Toxic Caynobacterial Bloom & Its Monitoring Perspectives

Toxic Caynobacterial Bloom & Its Monitoring Perspectives

Cyanobacterial blooms can alter the quality of water by releasing toxic & odorous compounds which has adverse impacts on animal and human health. Among the cyanobacteria, Microcystis sp. is frequently detected which produce hepatotoxins called microcystins (MCs). Ecological parameters which regulates Microcystis blooms, is complicated & diverse. This makes the local authorities difficult in controlling and managing cyanobacterial blooms. As a consequence, development of suitable monitoring approach to characterize cyanobacterial blooms is an important requirement. The main objective of this article is to evaluate monitoring approaches that are applicable to cyanobacterial blooms, especially focusing for Microcystis.

There are diverse and complex interactions among physical, chemical and biological variables leading to the proliferation of cyanobacteria and the regulation of MC synthesis at the cellular level. The lack of definitive correlation between a cyanobacterial bloom formation and MCs production necessitates the need for the development of rapid and more reliable methods for routine monitoring of Microcystis as well as MCs and their utilization in environmental sampling. The development of a cyanobacterial bloom mainly depends on the available nutrients, thus the environmental monitoring of nutrient concentrations provides valuable information for assessing bloom-forming potential.

Some of the techniques involved in monitoring Microcystis and MCs include: microscopy, photopigments, physiochemical analysis, enumeration of Microcystis cells, spectrophotometric , fluorometric analysis  remote sensing, ELISA, protein phosphatase inhibition assays (PPIA), HPLC and liquid chromatography-mass spectrometry (LCMS). Molecular techniques such as real-time PCR are important tool for early warning of cyanobacterial blooms. The current biological and physiochemical methods and their advantages and limitations for monitoring Microcystis and MCs are summarized in the below given figure:

Biological methods for monitoring Microcystis and MCs:

1. Cell Counting:

Flow cytometry and microscopic analysis, can be used for monitoring blooms but does not provide detailed information on species composition. Cell counting helps in the direct assessment of those organisms present in a cyanobacterial bloom; however, it is very laborious, time-consuming and requires skilled analysts for species identification.

2. Pigment analysis:

Spectrophotometric and fluorometric analysis are the preferred methods for the determination of chlorophyll a (Chl a). The presence of other accessory pigments and degradation products may interfere with its determination. As a consequence, submersible probe (FluoroProbe), containing five light emitting diodes, was developed for the estimation of total Chl a. Two- channel fluorometric sensor was used to detect phycocyanin (PC) of cyanobacteria. A three-stage alert system was proposed to monitor Microcystis bloom in Korean lakes and was based on PC levels of 0.1 (caution), 30 (warning), and 700 μg/L (outbreak), respectively. In vivo PC fluorescence measurements using field probes can provide early detection of cyanobacteria in drinking water intakes but, pigment contents (Chl a or PC) may vary with species and metabolic state of the cells which linits its use in monitoring. These probes also possess other limitations such as light source, turbidity of water, etc. Airborne sensors have been used by many investigators for mapping cyanobacterial blooms. Forecasting cyanobacterial bloom using remote sensing devices suffers from high cost, dependency on meteorological conditions with long monitoring intervals, which limits their use for routine monitoring.

3. Molecular Techniques:

Molecular-based techniques offer several advantages over other conventional monitoring methods. Real-time PCR is useful for quantitative analysis of cyanobacterial strains and denaturing gradient gel electrophoresis can be used to evaluate variation in community dynamic of cyanobacteria.  DNA chip/microaaray, using gene specific oligonucleotide probes is a technique suitable for high-throughput analysis and to study cyanobacterial community composition. Microarray can be used for the rapid identification of caynobacterial groups undetectable or present in low quantities, making it suitable for monitoring toxic as well as non-toxic strains in large number of environmental samples. Various genes for example, 16S rRNA, internal transcribed spacer (ITS) and a phycocyanin intergenic spacer (PC-IGS) have been used for characterizing toxic and non-toxic Microcystis. Overall, molecular techniques are expensive, but useful for early detection of potentially toxic organism.

Biochemical and Physiochemical methods for monitoring Microcystis and MCs:

Nutrients and other variable parameters such as water, temperature, pH, salinity etc can regulate the growth of cyanobacteria. These parameters are considered valuable in assessing the potential for future bloom development. The various physiological techniques used for monitoring Microcystis bloom and its toxins are given below:

ELISA and PPIA are rapid, sensitive, and easy to operate. These techniques are useful for quantification of MCs in drinking water before and after treatment. However, ELISA is prone to interferences and PPIA may respond to other protein phosphatase inhibitors. HPLC is the most commonly used analytical method for the detection and identification of MCs, but this method also lacks specificity for MCs. LC-MS is suitable when further conformation and identification of MCs is required. MALDI-TOF is useful for identification of MCs with very small sample volumes. GC-MS is highly sensitive and suitable for screening and accurate quantification of MCs in complex sample matrix but it is time-consuming and costly. NMR requires relatively large amounts of pure sample and expert and therefore it is not suitable for routine monitoring. Various portable biosensors have also been developed for rapid MCs detection in environmental samples. Enzymes, antibodies or nucleic acids based biosensors are highly specific and sensitive. However, most of the biosensors have been specifically developed for the MC-LR variant.

Conclusion:

The ultimate aim of monitoring programs is to predict cyanobacterial bloom events. Monitoring cyanobacterial blooms depend on various local aspects such as the intended use (drinking or recreation) & the types (Ponds, lakes, river or oceans) of water bodies. The factors regulating the cyanobacterial growth and toxin production are still not properly understood. The genetic regulation of cyanotoxin production is an important area for further study and exploration. Various analytical methods are limited in terms of their application in routine monitoring. Microscopic identification, cell counting and pigment analysis have traditionally been employed in monitoring programs of water bodies having Microcystis blooms. The data obtained from these methods can be employed as an early warning system by comparing with the threshold limits set by WHO. The development of a bloom is regulated by various nutrients; therefore, the source of such nutrients should be taken into account for early warning. Toxic and nontoxic strains may be present together in blooms of Microcystis sp. At a particular time, toxins may be released after cell lysis and sometimes cells might be present without any detectable toxins. In the absence of cyanobacterial cells, the monitoring of toxins by the more sensitive and rapid ELISA or PPIA methods is suggested. Subsequently, molecular approaches are useful for providing the information about the abundance and dominance of key toxic species and thus furnish the best monitoring/forecasting tool available for source water control strategy.