a Department of Pharmacy Practice, University of Nebraska Medical Center, Omaha, NE 68198-6045; b Section of Pulmonary and Critical Care Medicine, Louisiana State University Medical Center, Shreveport, LA 71130; c Division of Infectious Diseases, University of Nebraska Medical Center, Omaha, NE 68198, USA
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Abstract |
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Introduction |
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Previous studies examined lidocaine's antibacterial effects in various types of fluid, including 0.9% normal saline, bacterial and fungal growth media, Lactated Ringer's solution and distilled water.1,3,6 Although these fluids are used as laboratory standards, they do not duplicate the clinical practice of BALf collection and culture. No study to date has examined lidocaine's anti-infective potential in BALf. The effect of bacteria and fungal growth in BALf and its contents of normal saline, respiratory secretions, proteins, cells and other cellular components found in the lung airways remains unknown. This study determined the effect of lidocaine in BALf on the growth of Haemophilus influenzae, Moraxella catarrhalis, Candida albicans, Pseudomonas aeruginosa and Streptococcus pneumoniae.
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Materials and methods |
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Clinical respiratory isolates were obtained from the hospital laboratory and included four isolates of S. pneumoniae, two isolates of M. catarrhalis, and one isolate of P. aeruginosa, C. albicans and H. influenzae. Clinical isolates were initially inoculated upon the following agar plates: MuellerHinton agar (Difco, Detroit, MI, USA) for P. aeruginosa, chocolate agar (Becton Dickinson, Rutherford, NJ, USA) for H. influenzae, Sabouraud agar (Becton Dickinson) for C. albicans, and blood agar (Remel, Lenexa, KS, USA) for S. pneumoniae and M. catarrhalis. After growth, the isolates were transferred to appropriate broth solutions and were grown at 37°C with continuous shaking for 24 h before their final preparation. S. pneumoniae was grown at 37°C and 5% CO2 for 24 h.
Collection of BALf media
A BALf sample from a healthy donor was obtained during bronchoscopy. Bronchoscopy was performed using an estimated 1520 mL of 2% lidocaine (Abbott, Abbott Park, IL, USA) for anaesthesia followed by injection and aspiration of 100 mL of normal saline by standard technique. The lidocaine concentration (284 mg/L) in the BALf sample was determined by fluorescent polarization immunoassay (Abbott Diagnostic, Irving, TX, USA). BALf was filter sterilized using a 0.2 µm filter (Lida Corp., Kenosha, WI, USA) to remove potential bacterial contaminants, and the filtered BALf was cultured to confirm sterility before pathogen inoculation.
Study design
Following overnight growth, each organism was inoculated separately into fresh broth and incubated for 2 h with continuous shaking. The resulting bacterial suspensions were centrifuged at 1800g for 4 min and the pellet was resuspended in 10 mL of phosphate-buffered solution (Sigma, St Louis, MO, USA) to a predetermined absorbance that would equate to a concentration equal to 106 cfu/mL. Bacterial and fungal concentrations (cfu/mL) were quantified by plate count technique. Samples of the bacterial suspension (106 cfu) were added to test media to produce suspensions at the following final concentrations: 0.9% normal saline (NS) (Abbott), BALf with lidocaine (284 mg/L) and BALf supplemented with lidocaine (BALs) (3081 mg/L) (ElkinsSinn, Cherryhill, NJ, USA). The bacterial suspensions were of similar concentration to those found in clinical BALf samples from pneumonia patients.
To simulate clinical laboratory practice, BALf test suspensions remained at room temperature (25°C), under fluorescent lighting and ambient humidity after inoculation. Triplicate 50 µL samples from NS, BALf and BALs were pipetted and plated at time zero, 30 min and 60 min. After inoculation, agar plates were incubated for 24 h under previously described conditions and cfu counted.
Statistical analysis
Mean cfu/mL counts and the difference between the log10 cfu/mL were compared at each time period by ANOVA and Tukey's Test for multiple comparisons. The level of significance was set at P < 0.05.
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Results |
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Discussion |
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Our findings demonstrate that in at least some clinical isolates of S. pneumoniae growth is affected by lidocaine, resulting in a decrease by 1 log in the bacterial counts. Although other studies have not demonstrated similar results,1,5,8 differences in methods may explain our findings. Our study inoculated the isolates into BALf in contrast to using bacterial growth media or Lactated Ringer's solution as previously reported.1,5 The specific effect of BALf containing lidocaine upon bacterial isolates is unknown; however, growth media should provide better growth conditions than BALf, considering collection and laboratory transit time.
The clinical significance of these findings demonstrates that lidocaine may decrease the number of viable S. pneumoniae by a factor of 1 log in BALf over 1 h. As quantitative cultures are performed on the BALf specimens, a relatively modest antimicrobial effect of lidocaine could cause the concentration of bacteria to fall below a diagnostic threshold.9,10
Although these results are provocative, there were certain limitations to this study. First, only a limited number of isolates were tested. As only one isolate of H. influenzae, P. aeruginosa and C. albicans was tested, it is unknown whether testing more isolates would have changed the results. Multiple isolates of S. pneumoniae and M. catarrhalis were tested only after we demonstrated the reduction in growth with the first isolate.
The growth of two isolates of M. catarrhalis was significantly reduced in the presence of 0.9% normal saline. Reasons for this remain speculative. M. catarrhalis is a fastidious organism and it is possible that certain nutrients in BAL fluid supported its growth. Alternatively, M. catarrhalis is known to clump and the reduction in cfu/mL might have simply been an artefact of clumping, which did not occur in the protein-rich BALf. It is unknown whether this would have any clinical implications, as M. catarrhalis growth was not reduced in BAL fluid.
Bronchoalveolar lavage fluid had been frozen and filtered before use. It is unknown if potential antimicrobial products were removed by this process. There is also the possibility that freezing the BAL fluid at 70°C destroyed potentially harmful products. We acknowledge that these steps are not normal practice when processing BAL fluid samples.
Lidocaine appears to have a deleterious effect upon some isolates of S. pneumoniae. Future investigations should examine larger numbers of isolates and aim to determine whether this effect parallels antibiotic susceptibility or other bacterial properties.
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Notes |
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References |
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Received 4 January 1999; returned 16 June 1999; revised 6 August 1999; accepted 13 October 1999