Project Grant R21AI176089

MOLECULAR MECHANISMS FOR CLINICAL DOMINANCE OF P. AERUGINOSA ST111 ISOLATES - PROJECT SUMMARY PSEUDOMONAS AERUGINOSA IS AN OPPORTUNISTIC HUMAN PATHOGEN RATED AS A "SERIOUS THREAT" BY THE CENTERS FOR DISEASE CONTROL AND PREVENTION. IT IS PARTICULARLY DANGEROUS TO IMMUNOCOMPROMISED PATIENTS, INCLUDING PATIENTS WITH HEMATOLOGIC MALIGNANCIES (HM) AND THOSE RECOVERING FROM HEMATOPOIETIC STEM CELL TRANSPLANTATION (HCT). RECENT EPIDEMIOLOGICAL STUDY DISCOVERED THAT FLUOROQUINOLONE (FQ) NEUTROPENIC PROPHYLAXIS IN HM/HCT PATIENTS WAS ASSOCIATED WITH BREAKTHROUGH BLOODSTREAM INFECTIONS WITH P. AERUGINOSA. THESE BLOODSTREAM INFECTIONS WERE DOMINATED BY ISOLATES FROM THE ST111 MULTILOCUS SEQUENCE TYPE, A GROUP OF HIGH-RISK P. AERUGINOSA CLONES KNOWN TO FREQUENTLY CAUSE OUTBREAKS OF MULTIDRUG OR EXTREMELY DRUG RESISTANT INFECTIONS IN HEALTHCARE SETTINGS, INCLUDING HCT UNITS. OUR PRIOR STUDIES DEMONSTRATED THAT THE VAST MAJORITY OF ST111 ISOLATES FROM HM/HCT PA- TIENTS AT OREGON HEALTH SCIENCES UNIVERSITY UNDERGOING FQ PROPHYLAXIS WERE NON-SUSCEPTIBLE TO BOTH FQ AND MEROPENEM, DESPITE ONLY A SMALL FRACTION (5%) OF PATIENTS RECEIVING MEROPENEM IN THE PREVIOUS 6 MONTHS. CAR- BAPENEMS ARE A CRUCIAL, FRONT-LINE THERAPY IN THIS PATIENT POPULATION, AND NON-SUSCEPTIBILITY IS ASSOCIATED WITH SUBSTANTIAL INCREASES IN MORBIDITY AND MORTALITY. DESPITE THIS, A KNOWLEDGE GAP REMAINS IN THE MECHA- NISTIC UNDERSTANDING OF THE FACTORS THAT CONTRIBUTE TO ST111 DOMINANCE. OUR PRELIMINARY DATA INDICATED THAT AT LEAST TWO DIFFERENT FACTORS MAY DRIVE THIS DOMINANCE. FIRST, WE HAVE EVIDENCE THAT ST111 SECRETES AN INHIBITORY PROTEIN COMPLEX THAT PREVENTS THE GROWTH OF SOME OTHER STRAINS OF P. AERUGINOSA. THIS ALLOWS ST111 STRAINS TO DOMINATE OTHER STRAINS, DESPITE GROWING WORSE IN ISOLATION. OUR PRELIM- INARY STUDIES HAVE IDENTIFIED THIS FACTOR AND PINPOINTED A DEFENSE MECHANISM USED BY RESISTANT P. AERUGINOSA STRAINS TO OVERCOME GROWTH INHIBITION. SECOND, WHOLE GENOME SEQUENCING HAS DEMONSTRATED THAT ALL OF THE DOMINANT ST111 ISOLATES HAVE DISRUP- TIONS TO THE QUORUM SENSING REGULATOR LASR. THESE QUORUM SENSING MUTANTS ARE PREDICTED TO BE ABLE TO UTILIZE THE NORMAL SIGNALING PATHWAYS, BUT DO NOT EXPEND THE RESOURCES TO CONTRIBUTE TO THEM (A PHENOMENON OFTEN OBSERVED IN QUORUM SENSING MUTANTS). IMPORTANTLY, SOME OF THE ST111 ISOLATES HAD A FITNESS ADVANTAGE COM- PARED TO PA14LASR, WHILE OTHERS DID NOT, SUGGESTING THAT THESE FACTORS MAY BE ADDITIVE. IN THIS GRANT, WE PROPOSE TO STUDY EACH OF THESE MECHANISMS AND ESTIMATE THEIR IMPACT IN A WIDER PANEL OF ST111 AND SENSITIVE ISOLATES. GENETIC AND CELL BIOLOGICAL TOOLS WE HAVE GENERATED WILL BE USED TO STUDY EACH OF THESE FACTORS, INCLUDING PREVIOUSLY UNRECOGNIZED PLAYERS IN THE PRODUCTION OF THE INHIBITORY FACTOR. WE WILL ALSO STUDY LASR AND ITS DOWNSTREAM COMPONENTS TO DETERMINE WHAT ROLE THEY PLAY IN ST111 DOMINANCE. COMPLETION OF THE PROPOSED EXPERIMENTS WILL HELP CONFIRM FACTORS THAT DRIVE THE DOMINANCE OF ST111 AND MAY HELP IDENTIFY METHODS TO LIMIT THIS DOMINANCE SO THAT THE SPREAD OF ST111 CAN BE REDUCED, SIGNIFICANTLY IMPROVING CLINICAL OUTCOMES FOR HM/HCT PATIENTS.