Oscillatoria sp. Bloom and the Occurrence of Microcystin in the River Buriganga, Dhaka, Bangladesh
Md. Sagir Ahmed,
Potentially toxic cyanobacterial blooms are becoming common in the freshwater reservoirs in all regions of Bangladesh. The River Buriganga is a eutrophic urban river in the country and its water is utilized as drinking water supply and other recreational purposes. A bloom of Oscillatoria sp. occurred in the river Buriganga during May 2002. 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, detection and Mass Spectrum (MS) detected Microcystin-RR. The concentration of microcystin RR was 0.235 μg L-1. As this river water is supplied to city dweller for drinking and other domestic usage there should have a regular monitoring system for cyanobateria blooms and microcystins to ensure the public health safety. To avoid the microcystins health risk to humans through drinking water, it is advised not to use this river water for drinking or any other domestic purposes until the provisional safety levels of microcystins are reported through the concerned authority.
Eutrophication is an increasing problem for aquatic ecosystems in Bangladesh,
as in many other countries around the world. Cyanobacterial blooms are very
frequent episodes in our reservoirs, most of which are eutrophic or hypereutrophic.
This scenario is widespread in all regions of the country, partially as a consequence
of favorable climatic conditions, but mainly due to the over-enrichment of rivers,
reservoirs and estuaries. The production and release of cyanotoxins is often
associated with these cyanobacteria blooms (Ahmed, 2009;
Ahmed et al., 2008a, b).
Most of the toxic cyanobacteria genera have been recognized to produce a range
of hepatotoxic toxins- microcystins (Carmichael et al.,
1988). In recent years, for a variety of reasons, the harmful impact of
cyanobacteria on human health has been more reported by Figueiredo
et al. (2004). The most dramatic example is the human poisoning case
attributed to direct exposure to high concentrations of microcystins which occurred
in Brazil in 1996 (Jochimsen et al., 1998). Concern
about the microcystins health risk to humans through drinking water, led the
World Health Organization (WHO) to develop and suggest a provisional guideline
level of microcystin-LR at 1 μg L-1. Up to now this value has
been considered as a safe level in drinking water (Falconer
et al., 1999).
The supply of clean and safe drinking water is one of the main challenges of
public health care in Bangladesh. Traditionally, surface water is the main source
of drinking water and consumed without any treatment or after boiling when fuel
is available. Resulting, about 4.5 million people suffered from watery diarrhea
every year, among them a considerable number affected with cholera (Siddique
et al., 1996). In many of these intensively used eutrophic ponds/lakes
cyanobacteria blooms are common and microcystins have been detected occasionally
in pond water from several regions, mainly associated with high abundance of
Microcystis sp. (Ahmed, 2009; Ahmed
et al., 2008a, b; Aziz,
1974; Welker et al., 2005).
Bangladesh has about 700 rivers including tributaries and distributaries. Almost
all settlements of the country are established besides river. The Buriganga,
a tide-influenced river is passing through West and South of the capital city,
Dhaka. About 1.2 million, peoples live in both side of the river and use its
water for washing, bathing and other domestic purposes. Water of this river
is also supplied to a portion of city dwellers through Chandi Ghat water treatment
plant. Recently, there has been an increased use of surface water for human
consumption due to arsenic contamination in ground waters. In the case of Dhaka
district and perhaps other regions of Bangladesh such a practice could amount
of replacing one health hazard with another. Nevertheless, studies dealing with
toxins and toxicology of cyanobacteria in Bangladesh waters are not very abundant
(Ahmed, 2009; Ahmed et al.,
2008a, b; Welker et al., 2005; Ahmed
et al., 2007).
This study deals with isolation and characterization of microcystins from a natural bloom of Oscillatoria sp. occurring in the river Buriganga, Dhaka.
MATERIALS AND METHODS
The river Buriganga encompasses the West and Southern periphery of Dhaka, the capital city of Bangladesh. It is considered as one of the most polluted urban river in the country. The present study was conducted in the upper reaches of this river (Islambagh zone) from January to December 2002.
Collection and Processing of Samples
Samples of water and phytoplankton were collected fortnightly intervals
throughout the study period. The plankton samples were collected using plankton
net of 20 μm mesh size and kept in 20 mL glass bottle. All samples were
preserved and transported to the laboratory for further analysis. Physico-chemical
parameters were recorded by using an ecological HACH fresh water, aquaculture
ecological manual, test kit model FF-2. Qualitative and quantitative analysis
of phytoplankton were performed using taxonomic keys following Ward
and Whipple (1959) and Needham and Needham (1966)
methods by Welch (1952).
A bloom of Oscillatoria sp. was initiated in the first week of May 2002 and the highest cell density (80% Oscillatoria) was recorded on 20 May 2002. Three samples were collected with a plankton net of 20 μm mesh size from different locations of the bloom forming area in the river and the Oscillatoria sp. cell density was stated as the average of three counts. A portion of the concentrated samples were filtered through an 0.45 μm glass fiber filter (Whatman GF/C, 47 mm diameter) and dried in an oven at 60-80°C. Dried filters covered with algae cells, were transported to the Institute of Nutrition, University of Jena, Germany for analysis.
The algae on the GF/C filter was immersed in 1.0 mL of a mixture of water
and methanol (50:50; v:v), sonicated for 20 min and was finally centrifuged
(3000 g). The supernatant was filtered through a nylon filter with 0.45 μm
The HPLC/UV determination of microcystins was carried out following the method
by Lawton et al. (1994) with some modification
(Hummert et al., 2001) (C18 column: Phenomenex
prodigy, ODS (3), 250x4.6 mm, 5 μm, mobile phases: acetonitrile /water/0.05%TFA).
Detection of microcystins was done by an UV detector (Shimadzu SPD-10AV; 238
nm) and a diode array detector (Shimadzu SPD-M6A). The HPLC was coupled by means
of an electrospray interface to a single quadrupol mass spectrometer (PE/Sciex
API 150EX, Perkin-Elmer,Germany). The detection was carried out in Selected
Ion Monitoring (SIM) mode (Hummert et al., 2001).
Microcystins and Nodularin Standards
Standards of Microcystin-RR, Microcystin-LR, Microcystin-YR, Microcystin-LA
and nodularin were purchased from Calbiochem/Novabiochem (La Jolla, USA).
HPLC grade acetonitrile and methanol were collected from Baker (Deventer,
Netherlands). Water was purified to HPLC grade with a Millipore-Q RG Ultra Pure
Water System (Millipore, Milford, USA).
The River Buriganga is increasingly being polluted with citys thousands
of industrial units and sewerage wastes. There are three main pollutant discharge
routes into the Burigonga; viz., Hazaribugh Tanneries, City drains along
the river and Pagla sewerage treatment plant outfall. The physico-chemical parameters
of the study area (Station, Islambagh) are shown in Table 1.
The details of physico-chemical parameters, phytoplankton abundance and pollution
status of the River Buriganga are described by Ahmed et
al. (2007). Recent investigation also found that the monthly average
faecal coliform in the water, zooplankton and phytoplankton sample of Buriganga
were 3.99x109, 4.54x103 and 4.28x102 (cfu L-1),
respectively. Vibrio cholerae 01 and V. cholerae 0139 were isolated
from water, zooplankton and phytoplankton samples (Ahmed
et al., 2007).
In the original bloom sample the cell density of Oscillatoria sp. was 1.6x104 cells L-1. During the bloom the dissolved oxygen, free carbon dioxide, nitrite nitrogen, alkalinity and ammonia, of river water were recorded as 1.5, 63.0, 0.86, 175.0 and 2.05 mg L-1, respectively. The pH was 9.5 and the water temperature was between 28-30°C. The HPLC analysis of Oscillatoria sp. bloom extract showed a peak corresponding to the retention time of standard microcystin-RR (Fig. 1). The concentration of microcystin-RR was 0.235 μg L-1 and 14.68 pg cell-1.
||(A) HPLC separation of standards containing Microcystins and
Nodularin: Microcystin-RR, Micricystin-LR, Microcystin-LA, Microcystin-YR,
Microcystin-LW, Microcystin-LF. (B) Chromatograms of Oscillatoria sp.
Blooms extracted with water:methanol (50:50; v:v), UV detection 238 nm wave
The occurrence of cyanobacteria blooms are very common phenomenon in the freshwater
environments in Bangladesh. The main factors leading to periodic cyanobacteria
proliferations were pointed out as increased dissolved organic nutrients, long
sunshine hours and favorable water temperature (Ahmed et
al., 2007). As in other tropical country, cyanobacteria blooms phenomena
will often last year round occurring in many eutrophic lakes and reservoirs
in Bangladesh when the climate permits.
The HPLC analysis of Oscillatoria sp. extract showed one peak, the retention
time of which agreed well with standard Microcystin-RR (Fig. 1).
The results of HPLC-MS revealed the identification of microcystin-RR according
to its corresponding molecular weight. Further structural variants of microcystins
were not detected. The concentration of microcystins detected in the present
study was (0.235 μg L-1) less than the previously detected microcystins
from a lake and ponds in Bangladesh (Ahmed et al.,
2008a, b; Welker et al., 2005). And that should
be due to the differences in location, cell type, cell densities and environmental
factors. Welker et al., 2005 in a study at three different regions in
Bangladesh detected microcystins in 39 ponds, mostly together with varying abundance
of potentially microcystin-producing genera such as Microcystis, Planktothrix
and Anabaena. Total microcystin concentrations in their study ranged
between <0.1 and up to >1000 μg L-1 and more than half
of the positive samples contained high concentrations of more than 10 μg
L-1 . In Australia, a safety factor for tumor promotion is 1.0 μg
microcystins or nodularins L-1 (Falconer et
al., 1999). In Canadian drinking water maximum accepted concentration
for MC-LR is 0.5 mg L-1 and for other microcystins, 1 μg L-1
of total microcystins (Carmichael, 1995).
The occurrence of Oscillatoria sp. blooms in ponds, lakes or rivers
that produce hepatotoxic microcystins is a problem, especially if the water
is utilized as a drinking supply and/or for recreational purposes. Epidemiological
investigations have demonstrated that microcystins cause stomach and intestinal
inflammation, liver cancer and disease of the spleen in humans who drink water
containing microcystins (McDermott et al., 1998;
Ding et al., 2000; Zhou et
al., 2002). The Dhaka Water Supply And Sewerage Authority (DWASA) supplied
39.1 million liter of waters per day from the river Buriganga as drinking water
to the residents of Dhaka city through its Chandi Ghat water treatment plant.
Recently, it is also evident that microcystins does accumulate in the liver
and muscle of fish when they are exposed to toxic cyanobacteria bloom (Magalhaes
et al., 2001; Deblois et al., 2008).
In tropical and subtropical climates, where blooms can be permanent, chronic
year-round exposure is likely to occur, leading to potentially high levels of
microcystin contamination. So, the effect of cyanobacteria blooms (microcystins)
on human through direct exposure or food chain in Bangladesh waters remains
to be identified. Moreover, it is necessary to have regular monitoring on abundance
of cyanobacteria in waters for public health safety.
Authors would like to extend their thanks to Prof. Dr. Bernd Luckas, University of Jena, Germany for his support during analysis of the samples.
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