Biological and physico-chemical assessment of hydroxyapatite (HA) with different porosity

HA with specific internal porosities was loaded with different antibiotics (ATBs) and then tested on its microbiological effectiveness. The HA purity was controlled with X-ray diffraction, IR and Raman spectrometry. Varying the sintering temperature and/or adding graphite and PMMA as porogenous agents lead to obtained micro- and meso-porosities. The biological tests concerned cell viability, proliferation and morphology (SEM), and the cytochemical staining of actin and vinculin. The micro- and meso-porous HA samples had an internal pore size of 1-10 microm and 10-50 microm, respectively. X-ray diffraction and FTIR confirmed the high purity of the HA. The cell viability tests with L132 cells confirmed the excellent cytocompatibility of HA, the graphite powder and the ATB vancomycine. Proliferation rate was assessed with MC3T3-E1 osteoblasts. All HA samples produced a higher proliferation than the controls; the micro-porous HA inducing the highest cell growth. The ATB impregnated HA also stimulated cell proliferation but in lower extend. Cytochemical staining of osteoblasts revealed a well-developed cytoskeleton with strong stress fibres. Labelling of the focal adhesion contacts with anti-vinculin showed a less developed adhesion process in the cells on the different HA substrates. It was possible to realize a highly pure hydroxyapatite with different but controlled porosities by varying the sintering temperature and/or addition of a porogenous agents. This purity and the micro-porosity stimulate significantly cell growth.     

新生儿耐甲氧西林凝固酶阴性葡萄球菌败血症40例分析

目的：探讨耐甲氧西林凝固酶阴性葡萄球菌（MRCNS）在新生儿败血症中的分布,临床特征及耐药性特点。方法：应用全自动血培养系统（BACTE/9050）,ATB细菌鉴定仪,从80例新生儿血培养阳性瓶中分离出40株MRCNS。应用ATB药敏系统,测定了青霉素等17种抗生素的耐药性。结果：80株细菌中,分离出40株MRCNS（占50%）,其中溶血葡萄球菌17株（42.5%)居首位,表皮葡萄球菌15株（37.5%),其他葡萄球菌8株（20%）。药敏结果显示多重耐药。结论：凝固酶阴性葡萄球菌（CNS）已成为小儿血培养的第一位检出菌,主要为溶血葡萄球菌和表皮葡萄球菌,MRCNS占主导地位（达50%左右）,成为新生儿病房院内感染的主要致病菌。药敏结果显示MRCNS呈多重耐药,万古霉素及其他糖肽类抗生素是治疗MRCNS感染的首选药物,MRCNS在新生儿病房造成院内感染的危害性以及耐药性应引起广泛关注。     

2008-2010年舟山某院金黄色葡萄球菌的分布及耐药性变迁

目的 分析舟山医院三年来金黄色葡萄球菌分布及耐药性变迁,并对耐甲氧西林金黄色葡萄球菌(MRSA)与甲氧西林敏感金黄色葡萄球菌(MSSA)的耐药性差异做对比.方法 用ATB Expression半自动微生物分析仪进行菌株鉴定及药敏试验,用K-B法测红霉素、克林霉素、头孢西丁、苯唑西林直径,比较耐甲氧西林金黄色葡萄球菌(MRSA)与甲氧西林敏感金黄色葡萄球菌(MSSA)的耐药性差异.结果 金黄色葡萄球菌对苯唑西林、庆大霉素、红霉素、四环素和克林霉素的耐药率有上升的趋势;MRSA对苯唑西林、庆大霉素、复方新诺明、克林霉素、红霉素、青霉素、喹奴普汀-达福普汀、利福平和四环素的耐药率都明显高于MSSA的耐药率,二者间差异有统计学意义(P＜0.01),D-试验阳性71株,占72.45％.结论 金黄色葡萄球菌的耐药性逐渐升高,特别是对MRSA应引起临床的重视,检测克林霉素诱导型耐药具有重要的临床应用价值.     

7岁以下儿童细菌感染耐药分析

目的观察大连地区5家医院（大连医科大学附属一院、大连医科大学附属二院、大连儿童医院、大连市第六人民医院、大连开发区医院）2012年7岁以下儿童感染病原菌分布及其耐药情况。方法临床分离菌株,采用细菌分离鉴定方法（API、VITEK、Microsean系统）进行目标细菌的鉴定,药物敏感性试验用K—B纸片扩散法测定药物的敏感性。结果共收集细菌1235株,其中革兰阴性菌725株占58．7％,革兰阳性菌510株占41．3％;分离细菌前5位依次为大肠埃希菌占14．9％、凝固酶阴性葡萄球菌占13．6％、金黄色葡萄球菌占11．7％、肺炎克雷伯菌占10．8％、铜绿假单胞菌占7．4％。结论7岁以下儿童感染致病菌对抗生素均存在不同程度耐药情况,期望在临床治疗感染时有所帮助。     

Influence of testing conditions on the susceptibility results of Staphylococcus cohnii to beta-lactams

The high occurence of coagulase-negative staphylococci among bacteria responsible for hospital infections is unquestioned. Studies on the poorly-known novobiocin-resistant, coagulase-negative Staphylococcus cohnii were undertaken. The possibilities of optimizing conditions for determination of susceptibility to beta-lactam antibiotics of this species were researched. In the case of S. cohnii the new cefoxitin test for detection of methicillin resistant strains, introduced by the National Reference Centre for Antibiotics in Poland was found as a good and of credible quality. It was also shown, that application in in vitro examination conditions stimulating the mechanisms of resistance to beta-lactam antibiotics, supplies credible results relating to their true susceptibility. The necessity of establishing individual conditions for susceptibility determination in different species of coagulase-negative staphylococci was suggested.     

Microbiologic analysis of results from blood cultures

The aim of our investigations was the microbiological analysis together with the evaluation of sensitivity of bacteria frequently isolated from blood cultures. Blood samples were taken from patients with symptoms suggesting bacteremia in Rydygier's Hospital in Cracow. A total of 11,170 blood samples taken from 1997 to 2000 were tested. Automatic VITAL system (bioMerieux) was applied to culture and detect microorganisms. Bacteria were identified by ATB system (bioMerieux). Susceptibility was detected by ATB and disc diffusion method. Percentage of positive results relating to detection of microorganisms of clinical significance was 16.9% (1891 cultures). Staphylococcus spp. (Staph. epidermidis in range 22.8% to 21.9%), Enterococcus spp. and Streptococcus spp. were most frequently isolated species among aerobic Gram-positive bacteria. In 2000, compared to 1997 the number of isolates of MRSA increased considerably (from 1.8% to 6.8%). In blood infections the increase of frequency of E. coli bacteria was also noted: 6.1% and 11.4% in 1997 and 2000, respectively. Among non-fermentant bacilli the percentage of occurrence of P. aeruginosa in the period of 4 years was comparable in the range 7.3% in 1997 to 7.2% in 2000. The increase in the frequency of blood infections of A. baumanii was also noticed (respectively from 4.8% to 9.9%). Susceptibility of P. aeruginosa strains to selected beta-lactame antibiotics and aminoglycosydes increased in 2000 in comparison to 1999. A. baumanii strains were 100% sensitive only to imipenem.     

The Pharmaco –, Population and Evolutionary Dynamics of Multi-drug Therapy: Experiments with S. aureus and E. coli and Computer Simulations

There are both pharmacodynamic and evolutionary reasons to use multiple rather than single antibiotics to treat bacterial infections; in combination antibiotics can be more effective in killing target bacteria as well as in preventing the emergence of resistance. Nevertheless, with few exceptions like tuberculosis, combination therapy is rarely used for bacterial infections. One reason for this is a relative dearth of the pharmaco-, population- and evolutionary dynamic information needed for the rational design of multi-drug treatment protocols. Here, we use in vitro pharmacodynamic experiments, mathematical models and computer simulations to explore the relative efficacies of different two-drug regimens in clearing bacterial infections and the conditions under which multi-drug therapy will prevent the ascent of resistance. We estimate the parameters and explore the fit of Hill functions to compare the pharmacodynamics of antibiotics of four different classes individually and in pairs during cidal experiments with pathogenic strains of Staphylococcus aureus and Escherichia coli. We also consider the relative efficacy of these antibiotics and antibiotic pairs in reducing the level of phenotypically resistant but genetically susceptible, persister, subpopulations. Our results provide compelling support for the proposition that the nature and form of the interactions between drugs of different classes, synergy, antagonism, suppression and additivity, has to be determined empirically and cannot be inferred from what is known about the pharmacodynamics or mode of action of these drugs individually. Monte Carlo simulations of within-host treatment incorporating these pharmacodynamic results and clinically relevant refuge subpopulations of bacteria indicate that: (i) the form of drug-drug interactions can profoundly affect the rate at which infections are cleared, (ii) two-drug therapy can prevent treatment failure even when bacteria resistant to single drugs are present at the onset of therapy, and (iii) this evolutionary virtue of two-drug therapy is manifest even when the antibiotics suppress each other''s activity.     

Synergistic effect of ayurvedic pearl preparation on enhancing effectiveness of antibiotics

Studies were carried out with ayurvedic preparations derived from pearl, which include preparations bhasma and pishti. The synergistic effect to reduce the dose of antibiotic was tested against E. coli the test bacterium with ampicillin antibiotic by bore well and disks diffusion methods. It was observed that pearl preparations do not show any antibacterial activity but when used at 200 microg/ml concentration with antibiotics, then even at sub-lethal dose, the antibiotic has effectively shown the results with reduced contact time. The protocol was also tested with the other bacteria like, Pseudomonas aeruginosa. Vibrio cholarae, Salmonella typhi, and Staphylococcus aureus and has shown similar results. The pearl bhasma synergistic effect was also tested with other antibiotics such as erythromycin, kanamycin, and ampicillin.     

尿培养中病原菌分布及耐药性分析

目的研究大连市中心医院2007年1月至2012年9月尿培养中常见病原菌（真菌除外）的分布及其耐药性,为临床治疗尿路感染提供诊断及用药依据。方法采用MicroScan—Walkaway40全自动微生物鉴定药敏分析仪和ATBExpression自动细菌鉴定仪及K—B纸片法药敏试验,应用WHONET5．4软件对结果进行回顾性统计分析。结果五年间尿培养中共分离出病原菌1012株（真菌除外）,其中革兰阴性菌802株（79．2％）,主要是大肠埃希菌538株（53．2％）、肺炎克雷伯菌95株（9．5％）、铜绿假单胞菌51株（5．1％）、奇异变形杆菌30株（2．9％）、鲍曼不动杆菌26株（2．6％）。革兰阳性菌210株（20．8％）,主要是粪肠球菌86株（8．6％）,屎肠球菌83株（8．2％）,凝固酶阴性葡萄球菌21株（2．1％）,金黄色葡萄球菌13株（1．3％）。抗生素敏感试验中碳青霉烯类（亚胺培南及美洛培南）对肠杆菌科细菌大肠埃希菌、肺炎克雷伯菌及奇异变形杆菌抗菌活性最强,无耐药菌株,对非发酵菌中铜绿假单胞菌和鲍曼不动杆菌耐药率为30％～40％,革兰阳性菌对糖肽类（万古霉素、替考拉宁）和利奈唑胺等抗菌药物高度敏感。结论应当根据主要易感菌种类及其抗生素敏感试验结果指导临床合理使用抗生素。     

74例金黄色葡萄球菌乳腺炎的耐药性分析

目的分析哺乳期乳房脓肿患者的金黄色葡萄球菌的耐药情况,指导临床合理使用抗菌药物。方法将厦门市妇幼保健院乳腺门诊及病房2009年2月至2011年1月抽取的128例哺乳期乳房脓肿患者的乳腺脓液接种于哥伦比亚血平板,置35℃培养箱,过夜孵育18~24 h。革兰阳性球菌,触酶阳性,血浆凝固酶试验(+),必要时采用ATB-Expression细菌鉴定系统确认。用K-B纸片法检测其对苯唑西林等10种抗菌药物的敏感度。结果 128例检出细菌86例,阳性率67.2%,其中检出74株金黄色葡萄球菌(86.0%),其对青霉素、红霉素和克林霉素有较强的耐药性,耐药率分别为94.6%、64.9%和59.5%;耐苯唑西林19株(25.7%),对复方新诺明、磷霉素和庆大霉素较为敏感,对万古霉素、替考拉宁和左旋氧氟沙星100%敏感。结论哺乳期乳房脓肿致病菌主要为金黄色葡萄球菌,耐甲氧西林金黄色葡萄球菌(MRSA)占25.7%,其耐药情况不容忽视,青霉素类、大环内酯类等耐药率较高不能作为经验用药。建议临床使用抗菌药物前及时送检乳腺脓液细菌培养,以药敏结果指导临床合理经济用药。     

Tolerance towards gentamicin is a function of nutrient concentration in biofilms of patient-isolated <Emphasis Type="Italic">Staphylococcus epidermidis</Emphasis>

Staphylococcus epidermidis is a biofilm-forming bacterial strain that can cause major problems as an agent of nosocomial infections. Bacteria in biofilms are shielded from the environment and can survive high doses of antibiotics. We here test the antibiotic susceptibility of Staphylococcus epidermidis to rising gentamicin concentrations in optimal growth conditions as used in routine bacteriology laboratories with low nutrient situations as suggested to be found in clinical situations. We found that gentamicin-resistant Staphylococcus epidermidis biofilms survived in the absence of external nutrient supply in PBS. While addition of gentamicin sulfate significantly reduced the pH value of all used media and solutions, this acidification did not alter survival of bacteria in the biofilm. We found a statistically significant and dose-dependent reduction of survival in low nutrient situations using gentamicin sulfate in three out of four patient isolates of Staphylococcus epidermidis which have been tested to be gentamicin-resistant under optimal growth conditions. Supporting the original profiling, survival in full media under the same antibiotic dosages was not significantly reduced. Our data here show that antibiotic resistance is a function of the provided nutrient concentration. Antibiotic resistance profiling should consider variations in nutrient availability.     

Participation of gram-negative anaerobes in infections in hospitalized patients

The aim of the study was to assess the prevalence and antibiotic susceptibility profiles of strictly anaerobic Gram-negative bacteria isolated from clinical samples taken from hospitalized patients from 01.01.2007 to 31.12.2008. The specimens were cultured using media, incubated at 37 degrees C under anaerobic conditions. Biochemical identification and antibiotic susceptibility were done in an automated system ATB Expression (bioMerieux S.A, France). For selected strains of Bacteroides sp. sensitivity was determined using E-test (AB BIODISK, Sweden). Overall 1274 strains of obligate anaerobes were isolated. Gram-negative bacteria were cultured in number of 333 strains. Most frequently isolated was Bacteroides sp. (46,9%) and Prevotella sp. (29,7%). Isolated bacteria are still susceptible to imipenem (100%), metronidazole (100%) and beta-lactam antibiotics with beta-lactamase inhibitors: amoxicillin/clavulanate (97,8%) piperacillin/tazobactam (99,1%), ticarcillin/clavulanate (99,1%).     

不同气候条件下慢性阻塞性肺疾病的病原学和免疫学研究

目的：探讨COPD患者不同气候条件下（雷暴、阴雨、台风）的病原学和免疫功能。方法：细菌鉴定采用VITEK全自动微生物鉴定系统。K．B纸片扩散法测定药敏。T淋巴细胞亚群测定采用单克隆抗体免疫组化荧光染色法。血清免疫球蛋白（IgG,IgA,IgM）用琼脂单向扩散法。结果：台风、雷暴、阴雨、天气气候条件60例患者痰培养阳性率分别为45％、40％、30％,占革兰阴性菌前2位分别为肺炎克雷伯杆菌、流感嗜血杆菌和肺炎克雷伯杆菌、大肠埃希杆菌及流感嗜血杆菌、铜绿假单胞菌。革兰阳性菌中分别以肺炎链球菌、金黄色葡萄球菌为主及肺炎链球菌、金黄色葡萄球菌为主和金黄色葡萄球菌、表皮葡萄球菌为主。铜绿假单胞菌、肺炎克雷伯杆菌、流感嗜血杆菌对常用抗生素具有较高的耐药性。台风、雷暴、阴雨气候条件下3组AECOPD患者CD3＋ CD4＋、CD4＋／CD8＋、IgG、IgA、IgM均明显低于健康对照组（P〈0．05）,CD8＋高于健康对照组（P〈0．05）,而发作期和缓解期无显著差异（P〉0．05）。三种气候条件下相互之间比较的细胞和体液免疫指标比较无显著差剐（P〉0．05）。结论：雷暴、阴雨、台风条件下AECOPD的患者存在病原学分布差异,对常用抗生素具有较高的耐药性,细胞和体液免疫功能进一步下降。但三者相互之间的免疫功能比较无明显差异。     

抗生素诱导革兰阴性菌外膜囊泡产生及发挥生理作用的机制

外膜囊泡是革兰阴性菌分泌的一种球形纳米颗粒,由外膜及其所含成分组成,是细菌在外界压力条件下分泌的具有生理活性的特殊结构。外界压力如抗生素、缺氧等可触发细菌释放外膜囊泡,甚至在正常生长周期中,一些革兰阴性菌也会释放囊泡。外膜囊泡与细菌的多种生理过程相关,如应激反应、毒素传递、致病、细胞间通讯、免疫调节、基因水平转移及维持微生物群稳态等。在使用抗生素治疗过程中,尤其是当人体微生物群处于低剂量抗生素环境时,细菌会大量分泌外膜囊泡。在肠道中,外膜囊泡释放后会通过多种机制刺激肠道而引发多种炎症。本文综述了外膜囊泡的产生、结构及生理作用,提出抗生素治疗不但会破坏人体正常菌群而导致菌群失调,还会诱导细菌产生大量外膜囊泡而引发慢性炎症。噬菌体治疗不破坏正常菌群,特异性杀灭细菌时也不引起外膜囊泡的产生,因此开发使用噬菌体靶向治疗细菌感染将大大减少不良反应。     

临床常见病原性细菌对抗生素的耐药性分析

目的了解长春地区部分医院临床分离的致病菌对抗菌药物的耐药情况。方法收集吉林大学中日联谊医院和长春中医药大学附属医院临床分离的致病菌,采用K—B法进行抗生素敏感性试验,参照CLSI2009年版的标准判断结果。结果从这两家医院共分离细菌447株,其中革兰阴性菌353株,占78．9％,革兰阳性菌94株,占21．1％。肠杆菌科细菌对氨苄西林的耐药率在70％以上,对庆大霉素的耐药率超过了50％,但对亚胺培南保持较高的敏感性（耐药率〈5％）。金黄色葡萄球菌和表皮葡萄球菌对红霉素耐药率分别为65．2％和82．1％,对万古霉素的耐药率较低,分别为4．5％和3．6％。非发酵革兰阴性杆菌对头孢他啶的耐药率超过50％,铜绿假单胞菌对氨曲南的耐药率达到了62．9％,对他唑巴坦复合制剂、亚胺培南、美罗培南和阿米卡星较为敏感,而鲍曼不动杆菌对各类抗生素的耐药率都较高。结论细菌对抗生素的耐药性仍表现出增长趋势,加强细菌耐药性的检测,对于指导临床正确、合理使用抗生素,减少耐药菌的产生和传播具有重要意义。     

结核病房医院感染与抗生素使用相关性分析

目的：探讨临床抗菌药物应用与结核病房医院感染严重程度的相关性。方法：分析3年来我院结核病房医院感染209例共计221株病原菌,细菌鉴定采用法国梅里埃ATB细菌鉴定药敏分析仪,药敏试验联合应用微量测定法与K—B纸片法。结果：在221株病原菌中革兰阴性杆菌最多为112株,占50．7％,念珠菌75株,占33．9％。主要致病菌依次为白色念珠菌61株(27．6％),铜绿假单胞菌37株(16．8％),肺炎克雷伯菌25株(10．9％)。药敏试验表明细菌耐药明显增加,大多数革兰阴性杆菌对亚胺培南、阿米卡垦、头孢他啶较敏感,金黄色葡萄球菌对万古霉素100％敏感。每例结核病患者除抗结核药物外平均使用2．67种抗菌药物。结论：结核病房医院感染主要来源于内源性感染,其严重性与抗菌药物大量、重复使用密切相关。     

微生物鉴定系统在肠道病原菌检测中的应用

目的建立简便、快速、系统的病原菌生化检测方法。方法通过应用ATB微生物鉴定系统对112株可疑肠道病原菌菌株进行检测分析,并与国家标准法结合鉴定。结果ATB微生物鉴定系统在112株可疑肠道病原菌菌株中检测出肠道病原菌66株,鉴定结果与国家标准法结果一致,而对常规检测不能确定的37菌株也能给出明确的鉴定结果。结论该系统操作简便、快速,缩短了鉴定周期,提高了鉴定效率,检测范围广,可以作为肠道致病菌检测常规方法的辅助工具,有利于病原菌的快速检出。     

Alterations in rat gut bacteria and intestinal epithelial cells following experimental exposure of antimicrobials

The intestinal bacteria are known to play a significant role in intestinal homoeostasis and the mucosal immune system. In vitro interactions of Ampicillin (0.5-2.0 microg mL(-1)), Amphotericin-B (25-200 microg mL(-1)) and Ciprofloxacin (50-500 ng mL(-1)) with Escherichia coli, Pseudomonas sp. (Gram-negative), Lactobacillus sp., Staphylococcus sp. (Gram-positive), total mixed population of gut bacteria and intestinal epithelial cells were studied. In vitro exposure of Ciprofloxacin showed significant dose-dependent inhibition throughout the growth phase in bacteria. Similar patterns of concentration-dependent changes in membrane transport enzymes and structural constituents, dehydrogenase activity associated with respiratory and energy-producing processes and esterase activity test linked to the general heterotrophic activity of the cell were observed in both bacteria and epithelial cells. The antibiotic effects were in the order of Amphotericin-B相似文献   

血培养中病原菌的分布及其耐药状况分析

目的:了解血培养中病原菌的菌群分布及其耐药状况.方法:血培养标本用Bact/Alert-120自动血液分析系统和Vitek60鉴定仪进行血培养及鉴定,药敏用K-B法,用Whonet 5软件进行分析.结果:在3 680份血培养标本中分离出细菌348株,阳性检出率为9.4%,病原菌中以革兰阴性杆菌为主共分离出189株,占54.3%,其中主要为大肠埃希菌占15.5%,肺炎克雷伯菌占12.4%;革兰阳性菌138株,占39.7%,主要为凝固酶阴性葡萄球菌占14.9%,金黄色葡萄球菌占12.1%;链球菌占6.6%.血培养中的革兰阴性杆菌对亚胺培南、阿米卡星、头孢哌酮/舒巴坦治疗较为敏感:革兰阳性球菌对万古霉素和替考拉宁较为敏感.结论:肠杆菌科细菌和葡萄球菌是血培养中的主要病原菌,而产ESBLs菌株、MRSA、MRCNS耐药严重,提示应高度重视合理使用抗生素,减少细菌耐药性产生,以提高临床治疗效果.     

BioTimer Assay, a new method for counting Staphylococcus spp. in biofilm without sample manipulation applied to evaluate antibiotic susceptibility of biofilm

The medical device-related infections are frequently a consequence of Staphylococcus biofilm, a lifestyle enhancing bacterial resistance to antibiotics. Antibiotic susceptibility tests are usually performed on planktonic forms of clinical isolates. Some methods have been developed to perform antibiotic susceptibility tests on biofilm. However, none of them counts bacterial inoculum. As antibiotic susceptibility is related to bacterial inoculum, the test results could be mistaken. Here, a new method, BioTimer Assay (BTA), able to count bacteria in biofilm without any manipulation of samples, is presented. Moreover, the BTA method is applied to analyze antibiotic susceptibility of six Staphylococcus strains in biofilm and to determine the number of viable bacteria in the presence of sub-inhibitory doses of four different antibiotics. To validate BTA, the new method was compared to reference methods both for counting and antibiotic susceptibility tests. A high agreement between BTA and reference methods is found on planktonic forms. Therefore, BTA was employed to count bacteria in biofilm and to analyze biofilm antibiotic susceptibility. Results confirm the high resistance to antibiotics of Staphylococcus biofilm. Moreover, BTA counts the number of viable bacteria in the presence of sub-inhibitory doses of antibiotics. The results show that the number of viable bacteria depends on sub-inhibitory doses, age of biofilm and type of antibiotic. In particular, differently to gentamicin and ampicillin, sub-inhibitory doses of ofloxacin and azithromycin reduce the number of viable bacteria at lower extent in young than in old biofilm. In conclusion, BTA is a reliable, rapid, easy-to-perform, and versatile method, and it can be considered a useful tool to analyze antibiotic susceptibility of Staphylococcus spp. in biofilm.