血液感染优势细菌和真菌种类分析及Verigene-DNA芯片快速检测效果的研究

发布时间：2019-06-26 15:18

【摘要】：目的了解杭州市第三人民医院2008～2014年血液感染细菌和真菌的种类及其耐药性,探讨Verigene-DNA芯片快速检测血液感染革兰阳性菌的效果。方法采用BacT/ALERT3D血液标本培养系统及单侧单瓶、单侧双瓶和双侧双瓶法,分别对临床送检19789人份外周血标本中细菌和真菌进行分离培养,VITEK2Compact系统对获得的纯培养菌株进行鉴定,分别采用VITEK2Compact系统和纸片扩散法检测各菌株对不同抗生素的敏感性。采用CLSI推荐的纸片扩散法确证试验检测细菌分离株p-内酰胺酶活性。采用统计学软件对不同血培养模式的分离培养阳性率、优势感染细菌或真菌种类、分离菌株药物敏感性进行分析。另采用Verigene-DNA芯片对分离的98株革兰阳性菌进行鉴定并检测其耐药基因,菌种鉴定结果与VITEK2Compact系统进行比较,耐药基因检测结果用PCR复核。结果上述19789外周血标本中,双侧双瓶法送检的细菌和真菌血培养阳性率(29,22%)显著高于单侧单瓶法(8.28%)和单侧双瓶法(12.34%)(p0.01)。分离的124种1876株细菌中,人葡萄球菌(Staphylococcus hominis)、大肠埃希菌(Escherichia coli)、金黄色葡萄球菌(Staphylococcus aureus)、表皮葡萄球菌(Staphylococcus epidermidis)、溶血葡萄球菌(Staphylococcus haemolyticus)、肺炎克雷伯菌(Klebsiella pneumonia)、头状葡萄球菌(Staphylococcus capitis)、屎肠球菌(Enterococcus faecium)、鲍曼不动杆菌(Acinetobacter baumannii)和粪肠球菌(Enterococcus faecalis)分离率为11.46%-3.00%,其余114种细菌分离率均在2.3%以下。分离的18种136株真菌中,近平滑念珠菌(Candida parapsilosis)、白色念珠菌(Candida albicans)、光滑念珠菌(Candida glabrata)和季也蒙念珠菌(Candida guilliermondii)分离率分别为25.00%、21.32%、16.91%和11.76%,其余14种真菌分离率均在8%以下。所有葡萄球菌属菌株均对利奈唑胺、万古霉素和替加环素敏感,但耐甲氧西林凝固酶阴性葡萄球菌(methicillin-resistant CoNS, MRCoNS)检出率(87.88%,116/132)明显高于耐甲氧西林金黄色葡萄球菌(methicillin-resistant S. aureus, MRS A)(54.77%,109/199)(p0.01)。92株屎肠球菌和56株粪肠球菌中,替加环素敏感率均为100%,利奈唑胺敏感率分别为97.01%和95.56%,万古霉素耐药肠球菌(vancomycin-resistant enterococcus, VRE)检出率分别为15.22%(14/92)和8.93%(5/56)。208株大肠埃希菌对亚胺培南、厄他培南、替加环素、头孢替坦、阿米卡星、哌拉西林/他唑巴坦和呋喃妥因的敏感率为92.31%-99.52%,但有1.03%和0.48%菌株对碳青霉烯类抗菌药物厄他培南和亚胺培南耐药,其中54.31%菌株检出有β-内酰胺酶活性。122株肺炎克雷伯菌对阿米卡星、头孢替坦、替加环素和复方新诺明敏感率为77.05%-79.51%,但有32.71%和31.15%分别对碳青霉烯类抗菌药物厄他培南和亚胺培南耐药,其中28.21%菌株检出有β-内酰胺酶活性。71株鲍曼不动杆菌对多粘菌素B、替加环素、阿米卡星和头孢哌酮/舒巴坦的敏感率为60%-100%。29株白色念珠菌对氟康唑、伊曲康唑、克霉唑、5-氟胞嘧啶、制霉菌素和两性霉素B的敏感率均为100%；34株近平滑念珠菌对5-氟胞嘧啶和制霉菌素敏感率均为100%,但对两性霉素B、伊曲康唑、氟康唑和克霉唑敏感率为85.71%～97.06%；23株光滑念珠菌对5-氟胞嘧啶、两性霉素B和制霉菌素敏感率均为100%,但对克霉唑、伊曲康唑和氟康唑敏感率为56.52%～85.71%；16株季也蒙念珠菌对5-氟胞嘧啶和两性霉素B敏感率均为93.75%,但对氟康唑和伊曲康唑敏感率均仅为6.25%。Verigene-DNA芯片对16种98株革兰阳性菌鉴定结果与VITEK2Compact系统鉴定结果总符合率为91.84%(90/98),其中对葡萄球菌属、肠球菌属和链球菌属菌株检测符合率分别为91.67%(55/60),92.59%(25/27)和90.00%(9/10)。Verigene-DNA芯片法鉴定需时约3.1h,但VITEK2Compact系统鉴定需时约43h。22株金黄色葡萄球菌和12株表皮葡萄球菌中,Verigene-DNA芯片法分别检出21和8株携带甲氧西林耐药相关mecA基因,PCR检测结果与其完全一致；9株粪肠球菌和16株屎肠球菌中,Verigene-DNA芯片法分别检出1和5株携带万古霉素耐药相关vanA基因,PCR检测结果与之相同,但PCR检测结果显示有2株屎肠球菌携带vanM基因。结论双侧双瓶法可提高血标本中细菌和真菌分离培养阳性率。优势血液感染细菌依次为人葡萄球菌、大肠埃希菌、金黄色葡萄球菌和表皮葡萄球菌(10%),优势血液感染真菌依次为近平滑念珠菌、白色念珠菌、光滑念珠菌和季也蒙念珠菌(10%)。尽管不同种类的细菌和真菌临床菌株对不同抗菌药物的敏感性有差异,但革兰阳性菌株对利奈唑胺和替加环素有较高敏感率,革兰阴性菌株对替加环素和阿米卡星有较高敏感率,真菌菌株对5-氟胞嘧啶、两性霉素B和制霉菌素有较高敏感率。与VITEK2Compact系统比较,Verigene-DNA芯片法鉴定血液感染革兰阳性菌时具有快速、准确、简便并能同时检测细菌耐药基因的优点,值得在临床实验室中推广与应用。
[Abstract]:Purpose To understand the species and drug resistance of the blood-infected bacteria and fungi in the third People's Hospital of Hangzhou from 2008 to 2014, and to study the effect of the Veriene-DNA chip on the rapid detection of the Gram-positive bacteria in the blood Fruit. Methods The bacteria and fungi were isolated and cultured in 19789 human peripheral blood samples by using BacT/ ALERT3D blood specimen culture system and one-sided single-bottle, one-sided double-bottle and two-sided double-bottle method. The strains were identified by the VITEK2Compact system and the paper-paper diffusion method, respectively. Sensitivity of the element. The test-tested bacterial isolates, p-, were confirmed using the CLSI-recommended method of paper diffusion. The positive rate of isolation and culture of different blood culture modes, the superiority of the bacteria or the fungi, and the drug sensitivity of the isolated strains were investigated by using the statistical software. Sex analysis was carried out. The isolated 98 strains of Gram-positive bacteria were identified by the Veriene-DNA chip and their drug-resistant genes were tested. The results of the strain identification were compared with the VITEK2Compact system. PC Results In the above 19789 peripheral blood samples, the positive rate of bacterial and fungal blood culture (29,22%) of the two-sided double-bottle method was significantly higher than that of one-sided single-bottle (8.28%) and one-sided double-bottle (12.34 %) (p0.01). Among the 124 isolated strains of 1876, Staphylococcus hominis, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Klebsiella pneumoniae, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Klebsiella pneumoniae, Staphylococcus, Staphylococcus, Staphylococcus, Klebsiella pneumoniae, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Staphylococcus, Klebsiella pneumoniae, Staphyloc@@ S (s capitis), Enterococcus faecium, Acinetobacter baumannii and Enterococcus faecalis were 11.46%-3.00% and the remaining 114 species were isolated. The separation rates of Candida parapsilosis, Candida albicans, Candida glabrata and Candida guillimondii were 25.00%, 21.32%, 16.91% and 11.76%, respectively, and the other 14 fungi were isolated. The isolation rate was less than 8%. All of the strains of Staphylococci were sensitive to rilenalidomide, vancomycin and tigecycline, but the rate of methicillin-resistant coagulase-negative Staphylococci (MRCoNS) (87.88%,116/132) was significantly higher than that of methicillin-resistant S. aureus, MRS A) (54.77%,109/199) (p0.01). The sensitive rate of tigecycline was 100% in 92 strains of Enterococcus faecium and 56 strains of Enterococcus faecium, and the sensitive rate of the rilenalamine was 97.01% and 95.56%, respectively. The detection rates of vancomycin-resistant enterococci (VRE) were 15.22% (14/92) and 8.93% (5/56) respectively. The sensitivity of imipenem, ertapenem, tigecycline, cefotitan, amikacin, zacillin/ tafluvial/ tafluvial and babbitum was 92.31%-99.52%, but 1.03% and 0.48% of the strains were resistant to carbapenem The sensitivity of 122 strains of Klebsiella pneumoniae to amikacin, cefotitan, tigecycline and compound neomycin was 77.05%-79.51%, but 32.71% and 31.15% respectively applied to carbapenem antibacterial drug. The sensitivity of the 71 strains of Acinetobacter baumannii to polymyxin B, tigecycline, amikacin, and ceftriaxone/ sulbactam was 60% to 100%, and 29 strains of Candida albicans were sensitive to fluconazole, Iqu concha, g. The sensitive rate of mycotoxin,5-fluorophonate, nystatin and amphotericin B was 100%, and the sensitivity of 34 strains of near-smooth Candida to 5-fluorophonate and myxin B was 100%. The sensitive rate of the 5-fluorophonate, the amphotericin B and the myxin B was 100%, but the sensitive rate was 56.52% ~ 85.71%, and the sensitive rate was 56.52% ~ 85.71%. The 16 strains were also sensitive to 5-fluoro-cytomycin and amphotericin B. The results of the identification of 16 98 strains of Gram-positive bacteria and VITEK2Compact system were 91.84% (90/98), 91.67% (55/60), 92.59% (25/27) and 90.00% (9/10), respectively. At the time of the identification, about 3.1 h, but when the VITEK2Compact system was identified for about 43 h.22 strains of S. aureus and 12 S. epidermidis, the Verigene-DNA chip method was used to detect 21 and 8 strains of methicillin-resistant related mecA gene, and the result of the PCR was completely consistent with that of the 9 strains of Enterococcus faecalis. And 1 and 5 strains of vancomycin-resistant vanA gene were detected by the Veriene-DNA chip method, and the results of PCR were the same, but the results of PCR showed that 2 strains of enterococcus faecium were found. bacteria-carrying Conclusion The two-sided double-bottle method can increase the blood sample. The positive rate of bacterial and fungal isolates was positive. The dominant blood-infected bacteria were Staphylococcus, E. coli, S. aureus and Staphylococcus epidermidis (10%), and the dominant blood-infected fungi were Candida, Candida albicans, and Nostoglobus glabrata. In spite of the differences in the sensitivity of different kinds of bacteria and fungal clinical strains to different antibacterial drugs, the Gram-positive strain has a high sensitivity to the tigecycline and tigecycline, and the Gram-negative strains are highly sensitive to tigecycline and Amicka. 5-fluoro-1-(5-fluoro-1)-(2-)-(2-)-(5-fluorophenyl) in comparison with that VITEK2compact system, the Veriene-DNA chip method has the advantages of fast, accurate, simple and convenient and simultaneous detection of the bacterial drug resistance gene,
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R446.5

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