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Anabaena Sp. Bloom and the Occurrence of Microcystin- Lr from a Eutrophic Pond in Bangladesh

Md. Sagir Ahmed
University of Dhaka, Department of Zoology, Advanced Fisheries and DNA Barcoding Laboratory, Dhaka 1000, Bangladesh

Bernd Luckas
University of Jena, Institute of Nutrition, Department of Food Chemistry, Dornburger Street 25,07743 Jena, Germany.

Thomas Krueger
University of Jena, Institute of Nutrition, Department of Food Chemistry, Dornburger Street 25,07743 Jena, Germany.

Abstract/ Executive Summary:

A bloom of Anabaena sp. occurred in a freshwater pond in Brahmanbaria. Bloom sample was collected and filtered through a glass fiber filter. Methanol-water extract of filtered cells were analyzed by high performance liquid chromatography (HPLC) with UV, MS and MS-MS detection detected three variants of microcystins viz, [D-Asp³, Dha⁷] Microcystin-LR, [Dha⁷] Microcystin-LR and Microcystin-LR. The total concentration of microcystins was 4.0 μg l^-1, well above the WHO provisional guideline value for drinking water. So, the cyanotoxin risk assessment has become important to protect public health in Bangladesh where surface water is used as drinking-water source.

Keywords: Anabaena, Cyanobacteria bloom, Microcystin, Bangladesh

Location: Brahmanbaria district, Bangladesh

Start Date: 01-04-2008

End Date: 14-04-2008

Program Area: Conservation and Biodiversity

Commodity: Aquatic animal

Objectives:

To describes the first report of microcystin-LR from a natural bloom of Anabaena sp. occurring in a freshwater pond.

Methodology:

The study pond was 0.74 ha in size and located in Brahmanbaria district (23⁰58' N latitude 91⁰6' E longitude) 98 km east from Dhaka. Anabaena sp. bloom was initiated in the first week of April, 2008 and the highest cell density (90% Anabaena) was recorded on 14 April 2008. A portion of the concentrated samples were filtered through a 0.45 micro m glass fiber filter (Whatman GF/C, 47 mm diameter) and dried in an oven at 60-80⁰C. Extraction: The GF/C filters were extracted with 2.0 ml of mixture water and methanol (50:50; v:v) per filter by ice-cooled sonication for 4 min with an ultrasonic probe GM 70 and subsequent treatment for 15 min in an ultrasonic bath. Extracts were centrifuged (10000 g, 15 min) and the supernatants were filtered using 0.22 micro m nylon syringe filters. The extracts were directly subjected to the liquid chromatography. Chemical analysis: The HPLC/UV determination of microcystins was carried out with some modifications C18 column: Phenomenex prodigy, ODS (3), 250 x 4.6 mm, 5 µm, mobil phases: acetonitrile /water/0.05% TFA). Detection of microcystins was done by the use of an UV detector (Shimadzu SPD-10AV; λ=238 nm). HPLC/MS and HPLC/MS-MS analysis were applied to ensure the identity of the toxin peaks in the chromatograms. Liquid Chromatography was performed with a PE Series 200 Quaternary Pump and a PE Series 200 autosampler (Perkin- Elmer, Shelton, CT, USA). The chromatographic separation was carried out on a reversed-phase column (Luna C18(2), 5 m 250 x 4.6 mm I.D., Phenomenex, Torrance) using gradient elution (0 min 20% B, 15-17 min 90% B, 18-30 min 20% B) with a flow rate of 1 mL min^-1, throughout. Mobile phases consisted of 5 mM ammonium formate and 53 mM formic acid in (A) water and (B) water-acetonitrile (10:90, v:v), respectively. The HPLC was coupled by means of an electrospray interface to a single quadrupol mass spectrometer (API 150, PE Sciex Instruments, Canada) and additionally to a triple quadrupol mass spectrometer (API 365, PE Sciex Instruments, Canada). The detection was carried out in selected ion monitoring (SIM) mode using LC/MS and multiple reactions monitoring mode (MRM) using LC/MS-MS. Quantification: Since reference materials for desmethylated microcystins are not available commercially, determination of the concentrations of desmethyl-MCs, [D-Asp³^, Dha⁷] MC-LR, and [Dha7] MC-LR, was performed using the standard calibration curves of MC-LR. This approach should be kept in mind looking at the toxin values given in this paper. Chemicals: Reference standards of Microcystin-RR, -LR, -YR, -LA, LF and –LW were purchased from Calbiochem/Novabiochem (La Jolla, CA, USA). Acetonitrile and methanol obtained from VWR were HPLC grade. Water was purified to HPLC grade quality with a Millipore-Q RG Ultra Pure Water System (Millipore, Milford, USA). All chemicals were at least analytical grade.

Source of Information: International Journal of Fisheries and Aquatic Studies 2014; 2(3): 14-17

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

Accumulation of microcystin in fish is a potentially important route of exposure for humans. The freshwater fish ?Oreochromis niloticus accumulates microcystins in the guts, liver and kidneys but there are fish that can also accumulate these toxins in the muscle tissue, posing high risks to humans that consume contaminated fish. So, the effect of cyanobacteria blooms (microcystins) on aquatic animals and human through direct exposure or food chain in Bangladesh waters remains to be identified.

Knowledge Management: Journal