Development and antimicrobial susceptibility studies of in vitro monomicrobial and polymicrobial biofilm models with Aspergillus fumigatus and Pseudomonas aeruginosa.

BACKGROUND: Mixed microbial infections of the respiratory tracts with P. aeruginosa and A. fumigatus capable of producing biofilms are commonly found in cystic fibrosis patients. The primary objective of this study was to develop an in vitro model for P. aeruginosa and A. fumigatus polymicrobial biofilm to study the efficacy of various antimicrobial drugs alone and in combinations against biofilm-embedded cells. Simultaneous static cocultures of P. aeruginosa and sporelings were used for the development of in vitro P. aeruginosa-A. fumigatus polymicrobial biofilm in SD broth in 24-well cell culture plates at 35°C, and the biofilm formation was monitored microscopically and spectrophotometrically. Using P. aeruginosa-A. fumigatus sporelings cocultures we examined the effects of various antimicrobial drugs alone and in combination against polymicrobial biofilm by CFU and tetrazolium reduction assays. RESULTS: In simultaneous static cocultures P. aeruginosa cells killed A. fumigatus conidia, whereas the bacterial cells showed no substantial fungicidal effect on sporelings grown for 12 h or longer at 35°C. Monospecies cultures of P. aeruginosa produced loosely adhered monomicrobial biofilm and addition of 10% bovine serum to the growth medium inhibited the formation of monomicrobial biofilm by P. aeruginosa whereas it produced tightly adhered polymicrobial biofilm in the presence of A. fumigatus mycelial growth. A. fumigatus produced firmly adherent monomicrobial and polymicrobial biofilms. A comparison of CFU and MTT assays showed that the latter is unsuitable for studying the effectiveness of antimicrobial treatment against polymicrobial biofilm. Tobramycin alone and in combination with posaconazole was highly effective against monomicrobial and polymicrobial biofilms of P. aeruginosa whereas cefepime alone and in combination with posaconazole showed excellent activity against monomicrobial biofilm of P. aeruginosa but was less effective against polymicrobial biofilm. Monomicrobial and polymicrobial biofilms of A. fumigatus showed similar susceptibility to posaconazole with and without the antibacterial drug. CONCLUSIONS: Simultaneous static coculture of A. fumigatus sporelings grown for 12 h or longer was superior to ungerminated conidia with P. aeruginosa for the development of A. fumigatus-P. aeruginosa biofilm. P. aeruginosa-A. fumigatus polymicrobial biofilm shows differential susceptibility to antimicrobial drugs whereas the susceptibility of A. fumigatus to antimicrobial drugs was unchanged.

Molecular epidemiology of methicillin-resistant Staphylococcus aureus bloodstream isolates in urban Detroit.

To gain a better understanding of epidemiology of resistance in Staphylococcus aureus, we describe the molecular epidemiology of methicillin-resistant Staphylococcus aureus bloodstream isolates in urban Detroit. Bloodstream isolates from July 2005 to February 2007 were characterized. Two hundred ten bloodstream isolates from 201 patients were evaluated. Patient characteristics were as follows: median age, 54 years; 56% male; and 71% African-American. Seventy-six percent of infections were health care associated, with 55% being community-onset infections and 21% hospital acquired, and 24% were community associated. The most common sources were skin/wound (25%), central venous catheters (24%), unknown source (20%), and endocarditis (9%). Ninety percent and 5% of isolates had a MIC of vancomycin of or=1.5 mg/liter. Results of pulsed-field gel electrophoresis showed 17 strain types. The predominant strains were USA100 (104 isolates) and USA300 (74 isolates). Forty-nine percent of the isolates had staphylococcal cassette chromosome mec II, and 56% had agr II. All USA300 isolates were positive for the Panton-Valentine leukocidin toxin genes and agr I. Forty-seven percent of USA300 bloodstream infections were health care associated (35% community onset and 12% hospital onset). USA300 strains were more common in injection drug users with skin/wound as the predominant source of infection. Thirty percent of the USA100 strains were closely related to vancomycin-resistant Staphylococcus aureus isolates. The results of this study show that vancomycin MICs using automated dilution testing with Vitek-2 and E-test were highly discordant. Most methicillin-resistant S. aureus strains causing bacteremia are health care associated, commonly have MICs of vancomycin that are high within the susceptible range are not detected by routine automated dilution testing, and have significant diversity of molecular characteristics. USA100 strains that are closely related to vancomycin-resistant S. aureus (VRSA) isolates and USA300 strains are common as causes of both hospital and community-onset infection. Infection control measures should focus not only on prevention of the spread of community strains in the hospital but also prevention of the spread of hospital strains associated with VRSA into the community.

Characterization of Salmonella Isangi possessing a CTX-M15 ESBL associated with an outbreak in a US Hospital.

Over an approximately 50-day period in 2015, an outbreak of CTX-M-15 extended spectrum ß-lactamase-(ESBL)-possessing Salmonella Isangi occurred among 19 adult surgical patients and one healthcare worker (HCW) at a large urban tertiary care hospital in the United States. A total of 45 S. Isangi isolates were isolated from stool (35), blood (4), urine (3), respiratory (2), and wound (1) cultures. Phenotypically, all but three isolates demonstrated resistance to ampicillin, ampicillin/sulbactam, ceftriaxone, and cefepime, and one isolate was resistant to ertapenem. Genotypically, a single CTX-M-15 ESBL was identified in all but three isolates by real-time PCR. Interestingly, two of the CTX-M-15 negative, susceptible isolates were isolated from a single patient who initially had a CTX-M positive, resistant strain. Isolates were clonally related, including both resistant and susceptible strains, as confirmed by pulse field gel electrophoresis (PFGE). This is the first case of a novel Salmonella outbreak at this hospital, and we believe it to be the first case of an S. Isangi serotype outbreak in the United States.

Nosocomial Outbreak of a Novel Extended-Spectrum ß-Lactamase Salmonella enterica Serotype Isangi Among Surgical Patients.

OBJECTIVE Nosocomial outbreaks caused by Salmonella are rare. We describe the investigation and control of a cluster of novel extended-spectrum ß-lactamase (ESBL) Salmonella enterica serotype Isangi in a hospital in southeastern Michigan. METHODS An epidemiologic investigation, including case-control study, assessment of infection control practices and environmental cultures, was performed to identify modes of transmission. Healthcare workers (HCWs) exposed to case patients were screened. Strain relatedness was determined using pulsed-field gel electrophoresis (PFGE); ESBL confirmation was conducted using real-time PCR. Control measures were implemented to prevent further transmission. RESULTS Between September 2 and October 22, 2015, 19 surgical patients, including 10 organ transplant recipients and 1 HCW, had positive S. Isangi cultures. Of these case patients and HCW, 13 had gastroenteritis, 2 had bacteremia, 1 had surgical-site infection, and 4 were asymptomatic. Pulsed-field gel electrophoresis (PFGE) showed 89.5% similarity among the isolates in these cases. Isolates with resistant-phenotypes possessed plasmid-mediated CTX-M15 ESBL. A total of 19 case patients were compared with 57 control participants. Case patients had significantly higher odds of exposure to an intraoperative transesophageal (TEE) probe (adjusted odds ratio 9.0; 95% confidence interval, 1.12-72.60; P=.02). Possible cross-transmission occurred in the HCW and 2 patients. Cultures of TEE probes and the environment were negative. The outbreak ended after removal of TEE probes, modification of reprocessing procedures, implementation of strict infection control practices, and enhanced environmental cleaning. CONCLUSIONS We report the first nosocomial ESBL S. Isangi outbreak in the United States. Multiple control measures were necessary to interrupt transmission of this gastrointestinal pathogen. Exposure to possibly contaminated TEE probes was associated with transmission. Periodic monitoring of reprocessing procedures of TEE probes may be required to ensure optimal disinfection. Infect Control Hosp Epidemiol 2016;37:954-961.

Comparative evaluation of epidemiology and outcomes of methicillin-resistant Staphylococcus aureus (MRSA) USA300 infections causing community- and healthcare-associated infections.

Methicillin-resistant Staphylococcus aureus (MRSA) USA300 clone is commonly found in the community and is being increasingly reported in the healthcare setting. A retrospective analysis was conducted to compare the epidemiology and outcomes between community-associated (CA) and healthcare-associated (HA) USA300 MRSA infections. The study enrolled 160 subjects with USA300 MRSA infections (47.5% CA-MRSA and 52.5% HA-MRSA). Failure in the HA group was higher (38.1%) compared with the CA group (23.7%) (P=0.05). Predictors of failure included male gender, age, presence of any co-morbidity, coronary artery disease, chronic kidney disease, history of MRSA, previous admission, fluoroquinolone exposure, HA infection and osteomyelitis (P