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Real time human micro-organisms biotyping based on Water-Assisted Laser Desorption/Ionization

Benoit Fatou

Benoit Fatou

Université de Lille, Laboratoire Protéomique, Réponse Inflammatoire et Spectrométrie de Masse (PRISM)Villeneuve d’Ascq Cedex, France
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Fatou, Benoit
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Michel Salzet

Michel Salzet

Université de Lille, Laboratoire Protéomique, Réponse Inflammatoire et Spectrométrie de Masse (PRISM)Villeneuve d’Ascq Cedex, France
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Salzet, Michel
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Isabelle Fournier

Isabelle Fournier

Université de Lille, Laboratoire Protéomique, Réponse Inflammatoire et Spectrométrie de Masse (PRISM)Villeneuve d’Ascq Cedex, France
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Fournier, Isabelle
  
Apr 24, 2019
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Article Category: Research Article
Published Online: Apr 24, 2019
Page range: 97 - 104
DOI: https://doi.org/10.2478/ebtj-2019-0011
Keywords
© 2019 Benoit Fatou, Michel Salzet, Isabelle Fournier, published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 License.

Figure 1

Schematic representation of the tests proceed for pathogen identification using SpiderMass technology. 1) E. Coli were cultivated in poor broth medium. Microbial growth was assessed by the increase in A 620 nm after 8-hour incubation at 37°C . At different time of bacteria growth, 10 μl of the suspension are harvested and dropped either on agarose of a petri dish or smeared on glass slides. Direct analyses with SpiderMass were then conducted directly on the top of petri dish or smear. Direct m/z spectra profiles are then recorded.
Schematic representation of the tests proceed for pathogen identification using SpiderMass technology. 1) E. Coli were cultivated in poor broth medium. Microbial growth was assessed by the increase in A 620 nm after 8-hour incubation at 37°C . At different time of bacteria growth, 10 μl of the suspension are harvested and dropped either on agarose of a petri dish or smeared on glass slides. Direct analyses with SpiderMass were then conducted directly on the top of petri dish or smear. Direct m/z spectra profiles are then recorded.

Figure 2

Time course SpiderMass analyses from T0 to T24 hours of E. Coli growths in liquid suspension were performed on agarose of a petri dish. I 10 μl of the suspension collected at T0 and at T24h are harvested and dropped on agarose of a petri dish SpiderMass analyses were performed in negative and positive modes. (Duplicates are presented for each conditions).
Time course SpiderMass analyses from T0 to T24 hours of E. Coli growths in liquid suspension were performed on agarose of a petri dish. I 10 μl of the suspension collected at T0 and at T24h are harvested and dropped on agarose of a petri dish SpiderMass analyses were performed in negative and positive modes. (Duplicates are presented for each conditions).

Figure 3

Comparison of SpiderMass spectra A) collected either from culture medium directly , agarose (petri dish) and on smears on glass slides for bacteria collected at T24, B) represent the PCA analyses in negative mode of the spectra collected from petri dish a); Smear b) and Comparison between direct medium, agarose and smear by PCA in negative and positive modes c). C) Highlights of specific common ions detected in positive mode ranged from 549.432 to 564.453 from the 3 methods.
Comparison of SpiderMass spectra A) collected either from culture medium directly , agarose (petri dish) and on smears on glass slides for bacteria collected at T24, B) represent the PCA analyses in negative mode of the spectra collected from petri dish a); Smear b) and Comparison between direct medium, agarose and smear by PCA in negative and positive modes c). C) Highlights of specific common ions detected in positive mode ranged from 549.432 to 564.453 from the 3 methods.

Figure 4

Reproducibility experiments were conducted on 3 different slides containing smears of bacteria collected at T24. Experiments were realized on the same slides analyzed 3 times during 3 consecutive days in negative A) and positive B) modes.
Reproducibility experiments were conducted on 3 different slides containing smears of bacteria collected at T24. Experiments were realized on the same slides analyzed 3 times during 3 consecutive days in negative A) and positive B) modes.

Figure 5

Biotyping studies in negative mode on smear of different species of pathogens i.e Gram negative (E. coli D31, Pseudomonas aeruginosa); Gram positive (Staphylococcus aureus, Enterococcus faecalis) and Yeast (Candida Albicans).
Biotyping studies in negative mode on smear of different species of pathogens i.e Gram negative (E. coli D31, Pseudomonas aeruginosa); Gram positive (Staphylococcus aureus, Enterococcus faecalis) and Yeast (Candida Albicans).

Figure 6

MS/MS analyses directly by SpiderMass in real time of selected ions from E. Coli corresponding to : m/z 688.35 to PE (16:0/cy17:0), m/z of m/z 719.35 to PG (16:0/16:1) and m/z 733.36 to PG (16:0/cys17:0), 773.38 to PG (18:1/18:1).
MS/MS analyses directly by SpiderMass in real time of selected ions from E. Coli corresponding to : m/z 688.35 to PE (16:0/cy17:0), m/z of m/z 719.35 to PG (16:0/16:1) and m/z 733.36 to PG (16:0/cys17:0), 773.38 to PG (18:1/18:1).
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