MALDI-TOF-MS技术快速鉴定丝状真菌的临床评价

目的探讨基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)技术在丝状真菌快速鉴定中的临床应用价值。方法对西安交通大学第一附属医院2015年3月至2017年1月分离自临床的丝状真菌进行聚合酶链式反应(PCR)测序鉴定与MALDI-TOF-MS鉴定,同时回顾形态学方法鉴定结果,对丝状真菌菌种分布、标本来源进行分析,采用Kappa值一致性评价方法对质谱鉴定与形态学方法鉴定结果进行统计分析。结果本研究共包含分离自临床的丝状真菌102株,非重复分离株83株。烟曲霉50株(60.24%),黄曲霉26株(31.33%)。标本来源主要为呼吸道标本,其中痰液68株(81.93%),支气管灌洗液11株(13.25%)。MALDI-TOF-MS技术与形态学方法鉴定结果比对,Kappa值为0.141 0。结论 MALDI-TOF-MS技术在丝状真菌鉴定正确性与时效性优于形态学方法。     

两种酵母样真菌培养鉴定方法评价

目的 选择和评价实用性强、快速、准确酵母样真菌培养鉴定方法。为临床诊断和治疗真菌感染性疾病提供可靠的病原学依据。方法 将API酵母样真菌鉴定试条鉴定出的白色念珠菌（48株），热带念珠菌（15株），克柔氏念珠菌（19株）。光滑念珠菌（6株）转种科玛嘉酵母样真菌显色平板，经35℃，24—48小时培养，分析科玛嘉酵母样真菌显色平板鉴定结果正确率。结果 科玛嘉显色平板鉴定出白色念珠菌43株，热带念珠菌13株，克柔氏念珠菌8株，光滑念珠菌5株。科玛嘉显色平板法与酵母样真菌鉴定符合率为80％-90％．结论 科玛嘉显色平板法操作简单、快速。价格低廉，可作为酵母样真菌鉴定的初筛方法。     

RapID真菌快速鉴定系统临床应用评价

由于广谱抗生素及免疫抑制剂、静脉置管的广泛使用，使得真菌的机会感染大大地增加。传统的鉴定真菌的方法主要是依据形态学的观察和碳水化合物的同化试验，以后在此基础上又发展了商品化的鉴定系统，主要有法国Biomerieux公司的AP120C、ATB、ID32C、VITEKl的YBC卡、VITEK2的ID-YST卡、美国REMEL公司的RapiD真菌鉴定系统以及用分子生物学的方法来鉴定酵母样真菌，而RapiD真菌鉴定系统是鉴定速度最快（4h）的一种系统。本实验就是将快速真菌鉴定系统与AP120C方法进行比较，并对其结果进行评价。     

常见酵母样真菌的纸片鉴定法

近年来,由于广谱抗生素、肾上腺皮质激素及其它免疫抑制剂与细胞毒性药物的广泛应用,各种外科手术的广泛开展,深部真菌感染不断增多,早期病原学诊断和及早采取针对性治疗是临床亟待解决的问题。传统的培养鉴定法程序较复杂,费时较长,尚不能完全满足临床及早诊治的要求。Sobczak 从80余种化学药     

酵母样真菌的鉴定及耐药分析

目的：探讨本地区医院感染酵母样真菌的种类、样本分布，及各菌种对常用药物敏感程度。方法：真菌培养采用常规方法，酵母样真菌鉴定卡进行鉴定，用ATB Fungus药敏板测定对六种抗真菌药物的耐药性。结果：分离出233株酵母样真菌中，白色念珠菌占66．1％，热带念珠菌占18．0%；对唑类抗真菌药物耐药率相对高，克柔念珠菌对两性霉素敏感，对另几种抗真菌药物显示较高的耐药性。结论：非白色念珠菌的检出率有所增加；临床应根据实验室提供的药敏结果合理使用抗真菌药物。     

临床酵母样真菌实验室检查

酵母样真菌是一类以单细胞形式存在的真菌,临床上重要的酵母样真菌多属于子囊菌门和担子菌门。精准的实验室诊断是实现精准临床诊疗的前提和基础。该文介绍了酵母样真菌的实验室检查方法及临床应用,包括基于培养的检查方法,如菌落形态、镜下形态、显色培养基、芽管形成试验、生化反应、DNA测序和基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)菌落鉴定;以及非培养检查方法,包括标本直接镜检、组织病理、抗原检测、核酸检测和MALDI-TOF MS对临床标本的直接检测等,并分析各方法性能特点。     

临床标本酵母样真菌的分离鉴定及药敏分析

随着器官移植、心血管及人工瓣膜手术的开展,置留在体内的各种导管的增加,肿瘤化疗和放疗,大剂量广谱抗菌素、激素、免疫抑制剂的广泛应用,使病人的机体免疫力下降。条件致病菌包括酵母菌便乘机而入,使机体由带菌状态发展为大量繁殖,导致严重感染。我们将我院一年中临床标本分离出的酵母样真菌进行鉴定及药敏试验。报告如下:     

酵母样真菌的鉴定与结果分析

酵母样真菌是一种呈酵母样菌落形态的单细胞芽生真菌,是临床上重要的条件致病菌,它可以引起全身组织和器官的条件感染,特别是院内继发性感染.近年来,由于广谱抗生素.免疫抑制剂及皮质类固醇激素的广泛应用,导致酵母样真菌感染日益增多,其中主要是白色念珠菌.现将本院1993年以来鉴定的酵母样真菌的结果报道如下.     

痰中酵母样真菌分离鉴定的体会

我们对本院近3年来临床送检痰标本中，分离出的酵母样真菌进行了分类鉴定。现将菌种的分类与分离鉴定的体会报告如下，以供参考。     

Analysis of minor hemoglobins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

BACKGROUND: Hemoglobin (Hb) heterogeneity arises mainly from posttranslational modifications of the globin chains, and cation-exchange chromatography reveals falsely increased concentrations of some minor Hbs in the presence of abnormal Hbs. Here we describe a method for identification of the globin chains and their posttranslational modifications contained in the Hb fractions. METHODS: We used cation-exchange HPLC (PolyCAT A column) for separation of Hb fractions and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for analysis of the separated globin chains. Globin chains were identified by their molecular masses. Posttranslational modifications of globin chains were identified by digestion of the proteins with endoproteinase V8 before MALDI-TOF MS of the resulting peptides. RESULTS: Analysis of the HbA2 fractions of patients with HbS revealed 4 different globin chains. We found, in addition to the expected alpha- and delta-chains, the carbamylated alpha- and the betaS-chains. Additionally, we analyzed HbH, Hb Barts, HbA 1b, pre-HbA 1c, HbA 1c, HbF1, HbF, HbA 1d3a, HbA 1d3b, HbA2, and HbC1 fractions from control and pathologic blood samples. We identified several posttranslational modifications of the globin chains, such as pyruvatization, glycation, acetylation, carbamylation, and acetaldehyde adduct formation. CONCLUSIONS: The native and posttranslationally modified globin chains in minor and major Hbs are unambiguously identified by MALDI-TOF MS. A minor Hb containing the carbamylated alpha- and the betaS-chain elutes at the same time as normal HbA2 (alpha2delta2) and thus leads to falsely increased HbA2 values in patients with HbS when blood is analyzed with PolyCAT A chromatography.     

感染性疾病多学科交叉研究专题基质辅助激光解析电离飞行时间质谱技术在病原菌鉴定中的应用

目的 探讨基质辅助激光解析电离飞行时间质谱技术(MALDI-TOF MS)在快速鉴定血流感染病原菌的应用价值,并分析MALDI-TOF MS方法鉴定病原菌的准确性.方法 收集321例血流感染患者的血液标本进行血培养,微生物室细菌培养结果均为阳性.从血培养阳性瓶中取病原菌至分离胶促凝管中,应用MALDI-TOF MS对富...     

Application of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in Alzheimer's disease

Alzheimer's disease is becoming an increasing problem in our aging society. According to our knowledge, so far, no effective pharmacotherapy to cure the cause of the disease has been developed. Therefore, early diagnosis is needed, which will result in implementation of a drug therapy aimed at decreasing and/or inhibiting disease development. Mass spectrometry techniques (MS) have a wide range of applications in proteomics and the search for biomarkers of neurodegenerative disorders, opening new possibilities in diagnostics. Identification of proteins in body fluids (like cerebrospinal fluid or blood) is possible due to MS spectra analysis. The detected changes in protein concentrations are connected with pathological states in an organism and, therefore, can be regarded as biomarkers. Developing procedures for proteome analysis might result in fast diagnosis, as well as creating better suited pharmaceuticals. This paper reviews the search of biomarkers in cerebrospinal fluid and blood. Later on, the use of matrix-assisted-laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in proteomics, focusing on blood-related biomarkers, is discussed. The aim of the work is also to highlight the advantages and disadvantages of MALDI-TOF-based analyses.     

New serum biomarkers for detection of endometriosis using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

This study used proteomic fingerprint technology, combining nano-sized magnetic beads with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), to screen for potential protein biomarkers for the diagnosis of endometriosis. Serum proteins from 126 patients with endometriosis and 120 healthy controls were profiled and compared. Biomarker pattern software identified 46 discriminating mass-to-charge m/z ratio peaks that were related to endometriosis. The model constructed by the software, based on three of these peaks (m/z 5988.7, 7185.3 and 8929.8), generated excellent separation between the endometriosis and control groups. The sensitivity was 91.4% and the specificity 95.0%. Blind testing on a second series of serum samples from patients with endometriosis and healthy controls indicated a sensitivity of 89.3% and a specificity of 90.0%. Biomarkers for endometriosis can be discovered in serum by MALDI-TOF-MS in combination with nano-sized magnetic beads. The pattern of combined markers provides a powerful and reliable diagnostic method for endometriosis, with high sensitivity and specificity.     

基于基质辅助激光解吸电离飞行时间质谱的克柔念珠菌遗传分化研究

目的在蛋白质水平明确克柔念珠菌的遗传分化特征，建立基于基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）的不同谱系克柔念珠菌的鉴定方法。方法选取20株克柔念珠菌，采用MALDI-TOF MS结合分析软件ClinProTools，确定克柔念珠菌不同谱系的质谱标志峰，构建基于遗传算法（GA）的分类模型，用于区分不同谱系的克柔念珠菌菌株，选择25株克柔念珠菌对该模型进行验证。结果利用20株克柔念珠菌构建的GA分类模型，反映模型正确鉴别能力的识别能力值（RC）为100%，反映模型处理测试谱图变异能力的交叉验证值（CV）为97.89%。 25株克柔念珠菌、150张谱图外部验证模型显示其分类能力为95.30%。 确定13个克柔念珠菌谱系相关的特征峰（m/z 3 971.40、3 136.95、3 427.33、2 405.28、2 996.73、2 913.95、3 376.97、6 736.13、5 819.03、4 045.16、5 869.00、3 618.10及3 946.14），可用于正确区分来自不同谱系的克柔念珠菌。结论本研究论证并确认了克柔念珠菌的群体遗传分化事件，同时也建立了一种简便而有效的快速谱系分型方法。     

Characterization of glycation adducts on human serum albumin by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

BACKGROUND: Non-enzymatic glycation of human serum albumin (HSA) is associated with the long-term complications of diabetes. We examined the structure and location of modifications on minimally-glycated HSA and considered their possible impact on the binding of drugs to this protein. METHODS: Minimally-glycated and normal HSA (used as a control) were digested with trypsin, Glu-C or Lys-C, followed by fractionation of the resulting peptides and their analysis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to determine the structures and locations of glycation adducts. RESULTS: Several specific lysine and arginine residues were identified as modification sites in minimally-glycated HSA. Residues K12, K51, K199, K205, K439 and K538 were found to be modified through the formation of fructosyl-lysine, while the modification of K159 and K286 involved the formation of pyrraline or N(epsilon)-carboxymethyl-lysine, respectively. Lysine K378 was found to give N(epsilon)-carboxyethyl-lysine in some forms of glycated HSA but fructosyl-lysine in other forms. Residues R160 and R472 produced a modification based on N(epsilon)-(5-hydro-4-imidazolon-2-yl)ornithine. Lysine R222 was modified to produce argpyrimidine, N(epsilon)-[5-(2,3,4-trihydroxybutyl)-5-hydro-4-imidazolon-2-yl]ornithine or tetrahydropyrimidine. CONCLUSIONS: With the exception of K12, K199, K378, K439 and K525, all of the observed sites of modification for minimally-glycated HSA were new to this current study. The fact that many of these glycation-related modifications are located at or near known drug binding sites on HSA explains why some differences have been previously noted in the binding of certain drugs to normal vs glycated HSA.     

用MALDI-TOF MS技术快速鉴定临床分离棒状杆菌属细菌

目的评价基质辅助激光解吸/电离飞行时间质谱(MALDI-TOF MS)快速鉴定临床分离的非白喉棒状杆菌的价值。方法收集本院58株非白喉棒状杆菌,用MALDI-TOF MS和16S rRNA基因测序两种方法进行鉴定和比较。用MALDI-Biotyper软件构建不同棒状杆菌MALDI-TOF MS蛋白系统发育树。结果 58株非白喉棒状杆菌中,54株MALDI-TOF MS与16S rRNA基因测序鉴定结果一致,包括34株纹带棒状杆菌(Corynebacterium straitum)、11株杰氏棒状杆菌(C.jeikeium)、3株C.resistens、2株解葡萄糖苷棒状杆菌(C.glucuronolyticum)、2株黏金色棒状杆菌(C.aurimucosum)和2株无枝菌酸棒状杆菌(C.amycolatum)。4株16S rRNA基因测序无法鉴定到种的菌株中,MALDI-TOF MS鉴定为2株产黏棒状杆菌(C.mucifaciens)、1株C.singulare和1株假白喉棒状杆菌(C.commune)。结论 MALDI-TOF MS可将棒状杆菌属细菌快速、准确地鉴定到种。     

对《MALDI-TOF-MS在临床常见菌快速鉴定中的应用研究》一文的商榷

编辑同志:拜读了贵刊2010年33卷第6期蒋颜等[1]的研究报道,令人兴奋.应用MALDI-TOF-MS技术鉴定细菌,近年来国外才有开展,作者能在国内迅速开展并取得研究成果,笔者深感敬佩.但也许由于文章字数的限制,使笔者对文中的一些内容不太理解,希作者指教.