Sugarcane accounts for around 80% of global sugar production. However, the sugar industry is known for producing significant amounts of organic wastewater with a high COD (5000–8000 mg/L) that severely pollutes the environment. A lab-scale trial was conducted to evaluate the efficacy of a horizontal subsurface flow wetland planted with Typha latifolia and Phragmites australis in removing pollutants from sugar industry wastewater. The wetland system was subjected to rigorous testing, operating at a high flow rate of 2.166 gallons per day and exposed to a high organic loading rate (3800 mg/L COD and 2470 mg/L BOD), as well as elevated levels of inorganic nitrogen, sulfate, and phosphate (100 mg/L, 80 mg/L, and 10 mg/L, respectively). Our findings indicate significant removal efficiencies, with the wetland system achieving removal rates of 88% for COD, 97% for BOD, 96% for total nitrogen, and 95% for sulfate. Remarkably, the system exhibited enhanced removal efficiency when exposed to domestic wastewater compared to tap water, owing to the abundance of microbial populations. Moreover, toxicity assessments conducted on the treated water revealed no adverse effects on the germination of wheat seeds and on the survival of fish over a week-long observation period. In conclusion, our study underscores the promising potential of horizontal subsurface flow wetlands as an effective and sustainable approach for mitigating the adverse environmental impacts associated with sugar industry wastewater. The findings offer valuable insights for policymakers and stakeholders in devising strategies to promote environmental sustainability and safeguard vital ecosystems in the Sindh region of Pakistan and beyond.