An in vitro experiment was conducted to understand the nature of biofilm growth on polyvinyl chloride (PVC) surface when exposed to suboptimal-quality microbial water (>4 log cfu/mL) obtained from a poultry drinking water source mimicking water in waterlines during the first week of poultry brooding condition. PVC sections (internal surface area of 15.16 cm) were utilized in the study to grow biofilm. After a 7-d test period, test coupons with 7-day-old biofilm were transferred into autoclaved municipal water and then treated with either chlorine-based or hydrogen peroxide-based sanitizer at bird drinking water rate, to see the impact on removal of biofilm formed on test coupons. Two trials (T1 and T2) were conducted. Test coupons used in T1 and T2 had the bacterial growth of 3.67 (SEM 0.04) and 3.97 (SEM 0.11) log cfu/cm on d 7. After sanitizer application, chlorine-based sanitizer removed bacteria in biofilm completely (0 cfu/cm) within 24 h post treatment whereas hydrogen peroxide-based sanitizer reduced the counts to 1.68 log cfu/cm (P < 0.05) by 48 h post sanitizer application. Control remained the same (P > 0.05). Results indicated that biofilm formation can occur quickly under suboptimal water condition on PVC surface, and sanitizer application helped mitigate already formed biofilm, yet chlorine proved to be more effective than hydrogen peroxide.