A rapid field-based protocol to assess the heavy metals bioindicator potential of Azolla pinnata in diverse...

This study presents a quick, cost-effective field protocol to assess Azolla potential as a bioindicator of heavy metal pollution in various wastewaters (blackwater (BW), graywater (GW), and two industrial effluents (IE1 and IE2). Hierarchical cluster analysis (HCA) and pprincipal component analysis (PCA) were deduced for the differentiation of wastewaters, with the results confirming the domestic-industry wastewater separation. Azolla pinnata was treated with all types of wastewaters for 15-20 days, with high-resolution morphological alterations detected. Stress-related parameters, viz., percentage cover, greenness index, and necrotic/chlorotic tissue percentage, were analyzed quantitatively. The maximum baseline levels (mg/L) of cadmium (0.01), lead (0.030), and zinc (0.50) were found in BW, GW, and IE1 and IE2, respectively. Results showed progressive reductions in cover and greenness, together with enhanced necrosis and chlorosis during exposure to industrial effluents, which was positively correlated with heavy metal concentrations, hence, confirming the bioindicator assay. This pairing of quantitative statistical wastewater profiling and morphological stress indices demonstrates A. pinnata as a viable bioindicator for preliminary wastewater screening. The protocol addresses a community in a developing country, spinning the nature-based solutions wheel in wastewater management. In addition to diagnostics, it further emphasizes A. pinnata in metal sequestration as well as stress biomonitoring which synergizes with the United Nation's Sustainable Development Goals (UNSDGs) 6, 12, and 15. Future work involving physiological and biochemical parameters and long-term validation in the field would further enhance the potential of the protocol for use in environmental monitoring and risk management.