肺炎支原体突变敏感性分子开关检测对抗菌药物敏感性检测的作用研究

目的分析肺炎支原体突变敏感性分子开关检测对抗菌药物敏感性检测的价值。方法选择2015年1月至2016年2月在河北省儿童医院呼吸科住院收治的450例社区获得性肺炎患儿的肺泡灌洗液标本,进行MP培养与药敏分析,抗菌药物选择红霉素与阿奇霉素,对MP培养阳性菌株使用分子开关技术检测23SrRNA V区2063/2064基因的突变情况,并进行测序分析,最后明确分子开关检测MP耐药基因与其对抗菌药物敏感性的相关性。结果在450例患儿中,MP检测阳性80例,阳性率为17.8%。在80株MP中,72株对红霉素耐药,耐药率为90.0%;31株对阿奇霉素耐药,耐药率为38.8%,为此MP对于红霉素的耐药率明显高于阿奇霉素(χ2=9.134,P=0.000)。76株MP株的23SrRNA的2063/2064位发生了基因突变,其中有70株检测出A2063G的突变,6株有A2064G的突变。而在76株23S rRNA 2063/2064基因位点突变的MP中,有72株MP对红霉素产生高水平耐药,并对阿奇霉素发生交叉耐药,未发生突变的4株MP均对红霉素与阿奇霉素敏感。结论肺炎支原体对红霉素与阿奇霉素都有高度的耐药性,分子开关检测可识别23SrRNA V区基因2063/2064位点突变,有利于反映MP对红霉素与阿奇霉素的敏感性,具有很好的检测价值。     

菌丝霉素MP1106融合蛋白的复性及纯化方法

旨在建立高效、快捷的菌丝霉素衍生物MP1106大肠杆菌表达系统。通过基因融合的方式构建生物表面活性剂-菌丝霉素衍生物(DAMP_4-MP1106)融合蛋白表达载体,在大肠杆菌中进行表达;并对目的蛋白MP1106进行分离纯化和分子内二硫键鉴定。结果显示,融合蛋白DAMP_4-MP1106在大肠杆菌中以包涵体的形式成功表达,表达产物在变性条件下经Ni~(2+)-NTA亲和层析纯化;经检测分析,摇瓶中DAMP_4-MP1106的发酵产量为118 mg/L,纯度为94.7%;采用96孔板筛选并建立复性方法,获得水溶性融合蛋白DAMP_4-MP1106;并经TEV蛋白酶酶切以及二次Ni~(2+)-NTA亲和层析纯化,可获得纯度为99%的抗菌肽MP1106 18mg/L,回收率达到了38.4%。通过简捷方法快速鉴定分子内的二硫键,初步证实了抗菌肽MP1106完成了分子内结构正确折叠。建立了高效快捷的菌丝霉素大肠杆菌表达系统。     

女性生殖道支原体感染与不孕症的关系及药敏分析

目的探讨女性生殖道支原体感染与不孕症之间的关系及对11种抗生素的敏感率,指导临床明确诊断,合理用药。方法随机选择2007年至2009年来长治市妇幼保健院就诊的女性不孕症患者360例,进行宫颈分泌物支原体培养和药敏试验。结果 360例宫颈分泌物标本中检出解脲支原体(Uu)阳性者161例、人型支原体(Mh)阳性者3例、解脲支原体(Uu)+人型支原体(Mh)混合阳性者29例;对阳性标本都做了11种抗生素的药敏试验,其中交沙霉素敏感率为93.47%、美满霉素敏感率为92.00%、强力霉素敏感率为90.21%、克拉霉素敏感率为83.69%、甲砜霉素敏感率为73.91%、环脂红霉素敏感率为61.95%和阿奇霉素敏感率为53.26%等。结论女性不孕症伴随高发的生殖道支原体感染,支原体感染与不孕症可能有关;长治地区生殖道支原体对交沙霉素、美满霉素、强力霉素、克拉霉素、甲砜霉素敏感性较高,环脂红霉素、阿奇霉素次之,对红霉素、罗红霉素敏感性较低,对环丙沙星、左氧氟沙星敏感性很低。     

湛江地区小儿肺炎支原体感染调查分析

目的:探讨湛江地区小儿肺炎支原体(MP)感染情况.方法:采用日本富士瑞必欧株式会社肺炎支原体抗体检测试剂(SERODI-A-MYCOII),对2005年1月至2008年12月在本院就诊的肺炎患儿进行血清MP抗体检测,对不同年度、不同季节、不同年龄及性别MP肺炎的发病情况进行统计.结果:受检人数2825例,MP抗体阳性率为40.2%.阳性率的多少与不同的年龄段、不同性别有明显区别.0～1岁婴儿期MP感染率为9.5%;1～3岁组幼儿MP感染率为40.4%;4～6岁学龄前期MP感染发病率为45.4%;7～14学龄期MP感染率为48.3%.0～1岁组MP阳性率明显低于其他年龄组,差异有统计学意义(x2=110.5523,P＜0.01).男、女性肺炎患儿阳性率分别为36%、49.4%,两者比较差异有统计学意义(x2=44.9891,P＜0.01).一年四季均可发病.结论:MP肺炎的发病与年龄、性别、季节和年度有密切关系.     

95例肺炎支原体快速液体培养阳性标本分析

目的:采用巢式PCR法(nPCR)及全自动微生物鉴定仪对肺炎支原体(MP)快速液体培养阳性标本进行分析,探讨MP快速培养假阳性情况及原因.方法:收集95例MP快速液体培养阳性标本,巢式PCR检测MP核酸,用全自动微生物鉴定仪检测导致培养假阳性的微生物.结果:95例MP快速液体培养阳性标本中,90例巢式PCR结果阴性,假阳性率94.7%;经全自动细菌鉴定仪鉴定,97.8%为真菌所致.结论:普通的敏感细菌在MP快速液体鉴定培养基中可以被有效抑制;真菌是引起的快速培养假阳性的主要原因.     

女性泌尿生殖道支原体感染患者药敏状况

目的探讨女性泌尿生殖道感染患者中支原体的感染状况及其对不同药物的敏感性,为临床合理用药提供参考。方法对516例女性泌尿生殖道拭子进行支原体培养,并对阳性菌株进行10种常用抗生素药敏试验。结果 516例患者中,支原体培养阳性263例,检出率为50.97%,其中解脲支原体(Ureaplasma urealyticum,UU)为237例(45.93%),人型支原体(Mycoplasma hominis,MH)仅9例(1.74%),UU+MH 17例(6.46%);三种类型感染均对交沙霉素、强力霉素、美满霉素比较敏感。结论 UU为沈阳医学院附属第二医院泌尿生殖道支原体感染的主要病原体,交沙霉素、强力霉素及美满霉素可作为治疗的经验用药,其中交沙霉素为首选抗生素。     

2709例泌尿生殖道支原体培养及耐药性分析

目的对门诊2 709例病例的支原体培养标本进行泌尿生殖道解脲脲原体(Ureaplasma urealyticum,Uu)与人型支原体(Mycoplasma hominis,Mh)感染情况及耐药性的分析,为临床的诊断及治疗提供参考依据。方法采用商品化试剂盒对2 709例标本进行Uu、Mh半定量培养及药物敏感试验。结果 2 709例标本中总阳性率为56.15%,其中Uu占比39.87%、Mh占比8.08%、Uu和Mh共同阳性者占比8.19%,男性标本阳性率为47.78%、女性标本阳性率为66.26%。药敏结果显示,单独Uu感染者对喹诺酮类(5.00%~34.40%)、红霉素(42.80%)敏感率较低,而对四环素类(88.90%~95.00%)、交沙霉素(97.30%)敏感率较高。单独Mh感染者与Uu和Mh混合感染者耐药情况均较为严重,除四环素、强力霉素、交沙霉素、美满霉素敏感率在50%以上,对其余药物的敏感率均在50%以下。结论泌尿生殖道支原体感染以Uu感染为主,其中女性感染率高于男性;药敏结果显示感染者对四环素、强力霉素、交沙霉素、美满霉素敏感率较高,单独Mh感染者与Uu和Mh混合感染者耐药情况更为严重。     

523例宫颈上皮内瘤变患者支原体属感染情况及药敏分析

目的研究宫颈上皮内瘤变(CIN)患者支原体感染情况及耐药分析,为临床诊治提供依据。方法采用回顾性方法对523例宫颈上皮内瘤变(CIN)患者的分泌物进行解脲支原体(Uu)和人型支原体(Mh)培养及药敏试验。结果 523例宫颈上皮内瘤变患者Uu的检出率为55.25%(289例),Uu和Mh混合感染的检出率为13.58%(71例),检出5例单一Mh感染的患者。对抗菌药物敏感率高的是强力霉素(91.16%)、美满霉素(90.48%),其次是交沙霉素(81.29%)、克拉霉素(75.85%),对氧氟沙星的敏感率最低,仅为10.20%。结论宫颈上皮内瘤变患者中支原体感染存在普遍,应合理使用抗菌药物治疗,支原体感染与宫颈上皮内瘤变的发生可能密切相关。     

3468例支原体培养及耐药监测结果分析

目的调查分析大连市妇幼保健院支原体感染及耐药情况,为临床治疗提供依据。方法采用支原体培养/药敏试剂盒对3 468例来我院门诊就诊的患者,做支原体培养及药物敏感试验,统计支原体阳性率,并对13种抗菌药分析其耐药性。结果支原体阳性率为57.5%,其中Uu感染阳性率为67.7%,Uu+Mh混合感染阳性率为29.4%,Mh感染阳性率为2.9%。女性感染率为59.6%,男性感染率为31.4%。药敏结果显示对Uu较敏感的抗菌药物为强力霉素(92.9%)、美满霉素(89.8%)、克拉霉素(86.0%);对Mh较敏感的抗菌药物为强力霉素(86.2%)、美满霉素(84.5%);对Uu+Mh混合感染者较敏感的抗菌药物为强力霉素(54.8%)、美满霉素(55.1%)。结论大连市妇幼保健院支原体感染以Uu为主,但Uu+Mh混合感染也不容忽视;不同生物型支原体感染对抗菌药耐药性不同,特别是Uu+Mh混合感染耐药情况比较严重,临床医师应根据药敏试验正规合理选用抗生素,以有效的治疗感染。     

沙井地区384例支原体药敏试验结果及分析

目的:了解本地区社区感染支原体的药物敏感情况。方法:对门诊标本培养出的384例支原体进行药敏试验(微量肉汤法)。结果:Uu对交沙霉素、克拉霉素及罗红霉素敏感性高,敏感率分别为98%、94%及93%;对环丙沙星、大观霉素耐药性高,敏感率分别为8%及11%。Mh对交沙霉素敏感率为89%、对大现霉素、多西环素、美满霉素的敏感率均为78%;对罗红霉素、红霉素耐药性高,敏感率分别为0%及11%。Uu合并Mh对交沙霉素、多西环素、美满霉素敏感性高,敏感率分别为85%、73%及69%,对红霉素、克拉霉素、大观霉素的敏感性均为0%,对罗红霉素、阿奇霉素、环丙沙星的敏感率均为4%。结论:支原体的敏感性存在时空差异,且不同类型的支原体对抗菌素的敏感性是不同的,应采用实验室结果而非经验指导用药。     

Latex agglutination test for diagnosing pneumococcal pneumonia in children in developing countries.

OBJECTIVE--To prepare and assess the sensitivity and specificity of a latex agglutination test specific for the serotype of antigen in diagnosing pneumococcal pneumonia in Gambian children. DESIGN--Comparison of agglutination test specific for serotype with culture of blood and lung aspirates, countercurrent immunoelectrophoresis, and commercial latex agglutination tests in diagnosing pneumococcal pneumonia. Cross reaction studies and investigation of 102 control children to determine specificity of agglutination test specific for serotype. SETTING--General medical ward of Medical Research Council laboratories, The Gambia. PATIENTS--101 Gambian children aged between 2 months and 10 years admitted with severe pneumonia. INTERVENTIONS--Serum samples were boiled and treated with edetic acid, and urine samples were boiled and concentrated 25 times before testing. END POINT--A latex agglutination test specific for the serotype of pneumococcal antigen that is sensitive and highly specific for detecting pneumococcus in the urine of patients with pneumococcal pneumonia. MEASUREMENTS AND MAIN RESULTS--Concentrated urine samples from 16 of the 21 children (76%) with pneumococcal pneumonia established by results of culture of blood or lung aspirates gave a positive result with the agglutination test specific for serotype, whereas only four of the 102 urine samples obtained from control children without pneumonia gave positive results. The serotypes of antigens detected in the urine of children with pneumococcal pneumonia and the serotypes of pneumococci isolated from cultures of blood or lung aspirates were the same in all cases. CONCLUSIONS--When performed on urine samples the agglutination test specific for serotype has a high specificity and is more sensitive than culture of blood or lung aspirates, commercial agglutination tests, or countercurrent immunoelectrophoresis in identifying pneumococcal pneumonia. It is easy to use and should be especially useful in communities with limited laboratory facilities.     

131例住院肺炎患儿细菌感染现况及耐药谱分析

目的：分析住院肺炎惠儿的病原菌分布及其耐药性。方法：选择2009年6月至2010年5月,新疆维吾尔自治区人民医院儿科住院的791例肺炎患儿,采取下呼吸道痰液标本,进行细菌培养及药敏试验。结果：16．56％（131／791）患儿被确诊为细菌性肺炎并且有明确的病原,其中,革兰阴性菌感染为75．57％（99／131）,且以肺炎克雷伯氏菌、大肠埃希菌、阴沟肠杆菌多见;革兰阳性菌感染为21．37％（28／131）,以肺炎链球菌多见;真菌感染为6．87％（9／131）,均为白色假丝酵母;9．92％（13／131）的患儿存在两种及以两种上病原菌感染。鲍曼不动杆菌和肠杆菌对头孢类抗生素耐药严重,部分肺炎克雷伯菌、大肠埃希菌产伊内酰胺酶（BLA）,葡萄球菌属耐青霉素G,肠球菌对氯洁霉素、红霉素和复方新诺明耐药率100％,而肺炎链球菌对红霉素耐药率100％。真菌对常用抗菌药的耐药率为0。结论：我院住院肺炎患儿细菌性为16．56％。病原菌构成以革兰阴性菌为主,并且大多数病原菌耐药。临床应根据痰液细菌培养和药敏结果,合理选择抗菌药物,以减少细菌耐药性,防止滥用抗生素。     

血培养中病原菌分布及药敏调查分析

目的了解血培养中病原菌的分布及药敏情况,供临床借鉴。方法对中山大学附属第三医院血培养标本中所分离到的细菌及药敏结果进行统计分析。结果从血培养标本中分离到细菌239株,其中其中革兰阴性杆菌(G-b)131株,占52.6%,革兰阳性球菌(G+c)99株,占39.8%(99/236),真菌占6.8%,大肠埃希菌和肺炎克雷伯菌产ESBLs菌检出率分别是54.2%和48.6%,只对亚胺培南、特治星和阿米卡星敏感,敏感率超过85%,G+c中,耐甲氧西林金黄色葡萄球菌(MRSA)和耐甲氧西林凝固酶阴性葡萄球菌(MRCNS)的检出率分别是42.9%和89.2%,只对万古霉素敏感(100%),检出的肠球菌已出现对万古霉素耐药。结论血培养分离的病原菌分布复杂,产ESBLs菌和MRS菌株检出率高,临床应重视血培养检测结果.合理用药。     

A Prospective Study of the Prevalence of Tuberculosis and Bacteraemia in Bangladeshi Children with Severe Malnutrition and Pneumonia Including an Evaluation of Xpert MTB/RIF Assay

Background

Severe malnutrition is a risk factor for pneumonia due to a wide range of pathogens but aetiological data are limited and the role of Mycobacterium tuberculosis is uncertain.

Methods

We prospectively investigated severely malnourished young children (<5 years) with radiological pneumonia admitted over a 15-month period. Investigations included blood culture, sputa for microscopy and mycobacterial culture. Xpert MTB/RIF assay was introduced during the study. Study children were followed for 12 weeks following their discharge from the hospital.

Results

405 eligible children were enrolled, with a median age of 10 months. Bacterial pathogens were isolated from blood culture in 18 (4.4%) children, of which 72% were Gram negatives. Tuberculosis was confirmed microbiologically in 7% (27/396) of children that provided sputum - 10 by culture, 21 by Xpert MTB/RIF assay, and 4 by both tests. The diagnostic yield from induced sputum was 6% compared to 3.5% from gastric aspirate. Sixty (16%) additional children had tuberculosis diagnosed clinically that was not microbiologically confirmed. Most confirmed tuberculosis cases did not have a positive contact history or positive tuberculin test. The sensitivity and specificity of Xpert MTB/RIF assay compared to culture was 67% (95% CI: 24–94) and 92% (95% CI: 87–95) respectively. Overall case-fatality rate was 17% and half of the deaths occurred in home following discharge from the hospital.

Conclusion and Significance

TB was common in severely malnourished Bangladeshi children with pneumonia. X-pert MTB/RIF assay provided higher case detection rate compared to sputum microscopy and culture. The high mortality among the study children underscores the need for further research aimed at improved case detection and management for better outcomes.     

Detection of Mycoplasma pneumoniae in simulated and true clinical throat swab specimens by nanorod array-surface-enhanced Raman spectroscopy

The prokaryote Mycoplasma pneumoniae is a major cause of respiratory disease in humans, accounting for 20% of all community-acquired pneumonia and the leading cause of pneumonia in older children and young adults. The limitations of existing options for mycoplasma diagnosis highlight a critical need for a new detection platform with high sensitivity, specificity, and expediency. Here we evaluated silver nanorod arrays (NA) as a biosensing platform for detection and differentiation of M. pneumoniae in culture and in spiked and true clinical throat swab samples by surface-enhanced Raman spectroscopy (SERS). Three M. pneumoniae strains were reproducibly differentiated by NA-SERS with 95%-100% specificity and 94-100% sensitivity, and with a lower detection limit exceeding standard PCR. Analysis of throat swab samples spiked with M. pneumoniae yielded detection in a complex, clinically relevant background with >90% accuracy and high sensitivity. In addition, NA-SERS correctly classified with >97% accuracy, ten true clinical throat swab samples previously established by real-time PCR and culture to be positive or negative for M. pneumoniae. Our findings suggest that the unique biochemical specificity of Raman spectroscopy, combined with reproducible spectral enhancement by silver NA, holds great promise as a superior platform for rapid and sensitive detection and identification of M. pneumoniae, with potential for point-of-care application.     

Evaluation of a novel monoclonal-based antigen-in-stool enzyme immunoassay (Premier Platinum HpSA PLUS) for diagnosis of Helicobacter pylori infection in Vietnamese children

Background: Helicobacter pylori infection is difficult to diagnose in children, especially in developing countries where noninvasive methods such as urea breath test are often not available. We evaluate the sensitivity and specificity of a new monoclonal antibody-based antigen-in-stool enzyme immunoassay (Premier Platinum HpSA PLUS) for diagnosis of H. pylori infection in Vietnamese children.
Materials and Methods: Sensitivity of the antigen-in-stool test was evaluated in 232 children, 3–15 years of age, who were positive for H. pylori infection by culture from biopsies. For evaluation of the specificity 98 children of similar age with nongastrointestinal conditions and who were negative for H. pylori infection by serologic assays were included with blood and stool samples.
Results: Of the 232 culture-positive children, 224 were also positive by Premier Platinum HpSA PLUS. Of the 98 control children, 93 were H. pylori negative also in the stool test. The sensitivity of Premier Platinum HpSA PLUS was thus 96.6% (95% CI 93.3–98.5) and the specificity was 94.9% (95% CI 88.5–98.3).
Conclusions: The findings have demonstrated Premium Platinum HpSA PLUS to be a reliable method for detection of H. pylori infection also in children in our area.     

新生儿呼吸机相关性肺炎临床分析与防治对策

目的探讨新生儿呼吸机相关性肺炎的临床特点以及防治措施。方法59例患儿使用呼吸机机械通气进行治疗,分析与呼吸机相关性肺炎发生率情况、病原学检查结果、药敏试验结果以及转归情况。结果与呼吸机相关性肺炎者有30例,发生率为50．85％。从下呼吸道分泌物培养或管端培养出细菌29例。分析了病原菌种类与所占的比例,并给出了药敏试验结果。两组患儿治愈率相比具有统计学差异。结论采取综合的防治措施是降低呼吸机相关性肺炎发生的最佳方法。     

重症监护病房呼吸机相关肺炎的病原菌分布及耐药性分析

目的:研究重症监护病房(ICU)呼吸机相关肺炎的病原菌分布及耐药性情况。方法:采集2016年4月-2017年8月于我院ICU住院治疗的58例呼吸机相关肺炎患者痰样本进行病原菌培养,观察病原菌分布情况。同时对主要病原菌进行药敏试验,分析病原菌对头孢他啶、头孢曲松、环丙沙星、奈替米星、妥布霉素、氨曲南、亚胺培南、阿米卡星八种常见抗生素的耐药性情况。结果:58例呼吸机相关肺炎患者共培养204株病原菌,204株病原菌中根据占比分别为革兰氏阴性菌69.61%(142/204)、真菌15.20%(31/204)以及革兰氏阳性菌15.20%(31/204),且革兰氏阴性菌占比均明显高于真菌以及革兰氏阳性菌(P0.05),其中革兰氏阴性菌中肺炎克雷伯菌占比21.08%(43/204)、铜绿假单胞菌占比18.14%(37/204)、鲍氏不动杆菌占比10.78%(22/204)、产气肠杆菌占比9.31%(19/204)。呼吸机相关肺炎患者病原菌中肺炎克雷伯菌、产气肠杆菌、金黄色葡萄球菌对亚胺培南的耐药性均低于其他七种抗生素,差异有统计学意义(P0.05),铜绿假单胞菌对奈替米星的耐药性均低于其他七种抗生素,差异有统计学意义(P0.05),鲍氏不动杆菌耐药性均较高。结论:ICU中呼吸机相关肺炎主要是由革兰氏阴性菌引发,且耐药情况不容乐观,其中革兰氏阴性菌对亚胺培南最为敏感,值得临床重点关注。     

Urinary detection of Streptococcus pneumoniae antigen for diagnosis of pneumonia

Streptococcus pneumoniae is one of most common causes of community-acquired pneumonia. We evaluated a newly available rapid immunochromatographic test to detect S. pneumoniae in urine samples verifying its importance in the diagnosis of pneumococcal pneumonia. Our data, obtained from 104 patients with community-acquired pneumonia, show that Now S. pneumoniae Urinary Test is characterized by a sensitivity value of 77.7%, a specifity of 98.8%: positive and negative predictive values are 93.3% and 95.5%, respectively. In conclusion, Now S. pneumoniae Urinary Test should be a useful test to establish the etiology of community-acquired pneumonia.     

小儿呼吸道感染常见病原菌及药敏结果分析

目的:了解小儿下呼吸道感染的常见病原菌及其对抗菌药物的敏感性.方法:对北京市和平里医院儿科两年来确诊治疗的156例小儿呼吸道感染患儿的痰标本进行培养及药敏试验.结果:156例标本检出致病菌98株,其中G-杆菌71株(72.4％),G+球菌23株(23.5％),真菌4株(4.1％),主要致病菌依次为肺炎克雷伯菌26株,大肠埃希菌20株.流感嗜血杆菌13株,金黄色葡萄球菌7株,肺炎链球菌5株.药敏结果显示,G-杆菌对亚胺培南均敏感,其次为头孢西丁、庆大霉素,G+球菌对万古霉素均敏感,对头孢西丁、庆大霉素、环丙沙星、阿米卡星有较高的敏感性.结论:革兰阴性杆菌为本地区小儿呼吸道感染的主要致病菌,真菌的感染率也呈上升趋势,临床上应及时检测,合理用药.