The growth and composition of periphyton in low saline groundwater (5 ppt) were evaluated using 5 different substrates: split bamboo poles (SBP), green shade net (GSN), high density polyethylene (HDPE) sheets, low density polyethylene (LDPE) sheets, and polyvinyl chloride (PVC) pipes). The effect of substrate type, submersion time, and depth was evaluated. The average submersion depth of all the substrates was maintained at 88.27 cm, and this trial was conducted for 60 days. Chlorophyll a, dry weight, ash, ash free dry weight, and periphyton diversity were analyzed. The growth of periphyton in terms of chlorophyll a value on different substrates has revealed a higher mean value for bamboo substrate (6.82 ± 2.79 µg/cm 2 ), followed by green shade net, HDPE, LDPE, and PVC with values of 5.24 ± 2.27 µg/cm 2 , 4.64 ± 2.06 µg/cm 2 , 4.51 ± 2.08 µg/cm 2 , and 3.10 ± 1.57 µg/cm 2 , respectively. The mean values of dry weight recorded were 1.81 ± 0.78 mg/cm 2 , 1.65 ± 0.70 mg/cm 2 , 1.29 ± 0.56 mg/cm 2 , 1.32 ± 0.57 mg/cm 2 , and 0.78 ± 0.45 mg/cm 2 for SBP, GSN, LDPE, HDPE, and PVC substrates, respectively. In all substrates, there was a significant difference observed in the development of periphyton over time (p < 0.05). There was no significance (p > 0.05) observed in Chlorophyll a value with depth in all 5 different substrates.