Mass Analyzers and MS/MS Methods for Microbial Detection and Identification

2013 ◽  
pp. 11-38
Author(s):  
Michael Stanford
Keyword(s):  
Microbial Detection ◽  
Detection And Identification

scholarly journals Guidelines for Optimisation of a Multiplex Oligonucleotide Ligation-PCR for Characterisation of Microbial Pathogens in a Microsphere Suspension Array

2015 ◽  
Vol 2015 ◽  
pp. 1-10 ◽  
Author(s):  
Véronique Wuyts ◽  
Nancy H. C. Roosens ◽  
Sophie Bertrand ◽  
Kathleen Marchal ◽  
Sigrid C. J. De Keersmaecker
Keyword(s):  
Dna Isolation ◽  
Bacterial Species ◽  
Microbial Pathogens ◽  
Pcr Assay ◽  
Microbial Detection ◽  
Suspension Array ◽  
Negative Results ◽  
Detection And Identification ◽  
Probe Concentration ◽  
Ligation Step

With multiplex oligonucleotide ligation-PCR (MOL-PCR) different molecular markers can be simultaneously analysed in a single assay and high levels of multiplexing can be achieved in high-throughput format. As such, MOL-PCR is a convenient solution for microbial detection and identification assays where many markers should be analysed, including for routine further characterisation of an identified microbial pathogenic isolate. For an assay aimed at routine use, optimisation in terms of differentiation between positive and negative results and of cost and effort is indispensable. As MOL-PCR includes a multiplex ligation step, followed by a singleplex PCR and analysis with microspheres on a Luminex device, several parameters are accessible for optimisation. Although MOL-PCR performance may be influenced by the markers used in the assay and the targeted bacterial species, evaluation of the method of DNA isolation, the probe concentration, the amount of microspheres, and the concentration of reporter dye is advisable in the development of any MOL-PCR assay. Therefore, we here describe our observations made during the optimisation of a 20-plex MOL-PCR assay for subtyping ofSalmonellaTyphimurium with the aim to provide a possible workflow as guidance for the development and optimisation of a MOL-PCR assay for the characterisation of other microbial pathogens.

scholarly journals Preprototype of an Automated Microbial Detection and Identification System: a Developmental Investigation

1977 ◽  
Vol 6 (4) ◽  
pp. 400-405
Author(s):  
Alex C. Sonnenwirth
Keyword(s):  
Species Group ◽  
Identification System ◽  
Culture Methods ◽  
Microbial Detection ◽  
System A ◽  
Detection And Identification ◽  
Or Groups ◽  
Colony Forming Units ◽  
Identification Of Bacteria ◽  
System 1

The AutoMicrobic System is an automated, computerized instrument that uses highly selective media and an optical system for detection, enumeration, and identification of bacteria and some yeasts in 13 h. A preprototype instrument (AutoMicrobic System-1) and its urine culture kit (Identi-Pak), developed for the detection, enumeration, and identification of eight species or groups of bacteria and of Candida species and Torulopsis glabrata in urine specimens, was evaluated during its development. An overall agreement of approximately 90% between the preprototype instrument and conventional (manual) culture methods has been obtained both with 1,473 seeded (simulated) and 1,688 clinical (mono- or polymicrobial) specimens containing 70,000 (or more) colony-forming units per ml of Escherichia coli, Klebsiella-Enterobacter species, Proteus species, Citrobacter freundii, Serratia species, group D enterococci, or yeasts ( Candida species and T. glabrata ). Lower agreements in identification were obtained with Pseudomonas aeruginosa -containing (average of 75% in clinical specimens) and Staphylococcus aureus -containing (76%) specimens. Comparison of specimens tested simultaneously in two preprototype systems resulted in ⋜4% disagreement; true negativity agreements in all specimen groups tested were at least 94%. Among problems remaining are adaptation of system for specimens other than urine, improvement of sensitivity for P. aeruginosa and S. aureus , and standardization of manual methods used for comparison and validation.

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