RESUMEN
The cyanobacterial cytotoxin cylindrospermopsin (CYN) has become increasingly common in fresh waters worldwide. It was originally isolated from Cylindrospermopsis raciborskii in Australia; however, in European waters, its occurrence is associated with other cyanobacterial species belonging to the genera Aphanizomenon and Anabaena. Moreover, cylindrospermopsin-producing strains of widely distributed C. raciborskii have not yet been observed in European waters. The aims of this work were to assess the occurrence of CYN in lakes of western Poland and to identify the CYN producers. The ELISA tests, high-performance liquid chromatography (HPLC)-DAD, and HPLC-mass spectrometry (MS)/MS were conducted to assess the occurrence of CYN in 36 lakes. The cyrJ, cyrA, and pks genes were amplified to identify toxigenic genotypes of cyanobacteria that are capable of producing CYN. The toxicity and toxigenicity of the C. raciborskii and Aphanizomenon gracile strains isolated from the studied lakes were examined. Overall, CYN was detected in 13 lakes using HPLC-MS/MS, and its concentrations varied from trace levels to 3.0 µg L(-1). CYN was widely observed in lakes of western Poland during the whole summer under different environmental conditions. Mineral forms of nutrients and temperature were related to CYN production. The molecular studies confirmed the presence of toxigenic cyanobacterial populations in all of the samples where CYN was detected. The toxicity and toxigenicity analyses of isolated cyanobacteria strains revealed that A. gracile was the major producer of CYN.
Asunto(s)
Aphanizomenon/aislamiento & purificación , Lagos/microbiología , Uracilo/análogos & derivados , Contaminantes del Agua/análisis , Alcaloides , Anabaena/genética , Anabaena/aislamiento & purificación , Aphanizomenon/genética , Aphanizomenon/metabolismo , Toxinas Bacterianas , Toxinas de Cianobacterias , Cylindrospermopsis/genética , Cylindrospermopsis/aislamiento & purificación , Monitoreo del Ambiente , Genes Bacterianos/genética , Fitoplancton/genética , Fitoplancton/aislamiento & purificación , Fitoplancton/metabolismo , Polonia , Análisis de Secuencia de ADN , Temperatura , Uracilo/análisis , Uracilo/biosíntesis , Microbiología del Agua , Contaminantes del Agua/metabolismoRESUMEN
Blooms of toxic cyanobacteria, associated with illness and mortality in humans and animals, are becoming increasingly common worldwide. The safe use of surface waters for drinking water production and recreation necessitates assessment of toxigenic cyanobacteria. We have developed simple and reliable sample preparation and qPCR methods to detect microcystin-producing strains of three major bloom-forming genera, Anabaena, Microcystis and Planktothrix. The mcyB second thiolation motif, previously not recognized as a potential target for qPCR, was used as a basis for primer and genus-specific probe design. Assay specificity and sensitivity was confirmed with cultured cyanobacterial strains and the effect of different sample preparation methods on quantification was investigated. Sample filtration and cell lysis reduced assay time and resulted in more efficient amplification compared to DNA extraction. Positive correlation (p<0.005) between mcyB copy numbers and microcystin concentrations was observed in environmental samples. The results encourage the use of qPCR in water risk management.