Abstract: [Objective] To identify the microbial species and protease-producing activity that cause the deterioration of raw milk. [Method] The proteolyzed and fat-floating metamorphic raw materials provided by a large-scale ranch in Hengyuan City，Guangdong Province，were diluted and coated on MPC agar plates. The plates were incubated in a 6.5 ℃ thermostatic incubator for ten days， and the monobacterial colonies with different morphologies were picked up and reinvigorated in LB liquid medium， their DNA was extracted，and the isolated strains were identified by the 16S rDNA method. The protease activity and enzyme residue rate of this strain were also determined. [Result] MPC agar plate colony form is single，and the two strains isolated and identified in the raw milk are Pseudomonas rhodesiae. The hydrolysis circle of the milk plate at 28 ℃ was （15.35±0.56）mm，and the protease residue rate was 43.63% after 5 s treatment at 135 ℃.[Conclusion] Two strains of Pseudomonas rhodesiae were isolated from spoiled raw milk，and the bacterium produces heat-insensitive proteases. The presence of this enzyme will cause protein hydrolysis in raw milk，resulting in quality problems such as proteolysis and swelling of UHT milk during the shelf life.

Key words: deteriorated raw milk, pseudomonas, isolation and identification, protease activity