Opportunistic pathogens (OPs) in drinking water, like Legionella spp., mycobacteria, Pseudomonas aeruginosa and free-living amobae (FLA) are a risk to human health, due to their post-treatment growth in water systems. To assess and manage these risks, it is necessary to understand their variations and environmental conditions for the water routinely used. We sampled premise tap (N cold=26, N hot=26) and shower (N shower=26) waters in a bathroom and compared water temperatures to levels of OPs via qPCR, and identified Legionella spp. by 16S rRNA gene sequencing. The overall occurrence and cell-equivalent quantities (CE L −1) of Mycobacterium spp. were highest (100%, 1.4×10 5), followed by Vermamoeba vermiformis (91%, 493), Legionella spp. (59%, 146), P. aeruginosa (14%, 10), and Acanthamoeba spp. (5%, 6). There were significant variations of OP’s occurrence and quantities, and water temperatures were associated with their variations, especially for Mycobacterium spp., Legionella spp. and V. vermiformis. The peaks observed for Legionella, mainly consisted of L. pneumophila sg1 or L. anisa, occurred in the temperature ranged from 19 to 49°C, while Mycobacterium spp. and V. vermiformis not only co-occurred with Legionella spp. but also trended to increase with increasing temperatures. There were higher densities of Mycobacterium in 1 st than 2 nd draw water samples, indicating their release from faucet/shower head biofilm. Legionella spp. were mostly at detectable levels and mainly consisted of L. pneumophila, L. anisa, L. donaldsonii, L. tunisiensis and an unknown drinking water isolate based on sequence analysis. Results from this study suggested potential health risks caused by opportunistic pathogens when exposed to warm shower water with low chlorine residue and the use of Mycobacterium spp. as an indicator of premise pipe biofilm and the control management of those potential pathogens.