Effect of gemifloxacin on viability of Chlamydia pneumoniae (Chlamydophila pneumoniae) in an in vitro continuous infection model

Persistent infection with Chlamydia pneumoniae (Chlamydophila pneumoniae) has been implicated in the development of atherosclerosis, asthma and other chronic diseases. However, data on treatment of C. pneumoniae infections are limited. Microbiological failure of antimicrobial therapy has been described, even after prolonged courses of treatment with azithromycin, doxycycline and erythromycin. Gemifloxacin is an enhanced-affinity fluoroquinolone with excellent activity against most common respiratory pathogens, including C. pneumoniae. The effect of prolonged treatment with gemifloxacin, compared with azithromycin, on viability of C. pneumoniae was investigated in a continuous infection model. Gemifloxacin at final con-centrations of 0.25 and 2.5 mg/L reduced the viability of C. pneumoniae by 5 log(10), which was similar to the effect of azithromycin. However, both antimicrobials failed to completely eliminate C. pneumoniae from continuously infected cells, even after 30 days of treatment. Both antibiotics decreased levels of interleukin-6 and interleukin-8 in this model, but this effect appeared to be secondary to the antichlamydial activity, as the cytokine levels correlated with the concentrations of microorganisms.     

Subinhibitory concentrations of azithromycin decrease nontypeable Haemophilus influenzae biofilm formation and Diminish established biofilms

Nontypeable Haemophilus influenzae (NTHi) commonly causes otitis media, chronic bronchitis in emphysema, and early airway infections in cystic fibrosis. Long-term, low-dose azithromycin has been shown to improve clinical outcomes in chronic lung diseases, although the mechanism of action remains unclear. The inhibition of bacterial biofilms by azithromycin has been postulated to be one mechanism mediating these effects. We hypothesized that subinhibitory concentrations of azithromycin would affect NTHi biofilm formation. Laboratory strains of NTHi expressing green fluorescent protein and azithromycin-resistant clinical isolates were grown in flow-cell and static-culture biofilm models. Using a range of concentrations of azithromycin and gentamicin, we measured the degree to which these antibiotics inhibited biofilm formation and persistence. Large biofilms formed over 2 to 4 days in a flow cell, displaying complex structures, including towers and channels. Subinhibitory concentrations of azithromycin significantly decreased biomass and maximal thickness in both forming and established NTHi biofilms. In contrast, subinhibitory concentrations of gentamicin had no effect on biofilm formation. Furthermore, established NTHi biofilms became resistant to gentamicin at concentrations far above the MIC. Biofilm formation of highly resistant clinical NTHi isolates (azithromycin MIC of > 64 microg/ml) was similarly decreased at subinhibitory azithromycin concentrations. Clinically obtainable azithromycin concentrations inhibited biofilms in all but the most highly resistant isolates. These data show that subinhibitory concentrations of azithromycin have antibiofilm properties, provide mechanistic insights, and supply an additional rationale for the use of azithromycin in chronic biofilm infections involving H. influenzae.     

Azithromycin exhibits bactericidal effects on Pseudomonas aeruginosa through interaction with the outer membrane

The aim of the present study was to elucidate the effect of the macrolide antibiotic azithromycin on Pseudomonas aeruginosa. We studied the susceptibility to azithromycin in P. aeruginosa PAO1 using a killing assay. PAO1 cells at the exponential growth phase were resistant to azithromycin. In contrast, PAO1 cells at the stationary growth phase were sensitive to azithromycin. The divalent cations Mg2+ and Ca2+ inhibited this activity, suggesting that the action of azithromycin is mediated by interaction with the outer membranes of the cells, since the divalent cations exist between adjacent lipopolysaccharides (LPSs) and stabilize the outer membrane. The divalent cation chelator EDTA behaved in a manner resembling that of azithromycin; EDTA killed more PAO1 in the stationary growth phase than in the exponential growth phase. A 1-N-phenylnaphthylamine assay showed that azithromycin interacted with the outer membrane of P. aeruginosa PAO1 and increased its permeability while Mg2+ and Ca2+ antagonized this action. Our results indicate that azithromycin directly interacts with the outer membrane of P. aeruginosa PAO1 by displacement of divalent cations from their binding sites on LPS. This action explains, at least in part, the effectiveness of sub-MICs of macrolide antibiotics in pseudomonal chronic airway infection.     

Uptake of azithromycin by human monocytes and enhanced intracellular antibacterial activity against Staphylococcus aureus.

The uptake of azithromycin by human monocytes and the intracellular antibacterial activity of azithromycin against Staphylococcus aureus were investigated. With an extracellular pH of 6.9, the maximum intracellular concentration of azithromycin in monocytes was about six times the extracellular concentration. The half-life for diffusion was 44 min. The results support the view that no active transport is involved in the intracellular accumulation of azithromycin. In cell-free medium, the maximum effect of azithromycin on S. aureus was bacteriostasis, which was achieved at a concentration of 5 mg/liter. In contrast, concentrations greater than 1.5 mg of azithromycin per liter were bactericidal for S. aureus ingested by monocytes. The difference in maximum growth inhibition on S. aureus for the two conditions was 0.1.68 h-1 (95% confidence interval, 0.128 to 0.208). The concentration of the drug that achieved 50% of the maximum effect was 0.434 mg/liter for both conditions. The enhancement of the effect on S. aureus ingested by monocytes suggests that the intracellular environment in human monocytes favors the antibacterial action of azithromycin. Enhancement of the antibacterial activity of azithromycin was not observed with granulocytes.     

Effect of Prolonged Treatment with Azithromycin, Clarithromycin, or Levofloxacin on Chlamydia pneumoniae in a Continuous-Infection Model

Persistent infections with Chlamydia pneumoniae have been implicated in the development of chronic diseases, such as atherosclerosis and asthma. Although azithromycin, clarithromycin, and levofloxacin are frequently used for the treatment of respiratory C. pneumoniae infections, little is known about the dose and duration of therapy needed to treat a putative chronic C. pneumoniae infection. In this study, we investigated the effect of prolonged treatment with azithromycin, clarithromycin, or levofloxacin on the viability of C. pneumoniae and cytokine production in an in vitro model of continuous infection. We found that a 30-day treatment with azithromycin, clarithromycin, and levofloxacin at concentrations comparable to those achieved in the pulmonary epithelial lining fluid reduced but did not eliminate C. pneumoniae in continuously infected HEp-2 cells. All three antibiotics decreased levels of interleukin-6 (IL-6) and IL-8 in HEp-2 cells, but this effect appeared to be secondary to the antichlamydial activity, as the cytokine levels correlated with the concentrations of microorganisms. The levels of IL-1β, IL-4, IL-10, tumor necrosis factor alpha, and gamma interferon were too low to assess the effect of antibiotics. These data suggest that the dosage and duration of antibiotic therapy currently being used may not be sufficient to eradicate a putative chronic C. pneumoniae infection.     

Bactericidal activity of first-choice antibiotics against gamma interferon-induced persistent infection of human epithelial cells by Chlamydia trachomatis

Chlamydia trachomatis is responsible for clinically important chronic inflammatory diseases of humans, including trachoma and pelvic inflammatory disease. Persistent infection of mucosal sites may contribute to the development of these chronic inflammatory diseases. Standard clinical therapy results in satisfactory cure rates of acute infections; however, chronic infection associated with persistence has been suggested to be less responsive to antibiotic therapy. We report the efficiency of two first-line chlamydial antibiotics, azithromycin and doxycycline, under conditions of eradication of C. trachomatis persistent infection using the in vitro model of gamma interferon (IFN-gamma)-mediated persistence and reactivation from persistence. Doxycycline was superior in eradicating acute (minimal bactericidal concentration [MBC](100) = 2.5 to 5.0 microg/ml) compared to persistent (MBC(100) = 10 to 50 mirog/ml) infection. In contrast, azithromycin was significantly more effective in eradicating persistent infection (MBC(100) = 2.5 to 5.0 microg/ml) than acute infection (MBC(100) = 10 to 50 microg/ml). The superior bactericidal effect of azithromycin against persistent infection was found to correlate with the enhanced uptake of the drug by IFN-gamma-treated infected epithelial cells. Based on these findings, we hypothesize that azithromycin should be a particularly efficacious anti-infective agent for the eradication of IFN-gamma-induced chlamydial persistent infection in vivo.     

In vitro activity of azithromycin in combination with amikacin,ceftazidime, ciprofloxacin or imipenem against clinical isolates of Acinobacter baumannii

The in vitro activity of the two-drug combinations of azithromycin with amikacin, ceftazidime, ciprofloxacin or imipenem against five clonally unrelated strains of Acinobacter baumannii were evaluated. Synergy studies were performed by the checkerboard microtiter method. The fractional inhibitory concentration (FIC) index was calculated for each drug combination. None of the four combinations tested was antagonistic. The combination of azithromycin and ceftazidime was synergistic (FIC index 相似文献   

Meta-Analysis of the Adverse Effects of Long-Term Azithromycin Use in Patients with Chronic Lung Diseases

The adverse effects of azithromycin on the treatment of patients with chronic lung diseases (CLD) were evaluated in the present study. MEDLINE and other databases were searched for relevant articles published until August 2013. Randomized controlled trials that enrolled patients with chronic lung diseases who received long-term azithromycin treatment were selected, and data on microbiological studies and azithromycin-related adverse events were abstracted from articles and analyzed. Six studies were included in the meta-analysis. The risk of bacterial resistance in patients receiving long-term azithromycin treatment was increased 2.7-fold (risk ratio [RR], 2.69 [95% confidence interval {95% CI}, 1.249, 5.211]) compared with the risk in patients receiving placebo treatment. On the other hand, the risk of bacterial colonization decreased in patients receiving azithromycin treatment (RR, 0.551 [95% CI, 0.460, 0.658]). Patients receiving long-term azithromycin therapy were at risk of increased impairment of hearing (RR, 1.168 [95% CI, 1.030, 1.325]). This analysis provides evidence supporting the idea that bacterial resistance can develop with long-term azithromycin treatment. Besides the increasingly recognized anti-inflammatory role of azithromycin used in treating chronic lung diseases, we should be aware of the potential for adverse events with its long-term use.     

Intracellular activity of azithromycin against bacterial enteric pathogens.

Azithromycin, a new azalide antibiotic, is active in vitro against a variety of enteric bacterial pathogens. Since it is concentrated inside human neutrophils and other cells, it might be particularly useful in the treatment of infections caused by enteropathogens that invade host tissues. The intracellular activity of azithromycin against several enteric pathogens that had been phagocytosed by neutrophils was determined. Azithromycin was effective in reducing the intracellular viabilities of almost all strains tested, including representative strains of Salmonella, Shigella, and enteroinvasive, enteropathogenic, enterotoxigenic, and enterohemorrhagic Escherichia coli. Erythromycin was also effective in this model system, although azithromycin was generally more effective than erythromycin against strains of invasive enteric pathogens. Cefotaxime reduced intracellular bacterial viability to a lesser extent than either azithromycin or erythromycin. The presence of neutrophils did not significantly affect the activity of azithromycin in this system. Azithromycin may be a useful agent for the treatment of bacterial diarrhea, and clinical trials should be considered.     

Anti-inflammatory effects of macrolides--an underappreciated benefit in the treatment of community-acquired respiratory tract infections and chronic inflammatory pulmonary conditions?

BACKGROUND: It has been recognized for more than 20 years that the macrolides have immunomodulatory effects that are beneficial for those suffering from chronic pulmonary inflammatory syndromes, such as diffuse panbronchiolitis, cystic fibrosis, asthma and bronchiectasis. The macrolides have consistently been associated with decreased length of stay and mortality when used alone or in combination with beta-lactam antibiotics. This effect can be demonstrated against combinations consisting of beta-lactams and other antibiotics active against 'atypical chest pathogens' when treating community-acquired pneumonia (CAP) in hospitalized patients. As such, it appears that the macrolides' effects in CAP patients are more than just antibacterial in nature. AIMS OF THIS REVIEW: This review aims: to give the reader information on the background areas described, as well as related areas; to review the CAP benefits with macrolides and how they may be related to the immunomodulatory properties they demonstrate, albeit in a shorter period of time than previously demonstrated with chronic pulmonary disorders; to use ex vivo data to support these extrapolations. LITERATURE SEARCH: A literature search using Medline was conducted from 1966 onwards, searching for articles with relevant key words such as macrolide, diffuse panbronchiolitis, community-acquired pneumonia, biofilm, immunomodulation, cystic fibrosis, erythromycin, clarithromycin, roxithromycin and azithromycin, bronchiectasis and asthma. When appropriate, additional references were found from the bibliographies of identified papers of interest. Any relevant scientific conference proceedings or medical texts were checked when necessary. CONCLUSIONS: (1) Research into macrolide immunomodulation for chronic pulmonary disorders demonstrates consistent positive effects, although of types other than seen with diffuse panbronchiolitis. These effects, together with their inhibitory activity on biofilms, have the potential to make them a useful option. (2) The benefits for CAP are consistent, and higher when a macrolide is given with another atypical agent than if the other atypical agent is given alone, suggesting a non-antibacterial benefit. (3) Recent research of the immunomodulatory properties of azithromycin imply that azithromycin may have a previously unknown short-term biphasic effect on inflammation modulation: enhancement of host defence mechanisms shortly after initial administration followed by curtailment of local infection/inflammation in the following period. (4) Additional in vivo research is needed prior to developing any firm conclusions.     

Azithromycin blocks autophagy and may predispose cystic fibrosis patients to mycobacterial infection

Azithromycin is a potent macrolide antibiotic with poorly understood antiinflammatory properties. Long-term use of azithromycin in patients with chronic inflammatory lung diseases, such as cystic fibrosis (CF), results in improved outcomes. Paradoxically, a recent study reported that azithromycin use in patients with CF is associated with increased infection with nontuberculous mycobacteria (NTM). Here, we confirm that long-term azithromycin use by adults with CF is associated with the development of infection with NTM, particularly the multi-drug-resistant species Mycobacterium abscessus, and identify an underlying mechanism. We found that in primary human macrophages, concentrations of azithromycin achieved during therapeutic dosing blocked autophagosome clearance by preventing lysosomal acidification, thereby impairing autophagic and phagosomal degradation. As a consequence, azithromycin treatment inhibited intracellular killing of mycobacteria within macrophages and resulted in chronic infection with NTM in mice. Our findings emphasize the essential role for autophagy in the host response to infection with NTM, reveal why chronic use of azithromycin may predispose to mycobacterial disease, and highlight the dangers of inadvertent pharmacological blockade of autophagy in patients at risk of infection with drug-resistant pathogens.     

A comparison of 5-day courses of dirithromycin and azithromycin in the treatment of acute exacerbations of chronic obstructive pulmonary disease

BACKGROUND: Short-term use of antibiotics has become a common component of the management of acute exacerbations of chronic bronchitis (AECB), particularly in complex cases with productive cough or purulent phlegm. The macrolide antibiotics, particularly second-generation agents such as dirithromycin and azithromycin, are among the antibiotic classes frequently recommended and used to treat upper and lower respiratory infections, including AECB. OBJECTIVE: This study compared the clinical efficacy and tolerability of 5-day courses of dirithromycin and azithromycin given once daily for the treatment of acute exacerbations of chronic obstructive pulmonary disease (COPD). METHODS: This randomized, investigator-blinded, parallel-group clinical trial was conducted at 5 centers in the United States. Eligible patients were adult (age >35 years) smokers or ex-smokers (smoking history of at least 10 pack-years) with chronic bronchitis and an acute exacerbation, defined by the occurrence of increased dyspnea and/or productive cough and feverishness within 48 hours of enrollment. Before randomization, an attempt was made to obtain a sputum specimen from each patient for Gram's staining and culture. Patients were randomized to receive dirithromycin 500 mg QD for 5 days or azithromycin 500 mg QD on day 1 and 250 mg QD on days 2 to 5. Clinical efficacy was assessed separately by patients and physicians at early (days 7-10) and late (days 25-35) posttreatment visits. RESULTS: Eighty-six patients (48 women, 38 men; mean age, 55 years) with a mean smoking history of 31 pack-years were included in the intent-to-treat analysis. Forty-six (54%) patients were randomized to dirithromycin and 40 (47%) patients to azithromycin. Clinical efficacy was reported in a high proportion of patients in both treatment groups, both at the early posttreatment visit (84.8% dirithromycin, 75.7% azithromycin; difference dirithromycin - azithromycin, 9.1%; 95% CI, -8.2 to 26.4) and the late posttreatment visit (95.5% and 86.5%, respectively; difference dirithromycin - azithromycin, 9.0%; 95% CI, -3.7 to 21.6). A similar proportion of patients required a second course of antibiotics over the study period (20.5% dirithromycin, 27.0% azithromycin; difference dirithromycin - azithromycin, -6.6%; 95% CI, -25.2 to 12.1). Only 42 (48.8%) patients were able to produce a sputum sample before receiving study treatment, and of these, only 20 (47.6%) demonstrated a preponderance of neutrophils on Gram's staining. Both treatments were well tolerated. CONCLUSIONS: The results of this study suggest comparable clinical efficacy between 5-day courses of once-daily dirithromycin and azithromycin in acute exacerbations of COPD. There were insufficient data to permit meaningful comparison of the bacteriologic efficacy of these macrolide antibiotics.     

阿奇霉素对特发性肺间质纤维化干预作用的临床研究

目的探讨阿奇霉素治疗特发性肺间质纤维化（IPF）的临床疗效。方法IPF患者34例，随机分为阿奇霉素组20例（阿奇霉素0．5g/次，1次／d，静脉滴注；症状好转后改为片剂，0125g／次，1次／d，口服）和泼尼松组14例（泼尼松30mg／次，1次／d，口服；4周后减为15mg／次，1次／d，口服；8周后改为隔日1次，口服）；两组疗程均为3个月。观察两组患者临床表现、肺功能改变情况及副作用。结果两组患者的临床表现、肺功能均有不同程度的改善，但差异无显著性（P〉0．05），阿奇霉素组副作用显著少于泼尼松组（P〈0．05）。结论阿奇霉素治疗IPF具有与口服泼尼松相当的疗效且副作用少．临床值得推广。     

Single-dose azithromycin for acute otitis media: a pharmacokinetic/pharmacodynamic rationale

The pharmacokinetic (PK) and pharmacodynamic (PD) properties of the azalide azithromycin distinguish it from other antibiotics. The PK profile of azithromycin features high tissue-to-serum ratios, including high concentrations in the middle ear, and a prolonged elimination half-life. These characteristics result from the accumulation of drug within cells and its subsequent slow, sustained release from cells and tissues into the bloodstream. The PD properties of azithromycin include bactericidal activity against key respiratory tract pathogens and a prolonged postantibiotic, or persistent, effect. In addition, white blood cells deliver the drug to infected foci, thereby enhancing local tissue concentrations and improving in vivo efficacy. Recent PK studies in mice suggest that a single, large dose of azithromycin achieves higher tissue concentrations than do multidose regimens. Other studies in animal infection models, in particular, a gerbil model of acute otitis media, have demonstrated improved bacterial eradication when azithromycin is administered as a single dose rather than divided over 2 or 3 days. Taken together, the results from these preclinical studies provide a PK/PD rationale for the use of single-dose azithromycin in the treatment of acute otitis media. Clinical data on the efficacy and safety of single-dose azithromycin for the treatment of acute otitis media in children are presented in 2 accompanying articles in this supplement.     

Possible involvement of the drug transporters P glycoprotein and multidrug resistance-associated protein Mrp2 in disposition of azithromycin

P glycoprotein and multidrug resistance-associated protein 2 (Mrp2), ATP-dependent membrane transporters, exist in a variety of normal tissues and play important roles in the disposition of various drugs. The present study seeks to clarify the contribution of P glycoprotein and/or Mrp2 to the disposition of azithromycin in rats. The disappearance of azithromycin from plasma after intravenous administration was significantly delayed in rats treated with intravenous injection of cyclosporine, a P-glycoprotein inhibitor, but was normal in rats pretreated with intraperitoneal injection erythromycin, a CYP3A4 inhibitor. When rats received an infusion of azithromycin, cyclosporine and probenecid, a validated Mrp2 inhibitor, significantly decreased the steady-state biliary clearance of azithromycin to 5 and 40% of the corresponding control values, respectively. However, both inhibitors did not alter the renal clearance of azithromycin, suggesting the lack of renal tubular secretion of azithromycin. Tissue distribution experiments showed that azithromycin is distributed largely into the liver, kidney, and lung, whereas both inhibitors did not alter the tissue-to-plasma concentration ratio of azithromycin. Significant reduction in the biliary excretion of azithromycin was observed in Eisai hyperbilirubinemic rats, which have a hereditary deficiency in Mrp2. An in situ closed-loop experiment showed that azithromycin was excreted from the blood into the gut lumen, and the intestinal clearance of azithromycin was significantly decreased by the presence of cyclosporine in the loop. These results suggest that azithromycin is a substrate for both P glycoprotein and Mrp2 and that the biliary and intestinal excretion of azithromycin is mediated via these two drug transporters.     

Enhanced hypoprothrombinemia with warfarin due to azithromycin

OBJECTIVE: To report a case of probable azithromycin-warfarin drug interaction with enhanced hypoprothrombinemic effect of warfarin. CASE SUMMARY: An 83-year-old African American man stabilized on warfarin therapy (10 mg on Wednesdays, 7.5 mg on other days) developed a prolonged prothrombin time one day after starting azithromycin 500 mg. The elevated prothrombin time normalized 3 days after azithromycin was discontinued. After the initial increase in the international normalized ratio, the absence of any significant confounding factors affecting the anticoagulant effect of warfarin in our patient and the numerous reports of such interactions indicate that an interaction between azithromycin and warfarin may have been responsible for the elevated prothrombin time seen in this patient. An objective causality assessment revealed that the adverse event was probably related to the combination of these drugs. DISCUSSION: Azithromycin, unlike erythromycin and clarithromycin, is not known to inhibit the cytochrome P450 enzyme system and is presumed to be the macrolide of choice in patients already on warfarin. However, previously reported cases of azithromycin-warfarin interactions support the possibility that azithromycin does interact with warfarin, although the exact mechanism is not understood. CONCLUSIONS: Azithromycin may interact with warfarin and enhance its hypoprothrombinemic effects. This effect may be delayed for 4-8 days after a course of azithromycin has been completed. Periodic monitoring of the prothrombin time is recommended when using azithromycin in patients taking warfarin.     

阿奇霉素和红霉素转换治疗儿童支原体肺炎疗效比较

目的用阿奇霉素和红霉素分别转换治疗儿童支原体肺炎,比较两者临床疗效及不良反应。方法66位患儿随机分两组,阿奇霉素组36例,静滴阿奇霉素10mg／（kg·d）,1次／d,连用5d,停用4d,转换为阿奇霉素糖浆10mg／（kg·d）,1次／d,连用3d。红霉素组30例,红霉素针剂25～30mg／（kg·d）,1次／d,连用5d,转换为红霉素片25—30mg／（kg·d）,分3次口服,连用7d。临床观察治疗效果和不良反应发生率,并对肺外并发症进行半年随访。结果阿奇霉素组优良率为94．44％,红霉素组优良率为83．33％,优良率阿奇霉素组高于红霉素组,但无统计学意义。胃肠道反应、注射部位疼痛和肝功能损害三个主要的不良反应发生率阿奇霉素组出现8,3,1例,红霉素组出现20,12,6例,二组有统计学意义。随访半年,在慢性咳嗽、头痛伴脑电图异常、关节疼痛、粒细胞减少、荨麻疹等并发症,二组无明显差异。结论阿奇霉素转换治疗儿童支原体肺炎疗效肯定,不良反应少,儿童支原体肺炎可优先考虑使用阿奇霉素。     

Activity of azithromycin against Mycobacterium avium infection in beige mice.

The comparative activities of azithromycin and clarithromycin and the activities of azithromycin alone and in combination with other antimycobacterial agents were evaluated in the beige mouse model of disseminated Mycobacterium avium complex infection. Azithromycin was similar in activity to clarithromycin. Azithromycin plus clofazimine plus ethambutol reduced the number of splenic organisms more than azithromycin alone, while the combination was less active than azithromycin alone for bacteria in lungs. Rifabutin had activity similar to that of azithromycin for organisms in spleens and lungs. Rifabutin plus azithromycin was more active than either agent alone for organisms in spleens, but the combination's activity was not significantly different from that of rifabutin for organisms in lungs. The activity of azithromycin against several M. avium complex isolates was evaluated. The reduction of viable cell counts in spleens ranged from 1.7 to 0.8 log units. For the three isolates studied, there was little correlation between the in vitro MIC and the in vivo activity.     

The effect of azithromycin on intracellular Legionella pneumophila in the Mono Mac 6 cell line at serum concentrations attainable in vivo

We investigated the antimicrobial efficacy of clinically meaningful, low concentrations of azithromycin against intracellular growth of two clinical isolates of Legionella pneumophila. The mature monocytic cell line Mono Mac 6 was used as a model to investigate the effects of antimicrobial agents on L. pneumophila. Extracellular susceptibility was determined by microdilution susceptibility testing in BYEalpha broth after 48 h of incubation. Mono Mac 6 cells infected with L. pneumophila were incubated with various concentrations of azithromycin. After 2 days of incubation, intracellular bacteria were released from the phagocytes and plated on to BCYEalpha agar. Addition of the intracellular-acting antibiotics azithromycin or ciprofloxacin at their MICs (0.5 and 0. 015 mg/L, respectively) resulted in a significant decrease in cfu, of up to approximately 1 log(10) after 48 h of incubation. In contrast, incubation of intraphagocytic L. pneumophila in the presence of antibiotics without intracellular activity (ceftizoxime, imipenem or amoxycillin-clavulanic acid) did not have any effect. Azithromycin inhibited intracellular replication at concentrations as low as 0.125 mg/L, approximately one-quarter of the extracellular MIC. The Mono Mac 6 cell line is a useful infection model for investigating the intracellular activity of antimicrobial agents in vitro. In accordance with clinical data and animal experiments, azithromycin and ciprofloxacin inhibited the intraphagocytic replication of L. pneumophila. In particular, azithromycin killed ingested legionellae in vitro at concentrations below the peak serum concentrations and below the MIC.