Bacterial fingerprint

Woolfitt, A. Moura, H. Barr, J. De, B. Popovic,T. Satten, G. Jarman, K. H. Wahl, K. L., Differentiation of Bacillus spp. by MALDI-TOF mass spectrometry using a bacterial fingerprinting algorithm and a random forest classification algorithm. Presented at 5th ISIAM Meeting, Richland, WA 2004. 

Figure 12.9 Colorimetric bacterial fingerprinting. The color combination for each bacterium reflects the chromatic transitions (RCS) recorded 7 h after the start of growth at a bacterial concentration of 1 x lO /niL. The RCS color key is shown on the left (i) 1-a-dioleoylpho-sphatidylethanolamine (DOPE)/PDA (1 9 mole ratio), (ii) sphingomyelin/cholesterol/PDA (7 3 90), (iii) DMPC/PDA (1 9), and (iv) l-palmitoyl-2-oleoyl-j n-glycero-3-[phospho-rac-(l-glycerol)] (POPG)/PDA (1 9). Reprinted from Scindia et al. (2007). Copyright 2007 American Chemical Society. (See color insert.)...

Ideally, scientists would like to be able to perform laser desorption and analysis directly, but typical laser wavelengths cause fragmentation of bacteria and other particles. Due to the low energy produced by infrared lasers, however, bacterial fingerprints can indeed be obtained as shown by researchers at Lawrence Livermore National Laboratory. It is also possible to detect much larger species, an impossible task with earlier technology. Infrared laser desorption techniques are undergoing constant improvement. 

A good reference on the flow cytometry of DNA fragments is Kim Y, Jett JH, Larson EJ, et al. (1999). Bacterial fingerprinting by flow cytometry Bacterial species discrimination. Cytometry 36 324-332. The proposed technique for sequencing DNA is described in a paper by Jett JH, Keller RA, Martin JC, et al. (1989). High-speed DNA sequencing An approach based upon fluorescence detection of single molecules. J. Biomol. Struct. Dynamics 7 301-309. 

Fig. 11.14. Reprinted with permission of John Wiley Sons, Inc. 1999 from Kim Y, et al. (1999). Bacterial fingerprinting by flow cytometry bacterial species discrimination. Cytometry 36 324-332.

Py-MS is commonly applied for bacterial fingerprinting, i.e. classification of characteristic signal patterns followed the first reports published in the 1960s on the applicability of analytical pyrolysis techniques to clinical and pharmaceutical microbiology. Application of Py-MS as an independent tool for the char-... 

J. Versalovic, T. Koeuth, and J. R, Lupski, Distribution of repetitive DNA sequences in eubacteria and application to fingerprinting of bacterial genomes. Nucleic Acid Re.K. 79 6832 (1991). 

Waters Corporation has now introduced the MicrobeLynx , a MALDI-TOF MS system developed for fingerprint bacterial ID. The system and its applications are described thus ... 

Variations on the spectral peaks from different species of the same genus were also observed. Three species of Pseudomonas produced the spectra shown in Figure 14.2. These spectra are clearly unique and were used to correctly identify unknown samples. Because of peak ratio reproducibility issues in bacterial protein profiles obtained by MALDI MS,11 a fingerprint approach that had been used for other mass spectrometry approaches has not been used. The profile reproducibility problem was first recognized by Reilly et al.12,13 and later researched by others in the field.14,15 As a later alternative, a direct comparison of the mass-to-charge ratio (m/z) of the unknown mass spectral peaks with a database of known protein masses has been used to identify unknown samples.14... 

Electrospray (ESI) ionization mass spectrometry also plays in important role in bacterial characterization. Because it typically includes a chromatographic separation step, the approach is not considered as rapid as MALDI approaches, which do not incorporate a separation. However, compared to the times needed to grow bacteria in culture prior to analysis, the time frame is not lengthy, and the addition of chromatographic separation provides many opportunities to increase specificity. ESI/MS has been used to characterize cellular biomarkers for metabolic, genomic, and proteomics fingerprinting of bacteria, and these approaches are reported in two chapters. 

The i-poly(3HB) depolymerase of R. rubrum is the only i-poly(3HB) depolymerase that has been purified [174]. The enzyme consists of one polypeptide of 30-32 kDa and has a pH and temperature optimum of pH 9 and 55 °C, respectively. A specific activity of 4 mmol released 3-hydroxybutyrate/min x mg protein was determined (at 45 °C). The purified enzyme was inactive with denatured poly(3HB) and had no lipase-, protease-, or esterase activity with p-nitro-phenyl fatty acid esters (2-8 carbon atoms). Native poly(3HO) granules were not hydrolyzed by i-poly(3HB) depolymerase, indicating a high substrate specificity similar to extracellular poly(3HB) depolymerases. Recently, the DNA sequence of the i-poly(3HB) depolymerase of R. eutropha was published (AB07612). Surprisingly, the DNA-deduced amino acid sequence (47.3 kDa) did not contain a lipase box fingerprint. A more detailed investigation of the structure and function of bacterial i-poly(HA) depolymerases will be necessary in future. 

Yang CH, Crowley DE, Menge JA (2001) 16S rDNA fingerprinting of rhizosphere bacterial communities associated with healthy and phytophthora infected avocado roots. FEMS Microbiol Ecol 35 129-136 Yemefack M, Jetten VG, Rossiter DG (2006) Developing a minimum data set for characterizing soil dynamics in shifting cultivation systems. Soil Tillage Res 86 84-98... 

R.T. Vinopal, J.R. Jadamec, P. deFur, A.L. Demars, S. Jakubielski, C. Green, C.P. Anderson and J.E.D.R.F. Dugas, Fingerprinting bacterial strains using ion mobility spectrometry, Anal. Chim. Acta, 457 (2005) 83-95. 

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