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SUPPLEMENT ARTICLE

Bacterial Resistance: Origins, Epidemiology,


and Impact
David M. Livermore
Antibiotic Resistance Monitoring and Reference Laboratory, Central Public Health Laboratory, London, United Kingdom

Most bacteria have multiple routes to resistance to any


drug and, once resistant, can rapidly give rise to vast
numbers of resistant progeny. Natural selection favors
mechanisms that confer resistance with the least fitness
cost and those strains that are least burdened by their
resistance. Selection may also favor determinants that
prevent their own counterselection and resistant strains
with enhanced survival ability or virulence. To this genetic and biochemical potential must be added the wide
variety of bacteria that cause opportunistic infections
in vulnerable human patients and the fact that the numbers of vulnerable patients grow steadily with advances
in other fields of medicine. In short, the emergence of
resistance is profoundly unsurprising; what is remark-

able is how long it has taken for the problem to become


a source of public, as well as scientific, concern.
Resistance can result from modification of an antibacterials target or from functional bypassing of that
target, or it can be contingent on impermeability, efflux,
or enzymatic inactivation. All members of a species may
be resistant. Alternatively, resistance may arise in hitherto
susceptible organisms via mutation or DNA transfer. The
aim of this article is not to catalog individual mechanisms
or their prevalencethat has been done elsewherebut
to emphasize the continuing dynamism of resistance, its
impact on therapy, and the difficultybut also the potentialfor combating the problem.

SELECTION OF SPECIES WITH INHERENT


RESISTANCE
Reprints or correspondence: Dr. David M. Livermore, Antibiotic Resistance
Monitoring and Reference Laboratory, Central Public Health Laboratory, 61
Colindale Ave., London NW9 5HT, United Kingdom (DLivermore@phls.nhs.uk).
Clinical Infectious Diseases 2003; 36(Suppl 1):S1123
 2003 by the Infectious Diseases Society of America. All rights reserved.
1058-4838/2003/3602S1-0004$15.00

Antibacterial use disrupts the microbial ecology of the


patient, unit, or population. Entire species may be selected. The increasing role of enterococci as opportunist
pathogens in the past 20 years partly reflects increasing
Bacterial Resistance CID 2003:36 (Suppl 1) S11

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The basic mechanisms of antibacterial resistance are well known, but critical new aspects continue to be discovered.
Recently discovered factors with major implications for the emergence, dissemination, and maintenance of
resistance include multidrug efflux, hypermutability, integrons, and plasmid addiction. Some resistances are
widespread and others local, with prevalence rates often worst in newly prosperous countries and in those
specialist units where antibacterial use is heaviest. Multidrug-resistant epidemic strains are critical to the total
accumulation of resistance (e.g., among Streptococcus pneumoniae, methicillin-resistant Staphylococcus aureus,
Klebsiella pneumoniae), but it remains unclear why some bacterial lineages achieve epidemic spread whereas
others that are equally resistant do not. The correlation between in vitro resistance and treatment failure is
imperfect, but resistance undoubtedly increases mortality, morbidity, and costs in many settings. Recent concern
has led to a plethora of governmental and agency reports advocating less antibacterial use, better antibacterial
use, better infection control, and the development of new antibacterials. The evidence that better prescribing
can reduce resistance rates is mixed, and although changes to hospital regimens may reduce one resistance
problem, other opportunistic bacteria may fill the vacant niche. Overall, the best that can reasonably be anticipated
is an improved balance between the accumulation of resistance and new antibacterial development.

RESISTANCE VIA MUTATION


As DNA is replicated, uncorrected base substitutions occur randomly, at a frequency of 109 to 1010 per gene [8]. In addition, copying errors may lead to the partial or complete deletion of individual genes [8]. As a result, the targets of
antibacterials may be altered, drug-inactivation or efflux systems may be up- or downregulated, and uptake pathways (porins and active transporters) may be lost or activated. Resistance
genes or their repressors also can be activated or inactivated
by the migration of insertion sequences. Approximately 3% of
Bacteroides fragilis isolates have the carbapenemase gene ccrA
(cfiA), but its enzyme product is expressed only if an insertion
sequence has migrated upstream of this structural gene [9].
Classical experiments showed that antibacterials cause the
selection of preexisting variants, not the emergence of new
mutants. This observation entirely agrees with the precepts of
Darwinian evolution, but a twist is given by the observations
that bacteria can become hypermutable through inactivation
of the proofreading and DNA mismatchrepair systems that
normally correct DNA copying errors [8]. Hypermutators have
up to 200-fold higher mutation rates than normal cells and so
are more likely to become resistant to a first antibacterial by
mutation. Once selected by this first drug, they are then
primed to develop resistance to further agents. To this extent,
antibacterials may cause the emergence of variants with an
increased propensity to develop further resistance. HypermutS12 CID 2003:36 (Suppl 1) Livermore

ability also may arise transiently through induction of the SOS


system, a stress response that involves the expression and function of alternative DNA polymerases with reduced copying fidelity [8, 10]. The SOS system is induced, inter alia, by starvation, and it is also notable that the spectrum of mutations
seen in non- or slow-growing cells differs from those among
logarithmic-phase organisms. Also relevant is the fact that
DNA-damaging forms of reactive oxygen accumulate in nongrowing cells [8, 11], perhaps acting as mutagens.
The selection or transient induction of hypermutability may
explain why variants with multiple mutations have emerged
more rapidly than was predicted from laboratory studies. Nonclonal strains of fluoroquinolone-resistant Escherichia coli have
emerged worldwide and have become highly prevalent, for example, in India [12], Spain [13], and China [14]. This is despite
the fact that substantive fluoroquinolone resistance in Enterobacteriaceae requires mutations to the genes for subunits of
topoisomerases II and IV (gyrA and parC, respectively), together with further mutations that upregulate efflux and reduce
permeability, or both [1417]. Perhaps the emergence of these
multiple mutants is favored by the quinolone-mediated induction of the SOS response, with its contingent hypermutability [18]. This is speculation, but there is hard evidence of
the role of hypermutators in the cystic fibrosis lung, where
Oliver et al. [19] found that 36% of 30 chronically colonized
patients carried hypermutable Pseudomonas aeruginosa. Hypermutators were not found in 75 control (noncystic) patients
with acute P. aeruginosa infections.
Most mutations affect only a single antibacterial class, but
those affecting impermeability or efflux may have a pleiotropic
effect. The potential for porin loss to affect multiple drug classes
by restricting nonspecific permeability is self-evident, but the
role of broad-spectrum efflux has come as a surprise. Its function is best understood in P. aeruginosa, where mutation at
mexR upregulates the mexA-mexB-oprM operon and raises the
MICs of b-lactams (except imipenem), fluoroquinolones, tetracyclines, chloramphenicol, macrolides, various disinfectants,
detergents, and organic solvents [20]. MexAB-OprM also plays
a roleperhaps coincidentallyin excretion of the quorumsensing mediator, homoserine lactone. Even in typical P. aeruginosa isolates, MexAB-OprM is expressed to some degree and
accounts for much of the resistance previously ascribed to impermeability [21]. Isolates with further upregulation have additional resistance to substrate drugs (table 1). P. aeruginosa
strains have 3 further well-characterized efflux systems, 2 of
which (MexCD-Opr J and MexEF-Opr N) are normally repressed but may be activated by mutation (table 1). The genomic sequence for P. aeruginosa suggests the presence of at
least 5 further efflux systems, which await even preliminary
characterization [20]. Multidrug efflux systems are also being
reported in many other gram-negative bacteria, including En-

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use of fluoroquinolones and cephalosporins, to which these


organisms are inherently resistant [1, 2]. The increasing role
of coagulase-negative staphylococci (especially) and a-hemolytic streptococci in hematology patients also may reflect antimicrobial use: a-hemolytic streptococci are resistant to fluoroquinolones, which are often used as prophylaxis in these
patients, and coagulase-negative staphylococci frequently have
acquired multidrug resistance [3]. A greater factor behind the
rise of coagulase-negative staphylococci is, however, the increased use of indwelling lines, which provide entry portals for
the skin microflora. Among gram-negative pathogens, Acinetobacter baumannii and Stenotrophomonas maltophilia are increasingly prevalent in many intensive care units (ICUs), with
A. baumannii notoriously associated with ventilator-associated
pneumonias [4]. A. baumannii commonly has acquired resistance to all antibacterials except carbapenems, minocycline,
sulbactam, and colistin, whereas S. maltophilia is often resistant
to all antibacterials except co-trimoxazole (and perhaps ticarcillin-clavulanate). Some publications associate an increasing
incidence of S. maltophilia infections with carbapenem use [5],
but others find a less specific relationship to the use of multiple
antibacterials, including those (e.g., erythromycin) with predominantly antigram-positive spectra [6, 7].

Table 1.

Drug efflux systems of Pseudomonas aeruginosa.


Regulatory
gene

Mutation causing
upregulation

MexAB-OprM

mexR

nalB (affects mexR) and nalC


(lies outside mexR)

MexCD-OprJ

nfxB

nfxB

b-Lac (not imipenem), Cm, Em, Fq, Nv, Tm, Tc, ethidium bromide, acriflavine,
SDS, aromatic hydrocarbons, triclosan

MexEF-OprN

mexT

nfxC

Cm, Fq, Tm, aromatic hydrocarbons, triclosan

MexXY-OprM

mexZ

System

Substrates
b-Lac (not imipenem), Cm, Em, Fq, Nv, Tm, Su, Tc, ethidium bromide,
acriflavine, SDS, aromatic hydrocarbons, irgasan, triclosan, homoserine
lactone

Agl, b-lac (not carbenicillin, ceftazidime or imipenem), Cm, Em, Fq, Nv, Tc

NOTE. Agl, aminoglycosides; b-lac, b-lactam; Cm, chloramphenicol; Em, erythromycin; Fq, fluoroquinolones; Nv, novobiocin; SDS, sodium dodecyl sulfate;
Su, sulfonamides; Tc, tetracycline; Tm, trimethoprim. Data from [20, 21].

there has been concern about linezolid resistance emerging in


enterococci or, more rarely, in Staphylococcus aureus via modification of the domain V of 23S rRNA, which contains the
drugs binding site. Mutational resistance to linezolid is difficult
to obtain in vitro, apparently because the bacteria have multiple
copies of the encoding gene, with modification of more than
one copy required for resistance. Nevertheless, linezolid resistance can be selected in vivo, particularly in difficult-to-reach
infections, those requiring prolonged therapy and if the drug
is underdosed [31, 32]. It is not clear whether DNA recombination events follow the mutation of one gene copy or
whether multiple gene copies undergo sequential mutations,
perhaps by induction of a transient hypermutability (above).
ACQUISITION OF RESISTANCE BY DNA
TRANSFER
DNA transfer among bacteria is critical to the dissemination
of resistance and has recently been reviewed [33]. Transfer of
DNA is most often via plasmids. These existed long before
humans used antibacterials but did not then carry resistance
determinants, or rarely did so [34]. Since the advent of the
antibacterial era, plasmids have, however, proved to be the ideal
vehicles for recruitment and dissemination of resistance genes.
Within plasmids, resistance genes are often carried by transposons, which can shuttle determinants between more and less
promiscuous plasmids, or into and out of the chromosome.
Some transposons are directly transmissible between bacteria,
particularly (but not exclusively) among gram-positive species.
Resistance genes also may be transferred by lysogenic bacteriophage. This latter mechanism seems likely with the mecA
determinant staphylococci, which has never been located on
mobile DNA but which has spread among a few S. aureus
lineages, and among different coagulase-negative species [35].
Integrons are natural recombination systems that facilitate
the acquisition and expression of resistance determinants behind a single promoter. They are widely distributed among
gram-negative bacteria, often occurring within plasmids and
Bacterial Resistance CID 2003:36 (Suppl 1) S13

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terobacteriaceae as well as other nonfermenters [20, 22]. At a


functional level, the role of broad-spectrum efflux pumps may
be to remove amphipathic substances (i.e., those with hydrophilic and hydrophobic parts) from membranes, preventing
disorganization. Such a cleaning role would account for their
broad activity and wide distribution of these pumps, but their
substrate recognition sites await elucidation
Efflux systems can be coregulated with porin expression. In
P. aeruginosa, the nfxC mutation at mexT upregulates MexEFOprNmediated efflux (table 1), and also reduces expression
of OprD (D2 porin in older literature), thereby reducing
permeability to carbapenems [23]. In E. coli (and, probably,
other Enterobacteriaceae), mutation at mar simultaneously affects the expression of 160 genes [24] reducing, inter alia, expression of porin OmpF and upregulating AcrAB, which is
implicated in the efflux of b-lactams, fluoroquinolones, chloramphenicol, tetracycline, dyes, and detergents [25].
If mutational resistance emerges at high frequency without
deleterious side effects, it can swiftly undermine an antibacterials utility by allowing multifocal emergence of resistance
wherever the drug is used. Several antimicrobial groups introduced in the 1980s and 1990s are prone to mutational resistance, at least with some species. Early optimism about fluoroquinolones against staphylococci was dispelled by the discovery
that resistance emerged by upregulation of NorA-mediated efflux [26]. Oxyimino-aminothiazolyl cephalosporins at first appeared almost universally active against Enterobacter, Citrobacter, and Serratia spp., but it soon became apparent that activity
depended on a failure to induce chromosomal AmpC b-lactamases [27]. Activity was lost against derepressed mutants,
which hyperproduce AmpC b-lactamases independently of induction. Such mutants arise at frequencies of 105 to 107 and
are selected in 20% of patients treated with cephalosporins
for Enterobacter bacteremia, with this selection often leading to
clinical failures [28]. Although imipenem evades this mechanism, imipenem resistance emerges readily in P. aeruginosa by
loss of the OprD porin, which provides carbapenem-specific
pores through the outer membrane [29, 30]. Most recently,

Table 2.
DNA.

Sources of b-lactamase resistance genes now found on transferable

Gene(s) or product(s)

Source

SHV b-lactamases

Klebsiella pneumoniae chromosome

CTX-M2,4,5,6,7 and Toho-1 b-lactamases

Kluyvera spp. chromosome

CMY-2, 3, 4, 5, 6, 7, LAT-1, -2, -3, -4, BIL-1


AmpC b-lactamases

Citrobacter freundii chromosome

ACC-1 AmpC b-lactamases

Hafnia alvei chromosome

DHA-1 and -2 AmpC b-lactamases

Morganella morganii chromosome

TEM, OXA,a PSE, staphylococcal


penicillinase

Unknown

Although source organisms for plasmid-mediated OXA enzymes have not been identified,
some Aeromonas spp. have chromosomal enzymes belonging to this family.

S14 CID 2003:36 (Suppl 1) Livermore

and has been known for almost as long. Nevertheless, for no


obvious reason, it is 10-fold less prevalent that TEM-1 b-lactamase in every country and species surveyed [27].
Many of the resistance determinants now found on plasmids
are believed to have originated in the chromosomes of other
species, although only a few of their source organisms have
been identified definitively. The plasmid-mediated SHV b-lactamases are derived from the chromosomal b-lactamases of
Klebsiella pneumoniae; the plasmid-borne AmpC enzymes
emerging in Klebsiella spp. and E. coli are chromosomal escapes
from Citrobacter freundii, Hafnia alvei, Morganella morganii,
and Enterobacter cloacae [42, 43], and several CTX-M cefotaximases are chromosomal escapes from Kluyvera spp. [44] (table
2). Some resistance determinants to nonb-lactam drugs seemingly originated in antibiotic-producing streptomycetes, which
must protect themselves against their own products. Examples
include several aminoglycoside-modifying enzymes and the erm
determinants, which have products that methylate 23S rRNA
so as to block binding of macrolides, lincosamides, and group
B streptogramins [45]. Also, the genes that encode the d-alad-ala ligases are critical to glycopeptide resistance in enterococci
(and now also S. aureus [46, 47]). Many plasmids and transposons carry multiple resistance genes conferring resistance to
different antibacterials, and selection for any one determinant
may conserve the entire plasmid and its resistances. The frequent consequence is multidrug resistance, as illustrated in table
3, which shows that klebsiellae with extended-spectrum b-lactamases (ESBL) were more often resistant to aminoglycosides
and fluoroquinolones than those without ESBLs [48, 49]. ESBLs
and aminoglycoside-modifying enzymes may be encoded by
single plasmids, but the fluoroquinolone resistance, also seen
in an excess of the ESBL producers, is chromosomal and
independent.
The problems of multiresistance are increasing and becoming
more complex. When plasmid electrophoresis was developed
in the 1970s, it was uncommon to see isolates with 123 plasmids, and plasmid profiles were used to define strain epide-

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transposons, and are particularly important in the dissemination of resistance genes to sulfonamides (sul1), and streptomycin (aadA3) [36]. Other genes often located in integrons
include those for various OXA, PSE, VIM, and IMP b-lactamases and for many aminoglycoside-modifying enzymes [36,
37]. In principle, integrons have a fearsome capacity for the
recruitment, spread, and expression of resistance genes, and
surveys show that they are widespread among gram-negative
bacteria in countries as far apart as The Netherlands and Taiwan
[38, 39]. Nevertheless, it is striking that the most successful blactamase genes (i.e., blaTEM derivatives) are carried directly by
transposons, not within integrons, whereas the integron-associated OXA and PSE b-lactamases are considerably rarer [27].
Similarly with sulfonamide resistance: sul2, which is not integron-associated, is increasing in prevalence among E. coli in
the UK, whereas sul1, which is integron-determined, is stable
in prevalence, or declining [40]. It is notable also that the
composition of integrons is more stable over time than might
be expected [38]. In short, integrons are important, but their
importance should not be overplayed relative to that of other
vehicles of resistance.
The dissemination of plasmids, transposons, and integrons
among bacteria and species give rise to so-called gene epidemics. Plasmids encoding the TEM plasmid-mediated b-lactamases were first recognized in 1965 but have since spread
varying with the country, unit, and speciesto 30%60% of
clinical Enterobacteriaceae, to a few P. aeruginosa, and to anywhere between 1%50% of Haemophilus influenzae and Neisseria gonorrhoeae isolates [27]. Other determinants that have
spread extremely widely include erm, sul1, sul2 strA, strB, aad3,
tetA, and tetM. tetM has spread in both gram-positive and
gram-negative organisms, but more generally, the genes of
gram-positive and gram-negative species are distinct [41]. The
factors that determine whether a mobile gene will spread widely
are poorly understood. TEM-2 b-lactamase differs from TEM1 by a single amino acid substitution, confers similar resistance,
is coded by similarly promiscuous plasmids and transposons,

Table 3. Multidrug resistance among Klebsiella spp. collected in 2 European surveys


of isolates from intensive care units.
1994 survey

19971998 survey

ESBL
positive
(n p 220)

ESBL
negative
(n p 746)

ESBL
positive
(n p 110)

ESBL
negative
(n p 323)

Klebsiellae with the indicated ESBL


phenotype

23

77

25

75

Gentamicin resistance

76

72

10

Amikacin resistance

52

61

Ciprofloxacin resistant

31a

31

Resistance

NOTE.
a

Data are percentage of isolates. ESBL, extended-spectrum b-lactamase. Data from [42, 43].

Declined to 10% if members of a single widespread clone were discounted.

EPIDEMIOLOGY OF RESISTANCE: LOCAL,


NATIONAL, AND INTERNATIONAL
At one level, the epidemiology of resistance is extremely local.
Most outbreaks and clusters involve a few patients in a unit,
and the prevalence of resistance is often highest in those units
where the most vulnerable patients are congregated and where
antibacterial use consequently is heaviest. Archibald et al. [54]
found 2-fold higher rates of methicillin resistance among
staphylococci, ceftazidime resistance among E. cloacae and P.
aeruginosa, imipenem resistance among P. aeruginosa, and vancomycin resistance among enterococci in patients in ICUs than

in patients in general wards or outpatients at the same hospitals.


In virtually all European countries, the prevalence of methicillinresistant S. aureus is higher in ICUs than in general wards [55].
At another level, the epidemiology of resistance is national.
In Europe, the common pattern is for resistance to increase in
prevalence as one moves southward: it is lowest in Scandinavia
and highest in the Mediterranean countries. In North America,
resistance rates are mostly higher in the United States than in
Canada. Some of the worst resistance rates are in the newly
prosperous countries of East Asia and South America. A few
examples: methicillin-resistant strains comprise 30%45% of
all S. aureus from bacteremias in Spain, Portugal, Italy, France,
and the United Kingdom and 10%15% in Germany and Austria, but !1% in the Netherlands and Scandinavia (see, e.g.,
European Animicrobial Resistance Surveillance System, http://
www.earss.rivm.nl). In Korea, Japan, Taiwan, and Vietnam,
70%80% of S. pneumoniae are resistant or intermediately resistant to penicillin, compared with 30%40% in France and
Spain, 5%10% in the United Kingdom, and 1%2% in Scandinavia [5658]. As a final example, gentamicin resistance in
E. coli remains considerably more frequent in most Southern
European countries and the United States than in the United
Kingdom, where it occurs in !3% of isolates [57].
Finally, the epidemiology of resistance is partly international,
with some transferable determinants prevalent worldwide. The
epidemiology is also international to the extent that some resistant strains spread between countries and continents. Multidrug-resistant pneumococci of serotype 6B were imported from
Spain into Iceland, apparently by nasopharyngeal carriage in
the children of returning holidaymakers [59]. These pneumococci then became established in child care centers in Iceland, causing an increase in the penicillin resistance rate from
1% in 1988 to 17% in 1993. Other penicillin-resistant pneumococci of serotype 23F have spread from Spain to the Far
East, the Americas, and South Africa [60]. On a smaller scale,
many of the few E. coli and Klebsiella spp. with plasmid-mediated AmpC b-lactamases in the United Kingdom are epiBacterial Resistance CID 2003:36 (Suppl 1) S15

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miology. Nowadays, it is common to encounter Enterobacteriaceae with 56 plasmids; moreover, plasmid profiles often vary
greatly within the strains defined by PFGE [50]. These observations (which complicate epidemiological analysis) reflect
the gain and loss of plasmids by different representatives of the
same strain and the gain and loss of genes among different
representation of the same plasmid. Even when genes are not
gained or lost, multiple copies of the same plasmid may carry
different variants of a b-lactamase gene, reflecting mutation.
Thus, for example, some members of a K. pneumoniae strain
from Turkey had SHV-3 b-lactamase, whereas others had SHV5 (table 4) [50]. More strikingly, 84 blaTEM and blaSHV copies
were found among just 25 K. pneumoniae isolates collected at
one hospital in Durban, South Africa [51].
A few speciesNeisseria spp., Haemophilus spp., and a-hemolytic streptococcican absorb and incorporate DNA released by dead cells of related organisms, allowing the generation of mosaic genes [52]. This mechanism is absent from
most other species, which restrict incoming DNA. Mosaic gene
formation, acting in combination with mutation, is the basis
of emerging penicillin resistance in pneumococci and of blactamaseindependent ampicillin or penicillin resistance in H.
influenzae and Neisseria spp.; it is also implicated in sulfonamide resistance in H. influenzae [53].

Table 4. Diverse b-lactamases and resistances of multiple representatives of a serotype


K62 Klebsiella pneumoniae strain collected at an intensive care unit in Istanbul, Turkey.
MIC, mg/L
Isolate

Gentamicin

Aztreonam

Ceftazidime

1951

512

1024

256

1953

16

512

1024

256

1954

32

1959

16

512

512

1964

16

512

512

NOTE.

Ceftriaxone

Piperacillintazobactam

b-Lactamase

SHV-5

SHV-3 TEM

1024

SHV-3 TEM

512

1024

SHV-5 TEM

256

512

SHV-3 TEM

0.5

Data from [50]. All 5 representatives had the same DNA profile as investigated by PFGE.

EPIDEMIC RESISTANT STRAINS


Successful epidemic strains are critical to the accumulation of
many resistances. Common vectors in hospitals are contact with
staff members, contact with nonsterile devices, or procedures.
Spread in the community is favored by those factors that have
aided epidemics throughout history: crowding and travel. Many
strains, resistant or otherwise, spread locally, but a few achieve
a much wider distribution. The international spread of penicillin-resistant pneumococcal lineages was mentioned above,
but further examples abound. In England and Wales, the proportion of S. aureus bacteremias caused by methicillin-resistant
S. aureus (MRSA) was steady at 1%3% from 1989 to 1993
but increased rapidly afterward, reaching 42% by 2000 [57].
The beginning of this increase coincided with the emergence
of 2 new epidemic (E) strains, EMRSA 15 and 16, and recent
analysis shows that these now account for 193% of all S. aureus
bacteremias in England and Wales [66]. In France, a serotype
K25 K. pneumoniae strain with SHV-4 b-lactamase and crossresistance to amikacin and ciprofloxacin has disseminated
widely, having been reported repeatedly since 1988, first around
Paris and subsequently in hospitals from the Atlantic coast to
the Mediterranean, with reports also from Ghent in Belgium
[50, 67, 68]. A survey of ESBLs among 966 klebsiellae in 1994
included 35 centers, only 5 of them in France, yet this single
strain accounted for 52 of 220 ESBL producers collected [50].
Also in France and Belgium, an Enterobacter aerogenes strain
with TEM-24 b-lactamase and multiresistance to aminoglycosides, quinolones, and, occasionally (via porin loss), carbapenems has become widely established [69]. Other examples
S16 CID 2003:36 (Suppl 1) Livermore

where resistance has a major clonal element include Burkholderia


cepacia from cystic fibrosis patients [70] and Salmonella typhimurium, where major recent problems (now perhaps declining)
have been associated with the intercontinental spread of multidrug-resistant lineages of definitive type DT104 [71, 72].
In the case of vancomycin-resistant Enterococcus faecium, recent studies by amplified DNA restriction fragment-length
polymorphism suggest that epidemic strains from hospitals in
Europe, the United States, and Australia, although differing
from place to place, are more closely related to each other than
to sporadic and agricultural isolates [73].
Other strains with similar resistances to these epidemic organisms are recorded, often in the same hospitals or patient
groups, but these fail to spread extensively or fail to spread at
all. The reason for epidemic success remain obscure, but potential factorsnot mutually exclusiveinclude the following:
(1) increased adherence to host cells or prosthetic materials,
(2) greater tolerance of desiccation, (3) elevated resistance to
disinfectants, (4) faster growth rates, and (5) better adaptation
to the fitness cost of resistance. Hard evidence for the role of
any of these factors is scanty for many particular strains, and
studies are confounded by the fact that researchers, having
found one fact that may contribute to epidemic success, then
concentrate on that factor in isolation. In the case of the successful serotype K25 K. pneumoniae strain from France and
Belgium, one report suggests a plasmid-mediated fimbrial antigen that aids adherence to the gut mucosa [74].
Local variation, national variation, and role of epidemic
strains should be borne in mind when undertaking and assessing prevalence surveys. Grandiloquent statements about resistance rates in Europe or the Western Pacific abound in
the literature but often are based on surveys with a very few
centers per country, and with the extent of resistance differing
greatly among the countries included within a region. The variation in resistance prevalence between Stockholm and Madrid,
it should be noted, is vastly greater than between Boston and
San Francisco. Inferences on the general distribution of resistance likewise should not be made on the basis of data for

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demiologically linked to the Indian subcontinent, where there


is evidence of local frequency in Punjab [61, 62]. Last, PER-1
ESBL was first recorded from a P. aeruginosa isolate collected in
France [63] and shortly afterward was found in numerous P.
aeruginosa, Salmonella, and Acinetobacter spp. isolates from several cities in Turkey [64]. An inquiry revealed that the original
source patient in France was a Turk, visiting for treatment [65].

isolates from ICUs, where resistance rates are commonly higher


than among isolates overall [54, 55]. A finaland biggersource
of bias, particularly for surveys of community isolates, is that
microbiological investigations are more likely to be performed
for recalcitrant infections, which may be recalcitrant because
they are caused by resistant bacteria. Thus, MacGowan et al.
[75] found that only 3% of patients presenting with respiratory
symptoms to general practitioners in Bristol received microbiological investigation and that the apparent prevalence of
ampicillin resistance among H. influenzae isolates decreased
from 22% to 11% when all the presenting patients had sputum
culture performed.

IMPACT OF RESISTANCE
The consequences of resistance are harder to define than microbiologists, health care managers, and politicians might wish.
Some patients recover despite inadequate treatment, exactly as
some recovered before antibacterials were available. The infection of others failed to respond despite appropriate therapy. In
compromised patients, it often remains debatable whether infection or an underlying disease was ultimately fatal. Perhaps
the clearest link between in vitro resistance and in vivo responses for penicillin and gonorrhea. Classical strains with
MIC 0.06 mg/L respond to penicillin at low doses; those with
chromosomally mediated resistance (MIC 0.252 mg/L) respond to high-dose penicillin; and those with b-lactamase cannot be cured by any achievable level of penicillin. However,
gonorrhea is an uncomplicated infection mostly caught by
those in otherwise good health! Elsewhere, relationships are less
clear and pseudomonal infection in cystic fibrosis presents the

opposite extreme. Drugs that are active in vitro consistently fail


to eliminate infection, whereas some of those that entirely lack
in vitro antipseudomonal activity (e.g., erythromycin) often
ameliorate the symptoms of infection [76].
Most correlations fall between these extremes. Analysis by
Mosdell et al. [77] showed that the incidence of complications,
including reoperation, abscess formation, and wound infection,
increased 2-fold if empirical therapy for intra-abdominal sepsis failed to cover all the pathogens subsequently isolated and
that the length of hospital stay was likewise extended (figure
1). The incidence of complications rose further if inadequate
empirical regimen was not modified when resistant pathogens
were isolated. In recent analyses, Kollef and Ward [78] and
Ibrahim et al. [79] found 2-fold higher mortality among ICU
patients and those with ventilator-associated pneumonia when
the pathogens proved resistant to the antibacterials used empirically (figure 2). Many of the failures in these series reflected
infection by P. aeruginosa or MRSA, which were not well covered by the empirical regimens routinely used. Risk factors for
isolation of these pathogens and so for poor outcomes included
previous use of antibacterials and previous hospitalization. Such
factors, as well as the likely pathogens and their likely local
resistance patterns, should always be taken into account when
designing empirical regimens for hospital units. It should be
added that the science of pharmacodynamics is allowing more
precise modeling of the probability of cure of a given pathogen
in a given infection, underscoring the fact that for many infection types, there is a strong relationship between in vitro
susceptibility and outcome. These aspects are addressed elsewhere in this supplement by Drusano [80].
Increased morbidity and mortality are the most dramatic
Bacterial Resistance CID 2003:36 (Suppl 1) S17

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Figure 1. Complications among patients with peritonitis, in relation to whether initial empirical treatment was appropriate (group 1, white); was
changed from inappropriate to appropriate after isolation of resistant bacteria (group 2, gray); or was not changed despite the isolation of resistant
bacteria (group 3, black). Data are replotted from Mosdell et al. [77]. For total complications and reoperation, there were significant differences
(P ! .01, x2 test) between groups 1 and 3, and for abscess formation between groups 1 and 2 and between 1 and 3. Differences in wound infection
rates do not achieve statistical significance. Average lengths of stay were 10.9, 14.8, and 19.0 days for group 1, 2, and 3 patients, respectively, with
a significant difference (P ! .01, paired t test) between groups 1 and 3.

Figure 2. Mortality among critically ill patients in intensive care (n p 655 ) in relation to whether empirical therapy was appropriate (gray) or
inappropriate (black) in relation to the resistances of the bacteria subsequently isolated. Reproduced with permission from Ibrahim et al. [79].

RESPONSES TO RESISTANCE
Concern about resistance increased in the late 1990s. Since then,
many governmental and agency reports have been published,
adding to those of professional societies [8284]. These reports
vary in emphasis, especially as regards the agricultural use of
antibacterials, but all advise (1) less use of antibacterials, (2)

S18 CID 2003:36 (Suppl 1) Livermore

more appropriate choices of antibacterials and regimens, (3)


prevention of cross-infection, and (4) development of new antibacterials. Measuring the effect of these charges demands better surveillance of resistance prevalence and of prescribing. Optimists hold that it may be possible to reverse resistance trends,
and pessimists hold that it may only be possible to slow the
accumulation of resistance sufficiently to keep one step ahead
of bacterial evolution.
EFFECTS OF REDUCED PRESCRIBING
In a few cases, reductions in prescribing at a national level have
been followed by a reduced prevalence of resistance. In one
example of success, the prevalence of penicillin-resistant pneumococci in Iceland was reduced from 19.3% (1993) to 14%
(19982000) after a 12.9% reduction in drug use [85]. In Finland, a national advisory to reduce prescribing of macrolides
was followed by a decline in use from 3 doses/1000 population
per month in 1988 to 1.1 doses/1000 population per month in
1994 and by a decrease in the prevalence of erythromycinresistant Streptococcus pyogenes from 19% in 1993 to 8.5% in
1996 [86]. By 1998, however, macrolide use had increased to
2.1 doses/1000 population per month, and the resistance rate
among S. pyogenes was back to 18% (Bacterial Resistance to
Antimicrobial Agents in Finland FINRES 1999; http://www
.mmm.fi/el/julk/finres99en.htm). In both these cases, the resistances displaced were associated with clonal strainstwo
widely disseminated S. pyogenes lineages in Finland [87]and,
in Iceland, the serotype 6B S. pneumoniae strain mentioned
earlier. Displacement seems more difficult when resistances are
multiple and linked and when they have disseminated among
different strains. Resistance to streptomycin and chloramphenicol remains frequent in gram-negative bacteria, although these
drugs have fallen into virtual disuse in humans, and are compromised by mechanisms that do not directly affect any antibacterial that remains in extensive human use [88]. As a further

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consequence of resistance. Other effects are more insidious.


Physicians and surgeons are forced to use previously reserved
agents as first-line therapy. These may be inherently less potent
or more toxic that classical regimens: vancomycin is increasingly used as a first-line antistaphylococcal (and for prophylaxis) but is less convenient to administer safely and less bactericidal than the semisynthetic antistaphylococcal penicillins,
which themselves are 100-fold less active than benzylpenicillin
against fully susceptible staphylococci. Previously reserved
agentsnow used earliermay be undermined by resistance.
The accumulation of cephalosporin-resistant bacteria is driving
the earlier clinical use of carbapenems and is a reasonable justification for the development of oral and long half-life carbapenems. Nevertheless, the selection pressure that mass use
of these will cause is disturbing, coming precisely when the
number of reported carbapenemases is growing sharply [81].
Finally, resistance adds cost: treatment failures extend the
length of hospital stay or demand repeated physician visits;
hospital beds are blocked to new patients, and productive time
is lost. If new or hitherto reserved antibacterials are needed as
therapy, these are usually more expensive than previous regimens. These costs seem unlikely to decline in the future, especially with the growing demand of regulators and the new
costs of genomics-based drug discovery. A new antibacterial
already costs $0.5 billion to develop; this sum and the financing costs (for the $0.5 billion is spent before income is
generated) must be recouped in the 1012 years of patent life
remaining after the compound is launched.

Table 5. Incidence of colonization and infection with vancomycin-resistant enterococci in a London


hematology unit in relation to preferred empirical therapy

Phase

Empirical therapy for fever

Colonized with
vancomycin-resistant
enterococci, %

No. of
infections with
vancomycin-resistant
enterococci/total
no. of patients

Ceftazidime

57

5/75

2a

Pip/tazo infection control reinforced

29

0/70

2b

Pip/tazo infection control reinforced

0/59

Ceftazidime infection control reinforced

36

3/58

Each phase was defined as a 4-month period; identical regimens were used in phases 2a and 2b.

ing. Since an annual peak in 19961997 (the British financial


year runs from April to March), antibacterial prescriptions in
the community have decreased by 23.4% and veterinary use
by 25%.
FORMULARY CHANGES AND RESISTANCE
PATTERNS
Although there are few examples of reducing resistance on any
large scale, there are many examples of altered (rather than
reduced) prescribing being followed by changes in the local
epidemiology of resistance. A formulary switch to amikacin
was followed, over several years, by declining gentamicin resistance among opportunistic gram-negative bacteria [92, 93].
More recently, Bradley et al. [94] found that a formulary switch
from empirical ceftazidime to piperacillin-tazobactam in neutropenic fevers, together with reinforced infection control, was
associated with reduced colonization and infection by vancomycin-resistant enterococci (table 5). A return to empirical
ceftazidime was followed by a reemergence of vancomycinresistant enterococci, despite continued emphasis on infection
control. Other examples of apparent success include reductions
in the prevalence of AmpC-derepressed Enterobacter spp. after
a formulary switch from ceftazidime to cefepime [95] and of
cephalosporin-resistant Enterobacteriaceae after switches to
piperacillin-tazobactam or carbapenems [9698].
These examples highlight positive consequence, but caveats
should be noted. First, changes that achieve positive consequences are perhaps more likely to be reported than those that
fail to do so. Second, others who followed these strategies were
not always so successful. Formulary switches from gentamicin
to amikacin have been associated with increasing amikacin resistance [99]. Third, authors often concentrate on one pathogen
and demote other effects; thus, deaths owing to multidrugresistant gram-negative bacteria occurred in both the ceftazidime and piperacillin-tazobactam arms of the study outlined
in table 5 [94], and a formulary switch from cephalosporins

Bacterial Resistance CID 2003:36 (Suppl 1) S19

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example of the difficulty of displacing resistance, sulfonamide


use in the United Kingdom declined by 97% during
19911999 after national advice to prescribe trimethoprim instead of co-trimoxazole, but the prevalence of sulfonamide resistance among E. coli in London remained virtually constant,
at 39%45% [40].
The continued prevalence of resistance in disused antibacterials begs the question, Why? Potential contributory factors
include continued agricultural use andas was shown with the
sulfonamide resistance in E. coli from London [78] linkage
of resistance determinants within large multiresistance plasmids. More generally, evolution acts to favor those determinants that exert the least fitness cost on their host bacteria and
those strain variants in which this cost is minimized. Thus,
once antibacterials have been used heavily for long periods,
evolution seems likely to have honed the resistant strains. Laboratory experiments illustrate how this process may arise. An
E. coli strain with a tetracycline resistance plasmid initially grew
more slowly than its plasmid-free parent, but after 500 generations, it grew 6% more rapidly [89]. Similarly, the growth
rate of a streptomycin-resistant E. coli mutant was initially suppressed by 12%14%, but after repeated subculture, it grew
only 6% more slowly than the parent strain [90]. Such welladapted strains are unlikely to be swiftly displaced. Plasmids
themselves should perhaps be seen as selfish genomes, seeking
their own perpetuation, and not as mere possessions of their
host bacteria. Some plasmids have specific binding sites to the
chromosome, favoring segregation with each daughter cell.
Others encode plasmid addition systems determining both a
long half-life toxin and a homologous but less stable antitoxin.
So long as the plasmid is present, antitoxin is manufactured
and the cell survives, but residual toxin kills any daughter cell
that fails to inherit a plasmid copy [91]. Such factors maintain
plasmid carriage within populations, increasing the pressure to
minimize the fitness cost. It remains plausible (but unproven)
that a generalized reduction in prescribing may lead to a reduction in resistance prevalence. An answer may be forthcom-

SOCIAL CONTEXT
The scientific answers to these questions on antibacterial use
are uncertain, and to complicate matters, the whole problem
of resistance is intertwined with moral, social, political, and
commercial issues. Concern about resistance is used as ammunition for other agendas, most obviously including marketing by the pharmaceutical industry and cost containment
within managed or socialized health care. To some extent, the
individual patient gains when powerful antibacterials are used
early, but the resistance risks for society are raised. In reality,
matters are complex. Failed treatments with old and simple
drugs may lead to more severe disease or to the spread of
infection, resulting in a demand for further therapy, along with
its contingent selection pressure. Hungary, before 1989, had a
restricted list of antibacterials for community prescription, yet

S20 CID 2003:36 (Suppl 1) Livermore

achieved one of the worlds highest prevalence rates for penicillin-resistant pneumococci [105]. Moreover, the argument assumes a vacuum in which no new drugs are developed. This
assumption, made in many reports on resistance, is already
untrue for gram-positive pathogens [106].
Concerns about resistance have led to the banning of most
agricultural growth promoters in Europe. Such concerns are
also used to support wider objections to intensive farming and
to the genetic modification of crop plants (where unexpressed
resistance genes remain within the cloning vectors). Less is said
on the other aspects of modern life that potentially exacerbate
resistance: large hospitals; the concentration of the very young
and very old in socialized care; and increasing travel. Action
on these would be socially and politically impossible, even if
they are more pertinent to the sum total of resistance than the
use (recently banned in the European Union) of zinc bacitracin
as a agricultural growth promoter! Perhaps the contradictions
are best exemplified by the World Health Organization, which
argues on the one hand for effective, affordable antibacterials
for all the worlds population and on the other hand expresses
concern about the worldwide accumulation of resistance
(World Health Organization Report on Infectious Diseases
2000, http://www.who.int/infectious-disease-report/2000/
index.html). Both concerns are ethical, humane, and
honorablebut counterpoised.

CONCLUSIONS
Antibacterial resistance is complex and dynamic. Although the
major genetic and biochemical mechanisms have long been
recognized, new factors continue to be discovered, including
integrons, multidrug efflux, hypermutability, and plasmid addiction. Within many individual isolates, the complexity of
resistance is increasing, with multiple determinants carried, and
with genes being gained, amplified, and lost. Many international
resistance problems reflect the spread of a few multidrug-resistant strains, but the reasons underlying the success of particular
lineages remain almost universally obscure. Resistance is a significant cause of excess morbidity, mortality, and cost. Numerous reports have emphasized the need for less and better
use of antibacterials, improved infection control, and the development of new agents. However, reductions in antibacterial
use do not always lead to reduced resistance, perhaps because
bacteria are now well adapted to the carriage of resistance. It
is probably naive to anticipate reaching a grand control over
resistance, and attempts should center on management rather
than elimination, with the objective of slowing the development
of new resistance while continuing to develop new agents at a
sufficient rate to keep ahead of the bacteria.

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to imipenem was followed by the emergence of imipenemresistant Acinetobacter and P. aeruginosa as well as by a decline
in cephalosporin-resistant klebsiellae [96, 100]. Among opportunistic infections in the seriously ill, the cynic can argue
that resistance is like a balloon: squeeze it on one side, and it
bulges on the other.
It is salutary to emphasize how much remains unknown.
Does drug cycling have positive effects, or does it lead to the
accumulation of multidrug-resistant strains [101]? At what
prevalence of resistance should empirical therapy be changed
in different types of infection? To what extent does combination
therapy militate against resistance (except in the case of tuberculosis, where its value is beyond dispute)? What are the
relative selectivities of different antibacterials, allowing that a
recent Finnish study found a correlation between macrolide use
and resistance in S. pneumoniae but not between penicillin use
and resistance [102]? Is it in any way desirable to encourage
all community physicians to use the same therapies in the same
indications, or is it wiser to make the selection pressure more
diffuse [103]? What is the ideal duration of therapy, allowing
that underdosing may fail to eliminate the least susceptible
members of the original population and that excessive duration
may exacerbate disruption of the normal flora? Are drugs with
unlinked resistances (e.g., fosfomycin, nitrofurantoin, rifampin,
and fucidin) to be favored as unlikely to select multiresistance
plasmids or avoided because they may select hypermutable
strains primed to develop further resistances? The new respiratory quinolones exemplify a further conundrum. As a result
of greater antipneumococcal activity, they are less likely to than
ciprofloxacin to select first-step quinolone-resistant mutants of
S. pneumoniae; however, they are less active than ciprofloxacin
against Enterobacteriaceae and may be selective for resistance
in the gut microflora [104], which is gratuitously exposed.

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