The first evidence of bacterial TTX-producing in amphibians

SCHWARTZ, C. A., PIRES JR., O. R., SEBBEN, A.

Laboratório de Toxinologia, Departamento de Ciências Fisiológicas, Instituto de Ciências Biológicas, Universidade de Brasília, Brasília, DF, Brasil.

For marine animals is assumed an exogenous origin of TTX, many works report several bacteria genus producing TTX and related substances The same has been suggested for amphibians, although, there is no support to this hypothesis. Seven bacterial strains isolated of B. ephippium intestinal contents, were cultured in ISP-4 liquid media (for 5 days) and substituted for modified liquid media ISP-4 low phosphate concentration, Gallacher and Birkbeck, 1993 (Appl. Env. Microb. 59(11), 3981-3983) demonstrated that the stress induced for depletion of phosphate, in the culture media, increases the TTX production in marine bacteria. The cultures were centrifugated to separate bacteria from culture media. To the cells were added 100 ml of 0.1 % acetic acid. The mixture was heated to boiling and filtered. All bacteria extracts and culture media were semi-purified by a ion-exchange column (Amberlite GC-50, NH₄⁺) and active charcoal treatment (Norit-A). The semi-purified extracts were submitted to an HPLC system using a post-column fluorescent detection system (LC-FLD). Neither bacteria extracts or ISP-4 culture media showed fraction similar to TTX. Only low phosphate ISP-4 culture media presented a fraction with retention time equal to 14,793 and 14,839 minutes,  similar to of TTX standard, 14,425 minutes. Bacteria, as TTX source in amphibian was suggested by analysis in LC-FLD system, however it is not discarded the origin of this kind of compounds from diet. Although the mechanism of absorbing and accumulating this toxin or related substances remains indefinite, the identification of TTX-producing strains in intestinal microbiota in other amphibians must be done to elucidated the amount of bacteria involved in TTX production in terrestrial environment.

CORRESPONDENCE TO:

C.A. Schwartz, Laboratório de Toxinologia, Departamento de Ciências Fisiológicas – Instituto de Ciências Biológicas – Universidade de Brasília, Brasília, 70910-900, DF, Brasil, E.mail: schwartz@unb.br