Assessment of bacterial endospore viability with fluorescent dyes.
OBJECTIVE
To validate three fluorescence viability assays designed primarily for vegetative cells on pure Bacillus endospores.


RESULTS
Purified fresh and gamma-irradiated Bacillus endospores (Bacillus cereus, B. coagulans and two strains of B. subtilis) were used. The viability assays were: 5-cyano-2,3-diotolyl tetrazolium chloride (CTC) to test respiratory activity and early germination, DiBAC4(3) and Live/Dead BacLight to measure membrane energization and permeabilization, respectively. Gamma irradiation treatment completely eliminated spore culturability and was used as negative control. The untreated spores showed respiratory activity after 1 h of incubation and this was characteristic of almost 100% of spores after 24 h. The membrane potential assessment gave no answer about spore viability. A lower proportion of untreated spores had permeabilized membrane compared with gamma-irradiated spores using Live/Dead BacLight (P < 0.02).


CONCLUSIONS
It is possible to use CTC and Live/Dead BacLight to rapidly test endospore viability and evaluate the proportion of spores in a preparation that could not be recovered with plate count.


CONCLUSIONS
This study shows that fluorescence tests could be applied to assess viability in potentially pathogenic Bacillus spore preparations within 1 h.
Laflamme
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C
Institut Universitaire de cardiologie et de pneumologie, Hôpital Laval, Université Laval, Québec, Québec, Canada.
Lavigne
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Ho
J
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Duchaine
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D001407:Bacillus / Q000502:physiology* / Q000648:ultrastructure; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D007091:Image Processing, Computer-Assisted; D008856:Microscopy, Fluorescence; D013171:Spores, Bacterial / Q000648:ultrastructure
D005456:Fluorescent Dyes