The present study was aimed to investigate the two aquaculture pond waters of physio-chemcial parameters and microbial analysis at Alwarkurichi village, Tirunelveli district. The physio-chemcial parameters are pH, conductivity, dissolved oxygen, BOD (Biological oxygen demand), calcium, magnesium, total hardness, chloride, phosphate, nitrate, and total dissolved solid were tested by Tamilnadu water supply and Drainage broad, District water testing laboratory, Palayamkottai. Subsequently, the water samples microbial colonies were identified with standard methods. They are Klebsiella pneumoniae, Pseudomonas aeruginosa and Vibrio cholera as more dominant in the two ponds. Hence, the identified three pathogenic bacteria were treated with some commercially popular chemicals at various concentrations. The antibacterial activity of three compounds Sodium chloride, Formalin and Copper sulfate was studied against three pathogenic bacteria using agar well diffusion method. The growth suppression rate exhibited by Copper sulfate vial higher at 0.08mg/10ml when equivalence to others. When compared to copper sulfate, size of the acquired growth distinct was less when examined with ampicillin. Copper sulfate was identified as the most efficacious against absolutely the three pathogens.