New method quickly detects toxins produced by bacteria in water

A technique applied at Unesp using mass spectrometry identifies toxic compounds directly in cyanobacterial cells and in water reservoirs

By Elton Alisson

Agência FAPESP – Certain species of bacteria that occur in aquatic environments are photosynthetic. Because they contain chlorophyll and other pigments, these are frequently confused with microalgae.

However, unlike the tiny microalgae, which have many industrial applications, these bacteria, called cyanobacteria, produce toxins (microcystins and nodularins) that are highly harmful to human health. These toxins must be detected, identified and quantified rapidly, especially in water reservoirs, to avoid serious health risks.

To meet the demand for these tests in Brazil, researchers at the Universidade Estadual Paulista (Unesp) Biosciences Institute in Rio Claro, SP, are developing rapid and sensitive methods to detect cyanotoxins directly from cyanobacterial cells and in water reservoirs through mass spectrometry.

Results from the research project, conducted with FAPESP funding under the Foundation’s Young Researchers in Emerging Centers Program, were presented at the 4th BrMASS Conference, held by the Brazilian Mass Spectrometry Society from December 10-13 in Campinas, SP.

According to project coordinator Humberto Márcio Santos Milagre, one of the goals of the study was to verify whether it was possible to identify microcystin variations through the use of thin layer chromatography (TLC) together with the MALDI ionization method.

The chromatographic technique is considered one of the most simple and economical for the separation and visual identification of the components of a mixture. However, the unequivocal identification of compounds is not possible through traditional methods of detection and comparison with reference patterns.

For this reason, TLC has been combined with mass spectrometry to show the components of complex mixtures and identify these compounds.

Using the TLC and MALDI combination, the Unesp researchers managed to identify and characterize microcystins in commercial samples of bacteria and in different water samples from the Billings Reservoir in São Paulo following the proliferation of cyanobacteria. They were able to prove the technique’s effectiveness in detecting the toxins produced by the cyanobacteria.
 
 “Our expectation is to use the technique for environmental analyses,” said Milagre. Following the initial study using this technique, the group began identifying cyanotoxins directly from cyanobacterial cells.

Using strains of two toxin-producing bacteria obtained from the Pasteur Institute and grown in bioreactors at the Unesp Rio Claro Mass Spectrometry Laboratory, the researchers identified microcystins and nodularins a few days after the cyanobacteria were placed in the culture.

“These bacteria proliferate quickly under favorable conditions and, in this process, produce a large quantity of metabolites and microcystins, which are difficult to degrade and highly toxic,” said Milagre.
 
Milagre remembers one of the greatest tragedies in Brazil caused by microcystins, which occurred in Caruaru, Pernambuco. In 1996, 60 patients undergoing hemodialysis at a clinic in the northeastern city died of microcystin hepatotoxin poisoning from water used in the medical treatment that was collected by a water truck from a contaminated reservoir.

Medal

At the opening of the 4th BrMASS Conference, FAPESP’s scientific director, Carlos Henrique de Brito Cruz, received the BrMASS medal for his work with FAPESP for the promotion of science, technology and the field of mass spectrometry in Brazil.