Objective：To determine the contamination of Pseudomonasa spp. in dental unit waterlines(DUWLs) and water samples using bacterial culture technique as well as Miseq sequencing technology.Methods：Water samples were collected from 25 dental units(DU) at the Affiliated Stomatology Hospital of Nanjing Medical University.Before the opening of the clinic，a water sample of 400 mL was collected from each DU using triple change syringe.Each sample was divided into two parts.One part of the water samples was used for bacterial culture.The other one was used for extraction of the total bacterial DNA，which was subjected to PCR amplification using bacterial universal primers.The PCR products of each sample were constructed using Miseq sequencing technology to complete the sequencing analysis and biological information analysis.Results：No Pseudomonas spp. was detected by bacterial culture in 25 samples under the condition of different concentrations.The metagenomic libraries were successfully constructed by 16 samples.The detection rate of Pseudomonas spp. was 100%.The relative abundance of Pseudomonas spp. in 16 samples was accounted for(3.02± 2.12)%.Conclusion：Low level of contamination of Pseudomonas spp.existed in DUWLs. Most clones of the Pseudomonas spp. in DUWLs were dead or in viable but nonculturable(VBNC) condition.Thus，Pseudomonas spp. presented a low biohazard risk in DUWLs.When compared to bacterial culture，Miseq sequencing technology is more accurate in detecting particular pathogens. However，Miseq sequencing technology has limitation on identification of viable bacterial from dead ones，which may lead to false positive results.It needs further researches using high-throughput sequencing technology for selectively detecting viable pathogens.