Diagnosis and Stratification of Pseudomonas aeruginosa Infected Patients by Immunochemical Quantitative Determination of Pyocyanin From Clinical Bacterial Isolates

Frontiers in Cellular and Infection Microbiology

Nuria Pascual¹  Lluïsa Vilaplana¹  Maria-Pilar Marco¹  Carme Pastells¹  Barbara Rodriguez-Urretavizcaya¹  Maria Teresa Martin-Gomez² 

[1] Centro de Investigación Biomédica en Red (CIBER) de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Madrid, Spain;Microbiology Department, Hospital Universitari Vall d’Hebron, Barcelona, Spain;Nanobiotechnology for Diagnostics (Nb4D), Institute of Advanced Chemistry of Catalonia, Institute for Advanced Chemistry of Catalonia (IQAC)-Spanish National Research Council (CSIC), Barcelona, Spain;

关键词: quorum sensing;    pyocyanin;    ELISA;    Pseudomonas aeruginosa;    monoclonal antibody;    diagnostic;   

DOI  :  10.3389/fcimb.2021.786929

来源: DOAJ

【 摘 要 】

The development of a highly sensitive, specific, and reliable immunochemical assay to detect pyocyanin (PYO), one of the most important virulence factors (VFs) of Pseudomonas aeruginosa, is here reported. The assay uses a high-affinity monoclonal antibody (mAb; C.9.1.9.1.1.2.2.) raised against 1-hydroxyphenazine (1-OHphz) hapten derivatives (PC1; a 1:1 mixture of 9-hydroxy- and 6-hydroxy-phenazine-2-carobxylic acids). Selective screening using PYO and 1-OHphz on several cloning cycles allowed the selection of a clone able to detect PYO at low concentration levels. The microplate-based ELISA developed is able to achieve a limit of detection (LoD) of 0.07 nM, which is much lower than the concentrations reported to be found in clinical samples (130 μM in sputa and 2.8 μM in ear secretions). The ELISA has allowed the investigation of the release kinetics of PYO and 1-OHphz (the main metabolite of PYO) of clinical isolates obtained from P. aeruginosa-infected patients and cultured in Mueller–Hinton medium. Significant differences have been found between clinical isolates obtained from patients with an acute or a chronic infection (~6,000 nM vs. ~8 nM of PYO content, respectively) corroborated by the analysis of PYO/1-OHphz levels released by 37 clinical isolates obtained from infected patients at different stages. In all cases, the levels of 1-OHphz were much lower than those of PYO (at the highest levels 6,000 nM vs. 300 nM for PYO vs. 1-OHphz, respectively). The results found point to a real potential of PYO as a biomarker of P. aeruginosa infection and the possibility to use such VF also as a biomarker for patient stratification[2] and for an effective management of these kinds of infections.

【 授权许可】

Unknown