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MALDI-TOF MS as a method for rapid identification of Phytophthora de Bary, 1876.

ABSTRACT: The number of described species of the oomycete genus Phytophthora is growing rapidly, highlighting the need for low-cost, rapid tools for species identification. Here, a collection of 24 Phytophthora species (42 samples) from natural as well as anthropogenic habitats were genetically identified using the internal transcribed spacer (ITS) and cytochrome c oxidase subunit I (COI) regions. Because genetic identification is time consuming, we have created a complementary method based on by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Both methods were compared and hypothesis that the MALDI-TOF MS method can be a fast and reliable method for the identification of oomycetes was confirmed. Over 3500 mass spectra were acquired, manually reviewed for quality control, and consolidated into a single reference library using the Bruker MALDI Biotyper platform. Finally, a database containing 144 main spectra (MSPs) was created and published in repository. The method presented in this study will facilitate the use of MALDI-TOF MS as a complement to existing approaches for fast, reliable identification of Phytophthora isolates.

SUBMITTER: Bozik M

PROVIDER: S-EPMC8297470 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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MALDI-TOF MS as a method for rapid identification of Phytophthora de Bary, 1876.

Božik Matěj M Mrázková Marcela M Novotná Karolína K Hrabětová Markéta M Maršik Petr P Klouček Pavel P Černý Karel K

PeerJ 20210719

The number of described species of the oomycete genus Phytophthora is growing rapidly, highlighting the need for low-cost, rapid tools for species identification. Here, a collection of 24 Phytophthora species (42 samples) from natural as well as anthropogenic habitats were genetically identified using the internal transcribed spacer (ITS) and cytochrome c oxidase subunit I (COI) regions. Because genetic identification is time consuming, we have created a complementary ...[more]

PMID: 34322319

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Project description:BackgroundIn the last decade, an innovative approach has emerged for arthropod identification based on matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Increasing interest in applying the original technique for arthropod identification has led to the development of a variety of procedures for sample preparation and selection of body parts, among others. However, the absence of a consensual strategy hampers direct inter-study comparisons. Moreover, these different procedures are confusing to new users. Establishing optimized procedures and standardized protocols for mosquito identification by MALDI-TOF MS is therefore a necessity, and would notably enable the sharing of reference MS databases. Here, we assess the optimal conditions for mosquito identification using MALDI-TOF MS profiling.MethodsThree homogenization methods, two of which were manual and one automatic, were used on three distinct body parts (legs, thorax, head) of two mosquito laboratory strains, Anopheles coluzzii and Aedes aegypti, and the results evaluated. The reproducibility of MS profiles, identification rate with relevant scores and the suitability of procedures for high-throughput analyses were the main criteria for establishing optimized guidelines. Additionally, the consequences of blood-feeding and geographical origin were evaluated using both laboratory strains and field-collected mosquitoes.ResultsRelevant score values for mosquito identification were obtained for all the three body parts assayed using MALDI-TOF MS profiling; however, the thorax and legs were the most suitable specimens, independently of homogenization method or species. Although the manual homogenization methods were associated with a high rate of identification on the three body parts, this homogenization mode is not adaptable to the processing of a large number of samples. Therefore, the automatic homogenization procedure was selected as the reference homogenization method. Blood-feeding status did not hamper the identification of mosquito species, despite the presence of MS peaks from original blood in the MS profiles of the three body parts tested from both species. Finally, a significant improvement in identification scores was obtained for field-collected specimens when MS spectra of species from the same geographical area were added to the database.ConclusionThe results of the current study establish guidelines for the selection of mosquito anatomic parts and modality of sample preparation (e.g. homogenization) for future specimen identification by MALDI-TOF MS profiling. These standardized operational protocols could be used as references for creating an international MS database.

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MALDI-TOF MS as a method for rapid identification of Phytophthora de Bary, 1876.

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MALDI-TOF MS as a method for rapid identification of <i><i>Phytophthora</i></i> de Bary, 1876.

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