Richard A. Venezia

Peptide Nucleic Acid Fluorescence In Situ Hybridization-Based Identification of Candida albicans and Its Impact on Mortality and Antifungal Therapy Costs

ABSTRACT The impact of rapid identification of Candida albicans blood isolates by peptide nucleic acid fluorescence in situ hybridization (PNA FISH) on the selection and expenditure of antifungal therapy was evaluated. PNA FISH was 100% sensitive and specific in the rapid identification of 31 out of 72 candidemias as C. albicans and resulted in a significant reduction of caspofungin usage, with an overall cost savings of

Peptide Nucleic Acid Fluorescent In Situ Hybridization for Hospital-Acquired Enterococcal Bacteremia: Delivering Earlier Effective Antimicrobial Therapy

1,729 per patient.

Multicenter evaluation of the Candida albicans/Candida glabrata peptide nucleic acid fluorescent in situ hybridization method for simultaneous dual-color identification of C. albicans and C. glabrata directly from blood culture bottles

ABSTRACT Hospital-acquired vancomycin-resistant enterococcal bacteremia has been associated with increased hospital costs, length of stay, and mortality. The peptide nucleic acid fluorescent in situ hybridization (PNA FISH) test for Enterococcus faecalis and other enterococci (EFOE) is a multicolor probe that differentiates E. faecalis from other enterococcal species within 3 h directly from blood cultures demonstrating gram-positive cocci in pairs and chains (GPCPC). A quasiexperimental study was performed over two consecutive years beginning in 2005 that identified GPCPC by conventional microbiological methods, and in 2006 PNA FISH was added with a treatment algorithm developed by the antimicrobial team (AMT). The primary outcome assessed was the time from blood culture draw to the implementation of effective antimicrobial therapy before and after PNA FISH. The severity of illness, patient location, and empirical antimicrobial therapy were measured. A total of 224 patients with hospital-acquired enterococcal bacteremia were evaluated, with 129 in the preintervention period and 95 in the PNA FISH period. PNA FISH identified E. faecalis 3 days earlier than conventional cultures (1.1 versus 4.1 days; P < 0.001). PNA FISH identified Enterococcus faecium a median 2.3 days earlier (1.1 versus 3.4 days; P < 0.001) and was associated with statistically significant reductions in the time to initiating effective therapy (1.3 versus 3.1 days; P < 0.001) and decreased 30-day mortality (26% versus 45%; P = 0.04). The EFOE PNA FISH test in conjunction with an AMT treatment algorithm resulted in earlier initiation of appropriate empirical antimicrobial therapy for patients with hospital-acquired E. faecium bacteremia.

National Survey on the Susceptibility of Bacteroides fragilis Group: Report and Analysis of Trends in the United States from 1997 to 2004

ABSTRACT We evaluated the performance of the Candida albicans/Candida glabrata peptide nucleic acid fluorescent in situ hybridization (PNA FISH) method, a rapid two-color assay for detection of C. albicans and C. glabrata, in a multicenter study. The assay is designed for use directly from positive blood culture bottles in a FISH format. Intact, fixed cells are labeled fluorescent green (C. albicans) or fluorescent red (C. glabrata) by rRNA hybridization of fluorophore-labeled PNA probes. Results are available <3 h after cultures signal positive. An evaluation of 197 routine blood culture bottles newly positive for yeast by Gram staining was performed at five hospitals. The sensitivities of detection for C. albicans, and C. glabrata were 98.7% (78/79) and 100% (37/37), respectively, and the specificity for both components of the assay was 100% (82/82). The assay was also evaluated with 70 fungal reference strains and was challenged in the BacT/ALERT microbiological detection system with spiked blood culture bottles. These results support the use of the assay for rapid, simultaneous identification of C. albicans and C. glabrata in positive blood culture bottles. This rapid assay may aid in the selection of initial antifungal drugs, leading to improved patient outcomes.

Comparison of Multiplex PCR Assay with Culture for Detection of Genital Mycoplasmas

ABSTRACT The susceptibility trends for the species of the Bacteroides fragilis group against various antibiotics from 1997 to 2004 were determined by using data for 5,225 isolates referred by 10 medical centers. The antibiotic test panel included ertapenem, imipenem, meropenem, ampicillin-sulbactam, piperacillin-tazobactam, cefoxitin, clindamycin, moxifloxacin, tigecycline, chloramphenicol, and metronidazole. From 1997 to 2004 there were decreases in the geometric mean (GM) MICs of imipenem, meropenem, piperacillin-tazobactam, and cefoxitin for many of the species within the group. B. distasonis showed the highest rates of resistance to most of the β-lactams. B. fragilis, B. ovatus, and B. thetaiotaomicron showed significantly higher GM MICs and rates of resistance to clindamycin over time. The rate of resistance to moxifloxacin of B. vulgatus was very high (MIC range for the 8-year study period, 38% to 66%). B. fragilis, B. ovatus, and B. distasonis and other Bacteroides spp. exhibited significant increases in the rates of resistance to moxifloxacin over the 8 years. Resistance rates and GM MICs for tigecycline were low and stable during the 5-year period over which this agent was studied. All isolates were susceptible to chloramphenicol (MICs < 16 μg/ml). In 2002, one isolate resistant to metronidazole (MIC = 64 μg/ml) was noted. These data indicate changes in susceptibility over time; surprisingly, some antimicrobial agents are more active now than they were 5 years ago.

Risk factors for surgical-site infections following cesarean section.

ABSTRACT Ureaplasma, spp. Mycoplasma genitalium, and Mycoplasma hominis are associated with infection of the genitourinary tract, reproductive failure, and neonatal morbidity and mortality. We have developed a multiplex PCR for the detection of these Mycoplasmatales in a single amplification reaction. The analytical sensitivities of this assay were 10.8, 10.8, and 8.8 CFU for each organism, respectively. This multiplex PCR was compared to culture on 26 cervical swabs, 2 vaginal swabs, 4 female urine specimens, 49 semen samples, 2 male urine specimens, and 1 nonspecified sample. A total of 21 specimens were culture positive (25%); 17 of these were PCR positive. An additional 11 specimens were PCR positive but culture negative. Of the 21 culture-positive specimens, 17 (81%) grew Ureaplasma spp. and 4 (19%) grew Mycoplasma spp. Of the 28 PCR-positive specimens, Ureaplasma spp. was detected in 23 (82%), M. hominis was detected in 3 (11%), and both were detected in 2 (7%). In a confirmatory analysis, all samples were tested by amplification of a second target of the ureaplasma genome. True-positive cases were defined as a positive result by culture or by both amplification assays. The multiplex PCR detected organisms in 26 of the 30 true-positive specimens, as well as in 2 other specimens. Based on a 36% prevalence of infection, the sensitivity, specificity, and positive and negative predictive values of multiplex PCR analyses were 87, 96, 94, and 93%, respectively. Multiplex PCR offers a rapid, sensitive, and easy method to detect genital mycoplasmas.

Lessons Learned from the Anaerobe Survey: Historical Perspective and Review of the Most Recent Data (2005–2007)

OBJECTIVE To identify risk factors associated with surgical-site infections (SSIs) following cesarean sections. DESIGN Prospective cohort study. SETTING High-risk obstetrics and neonatal tertiary-care center in upstate New York. PATIENTS Population-based sample of 765 patients who underwent cesarean sections at our facility during 6-month periods each year from 1996 through 1998. METHODS Prospective surgical-site surveillance was conducted using methodology of the National Nosocomial Infections Surveillance System. Infections were identified during admission, within 30 days following the cesarean section, by readmission to the hospital or by a postdischarge survey. RESULTS Multiple logistic-regression analysis identified four factors independently associated with an increased risk of SSI following cesarean section: absence of antibiotic prophylaxis (odds ratio [OR], 2.63; 95% confidence interval [CI95], 1.50-4.6; P=.008); surgery time (OR, 1.01; CI95, 1.00-1.02; P=.04); <7 prenatal visits (OR, 3.99; CI95, 1.74-9.15; P=.001); and hours of ruptured membranes (OR, 1.02; CI95, 1.01-1.03; P=.04). Patients given antibiotic prophylaxis had significantly lower infection rates than patients who did not receive antibiotic prophylaxis (P=02), whether or not active labor or ruptured membranes were present. CONCLUSION Among the variables identified as risk factors for SSI, only two have the possibility to be changed through interventions. Antibiotic prophylaxis would benefit all cesarean patients regardless of active labor or ruptured membranes and would decrease morbidity and length of stay. Womens healthcare professionals also must continue to encourage pregnant women to start prenatal visits early in the pregnancy and to maintain scheduled visits throughout the pregnancy to prevent perinatal complications, including postoperative infection.

National Survey on the Susceptibility of Bacteroides fragilis Group: Report and Analysis of Trends for 1997-2000

BACKGROUND The rationale and lessons learned through the evolution of the National Survey for the Susceptibility of Bacteroides fragilis Group from its initiation in 1981 through 2007 are reviewed here. The survey was conceived in 1980 to track emerging antimicrobial resistance in Bacteroides species. METHODS Data from the last 11 years of the survey (1997-2007), including 6574 isolates from 13 medical centers, were analyzed for in vitro antimicrobial resistance to both frequently used and newly developed anti-anaerobic agents. The minimum inhibitory concentrations of the antibiotics were determined using agar dilution in accordance with Clinical and Laboratory Standards Institute recommendations. RESULTS The analyses revealed that the carbapenems (imipenem, meropenem, ertapenem, and doripenem) and piperacillin-tazobactam were the most active agents against these pathogens, with resistance rates of 0.9%-2.3%. In the most recent 3 years of the survey (2005-2007), resistance to some agents was shown to depend on the species, such as ampicillin-sulbactam against Bacteroides distasonis (20.6%) and tigecycline against Bacteroides uniformis and Bacteroides eggerthii ( approximately 7%). Very high resistance rates (>50%) were noted for moxifloxacin and trovafloxacin, particularly against Bacteroides vulgatus. During that period of study, non-B. fragilis Bacteroides species had >40% resistance to clindamycin. Metronidazole-resistant Bacteroides strains were also first reported during that period. CONCLUSIONS In summary, resistance to antibiotics was greater among non-B. fragilis Bacteroides species than among B. fragilis and was especially greater among species with a low frequency of isolation, such as Bacteroides caccae and B. uniformis. The emergence of resistance among the non-B. fragilis Bacteroides species underscores the need for speciation of B. fragilis group isolates and for clinicians to be aware of associations between species and drug resistance.

The Etiology of Severe Acute Gastroenteritis Among Adults Visiting Emergency Departments in the United States

The results of a multicenter US survey using the National Committee for Clinical Laboratory Standards currently recommended methodology for measuring in vitro susceptibility of 2673 isolates of Bacteroides fragilis group species were compared from 1997 to 2000. The test panel consisted of 14 antibiotics: 3 carbapenems, 3 beta-lactam-beta-lactamase inhibitors, 3 cephamycins, 2 fluoroquinolones, clindamycin, chloramphenicol, and metronidazole. Declines in the geometric mean minimum inhibitory concentrations were seen with imipenem, meropenem, ampicillin-sulbactam, and the cephamycins. Increased geometric means were observed with the fluoroquinolones and were usually accompanied by an increase in resistance rates. Bacteroides distasonis shows the highest resistance rates among beta-lactam antibiotics, whereas Bacteroides vulgatus shows the highest resistance levels among fluoroquinolones. B. fragilis shows the lowest resistance rates for all antibiotics. All strains were susceptible to chloramphenicol and metronidazole concentrations <8 microgram/mL. The data underscore the need for species identification and continued surveillance to monitor resistance patterns.

Prevention of gram-positive sepsis in neonates weighing less than 1500 grams

BACKGROUND Acute gastroenteritis (AGE) remains a common cause of clinic visits and hospitalizations in the United States, but the etiology is rarely determined. METHODS We performed a prospective, multicenter emergency department-based study of adults with AGE. Subjects were interviewed on presentation and 3-4 weeks later. Serum samples, rectal swab specimens, and/or whole stool specimens were collected at presentation, and serum was collected 3-4 weeks later. Fecal specimens were tested for a comprehensive panel of viral, bacterial, and parasitic pathogens; serum was tested for calicivirus antibodies. RESULTS Pathogens were detected in 25% of 364 subjects, including 49% who provided a whole stool specimen. The most commonly detected pathogens were norovirus (26%), rotavirus (18%), and Salmonella species (5.3%). Pathogens were detected significantly more often from whole stool samples versus a rectal swab specimen alone. Nine percent of subjects who provided whole stool samples had >1 pathogen identified. CONCLUSIONS Viruses, especially noroviruses, play a major role as agents of severe diarrhea in adults. Further studies to confirm the unexpectedly high prevalence of rotaviruses and to explore the causes of illness among patients from whom a pathogen cannot be determined are needed. Studies of enteric pathogens should require the collection of whole stool samples.