Analysis of Single Nucleotide Polymorphism in Human Paraoxonase 1 Gene(Q192R) with Matrix-assisted Laser Desorption/Ionization Time-of-flight Mass Spectrometry

Introduction Single nucleotide polymorphisms （SNPs） are the most abundant DNA markers in the human genome occurring at a frequency of one in every 500--1000 nucleotides. A variety of methods have been used for the analysis of single nucleotide polymorphisms, including restriction fragment length polymorphism （RFLP）, direct sequencing by using laser-induced fluorescence detectionTM, fluorescence energy transfer, MALDI-TOF MS combined with primer extension or invasive cleavage, and fluorescence polarization. During the past two decades, mass spectrometry has become a very popular tool in the analysis of biomolecules and is perfectly suited to the analysis of single nucleotide polymorphisms （SNPs） due to its speed, low cost, and accuracy. In this work, we used MALDI TOF mass spectrometry to detect the fragments of restriction endonuclease hydrolysis of PCR products flanking a SNP located at paraoxonase 1（Q192R）. Compared with electrophoresis, this method requires less time of analysis and possess a higher accuracy.

A novel sample preparation method of matrix-assisted laser desorption/ionization time of flight mass spectrometry for polystyrene

A novel sample preparation method of matrix-assisted laser desorption/ionization mass spectrometry for polystyrene was reported. Compared to the conventional dried-droplet method, the efficiency of ionization and signal intensity of mass spectra were improved. The mechanism was also analyzed.

MicrobMatcher: a microbial comparison software based on matrix-assisted laser desorption/ionization with time-of-flight mass spectrometry

Matrix-assisted Laser Desorption/Ionization with Time-of-flight Mass Spectrometry (MALDI-TOFMS) was investigated as a method for the rapid identifica-tion of species. Current demand in microbial identi-fication is how to compare unknown strains to the known one quickly, semi-automatically and accurately. In this paper, we present a software tool that allows flexibly microbial matching in a user-friendly way, by letting the users to customize comparison parameters including: in vitro transcription enzyme, mass tolerance,minimum fragment length, intensity threshold and corresponding weights. We provide three spectral scoring functions to compute the affin-ity between the species. Therefore, the precision of microbial comparison increases. To test and verify this tool, we employed experimental spectral data based on MALDI-TOFMS and the gene sequences of E.coli and Salmonella. This software is written in Java for cross-platform intention.

Two-dimensional gel electrophoresis and surface-enhanced laser desorption ionization-time of flight-mass spectrometry for detection of protein expression profiles in the hippocampus following closed brain injury

Gene expression profile changes in brain regions following traumatic brain injury at the gene level cannot sufficiently elucidate gene expression time, expression amount, protein post-translational processing or modification. Therefore, it is necessary to quantitatively analyze the gene expression profile using proteomic techniques. In the present study, we established a rat model of closed brain injury using Marmarou＇s weight-drop device, and investigated hippocampal differential protein expression using two-dimensional gel electrophoresis and surface-enhanced laser desorption ionization-time of flight-mass spectrometry. A total of 364 protein peaks were detected on weak cation exchange-2 protein chips, including 37 differential protein peaks. 345 protein peaks were detected on immobilized metal affinity capture arrays-Cu, including 12 differential protein peaks Further examination of these differential proteins revealed that glucose-regulated protein and proteasome subunit alpha type 3 expression were significantly upregulated post-injury. These results indicate that brain injury can alter protein expression in the hippocampus, and that glucose-regulated protein and proteasome subunit alpha type 3 are closely associated with the occurrence and development of traumatic brain injury.

Detection of lung adenocarcinoma using magnetic beads based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry serum protein profiling

Background Recently, due to the rapid development of proteomic techniques, great advance has been made in many scientific fields. We aimed to use magnetic beads （liquid chip） based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS） technology to screen distinctive biomarkers for lung adenocarcinoma （adCA）, and to establish the diagnostic protein profiles. Methods Using weak cation exchange magnetic beads （MB-WCX） to isolate and purify low molecular weight proteins from sera of 35 lung adCA, 46 benign lung diseases （BLDs） and 44 healthy individuals. The resulting spectra gained by anchor chip-MALDI-TOF-MS were analyzed by ClinProTools and a pattern recognition genetic algorithm （GA）. Results In the working mass range of 800-10 000 Da, 99 distinctive peaks were resolved in lung adCA versus BLDs, while 101 peaks were resolved in lung adCA versus healthy persons. The profile gained by GA that could distinguish adCA from BLDs was comprised of 4053.88, 4209.57 and 3883.33 Da with sensitivity of 80%, specificity of 93%, while that could separate adCA from healthy control was comprised of 2951.83 Da and 4209.73 Da with sensitivity of 94%, specificity of 95%. The sensitivity provided by carcinoembryonic antigen （CEA） in this experiment was significantly lower than our discriminatory profiles （P 〈0.005）. We further identified a eukaryotic peptide chain release factor GTP-binding subunit （eRF3b） （4209 Da） and a complement C3f （1865 Da） that may serve as candidate biomarkers for lung adCA. Conclusion Magnetic beads based MALDI-TOF-MS technology can rapidly and effectively screen distinctive proteins/polypeptides from sera of lung adCA patients and controls, which has potential value for establishing a new diagnostic method for lung adCA.

Studies on extracting solutions of endohedral rare-earth metallofullerenes by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Thirteen extracting solutions of rare-earth metallofullerenes containing La,Ce,Pr,Nd Sm,Eu,Gd,Tb,Dy,Ho,Er,Tm and Yb respectively have been investigated by means of matrix-assisted laser desorpuon/ ionization time-of-flight mass spectrometry.The influences of the positive-ion/negative-ion mode,laser intensity,ma trix and mass discrimination to the analytical results are studied,based on which the optimal analytical conditions have been determined.The results show that the extracting solutions contain large quantities of rare-earth metallofullerenes besides empty fullerenes.On the basis of comparing their relative intensities,the different structure stabilities and solubilities of metallofullerenes with different rare-earth metals encapsulated into the fullerene cages,as well as some possible reasons to those differences,are discussed.

Detection of Sialylated N-Linked Glycans by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone（THAP）and 2,5-dihydroxybenzoic acid（DHB）as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer（MALDI-TOF MS）.High quality negative-ion spectra of commercial sialylated glycan were obtained with THAP as matrix.Detection limit of the glycan was less than 0.1 pmol.After methyl esterification of sialic acid（SA）residue,sialylated glycans were detected sensitively in the positive-ion mode using DHB as matrix.Neutral and sialylated glycans from the mixture of asialofetuin and fetuin were methylesterified and simultaneously recognized in one manipulation.Methyl esterification of SA residue offers a convenient and sensitive way to identify the structure of N-linked glycans for glycan profiling.

Identification of serum biomarkers for diagnosing stage Ⅰ lung adenocarcinoma by MALDI-TOF mass spectrometry

Objective To identify specific biomarkers that could improve early diagnosis of lung adenocarcinoma using matrix-assisted laser desorption/ionization (MALDI) technology. Methods Serum samples were isolated from 17 patients with stage Ⅰ lung adenocarcinoma and 17 age-and sex-matched healthy controls,and the serum proteomic profiles were obtained by matrix-assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry. Results Compared with healthy control group,two highly expressed potential biomarkers were identified with the relative molecular weights of 6 631.64 Da and 4 964.21 Da. The two best novel protein peaks were automatically chosen for the system training and the development of the constructed model. The constructed model was then used to test an independent set of masked serum samples from 15 lung adenocarcinoma patients and 22 healthy individuals. The analysis yielded a sensitivity of 93.3%,and a specificity of 95.5%. Conclusion These results suggest that MALDI-TOF-MS ProteinChip technology is a quick,convenient,and high-output analyzing method that is capable of selecting several relatively potential biomarkers from the serum of lung adenocarcinoma patients and may have a clinical value in the future,and will provide clues to identifying new serologic biomarkers of lung adenocarcinoma.

Establishment and Preliminary Application of Two dimensional Electrophoresis and Mass Spectrometry in Ovary Study

Objective To apply two-dimensional electrophoresis and mass spectrometry in the ovary proteome researchMethods Protein extractions from mouse ovaries were run in IPGphor isoelectric focus system with 11 cm and 24 cm IPG strips respectively (pH 3～10, 0.3 mm thick), then the protein spots were identified by mass spectrometry.Results The ovary protein exactions separated by two-dimensional electrophoresis have got high resolution, and identifing protein by mass spectrometry was highly efficient and facilitly. These two techniques should facilitate further investigation of female reproduction proteome research.Conclusion These two rapid high resolutions and efficient techniques have a variety of applications foreground in female reproduction proteome pattern research.

核酸MALDI-TOF MS检测在肺结核诊断中的价值

目的探讨基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)在疑似肺结核(PTB)诊断中的应用价值及其阳性结果的影响因素,以期为临床治疗提供参考。方法选取甘肃省第二人民医院(西北民族大学附属医院)2022年8月—2023年3月收治的疑似PTB患者77例为研究对象。所有患者肺泡灌洗液(BALF)均接受MALDI-TOF MS检测和结核分枝杆菌(MTB)培养。根据MTB培养结果,对核酸MALDI-TOF MS检测进行诊断效能分析,并探讨核酸MALDI-TOF MS检测阳性的影响因素。结果核酸MALDI-TOF MS检测结果显示,MTB阳性共37例(48.1%,37/77),40例阴性(51.9%,40/77)。以培养结果为金标准,核酸MALDI-TOF MS的灵敏度、特异度、阳性预测值(PPV)、阴性预测值(NPV)和准确度分别为90.9%、84.1%、81.1%、92.5%和87.0%,采用Kappa检验法评价核酸MALDI-TOF MS检测和MTB培养法的一致性,结果显示Kappa值为0.739(P<0.05)。2组患者在是否有糖尿病、身体质量指数(BMI)及降钙素原(PCT)方面比较,差异均有统计学意义(P<0.05)。多因素Logistic回归分析结果显示:有糖尿病和PCT异常为核酸MALDITOF MS阳性的独立危险因素(P<0.05)。结论核酸MALDI-TOF MS检测对PTB的早期诊断具有重要价值,可为患者的规范用药提供更好的指导,即为患者的治疗制定更为精准、有效的治疗方案,从而提高治疗效果、改善患者生活质量。

MALDI-TOF MS技术在临床微生物检验中的应用

随着科学技术的发展,新技术在医学检验中的应用越来越广泛。相较于传统微生物鉴定繁琐、耗时的工作流程,基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS,简称MS)技术因其快速、灵敏、简便、省时和特异性高等特点,在菌种鉴定、耐药监测、突变菌株鉴别等方面发挥巨大的作用,逐渐成为临床微生物实验室不可或缺的微生物鉴定工具。虽然MS技术鉴定菌种高度依赖数据库,也在一定程度上受限于临床样本质量,但其在临床微生物检验中仍有较大的空间待开发。文章对MS技术在临床微生物检验中的应用进行综述。

MALDI-TOF MS直接靶板微滴生长测定法在CRKP快速检测中的应用

目的探讨基于基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)的直接靶板微滴生长测定(DOT-MGA)法快速筛查碳青霉烯类耐药肺炎克雷伯菌(CRKP)的准确性和临床应用价值。方法收集2020—2021年连云港市第一人民医院肺炎克雷伯菌临床分离株251株,其中美罗培南敏感123株,CRKP128株。采用DOT-MGA法对4和6μL 2种取样量的所有菌株进行美罗培南耐药性检测。采用微量肉汤稀释法测定菌株的美罗培南最小抑菌浓度(MIC)。以微量肉汤稀释法耐药性分析结果为标准,评价DOT-MGA法耐药性检测结果的准确性。结果以微量肉汤稀释法为标准,孵育生长4 h后,DOT-MGA法2种取样量的敏感性和阴性预测值均为100.0%,其中4μL取样量检测CRKP的特异性为92.5%,阳性预测值为92.8%;6μL取样量检测CRKP的特异性为89.8%,阳性预测值为90.1%。123株美罗培南敏感株的美罗培南MIC_(50)为0.125μg·mL^(-1),MIC_(90)为0.25μg·mL^(-1);128株美罗培南耐药株的MIC_(50)和MIC_(90)均为≥128μg·mL^(-1)。DOT-MGA法与微量肉汤稀释法的一致性分析结果显示,4μL加样量Kappa值为0.88,6μL加样量Kappa值为0.83。结论基于MALDITOF MS的DOT-MGA法筛查CRKP具有较高的准确性,且简便、快捷,可用于快速检测CRKP。

MALDI-TOF MS法快速鉴定散装即食食品中3种食源性致病菌

本研究旨在利用基质辅助激光解吸电离飞行时间质谱法(Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry,MALDI-TOF MS)快速检测并鉴定散装即食食品中的沙门氏菌、金黄色葡萄球菌和蜡样芽孢杆菌3种食源性致病菌。通过对点样方式、前处理方式、不同培养基及培养时间等各项实验条件的优化筛选确定最佳方法,并考察其检测限和稳定性,同时用所建立的方法与多重PCR法、VITEK 2等生化鉴定法进行结果比对,比较三者鉴定结果的一致性,最后进行批量食品样本检测来探寻MALDI-TOF MS鉴定法在散装即食食品中的适用性。结果表明,三种目标菌同一批次及不同传代次数的鉴定得分均大于95%,变异系数均小于2%,所建立的方法重复性和稳定性良好,且检测限为10^(5)CFU/mL,MALDI-TOF MS菌株鉴定结果与多重PCR法、生化鉴定法结果高度一致,且比二者检测用时更短,更加特异高效;在点样方式上,相较于夹心法与混合干滴法,三种覆盖法点样平均鉴定得分更高,均为99.9%,其中1μL+1μL覆盖法点样可获得的平均峰数目更多,特征峰更复杂且强度更高;而在前处理方式上,甲酸乙腈法与其他两种菌体处理方法相比可获得更多的出峰量和更高的峰强度,平均鉴定得分均在98%以上,且与数据库中图谱匹配度更高。综上所述,MALDI-TOF MS法在致病菌鉴定方面不仅灵敏特异、准确快速,且操作简便、结果直观,作为国标法的有力补充,为食源性致病菌的分析鉴定与分型溯源提供了新思路。

Mode of interaction of calcium oxalate crystal with human phosphate cytidylyltransferase 1: a novel inhibitor purified from human renal stone matrix

Nephrolithiasis is a common clinical disorder, and calcium oxalate (CaOx) is the principal crystalline component in approximately 75% of all renal stones. It is widely believed that proteins act as inhibitors of crystal growth and aggregation. Acidic amino acids present in these proteins play a significant role in the inhibition process. In this study, interaction of cal-cium oxalate with human phosphate cytidylyltrans-ferase 1(CCT), a novel calcium oxalate crystal growth inhibitor purified from human renal stone matrix has been elucidated in silico and involvement of acidic amino acids in the same. As only sequence of CCT is available, henceforth its 3-D structure was modeled via Homology modeling using Prime module of Schrodinger package. Molecular dynamic simulation of modeled protein with solvation was done by mac-romodel (Schrodinger). The quality of modeled pro-tein was validated by JCSG protein structure valida-tion (PROCHECK & ERRAT) server. To analyze the interaction of modeled protein CCT with calcium oxalate along with role played by acidic amino acids, ‘Docking simulation’ was done using MOE–Dock. Interaction between calcium oxalate and CCT was also studied by substituting acidic amino acid in the active sites of the protein with neutral and positively charged amino acids. The in silico analysis showed the bond formation between the acidic amino acids and calcium atom, which was further substantiated when substitution of these acidic amino acids with alanine, glycine, lysine, arginine and histidine com-pletely diminished the interaction with calcium ox-alate.

Autof ms 1000与MALDI Biotyper对临床常见丝状病原真菌鉴定的比较分析

目的:研究比较两种基质辅助激光解吸电离飞行时间质谱仪(Autof ms 1000与MALDI Biotyper)对临床常见丝状病原真菌鉴定的准确性和效率。方法:共纳入32株丝状病原真菌标准菌株和120株临床分离菌株,按照标准操作方法分别用Autof ms 1000与MALDI Biotyper对相关菌株进行鉴定,对相关结果及测序分子鉴定结果进行对比分析。结果:在种水平上,Autof ms 1000质谱仪共检出140株丝状病原真菌,检出正确率为92.1%(140/152),其中126株分值在9.0以上,2株出现错误鉴定,误鉴定比例为1.3%(2/152);MALDI Biotyper质谱仪共正确检出98株,检出正确率为64.4%,2.0分以上的菌株47株,仅3株鉴定分值大于2.3分,7株出现误鉴定。结论:Autof ms 1000与MALDI Biotyper这两种质谱鉴定系统均能用于临床常见丝状病原真菌的快速鉴定,而Autof ms 1000质谱仪更具优势。

Autof MS1000质谱鉴定系统快速检测肺炎克雷伯菌和铜绿假单胞菌产碳青霉烯酶的临床价值评估

目的探讨基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)快速检测肺炎克雷伯菌和铜绿假单胞菌产碳青霉烯酶的临床价值。方法收集2022年1月至2023年10月徐州医科大学附属邳州市人民医院分离的肺炎克雷伯菌60株和铜绿假单胞菌80株,其中碳青霉烯类耐药肺炎克雷伯菌(CRKP)30株,碳青霉烯类敏感肺炎克雷伯菌(CSKP)30株,碳青霉烯类耐药铜绿假单胞菌(CRPA)50株,碳青霉烯类敏感铜绿假单胞菌(CSPA)30株。分别采用改良碳青霉烯灭活试验(mCIM)、胶体金免疫层析法和Autof MS 1000质谱鉴定系统进行检测,评估Autof MS 1000质谱鉴定系统检测肺炎克雷伯菌和铜绿假单胞菌产碳青霉烯酶的能力。结果Autof MS 1000质谱鉴定系统检测结果与mCIM和胶体金免疫层析法结果完全一致,30株CRKP中,有28株检测到碳青霉烯酶,2株阴性;50株CRPA中,有15株检测到碳青霉烯酶,35株阴性;30株CSKP和30株CSPA均为阴性。3种方法检测碳青霉烯酶的结果符合率为100%。结论Autof MS 1000质谱鉴定系统检测碳青霉烯酶的结果与mCIM法和胶体金免疫层析法一致,且既有mCIM法低成本的特点,又有胶体金免疫层析技术速度快、准确率高的优点,可用于临床肺炎克雷伯菌和铜绿假单胞菌产碳青霉烯酶菌株的快速鉴定。

MALDI-TOF-MS技术在肉芽肿性乳腺炎脓液棒状杆菌检测中的应用研究

目的:研究拟通过基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)技术在脓液克氏棒状杆菌(CK)鉴定过程中的应用探索,为临床微生物实验室对CK的准确鉴定提供帮助。方法:无菌条件下分别采集2019年1月-2021年7月镇江市妇幼保健院乳腺科门诊及病房30例经病理确诊为肉芽肿性乳腺炎(GM)患者手术中获取的新鲜脓肿穿刺液标本,用于细菌、真菌、结核菌涂片检查及需氧菌、厌氧菌培养。结果:采用梅里埃VITEK2compact全自动细菌鉴定仪进行生化鉴定,结果表明,23株菌鉴定为CK,其余7株菌均鉴定失败或错误。布鲁克MALDIBiotyper质谱仪鉴定结果27株为CK,仅3株菌鉴定失败,该结果与16SrRNA测序结果一致性较高,但是27份标本中鉴定为CK的质量分数相对较低。结论:MALDI-TOF-MS技术在棒状杆菌属细菌鉴定方面的应用可以为临床诊治GM方面提供病原学理论依据。

MALDI-TOF MS与VITEK2在血流感染致病菌快速鉴定及药敏试验中的应用

目的:探究基质辅助激光解吸/电离飞行时间质谱(Matrix-assisted laser desorption/ionizationtime-of-flight mass spectrometry,MALDI-TOF MS)与VITEK2在血流感染致病菌药敏试验中的联合应用效果。方法:选取景德镇市第二人民医院检验科2022年1月1日至2022年12月31日血流感染阳性样本126份,分别行常规鉴定及MALDI-TOF MS鉴定,完成菌株鉴定后通过VITEK2系统行抗菌药物敏感性试验(Antimicrobial susceptibility testing,AST),比较2种鉴定方法的细菌菌株鉴定结果及细菌耐药情况。结果:常规鉴定方法对革兰氏阴性、革兰氏阳性菌在属、种水平鉴定符合率达到100%;MALDI-TOF MS鉴定革兰氏阴性菌在属水平的符合率达97.7%(85/87),在种水平的符合率达95.4%(83/87),2例无结果(1例阴沟肠杆菌阴沟亚种,1例少动鞘氨醇单胞菌);MALDI-TOF MS鉴定革兰氏阳性菌在属、种水平的符合率均达97.4%(38/39),1例无结果(科氏葡萄球菌解脲亚种)。常规鉴定+VITEK2对革兰氏阴性菌AST分析的分类一致(Categorical agreement,CA)、基本一致(Essential agreement,EA)分别达到98.6%(1086/1101)、99.5%(1096/1101),对革兰氏阳性菌AST分析的CA、EA分别达到98.6%(550/558)、99.6%(556/558);MALDI-TOF MS+VITEK2(VITEK MS)对革兰氏阴性菌AST分析的CA、EA分别达到98.3%(1082/1101)、99.5%(1095/1101),对革兰氏阳性菌AST分析的CA、EA分别达到97.8%(546/558)、99.5%(555/558)。进一步参考肉汤稀释法药敏结果行一致性分析。结果发现,常规鉴定+VITEK2在革兰氏阴性菌的药敏试验中,10份AST鉴定结果出现小误差(Minus errors,MIE),5份AST鉴定结果出现重大误差(Major errors,ME);在革兰氏阳性菌的药敏试验中,8份AST鉴定结果出现MIE,2份AST鉴定结果出现ME。VITEK MS在革兰氏阳性菌的药敏试验中,13份AST鉴定结果出现MIE,6份AST鉴定结果出现ME;在革兰氏阳性菌的药敏试验中,9份AST鉴定结果出现MIE,3份AST鉴定结果出现ME。结论:MALDI-TOF MS联合VITEK2在临床血流感染致病菌鉴定及药敏试验中可快速、精确实现诊断并提供AST报告。

分离胶促凝管联合MALDI-TOF MS直接检测血培养阳性细菌

目的建立用分离胶促凝管联合基质辅助激光解析电离飞行时间质谱（MALDI-TOF MS）直接检测血培养阳性待测菌的方法并评估其鉴定符合率。方法共收集阳性血培养491例,利用分离胶促凝管直接从血培养瓶中富集并提纯菌体,采用MALDI-TOF MS对待测菌进行菌种鉴定,同时对报阳性血培养瓶进行转种培养,纯菌落用Vitek 2 Compact全自动微生物分析系统（简称Vitek 2 Compact）进行鉴定。若两者鉴定结果不一致,则以基因测序结果予以确证。结果 491例阳性血培养瓶鉴定出的待测菌包括30个属和64个种。在462例单菌株感染血培养瓶中,菌株的种、属鉴定符合率分别为73.6%和3.7%。其中175例革兰阴性菌的种、属鉴定率分别为84.0%（147例）和2.9%（5例）;251例革兰阳性菌的种、属鉴定率分别为75.3%（189例）和4.4%（11例）;36例念珠菌的种、属鉴定率分别为19.4%（7例）和2.8%（1例）。在29例复数菌感染血瓶中,鉴定出其中一种细菌的种、属鉴定率分别为79.3%（23例）和10.3%（3例）。血流感染中常见病原菌的菌种鉴定符合率达83.3%～96.9%。结论本研究建立了分离胶促凝管联合MALDI-TOF MS直接检测血培养阳性待测菌的方法,与传统的培养鉴定方法相比,对血流感染中主要病原菌的鉴定符合率较高,且方法快速、简便,成本低廉,适合在临床微生物实验室中推广应用。

MALDI-TOF MS与16SrDNA方法对肠杆菌科微生物鉴定比较分析

为了研究基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)法鉴定肠杆菌科微生物及其对微生物系统分类学相关分析的准确性,采用MALDI-TOF MS对金桔表面分离的10种肠杆菌科微生物进行蛋白质图谱收集,通过比对图谱特征峰实现微生物的鉴定与系统进化学分析;同时对10种微生物进行16SrDNA提取与测序,得到分子生物化学水平鉴定,并采用邻近连接法对16SrDNA序列做系统进化树分析。结果表明:MALDI-TOF MS与16SrDNA测序对10种肠杆菌科微生物鉴定结果中,克雷伯菌属2株菌鉴定种级有偏差,其他8种一致;MALDI-TOF MS与16SrDNA测得的数据都可计算微生物相关性,从而得到微生物系统发育树,两株系统发育树中同属级微生物归类的亲缘位置与方向一致,但不同属肠杆菌科微生物的亲缘距离与位置差异较大。MALDI-TOF MS法鉴定农产品表面微生物具有快速、准确的特点,但准确建立系统发育相关性还需要扩大数据库和优化算法。