Accepted Manuscript

Title: Do orally administered antibiotics reach concentrations

in the middle ear sufficient to eradicate planktonic and biofilm
bacteria? A review
Author: Miss Katherine Belfield Roger Bayston J.P. Birchall
Matija Daniel
PII:
DOI:
Reference:

S0165-5876(15)00019-1
http://dx.doi.org/doi:10.1016/j.ijporl.2015.01.003
PEDOT 7427

To appear in:

International Journal of Pediatric Otorhinolaryngology

Received date:
Revised date:
Accepted date:

5-9-2014
7-1-2015
8-1-2015

Please cite this article as: M.K. Belfield, R. Bayston, J.P. Birchall, M. Daniel, Do
orally administered antibiotics reach concentrations in the middle ear sufficient to
eradicate planktonic and biofilm bacteria? A review, International Journal of Pediatric
Otorhinolaryngology (2015), http://dx.doi.org/10.1016/j.ijporl.2015.01.003
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Do orally administered antibiotics reach concentrations in the middle ear

sufficient to eradicate planktonic and biofilm bacteria? A review

Authors
(Corresponding author)

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Miss Katherine Belfielda

Tel : (+44) 0115 823 1113

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Email: katherine.belfield@nottingham.ac.uk

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Professor Roger Baystona

Tel: (+44) 0115 823 1115

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Email: roger.bayston@nottingham.ac.uk

Tel: (+44) 0115 924 9924 x65224

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Mr. Matija Daniel a,b,c

Email: Wendy.Phillips@nuh.nhs.uk

Professor JP Birchall b,c

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Tel: (+44) 0115 924 9924 x61224

Email: Matija.Daniel@nottingham.ac.uk

Affiliations

a. Biomaterials-Related Infection Group

School of Medicine

University of Nottingham
Queens Medical Centre
Derby Rd.
NG7 2UH
UK

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b. NIHR Nottingham Hearing Biomedical Research Unit

Nottingham University Hospitals
Ropewalk House
113 The Ropewalk

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Nottingham
NG1 5DU

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UK

c. Otology and Hearing Group

Division of Clinical Neuroscience

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University of Nottingham
Queens Medical Centre

Derby Rd.
NG7 2UH

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UK

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Abstract
Introduction: Infectious conditions of the middle ear are a common and significant cause of
morbidity and mortality worldwide. Systemic antibiotics are frequently used, but their
effectiveness will depend on whether an adequate antibiotic concentration is achieved in the

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middle ear; this is especially important in biofilm infections such as Otitis Media with Effusion
(OME), where high antibiotic concentrations are typically required for effective treatment.
This review examines what antibiotic levels can be reached in the middle ear with oral

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administration, as a means of guiding rational antibiotic choice in the clinic and future

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research, and to determine whether levels high enough for biofilm eradication are reached.

Methods: A literature search of studies measuring levels of antibiotics in the plasma and in

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the middle ear after oral administration was conducted. These levels were compared to the
minimum inhibitory concentrations (MIC) provided by the European Committee for
Antimicrobial Susceptibility Testing (EUCAST) to determine if antibiotic doses were reaching

sufficient levels to inhibit planktonic bacteria. The middle ear concentrations were then
calculated as a multiple of the MIC to determine if the concentrations were reaching biofilm

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eradication concentrations (typically up to 1000 xMIC).

Results: The highest antibiotic levels against Staphylococcus aureus reach 8.3 xMIC,

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against Moraxella catarrhalis 33.2 x MIC, against Haemophilus influenzae 31.2 xMIC, and
against S. pneumoniae 46.2 xMIC. The macrolide antibiotics reach higher levels in the
middle ear than in plasma.

Conclusions: Orally administered antibiotics reach levels above the MIC in the middle ear.
However, they do not reach levels that would be likely to eradicate biofilms.

Keywords:
Otitis media
Ear
Antibiotic

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Biofilm
MIC

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1.Introduction
Infections of the middle ear such as acute otitis media (AOM) and otitis media with effusion
(OME) are a significant cause of morbidity and mortality worldwide. Successful treatment

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with systemic antibiotics relies on being able to achieve a sufficiently high concentration in

the middle ear fluid to have the desired antimicrobial effect. This is important in the case of

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free-floating planktonic bacteria, and even more so when one is trying to deal with a biofilm
infection such as in OME. A variety of antibiotics are used to treat middle ear infections. This

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article reviews the antibiotic levels that can be achieved in the middle ear with systemic
administration, summarising the various published articles, with the aim of guiding rational
antibiotic use especially when biofilm infections are the target. In addition, this information

would be useful for researchers contemplating future studies of antibiotic use in middle ear
infectious conditions.

AOM and OME are related conditions [1], with AOM being an acute infection (viral or

bacterial) with signs and symptoms of inflammation [2], whereas OME is defined by the

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presence of an effusion with no signs or symptoms of acute inflammation [3]. At least 5085% of children will have one episode of AOM by three years of age [1], and the treatment of

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OME with ventilation tubes is one of the commonest surgical procedures in children [4].

1.1 Antibiotics for OME

The aetiology of OME is multifactorial. The main organisms responsible include

Streptococcus pneumoniae, Moraxella catarrhalis, and Haemophilus influenzae. Other
organisms such as Staphylococcus aureus, streptococci and gram-negative rod bacteria
have also been implicated [5-8]. Viruses such as respiratory syncytial virus may contribute to
pathogenesis, but are especially important in AOM [9]. Additional aetiological factors include
genetic and environmental factors, and Eustachian tube function [10].
Antibiotics are often prescribed for AOM (with clear guidelines in place), and are effective.
However, much research has examined the use of antibiotics in OME, but whilst they may
be effective in the short term, antibiotics do not appear to affect the long-term outcome of
OME [11-13]. This may be because bacterial biofilms are important in the aetiology of OME

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[14,15]. Biofilms are functional communities of microorganisms attached to a surface (such

as mucus or middle ear mucosa) and are surrounded by an exopolysaccharide matrix [16].
One of the defining features of a biofilm is that antibiotic levels 10-1000 higher than the
levels sufficient to eradicate free planktonic bacteria are required [17,18]. This is termed

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recalcitrance and is influenced by a variety of factors (Figure 1).

Topical antibiotic therapy is often the preferred route of administration when the tympanic
membrane is perforated or infection is in the ear canal, as it achieves a localised high

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antibiotic level with reduced systemic exposure. However, in OME the tympanic membrane
is intact, so if antibiotics are required then the route of delivery has to be systemic. This

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article discusses antibiotic levels that can be achieved with oral administration, as this route
is simplest, most convenient, and most acceptable to patients. Furthermore, there is no
advantage to using an antibiotic intravenously if it is available orally and has high

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bioavailability [19]. Therefore, although intravenous administration of some antibiotics may

achieve higher levels, the drawbacks of intravenous administration led us to focus on the

oral route.

Antibiotics will be effective only if they actually reach sufficiently high levels in the middle ear.
Inhibition of planktonic bacteria requires levels above the minimum inhibitory concentration

(MIC), which by definition is the minimum concentration of an antimicrobial required to inhibit

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growth of an isolate in planktonic culture. Simply, MIC is the lowest concentration of

antimicrobial that will inhibit visible growth of the bacterial isolate after 24 hours incubation

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on an agar plate in the microbiology laboratory. The MIC of an organism will either be below
or above a breakpoint, which correlates with sensitivity or resistance, respectively. MICs of
individual isolates of a particular species can be determined in the laboratory setting, but
several organisations have set species-specific MICs levels that divide resistant from
susceptible isolates; these levels are the MIC breakpoints.
Achieving levels above the MIC does not guarantee clearance of all bacteria, however.
Dagan et al, demonstrated that even though azithromycin reached levels above the MIC in
the middle ear for H. influenzae and S. pneumoniae, this did not correspond with
bacteriological cure [20]. In the case of a biofilm being present, achieving the MIC at the
target site may not be sufficient for eradicating biofilm. The minimum biofilm - eradicating
concentration (MBEC) is typically up to 1000 times higher than MIC. Importantly, antibiotic
levels below MBEC (in the case of biofilm infections) not only fail to eradicate biofilms but
can promote bacterial resistance. Therefore, this article aims to review articles in the
literature that have measured antibiotic levels in the plasma and in the middle ear to

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determine if the antibiotics prescribed for middle ear infections reach sufficient levels to
eradicate planktonic and biofilm bacteria.
2. Defining Minimum Inhibitory Concentrations of Otitis Media Pathogens

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2.1 Standardizing MIC values

Standardized MIC values (breakpoints) for different organisms have been developed by
several bodies worldwide, including the Clinical Laboratory Standards Institute (USA),

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European Committee on Antimicrobial Susceptibility Testing (EUCAST), and British Society

for Antimicrobial Chemotherapy (BSAC; United Kingdom). The breakpoints aim to correlate

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MIC with clinical outcome [21] by categorizing isolates as clinically susceptible, intermediate,
or resistant. The clinical breakpoints, which separate MIC values that indicate sensitivity and
resistance, distinguish between infections that are likely to respond (sensitive) to antibiotics

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and those that may not (resistant) [22]. MIC breakpoints are influenced by the
pharmacodynamics of the drug, the distribution of susceptibility of the microorganism (ie
bacterial populations that acquire resistance mechanisms or are innately resistant), and

clinical outcomes (ie how often does an isolate determined as susceptible, respond to
standard treatment?) [23]. Likewise, reaching antibiotic levels under the MIC, meaning levels
that will not inhibit sensitive bacteria, can lead to the development of resistance to the

antimicrobial [24] and even induction of the biofilm phenotype [25]. Therefore, achieving

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levels above the MIC is important for planktonic bacteria because this will most likely result
in a positive clinical outcome, will help prevent the development of resistance, and may

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prevent development of the biofilm phenotype.

2.2 Do concentrations cited in the literature reach levels above the MIC in the middle ear?
Table 1 summarises the concentrations of antibiotics in the serum and the middle ear that
can be achieved with oral administration.
With the exception of azithromycin, the concentrations of antibiotics reach their respective
MICs in the serum for S. pneumoniae, H. influenzae, M. catarrhalis, and S. aureus.
Azithromycin reached a serum concentration of 0.07 +/- 0.08 g/mL 24-48 hours after the
last dose was administered, which is below the MIC for all four organisms. The
pharmacokinetic profile of azithromycin demonstrates that the antibiotic is more
concentrated in tissue, particularly polymorphonuclear cells, than in the serum [33], hence
azithromycin levels do reach above the MIC for all four organisms in the middle ear.
Clarithromycin levels were higher in the middle ear than in the serum.

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Some of the antibiotics reach levels above 10X MIC such as azithromycin, ciprofloxacin, and
clarithromycin against S. pneumoniae, and cefaclor, ciprofloxacin and clarithryomycin
against M. catarrhalis. However, none of the antibiotics reached concentrations 10X MIC in
the middle ear against S. aureus and only one (ciprofloxacin) reached above 10X MIC
against H. influenzae. The highest antibiotic levels against S. aureus reach 8.3 xMIC,

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against M. catarrhalis 33.2 x MIC, against H. influenzae 31.2 xMIC, and against S.
pneumoniae 46.2 xMIC. Importantly, antibiotic levels in the middle ear with oral

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administration do not reach 100 xMIC for any antibiotic/organism.

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2.3 Do antibiotics reach levels that will eradicate biofilm in the middle ear?

The MBEC which is the minimum concentration of antibiotics needed to kill biofilm, is
influenced by many factors such as the antibiotic and the bacterial isolate. 10X MIC may be

sufficient to inhibit some biofilms whereas others are sensitive only to levels above 100X
MIC and 1000XMIC [34,35]. For example, in strains of nontypeable H. influenzae isolated
from the middle ear of patients with acute otitis media the MBEC for amoxicillin was greater

than 1,024 g/mL as compared to the MIC of 2.0 g/mL as reported by BSAC and EUCAST.
The concentration of amoxicillin needed to eradicate biofilm was 512X MIC, which has not

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been achieved in the middle ear with amoxicillin or any other antibiotic. However, in the
same study, the MBEC for clarithromycin was 0.25-1.0 g/mL which is equivalent to the

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EUCAST MIC of 1.0 [36]. In this example, clarithromycin reached levels in the middle ear
greater than the MIC and MBEC for nontypeable H. influenzae. The MBEC for amoxicillin
against M. catarrhalis in an in vitro model was greater than 64 g/mL compared to the
laboratory - determined MIC of 0.06 g/mL, which means approximately 100X MIC
amoxicillin is needed to eradicate M. catarrhalis biofilm [37].

It is also important to note that non-typeable H. influenzae and M. catarrhalis frequently

produce beta-lactamase enzymes and are thus resistant to amoxicillin and cefaclor [38,39].
The above levels are given for amoxicillin-susceptible isolates and these would differ greatly
from the isolates that are beta-lactamase producing. Likewise, the European Centre for
Disease Control and Prevention reports 8.8% of S. pneumoniae isolates from 27 European
countries as non-susceptible to penicillin [40], meaning eradication levels with beta-lactams
would also vary drastically.

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MBEC values for methicillin-resistant S. aureus were found to be 1280-2560 mg/L for
ciprofloxacin and between 1280-5120 mg/L for azithromycin in a recent study by Dosler and
Mataraci [41]. These MBEC values are 1280-2560X MIC of ciprofloxacin and 1280-5120X

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MIC of azithromycin, highlighting the importance of high levels reaching the site of biofilm
infection. Methicillin-sensitive S. aureus strains are comparable; the MBEC of clindamycin

for MSSA was greater than 512 g/mL [42] which is equivalent to 2048X MIC as reported by

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BSAC and EUCAST. The ciprofloxacin and azithromycin levels in the middle ear provided
here reach only 5.5X MIC and 3.9X MIC respectively: nowhere near these MBECs.

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Of course the antibiotic levels needed for biofilm eradication in the middle ear might be so
high that human toxicity would occur if one tries to achieve that level using systemic

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antibiotic administration. For example, S. pneumoniae, the most sensitive of the four
organisms discussed here, would require a local concentration of at least 12.5mg/L of
ciprofloxacin to achieve 100X MIC in vitro. However, the expected peak plasma level of a

normal dose ciprofloxacin (250mg twice daily) after oral administration is only 0.94 mg/L [43],
suggesting that attempts to eradicate biofilms using higher systemic doses might lead to
toxicity, because the 100X MIC level is 13.3 times greater than the plasma level achievable

with a normal oral dose of ciprofloxacin. Further, the MBECs for ciprofloxacin reported above
for MRSA and MSSA far exceed the higher toxic doses reported in man. This suggests that

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the MBEC may not be achievable using systemic administration without toxicity.

2.4 Factors influencing antibiotic concentrations

It is also important to take into account variables that may influence serum or middle ear
concentrations such as time since last dose, dose, constituents of the middle ear fluid and in
the case of detection, method used to quantify antibiotic concentrations. Here we cite a
cefaclor concentration of 11.2 g/mL in serum and 2.3 g/mL in the middle ear. This study
by Krause et al [26], provides higher cefaclor concentrations in the serum and middle ear
than those produced by Lildholdt et al, despite giving the same dose of cefaclor to subjects.
The Lildholdt et al study demonstrated 8.49 +/-7.79 g/mL in serum and 0.47 +/- 0.78 g/mL
in the middle ear [44]. Krause et al cites differences in methods in which the quantification
assay used by Lildholdt et al exposed the samples to greater levels of degradation [26].
While the dosing was the same by the previous two studies, the studies by Klutymans et al
and Keusch and Present gave their patients either a single dose of 150 mg of clindamycin
[31] or a single dose of 450 mg of clindamycin [45] respectively, resulting in different serum

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concentrations. Keusch and Present further find that when clindamycin is given orally, serum
concentration is directly related to oral dose; ie a serum level of approximately 3 g/mL
could be predicted if the oral dose were 300 mg of clindamycin, with repeated dosing

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increasing serum levels [45].

Furthermore, the middle ear fluid contains mucins, inflammatory cells, and other cell

constituents whose concentrations may vary between healthy patients and those with OME

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[46]. Therefore, the amount of drug available to bacteria is likely to vary, such as the

macrolide antibiotics being mainly found intracellularly in tissue and inflammatory cells [47].

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The levels in the middle ear fluid may not equate with levels that are actually able to
penetrate though the biofilm, as the biofilm may contain barriers to antibiotic diffusion.

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Finally, time since last dose influences serum and middle ear concentrations, based on
excretion of the drug and metabolism. For example, the co-amoxyclav concentration in the
middle ear decreased from AMX/CLV: 7.4/0.94 when sampled two hours since last dose to

AMX/CLV: 6.01/0.11 eight hours since last dose [32].

Thus, the middle ear concentrations of antibiotics achieved with systemic administration are
in general below the MBEC. This may explain why antibiotics fail to cure clinical biofilm

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infections such as OME. Much research has been done to investigate oral antibiotics against
OME, with a recent Cochrane review advising against their use [12]. The data presented

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here provide a sound theoretical basis to explain why oral antibiotics would not work against
OME. Local delivery of antibiotics would be a better alternative, but in OME this may require
a novel delivery strategy such as modified-release antibiotic formulations inserted at
myringotomy [17,48].

3. Conclusions

Understanding what antibiotic concentrations can be reached in the middle ear with oral
administration is essential when choosing an appropriate antibiotic, particularly in the case of
biofilm infections such as OME. The majority of antibiotic oral dosing regimens are reaching
middle ear concentrations greater than the MIC for S. pneumoniae, M. catarrhalis, H.
infleunzae, and S. aureus. However, none of these antibiotics reach levels that approach
100 xMIC, and this may explain why oral antibiotics fail to cure OME.

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Prevention and Control; 2012.
<http://www.ecdc.europa.eu/en/publications/antimicrobial-resistance-surveillanceeurope-2011.pdf. Accessed December 12, 2014.

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in combination with antibiotics against methicillin resistant Staphylococcus aureus
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43. W. Wingender, K.H. Graefe, W. Gau. D. Frster, D. Beermann, P. Schacht.

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delivery to the ear. Ther. Deliv. 4 (2013) 115-124

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ip
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cr
us

Ac
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p

te

an

Figure 1: Factors influencing biofilm recalcitrance. The main factor in recalcitrance is now
considered to be the phenotypic modification of bacterial metabolism resulting from enforced
slow replication. Energy generation and transport inside the bacteria is critically low, and
non-essential functions are down regulated. In this dormant state, bacterial targets for
common groups of antibiotics are no longer active, and anti cell wall synthesis drugs
(betalactams, vancomycin), anti-protein synthesis drugs (aminoglycosides, macrolides,
tetracyclines etc) and anti-DNA replication agents (quinolones) are orders of magnitude less
effective. Matrix refers to the exopolysaccharide matrix of the biofilm

Page 14 of 15

Ciprofloxacin
Clarithromycin
Clindamycin
Co-Amoxiclav

TrimethoprimSulfamethoxazole

0.07 +/- 0.08

11.2 +/- 3.6

5.6 +/- 0.4 26

20

25

7.27 +/- 0.5 27

3.96 +/- 1.19

MIC

xMIC

n/a

n/a

3.9+/- 3.1

0.25

15.6

n/a

n/a

0.125

2.3 +/-1.3

25

0.32

7.2

0.125

18.5

0.5

0.125

44.3

0.5

11.1

0.5

8.30 +/- 2.48

AMX:7.2

AMX:7.4

CLV: 2.032

CLV: 0.94 32

TMP:1.8 +/- 1.2

TMP: 1.0 +/-

17.2

MIC

xMIC
2.8

N/A

+/- 0.5

26

MIC
2

4.5 31

SFX:38.2

H. influenza

20

5.54 +/- 3.46 28

29

M. catarrhalis

0.25
0.5

SFX: 6.0 +/-

33.2

0.25

33.2

n/a

n/a

n/a

n/a

n/a

n/a

TMP: 1
SFX: 6

AMX:7.4
CLV:0.94
TMP: 2
SFX: 12

0.5

2.9 26

30

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Cefaclor

13.6 +/- 7.3 26

S. pneumoniae

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Azithromycin

ME level

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Amoxicillin

Serum level

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Antimicrobial

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Table 1: Common antimicrobials and the maximum levels achieved in serum and middle ear
with oral administration (Note: Only the highest concentrations in serum and middle ear were
reported here despite there being other studies with lower concentrations) The MIC (g/mL),
as determined by the European Committee for Antimicrobial Susceptibility Testing
(EUCAST) for common middle ear pathogens is given. The xMIC columns give the highest
middle ear antibiotic levels achieved with oral administration, described as a multiple of MIC.
ME Level: Concentration of antimicrobials in the middle ear, AMX: Amoxicillin, CLV:
Clavulanic acid, TMP: Trimethoprim, SFX: Sulfamethoxazole

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