Surgical Neurolog

Infection

Intrathecal baclofen does not inhibit the growth of different bacterial

species and Candida albicans

Alessandro Dario, MDa,4, Andrea Endimiani, MDb, Antonio Toniolo, MDb, Anna Iadini, MDc,

Simone Sangiorgi, MDa, Carlo Scamoni, MDa, Giustino Tomei, MDa

aNeurosurgical Clinic, Ospedale di Circolo e Fondazione Macchi and Insubria University, 21100 Varese, ItalybLaboratory of Microbiology, Ospedale di Circolo e Fondazione Macchi and Insubria University, 21100 Varese, ItalycLaboratory of Pharmacology, Ospedale di Circolo e Fondazione Macchi and Insubria University, 21100 Varese, Italy

Received 23 August 2006; accepted 24 October 2006

www.surgicalneurology-online.com

Abstract Background: The antimicrobial activity of intrathecal baclofen was investigated. Several different

0090-3019/$ – see fro

doi:10.1016/j.surneu.2

Abbreviations: C

acid; MBC, minimu

hibitory concentration

tory Standards.

This study has bee

Society for Stereotact

4 Corresponding

Fondazione Macchi, V

0332 261045.

E-mail address: d

microorganisms were used: Staphylococcus aureus (b-lactamase–positive and b-lactamase–negativestrains); S epidermidis; Enterococcus faecalis; Klebsiella pneumoniae; Escherichia coli; Pseudo-

monas aeruginosa; and Candida albicans.

Methods: Three experimental approaches were used to assess baclofen antimicrobial activity: (1)

determination of the MIC; (2) determination of the MBC; and (3) kinetic time-kill assay.

Experiments were performed according to current methods of the NCCLS.

Results: As compared with control organisms exposed to physiologic saline, organisms exposed to

baclofen over a 10-day period failed to reduce the number of viable cells by at least 3 log10, as

requested by NCCLS criteria.

Conclusions: Because the viability of the investigated organisms was not reduced over that of

microbial suspensions exposed to physiologic saline, we conclude that intrathecal baclofen has no

measurable activity against different bacterial species and C albicans.

D 2007 Elsevier Inc. All rights reserved.

Keywords: Intrathecal baclofen; Antimicrobial activity; Bacterial species

1. Introduction

Baclofen (b-chlorophenyl-GABA) is an agonist ligand

specific for bicuculline-insensitive GABA receptors of the

medulla posterior dorsal horn. It is one of the main drugs

used in the treatment of spasticity. Long-term infusion of

nt matter D 2007 Elsevier Inc. All rights reserved.

006.10.068

FU, colony-forming unit; GABA, g-aminobutyric

m bactericidal concentration; MIC, minimum in-

; NCCLS, National Committee for Clinical Labora-

n presented in part at the 14th Meeting of the World

ic and Functional Neurosurgery.

author. Neurosurgical Clinic, Ospedale di Circolo e

arese 21100, Italy. Tel.: +39 0332 278388; fax: +39

ario.alessandro@virgilio.it (A. Dario).

intrathecal baclofen (ie, the parenteral form of baclofen) is

extensively observed in patients with spasticity unrespon-

sive to treatment with oral drugs. Intrathecal baclofen is

delivered continuously via a totally implantable program-

mable pump and catheter directly into the cerebrospinal

fluid. At present, more than 15000 patients have been

treated with the implantable pump device [5]. Pump

implantations are also performed for the treatment of

chronic or malignant pain [21]: in patients with the latter

type of pain, opiates and/or local anesthetics are infused.

Local anesthetics have long been known to possess

antimicrobial [16,18] and antifungal [13] properties. In

particular, solutions for intraspinal infusion of bupivacaine

and diamorphine are endowed with antibacterial properties

[3]. We report on the first study on the antibacterial and

antifungal activity of intrathecal baclofen.

y 68 (2007) 168–171

A. Dario et al. / Surgical Neurology 68 (2007) 168–171 169

2. Methods

Baclofen dissolved in saline (concentration of 2 mg/mL)

was obtained from Novartis Pharma Stein (Stein, Switzer-

land). The followingmicrobial strains were obtained from the

ATCC (American Type Culture Collection, Promochem,

London, UK): Staphylococcus aureus no. 29213 (b-lacta-mase positive); S aureus no. 25923 (b-lactamase negative);

S epidermidis no. 12228; Enterococcus faecalis no. 25212;

Klebsiella pneumoniae no. 13883; Escherichia coli no.

35218; Pseudomonas aeruginosa no. 27853; and Candida

albicans no. 90028.

Fig. 1. Kinetics of the in vitro time-kill assay of different microorganisms expos

baclofen 2 mg/mL; (n), NaCl 0.85% used as negative control.

Three experimental methods were used: (1) determina-

tion of the MIC; (2) determination of the MBC; and (3)

kinetic time-kill assay. Experiments were performed accord-

ing to methods published by the NCCLS [10].

All tests were carried out in duplicate. Briefly, MIC and

MBC values for bacterial strains were determined with the

use of a microdilution test method using 96-U–well micro-

plates (Falcon, Oxnard, Calif) and cation-adjusted Mueller-

Hinton broth medium (Oxoid, Garbagnate Milanese, Milan,

Italy). RPMI-1640 medium (Sigma-Aldrich, St Louis, Mo)

was used for determining the MIC and MBC values of

C albicans. Serial dilutions of baclofen (2000 to 2 lg/mL)

ed to baclofen. Each point represents the mean of 2 determinations: (E)

,

A. Dario et al. / Surgical Neurology 68 (2007) 168–171170

were prepared in 96-well microplates using 100 lL per well.

Subsequently, each well was seeded with 100 lL of a viable

microbial suspension containing 5 � 105 CFU/mL. Micro-

plates were incubated at 358C for 24 hours (bacterial strains)

or 48 hours (C albicans). The MIC was defined as the

amount of drug required to inhibit the growth or multipli-

cation of an organism isolate [8,19]. The MBC was

determined using the same method described, except that,

at the end of the incubation time, 10-lL aliquots of the first

clear well of each microbial strain were subcultured on

sheep blood agar plates (Oxoid) to determine the number of

vital organisms. Colonies were counted after a 24-hour or a

48-hour incubation. The MBC end point was defined as the

lowest concentration of the drug that reduced the number of

viable organisms by 99.9% [8,19]. Measuring the rate of

bactericidal activity by time-kill analysis provides the

opportunity to assess the speed with which killing may

occur at a given drug concentration [19]. Organisms

in the log growth phase were used to prepare suspensions

of 5 � 106 CFU/mL in saline (0.85% NaCl). Glass

containers were inoculated with 10 mL of saline containing

106 CFU/mL and baclofen at a final concentration of

800 lg/mL. Samples were incubated for 10 days at 358C.Every 24 hours, 100-lL aliquots were removed from each

container. Serial 10-fold dilutions were prepared in Mueller-

Hinton broth and immediately plated on sheep blood agar

plates. Colonies were counted after a 24-hour or a 48-hour

incubation. The number of organisms remaining in each

sample was plotted over time to determine the rate of

microbial killing. The effect of organisms exposed to

baclofen was compared with that of control organisms

containing saline alone. According to NCCLS criteria, a

bactericidal response is obtained when the numbers of

viable organisms are reduced by at least 3 log10 [10]. Graphs

were produced using the Statistica PC software (StatSoft,

Tulsa, Ok).

3. Results

The MIC and MBC values for baclofen were 2 mg/mL or

higher for all the investigated organisms. The reduction rate

of viable cell counts over a 10-day incubation period is

shown in Fig. 1, where each point represents the mean of

2 determinations. Gram-positive cocci showed a nearly

linear reduction of viable cell counts within 6 days of

incubation. No significant difference was detected between

control and baclofen-exposed bacterial suspensions.

K pneumoniae and C albicans showed responses compara-

ble with those of Gram-positive bacteria. Environmental

Gram-negative bacteria (E coli and P aeruginosa) showed a

markedly prolonged decrease of viable cell counts. As

compared with control organisms exposed to physiologic

saline, organisms exposed to baclofen failed to reduce the

number of viable cells by more than 2 log10 over a 10-day

period. On the whole, none of the investigated organisms

showed a reduction of viable counts 3 log10 or greater over

control cell suspensions.

4. Discussion and conclusions

Several nonantibiotic drugs, including antidiuretics,

antidiabetics, b-blockers, psychotropics, and nonsteroidal

anti-inflammatory molecules, possess an antimicrobial

action [4]. In particular, benzydamine, lidocaine [12], and

bupivacaine are known to have antimicrobial properties

[13]. The mechanisms of the antimicrobial activity of

ropivacaine [9] have not been clarified [1]. The antimicro-

bial effects on bacterial cells include reduction in the

number of viable cells, growth inhibition, ultrastructural

alterations, inhibition of membrane-bound enzymatic activ-

ities, and permeability changes [1]. The antimicrobial

activity of local anesthetics is thought to explain, at least

in part, the very low incidence of epidural abscess formation

after epidural anesthesia administration or infusion [7]. The

infection rate of patients affected by spasticity and treated

with totally implanted pumps ranges from 0.7% [11] to

1.7% [17]. Meningitis is a very rarely reported event [11].

Investigations on the antimicrobial activity of intrathecal

baclofen may be of clinical relevance because this drug

remains to be in the pump reservoir for 2 or 3 months after

each refill. The results failed to show significant differences

between control and baclofen-exposed bacterial and fungal

suspensions. It is of interest that the 2 bacterial species that

are frequently recovered from the hospital environment

[15]—E coli and P aeruginosa—showed a modest decrease

of viable cell counts over the 10-day observation time and

that baclofen had no activity against bacterial species that

circulate in the environment.

Careful disinfection and protection of the skin are

mandatory during surgical pump implantation and at the

time of pump refills. The microbial agents investigated in

this study are reported to be the most frequent pathogens

related to skin, soft tissue [14,15], and pump-related [2,6]

infections. To this end, high-level skin disinfection is

mandatory (eg, with alcohol, chlorhexidine, or iodophor)

[20]. Moreover, close supervision should be performed; in

the case of baclofen, long-term tests should be carried out by

temporary spinal catheter implantation.

In conclusion, the in vitro studies indicate that intrathecal

baclofen has no inhibitory activity on common bacteria and

C albicans. Thus, it is particularly important to avoid

contamination during surgical implantation and refills of the

infusion pump.

References

[1] Aydin ON, Eyigor M, Aydin N. Antimicrobial activity of ropivacaine

and other local anaesthetics. Eur J Anaesthesiol 2001;18:687 -94.

[2] Boviatsis EJ, Kouyialis AT, Boutsikakis I, Korfias S, Sakas DE.

Infected CNS infusion pumps. Is there a chance for treatment without

removal? Acta Neurochir (Wien) 2004;146:463-7.

A. Dario et al. / Surgical Neurology 68 (2007) 168–171 171

[3] Cook TM, James PA, Stannard CF. Diamorphine and bupivacaine

mixtures: an in vitro study of microbiological safety. Pain 1998;76:

259 -63.

[4] Cederlund H, Mardh PA. Antibacterial activities of non-antibiotic

drugs. J Antimicrob Chemother 1993;32:355-65.

[5] Dario A, Tomei G. A benefit-risk assessment of baclofen in severe

spinal spasticity. Drug Saf 2004;27:799-818.

[6] Galloway A, Falope FZ. Pseudomonas aeruginosa infection in an

intrathecal baclofen pump: successful treatment with adjunct intra-

reservoir gentamicin. Spinal Cord 2000;38:126-8.

[7] Goodman EJ, Jacobs MR, Bajaksouzian S, Windau AR, Dagirmanjian

JP. Clinically significant concentrations of local anesthetics inhibit

Staphylococcus aureus in vitro. Int J Obstet Anesth 2002;11:95 -9.

[8] Hindler JA. Special antimicrobial susceptibility tests. In: Mahon CR,

Manuselis G, editors. Textbook of diagnostic microbiology. 2nd ed.

New York7 W.B. Saunders Company; 2000. p. 97 -104.

[9] Kampe S, Poetter C, Buzello S, et al. Ropivacaine 0.1% with

sufentanil 1 microg/mL inhibits in vitro growth of Pseudomonas

aeruginosa and does not promote multiplication of Staphylococcus

aureus. Anesth Analg 2003;97:409-11.

[10] National Committee for Clinical Laboratory Standards. Methods for

determining bactericidal activity of antimicrobial agents. Document

M26-A. Wayne (Pa)7 National Committee for Clinical Laboratory

Standards, NCCLS; 1999.

[11] Ordia JI, Fischer E, Adamski E, Kimberly G, Chagnon MS, Edward L.

Continuous intrathecal baclofen infusion by a programmable pump in

131 consecutive patients with severe spasticity of spinal origin.

Neuromodulation 2002;5:16 -24.

[12] Parr AM, Zoutman DE, Davidson JS. Antimicrobial activity of

lidocaine against bacteria associated with nosocomial wound infec-

tion. Ann Plast Surg 1999;43:239-45.

[13] Pina-Vaz C, Rodrigues AG, Sansonetty F, Martinez-De-Oliveira J,

Fonseca AF, Mardh PA. Antifungal activity of local anesthetics

against Candida species. Infect Dis Obstet Gynecol 2000;8:124 -37.

[14] Pirbudak L, Kasarsligil T, Zer Y, Oner U, Balci I. Antibacterial effect

of bupivacaine and ropivacaine; effect of adjuvant drugs. Pain Clin

2005;17:73 -80.

[15] Rennie RP, Jones RN, Mutnick AH, SENTRY Program Study Group

(North America). Occurrence and antimicrobial susceptibility patterns

of pathogens isolated from skin and soft tissue infections: report from

the SENTRY Antimicrobial Surveillance Program (United States and

Canada, 2000). Diagn Microbiol Infect Dis 2003;45:287-93.

[16] Sakuragi T, Ishino H, Dan K. Bactericidal activity of clinically used

local anesthetics on Staphylococcus aureus. Reg Anesth 1996;21:

239-42.

[17] Stempien L, Tsai T. Intrathecal baclofen pump use for spasticity.

A clinical survey. Am J Phys Med Rehabil 2000;79:536 -41.

[18] Stratford AF, Zoutman DE, Davidson JS. Effect of lido-

caine and epinephrine on Staphylococcus aureus in a guinea pig

model of surgical wound infection. Plast Reconstr Surg 2002;110:

1275-9.

[19] Swenson JM, Hindler JF, Jorgensen JH. Special phenotypic methods

for detecting antimicrobial resistance. In: Murray PR, Baron EJ,

Jorgensen JH, Pfaller MA, Yolken RH, editors. Manual of clinical

microbiology. 8th ed. Washington7 ASM Press; 2003. p. 1178-95.

[20] Widmer AF, Frei R. Decontamination, disinfection, and sterilization.

In: Murray PR, Baron EJ, Jorgensen JH, Pfaller MA, Yolken RH,

editors. Manual of clinical microbiology. 8th ed. Washington7 ASM

Press; 2003. p. 77 -108.

[21] Winkelmuller M, Winkelmuller W. Long-term effects of continuous

intrathecal opioid treatment in chronic pain of nonmalignant etiology.

J Neurosurg 1996;85:458-67.

Commentary

This brief report addresses an important topic related to

infections of intrathecal infusion systems. I was not

surprised to discover lack of antimicrobial activity of

baclofen. (Actually, owing to my infrequent reading of

pharmacology journals, I was surprised to learn about the

antimicrobial properties of lidocaine and bupivacaine.)

Indeed, the utmost attention to sterility and the mandatory

use of bacteriostatic filters during pump refill procedures are

the issues that are uniformly accepted.

However, this article opens a window of opportunity for

drug developers and device manufacturers. Would it be

possible to make intrathecal baclofen preparations bacteri-

cidal or at least bacteriostatic? Would it be feasible to coat

the pump reservoirs and catheters with antibiotics or

antiadhesion media? These would not eliminate the risk

for infection and the need for sterility, but when it comes to

the development of meningitis or hardware infection, one

may not be too cautious. Even a handful of cases prevented

by this innovation would make it worthwhile.

Konstantin V. Slavin, MD

Department of Neurosurgery

University of Illinois at Chicago

Chicago, IL, USA