Detection of hepatotoxic Microcystis strains by PCR with intact cells from both culture and environmental samples.
Microcystins are small hepatotoxic peptides produced by a number of cyanobacteria. They are synthesized non-ribosomally by multifunctional enzyme complex synthetases encoded by the mcy genes. Primers deduced from mcy genes were designed to discriminate between toxic microcystin-producing strains and non-toxic strains. Thus, PCR-mediated detection of mcy genes could be a simple and efficient means to identify potentially harmful genotypes among cyanobacterial populations in bodies of water. We surveyed the distribution of the mcyB gene in different Microcystis strains isolated from Chinese bodies of water and confirmed that PCR can be reliably used to identify toxic strains. By omitting any DNA purification steps, the modified PCR protocol can greatly simplify the process. Cyanobacterial cells enriched from cultures, field samples, or even sediment samples could be used in the PCR assay. This method proved sensitive enough to detect mcyB genes in samples with less than 2,000 Microcystis cells per ml. Its accuracy, specificity and applicability were confirmed by sequencing selected DNA amplicons, as well as by HPLC, ELISA and mouse bioassay as controls for toxin production of every strain used.
Pan
Hui
H
Department of Phycology, Institute of Hydrobiology, The Chinese Academy of Sciences, Luojiashan, Wuhan 430072, P. R. China.
Song
Lirong
L
Liu
Yongding
Y
Börner
Thomas
T
D000818:Animals; D001427:Bacterial Toxins / Q000235:genetics / Q000302:isolation & purification; D001681:Biological Assay; D002851:Chromatography, High Pressure Liquid; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D004784:Environmental Monitoring; D004797:Enzyme-Linked Immunosorbent Assay; D007928:Lethal Dose 50; D051379:Mice; D052998:Microcystins; D046931:Microcystis / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification*; D010456:Peptides, Cyclic / Q000032:analysis* / Q000235:genetics; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D014871:Water Microbiology
D001427:Bacterial Toxins; D004269:DNA, Bacterial; D052998:Microcystins; D010456:Peptides, Cyclic; C078588:microcystin