Application of PCR and Real-Time PCR for Monitoring Cyanobacteria, Microcystis spp. and Cylindrospermopsis raciborskii, in Macau Freshwater Reservoir

Weiying,; Zhang, Bo; Inchio,; Lou,; Wai,; Kin,; Ung,; Yijun,; Kong,; Kai, Kai; Meng,; Mok,

doi:10.1007/978-94-024-0933-8_5

Cited by 5 publications

(7 citation statements)

References 26 publications

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“…Among these metabolites, cyanotoxins may cause illness or death of domestic animals and humans when they are exposed to water contaminated with high cyanobacterial concentrations [ 3 ]. Microcystins (MCs) and cylindrospermopsin (CYN) are two well-known cyanotoxin groups found in the reservoirs of Taiwan [ 4 , 5 , 6 ] and the world [ 7 , 8 , 9 , 10 , 11 ]. MCs have been linked to the increased incidence of primary liver cancer [ 3 , 12 , 13 ] and acute cases of poisoning in humans and other animals [ 14 ].…”

Section: Introductionmentioning

confidence: 99%

“…To date, qPCR systems developed to monitor cyanotoxins are mostly uniplex, with limitation to a single genetic target [ 9 , 47 ]. However, toxic cyanobacterial blooms are usually complex, often involving more than one cyanotoxin [ 11 , 50 , 51 ], substantiating the importance of detecting multiple producers at the same time. Therefore, multiplex-qPCR, capable of amplifying several different targets in a single reaction well, may provide a simple approach to simultaneously detect multiple target genes.…”

Section: Introductionmentioning

confidence: 99%

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A qPCR-Based Tool to Diagnose the Presence of Harmful Cyanobacteria and Cyanotoxins in Drinking Water Sources

Chiu

Chen

Wang

et al. 2017

IJERPH

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Harmful cyanobacteria have been an important concern for drinking water quality for quite some time, as they may produce cyanotoxins and odorants. Microcystis and Cylindrospermopsis are two common harmful cyanobacterial genera detected in freshwater lakes and reservoirs, with microcystins (MCs) and cylindrospermopsin (CYN) as their important metabolites, respectively. In this study, two sets of duplex qPCR systems were developed, one for quantifying potentially-toxigenic Microcystis and Microcystis, and the other one for cylindrospermopsin-producing cyanobacteria and Cylindrospermopsis. The duplex qPCR systems were developed and validated in the laboratory by using 338 samples collected from 29 reservoirs in Taiwan and her offshore islands. Results show that cell numbers of Microcystis and Cylindorspermopsis enumerated with microscopy, and MCs and CYN concentrations measured with the enzyme-linked immuno-sorbent assay method, correlated well with their corresponding gene copies determined with the qPCR systems (range of coefficients of determination R2 = 0.392−0.740). The developed qPCR approach may serve as a useful tool for the water industry to diagnose the presence of harmful cyanobacteria and the potential presence of cyanotoxins in source waters.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

A qPCR-Based Tool to Diagnose the Presence of Harmful Cyanobacteria and Cyanotoxins in Drinking Water Sources

Chiu

Chen

Wang

et al. 2017

IJERPH

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“…Although the number of 16S rRNA gene copies differs between bacterial genomes and that difference can occasionally distort abundance profiles, the 16S rRNA gene remains as an important molecular marker for taxonomic and phylogenetic inferences since it is present in all bacteria, its mutation taxa is low, and it presents conserved regions that facilitate representations of bacterial diversity (Zhang et al 2014b). Under this light, qPCR may not necessarily quantify contaminants with precision, but it could still be considered a method for positive detection of contaminants in cyanobacterial cultures.…”

Section: Resultsmentioning

confidence: 99%

Evaluating methods for purifying cyanobacterial cultures by qPCR and high-throughput Illumina sequencing

Silva

Machineski

Alvarenga

et al. 2016

Journal of Microbiological Methods

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Cyanobacteria are commonly found in association with other microorganisms, which constitutes a great challenge during the isolation of cyanobacterial strains. Although several methods have been published for obtaining axenic cyanobacterial cultures, their efficiency is usually evaluated by observing the growth of non-cyanobacteria in culture media. In order to verify whether uncultured bacteria should be a concern during cyanobacterial isolation, this work aimed to detect by molecular methods sequences from cyanobacteria and other bacteria present before and after a technique for obtaining axenic cultures from plating and exposure of Fischerella sp. CENA161 akinetes to the Extran detergent and sodium hypochlorite. Solutions containing 0.5, 1, and 2% sodium hypochlorite were able to remove contaminant bacterial CFUs from the culture. However, qPCR pointed that the quantity of sequences amplified with universal bacteria primers was higher than the number of cyanobacteria-specific sequences before and after treatments. The presence of uncultured bacteria in post-hypochlorite cultures was confirmed by high-throughput Illumina sequencing. These results suggest that culturing may overlook the presence of uncultured bacteria associated to cyanobacterial strains and is not sufficient for monitoring the success of cyanobacterial isolation by itself. Molecular methods such as qPCR could be employed as an additional measure for evaluating axenity in cyanobacterial strains.

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“…Primers targeting the V3-V4 region of the cyanobacterial 16S rRNA gene for (Cya359F (5 0 -GGGGAATYTTCCGCAATGGG-3 0 ) and Cya781R (5 0 -GACTACWGGGGTATC-TAATCCCWTT-0 3)) (Nübel et al, 1997) were employed to quantify cyanobacterial 16S rRNA gene. Assays were performed at 64°C as described previously (Zhang et al, 2014). Primers for quantification for 18S rRNA were the same as those described above for amplicon sequencing and assays were performed at an annealing temperature of 42°C.…”

Section: Nucleic Acid Extractionmentioning

confidence: 99%

Carbon and nitrogen recycling during cyanoHABs in dreissenid-invaded and non-invaded US midwestern lakes and reservoirs

2019

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Lakes and reservoirs play key roles in global carbon cycling, especially as a carbon sink. Enrichment of nutrients in lakes and reservoirs (eutrophication) and rising global temperatures favors the proliferation of bloom-forming cyanobacteria. Harmful blooms of cyanobacteria (cyanoHABs) alter carbon and nutrient cycling in freshwater ecosystems. Some evidence suggests the introduction or establishment of invasive mussel species (i.e., Dreissena spp.) also favor cyanoHAB formation through selective filter feeding, a process through which they may also impact biogeochemical processes including carbon cycling and sequestration. However, few studies have considered the combined effects of invasive mussels and cyanoHABs on carbon and nitrogen cycling in freshwater ecosystems. Here, we examined microbial community composition and biogeochemical attributes (including carbon and nitrogen stable isotopes) in eutrophic lakes, reservoirs, and rivers in western Ohio, eastern Indiana, and northern Kentucky during the cyanobacterial bloom period of the summer of 2015. Our samples include both sites impacted by invasive mussels and those where invasive mussels have not yet been observed. Based on 16S and 18S rRNA gene sequence analysis, we found that cyanobacterial and algal communities varied across sites and were most closely related to habitat (sediment or water column sample) and site, regardless of the presence of invasive mussels or other environmental factors. However, we did find evidence that invasive mussels may influence both carbon and nitrogen cycling. While the results are based on a single time point sampling, they highlight the interactions of multiple environmental stressors in aquatic ecosystems and the critical need for more temporally intensive studies of carbon and nutrient cycling in bloom-and mussel-impacted waters.

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Application of PCR and Real-Time PCR for Monitoring Cyanobacteria, Microcystis spp. and Cylindrospermopsis raciborskii, in Macau Freshwater Reservoir

Cited by 5 publications

References 26 publications

A qPCR-Based Tool to Diagnose the Presence of Harmful Cyanobacteria and Cyanotoxins in Drinking Water Sources

A qPCR-Based Tool to Diagnose the Presence of Harmful Cyanobacteria and Cyanotoxins in Drinking Water Sources

Evaluating methods for purifying cyanobacterial cultures by qPCR and high-throughput Illumina sequencing

Carbon and nitrogen recycling during cyanoHABs in dreissenid-invaded and non-invaded US midwestern lakes and reservoirs

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