Development of a Dry Bonde Nutrient Medium for Detection of Pseudomonas aeruginosa in Environmental Samples

Introduction: P. aeruginosa belongs to ESKAPE pathogens, which are highly resistant to antimicrobials and pose a significant public health challenge. Pseudomonas is characterized by its ability to form biofilms on various environmental surfaces (especially in humid environments), which protect the pathogen from antibiotic exposure. To detect P. aeruginosa in environmental samples, current regulatory documents require the use of a culture method using laboratory-prepared Bonde’s culture medium.

Objective: To develop a dry Bonde nutrient medium to detect Pseudomonas aeruginosa in environmental samples.

Materials and Methods: The current regulatory and methodological documents regulating the use of Bonde nutrient media for the accumulation of P. aeruginosa are analyzed. The experiments used strains of test microorganisms obtained from the State Collection of Pathogenic Microorganisms and Cell Cultures (SCPMCC-Obolensk). The studies were carried out according to the methods described in MG. 4.2.2316–08. To assess sensitivity and determine the effectiveness of media, we used target strains of pseudomonads at a 10–7 dilution as well as non-target strains at 10–4 and 10–5 dilutions to evaluate the inhibitory properties of the media. Subculturing from the Bonde medium to Gloss (“Blesk”) medium and nutrient medium No. 1 was carried out using the Gould sector sowing method. The isolates from simulated samples were identified by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry.

Results: We created a dry modified Bonde medium promoting a faster growth of P. aeruginosa. It demonstrated high sensitivity enabling the detection of low amounts of the bacteria (10 CFU/mL) and high inhibitory capacity against associated microbes (10,000 CFU/mL). The modified Bonde medium was 1.7 to 4.8 times more effective than the laboratory-prepared Bonde medium in terms of accumulation of all P. aeruginosa strains under study.

Conclusion: Application of the improved Bonde growth medium will enhance the quality and reliability of detection of P. aeruginosa in environmental samples including those with low levels of contamination. 

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