CLINICAL ORTHOPAEDICS AND RELATED RESEARCH
Number 437, pp. 7–11
SESSION I: BIOFILMS IN
ORTHOPAEDIC INFECTIONS
Biofilm Theory Can Guide the Treatment of
Device-Related Orthopaedic Infections
J. William Costerton, PhD
Direct observations of the surfaces of orthopaedic prostheses
that have failed and of bone affected by osteomyelitis with
and without the presence of a prosthesis have shown that the
bacteria that cause these infections live in well-developed
biofilms. The cells within these matrix-enclosed surface-
associated communities are protected from host defenses and
antibiotics, and clinical experience has shown that they must
be removed physically before the infection can be resolved.
The biofilm etiology of these diseases demands new diagnos-
tic methods because biofilm cells typically do not grow on
agar plates when recovered by scraping or swabbing. I will
recommend new molecular and immunologic diagnostic
methods that have been useful in other biofilm infections.
These diseases progress through quiescent periods that al-
ternate with acute exacerbations, and clinicians must realize
that antibiotic therapy can control the acute phases but can-
not resolve the basic biofilm nidus of the infection. Now that
it has been realized that these orthopaedic infections are
caused by relatively common biofilm-forming bacterial
pathogens, new technologies that deliver very high concen-
trations of antibiotics locally and “on demand” and novel
molecular “mimics” that block the signals that control bio-
film formation need to be examined.
The powerful tools of modern microbial ecology and of
modern molecular microbiology have produced one rev-
elation after another when they have been applied to the
study of bacteria in natural and pathogenic ecosystems.
Bacteria have been found to grow predominantly in bio-
films.5 These sessile communities have been shown to be
From the University of Southern California, Center for Biofilms, School of
Dentistry, Los Angeles, CA.
The author certifies that he has no commercial associations (eg, consultan-
cies, stock ownership, equity interest, patent/licensing arrangements etc) that
might pose a conflict of interest in connection with the submitted article.
Correspondence to: J. William Costerton, PhD, University of Southern Cali-
fornia, Center for Biofilms, School of Dentistry, 925 West 34th St., Los
Angeles, CA 90089. Phone: 213-740-2538; Fax: 213-740-5715; E-mail:
costerto@usc.edu
DOI: 10.1097/01.blo.0000175128.44966.d9
7
© 2005 Lippincott Williams & Wilkins
highly structured,6 biofilm cells have been shown to adopt
a different phenotype,18 and the component cells of bio-
films have been shown to communicate by means of cell–
cell signals.7 Concepts and methods dating from the mid-
1850s, such as Koch’s postulates and bacterial enumera-
tion by recovery and plating, have now been superseded
by new concepts and methods that have proven to be es-
pecially useful in our understanding of chronic bacterial
infections. This radical transformation in microbiology
now is embraced universally by researchers in natural and
industrial environments, where direct microscopy has
shown bacteria to live predominantly in mixed-species
biofilm communities adherent to surfaces.20 Because these
microbial ecologists have perceived that individual float-
ing (planktonic) bacteria are rare in real ecosystems, they
have abandoned the recovery and culture of these cells in
favor of new methods that allow the examination of real
communities in situ. Bacterial cells in natural biofilm com-
munities grow poorly (if at all) when recovered from their
ecosystem and spread on agar plates,22 and the majority of
bacterial species in natural environments cannot be cul-
tured in any known medium. Bacterial cells from patho-
genic ecosystems grow more readily in media developed
for this specific purpose, but these in-vitro–grown organ-
isms differ from biofilms in the infected host in many
salient characteristics, including their susceptibility to an-
tibiotics.17
It is my objective to capture the new concepts and
methods of biofilm microbiology and to apply them di-
rectly and unequivocally to the diagnosis and treatment of
bacterial infections of bone and of orthopaedic prostheses.
Device-Related Infections and Other Chronic
Bacterial Infections
Very soon after it was discovered that chronic bacterial
infections associated with cardiac pacemakers15 and or-

8 Costerton
thopaedic devices are caused by bacteria growing in bio-
films, it was shown13,16 that osteomyelitis is caused by the
same matrix-enclosed bacterial communities. Bacterial
biofilms were seen, by direct microscopic techniques, on
orthopaedic prostheses that had been removed from pa-
tients with overt device-related infections.9 The recalci-
trance of osteomyelitis and of prosthesis-related infections
to antibiotic therapy was rationalized by authors of studies
that showed that cells within bioiflms are resistant to an-
tibiotic level 1000 times higher than those that will kill
their planktonic counterparts.17 The chronicity of osteo-
myelitis and of prosthesis-related infections was explained
by our finding that antibodies fail to penetrate biofilms and
kill the component organisms,12 and that even activated
phagocytes cannot kill bacteria in biofilms.14 The role
played by bacterial biofilms was summarized in a 1999
review,6 and osteomyelitis and infections of orthopaedic
prostheses were listed as examples of these chronic infec-
tions that now constitute 65% of bacterial and fungal in-
fections treated by physicians in the industrial world. The
pivotal concept in bone and joint infections is that bacteria
that are present in the surgical field form biofilms on
the surfaces of dead bone and of metal and plastic pros-
theses. Cells within these sessile communities go through
a phenotypic change that makes them resistant to conven-
tional antibiotics and that they adopt a matrix-protected
mode of growth that renders them almost immune to
attack by antibodies or by phagocytes. It is suggested
that much of the tissue damage in chronic biofilm infec-
tions is caused by “frustrated phagocytosis,” and it is
known now that inflammation in response to the presence
of biofilms is a major cause of tissue damage in chronic
infections.6
The gold standards in studies of human pathogenesis
are clinical observations and patient data. The notion that
biofilms are involved in the etiology of both osteomyelitis
and of prosthesis-related infections is supported by the
observations that these infections often develop months or
years after damage or surgical intervention, and that the
infections remain localized and often develop relatively
slowly. Antibiotics are effective in the relief of acute
symptoms, caused by the release of planktonic bacteria
from biofilms, but this therapy is notably ineffective in full
resolution of the infection, because it cannot kill all of the
bacteria within the biofilms.11 Clinical wisdom dictates
that inert materials and dead bone must be removed com-
pletely to allow the resolution of these infections, and we
see this as the physical removal of the causative biofilms.
Clinical wisdom also dictates that surgical revisions must
be covered by high doses of antibiotics, and this is seen as
providing enough of the agents to kill the large number of
planktonic bacteria that will inevitably be present in sites
from which large volumes of biofilm have been removed.
Clinical Orthopaedics
and Related Research
Chronic infections of bones and of orthopaedic prostheses
persist in spite of vigorous humoral and cellular immune
responses and these infections are now seen to be quint-
essential biofilm infections.6
Diagnosis of Orthopaedic Biofilm Infections
The diagnosis of biofilm infections is always complicated
by the fact that matrix-enclosed sessile bacteria elicit much
less inflammatory response than would be occasioned by
the present of a corresponding number of planktonic bac-
terial cells. This response typically is local, so a general
malaise is rare, but local changes in bone function may be
very revealing. We examined 10 cases of aseptic loosening
of acetabular cups as a subset of a very large number of
similar cases that never have yielded positive bacterial
cultures from either aspirates or from removed devices,
and eight of these 10 cases showed the presence of bac-
terial biofilms on the devices. This disturbing revelation
may mean that large numbers of low-grade biofilm infec-
tions currently are misdiagnosed as aseptic loosening and
are revised without the antibacterial precautions that
would be used if they were correctly attributed. Until a
prospective controlled study can be done, it may be pru-
dent to treat all prosthetic loosening as being potentially
infected.
Because of published evidence that cells of Staphylo-
coccus aureus growing in biofilms in the human vagina
cannot be detected by swab recovery and plating on ap-
propriate agar media,22 now it is obvious that we cannot
rely on cultures of aspirates to diagnose infections of or-
thopaedic prostheses. Because of this lack of sensitivity of
established microbiologic methods, molecular methods
[eg, polymerase chain reaction (PCR) and fluorescence in
situ hybridization (FISH)] are more suitable for the detec-
tion of biofilm infections. The humoral and cellular im-
mune systems are also being used to detect the presence of
developing biofilms in patients who have received various
kinds of prostheses.19 The humoral system reacts to the
presence of immunogenic epitopes on the surfaces of bac-
teria by producing specific antibodies, and biofilm bacteria
produce surface proteins that are very distinct from those
on the surfaces of planktonic cells of the same species.18
Selan et al19 have identified a surface epitope (SSPA) that
is produced by biofilm cells of S. aureus and S. epidermi-
dis, and they have developed an enzyme-linked immuno-
sorbent assay (ELISA) test to detect antibodies to this
epitope in patients who have received vascular grafts. This
ELISA test is very sensitive, and very specific, in the
detection of staphylococcal biofilms in these patients (p <
0.001) and the surgical revision of infected vascular grafts
has saved hundreds of lives. I see no obstacles in the
application of this same test to the detection of nascent
biofilm formation in patients who have received orthopae-
Number 437
August 2005
dic prostheses, and we think that the ELISA test may be
useful in guiding informed clinical decisions. The pres-
ence of bacterial biofilms in any bone or joint also will be
expected to trigger specific responses in other immune
cells, like the surface determinants of phagocytes in the
synovial space. It is encouraging that several potential mo-
lecular and immunologic tests already are available to re-
place bacterial cultures in the diagnosis of biofilm infec-
tions of bones and joints, and we are especially enthusi-
astic about technologies that can be adapted to imaging so
that these bacterial communities can be located with some
degree of accuracy. To be specific, anti-biofilm antibodies
can be tagged with specific “opacity markers” for many
types of scans, and these antibodies will react with their
specific antigens on biofilm cells to yield an image of the
bacterial communities, many of which attain macroscopic
proportions. A rise in the ELISA titer would designate an
orthopaedic patient for workup regarding infection, and
antibody-based imaging could locate the problem and
guide clinical treatment and revision.
Prevention of Colonization and Biofilm Formation
The biofilm concept has been accepted throughout natural
and industrial ecosystems, so that engineers who treat mi-
crobially influenced corrosion (MIC) on the basis of bac-
terial counts in the bulk fluid (instead of on the pipe sur-
face) now are in danger of lawsuits against them. The
general biofilm community has made great strides in the
detection of biofilms and, once a problem or disease is
recognized as being caused by biofilms, several companies
and large numbers of engineers and scientists swing into
action. Biofilms are detected by new technologies that
range from auto-fluorescence of co-enzymes to PCR, doz-
ens of coatings are nominated for use in biofilm preven-
tion, and dozens of antibiofilm agents are suggested to kill
sessile bacterial cells in these protected sessile communi-
ties.
The newest concern is that any plastic or metal bioma-
terial that is placed into the body must be perfectly clean.
Research in the water industry has shown that surfaces are
very similar in their tendencies to attract planktonic cells
and thus accrete biofilms,5 but that the contamination of
surfaces by organic materials (especially residual biofilm
matrices) accelerates this process at least tenfold. Because,
as Gristina et al10 said with haunting accuracy, the bacteria
and the tissues are engaged in a “race for the surface,” the
presence of any residual biofilm matrix on a biomaterial
surface favors colonization and infection. Orthopaedic
prostheses are manufactured by machining techniques that
use fluids that favor biofilm development to an extent that
sometimes leads to the physical plugging of macroscopic
apertures in the machine tool assembly. Simple steriliza-
tion (eg, ethylene oxide) kills the bacteria in these bio-
Biofilms in Orthopaedic Infections 9
films, but fails to remove the residue of their matrices, and
these invidious deposits must be removed before the de-
vices are implanted. Several enzyme treatments and plas-
mas are available for the removal of biofilm residues from
the surfaces of even the most delicate biomaterials.
One of the most practical strategies for the prevention
of colonization and consequent biofilm formation on bio-
materials is the use of materials and coatings that release
conventional antibiotics into the surrounding tissues and
fluids. Ideally, these materials will release antibiotics in
concentrations that kill any planktonic cells in the area of
the implant before they can initiate biofilm formation and
infection. The downside of this basic strategy is that re-
lease curves are typically abrupt at the outset, and “tail
off” after days or months, so that subinhibitory concentra-
tions may favor the development of resistant strains of
bacteria, and any survivors are provided with an inert and
attractive surface. This problem may have been solved by
researchers at the University of Washington who have
developed plastics with a “skin” that seals its surface and
limits the release of antibiotics and other organic mol-
ecules until it is made permeable by ultrasonic energy.
After an “on-demand” release of antibiotics, which can be
regulated by the duration of the ultrasonic pulse, the skin
reseals and the agent is conserved until the next pro-
grammed release. This controlled release coating may be a
useful addition to the current strategy of placing antibiotic-
loaded beads into the sites of orthopaedic infections, in
that even higher local concentrations of antibiotics may be
achieved.
The discovery7 that the development of microbial bio-
films is controlled by cell-cell signaling has opened a
whole new approach to the prevention of the colonization
of prostheses and to the progression to chronic device-
related infections. The signals that exercise this control are
simple acyl homoserine lactones (AHLs), in the case of
gram-negative bacteria,8 and gram-positive bacteria use
equally simple cyclic octapeptides for the same purpose.1
Soon after these and other signals were discovered, several
companies were formed to exploit the fact that biofilm
formation can be blocked by natural and synthetic mol-
ecules that mimic these signals, react with the cognitive
signal receptor proteins, and prevent biofilm formation.
Balaban et al2 presented very persuasive evidence that
their RIP inhibitor of the RAP signal that controls biofilm
formation in staphylococcal species, prevents biofilm for-
mation by cells of S. aureus. This inhibition of biofilm
formation was shown in elegant animal models of device-
related infection2 and the inhibitor was shown to be espe-
cially effective in infection control if it was combined with
an antibiotic (mupurocin). The fertile minds of orthopae-
dic practitioners soon will combine the dissolving bead
delivery system, the ultrasonic control of release, inhibi-

10 Costerton
tors of biofilm formation signals, and new combinations of
antibiotics, with the concept that surfaces must be scrupu-
lously clean when the device is implanted. Progress prob-
ably will be incremental, but spectacular success will at-
tend the inventive combination of these technologies, and
patients will be helped.
Rational Treatment of Biofilm Infections
If clinicians who deal with infections in orthopaedic prac-
tice will embrace the biofilm concept, this notion will
provide a rational basis for the prevention, diagnosis, and
treatment of these devastating diseases. The concept sees
the infecting bacteria as planktonic cells that rapidly form
matrix-enclosed biofilms on available surfaces, with some
preference for inert materials and dead tissues, and we can
be comforted by the fact that bacteria behave in this same
way in nature, and in other chronic infections. The slow
development, the ineffective immune response, the trig-
gering of inflammation, the inherent resistance to antibi-
otics, and the profound difficulty of culturing the causative
organisms, are all characteristics of biofilm infections of
other tissues and organ systems.6 The speed with which
biofilms develop on inert surfaces or on compromised tis-
sues (eg, dead bone) depends on the number and kind of
bacterial cells, on the cleanliness of the surfaces, and the
immune state of the patient. However, once the biofilms
have developed and matured, the clinician is faced with an
implacable foe. The low doses achieved by systemic an-
tibiotic therapy may alleviate symptoms caused by plank-
tonic bacteria that leave the biofilms during acute exacer-
bations, but the sessile cells of the biofilm are not affected,
and they continue to damage tissues by stimulating inflam-
matory reactions of the host. Aspirates from joints or from
overtly infected bone will not yield positive cultures, on
routine microbiological examination, but indices of im-
mune response and of phagocyte activation will be useful
in monitoring the progress of the infection.
Because biofilms profoundly are resistant to antibiotics,
and because the deleterious effects of sustained inflamma-
tion are cumulative, good treatment will dictate the early
removal of colonized prosthesis and the aggressive use of
antibiotics, even if the problem seems to be aseptic loos-
ening. The surgical axiom that dead tissue should be com-
pletely removed especially is true in device-related osteo-
myelitis, and the orthopaedic surgeon will be faced with
the unique problem of maintaining space for the implan-
tation of a new prosthesis when the infection has been
resolved. Very large doses of antibiotics can be used in
bones and joints without producing toxic circulating lev-
els, but these agents must be chosen for their efficacy
against biofilm cells (which may differ from their minimal
bactericidal concentration against planktonic cells in the
lab) and for their lack of toxicity to bone. The debridement
Clinical Orthopaedics
and Related Research
that accompanies the revision of an orthopaedic prosthesis
starts a new “race for the surface” of bone tissue, bone
fragments, and the beads used for antibiotic delivery. The
operative field will contain planktonic cells liberated from
biofilms as they were being removed, and some biofilm
cells on bone fragments that have not been removed com-
pletely. The planktonic cells can be killed by moderate
doses of antibiotics, and their natural tendency toward bio-
film formation soon may be thwarted by signal inhibitors,
but the residual biofilms present a special problem. Sessile
cells within mature biofilms can be killed by very high
doses of conventional antibiotics,21 delivery systems in
current use already can produce these high levels, and new
technologies may attain even higher concentrations. We
now are in a position to determine the exact concentrations
of specific antibiotics that will kill biofilm cells of S. au-
reus, in vivo. Infected animals with colonized prostheses
can be treated with antibiotics whose concentration in the
tissues can be determined, and we can assess the viability
of the biofilm bacteria by the use of the new “live/dead”
probe.4 Once the antibiotics have been screened for effi-
cacy and bone toxicity and their effective concentrations
have been determined, modern technologies can be mobi-
lized to deliver effective doses to the biofilms in question.
DISCUSSION
The biofilm etiology of device-related and other chronic
infections of importance in orthopaedic practice now has
been established. Biofilms of orthopaedic pathogens have
been examined in vitro and in animal models, and these
matrix-enclosed communities of bacteria have been shown
to be resistant to antibiotic therapy and to intact host de-
fenses. These observations rationalize our collective clini-
cal experience which teaches us that entrenched biofilm
infections are rarely resolved by antibiotic therapy alone,
but that resolution is often possible if the prosthesis and
any compromised bone are removed by careful debride-
ment.
Now that device-related and other chronic orthopaedic
infections have joined the ranks of the burgeoning number
of biofilm infections of humans, certain earmarks of these
infections are seen to apply. Diagnosis by recovery and
culture is not reliable, because biofilm cells grow poorly if
at all on agar plates, and all instances in which symptoms
are present should be treated as infections until we can
develop molecular and immunologic methods to replace
culture methods. Orthopaedic infections should be seen as
alternating between quiescent periods of biofilm growth
and acute exacerbations caused by the release of plank-
tonic bacteria, and the acute phases should be treated with
antibiotic therapy. Because orthopaedic infections share
many etiologic factors with other biofilm infections, pa-
Number 437
August 2005
tients may be helped by insights that have developed in
other medical specialties. Infections may be controlled by
locally very high concentrations of antibiotics delivered
“on demand” by means of a novel ultrasonic-triggered
release mechanism, and biofilm bacteria may be made
more susceptible to antibiotics by the imposition of weak
direct-current electric fields, or by ultrasonic energy de-
livered at particular wavelengths. Furthermore, like all
biofilm infections, orthopaedic infections may be pre-
vented by the judicious use of specific signal analogs that
block biofilm formation, and lock potential pathogens in
the planktonic mode-of-growth, in which they are suscep-
tible to routine antibiotic therapy.
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