Claudia K. Hoyen

Effect of antibiotic therapy on the density of vancomycin-resistant enterococci in the stool of colonized patients.

BACKGROUND Colonization and infection with vancomycin-resistant enterococci have been associated with exposure to antibiotics that are active against anaerobes. In mice that have intestinal colonization with vancomycin-resistant enterococci, these agents promote high-density colonization, whereas antibiotics with minimal antianaerobic activity do not. METHODS We conducted a seven-month prospective study of 51 patients who were colonized with vancomycin-resistant enterococci, as evidenced by the presence of the bacteria in stool. We examined the density of vancomycin-resistant enterococci in stool during and after therapy with antibiotic regimens and compared the effect on this density of antianaerobic agents and agents with minimal antianaerobic activity. In a subgroup of 10 patients, cultures of environmental specimens (e.g., from bedding and clothing) were obtained. RESULTS During treatment with 40 of 42 antianaerobic-antibiotic regimens (95 percent), high-density colonization with vancomycin-resistant enterococci was maintained (mean [+/-SD] number of organisms, 7.8+/-1.5 log per gram of stool). The density of colonization decreased after these regimens were discontinued. Among patients who had not received antianaerobic antibiotics for at least one week, 10 of 13 patients who began such regimens had an increase in the number of organisms of more than 1.0 log per gram (mean increase, 2.2 log per gram), whereas among 10 patients who began regimens of antibiotics with minimal antianaerobic activity, there was a mean decrease in the number of enterococci of 0.6 log per gram (P=0.006 for the difference between groups). When the density of vancomycin-resistant enterococci in stool was at least 4 log per gram, 10 of 12 sets of cultures of environmental specimens had at least one positive sample, as compared with 1 of 9 sets from patients with a mean number of organisms in stool of less than 4 log per gram (P=0.002). CONCLUSIONS For patients with vancomycin-resistant enterococci in stool, treatment with antianaerobic antibiotics promotes high-density colonization. Limiting the use of such agents in these patients may help decrease the spread of vancomycin-resistant enterococci.

High-Level Expression of Chromosomally Encoded SHV-1 β-Lactamase and an Outer Membrane Protein Change Confer Resistance to Ceftazidime and Piperacillin- Tazobactam in a Clinical Isolate of Klebsiella pneumoniae

ABSTRACT We describe Klebsiella pneumoniae 15571, a clinical isolate resistant to ceftazidime MIC = 32 μg/ml) and piperacillin-tazobactam (MICs = 1,024 and 128 μg/ml). K. pneumoniae 15571 expresses a single β-lactamase with a pI of 7.6. However, when cloned in a high-copy-number vector inEscherichia coli, this blaSHV-1gene did not confer resistance to ceftazidime, a spectrum consistent with the nucleotide sequence, which was nearly identical to those of previously described blaSHV-1 genes. Outer membrane protein (OMP) analysis of K. pneumoniae 15571 revealed a decrease in the quantity of a minor 45-kDa OMP in comparison to that in K. pneumoniae 44NR, a low-level ampicillin-resistant strain that also expresses a chromosomally determined blaSHV-1. Crude β-lactamase enzyme extracts from K. pneumoniae 15571 produced roughly 200-fold more β-lactamase activity than K. pneumoniae 44NR. Northern hybridization analysis revealed that this difference was explainable by quantifiable differences in transcription of theblaSHV-1 gene in the two strains. Primer extension analysis of blaSHV-1 mRNA fromK. pneumoniae 15571 and 44NR indicated that the transcriptional start sites were identical in the two strains. DNA sequencing of the promoter regions upstream of the ofblaSHV-1 open reading frames in the twoK. pneumoniae strains revealed an A→C change in the second position of the −10 region in K. pneumoniae 44NR compared to that in 15571. Site-directed mutagenesis of the clonedK. pneumoniae 15571 blaSHV-1, in which the A in the second position of the 15571 −10 region was changed to a C, resulted in a substantial lowering of the MIC of ampicillin. When the levels of β-lactamase enzyme expression inE. coli were compared, the blaSHV-1downstream of the altered −10 region produced 17-fold less β-lactamase enzyme. These results indicate that elevated levels of ceftazidime resistance can result from a combination of increased enzyme production and minor OMP changes and that levels of chromosomally encoded SHV-1 β-lactamase production can vary substantially with a single-base-pair change in promoter sequence.

Antianaerobic antibiotic therapy promotes overgrowth of antibiotic-resistant, gram-negative bacilli AND vancomycin-resistant enterococci in the stool of colonized patients

BACKGROUND AND OBJECTIVE Antianaerobic antibiotic therapy promotes persistent high-density growth of vancomycin-resistant enterococci (VRE) in the stool of colonized patients. We tested the hypothesis that antibiotic regimens with potent antianaerobic activity promote overgrowth of coexisting antibiotic-resistant, gram-negative bacilli in the stool of VRE-colonized patients. DESIGN Eight-month prospective study examining the effect of antibiotic therapy on the stool density of gram-negative bacilli resistant to ceftazidime, ciprofloxacin, or piperacillin/tazobactam. SETTING A Department of Veterans Affairs medical center including an acute care hospital and nursing home. PATIENTS All VRE-colonized patients with at least 3 stool samples available for analysis. RESULTS One-hundred forty stool samples were obtained from 37 study patients. Forty-nine (61%) of 80 stool samples obtained during therapy with an antianaerobic regimen were positive for an antibiotic-resistant, gram-negative bacillus, where-as only 14 (23%) of 60 samples obtained 4 or more weeks after completion of such therapy were positive (P < .001). Twenty-four (65%) of the 37 patients had one or more stool cultures positive for a gram-negative bacillus resistant to ciprofloxacin, ceftazidime, or piperacillin/tazobactam. The density of these organisms was higher during therapy with antianaerobic regimens than in the absence of such therapy for at least 2 weeks (mean +/- standard deviation, 5.6 +/- 1.4 and 3.9 +/- 0.71 log10 organisms/g; P < .001). CONCLUSION Limiting the use of antianaerobic antibiotics in VRE-colonized patients may reduce the density of colonization with coexisting antibiotic-resistant, gram-negative bacilli.

Factors That Predict Preexisting Colonization With Antibiotic-Resistant Gram-Negative Bacilli in Patients Admitted to a Pediatric Intensive Care Unit

Objective. To predict which patients hospitalized in a pediatric intensive care unit (ICU) are colonized with antibiotic-resistant Gram-negative rods on admission. Methods. Consecutive children admitted to a pediatric ICU over a 6-month period were entered into the study. A questionnaire soliciting information regarding the childs medical history and home environment was completed by the parent or guardian. Nasopharyngeal and rectal cultures were obtained on each of the first 3 days of ICU admission, and organisms resistant to ceftazidime or tobramycin were identified. Only clonally distinct organisms, as confirmed by pulsed field gel electrophoresis, were analyzed. The association between identification of colonization with an antibiotic-resistant Gram-negative rod within 3 days of ICU admission and factors included in the questionnaire was tested by χ2 or ttest. Results. In 64 (8.8%) of 727 admissions, an antibiotic-resistant Gram-negative bacillus was isolated within the first 3 ICU days. More than half were identified on the day of admission. Colonization was associated with two factors related to the patients medical history, namely, number of past ICU admissions (1.98 vs .87) and administration of intravenous antibiotics within the past 12 months (67.9% vs 28.2%). No association was found between colonization and exposure to oral antibiotics. In addition, factors related to the childs environment were also associated with presumed importation of an antibiotic-resistant Gram-negative rod into the ICU. Specifically, residence in a chronic care facility was strongly associated with colonization (28.3% vs 2.6%); exposure to a household contact who had been hospitalized in the past 12 months also predicted colonization (41.7% vs 18.5%). Conclusions. These data suggest that a profile can be established characterizing children colonized with resistant Gram-negative bacilli before admission to a pediatric ICU. Infection control measures may help to contain these potentially dangerous bacteria once they have been introduced into the unit.antibiotic resistance, nosocomial infections, Gram-negative aerobic bacteria, disease transmission, pediatric intensive care unit, pulsed field gel electrophoresis.

Recurrence of vancomycin-resistant Enterococcus stool colonization during antibiotic therapy

OBJECTIVE To test the hypothesis that antibiotic therapy may promote recurrence of vancomycin-resistant Enterococcus (VRE) stool colonization in patients who have previously had three consecutive negative stool cultures obtained at least 1 week apart. DESIGN One-year prospective cohort study examining the effect of antibiotic therapy on recurrence and density of VRE stool colonization in patients who have cleared colonization. Pulsed-field gel electrophoresis (PFGE) was performed to determine whether recurrent VRE strains were the same clone as the previous colonizing strain. SETTING A Department of Veterans Affairs medical center including an acute care hospital and nursing home. PATIENTS All patients with at least one stool culture positive for VRE who subsequently had three consecutive negative stool cultures obtained at least 1 week apart. RESULTS Of the 16 patients who cleared VRE colonization, 13 received antibiotic therapy during the study period. Eight (62%) of the 13 patients who received antibiotics developed recurrent high-density VRE stool colonization (range, 4.9 to 9.1 log10 colony-forming units per gram) during a course of therapy. Five patients had VRE strains available for PFGE analysis; recurrent strains were unrelated to the prior strain in 3 patients, closely related in 1 patient, and indistinguishable in 1 patient. CONCLUSIONS Antibiotic therapy may be associated with recurrent high-density VRE stool colonization in many patients who have previously had three consecutive negative stool cultures. These patients should be screened for recurrent stool colonization when antibiotic therapy is administered.

Geographic distribution of a large mobile element that transfers ampicillin and vancomycin resistance between Enterococcus faecium strains.

ABSTRACT In several clonally unrelated VanB-type vancomycin-resistantEnterococcus faecium strains, we demonstrated a common physical relationship between pbp5 and Tn5382as well as common mutations within pbp5. The majority of these strains transferred vancomycin and ampicillin resistance toE. faecium in vitro, suggesting the dissemination of similar transferable pbp5-vanB-containing mobile elements throughout the United States.

Effect of parenteral antibiotic administration on establishment of intestinal colonization in mice by Klebsiella pneumoniae strains producing extended-spectrum beta-lactamases

ABSTRACT A mouse model was used to test the hypothesis that antibiotics with activity against anaerobes promote overgrowth of extended-spectrum β-lactamase-producing Klebsiella pneumoniae strains in stool. Subcutaneous clindamycin consistently promoted establishment of high-density colonization, whereas piperacillin-tazobactam, ceftriaxone, and ceftazidime promoted colonization only when a large inoculum and/or more resistant strain was administered.

Colonization and infection with multiple nosocomial pathogens among patients colonized with vancomycin-resistant Enterococcus.

OBJECTIVE To test the hypothesis that patients colonized with vancomycin-resistant Enterococcus (VRE) have a higher frequency of colonization or infection with other nosocomial pathogens than do patients who are not colonized with VRE. DESIGN A rectal swab culture survey was conducted to determine the point-prevalence of stool colonization with ceftazidime-resistant gram-negative bacilli in hospitalized patients with or without VRE stool colonization. For a 6-month period, the frequency of Clostridium difficile diarrhea and isolation of antibiotic-resistant (ie, ceftazidime-, piperacillin/tazobactam-, levofloxacin-, or trimethoprim/sulfamethoxazole-resistant) gram-negative bacilli, methicillin-resistant Staphylococcus aureus (MRSA), and non-albicans Candida species from clinical specimens other than stool was examined. SETTING A Department of Veterans Affairs medical center. PATIENTS All patients hospitalized in the acute care facility and one nursing home unit during a 1-week period in February 2001. RESULTS VRE-colonized patients had a higher point-prevalence of rectal colonization with ceftazidime-resistant gram-negative bacilli than did patients not colonized with VRE (17% vs 4%; P = .026). During a 6-month period,the VRE-colonized patients were more likely to have Clostridium difficile-associated diarrhea (26% vs 2%; P = .001), MRSA infection (17% vs 4%; P = .017), or colonization or infection with gram-negative bacilli resistant to 4 different antibiotics. CONCLUSION VRE-colonized patients in our institution have a higher frequency of colonization or infection with other nosocomial pathogens than do patients who are not colonized with VRE. This suggests that isolation measures implemented to control VRE could help limit the dissemination of other, coexisting pathogens.

Use of real time pulsed field gel electrophoresis to guide interventions during a nursery outbreak of Serratia marcescens infection

BACKGROUND Pulsed field gel electrophoresis (PFGE) is a commercially available technique that can establish clonal relationships among many common hospital-derived organisms with a high degree of accuracy and can yield results in a sufficiently short time to guide interventions during an outbreak investigation. METHODS The CHEF Genomic Bacterial DNA Plug Kit (Bio-Rad) was applied to an unfolding nursery outbreak of Serratia marcescens infections according to the manufacturers guidelines. Bacterial genomic DNA was digested with XbaI or SpeI and separated on 1% agarose gels, and the isolates were grouped by restriction endonuclease patterns according to established standards. RESULTS S. marcescens was isolated from nine patients in an intensive care nursery during an 8-week period. Initial PFGE analysis performed after identification of the first eight patients, when closure of the nursery was imminent, revealed that the epidemic was caused by two groups of four isolates each. In both instances the group was geographically contained, and the nursery remained open. A second PFGE analysis indicated that a ninth S. marcescens isolate, recovered in Week 8, was genetically unrelated to the other two. Surveillance during an additional 6 weeks revealed no new cases, and the epidemic was declared over. No cases of invasive S. marcescens infection were identified during the subsequent 10 months. CONCLUSION Real-time PFGE determined that an apparent nursery outbreak of S. marcescens infection was, in fact, caused by three genetically distinct strains. This information allowed the nursery to remain open after other appropriate infection control measures had been imposed.

Undetected vancomycin-resistant Enterococcus stool colonization in a Veterans Affairs Hospital using a Clostridium difficile-focused surveillance strategy.

We examined the point prevalence of undetected vancomycin-resistant Enterococcus (VRE) stool colonization in an institution that screens stool samples submitted for Clostridium difficile testing. Of 112 patients not known to be colonized, 10 (9%) had rectal VRE colonization. A prospective algorithm was effective for identification of colonized patients.