Patricia L. Winokur

Variations in the Prevalence of Strains Expressing an Extended-Spectrum β-Lactamase Phenotype and Characterization of Isolates from Europe, the Americas, and the Western Pacific Region

To evaluate the prevalence of extended-spectrum beta-lactamase (ESBL)-producing strains among species of Enterobacteriaceae, a microdilution susceptibility test was performed with strains of Klebsiella pneumoniae, Escherichia coli, Proteus mirabilis, and Salmonella species that were isolated as part of the SENTRY project. The highest percentage of ESBL phenotype (defined as a minimum inhibitory concentration [MIC] > or =2 microg/mL for ceftazidime, ceftriaxone, or aztreonam) was detected among K. pneumoniae strains from Latin America (45%), followed by those from the Western Pacific region (25%), Europe (23%), the United States (8%), and Canada (5%). P. mirabilis and E. coli strains for which MICs of extended-spectrum cephalosporins or monobactams were elevated also were more prominent in Latin America. Testing with ceftazidime revealed more isolates with elevated MICs than did testing with ceftriaxone or aztreonam. ESBL strains showed high levels of co-resistance to aminoglycosides, tetracycline, trimethoprim-sulfamethoxazole, and ciprofloxacin. Imipenem remains highly effective against ESBL strains. Organisms expressing an ESBL are widely distributed worldwide, although prevalence rates are significantly higher in certain geographic regions.

Evidence for Transfer of CMY-2 AmpC β-Lactamase Plasmids between Escherichia coli and Salmonella Isolates from Food Animals and Humans

ABSTRACT Escherichia coli is an important pathogen that shows increasing antimicrobial resistance in isolates from both animals and humans. Our laboratory recently described Salmonellaisolates from food animals and humans that expressed an identical plasmid-mediated, AmpC-like β-lactamase, CMY-2. In the present study, 59 of 377 E. coli isolates from cattle and swine (15.6%) and 6 of 1,017 (0.6%) isolates of human E. coli from the same geographic region were resistant to both cephamycins and extended-spectrum cephalosporins. AnampC gene could be amplified with CMY-2 primers in 94.8% of animal and 33% of human isolates. Molecular epidemiological studies of chromosomal DNA revealed little clonal relatedness among the animal and human E. coli isolates harboring the CMY-2 gene. The ampC genes from 10 animal and human E. coli isolates were sequenced, and all carried an identical CMY-2 gene. Additionally, all were able to transfer a plasmid containing the CMY-2 gene to a laboratory strain of E. coli. CMY-2 plasmids demonstrated two different plasmid patterns that each showed strong similarities to previously describedSalmonella CMY-2 plasmids. Additionally, Southern blot analyses using a CMY-2 probe demonstrated conserved fragments among many of the CMY-2 plasmids identified in Salmonella andE. coli isolates from food animals and humans. These data demonstrate that common plasmids have been transferred between animal-associated Salmonella and E. coli, and identical CMY-2 genes carried by similar plasmids have been identified in humans, suggesting that the CMY-2 plasmid has undergone transfer between different bacterial species and may have been transmitted between food animals and humans.

Survey of Bloodstream Infections Due to Gram-Negative Bacilli: Frequency of Occurrence and Antimicrobial Susceptibility of Isolates Collected in the United States,Canada, and Latin America for the SENTRY Antimicrobial Surveillance Program, 1997

During 1997, a total of 4,267 nosocomial and community-acquired bloodstream infections due to gram-negative organisms were reported from SENTRY hospitals in Canada (8 sites), the United States (30 sites), and Latin America (10 sites). Escherichia coli was the most common isolate (41% of all gram-negative isolates), followed by Klebsiella species (17.9%), Pseudomonas aeruginosa (10.6%), and Enterobacter species (9.4%). For all gram-negative isolates combined, the most active antimicrobials tested were meropenem, imipenem, and cefepime. The quinolones levofloxacin (MIC90, 2 microg/mL), ciprofloxacin (MIC90, 1 microg/mL), gatifloxacin (MIC90, 2 microg/mL), sparfloxacin (MIC90, 2 microg/mL), and trovafloxacin (MIC90, 2 microg/mL) were also active against most isolates. Bloodstream infection isolates from Latin America were uniformly more resistant to all classes of antimicrobial agents tested than were isolates from Canada or the United States. Resistance phenotypes consistent with extended-spectrum beta-lactamase production were also most common among E. coli and Klebsiella species from Latin America. Further investigation of the reasons for regional differences in resistance patterns is needed, as is ongoing surveillance to detect resistance trends and to guide antimicrobial use.

Antimicrobial Drug–Resistant Escherichia coli from Humans and Poultry Products, Minnesota and Wisconsin, 2002–2004

Similarities were found between drug-resistant E. coli from humans and poultry products.

Characteristics of pathogens causing urinary tract infections in hospitals in North America: results from the SENTRY Antimicrobial Surveillance Program, 1997

Urinary tract infection (UTI) is common and involves pathogens with changing susceptibility patterns. The SENTRY Antimicrobial Surveillance Program evaluates international pathogen incidence patterns to detect and manage the emergence of resistant strains. We describe the antimicrobial resistance patterns among 1617 pathogens recovered from UTIs during the third-quarter of 1997 in North America (United States and Canada), as part of this worldwide program. The isolates were tested against more than 50 antimicrobial agents (20 reported) by reference broth microdilution methods, and selected isolates were characterized by pulsed-field gel electrophoresis (PFGE) and automated ribotyping. The five most frequently isolated species were Escherichia coli (48.6%), Enterococcus spp. (13.7%), Klebsiella spp. (12.0%), Pseudomonas aeruginosa (6.2%), and Enterobacter spp. or Proteus mirabilis (3.8% each). For each nation, imipenem and cefepime produced the widest spectrum of coverage among the beta-lactams and amikacin was best among the aminoglycosides. For Gram-negative species, high resistance among beta-lactam antimicrobial agents was noted especially for various penicillins against E. coli (37.9% to 42.8%) and for the cephalosporins tested against enterococci (99.4% and 100%). Approximately 7.0% of enterococci in the USA were vancomycin-resistant (88% with Van A). P. aeruginosa provided the most consistent levels of resistance, but the following agents were most active against these organisms: amikacin (96.6 to 97.4% susceptible), tobramycin (89.5 to 100.0%), piperacillin/tazobactam (89.5 to 100.0%), piperacillin (89.5 to 96.6%), imipenem (89.7 to 92.1%), cefepime (77.6 to 89.7%), and ceftazidime (82.9 to 86.2%). E. coli (2.2 to 2.7%), K. pneumoniae (6.2 to 6.4%), and a single Enterobacter cloacae strain produced extended-spectrum beta-lactamases; and five other Enterobacter spp. were likely to have expressed chromosomally mediated (Amp C) Stably derepressed cephalosporinases with associated resistance to ceftazidime (16.7 to 21.2% resistance). These data demonstrated that several UTI isolates in SENTRY hospitals have high levels of resistance to various classes of antimicrobial agents with little evidence of clonal dissemination.

Antimicrobial susceptibility patterns for pathogens isolated from patients in Latin American medical centers with a diagnosis of pneumonia: analysis of results from the SENTRY Antimicrobial Surveillance Program (1997)

Pneumonia is the most common fatal hospital-acquired infection, with attributable mortality rates ranging from 30 to 60%. Rapid initiation of optimal antimicrobial therapy is essential for obtaining treatment success. In this report the antimicrobial susceptibility of 556 strains from the lower respiratory tract were collected by the SENTRY Antimicrobial Surveillance Program (1997). These strains were isolated from hospitalized patients with pneumonia in 10 Latin American centers (6 countries) as part of this 68-center worldwide program. The isolates were susceptibility tested against more than 70 drugs (35 reported) by the reference broth microdilution method. Klebsiella pneumoniae and Escherichia coli phenotypically consistent with extended spectrum beta-lactamase (ESBL) production were characterized further by ribotyping and pulsed-field gel electrophoresis. The five most frequently isolated species were (n/%): Pseudomonas aeruginosa (149/26.8%), Staphylococcus aureus (127/22.8%), Acinetobacter spp. (66/11.9%), Klebsiella spp. (56/10.1%), and Enterobacter spp. (40/7.2%). P. aeruginosa demonstrated high rates of resistance to a majority of the antimicrobial drugs tested. Carbapenems, amikacin, and piperacillin/tazobactam demonstrated the highest susceptibility rates (73.8-77.2%) against P. aeruginosa, however the lowest resistance rate was observed for cefepime (6.7%). Acinetobacter spp. also showed very high rates of resistance and the most active compounds were imipenem and meropenem (89.0% susceptibility) followed by the tetracyclines. Cephalosporin susceptibilities among Klebsiella spp. were low: cefoxitin, 73.0%; ceftazidime, 69.4%; and ceftriaxone, 65.9%. Approximately 37% and 28% of K. pneumoniae and E. coli isolates, respectively, were considered ESBL producers based on NCCLS criteria. Ceftriaxone was active against only 52.5% of Enterobacter spp. isolates, whereas cefepime was active against 90.0% of isolates (MIC50, < or = 0.12 microgram/mL). Oxacillin resistance was detected in nearly 50% of S. aureus isolates. The most active drugs against S. aureus were vancomycin, teicoplanin, and quinupristin/dalfopristin (MIC90, 1 microgram/mL). In summary, our study of pneumonias in Latin American medical centers demonstrated a greatly increased prevalence of Acinetobacter spp. and higher resistance rates among Gram-negative bacilli when compared with similar controlled studies from North America.

Cathelicidin Peptides Inhibit Multiply Antibiotic-Resistant Pathogens from Patients with Cystic Fibrosis

ABSTRACT Endogenous peptide antibiotics are under investigation as inhaled therapeutic agents for cystic fibrosis (CF) lung disease. The bactericidal activities of five cathelicidin peptides (LL37 [human], CAP18 [rabbit], mCRAMP [mouse], rCRAMP [rat], and SMAP29 [sheep]), three novel alpha-helical peptides derived from SMAP29 and termed ovispirins (OV-1, OV-2, and OV-3), and two derivatives of CAP18 were tested by broth microdilution assays. Their MICs were determined for multiply antibiotic-resistant Pseudomonas aeruginosa(n = 24), Burkholderia cepacia(n = 5), Achromobacter xylosoxidans(n = 5), and Stenotrophomonas maltophilia (n = 5) strains isolated from CF patients. SMAP29 was most active and inhibited mucoid and nonmucoidP. aeruginosa strains (MIC, 0.06 to 8 μg/ml). OV-1, OV-2, and OV-3 were nearly as active (MIC, 0.03 to 16 μg/ml), but CAP18 (MIC, 1.0 to 32 μg/ml), CAP18-18 (MIC, 1.0 to >32 μg/ml), and CAP18-22 (MIC, 0.5 to 32 μg/ml) had variable activities. LL37, mCRAMP, and rCRAMP were least active against the clinical isolates studied (MIC, 1.0 to >32 μg/ml). Peptides had modest activities against S. maltophilia and A. xylosoxidans (MIC range, 1.0 to > 32 μg/ml), but none inhibited B. cepacia. However, CF sputum inhibited the activity of SMAP29 substantially. The effects of peptides on bacterial cell membranes and eukaryotic cells were examined by scanning electron microscopy and by measuring transepithelial cell resistance, respectively. SMAP29 caused the appearance of bacterial membrane blebs within 1 min, killed P. aeruginosa within 1 h, and caused a dose-dependent, reversible decrease in transepithelial resistance within 5 h. The tested cathelicidin-derived peptides represent a novel class of antimicrobial agents and warrant further development as prophylactic or therapeutic agents for CF lung disease.

Serological Responses to an Avian Influenza A/H7N9 Vaccine Mixed at the Point-of-Use With MF59 Adjuvant: A Randomized Clinical Trial

IMPORTANCE Human infections with avian influenza A/H7N9 have resulted in high morbidity and mortality in China. OBJECTIVE To compare safety and immunogenicity of different doses of influenza A/Shanghai/2/13 (H7N9) vaccine mixed with or without the MF59 adjuvant. DESIGN, SETTING, AND PARTICIPANTS Multicenter, randomized, double-blind, phase 2 trial at 4 US sites enrolled 700 adults aged 19 to 64 years beginning in September 2013; 6-month follow-up was completed in May 2014. INTERVENTIONS The H7N9 inactivated virus vaccine was administered intramuscularly on days 0 and 21 at nominal doses of 3.75, 7.5, 15, or 45 µg of hemagglutinin (actual doses approximately 50% higher) with or without the MF59 adjuvant. A total 99, 100, or 101 participants were randomized to each group (7 groups; N = 700). MAIN OUTCOMES AND MEASURES Proportions achieving day 42 antibody titer of 40 or greater or seroconversion (a minimum 4-fold increase to titer ≥40) with the hemagglutination inhibition assay; vaccine-related serious adverse events through month 13; and solicited postvaccination symptoms through day 7. RESULTS Hemagglutination inhibition antibodies were minimal after participants received an unadjuvanted vaccine. After receiving 2 doses of H7N9 vaccine at a dosage of 3.75 µg plus the MF59 adjuvant, day 42 seroconversion occurred in 58 participants (59%; 95% CI, 48%-68%). The peak seroconversion occurred at day 29 in 62 participants (62%; 95% CI, 52%-72%). The day 42 geometric mean titer was 33.0 (95% CI, 24.7-44.1). Higher antigen doses were not associated with increased response. For the neutralizing antibody assays, after receiving 3.75 µg of H7N9 vaccine plus the MF59 adjuvant, day 42 seroconversion occurred in 81 participants (82%; 95% CI, 73%-89%). The day 42 geometric mean titer was 81.4 (95% CI, 66.6-99.5). There was no statistically significant difference in day 42 hemagglutination inhibition seroconversion after mixing adjuvant with either the first or both 15 µg doses (n = 34 [35%; 95% CI, 25%-45%] vs n = 47 [47%; 95% CI, 37%-58%], respectively; P = .10). Recent receipt of seasonal influenza vaccination and older age were associated with attenuated response. No vaccine-related serious adverse events occurred. Solicited postvaccination symptoms were generally mild with more local symptoms seen in participants who received the adjuvant. CONCLUSIONS AND RELEVANCE Point-of-use mixing and administration of 2 doses of H7N9 vaccine at the lowest tested antigen dose with MF59 adjuvant produced seroconversion in 59% of participants. Although these findings indicate potential value in this approach, the study is limited by the absence of antibody data beyond 42 days and the absence of clinical outcomes. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT01938742.

A High Dosage Influenza Vaccine Induced Significantly More Neuraminidase Antibody than Standard Vaccine among Elderly Subjects

Antibody to the neuraminidase (NA) antigen of influenza viruses has been shown to correlate with immunity to influenza in humans and animal models. In a previous report, we showed that an inactivated influenza vaccine containing 60microg of the hemagglutinin (HA) of each strain induced significantly more serum anti-HA antibody among elderly persons than did the standard vaccine containing 15microg of the HA of each component. We developed a lectin-based assay for anti-NA antibody and used it to measure anti-NA antibody responses among subjects who had participated in that study. The high dosage vaccine contained eight times as much NA activity as the standard vaccine and induced a significantly higher frequency of antibody responses and higher mean postvaccination anti-NA titers to the N1 and N2 of the A/H1N1 and A/H3N2 viruses in the vaccines than did the standard vaccine. Ensuring an increased antibody response to the NA antigen in inactivated influenza virus vaccines should increase the protection against influenza. An increased quantity of the NA antigen in the vaccine will ensure an increased response.

Distribution and characteristics of Escherichia coli clonal group A

Among 1,102 recent Escherichia coli clinical isolates, clonal group A was identified in 17 of 20 (U.S. and non-U.S.) geographic locales, mainly among U.S. isolates (9% vs. 3%; p < 0.001) and those resistant to trimethoprim-sulfamethoxazole (10% vs. 1.7%; p < 0.001). The extensive antimicrobial resistance and virulence profiles of clonal group A may underlie its recent widespread emergence.