重庆市1264株粪肠球菌和屎肠球菌耐药性监测

【摘 要】 目的 了解2011年中国重庆市主要7所教学医院临床分离粪肠球菌和屎肠球菌对各类抗菌药物的耐药性。方法 重庆市主要7所教学医院(6所综合性医院,1所儿童医院)按统一方案、采用统一的材料、方法和判断标准(CLSI 2011年版)进行粪肠球菌和屎肠球菌的耐药性监测。数据用WHONET 5.5软件按照CLSI 2011年版折点进行分析。结果 共分离到非重复粪肠球菌589株、屎肠球菌675株,对利奈唑胺、万古霉素、替考拉宁仍极敏感,耐药率<2%,万古霉素耐药粪肠球菌和屎肠球菌检出率分别为0.3%、0.7%。粪肠球菌对青霉素、氨苄西林、呋喃妥因的耐药率较低,分别为14.8%、8.6%和5.1%,对高浓度庆大霉素的耐药率分别为46.9%;屎肠球菌耐药性明显高于粪肠球菌,对青霉素和氨苄西林耐药率接都在90％左右。儿童和成人耐药率存在一定差别。结论 本市医院肠球菌感染以屎肠球菌为主, 粪肠球菌次之,两者耐药性明显不同, 监测其耐药情况对指导临床用药具有重要意义。     

肠球菌属细菌的临床分布与耐药性分析

目的了解2012-2017年临床分离肠球菌的分布特征及耐药性,为临床合理用药提供依据。方法采用全自动微生物分析仪进行菌株鉴定及药敏试验,对肠球菌的临床分布与耐药情况进行统计分析。结果共分离出1 432株肠球菌,其中粪肠球菌为603株(42.11%),屎肠球菌为596株(41.62%)。肠球菌属细菌标本来源以尿液、胆汁和全血为主,分别占39.66%、34.50%和11.59%,其中粪肠球菌主要来自普外科、泌尿外科和ICU,而屎肠球菌主要来自ICU、普外科和消化内科。肠球菌总体对红霉素的耐药率最高(67.81%),其次为四环素(47.49%)、环丙沙星(47.00%)和左旋氧氟沙星(46.44%),对利奈唑胺和万古霉素的耐药率较低,分别为4.89%和1.19%。粪肠球菌对奎奴普丁/达福普汀、四环素的耐药率分别为83.91%和64.01%,明显高于屎肠球菌(均P0.05)。屎肠球菌对红霉素、青霉素G、氨苄西林、喹诺酮类的耐药率均超过85.00%,且均高于粪肠球菌(均P0.05)。粪肠球菌和屎肠球菌对利奈唑胺的耐药率分别为6.80%和2.18%,对万古霉素的耐药率分别为0.66%和0.67%。结论肠球菌感染病原菌以粪肠球菌和屎肠球菌为主,肠球菌属细菌对万古霉素和利奈唑胺仍然保持较高的敏感性,不同种的肠球菌其耐药性差异显著。     

肠球菌的临床感染与耐药性分析

目的了解温州医学院附属第一医院临床分离主要肠球菌的分布及其对常用抗菌药物的耐药现状,以指导临床合理用药。方法对2008年至2011年临床分离的635株粪肠球菌和屎肠球菌的标本来源和药敏结果进行回顾性分析。结果各种临床标本中两种肠球菌的分布比例存在差异,总体以尿液标本所占比例最多,且屎肠球菌的总体分离率高于粪肠球菌。粪肠球菌对利奈唑胺、氨苄西林、万古霉素、呋喃妥因和替考拉宁的耐药率都在5．0％以下,对莫西沙星和青霉素G的耐药率也仅为7．0％和6．7％;屎肠球菌对莫西沙星、左旋氧氟沙星、环丙沙星、氨苄西林、青霉素G和红霉素的耐药率都在90．0％以上,对利奈唑胺、万古霉素、替考拉宁和奎奴敏感。粪肠球菌的多重耐药株占总数的26．4％,屎肠球菌的多重耐药株占总数的78．2％。结论粪肠球菌和屎肠球菌对15种抗菌药物的耐药情况不同,屎肠球菌具有更高的耐药率和更广的耐药谱。临床应根据药敏试验的结果合理选择抗菌药物,以防止耐药菌株的产生和播散。     

粪肠、屎肠球菌及相近种部分持家基因的系统发育分析

【目的】利用16S rRNA、clpX和recA基因分子标记研究Enterococcus faecalis、Enterococcus faecium及相近种间的种系发育关系,并比较这些基因序列对E.faecalis、E.faecium及相近种的区分能力。【方法】以分离自传统乳制品中的9株E.faecium和1株E.durans分离株为研究对象,以clpX和recA基因片段为标记,通过PCR扩增、测序,结合已公布的近缘种相应序列构建系统发育树并与16S rRNA基因进行比较。【结果】在基于clpX和recA基因的进化树中,10株试验菌株与E.faecalis始终处于同一分支。与该物种这两个基因的平均相似性为99.6%和98.6%,与另一分支的Faecium-group(E.durans和E.faecium)的平均相似性仅为61.5%和33.5%。相近种E.durans和E.hirae间这两个基因的差异性为20.3%和39.0%;在基于16S rRNA基因的进化树中,试验菌株与Faecium-group(E.lactis、E.faecium、E.durans、E.hirae)处于同一分支。与这些成员间该基因的相似性大于99.6%,与E.faecalis基因的平均相似性可达98.4%。相近种间该基因相似性无明显差异。【结论】按照10株试验菌株clpX和recA基因的分析结果可将由传统生理生化和16S rRNA基因序列鉴定的9株E.faecium和1株E.durans归类为E.faecalis,clpX和recA基因可用于部分相近种的分类鉴定。     

屎肠球菌的临床分布与耐药性分析

目的了解医院屎肠球菌的临床分布和耐药情况,为临床抗感染的预防与治疗提供参考。方法回顾性分析1999年1月至2011年12月临床标本中分离的1161株屎肠球菌;用WHONET5．6软件分析耐药率变迁。结果临床分离的1161株屎肠球菌,在同期分离的1944株肠球菌属中占59．72％。主要分离自尿液和血液,分别占40．91％和26．87％;主要分离自外科病区、内科病区、ICU和儿科病区的菌株,分别占29．37％、25．15％、13．95％和13．53％;屎肠球菌对多种抗菌药物耐药,对万古霉素、替考拉宁和利奈唑胺的耐药率较低,分别为1．04％、0．94％和1．85％。结论屎肠球菌在临床的分离率逐年增加,已成为医院内感染的主要病原菌之一,其多药耐药和高耐药现象相当严重,目前万古霉素、替考拉宁和利奈唑胺仍然是治疗肠球菌属引起感染的有效药物。     

临床分离肠球菌的分布特点及耐药性变迁

目的分析临床分离肠球菌的分布特点及耐药性变迁,为临床合理应用抗生素提供依据。方法从中国医科大学附属盛京医院门诊和住院病人采集血液、尿液、手术切口分泌物、腹腔引流液等标本,应用法国生物梅里埃API细菌鉴定系统进行细菌的分离和种属鉴定,用K-B纸片琼脂扩散方法对不同肠球菌做药物敏感试验。结果肠球菌属的数量占全部分离到细菌种类的第6位。其中屎肠球菌的构成比,2002年为30.9%,2006年上升到71.6%,超过粪肠球菌的比例。肠球菌主要分布于ICU、普通外科、呼吸内科、肾内科、干诊、感染科和新生儿科病房。粪肠球菌对青霉素、氨苄西林和呋喃妥因的耐药率分别从2002年的53.6%、38.5%和30.4%下降到2006年的4.5%、0和4.5%,屎肠球菌对氯霉素的耐药率从2002年的37.1%下降到2006年的14.3%,两种菌对万古霉素和替考拉宁的敏感性均高,但临床出现耐万古霉素和耐替考拉宁的肠球菌。结论中国医科大学附属盛京医院临床肠球菌出现新的分布特点,屎肠球菌比例有增加趋势。调查肠球菌尤其是屎肠球菌感染的危险因素和采取相应的防治措施是当前研究的重要课题。     

肠球菌血流感染研究进展

近年来,随着肠球菌感染发生率的增加,有关肠球菌血流感染、尿路感染和腹腔感染的研究也逐渐增多,其中肠球菌血流感染病死率显著增加,预后不佳。临床上肠球菌感染以屎肠球菌和粪肠球菌为主。年龄、性别、致病菌种类及耐药情况、治疗方案的选择等均与肠球菌血流感染的发生和预后密切相关。此外,耐万古霉素肠球菌血流感染的治疗用药——达托霉素和利奈唑胺血药浓度及剂量的选择也被广泛关注。遵守临床操作规程,合理选择抗生素的种类及剂量浓度,严格监测药物使用情况,可减少血流感染的发生率,改善预后。现对肠球菌血流感染的发生及预后危险因素、治疗药物的选择进行综述,以期为肠球菌血流感染患者的临床治疗提供指导。     

屎肠球菌活菌制剂的研究

对屎肠球菌活菌制剂的特性及功效进行研究和分析。方法：通过分离菌种、筛选、鉴定、初步确定一株具有调节血脂作用的屎肠球菌菌株。经过毒理学试验、调血脂功能试验及人体试食试验来观察该制剂的作用。结果：毒理学试验结果证明屎肠球菌DM891129菌株属无毒、弱蓄积,无遗传毒性。功效试验表明高中低剂量组均只有降低实验高脂血症大鼠血胆固醇的作用。结论：屎肠球菌活菌制剂具有调节血脂作用。     

一株猪源屎肠球菌HDRsEf1益生特性研究

本研究旨在筛选一株具有优良生物特性的屎肠球菌,作为微生态制剂候选菌株;重点研究其与肠道相关的抗逆性能和对病原菌的抑菌活性。以自健康通城猪直肠内容物分离得到的屎肠球菌HDRsEf1为研究对象,通过耐受试验、抑菌试验研究该菌的益生特性。发现屎肠球菌HDRsEf1能够耐受浓度为0.5%的胆盐,并能在pH为2.0的酸中存活;对粘蛋白有显著的粘附作用(P0.05);能耐受70℃的温度。该菌发酵上清液能够抑制食源致病菌和畜禽临床主要致病菌的生长,结果表明屎肠球菌HDRsEf1可作微生态制剂菌株。     

粪肠球菌和屎肠球菌耐药性分析

目的 监测我院肠球菌中粪肠球菌株和屎肠球菌株的耐药性,为临床合理应用抗菌药物提供依据。方法 采用法国生物梅里埃公司的GPI板进行细菌鉴定及药敏试验,应用whonet5软件统计粪肠球菌和屎肠球菌的耐药率。结果 粪肠球菌和屎肠球菌对氯霉素、呋喃妥因、万古霉素有较好体外抗菌活性,耐药率都在50%以下,对万古霉素的耐药率在1%以下。粪肠球菌对青霉素、高水平庆大霉素、环丙沙星、利福平、红霉素等大部分抗菌素的耐药率有逐年下降趋势,而屎肠球菌对环丙沙星、利福平、呋喃妥因等抗菌素的耐药率则有上升趋势,屎肠球菌对大多数抗菌素耐药率都高于粪肠球菌。结论 粪肠球菌和屎肠球菌呈多重耐药,临床用药应结合药敏试验结果合理选择抗菌药物。     

一株粪肠球菌噬菌体的分离及其生物学特性研究

目的:利用临床耐药粪肠球菌分离裂解性噬菌体,为应用噬菌体治疗耐药粪肠球菌感染提供基础。方法:利用噬菌斑实验分离噬菌体并观察噬菌斑形态;双层平板培养法测定噬菌体效价、最佳感染复数及一步生长曲线;负染法电镜观察噬菌体形态;蛋白酶K/SDS法提取噬菌体基因组,酶切处理后琼脂糖凝胶电泳分析。结果:分离出一株噬菌体IME-EF1,该噬菌体能裂解多株临床分离的粪肠球菌;电镜观察呈蝌蚪形,最佳感染复数为1;通过绘制一步生长曲线,证明该噬菌体感染后的潜伏期为25 min,爆发期为35 min,裂解量为60 pfu。结论:研究结果表明利用临床分离的耐药粪肠球菌分离裂解性噬菌体是可行的,有望为耐药粪肠球菌的抗生素替代疗法奠定基础。     

水溶性蜂胶对感染根管内粪肠球菌的体外抑菌试验

目的 研究水溶性蜂胶对感染根管内常见菌粪肠球菌的体外抑菌活性.方法 采用液体稀释法测定水溶性蜂胶对粪肠球菌的最低杀菌浓度(Minimal bactericidal concentrations,MBC).结果 蜂胶对粪肠球菌的MBC为0.156%.结论 蜂胶对感染根管内粪肠球菌具有良好的抑菌活性,将其用于感染根管消毒具...     

The first molecular analysis of clinical isolates of VanA-type vancomycin-resistant Enterococcus faecium strains in Mainland China

AIMS: The aim of this study was to examine two VanA-type vancomycin-resistant Enterococcus faecium (VRE) strains that had been isolated from patients resident in mainland China. This is the first molecular analysis of clinical VRE strains being isolated in mainland China. METHODS AND RESULTS: Two VanA-type VRE isolates were isolated from in-patients at hospitals located in the Chinese cities Beijing and Dalian and were designated C264 and I125. The plasmids pC264V (40 kbp) and pI125V (370 kbp) that were isolated from C264 and I125, respectively, carried a Tn1546-like element encoding VanA resistance. The vancomycin-resistant plasmids pC264V and pI125V were transferred by filter mating at frequencies of 10(-7) and 10(-4) respectively. Sequence analysis of pC264V revealed that two IS1216V sequences and an IS1542 sequence were present within the Tn1546-like element. pI125V had two IS1216V insertions in the Tn1546-like element. CONCLUSIONS: The two VanA-type vancomycin-resistant E. faecium (VRE) strains C264 and I125 were isolated from in-patients in Chinese hospitals. The vancomycin-resistant conjugative plasmids pC264V and pI125V plasmids isolated from these strains carried the Tn1546-like element. The Tn1546-like element was found to contain the insertion sequences IS1216V and IS1542. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first molecular analysis of VanA-type VRE strains from patients resident in mainland China.     

Enterococcus faecalis: specific and non-specific interactions with human polymorphonuclear leukocytes

In previous studies we have demonstrated that the ability of Enterococcus faecalis to adhere to and to be internalized in human urinary tract epithelial cells, Girardi Heart cells and human polymorphonuclear leukocytes (PMNs), was dependent on whether the strain had been isolated from urinary tract infections (UTI) or endocarditis (EN) respectively. These properties were further modified by growth of the organism in human serum. In the present report, using competition assays we show that adhesins containing a D-glucose moiety play a role in mediating the interactions between human PMNs and E. faecalis strains isolated from UTI and grown in brain-heart infusion broth (BHIB). On the other hand, adhesins containing both D-glucose and D-galactose moieties were involved in the interactions between PMNs and serum grown UTI isolates or EN isolates grown in either BHIB or human serum. Moreover, the impairment in the association between both UTI and EN strains after growth in serum appears to be at least partially related to a decrease in enterococcal surface hydrophobicity.     

Characteristics and identification of enterocins produced by Enterococcus faecium JCM 5804T

AIMS: To screen bacteriocin-producing lactic acid bacteria (LAB) in 52 type and reference strains, which have not previously been studied, with respect to bacteriocins, and to characterize the presence of bacteriocins. METHODS AND RESULTS: Only Enterococcus faecium JCM 5804T showed bacteriocin-like activity. It inhibited the growth of Lactobacillus spp., Enterococcus spp., Clostridium spp., Listeria monocytogenes, and vancomycin resistant Enterococcus (VRE). However, it was not effective against Gram-negative strains, Weisella spp., Leuconostoc spp., Lactococcus spp., or methicillin resistant Staphylococcus aureus (MRSA). The inhibitory activity of Ent. faecium JCM 5804T was inactivated by proteinase K, trypsin, alpha-chymotrypsin, and papain, but not by lysozyme, lipase, catalase, or beta-glucosidase. The inhibitory activity was stable at 100 degrees C for 30 min, and had a pH range from 2 to 10. The molecular weight of the partially purified bacteriocin(s) was approx. 4.5 kDa, according to tricine-sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Polymerase chain reaction and direct sequencing methods identified three different types of bacteriocins produced by Ent. faecium JCM 5804T, enterocin A, enterocin B, and enterocin P-like bacteriocin. CONCLUSION: Enterococcus faecium JCM 5804T produced three different types of bacteriocins, and they inhibited LAB and pathogens. SIGNIFICANCE AND IMPACT OF STUDY: This is the first report of enterocin A, enterocin B, and enterocin P-like bacteriocin, detected in Ent. faecium JCM 5804T among LAB type and reference strains.     

Specific antibody promotes opsonization and PMN-mediated killing of phagocytosis-resistant Enterococcus faecium

Many clinical isolates of Enterococcus faecium are resistant to neutrophil (PMN)-mediated phagocytosis and killing in the presence of normal human serum. We have now examined the ability of specific polyclonal rabbit antibodies to promote opsonization and killing of phagocytosis-resistant E. faecium. Immune rabbit serum generated against formalin-killed E. faecium TX0016, a phagocytosis-resistant strain, markedly promoted binding of TX0016 organisms to PMNs and PMN-mediated killing. These effects were dramatically reduced by (a) adsorption of immune serum with E. faecium TX0016, but not by adsorption with a strain of E. faecium susceptible to phagocytosis, and (b) incubation of immune serum with carbohydrate purified from TX0016, but not by incubation with a surface protein extract from TX0016. IgG purified from immune serum was unable by itself to promote bacterial binding to PMNs. However, specific IgG was able to promote binding to PMNs and PMN-mediated killing in the presence of normal human serum as a complement source, as were F(ab')(2) and Fab fragments produced from it, and the alternative pathway of complement was sufficient to promote IgG- and F(ab')(2)-mediated opsonization. PMN complement receptor type 3, but not complement receptor type 1, was involved in bacterial binding to PMNs induced by the combination of F(ab')(2) fragments and normal human serum. These results suggest that opsonization by antibodies potentially directed against bacterial carbohydrate, in conjunction with complement activation, has an important role in the host defense against phagocytosis-resistant E. faecium.     

Cross‐transmission of clinical Enterococcus faecium in relation to esp and antibiotic resistance

Aims: To investigate clonality among clinical Enterococcus faecium isolates and normal intestinal microflora isolates as well as cross‐transmission between patients in relation to the presence of the esp gene and antibiotic resistance. Methods and Results: Blood‐culture isolates (n = 101) deriving from tertiary, secondary and primary hospitals were analysed. Antibiotic susceptibility was investigated. Polymerase chain reaction and pulsed‐field gel electrophoresis were used for detection of esp and genotyping, respectively. Nearly half (43%) of the patients included were involved in a cross‐transmission event with Ent. faecium. These strains disseminated both within and between all hospitals. The antibiotic resistance and presence of esp were highest in isolates from the tertiary hospital. Isolates harbouring esp showed less genetic diversity compared with esp negative ones. Conclusions: Cross‐transmission with Ent. faecium between patients was readily detected, indicating that hospital‐adapted clones circulate within and between hospitals. Acquired characteristics, such as antibiotic resistance and esp, seem to accumulate in the isolates disseminating in the tertiary hospital. Significance and Impact of the Study: It is important to characterize Ent. faecium isolates causing infections and to determine the extent of dissemination in order to prevent further spread of these pathogens.     

Characteristics of bacteriocin-like inhibitory substances from dochi-isolated Enterococcus faecium D081821 and D081833

AIMS: To characterize bacteriocin-like inhibitory substances (BLIS) from two dochi-isolated Enterococcus faecium. METHODS AND RESULTS: Enterococcus faecium D081821 and D081833 were isolated from dochi (a traditional fermented food in Taiwan) and found to produce BLIS with inhibitory activities against Listeria monocytogenes, Clostridium perfringens, and Bacillus cereus. Strains D081821 and D081833 showed different growth temperatures and their BLIS showed different sensitivities to heat, proteolytic enzymes, and antibacterial spectra. Both BLIS were collected, partially purified, and analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). SDS-PAGE showed that both partially purified BLIS were approximately 3.0 kDa in size. CONCLUSIONS: These results indicate that E. faecium D081821 and D081833 produce different BLIS with strong antibacterial actions against the tested pathogenic bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study suggest that two different BLIS from dochi-isolated lactic acid bacteria have potential for use as food preservatives.