5 g of soil 5 mL of PBS
A
B
C
D
E
Mixing
Sedimentation
5 min
30°C
Daily check until growth
(5-20 days)
(0.22
µm filtration if
contamination persists)
30°C
5 mL EMJH
(5-20 days)
+2 mL of supernatant
+2.5 mL of 2X EMJH
+0.5 mL of 10X STAFF
filtration 0.45
µm
100
µL of positive EMJH
culture
(1:100 & 1:1000 dilution)
Solid EMJH plate
Single colonies
isolation
16S molecular
typing and
MALDI-ToF
identification
Fig. 1
Leptospira isolation on EMJH agar plates. (a) Global isolation strategy. (b) Macroscopic aspect of colonies. (c) Closer view from (a) showing colonies of
different sizes. (d) Opaque (yellow arrow) and transparent (red arrow) colonies. (e) Translucent colonies. (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)
Leptospira Isolation and Culture
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