for publication purposes, we recommend employing the more
precise and time-consuming maximum likelihood method, which
is not a part of the BIGSdb function. Additional fields can be
selected to be included as metadata for use in coloring nodes—
select any fields you wish to include in the “iTOL datasets” list. For
detailed explanation of the iTOL function, see the following link:
https://itol.embl.de/help.cgi.
3.3.3 Species
Identification
If you are not sure which species of Leptospira you are working
with, you can check using iTOL plug-in by generating the phylogenetic tree based on the concatenated core gene sequences of your
unknown sample(s) together with the reference sequences of Leptospira species (n ¼ 64) [6] which are present in BIGSdb (Table 1
and Fig. 2). Selecting the BIGSdb IDs of reference strains and your
unknown sample(s) in the iTOL plugin will generate a phylogenetic
tree which will cluster your isolates together with one of the reference strains. If the clustering is not clear, the average nucleotide
identity (ANI) of draft genomes should be performed, for example,
using the ANI calculator [14] at the following link: https://www.
ezbiocloud.net/tools/ani.
3.3.4 Prediction
of Possible Serogroup
Strains belonging to the same serogroups are usually subdivided
into several cgCGs; however, when strains are part of the same
clonal group, they should belong to the same serogoup (based on
the all available isolates at the time of writing, n ¼ 1007); i.e., the
branching based on the concatenated cgMLST sequences could be
useful for determination of the potential serogroup.
3.3.5 GrapeTree
GrapeTree allows for exploration of the fine-grained population
structure and phenotypic properties of large number of genomes
(more than 200) in a web browser. It generates and displays the
minimum spanning tree (MSTree) based on cgSTs [9]. The GrapeTree algorithm is able to export large datasets and is compatible to
handle larger amount of missing sequences and thus is perfect for
handling cgMLST data. The datasets can include metadata, which
allows nodes in the result tree to be colored interactively. It can be
accessed from the contents page or following the query by clicking
the “GrapeTree” link. In the Leptospira setting, it could be very
useful in an easy identification of the potential source of infection
by determination of cgCGs which are shared among human and
animal isolates (Fig. 3). In Fig. 3, it is evident that several cgCGs are
unique to particular hosts (e.g., cgCG176 was found only in dogs
and cgCG81 was found only in patients), while other cgCGs were
shared among multiple hosts (e.g., cgCG6 was shared among
humans, dogs, and rats and cgCG5 was found in humans, cows,
hedgehogs, dogs, and other mammals). Another example of how
GrapeTree can be used in Leptospira molecular epidemiology is the
tracking of the different distributions of CGs over a specific time
period.
cgMLST Scheme for Leptospira
15
precise and time-consuming maximum likelihood method, which
is not a part of the BIGSdb function. Additional fields can be
selected to be included as metadata for use in coloring nodes—
select any fields you wish to include in the “iTOL datasets” list. For
detailed explanation of the iTOL function, see the following link:
https://itol.embl.de/help.cgi.
3.3.3 Species
Identification
If you are not sure which species of Leptospira you are working
with, you can check using iTOL plug-in by generating the phylogenetic tree based on the concatenated core gene sequences of your
unknown sample(s) together with the reference sequences of Leptospira species (n ¼ 64) [6] which are present in BIGSdb (Table 1
and Fig. 2). Selecting the BIGSdb IDs of reference strains and your
unknown sample(s) in the iTOL plugin will generate a phylogenetic
tree which will cluster your isolates together with one of the reference strains. If the clustering is not clear, the average nucleotide
identity (ANI) of draft genomes should be performed, for example,
using the ANI calculator [14] at the following link: https://www.
ezbiocloud.net/tools/ani.
3.3.4 Prediction
of Possible Serogroup
Strains belonging to the same serogroups are usually subdivided
into several cgCGs; however, when strains are part of the same
clonal group, they should belong to the same serogoup (based on
the all available isolates at the time of writing, n ¼ 1007); i.e., the
branching based on the concatenated cgMLST sequences could be
useful for determination of the potential serogroup.
3.3.5 GrapeTree
GrapeTree allows for exploration of the fine-grained population
structure and phenotypic properties of large number of genomes
(more than 200) in a web browser. It generates and displays the
minimum spanning tree (MSTree) based on cgSTs [9]. The GrapeTree algorithm is able to export large datasets and is compatible to
handle larger amount of missing sequences and thus is perfect for
handling cgMLST data. The datasets can include metadata, which
allows nodes in the result tree to be colored interactively. It can be
accessed from the contents page or following the query by clicking
the “GrapeTree” link. In the Leptospira setting, it could be very
useful in an easy identification of the potential source of infection
by determination of cgCGs which are shared among human and
animal isolates (Fig. 3). In Fig. 3, it is evident that several cgCGs are
unique to particular hosts (e.g., cgCG176 was found only in dogs
and cgCG81 was found only in patients), while other cgCGs were
shared among multiple hosts (e.g., cgCG6 was shared among
humans, dogs, and rats and cgCG5 was found in humans, cows,
hedgehogs, dogs, and other mammals). Another example of how
GrapeTree can be used in Leptospira molecular epidemiology is the
tracking of the different distributions of CGs over a specific time
period.
cgMLST Scheme for Leptospira
15