Clinical challenges of nosocomial infections caused by antibiotic-resistant pathogens in pediatrics

Clinical Challenges of Nosocomial Infections Caused by Antibiotic-Resistant Pathogens in Pediatrics Alice Pong, MD, and John S. Bradley, MD Antibiotic resistance in nosocomial infections is an ever-increasing problem as health care institutions provide care for children with more complicated medical and surgical problems. Several mechanisms of antibiotic resistance are reviewed for both gram-negative and gram-positive nosocomial pathogens. These adaptive resistance mechanisms allow organisms to survive in an environment of extensive antibiotic use and result in clinically significant infections. Mobile genetic elements have facilitated the rapid spread of antibiotic resistance within and among species. The clinical challenge faced by many practitioners is to understand these mechanisms of antibiotic resistance and to develop strategies for successfully treating infection caused by resistant pathogens. Nosocomial outbreaks caused by resistant organisms are described, and an approach to empiric therapy based on presumed pathogens, site of infection, and local resistance patterns is discussed. © 2004 Elsevier Inc. All rights reserved.

W

ith improvements in medical technology and infrastructure, increased numbers of infants and children with a variety of medical and surgical problems from cancer to trauma to community-acquired infections now experience survival in pediatric critical care units, neonatal intensive care units (ICUs), and oncology/transplant units.1-3 Their care often requires complicated hospitalizations. Hospital-acquired infections have become a serious problem and have promoted the development of pediatric nosocomial infection surveillance programs.4-6 Nosocomial infections that may be encountered by the clinician in hospitalized children include bloodstream infections related to intravenous and intraarterial catheters, ventilator-associated pneumonia, urinary tract infections, surgical wound infections, and infections related to implanted foreign devices. Many of these children are susceptible to infections that develop as a result of breaches in the integrity of the mucus membranes and skin caused by a variety of tubes and catheters. In addition, children who have sustained significant trauma or extensive surgery may have increased susceptibility to bacterial infections.7,8 Other patients prone to infection with antibiotic-resistant

From the Division of Infectious Diseases, Children’s Hospital and Health Center, 3020 Children’s Way, MC 5041, San Diego, CA. Address reprint requests to: Alice Pong, MD, Division of Infectious Diseases, Children’s Hospital and Health Center, 3020 Children’s Way, MC 5041, San Diego, CA 92123. © 2004 Elsevier Inc. All rights reserved. 1045-1870/04/1501-0004$30.00/0 doi:10.1053/j.spid.2004.01.005

organisms are those with well-defined immune compromising conditions, including those treated with chemotherapy or organ transplantation, those with congenital or acquired immune deficiencies, and neonates in intensive care nurseries. These patients are at risk for acquisition of infection not only because they may be physically debilitated but because they often have prolonged exposure to the hospital environment and antibiotics. Antibiotic resistance, a common feature of nosocomial infections, may lead to increased rates of morbidity and mortality, as well as increased healthcare costs.9 The mechanisms of antibiotic resistance found in nosocomial pathogens and an approach to therapy of infections caused by these bacteria, are discussed.

The Problems Antibiotic Resistance Mechanisms In the hospital environment, particularly in ICUs, antibiotic use is extensive, resulting in selective pressure for antibiotic-resistant pathogens. Antibiotic resistance certainly is not a new phenomenon, with bacteria having developed resistance mechanisms long before humans were using antibiotics therapeutically.10-12 However, the remarkable ability of bacteria to survive in an environment containing many “toxins” is a function of the expression of one or more of many different potential antibiotic-resistance mechanisms. To make matters worse, genetic information encoding antibiotic resistance may be shared between organisms within a species or between species. Exchange of plasmids is

Seminars in Pediatric Infectious Diseases, Vol 15, No 1 ( January), 2004: pp 21-29

21

22

Pong and Bradley

a common method by which resistance genes are shared between bacteria. The description of mobile genetic elements helps explain the rapid development and spread of antibiotic resistance within the pathogens responsible for nosocomial infections.11 Antibiotic resistance gene cassettes have the ability to assemble on integrons, which in turn may be associated with transposons, capable of “jumping” from one segment of nucleic acid to another (eg, plasmid to chromosome, or visa versa).11-13 Antibiotic-resistant mutants normally may exist at low frequencies in a given population of bacteria. Antibiotic exposure often is the selection pressure allowing these otherwise silent mutants to achieve significant numbers, leading to treatment failure. Indiscriminate use of broadspectrum antibiotics and poor infection control practices contribute to the increased prevalence of antibiotic-resistant pathogens. The most common mechanisms of resistance are: 1) enzymatic alteration of the antibiotic by bacteria; 2) alteration of the antibiotic target site within the pathogen (by mutation at the binding site or enzymatic alterations of the binding site); 3) removal of the antibiotic from within the organism by efflux pumps; or 4) alteration of the cell wall of the organism to prevent movement of the antibiotic into the organism. The regulation of resistance gene expression also may be altered to allow the organism to produce much greater amounts of the gene product that leads to resistance. Finally, nosocomial pathogens may express many of these mechanisms simultaneously, resulting in additive or synergistic resistance against an antibiotic. Enzymatic Antibiotic Alteration. The best known of the enzymes that inactivate antibiotics are the ␤-lactamases that hydrolyze the beta-lactam ring of penicillins, cephalosporins, and carbapenems. Many different classifications of ␤-lactamases have been proposed based on substrate specificity, structure, and sequence homology.14-16 The structure and substrate specificity of ␤-lactamases are quite varied and continue to evolve under antibiotic pressure.15,16 Certain ␤-lactamases are active only against penicillins, others are active against cephalosporins, whereas still others are active against a wide variety of beta-lactam antibiotics. An organism may carry chromosomally encoded ␤-lactamases, plasmid-mediated ␤-lactamases, or a combination of the two. Each enzyme may prefer a different beta-lactam substrate, increasing the spectrum of beta-lactam resistance for the organism. Although chromosomal ␤-lactamases generally are not transmissible, transposon-mediated movement of these enzymes into plasmids may facilitate their spread among bacteria.17 Although some of the earlier characterized plasmidmediated ␤-lactamases demonstrated activity against ampicillin (eg, TEM in Escherichia coli and Haemophilus influenzae and SHV in Klebsiella), chromosomal ␤-lactamases (class I, or AmpC) found in many enteric bacilli and Pseudomonas were capable of hydrolyzing second- and third-generation cephalosporins. Presumably with the selective pressure exerted from extensive use of third-generation cephalosporins, numerous mutations in plasmid-carried TEM and SHV enzymes have occurred and now confer resistance to

third-generation cephalosporins and some fourth-generation cephalosporins.15,16,18 More than 100 of these new, more potent ␤-lactamases have been described and sequenced. Some are highly active against the third-generation cephalosporins. These newly emergent TEM- and SHV-related enzymes, designated “extended-spectrum ␤-lactamases” (ESBLs) have spread rapidly through many parts of the world, rendering treatment with third-generation cephalosporins ineffective for infections caused by strains of E. coli and Klebsiella harboring these enzymes. ␤-lactamases of the CTX-M class are among the most active against cefotaxime, with some of these enzymes demonstrating activity against both third- and fourth-generation cephalosporins.18 Metallo-␤-lactamases, active against the carbapenems, have been described as chromosomal enzymes in Stenotrophomonas maltophilia and may be present in antibiotic resistance cassettes within integrons on plasmids in Pseudomonas aeruginosa and a variety of enteric bacilli.19 Plasmid-mediated resistance suggests that widespread resistance to imipenem and meropenem may not be far away. Regulation of the production of chromosomal ␤-lactamases also may become altered, giving rise to constitutive hyperproduction of AmpC enzymes.20 These “derepressed mutants,” with a mutation in the ampD locus, can be found in species of Enterobacter, Citrobacter, Serratia, Morganella, Providencia, and Pseudomonas.21 The quantity and location of these enzymes in the periplasmic space allow these pathogens to display a phenotype of high resistance to thirdgeneration cephalosporins, with well-documented spread occurring in the hospital setting.22 Enzymes capable of inactivating the aminoglycosides by adenylation, acetylation, or phosphorylation may be found in several species of bacteria, both gram-negative and gram-positive.23 These resistance determinants generally are present on plasmids and transposons and can be shared between organisms. As with ␤-lactamases, variations in substrate specificity exist among the aminoglycosides.24,25 For example, high-level enterococcal resistance to gentamicin is mediated by the enzyme 2⬙-phosphotransferase-6⬘acetyltransferase, an aminoglycoside-modifying enzyme with activity against tobramycin and amikacin but not against streptomycin.26 Target Binding Site Alteration. Antibiotics effectively inhibit the growth of bacteria by binding to essential structures and interfering with cell metabolism. For the betalactam antibiotics, the target sites are the transpeptidases (penicillin-binding proteins, or PBPs) that cross-link peptidoglycan in the formation of the cell wall. A mutation leading to an alteration in the beta-lactam binding site on the transpeptidase will decrease the affinity of antibiotic binding,10 which in turn will decrease the ability of the antibiotic to inhibit transpeptidase function and cell metabolism. This decrease in binding will result in an increase in the minimum inhibitory concentration (MIC) of the antibiotic against the organism. Bacteria may contain several transpeptidases; each may have mutations at the respective binding sites, leading to additive resistance. Some of the mutations may lead to

Nosocomial Infections Caused by Antibiotic-Resistant Pathogens small decreases in the binding affinity by the antibiotic and relatively little change in the inhibition of enzyme function; however, some changes will cause dramatic alterations in binding, effectively resulting in antibiotic resistance and treatment failure. Although numerous PBP-altered clones have been described, additional mutations continue to occur, leading to variations in binding affinity within each clone and to further resistance to beta-lactam antibiotics.27 For example, methicillin-resistant Staphylococcus aureus (MRSA) strains have developed resistance to semisynthetic penicillins by using an alternate penicillin-binding protein (PBP2⬘), which has a lower affinity to the beta lactam drugs.28 Enterococci utilize a similar low affinity beta lactam transpeptidase (PBP5), leading to intrinsic resistance to cephalosporin antibiotics.26 Macrolides and clindamycin (a lincosamide) bind to the 50S ribosomal subunit at the same critical target site, inhibiting the formation of proteins. An alteration of the binding site by methylation of an adenine residue (mediated by erm-related genes) prevents the association of the antibiotic with the binding site and yields an organism that is resistant to both macrolides (erythromycin, azithromycin, and clarithromycin) and lincosamides, resulting in the MLSB phenotype.29 Many strains of MRSA also carry these MLSB resistance genes (ermA, ermC).29 Vancomycin resistance in some gram-positive nosocomial pathogens is based on the alteration of the vancomycin binding site.30 The ability of vancomycin to bind to the D-Ala-D-Ala end of the peptidoglycan pentapeptide prevents this substrate from being utilized in the formation of the cell wall by inhibiting both transpeptidation and transglycosylation. The vanA resistance gene allows for the DAla-D-Ala terminus to be altered to D-Ala-D-Lactate, preventing vancomycin binding. The VanA resistance phenotype is found in strains of Enterococcus faecium and Enterococcus faecalis. This plasmid-mediated gene actually is part of a cluster of genes, which includes vanH (increases D-lac production), vanX (encodes for a dipeptidase which breaks down the normal D-ala-D-ala terminus), and regulatory genes vanR and vanS. Other resistance phenotypes include VanB (may be susceptible to teicoplanin), VanC (low level vancomycin resistance primarily in Enterococcus gallinarum, Enterococcus casselflavus, and Enterococcus flavescens), VanD, and VanE.26 More recently, strains of S. aureus have been found with decreased susceptibility to glycopeptides.31 The mechanism of resistance is unclear, although it is not by acquisition of vanA or vanB genes. Resistance appears to be related to a thickened cell wall and adherence of vancomycin to areas of cell wall that are not actively being created.30 Fluoroquinolones inhibit the growth of bacteria by interfering in DNA synthesis as a result of antibiotic binding to DNA gyrase and/or and topoisomerase IV. Alteration in the binding site of fluoroquinolones to one or both of these 2 target enzymes will produce an organism with decreased susceptibility.32 Efflux Pumps. One mechanism of bacterial survival in an environment containing lethal factors is the efflux pump, designed to transport specific types of molecules

23

from the intracellular to the extracellular environment.33-35 Many different kinds of efflux pumps exist, presumably designed for removing a wide variety of toxic molecules, including antibiotics, and thereby limiting the harmful impact on cellular function. Although some of the best-studied transport systems are involved in beta-lactam antibiotic transport, fluoroquinolone33 and macrolide29 efflux pumps also are clinically relevant causes of antibiotic resistance. Multidrug efflux pumps that bind multiple classes of antibiotics have been described recently, as have alterations in the genetic regulation of pump expression leading to increased antibiotic efflux. Multiple types of efflux pumps may coexist within the pathogen, increasing the observed in vitro antibiotic resistance.36 Both gram-negative and grampositive organisms may contain multiple antibiotic efflux pumps against a wide variety of antibiotics. Cell Wall Porin-Deficient Mutants. In order for an antibiotic to enter the cytoplasm of a gram-negative bacillus, it must pass through a complex cell wall, characterized by an outer membrane, a periplasmic space, and a cytoplasmic membrane. In order for an antibiotic to bind to its target proteins, it must enter the organism through porin proteins in the semipermeable outer membrane. Gram-negative pathogens, most notably Pseudomonas aeruginosa and the Enterobacteriaceae, may display resistance to antibiotics based on a deficiency of specific porin proteins in the cell wall.37,38 Some porins are known to be part of an efflux mechanism responsible for capturing the antibiotic, transporting it through the periplasmic space, and extruding it through a porin into the extracellular environment.38 Imipenem-resistant P. aeruginosa has been recognized for the past 20 years, with resistance linked to mutants demonstrating a deficiency of OprD porin proteins, as recently reviewed.38 In as many as 20 percent of all patients treated for Pseudomonas with imipenem, antibiotic-resistant mutants can be isolated following therapy.39 The same phenomenon also may occur with other carbapenems. Genetic Regulation Mutations. As mentioned in the preceeding sections, genes regulating production of proteins leading to antibiotic resistance, such as ␤-lactamases or efflux pumps, may undergo a mutational change, which can lead to excess production of these proteins and subsequent highly antibiotic-resistant phenotypes.16 Organisms with altered regulation of production of resistance factors may have a survival advantage in an environment of high antibiotic pressure. Many strains of enteric gram-negative bacteria have derepressed regulation of chromosomal AmpC class ␤-lactamase genes, leading to hyperproduction of beta lactamase, resulting in resistance to third-generation cephalosporins.20,40,41 Upregulation of other mechanisms of resistance, such as efflux pumps, also may contribute to overall antibiotic resistance.16 In gram-positive organisms, constitutive producers of ribosomal methylase are responsible for macrolide resistance.29 Multiple Simultaneous Resistance Mechanisms. The clinical expression of antibiotic resistance may involve several different mechanisms operating simultaneously within a pathogen, with each mechanism expressed to a different degree, based on the regulation of resistance at a molecular

24

Pong and Bradley

Figure 1. Major resistance mechanisms against ␤-lactam antibiotics for gram-negative bacilli include ␤-lactamases, cell membrane porin deficiencies, and efflux pumps. All 3 mechanisms may be present simultaneously, each contributing to the overall antibiotic resistance.

level. For example, an organism with a weak beta-lactamase that appears susceptible to an antibiotic in vitro may acquire an efflux pump or develop porin deficiency, or both. Either of these additional mechanisms will lead to a much lower concentration of antibiotic intracellularly, allowing the weak beta-lactamase the opportunity to degrade the antibiotic before significant cell injury can occur.15,16,42 Most often, when a laboratory tests for susceptibility of a pathogen to an antibiotic, the sum of all resistance mechanisms (except those that may require induction) leads to an in vitro determination of susceptibility (Fig 1), which the clinician must use in determining the best agent(s) for treatment.

Nosocomial Infections In Pediatrics Nosocomial bacterial infections and pathogens in pediatrics are summarized in Table 1. Richards and coworkers reviewed nosocomial infections in pediatric ICUs in the United States.43 The authors found that the most frequent infections were bloodstream, pneumonia, and urinary tract infection, with bloodstream infections seen more commonly in children younger than 2 months and urinary tract infections found more commonly in older children. The most common organisms isolated in the bloodstream were coagulase-negative staphylococci (37.8%), enterococci (11.2%), S. aureus (9.3%), and Enterobacter spp. (6.2%). S. aureus, P. aeruginosa, and H. influenzae were seen most commonly in pneumonia. Although E. coli remained the most frequent urinary tract pathogen, Candida albicans, P. aeruginosa, Enterobacter spp. and enterococci each comprised at least 10 percent of the urinary pathogens. S. aureus, P. aeruginosa and

coagulase-negative staphylococci were the primary pathogens for surgical site infections.43 In a similar review of nosocomial infections in intensive care nurseries, the bloodstream was found to be the most common site of infection (32-49% depending on birth weight), followed by pneumonia (12-18%) and ear, nose, and throat infections (8-21%). The most common pathogens were coagulase-negative staphylococci in the bloodstream and coagulase-negative staphylococci, S. aureus, and P. aeruginosa for pneumonia.44 Outbreaks of antibiotic-resistant organisms in neonatal and pediatric units certainly are not new, although the organisms and their antibiotic-resistance patterns, as well our tools to investigate outbreaks, continue to evolve (Jarvis 2004, in this issue). These outbreaks often reflect the same pathogens and tissues sites noted in adults. Current clinical challenges relate to pathogens that display newer resistance patterns. The ESBL-producing gram-negative pathogens are resistant to third-generation and some fourthgeneration cephalosporins, with a proportion of these organisms also demonstrating decreased susceptibility to aminoglycosides, carbapenems, and fluoroquinolones. Outbreaks caused by Klebsiella and E. coli have been reported widely.45,46 Outbreaks by enteric gram-negative bacilli that carry chromosomal AmpC ␤-lactamases (present in Enterobacter, Serratia, and Citrobacter) have been occurring for several years and continue to present challenges.47-49 Acinetobacter baumannii has caused recent outbreaks reported primarily from Asia and Europe.50,51 The most critical aspect of Acinetobacter for the clinician is the ability of the organism to acquire resistance to all classes of antibiotics. P. aeruginosa is a perennial problem on pediatric units,

Table 1. Most Commonly Reported Antibiotic-Resistant Bacterial Pathogens Infection

Bacteria

Ventilator-associated pneumonia Acinetobacter spp. Enterobacter spp. Escherichia coli Klebsiella spp. Pseudomonas aeruginosa Serratia spp. Staphylococcus spp. Intravenous catheter-associated Acinetobacter spp. bloodstream infection Enterobacter spp. Enterococcus spp. Klebsiella spp. Pseudomonas aeruginosa Staphylococcus spp. Urinary tract infection Escherichia coli Enterobacter spp. Enterococcus spp. Pseudomonas aeruginosa Wound infection Enterobacter spp. Klebsiella spp. Staphylococcus spp. Pseudomonas aeruginosa Data from

43,44,70

Nosocomial Infections Caused by Antibiotic-Resistant Pathogens particularly neonatal and pediatric ICUs, and hematology/ oncology wards.52,53 The nonfermenting gram-negative bacteria, including Stenotrophomonas, Comamonas, Chyseobacteria spp. (previously known as strains of Flavobacteria) and, of course, Acinetobacter spp., also are responsible for nosocomial infections.54,55 Although these organisms often are resistant to many antibiotics, they appear to be less virulent in normal hosts than are the aerobic enteric bacilli or P. aeruginosa. Although the prevalence of MRSA in pediatric institutions is lower than that in adult hospitals and nursing homes, reports of MRSA as a significant pathogen in children are increasing. Jarvis and coworkers reported increasing numbers of MRSA seen in pediatric hospitals throughout the 1970s.56 In 1986, Storch and coworkers reviewed pediatric patients with MRSA bacteremia compared with those with methicillin-susceptible S. aureus (MSSA) and found a higher mortality rate (50% versus 2% for MSSA) in patients with MRSA infections. Underlying disease and inappropriate therapy were thought to be contributing factors.57 In the adult population, results vary regarding greater virulence and mortality in patients with MRSA infection versus those infected with MSSA.58-61 Numerous instances of MRSA occurring in neonatal ICUs have been reported.62,63 The most common sites of infection were bloodstream and lung. Wound infections, meningitis, bone and joint infections, and urinary tract infections also were reported.77,79 MRSA also has been reported in pediatric burn units64 and long-term care facilities.65 MRSA also causes a significant problem in patients with cystic fibrosis (CF). Burns and coworkers reported 18.8 percent of S. aureus isolates in CF centers in the United States were oxacillin-resistant.66 Miall and coworkers suggested that patients with CF and MRSA have greater lung dysfunction and require more courses of antibiotics than do those without MRSA.67 To complicate matters further, MRSA now is being seen in children without known hospital risk factors. Reports of “community acquired” MRSA in otherwise healthy children are increasing.68-71 Vancomycin-resistant enterococci (VRE) strains have been reported in pediatric hospitals, particularly in neonatal ICUs and oncology wards, and in patients with gastrointestinal disease.72-78 Most of these strains are E. faecium expressing the vanA phenotype.73,77,78 Although VRE infections in pediatrics usually are reported as bloodstream infections,75,77 most isolates are found colonizing the gastrointestinal tract.78 Colonization may be a risk factor for later development of infection.75 Risk factors for the development of VRE infection in adult patients include prolonged hospitalization, intensive care stays, and use of antibiotics, including third-generation cephalosporins, vancomycin, and antibiotics with anaerobic activity.26 In the neonatal population, colonization with VRE is associated with low birth weight and a longer duration of antibiotic therapy.72,74 Singh-Naz and coworkers found VRE colonization associated with younger age, use of chemotherapy, and use of antibiotic therapy, specifically with cefotaxime, vancomycin, and ceftazidime.76 Henning and coworkers evaluated E. faecium colonization over the course of a 12-month

25

period. Forty-three percent of patients showed persistent carriage for a median duration of 112 days. Colonized patients had a higher rate of infection with VRE than did noncolonized patients (19% versus 4%).75

Treatment General Considerations for Antibiotic Therapy. Once an infection is suspected, based on the clinical, laboratory, and imaging characteristics of the child, appropriate cultures should be obtained. Broad-spectrum antibiotics should be administered empirically, based on the susceptibility patterns of nosocomial pathogens in the child’s hospital. The more severely ill the child, the more broadspectrum the antibiotics should be, as the physician may have only one chance for cure. Data suggest that using active antibiotics in the appropriate dose without delay will decrease the morbidity and mortality rates of the infection, decrease the overall costs of treating the infection, and decrease the emergence of resistance.79-82 The relative activity of antimicrobial agents against gram-negative (Table 2A) and gram-positive pathogens (Table 2B) is provided, although each clinician should consult the local antibiogram for the susceptibility pattern of the pathogens isolated in his/her own institution. Published data on resistance patterns comes from a wide variety of sources, including both government (Centers for Disease Control and Prevention) and industry-based (MYSTIC, The Surveillance Network, TRUST, SENTRY, Alexander Project). Each collection of isolates on which resistance is reported may include information for different geographic regions, population groups, patient comorbidities, tissue sites of infection, and previous antibiotic exposures, among others. These reports usually focus on susceptibilities of many antibiotics to a single pathogen or the susceptibilities of the pathogens isolated from a single infection (eg, nosocomial pneumonia83,84 or urinary tract infection85) to a wide range of antibiotics. As very few data collections published have specific information on children, a critical concern is that the conclusions reached in these manuscripts be taken with caution in applying them to pediatric patients. Once bacterial culture results are available, a decision regarding the role of the isolated organisms must be made. Contamination and colonization must be considered in the interpretation of positive cultures. For bloodstream infections, coagulase-negative staphylococci may represent contamination from the skin rather than true pathogens, although in the intensive care nursery, they represent the most frequent cause of nosocomial bacteremia. Klebsiella spp. isolated from a culture obtained from an endotracheal tube may represent colonization or true infection. Based on the overall clinical assessment, supported by laboratory and imaging data and the response (or lack of response) to empiric therapy, the physician needs to decide whether to continue therapy for a complete treatment course or to stop the antibiotics if data do not support an infection as the cause of the child’s clinical state. Once empiric therapy is started, the culture results hopefully will allow more appropriate antibiotic agents to be used for completion of therapy. An E. coli that is suscep-

Pong and Bradley

26

Table 2A. Antibiotic Activity for Gram-Negative Pathogens (0 to ⫹⫹⫹⫹⫹)a Antibiotic Ticarcillin- PiperacillinCeftazidime Ceftriaxone Cefepime Tobramycin clavulanate tazobactam Ciprofloxacin Meropenem

Organism E. coli Klebsiella spp. Enterobacter spp. Pseudomonas aeruginosab Acinetobacter spp.b Stenotrophomonasc

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹ ⫹⫹⫹ ⫹⫹ ⫹⫹⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹ ⫹ ⫹⫹ ⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹ ⫹⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹ ⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹ ⫹⫹⫹ ⫹⫹⫹ ⫹⫹⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹ ⫹⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹ ⫹⫹ ⫹⫹

⫹⫹⫹⫹⫹ ⫹⫹⫹⫹⫹ ⫹⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹

aSusceptibility bColistin

data are averaged,83,85,93,94 with local hospital data potentially much different than these values. may be effective in vitro against organisms resistant to all available agents, with limited data on efficacy and significant

toxicities.87,88 cTrimethoprim-sulfamethoxaxazole is the most active antibiotic against Stenotrophomonas in vitro.

tible to first-generation cephalosporins should not require therapy with second-, third-, or fourth-generation agents, or with carbapenems. However, if the organism is resistant to all cephalosporins based on ESBL production, then an aminoglycoside, carbapenem, or fluoroquinolone may be required. Data on safety of quinolones collected through compassionate use programs86 and passive reporting to the Food and Drug Administration have to date not demonstrated a confirmed case of quinolone-induced arthropathy in a child in the United States following the use of a product approved by the Food and Drug Administration. Therefore, the risk-benefit analysis that accompanies the decision regarding selection of an antibiotic for the child who has a serious infection caused by a multiresistant pathogen may well favor the use of a fluoroquinolone. Antibiotic Therapy for Specific Pathogens. Children with infections caused by enteric gram-negative bacilli that carry chromosomal AmpC ␤-lactamases (Enterobacter, Serratia, and Citrobacter), which may test as susceptible to thirdgeneration cephalosporins at the time the organisms initially are cultured, are at risk for having treatment failure if these antibiotics are used as sole therapy. Definitive therapy of these pathogens should take into account the selection of organisms that constitutively produce AmpC ␤-lactamase and may include the addition of an aminoglycoside to the third-generation cephalosporin, the use of cefepime (a fourth-generation cephalosporin), or the use of a carbapenem (meropenem, imipenem, or ertapenem). For gram-negative pathogens, including Acinetobacter, colistin

has been used as intravenous therapy in adult populations for multiresistant organisms87,88 and currently is being used in infants for whom no other options are available (unpublished observations). The Acinetobacter strains isolated from most of the geographic areas of the world fortunately have remained susceptible to carbapenems and aminoglycosides, as well as to the extended-spectrum cephalosporins (thirdand fourth-generation); however, the local resistance patterns are quite variable. For P. aeruginosa, ceftazidime (or an antipseudomonal penicillin) plus an aminoglycoside (tobramycin or amikacin) represents effective therapy in most institutions. However, in situations in which ceftazidime resistance is likely to occur, alternatives include cefepime (with or without an added aminoglycoside), meropenem or imipenem (with or without an added aminoglycoside), or, as a last resort due to safety considerations, a fluoroquinolone such as ciprofloxacin. For pediatric institutions, the rates of resistance to fluoroquinolones appear to be far less than those in comparable adult institutions, likely due to the lower rates of quinolone use in children. For MRSA infection, vancomycin remains the mainstay for treatment. However, clindamycin exhibits greater activity against “community acquired” strains than against MRSA strains previously documented to cause nosocomial infections and represents alternative effective therapy.68,70 Strains of MRSA also may be susceptible to trimethoprimsulfamethoxazole, rifampin, and selected fluoroquinolones. The oxazolidinone, linezolid, may be another option.

Table 2B. Antibiotic Activity for Gram-Positive Pathogens (0 to ⫹⫹⫹⫹⫹)a Antibiotic Organism

Ampicillin

Oxacillin

Cefazolin

Vancomycin

Linezolid

Methicillin-susceptible Staphylococcus spp. (S. aureus or coagulase-negative staphylococci) Methicillin-resistant Staphylococcus spp. Enterococcus faecalisb Enterococcus faeciumb

0 0 ⫹⫹⫹⫹ ⫹⫹

⫹⫹⫹⫹⫹ 0 0 0

⫹⫹⫹⫹⫹ 0 0 0

⫹⫹⫹⫹⫹ ⫹⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹

⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹ ⫹⫹⫹⫹

aSusceptibility bFor

data are averaged,28,83,91 with local hospital data potentially much different than these values. vancomycin susceptible strains.

Nosocomial Infections Caused by Antibiotic-Resistant Pathogens Treatment of VRE infection can be problematic. Removal of an associated foreign device is important and may be curative.89 Penicillin and ampicillin may be effective with certain strains of VRE that remain susceptible to these agents, although higher doses than required for therapy of other pathogens may be needed.26 Quinupristin-dalfopristin is a combination of 2 streptogramins with activity against E. faecium, but E. faecalis strains are inherently resistant.90 Perhaps the most promising agent with activity against VRE is the oxazolidinone, linezolid. Linezolid has been shown clinically to be effective in treating VRE infection as well as infections with MRSA.91 It is approved for use in both adults and children in the United States and is available both in parenteral and oral forms. Unfortunately, E. faecium with resistance to linezolid already has been reported.92

Summary As medical care becomes more sophisticated and children are hospitalized for longer periods of time, the risk of development of complicated infections caused by multidrug-resistant pathogens increases. The physician is constantly being challenged to deliver effective antibiotic therapy, while at the same time preventing the selection of antibiotic resistance. Knowledge of the pathogens most likely to be present and the potential resistance mechanisms in these organisms is important in selecting empiric antibiotic therapy. Appropriate collection of cultures to obtain susceptibility information on the pathogens is crucial to optimize subsequent therapy and minimize the development of antibiotic resistance.

References 1. Hallahan AR, Shaw PJ, Rowell G, et al: Improved outcomes of children with malignancy admitted to a pediatric intensive care unit. Crit Care Med 28:3718-3721, 2000 2. Booy R, Habibi P, Nadel S, et al: Reduction in case fatality rate from meningococcal disease associated with improved healthcare delivery. Arch Dis Child 85:386-390, 2001 3. Tenner PA, Dibrell H, Taylor RP: Improved survival with hospitalists in a pediatric intensive care unit. Crit Care Med 31:847-852, 2003 4. Stover BH, Shulman ST, Bratcher DF, et al: Nosocomial infection rates in US children’s hospitals’ neonatal and pediatric intensive care units. AM J Infect Control 29:152-157, 2001 5. Sohn AH, Garrett DO, Sinkowitz-Cochran RL, et al: Prevalence of nosocomial infections in neonatal intensive care unit patients: Results from the first national point-prevalence survey. J Pediatr 139:821-827, 2001 6. Girouard S, Levine G, Goodrich K, et al: Pediatric Prevention Network: a multicenter collaboration to improve health care outcomes. Am J Infect Control 29:158-161, 2001 7. Meert KL, Long M, Kaplan J, et al: Alterations in immune function following head injury in children. Crit Care Med 23:822-828, 1995 8. Tarnok A, Schneider P: Pediatric cardiac surgery with cardiopulmonary bypass: pathways contributing to transient systemic immune suppression. Shock 16:24-32, 2001 (suppl)

27

9. Howard DH, Scott RD II, Packard R, et al: The global impact of drug resistance. Clin Infect Dis 36:4-10, 2003 (suppl) 10. Koch AL: Penicillin binding proteins, ␤-lactams, and lactamases: offensives, attacks, and defensive countermeasures. Crit Rev Microbiol 26:205-220, 2000 11. Recchia GD, Hall RM: Origins of the mobile gene cassettes found in integrons. Trends Microbiol 5:389-394, 1997 12. Leibert CA, Hall RM, Summers AO: Transposon Tn21, flagship of the floating genome. Microbiol Mol Rev 63:507-522, 1999 13. Fluit AC, Schmitz FJ: Class 1 integrons, gene cassettes, mobility, and epidemiology. Eur J Clin Microbiol Infect Dis 18:761770, 1999 14. Bush K, Jacoby GA, Medeiros AA: A functional classification scheme for ␤-lactamases and its correlation with molecular structure. Antimicrob Agents Chemother 39:1211-1233, 1995 15. Bush K: New ␤-lactamases in Gram-negative bacteria: diversity and impact on the selection of antimicrobial therapy. Clin Infect Dis 32:1085-1089, 2001 16. Livermore DM: Bacterial resistance: origins, epidemiology, and impact. Clin Infect Dis 36:11-23, 2003 (suppl) 17. Odeh R, Kelkar S, Hujer AM, et al: Broad resistance due to plasmid-mediated AmpC beta-lactamases in clinical isolates of Escherichia coli. Clin Infect Dis 35:140-145, 2002 18. Bradford PA: Extended-spectrum ␤-lactamases in the 21st century: characterization, epidemiology, and detection of this important resistance threat. Clin Microbiol Rev 14:933-951, 2001 19. Nordmann P, Poirel L: Emerging carbapenemases in Gramnegative aerobes. Clin Microbiol Infect 8:321-331, 2002 20. Livermore DM: Beta-lactamases: quantity and resistance. Clin Microbiol Infect 3:10-19, 1997 (suppl) 21. Livermore DM: ␤-lactamases in laboratory and clinical resistance. Clin Microbiol Rev 8:557-584, 1995 22. Philippon A, Arlet G, Jacoby GA: Plasmid-determined AmpCType ␤-lactamases. Antimicrob Agents Chemother 46:1-11, 2002 23. Wright GD: Aminoglycoside-odifying enzymes. Curr Opin Microbiol 2:499-503, 1999 24. Miller GH, Sabatelli FJ, Hare RS, et al: The most frequent aminoglycoside resistance mechanisms— changes with time and geographic area: a reflection of aminoglycoside usage patterns? Aminoglycoside Resistance Study Groups. Clin Infect Dis 24:46-62, 1997 (suppl) 25. Sabtcheva S, Galimand M, Gerbaud G, et al: Aminoglycoside resistance gene ant(4⬘)-Iib of Pseudomonas aeruginosa BM4492, a clinical isolate from Bulgaria. Antimicrob Agents Chemother 47:1584-1588, 2003 26. Cetinkaya Y, Falk P, Mayhall CG: Vancomycin-resistant enterococci. Clin Microbiol Rev 13:686-707, 2000 27. Chambers HF: Penicillin-binding protein-mediated resistance in pneumococci and staphylococci. J Infect Dis 179:353-359, 1999 (suppl) 28. Livermore DM: Antibiotic resistance in staphylococci. Int J Antimicrob Agents 16:S3-S10, 2000 29. Leclercq R: Mechanisms of resistance to macrolides and lincosamides: nature of the resistance elements and their clinical implications. Clin Infect Dis 34:482-492, 2002 30. Pootoolal Neu J, Wright GD: Glycopeptide antibiotic resistance. Annu Rev Pharmacol Toxicol 42:381-408, 2002 31. Martin R, Wilcox KR: Staphylococcus aureus with reduced susceptibility to vancomycin—United States, 1997. MMWR 46:765-766, 1997 32. Hooper DC: Emerging mechanisms of fluoroquinolone resistance. Emerg Infect Dis 7:337-341, 2001

28

Pong and Bradley

33. Poole K: Efflux-mediated resistance to fluoroquinolones in Gram-negative bacteria. Antimicrob Agents Chemother 44: 2233-2241, 2000 34. Poole K: Multidrug efflux pumps and antimicrobial resistance in Pseudomonas aeruginosa and related organisms. J Mol Microbiol Biotechnol 3:255-264, 2001 35. Bambeke FV, Glupczynski Y, Ple´siat P: Antibiotic efflux pumps in prokaryotic cells: occurrence, impact on resistance and strategies for the future of antimicrobial therapy. J Antimicrob Chemother 51:1055-1065, 2003 36. Lee A, Mao W, Warren MS, et al: Interplay between efflux pumps may provide either additive or multiplicative effects on drug resistance. J Bacteriol 182:3142-3150, 2000 37. De´ E, Basle´ A, Jaquinod M, et al: A new mechanism of antibiotic resistance in Enterobacteriaceae induced by a structural modification of the major porin. Mol Microbiol 41:189-198, 2001 38. Hancock RE, Brinkman FS: Function of Pseudomonas porins in uptake and efflux. Annu Rev Microbiol 56:17-38, 2002 39. Calandra GB, Hesney M, Brown KR: Imipenem/cilastatin therapy of serious infections: a U. S. multicenter noncomparative trial. Clin Ther 7:225-238, 1985 40. Ehrhardt AF, Sanders CC: Beta-lactam resistance amongst Enterobacter species. J Antimicrob Chemother 32:1-11, 1993 (suppl) 41. Bagge N, Ciofu O, Hentzer M, et al: Constitutive high expression of chromosomal ␤-lactamase in Pseudomonas aeruginosa caused by a new insertion sequence (IS1669) located in ampD. Antimicrob Agents Chemother 46:3406-3411, 2002 42. Crowley B, Bened VJ, Domınech-Sa´nchez A: Expression of SHV-2 ␤-lactamase and of reduced amounts of OmpK36 porin in Klebsiella pneumoniae results in increased resistance to cephalosporins and carbapenems. Antimicrob Agents Chemother 46:3679-3682, 2002 43. Richards MJ, Edwards JR, Culver DH, et al: Nosocomial infections in pediatric intensive care units in the United States. Pediatrics 103:e39, 1999 44. Gaynes RP, Edwards JR, Jarvis WR, et al: Nosocomial infections among neonates in high-risk nurseries in the United States. Pediatrics 98:357-361, 1996 45. Kim YK, Pai H, Lee HJ, et al: Bloodstream infections by extended-spectrum ␤-lactamase-producing Escherichia coli and Klebsiella pneumoniae in childrenepidemiology and clinical outcome. Antimicrob Agents Chemother 46:1481-1491, 2002 46. Gupta A: Hospital-acquired infections in the neonatal intensive care unit—Klebsiella pneumoniae. Semin Perinatol 26:340345, 2002 47. Jang TN, Fung CP, Yang TL, et al: Use of pulsed-field gel electrophoresis to investigate an outbreak of Serratia marcescens infection in a neonatal intensive care unit. J Hosp Infect 48: 13-19, 2001 48. Liu SC, Leu HS, Yen MY, et al: Study of an outbreak of Enterobacter cloacae sepsis in a neonatal intensive care unitthe application of epidemiologic chromosome profiling by pulsedfield gel electrophoresis. Am J Infect Control 30:381-385, 2002 49. Manning ML, Archibald LK, Bell LM, et al: Serratia marcescens transmission in a pediatric intensive care unit: a multifactorial occurrence. Am J Infect Control 29:115-119, 2001 50. Huang YC, Su LH, Wu TL, et al: Outbreak of Acinetobacter baumannii bacteremia in a neonatal intensive care unitclinical implications and genotyping analysis. Pediatr Infect Dis J 21: 1105-1109, 2002

51. Melamed R, Greenberg D, Porat N, et al: Successful control of an Acinetobacter baumannii outbreak in a neonatal intensive care unit. J Hosp Infect 53:31-38, 2003 52. Brown DG, Baublis J: Reservoirs of pseudomonas in an intensive care unit for newborn infants: mechanisms of control. J Pediatr 90:453-457, 1977 53. Foca M, Jakob K, Whittier S, et al: Endemic Pseudomonas aeruginosa infection in a neonatal intensive care unit. N Engl J Med 343:695-700, 2000 54. Hoque SN, Graham J, Kaufmann ME, et al: Chryseobacterium (Flavobacterium) meningosepticum outbreak associated with colonization of water taps in a neonatal intensive care unit. J Hosp Infect 47:188-192, 2001 55. Higgens CS, Murtough SM, Williamson E, et al: Resistance to antibiotics and biocides among non-fermenting Gram-negative bacteria. Clin Microbiol Infect 7:308-315, 2001 56. Jarvis WR, Thornsberry C, Boyce J, et al: Methicillin-resistant Staphylococcus aureus at children’s hospitals in the United States. Pediatr Infect Dis 4:651-655, 1985 57. Storch GA, Rajagopalan L: Methicillin-resistant Staphylococcus aureus bacteremia in children. Pediatr Infect Dis 5:59-67, 1986 58. Hershow RC, Khayr WF, Smith NL: A comparison of clinical virulence of nosocomially acquired methicillin-resistant and methicillin-sensitive Staphylococcus aureus infections in a university hospital. Infect Control Hosp Epidemiol 13:587-593, 1992 59. Soriano A, Martinez A, Mensa J, et al: Pathogenic significance of methicillin resistance for patients with Staphylococcus aureus bacteremia. Clin Infect Dis 30:368-373, 2000 60. Gonza´lez C, Rubio M, Romero-Vivas J, et al: Bacteremic pneumonia due to Staphylococcus aureusa comparison of disease caused by methicillin-resistant and methicillin-susceptible organisms. Clin Infect Dis 29:1171-1177, 1999 61. Romero-Vivas J, Rubio M, Fernandez C, et al: Mortality associated with nosocomial bacteremia due to methicillin-resistant Staphylococcus aureus. Clin Infect Dis 21:1417-1423, 1995 62. Haley RW, Cushion NB, Tenover FC, et al: Eradication of endemic methicillin-resistant Staphylococcus aureus infections from a neonatal intensive care unit. J Infect Dis 171:614-624, 1995 63. Reboli AC, John JF Jr, Levkoff AH: Epidemic methicillingentamicin-resistant Staphylococcus aureus in a neonatal intensive care unit. Am J Dis Child 143:34-39, 1989 64. Sheridan RL, Weber J, Benjamin J, et al: Control of methicillin-resistant Staphylococcus aureus in a pediatric burn unit. Am J Infect Control 22:340-345, 1994 65. Stover BH, Duff A, Adams G, et al: Emergence and control of methicillin-resistant Staphylococcus aureus in a children’s hospital and pediatric long-term care facility. Am J Infect Control 20:248-255, 1992 66. Burns JL, Emerson J, Stapp JR, et al: Microbiology of sputum from patients at cystic fibrosis centers in the United States. Clin Infect Dis 27:158-163, 1998 67. Miall LS, McGinley NT, Brownlee KG, et al: Methicillin resistant Staphylococcus aureus (MRSA) infection in cystic fibrosis. Arch Dis Child 84:160-162, 2001 68. Fergie JE, Purcell K: Community-acquired methicillin-resistant Staphylococcus aureus infections in South Texas children. Pediatr Infect Dis J 20:860-863, 2001 69. Herold BC, Immergluck LC, Maranan MC, et al: Communityacquired methicillin-resistant Staphylococcus aureus in children with no identified predisposing risk. JAMA 279:593-598, 1998 70. Sattler CA, Mason EO Jr, Kaplan SL: Prospective comparison of risk factors and demographic and clinical characteristics of community-acquired, methicillin-resistant versus methicillin-

Nosocomial Infections Caused by Antibiotic-Resistant Pathogens

71.

72.

73.

74. 75.

76.

77. 78.

79.

80.

81.

82.

83.

susceptible Staphylococcus aureus infection in children. Pediatr Infect Dis J 21:910-916, 2002 Gorak EJ, Yamada SM, Brown JD: Community-acquired methicillin-resistant Staphylococcus aureus in hospitalized adults and children without known risk factors. Clin Infect Dis 29: 797-800, 1999 Malik RK, Montecalvo MA, Reale MR, et al: Epidemiology and control of vancomycin-resistant enterococci in a regional neonatal intensive care unit. Pediatr Infect Dis J 18:352-356, 1999 Lee HK, Lee WG, Cho SR: Clinical and molecular biological analysis of a nosocomial outbreak of vancomycin-resistant enterococci in a neonatal intensive care unit. Acta Paediatr 88: 651-654, 1999 Yu ˝ce A, Karaman M, Gu ˝lay Z, et al: Vancomycin-resistant enterococci in neonates. Scand J Infect Dis 33:803-805, 2001 Henning KJ, Delencastre H, Eagan J, et al: Vancomycin-resistant Enterococcus faecium on a pediatric oncology wardduration of stool shedding and incidence of clinical infection. Pediatr Infect Dis J 15:848-854, 1996 Singh-Naz N, Sleemi A, Pikis A, et al: Vancomycin-resistant Enterococcus faecium colonization in children. J Clin Microbiol 37:413-416, 1999 Gray JW, George RH: Experience of vancomycin-resistant enterococci in a children’s hospital. J Hosp Infect 45:11-18, 2000 Nourse C, Murphy H, Byrne C, et al: Control of a nosocomial outbreak of vancomycin resistant Enterococcus faecium in a paediatric oncology unitrisk factors for colonization. Eur J Pediatr 157:20-27, 1998 Ibrahim EH, Sherman G, Ward S, et al: The influence of inadequate antimicrobial treatment of bloodstream infections on patient outcomes in the ICU setting. Chest 118:146-155, 2000 Iregui M, Ward S, Sherman G, et al: Clinical importance of delays in the initiation of appropriate antibiotic treatment for ventilator-associated pneumonia. Chest 122:262-268, 2002 Drusano GL: Prevention of resistance: a goal for dose selection for antimicrobial agents. Clin Infect Dis 36:S42-S50, 2003 (suppl) Kollef MH: An empirical approach to the treatment of multidrug-resistant ventilator-associated pneumonia. Clin Infect Dis 36:1119-1121, 2003 Hoban DJ, Biedenbach DJ, Mutnick AH, et al: Pathogen of occurrence and susceptibility patterns associated with pneumonia in hospitalized patients in North America: results of the SENTRY antimicrobial surveillance study (2000). Diagn Microbiol Infect Dis 45:279-285, 2003

29

84. Mathai D, Lewis MT, Kugler KC, et al: Antibacterial activity of 41 antimicrobials tested against over 2773 bacterial isolates from hospitalized patients with pneumonia: I—results from the SENTRY antimicrobial surveillance program (North America, 1998). Diagn Microbiol Infect Dis 39:105-116, 2001 85. Mathai D, Jones RN, Pfaller MA, et al: Epidemiology and frequency of resistance among pathogens causing urinary tract infections in 1,510 hospitalized patients: a report from the SENTRY antimicrobial surveillance program (North America). Diagn Microbiol Infect Dis 40:129-136, 2001 86. Burkhardt JE, Walterspiel JN, Schaad UB: Quinolone arthropathy in animals versus children. Clin Infect Dis 25:1196-1204, 1997 87. Levin AS, Barone AA, Penco J, et al: Intravenous colistin as therapy for nosocomial infections caused by multidrug-resistant Pseudomonas aeruginosa and Acinetobacter baumannii. Clin Infect Dis 28:1008-1011, 1999 88. Garnacho-Montero J, Ortiz-Leyba C, Jime´nez-Jime´nez FJ, et al: Treatment of multidrug-resistant Acinetobacter baumannii ventilator associated pneumonia (VAP) with intravenous colistina comparison with imipenem-susceptible VAP. Clin Infect Dis 36:1111-1118, 2003 89. Lai KK: Treatment of vancomycin-resistant Enterococcus faecium infections. Arch Intern Med 156:2579-2584, 1996 90. Low DE: Quinupristin/dalfopristin: spectrum of activity, pharmacokinetics, and initial clinical experience. Microb Drug Resist 1:223-234, 1995 91. Birmingham MC, Rayner CR, Meagher AK, et al: Linezolid for the treatment of multidrug-resistant, Gram-positive infections: experience from a compassionate-use program. Clin Infect Dis 36:159-168, 2003 92. Herrero IA, Issa NC, Patel R: Nosocomial spread of linezolidresistant, vancomycin-resistant Enterococcus faecium. N Engl J Med 346:867-869, 2002 93. Karlowsky JA, Draghi DC, Jones ME, et al: Surveillance for antimicrobial susceptibility among clinical isolates of Pseudomonas aeruginosa and Acinetobacter baumannii from hospitalized patients in the United States, 1998 to 2001. Antimicrob Agents Chemother 47:1681-1688, 2003 94. Gales AC, Jones RN, Forward KR, et al: Emerging importance of multidrug-resistant Acinetobacter species and Stenotrophomonas maltophilia as pathogens in seriously ill patientsgeographic patterns, epidemiological features, and trends in the SENTRY antimicrobial surveillance program (1997-1999). Clin Infect Dis 32:104-113, 2001 (suppl 2)