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通过适体功能化金纳米颗粒比色法检测铜绿假单胞菌

Colorimetric detection of Pseudomonas aeruginosa by aptamer-functionalized gold nanoparticles.

作者信息

Schmitz Fernanda Raquel Wust, Cesca Karina, Valério Alexsandra, de Oliveira Débora, Hotza Dachamir

机构信息

Department of Chemical Engineering and Food Engineering (EQA), Federal University of Santa Catarina (UFSC), Florianópolis, SC, 88040-900, Brazil.

出版信息

Appl Microbiol Biotechnol. 2023 Jan;107(1):71-80. doi: 10.1007/s00253-022-12283-5. Epub 2022 Nov 24.

Abstract

Novel rapid methodologies for the detection of bacteria have been recently investigated and applied. In hospital environments, infections by pathogens are very common and can cause serious health problems. Pseudomonas aeruginosa is one of the most common bacteria, which can grow in hospital equipment such as catheters and respirators. Even at low concentrations, it can cause severe infections as it is resistant to antibiotics and other treatments. Based on this subject's relevance, this work aimed to develop a colorimetric biosensor using aptamer-functionalized gold nanoparticles for identifying P. aeruginosa. The detection mechanism is based on the color change of gold nanoparticles (AuNPs) from red to blue-purple through NaCl induction after bacteria incubation and aptamer-target binding. First, AuNPs were synthesized and characterized. The influence of aptamer and sodium chloride concentration on the agglomeration of AuNPs was investigated. Optimization of aptamer concentration and salt addition were performed. The best condition for detection was 5 µM aptamers and 200 mM of NaCl. In this case, P. aeruginosa was detected after 5 h for concentrations from 10 to 10 CFU mL, being 10 and 10 CFU mL the detection limit for color change by the naked eye and UV-Vis spectrometry, respectively. In addition, other bacteria such as E. coli, S. typhimurium, and Enterobacteriaceae bacterium were also detected with color changing from red to gray. Finally, it was confirmed that the salt incubation time can be 2 h, and that the ideal aptamer concentration is 5 µM. Thus, the colorimetric analysis can be a simple and fast detection method for P. aeruginosa in the range of 10 to 10 CFU mL to the naked eye. KEY POINTS: • A new method for rapid detection of Pseudomonas aeruginosa • Aptamers conjugated with gold nanoparticles allow pathogen detection by colorimetry • No need for previous surface modification of nanoparticles.

摘要

近年来,人们对新型快速细菌检测方法进行了研究和应用。在医院环境中,病原体感染非常普遍,会导致严重的健康问题。铜绿假单胞菌是最常见的细菌之一,可在导管和呼吸器等医院设备中生长。即使浓度很低,它也会引起严重感染,因为它对抗生素和其他治疗方法具有抗性。基于该课题的相关性,本研究旨在开发一种使用适体功能化金纳米颗粒的比色生物传感器来识别铜绿假单胞菌。检测机制基于细菌孵育和适体与靶标结合后,通过NaCl诱导金纳米颗粒(AuNPs)从红色变为蓝紫色的颜色变化。首先,合成并表征了AuNPs。研究了适体和氯化钠浓度对AuNPs团聚的影响。对适体浓度和盐添加量进行了优化。检测的最佳条件是5 μM适体和200 mM NaCl。在这种情况下,对于浓度为10⁵至10⁷ CFU/mL的铜绿假单胞菌,5小时后即可检测到,肉眼颜色变化和紫外可见光谱法的检测限分别为10⁵和10⁶ CFU/mL。此外,其他细菌如大肠杆菌、鼠伤寒沙门氏菌和肠杆菌科细菌也能检测到,颜色从红色变为灰色。最后,证实盐孵育时间可为2小时,理想的适体浓度为5 μM。因此,比色分析可以是一种简单快速的肉眼检测10⁵至10⁷ CFU/mL范围内铜绿假单胞菌的方法。要点:• 一种快速检测铜绿假单胞菌的新方法• 与金纳米颗粒共轭的适体可通过比色法检测病原体• 无需对纳米颗粒进行预先表面修饰

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