative analysis of cell-free DNA from a large number of laboratories have become a considerable pitfall, hampering its clinical application. METHODS: We designed a novel recombinant DNA fragment that could be applied as an internal standard in a newly developed and validated duplex real-time PCR assay for the quantitative analysis of total cell-free plasma DNA, which was tested in 5,442 healthy adults and 200 trauma patients. RESULTS: Compared with two traditional methods, this novel assay showed a lower detection limit of 0.1 ng/mL, lower intra- and inter-assay CVs, and higher accuracy in the recovery test. The median plasma DNA concentration of healthy males (20.3 ng/mL, n=3,092) was significantly higher than that of healthy females (16.1 ng/mL, n=2,350) (Mann-Whitney two-sample rank sum test, P<0.0001). The reference intervals of plasma DNA concentration were 0-45.8 ng/mL and 0-52.5 ng/mL for healthy females and males, respectively. The plasma DNA concentrations of the majority of trauma patients (96%) were higher than the upper normal cutoff values and were closely related to the corresponding injury severity scores (R²=0.916, P<0.0001). CONCLUSIONS: This duplex real-time PCR assay with a new internal standard could eliminate variation and allow for more sensitive, repeatable, accurate, and stable quantitative measurements of plasma DNA, showing promising application in clinical diagnosis.

Clinical application evaluation of Xpert detection system of Clostridium difficile / 临床检验杂志

Objective To evaluate the clinical application value of Xpert detection system of Clostridium difficile (C.difficile).Methods A total of 43 stool specimens from the patients with diarrhea were collected,and C.difficile in stool specimens were detected by the Xpert detection system,the toxigenic culture method,and the toxin detection method which detected the toxin of C.difficile by VⅥDAS automatic analyzer after anaerobic culture,respectively.The analytic performance of Xpert detection system was evaluted based on the toxigenic culture method as the gold standard.Meanwhile,the consistency of the results from different detection methods was compared.The ribotype 027 strain (ATCC BAA-1870) simulating the stool specimen was further used to verify the Xpert detection system.Results Based on the gold standard of the toxigenic culture method,the sensitivity,specificity,positive predictive value and negative predictive value of the Xpert detection system were 90.9％,93.8％,83.3％ and 96.8％,respectively.The Kappa values for the consistency between the Xpert detection system and the toxigenic culture method or the toxin detection method were 0.822 (P ＜ 0.05) and 0.419 (P ＜ 0.05),respectively.Moreover,the ribotype 027 strain simulating the stool specimen was verified by the Xpert detection system successfully.Conclusion The Xpert detection system may rapidly and accurately detect the C.difficile in stool specimens,especially the ribotype 027 strain with high toxicity.

Distribution and antimicrobial resistance of clinical gram-negative bacteria in the First Afifliated Hospital of Nanjing Medical University during 2014 / 中国感染与化疗杂志

Objective To investigate the distribution and antimicrobial resistance profile of clinical gram-negative bacterial isolates in the First Afifliated Hospital of Nanjing Medical University during 2014.Methods Bacteria identiifcation was performed by API system or the VITEK-2 Compact automatic identiifcation system. Disk diffusion susceptibility testing or VITEK-2 Compact automatic identification system was used to determine the susceptibility to antimicrobial agents. All data were analyzed using WHONET 5.6 software.Results Among the total 7 931 clinical isolates in 2014, gram-negative bacteria accounted for 64.2% (5 088/7 931). The top three pathogens wereE. coli,A. baumannii andK. pneumoniae. Notably, during the year 2014, 195 strains of carbapenem-resistantEnterobacteriaceaewere isolated, about 6.9% of all theEnterobacteriaceae isolates. Meanwhile, 613 (66.5%) strains of multiple drug resistantA. baumannii and 197 (28.7%) strains of multiple drug resistantP. aeruginosa were isolated.Conclusion During the year 2014, the resistance of the gram-negative bacteria in this hospital is mainly characterized by carbapenem-resistantEnterobacteriaceae, multiple drug resistant A. baumanniiand multiple drug resistantP. aeruginosa. Surveillance of antimicrobial resistance is beneifcial for rational use of antibiotics.

Clinical application of quantitative detection of plasma DNA in evaluating the effect of chemotherapy on chronic leukemia / 医学研究生学报

Objective Plasma circulating DNA can be em-ployed in place of bone marrow examination for the auxiliary diagnosis of leukemia.This study aimed to explore the clinical application of the plasma DNA level in evaluating the effect of chemotherapy on chronic leukemia. Methods We collected blood samples from 52 patients with chronic myelogenous leukemia (CML) (33 in the chronic phase, 7 in the acceleration phase, and 12 in the blast phase) , 85 with chron-ic lymphocytic leukemia (CLL) (28 with complete remission, 27 with partial remission, and 30 with no remission), 4 patients with hairy cell leukemia (HCL), and 80 healthy subjects.We simultaneously obtained plasma DNA and recombinant plasmid DNA using the BI-LATEST DNA Kit and examined the human β-actin gene and the level of plasmid DNA by real-time quantitative PCR. Results Before chemotherapy, the median value of plasma DNA was 149.46(30.63-496.91)ng/ml in the CML and 101.54(69.10-258.14) ng/ml in the CLL patients, both significantly higher than in the healthy controls (19.05[12.67-25.92]ng/ml) (P<0.01).After chemotherapy, the plasma DNA level of the CML patients was remarkably decreased, but still higher than that of the controls ( P<0.01).The CML patients in the chronic phase showed a markedly higher level of plasma DNA (302.89[93.33-541.52]ng/ml) than those in the blast phase (43.19[23.54-70.03]ng/ml) and acceleration phase (28.11[16.21-92.07]ng/ml) (P<0.05).The CLL patients with CR exhibited a significantly lower level of plasma DNA (24.29[14.64-30.74]ng/ml) than those with PR (106.88 [96.23-143.25]ng/ml) and NR (460.73[284.57-653.38〗ng/ml) (P<0.01), but all dramatically higher than that of the healthy controls (P<0.01) Conclusion The quantification of plasma DNA has a clinical application value in evaluating the effect of chemo-therapy on chronic leukemia.

Detection of drug resistance genes in integron of Shigella sonnei from six cities of Jiangsu Province in 2011 / 中华传染病杂志

Objective To investigate the resistance status of different integrons of Shigella sonnei (S.sonnei) and to analyze the distribution of resistant genes in integrons in Jiangsu Province.Methods A total of 32 strains of S.sonnei isolated from six cities of Jiangsu Province in 2011 were collected.The antibiotic susceptibility was tested by disk diffusion method.The molecular homology was analyzed by pulsed field gel electrophoresis (PFGE).The detection and classification of integrons were achieved by analyzing the positive polymerase chain reaction (PCR) products using restriction fragment length polymorphism (RFLP).RFLP and DNA sequencing were used to analyze the resistance genes in integrons.Results Multi-drug resistance (MDR) was detected in 28 (87.5％) S.sonnei strains.The resistant rates to ampicillin,nalidixic acid and tetracycline were highest (87.5％,respectively).However,it was sensitive to norfloxacin.PFGE analysis showed that there were 3 kinds of homologous clones involving 31 strains of the 32 S.sonnei strains.Among them,2,5 and 24 strains had the same clones,respectively.Accordingly,they spread within one,two and five different cities.The detection rates of class 1,class 2 and the atypical class 1 integrons in S.sonnei were 62.5％ (20/32),81.3％ (26/32) and 21.9％ (7/32),respectively,and no class 3 integron was detected.Sequence analysis of class 1 integron variable area revealed that it contained multiple resistant genes (aacA4-cmlA1 and dfrA1-aadA 1) ; dfrA1-sat 1-aadA 1 from class 2 integron and blara-30-aadA 1 from atypical class 1 integron were also identified.Conclusions In 2011,homologous S.sonnei strains spread among different cities in Jiangsu Province.MDR strains are prevalent and integrons are widespread which mediated the emergence of MDR strains.

Study of quantitative detection of circulating DNA in the plasma of patients with cervical lesion / 中华妇产科杂志

Objective To quantitatively detect circulating DNA levels in the plasma of patients withcervical lesion and to determine the value for diagnosis of cervical lesion and cervical cancer . Methods Preoperative blood samples were collected from 53 cases of low-grade lesions, 49 cases of high-grade lesions, 44 cases of cervical invasive cancer and 70 cases of healthy women. Plasma DNA was extracted by magnetic bead method (BILATEST DNA kit). The quantity of plasma DNA was determined by duplex real-time quantitative PCR. Results Median plasma DNA level of invasive cervical cancer patients was 61. 59 mg/L (32. 06 - 162. 16 mg/L) , which was significantly higher than that of healthy women [16. 35 mg/L(11. 98 -22.71 mg/L), P 0. 05). Median plasma DNA level of stage I patients was lower than that of stage Ⅱ- Ⅲ patients (46. 02 versus 71. 35 mg/L, P <0. 05). Conclusion Quantitatively detecting plasma circulating DNA may be with some application prospect in the diagnosis of cervical diseases.

Distribution and antimicrobial resistance change of blood culture isolates from the year 2004 to 2007 / 中华检验医学杂志

n positive blood culture, and they are resistant to a variety of antimicrobial agents, which should be called attention.

Assessment for hepatocyte injury of severe hepatitis patients by quantitative analysis of plasma DNA / 临床检验杂志

Objectives To evaluate accurately hepatocyte injury degree of severe hepatitis patients by quantifying plasma DNA of severe hepatitis patients and study its clinical application in diagnosis of severe hepatitis comparing with ALT.Methods Six milliliters of peripheral blood samples were collected from 185 patients with hepatitis B which are divided into four groups, severe hepatitis with 30 cases, acute hepatitis with 20 cases, chronic B hepatitis with 90 cases, and liver cirrhosis with 45 cases. 100 healthy controls were enrolled. Circulating DNA was extracted from plasma by the BILATEST virus DNA/RNA extraction kit and quantified with a novel duplex real-time PCR assay, respectively.Results Plasma DNA levels of hepatitis B patients were significantly higher than those of healthy controls (104.2 ng/ml vs. 23.4 ng/ml (median),P=0.0000).A significant difference of plasma DNA concentration was found between severe hepatitis and acute hepatitis (P=0.0018), and chronic B hepatitis (P=0.0000), and liver cirrhosis (P=0.0000).The median value of serum ALT of hepatitis B patients was 107.5 U/L, much higher than that of the healthy controls (24.1 U/L,P=0.0000).The levels of serum ALT were significantly different between severe hepatitis and acute hepatitis (P=0.0024), while there was no remarkable difference between severe hepatitis and chronic B hepatitis (P=0.0600), liver cirrhosis (P=1.0000). Moreover, for distinguishing severe hepatitis from liver cirrhosis and chronic B hepatitis, the plasma DNA assay was notably superior to ALT by the analysis of receive operating characteristic (ROC) curves (AUC, 0.95 vs. 0.51,P=0.0000; 0.86 vs. 0.34,P=0.0000).Conclusion The results by measuring plasma DNA of hepatitis B patients with the novel duplex real-time quantitative PCR showed that plasma DNA may be considered as a robust predictive marker for accurately evaluating hepatocyte injury degree of severe hepatitis patients.