Updating breakpoints in the United States: a summary from the ASM Clinical Microbiology Open 2022.

Accurate antimicrobial susceptibility testing (AST) and reporting are essential for guiding appropriate therapy for patients and direction for public health prevention and control actions. A critical feature of AST reporting is the interpretation of AST results using clinical breakpoints for reporting as susceptible, susceptible-dose dependent, intermediate, or resistant. Breakpoints are subject to continuous adjustment and updating to best reflect current clinical data. These breakpoint changes can benefit patients and public health only if adopted in a timely manner. A recent survey identified that up to 70% of College of American Pathologists (CAP)-accredited U.S. laboratories and 45% of CAP-accredited laboratories outside the U.S. use various obsolete clinical breakpoints to interpret AST results to guide patient care. The reason for the ongoing use of obsolete breakpoints is multifactorial, including barriers encountered by laboratories, commercial AST device manufacturers, standards development organizations, and regulatory bodies alike. To begin to address this important patient safety issue, CAP implemented checklist requirements for CAP-accredited laboratories to ensure up-to-date clinical breakpoint use. Furthermore, the topic was discussed at the June 2022 American Society for Microbiology Clinical Microbiology Open (CMO) with various stakeholders to identify potential solutions. This minireview summarizes the breakpoint setting process in the U.S. and highlights solutions to close the gap between breakpoint revisions and implementation in clinical and public health laboratories. Solutions discussed include clarification of data requirements and minimum inhibitory concentration only reporting for regulatory clearance of AST devices, clinical data generation to close breakpoints gaps, advocacy, education, and greater dialogue between stakeholders.

Assessing the in vitro impact of ceftazidime on aztreonam/avibactam susceptibility testing for highly resistant MBL-producing Enterobacterales.

BACKGROUND: Aztreonam/avibactam is a combination agent that shows promise in treating infections caused by highly antibiotic-resistant MBL-producing Enterobacterales. This combination can be achieved by combining two FDA-approved drugs: ceftazidime/avibactam and aztreonam. It is unknown whether ceftazidime in the combination ceftazidime/aztreonam/avibactam has a synergistic or antagonistic effect on the in vitro activity of aztreonam/avibactam by significantly increasing or decreasing the MIC. OBJECTIVES: To determine whether increasing ceftazidime concentrations affect the MICs of aztreonam/avibactam alone. METHODS: A custom 8 × 8 chequerboard broth microdilution (BMD) panel was made using a digital dispenser (Hewlett-Packard, Corvallis, OR, USA). The panel included orthogonal 2-fold dilution series of aztreonam and ceftazidime ranging from 0.5 to 64 mg/L. Avibactam concentration was kept constant at 4 mg/L throughout the chequerboard. Thirty-seven Enterobacterales isolates from the CDC & FDA Antibiotic Resistance Isolate Bank or CDC's internal collection with intermediate or resistant interpretations to aztreonam and ceftazidime/avibactam were included for testing. All isolates harboured at least one of the following MBL genes: blaIMP, blaNDM or blaVIM. RESULTS: Regardless of the concentration of ceftazidime, aztreonam/avibactam with ceftazidime MICs for all 37 isolates were within one 2-fold doubling dilution of the aztreonam/avibactam MIC. CONCLUSIONS: Ceftazidime, in the combination ceftazidime/avibactam/aztreonam, did not affect the in vitro activity of aztreonam/avibactam in this sample of isolates. These findings can help assure clinical and public health laboratories that testing of aztreonam/avibactam by BMD can act as a reliable surrogate test when the combination of ceftazidime/avibactam and aztreonam is being considered for treatment of highly antibiotic-resistant MBL-producing Enterobacterales.

Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing Position Statements on Polymyxin B and Colistin Clinical Breakpoints.

Recent data on polymyxin pharmacokinetics, pharmacodynamics, toxicity, and clinical outcomes suggest these agents have limited clinical utility. Pharmacokinetics-pharmacodynamics data show a steady-state concentration of 2 µg/mL is required for killing bacteria with colistin minimum inhibitory concentrations of 2 µg/mL. Less than 50% of patients with normal renal function achieve this exposure, and it is associated with high risk of nephrotoxicity. This exposure does not achieve bacterial stasis in pneumonia models. Randomized and observational studies consistently demonstrate increased mortality for polymyxins compared with alternative agents. The Clinical and Laboratory Standards Institute (CLSI) and European Committee on Antimicrobial Susceptibility Testing (EUCAST) are 2 global organizations that establish interpretive criteria for in vitro susceptibility data. CLSI has recently taken the step to eliminate the "susceptible" interpretive category for the polymyxins, whereas EUCAST maintains this interpretive category. This viewpoint describes the opinions of these organizations and the data that were used to inform their perspectives.

Rationale for a Neisseria gonorrhoeae Susceptible-only Interpretive Breakpoint for Azithromycin.

BACKGROUND: Azithromycin (AZI) is recommended with ceftriaxone (CRO) for treatment of uncomplicated gonococcal urethritis and cervicitis in the United States, and an AZI-susceptibility breakpoint is needed. Neither the Food and Drug Administration (FDA) nor the Clinical and Laboratory Standards Institute (CLSI) has set interpretive breakpoints for AZI susceptibility. As a result, AZI antimicrobial susceptibility testing (AST) cannot be interpreted using recognized standards. This has contributed to increasingly unavailable clinical laboratory AST, although gonorrhea is on the rise with >550 000 US gonorrhea cases reported to the Centers for Disease Control and Prevention in 2017, the highest number of cases since 1991. METHODS: This article summarizes the rationale data reviewed by the CLSI in June 2018. RESULTS: The CLSI decided to set a susceptible-only interpretive breakpoint at the minimum inhibitory concentration of ≤1 µg/mL. This is also the epidemiological cutoff value (ECV) (ie, the end of the wild-type susceptibility distribution). This breakpoint presumes that AZI (1-g single dose) is used in an approved regimen that includes an additional antimicrobial agent (ie, CRO 250 mg, intramuscular single dose). CONCLUSIONS: Having a breakpoint can improve patient care and surveillance and allow future development and FDA regulatory approval of modernized AST to guide treatment. The breakpoint coincides with a European Committee on AST decision to remove previously established, differing AZI breakpoints and use the ECV as guidance for testing. The CLSI breakpoint is now the recognized standard that defines AZI susceptibility for gonococcal infections.

Validation of Aztreonam-Avibactam Susceptibility Testing Using Digitally Dispensed Custom Panels.

Aztreonam-avibactam is a combination antimicrobial agent with activity against carbapenemase-producing Enterobacteriaceae (CPE) with metallo-ß-lactamases (MßLs). Although aztreonam-avibactam is not yet approved by the U.S. Food and Drug Administration (FDA), clinicians can administer this combination by using two FDA-approved drugs: aztreonam and ceftazidime-avibactam. This combination of drugs is recommended by multiple experts for treatment of serious infections caused by MßL-producing CPE. At present, in vitro antimicrobial susceptibility testing (AST) of aztreonam-avibactam is not commercially available; thus, most clinicians receive no laboratory-based guidance that can support consideration of aztreonam-avibactam for serious CPE infections. Here, we report our internal validation for aztreonam-avibactam AST by reference broth microdilution (BMD) according to Clinical and Laboratory Standards Institute (CLSI) guidelines. The validation was performed using custom frozen reference BMD panels prepared in-house at the Centers for Disease Control and Prevention (CDC). In addition, we took this opportunity to evaluate a new panel-making method using a digital dispenser, the Hewlett Packard (HP) D300e. Our studies demonstrate that the performance characteristics of digitally dispensed panels were equivalent to those of conventionally prepared frozen reference BMD panels for a number of drugs, including aztreonam-avibactam. We found the HP D300e digital dispenser to be easy to use and to provide the capacity to prepare complex drug panels. Our findings will help other clinical and public health laboratories implement susceptibility testing for aztreonam-avibactam.

Harnessing alternative sources of antimicrobial resistance data to support surveillance in low-resource settings.

One of the most pressing challenges facing the global surveillance of antimicrobial resistance (AMR) is the generation, sharing, systematic analysis and dissemination of data in low-resource settings. Numerous agencies and initiatives are working to support the development of globally distributed microbiology capacity, but the routine generation of a sustainable flow of reliable data will take time to establish before it can deliver a clinical and public health impact. By contrast, there are a large number of pharma- and academia-led initiatives that have generated a wealth of data on AMR and drug-resistant infections in low-resource settings, together with high-volume data generation by private laboratories. Here, we explore how untapped sources of data could provide a short-term solution that bridges the gap between now and the time when routine surveillance capacity will have been established and how this could continue to support surveillance efforts in the future. We discuss the benefits and limitations of data generated by these sources, the mechanisms and barriers to making this accessible and how academia and pharma might support the development of laboratory and analytical capacity. We provide key actions that will be required to harness these data, including: a mapping exercise; creating mechanisms for data sharing; use of data to support national action plans; facilitating access to and use of data by the WHO Global Antimicrobial Resistance Surveillance System; and innovation in data capture, analysis and sharing.

FDA-CDC Antimicrobial Resistance Isolate Bank: a Publicly Available Resource To Support Research, Development, and Regulatory Requirements.

The FDA-CDC Antimicrobial Resistance Isolate Bank was created in July 2015 as a publicly available resource to combat antimicrobial resistance. It is a curated repository of bacterial isolates with an assortment of clinically important resistance mechanisms that have been phenotypically and genotypically characterized. In the first 2 years of operation, the bank offered 14 panels comprising 496 unique isolates and had filled 486 orders from 394 institutions throughout the United States. New panels are being added.

Vital Signs: Containment of Novel Multidrug-Resistant Organisms and Resistance Mechanisms - United States, 2006-2017.

BACKGROUND: Approaches to controlling emerging antibiotic resistance in health care settings have evolved over time. When resistance to broad-spectrum antimicrobials mediated by extended-spectrum ß-lactamases (ESBLs) arose in the 1980s, targeted interventions to slow spread were not widely promoted. However, when Enterobacteriaceae with carbapenemases that confer resistance to carbapenem antibiotics emerged, directed control efforts were recommended. These distinct approaches could have resulted in differences in spread of these two pathogens. CDC evaluated these possible changes along with initial findings of an enhanced antibiotic resistance detection and control strategy that builds on interventions developed to control carbapenem resistance. METHODS: Infection data from the National Healthcare Safety Network from 2006-2015 were analyzed to calculate changes in the annual proportion of selected pathogens that were nonsusceptible to extended-spectrum cephalosporins (ESBL phenotype) or resistant to carbapenems (carbapenem-resistant Enterobacteriaceae [CRE]). Testing results for CRE and carbapenem-resistant Pseudomonas aeruginosa (CRPA) are also reported. RESULTS: The percentage of ESBL phenotype Enterobacteriaceae decreased by 2% per year (risk ratio [RR] = 0.98, p<0.001); by comparison, the CRE percentage decreased by 15% per year (RR = 0.85, p<0.01). From January to September 2017, carbapenemase testing was performed for 4,442 CRE and 1,334 CRPA isolates; 32% and 1.9%, respectively, were carbapenemase producers. In response, 1,489 screening tests were performed to identify asymptomatic carriers; 171 (11%) were positive. CONCLUSIONS: The proportion of Enterobacteriaceae infections that were CRE remained lower and decreased more over time than the proportion that were ESBL phenotype. This difference might be explained by the more directed control efforts implemented to slow transmission of CRE than those applied for ESBL-producing strains. Increased detection and aggressive early response to emerging antibiotic resistance threats have the potential to slow further spread.

Use of a Clinical Decision Support System Alert to Prevent Supratherapeutic Vancomycin Concentrations.

Background: Alerts issued by clinical decision support systems (CDSS) may be useful to identify and prevent the occurrence of acute kidney injury among patients on nephrotoxic drugs, particularly vancomycin. Objective: The purpose of this instructive study was to determine the effectiveness of using a pharmacist-run CDSS alert of early serum creatinine increases in patients receiving intravenous vancomycin to decrease the proportion of severely elevated vancomycin concentrations. Methods: This was a retrospective study of a prospectively reviewed CDSS alert that triggered in patients with an increase in serum creatinine by 25% from baseline within 24 hours. Severely elevated vancomycin concentrations were divided into a control group (before alert implementation) and a study group (after alert implementation) and considered for study inclusion. The proportion of severely elevated vancomycin concentrations (ie, >30 mg/L) were collected in the control and study groups. Results: There were 1290 and 1501 vancomycin concentrations in the control group and the study group, respectively. A total of 696 CDSS alerts triggered during the study period. The proportion of severely elevated vancomycin troughs decreased from 5.3% (n = 68, median = 36.6 mg/L, interquartile range = 33.75-43.2 mg/L) in the control group to 3.7% (n = 55, median = 34.7 mg/L, interquartile range = 31.3-39.3 mg/L) in the study group. This reflects a statistically significant decrease in the proportion of severely elevated vancomycin concentrations (P = .04). Conclusion: Overall, this instructive analysis on a novel use of CDSS software suggests that the implementation of an alert based on early detection of serum creatinine changes led to a significant decrease in the proportion of severely elevated serum vancomycin concentrations.

pSK41-like plasmid is necessary for Inc18-like vanA plasmid transfer from Enterococcus faecalis to Staphylococcus aureus in vitro.

Vancomycin-resistant Staphylococcus aureus (VRSA) is thought to result from the in vivo conjugative transfer of a vanA plasmid from an Enterococcus sp. to S. aureus. We studied bacterial isolates from VRSA cases that occurred in the United States to identify microbiological factors which may contribute to this plasmid transfer. First, vancomycin-susceptible, methicillin-resistant S. aureus (MRSA) isolates from five VRSA cases were tested for their ability to accept foreign DNA by conjugation in mating experiments with Enterococcus faecalis JH2-2 containing pAM378, a pheromone-response conjugative plasmid. All of the MRSA isolates accepted the plasmid DNA with similar transfer efficiencies (∼10(-7)/donor CFU) except for one isolate, MRSA8, for which conjugation was not successful. The MRSA isolates were also tested as recipients in mating experiments between an E. faecalis isolate with an Inc18-like vanA plasmid that was isolated from a VRSA case patient. Conjugative transfer was successful for 3/5 MRSA isolates. Successful MRSA recipients carried a pSK41-like plasmid, a staphylococcal conjugative plasmid, whereas the two unsuccessful MRSA recipients did not carry pSK41. The transfer of a pSK41-like plasmid from a successful MRSA recipient to the two unsuccessful recipients resulted in conjugal transfer of the Inc18-like vanA plasmid from E. faecalis at a frequency of 10(-7)/recipient CFU. In addition, conjugal transfer could be achieved for pSK41-negative MRSA in the presence of a cell-free culture filtrate from S. aureus carrying a pSK41-like plasmid at a frequency of 10(-8)/recipient CFU. These results indicated that a pSK41-like plasmid can facilitate the transfer of an Inc18-like vanA plasmid from E. faecalis to S. aureus, possibly via an extracellular factor produced by pSK41-carrying isolates.

Development of doxycycline MIC and disk diffusion interpretive breakpoints and revision of tetracycline breakpoints for Streptococcus pneumoniae.

A study was performed to derive susceptibility testing interpretive breakpoints for doxycycline with Streptococcus pneumoniae and to reassess breakpoints for tetracycline using the requirements defined in Clinical and Laboratory Standards Institute (CLSI) document M23-A3. Tetracycline and doxycycline MICs and disk diffusion zone sizes were determined on 189 isolates selected from the 2009-2010 CDC Active Bacterial Core surveillance strain collection according to the testing methods described in CLSI documents M07-A8 and M02-A10. Tetracycline and doxycycline MICs and zones were compared to each other directly, and the reproducibility of MICs and zone diameters for both drugs was determined. Scattergrams of tetracycline MICs versus corresponding zone diameters and doxycycline MICs versus zones were prepared, and analysis indicated that the present CLSI tetracycline MIC and disk breakpoints did not fit the susceptibility data for doxycycline. Doxycycline was 1 to 3 dilutions more potent than tetracycline, especially in strains harboring the tetM resistance determinant. tetM was detected in ≥ 90% of isolates having tetracycline MICs of ≥ 4 µg/ml and in ≥ 90% with doxycycline MICs of ≥ 1. Limited pharmacokinetic/pharmacodynamic (PK/PD) data coupled with application of the error-rate bounded method of analysis suggested doxycycline-susceptible breakpoints of either ≤ 0.25 µg/ml or ≤ 0.5 µg/ml, with intermediate and resistant breakpoints 1 and 2 dilutions higher, respectively. The disk diffusion zone diameter correlates were susceptible at ≥ 28 mm, intermediate at 25 to 27 mm, and resistant at ≤ 24 mm. Revised lower tetracycline MIC breakpoints were suggested as susceptible at ≤ 1 µg/ml, intermediate at 2 µg/ml, and resistant at ≥ 4 µg/ml. Suggested tetracycline disk diffusion zones were identical to those of doxycycline.

The impact of anti-infective drug shortages on hospitals in the United States: trends and causes.

Anti-infective shortages pose significant logistical and clinical challenges to hospitals and may be considered a public health emergency. Anti-infectives often represent irreplaceable life-saving treatments. Furthermore, few new agents are available to treat increasingly prevalent multidrug-resistant pathogens. Frequent anti-infective shortages have substantially altered patient care and may lead to inferior patient outcomes. Because many of the shortages stem from problems with manufacturing and distribution, federal legislation has been introduced but not yet enacted to provide oversight for the adequate supply of critical medications. At the local level, hospitals should develop strategies to anticipate the impact and extent of shortages, to identify therapeutic alternatives, and to mitigate potential adverse outcomes. Here we describe the scope of recent anti-infective shortages in the United States and explore the reasons for inadequate drug supply.

Carbapenem-resistant Enterobacteriaceae: epidemiology and prevention.

Over the past 10 years, dissemination of Klebsiella pneumoniae carbapenemase (KPC) has led to an increase in the prevalence of carbapenem-resistant Enterobacteriaceae (CRE) in the United States. Infections caused by CRE have limited treatment options and have been associated with high mortality rates. In the previous year, other carbapenemase subtypes, including New Delhi metallo-ß-lactamase, have been identified among Enterobacteriaceae in the United States. Like KPC, these enzymes are frequently found on mobile genetic elements and have the potential to spread widely. As a result, preventing both CRE transmission and CRE infections have become important public health objectives. This review describes the current epidemiology of CRE in the United States and highlights important prevention strategies.

Comparison of detection methods for heteroresistant vancomycin-intermediate Staphylococcus aureus, with the population analysis profile method as the reference method.

Staphylococcus aureus clinical isolates with vancomycin MICs of 2 µg/ml have been associated with vancomycin therapeutic failure and the heteroresistant vancomycin-intermediate S. aureus (hVISA) phenotype. A population analysis profile (PAP) with an area under the curve (AUC) ratio of ≥ 0.9 for the AUC of the clinical isolate versus the AUC for hVISA strain Mu3 is most often used for determining hVISA, but it is time-consuming and labor-intensive. A collection of 140 MRSA blood isolates with vancomycin MICs of 2 µg/ml by reference broth microdilution and screened for hVISA using PAP-AUC (21/140 [15%] hVISA) were tested by additional methods to detect hVISA. The methods included (i) Etest macromethod using vancomycin and teicoplanin test strips, brain heart infusion (BHI) agar, and a 2.0 McFarland inoculum; (ii) Etest glycopeptide resistance detection (GRD) using vancomycin-teicoplanin double-sided gradient test strips on Mueller-Hinton agar (MHA) with 5% sheep blood and a 0.5 McFarland inoculum; and (iii) BHI screen agar plates containing 4 µg/ml vancomycin and 16 g/liter casein using 0.5 and 2.0 McFarland inocula. Each method was evaluated using PAP-AUC as the reference method. The sensitivity of each method for detecting hVISA was higher when the results were read at 48 h. The Etest macromethod was 57% sensitive and 96% specific, Etest GRD was 57% sensitive and 97% specific, and BHI screen agar was 90% sensitive and 95% specific with a 0.5 McFarland inoculum and 100% sensitive and 68% specific with a 2.0 McFarland inoculum. BHI screen agar with 4 µg/ml vancomycin and casein and a 0.5 McFarland inoculum had the best sensitivity and specificity combination, was easy to perform, and may be useful for clinical detection of hVISA.

Comparison of Etest method with reference broth microdilution method for antimicrobial susceptibility testing of Yersinia pestis.

The utility of Etest for antimicrobial susceptibility testing of Yersinia pestis was evaluated in comparison with broth microdilution and disk diffusion for eight agents. Four laboratories tested 26 diverse strains and found Etest to be reliable for testing antimicrobial agents used to treat Y. pestis, except for chloramphenicol and trimethoprim-sulfamethoxazole. Disk diffusion testing is not recommended.

Genetic factors associated with elevated carbapenem resistance in KPC-producing Klebsiella pneumoniae.

In the United States, the most prevalent mechanism of carbapenem resistance among Enterobacteriaceae is the production of a Klebsiella pneumoniae carbapenemase (KPC). KPC-producing isolates often exhibit a range of carbapenem MICs. To better understand the factors that contribute to overall carbapenem resistance, we analyzed 27 KPC-producing K. pneumoniae isolates with different levels of carbapenem resistance, 11 with low-level (i.e., meropenem or imipenem MIC ≤ 4 µg/ml), 2 with intermediate-level (i.e., meropenem and imipenem MIC = 8 µg/ml), and 14 with high-level (i.e., imipenem or meropenem MIC ≥ 16 µg/ml) carbapenem resistance, that were received from throughout the United States. Among 14 isolates that exhibited high-level carbapenem resistance, Western blot analysis indicated that 10 produced an elevated amount of KPC. These isolates either contained an increased bla(KPC) gene copy number (n = 3) or had deletions directly upstream of the bla(KPC) gene (n = 7). Four additional isolates lacked elevated KPC production but had high-level carbapenem resistance. Porin sequencing analysis identified 22 isolates potentially lacking a functional OmpK35 and three isolates potentially lacking a functional OmpK36. The highest carbapenem MICs were found in two isolates that lacked both functioning porins and produced elevated amounts of KPC. The 11 isolates with low-level carbapenem resistance contained neither an upstream deletion nor increased bla(KPC) copy number. These results suggest that both bla(KPC) copy number and deletions in the upstream genetic environment affect the level of KPC production and may contribute to high-level carbapenem resistance in KPC-producing K. pneumoniae, particularly when coupled with OmpK36 porin loss.

Emergence of resistance among USA300 methicillin-resistant Staphylococcus aureus isolates causing invasive disease in the United States.

USA300 methicillin-resistant Staphylococcus aureus (MRSA) isolates are usually resistant only to oxacillin, erythromycin, and, increasingly, levofloxacin. Of these, oxacillin and levofloxacin resistances are chromosomally encoded. Plasmid-mediated clindamycin, mupirocin, and/or tetracycline resistance has been observed among USA300 isolates, but these descriptions were limited to specific patient populations or isolated occurrences. We examined the antimicrobial susceptibilities of invasive MRSA isolates from a national surveillance population in order to identify USA300 isolates with unusual, possibly emerging, plasmid-mediated antimicrobial resistance. DNA from these isolates was assayed for the presence of resistance determinants and the presence of a pSK41-like conjugative plasmid. Of 823 USA300 isolates, 72 (9%) were tetracycline resistant; 69 of these were doxycycline susceptible and tetK positive, and 3 were doxycycline resistant and tetM positive. Fifty-one (6.2%) isolates were clindamycin resistant and ermC positive; 22 (2.7%) isolates were high-level mupirocin resistant (mupA positive); 5 (0.6%) isolates were trimethoprim-sulfamethoxazole (TMP-SMZ) resistant, of which 4 were dfrA positive; and 7 (0.9%) isolates were gentamicin resistant and aac6'-aph2'' positive. Isolates with pSK41-like plasmids (n = 24) were positive for mupA (n = 19), dfrA (n = 6), aac6'-aph2'' (n = 6), tetM (n = 2), and ermC (n = 8); 20 pSK41-positive isolates were positive for two or more resistance genes. Conjugative transfer of resistance was demonstrated between four gentamicin- and mupirocin-resistant and three gentamicin- and TMP-SMZ-resistant USA300 isolates; transconjugants harbored a single pSK41-like plasmid, which was PCR positive for aac6'-aph2'' and either mupA and/or dfrA. USA300 and USA100 isolates from the same state with identical resistance profiles contained pSK41-like plasmids with indistinguishable restriction and Southern blot profiles, suggesting horizontal plasmid transfer between USA100 and USA300 isolates.

Dissemination of an Enterococcus Inc18-Like vanA plasmid associated with vancomycin-resistant Staphylococcus aureus.

Of the 9 vancomycin-resistant Staphylococcus aureus (VRSA) cases reported to date in the literature, 7 occurred in Michigan. In 5 of the 7 Michigan VRSA cases, an Inc18-like vanA plasmid was identified in the VRSA isolate and/or an associated vancomycin-resistant Enterococcus (VRE) isolate from the same patient. This plasmid may play a critical role in the emergence of VRSA. We studied the geographical distribution of the plasmid by testing 1,641 VRE isolates from three separate collections by PCR for plasmid-specific genes traA, repR, and vanA. Isolates from one collection (phase 2) were recovered from surveillance cultures collected in 17 hospitals in 13 states. All VRE isolates from 2 Michigan institutions (n = 386) and between 60 and 70 VRE isolates (n = 883) from the other hospitals were tested. Fifteen VRE isolates (3.9%) from Michigan were positive for an Inc18-like vanA plasmid (9 E. faecalis [12.5%], 3 E. faecium [1.0%], 2 E. avium, and 1 E. raffinosus). Six VRE isolates (0.6%) from outside Michigan were positive (3 E. faecalis [2.7%] and 3 E. faecium [0.4%]). Of all E. faecalis isolates tested, 6.0% were positive for the plasmid, compared to 0.6% for E. faecium and 3.0% for other spp. Fourteen of the 15 plasmid-positive isolates from Michigan had the same Tn1546 insertion site location as the VRSA-associated Inc18-like plasmid, whereas 5 of 6 plasmid-positive isolates from outside Michigan differed in this characteristic. Most plasmid-positive E. faecalis isolates demonstrated diverse patterns by PFGE, with the exception of three pairs with indistinguishable patterns, suggesting that the plasmid is mobile in nature. Although VRE isolates with the VRSA-associated Inc18-like vanA plasmid were more common in Michigan, they remain rare. Periodic surveillance of VRE isolates for the plasmid may be useful in predicting the occurrence of VRSA.