3.3 Creation of a MS
Database
Using the features of the software, create a MS database with wellidentified clearly defined strains of each species (e.g., type strains).
This database will then be used for pairwise comparison of mass
spectra of unknown isolates to allow identification (see Note 8).
Figure 2 shows normalized MSP of a representative subset of
Leptospira reference strains and isolates.
3.4 Identification
of an Unknown Isolate
Unknown isolates are prepared as described in Subheading 3.2. A
single spot is usually sufficient to obtain an identification but replicates might be preferred.
The MSP database is loaded into MALDI Biotyper 3.0 software
and used to compare the MS of the isolate with the MS database.
Using pairwise comparisons, a similarity score is calculated considering the proportion of matching peaks between the MS of the
unknown isolate and all MS in the reference database.
The similarity score ranging from 0 to 3 is interpreted as per
Bruker recommendations: a score !2.3 is considered as a valid
identification of the species, and a score in the range 2.0–2.3 is
considered as inconclusive for the species. Scores below 2.0 indicate
no reliable match to any of the MSP in the database (see Note 9).
Table 1 summarizes the recommendations for reliable identification
using identification scores.
Fig. 1 MSP-derived phylogeny of 35 species of the genus Leptospira. (Original figure published under Creative
Commons Attribution License in Thibeaux R, Girault D, Bierque E, Soupe ´ -Gilbert ME, Rettinger A, Douye ` re A,
Meyer M, Iraola G, Picardeau M, Goarant C. Biodiversity of environmental leptospira: improving identification
and revisiting the diagnosis. Front Microbiol. 2018 May 1;9:816. doi: https:/ /doi.org/10.3389/fmicb.2018.
00816)
26
Dominique Girault et al.
Database
Using the features of the software, create a MS database with wellidentified clearly defined strains of each species (e.g., type strains).
This database will then be used for pairwise comparison of mass
spectra of unknown isolates to allow identification (see Note 8).
Figure 2 shows normalized MSP of a representative subset of
Leptospira reference strains and isolates.
3.4 Identification
of an Unknown Isolate
Unknown isolates are prepared as described in Subheading 3.2. A
single spot is usually sufficient to obtain an identification but replicates might be preferred.
The MSP database is loaded into MALDI Biotyper 3.0 software
and used to compare the MS of the isolate with the MS database.
Using pairwise comparisons, a similarity score is calculated considering the proportion of matching peaks between the MS of the
unknown isolate and all MS in the reference database.
The similarity score ranging from 0 to 3 is interpreted as per
Bruker recommendations: a score !2.3 is considered as a valid
identification of the species, and a score in the range 2.0–2.3 is
considered as inconclusive for the species. Scores below 2.0 indicate
no reliable match to any of the MSP in the database (see Note 9).
Table 1 summarizes the recommendations for reliable identification
using identification scores.
Fig. 1 MSP-derived phylogeny of 35 species of the genus Leptospira. (Original figure published under Creative
Commons Attribution License in Thibeaux R, Girault D, Bierque E, Soupe ´ -Gilbert ME, Rettinger A, Douye ` re A,
Meyer M, Iraola G, Picardeau M, Goarant C. Biodiversity of environmental leptospira: improving identification
and revisiting the diagnosis. Front Microbiol. 2018 May 1;9:816. doi: https:/ /doi.org/10.3389/fmicb.2018.
00816)
26
Dominique Girault et al.