In vitro activities of new quinolones against Brucella melitensis isolated in a tertiary-care hospital in Turkey

We have evaluated the in vitro activities of seven fluoroquinolones against 69 strains of Brucella melitensis . According to their minimum inhibitory concentration for 90% growth (MIC₉₀) values, the most active agent was found to be sparfloxacin (MIC₉₀ 0.12 mg/L) followed by levofloxacin, ciprofloxacin, ofloxacin (MIC₉₀ 0.50 mg/L) and grepafloxacin (MIC₉₀ 1 mg/L), gemifloxacin (MIC₉₀ 2 mg/L) and gatifloxacin (MIC₉₀ 4 mg/L).     

Activity of telithromycin against 26 quinolone-resistant pneumococci with known quinolone-resistance mechanisms

NCCLS agar dilution was used to test activity of telithromycin compared to clarithromycin, penicillin G, ciprofloxacin, levofloxacin, sparfloxacin and moxifloxacin against 26 pneumococci with defined quinolone resistance (type II topoisomerase and efflux) mechanisms. Thirteen strains were penicillin susceptible, six intermediate and seven resistant. Clarithromycin resistance ( mef and/or erm ) was seen in eight strains. Ciprofloxacin MICs (mg/L) were 8–64 compared to 1–32 (levofloxacin), 0.5 ≥ 32 (sparfloxacin) and 0.125–4 (moxifloxacin). Telithromycin MIC₅₀ and MIC₉₀ values (mg/L) were 0.016 and 0.25, with only one strain having an MIC of 2 mg/L.     

The DUEL study: a multi-center in vitro evaluation of linezolid compared with other antibiotics in the Netherlands

Objective   To evaluate bacterial susceptibility to linezolid in the Netherlands in comparison with other antibiotics.
Methods   Bacterial strains were isolated between September 1999 and January 2000 from patients presumed to require antibiotic treatment. The in vitro activity of 1226 strains from 34 participating laboratories was tested against linezolid, vancomycin, teicoplanin, oxacillin, penicillin, erythromycin, ampicillin and other antibiotics against enterococci, coagulase-negative staphylococci, Staphylococcus aureus and Streptococcus pneumoniae . Minimal inhibitory concentrations (MIC) were obtained with the E test on Mueller-Hinton agar: every laboratory included control strains. For vancomycin and teicoplanin only, brain–heart infusion agar and an inoculum of 2.0 McFarland was used for Staphylococcus aureus , coagulase-negative staphylococci and enterococci to support a better growth and clear recognition of hetero-resistant colonies.
Results   The values of MIC₉₀ for linezolid were 1.5, 0.75, 0.75 and 1 mg/L for Staphylococcus aureus , coagulase-negative staphylococci, Streptococcus pneumoniae and enterococci, respectively. Six enterococcal strains with decreased susceptibility against vancomycin or teicoplanin were identified as Enterococcus faecium , E. gallinarum and E. casseliflavus (two strains each) and they were found to harbor vanA , vanC1 and vanC2/3 genes, respectively. Nine per cent of Streptococcus pneumoniae (an increase from 1% 4 years ago) showed decreased susceptibility to erythromycin, of both the ermB and mefE type; there was no cross-resistance with linezolid. Twelve coagulase-negative staphylococcal strains were resistant to teicoplanin.
Conclusion   Linezolid is a promising drug in the treatment of infections caused by Gram-positive cocci. Cross-resistance with other antibiotics tested was not found.     

CQ-397 and CQ-414: antimicrobial activity and spectrum of two fluoroquinolone–cephalosporin, dual-action compounds with carboxamido bonds

Objective: To evaluate the potential spectrum of activity of two novel dual-action compounds with carboxamido bonds (CQ-397 and CQ-414; Laboratories Aranda, San Rafael, Mexico) against human pathogens.
Method: Approximately 800 Gram-positive and Gram-negative aerobic clinical bacteria were tested in vitro using the Mueller-Hinton broth microdilution method of the National Committee of Clinical Laboratory Standards.
Results: CQ-397 (cefamandole+enrofloxacin) and CQ-414 (cefamandole+norfloxacin) were equally potent against Enterobacteriaceae (MIC₉₀ range, 0.06–0.5 μg/mL and 0.06-1 μg/mL, respectively). Citrobacter freundii (MIC₉₀, 4 μg/mL) and Providencia spp. (MIC₉₀,>32 μg/mL) exhibited elevated study drug MICs. Enterobacteriaceae resistant to fluoroquinolones generally remained resistant. CQ-397 and CQ-414 were active against Stenotrophomonas maltophilia (MIC₉₀, 4 μg/mL) and oxacillin-susceptible staphylococci (MIC₉₀, 0.25 μg/mL), but not oxacillin-resistant Staphylococcus aureus (MIC₉₀,>32 μg/mL), Staphylococcus epidermidis (MIC₉₀, 8 μg/mL), and enterococci (MIC₉₀s, 8 to>32 μg/mL). There was no difference in the dual-action drug activity (MIC₉₀, 2 μg/mL) between penicillin-susceptible and -resistant pneumococci. Haemophilus influenzae and Moraxella catarrhalis were very susceptible (MIC range, <0.015-0.06 μg/mL) to both compounds.
Conclusions: The activity of these novel dual-action compounds, formed from the bonding of older antimicrobials, warrants further investigation for potential human and/or animal health use, including toxicology and pharmacokinetics.     

In vitro anti-anaerobic activity of the cephalosporin derivative RWJ 54428, compared to seven other compounds

Agar dilution MIC was used to test the activity of RWJ 54428, a new cephalosporin derivative, compared to imipenem, meropenem, ceftriaxone, piperacillin, piperacillin–tazobactam, clindamycin and metronidazole against 363 anaerobes isolated from clinical specimens. RWJ 54428 had low MICs against most β -lactamase-negative Gram-negative rods, and all Gram-positive strains except Clostridium difficile . Imipenem and meropenem had the lowest MICs (MIC₅₀s of 0.125 mg/L and MIC₉₀s of 1.0 mg/L). Piperacillin–tazobactam, clindamycin and metronidazole were active against most strains, and ceftriaxone was active mainly against β -lactamase-negative organisms.     

Comparative in vitro activities of five quinolone antibiotics, including gemifloxacin, against clinical isolates

The in vitro activities of ciprofloxacin, ofloxacin, norfloxacin, levofloxacin and gemifloxacin against 343 clinical isolates were compared. Gemifloxacin showed the greatest activity, with MIC₉₀ values as low as 0.03–0.25 mg/L against Streptococcus pneumoniae , Haemophilus influenzae , Moraxella catarrhalis , methicillin-susceptible Staphylococcus aureus and Klebsiella pneumoniae , while methicillin-resistant Staphylococcus aureus , Enterococcus spp., Pseudomonas spp., Acinetobacter spp., Escherichia coli and Enterobacter spp. strains exhibited low rates of susceptibility to all five fluoroquinolones.     

Antimicrobial susceptibility of 840 clinical isolates of Haemophilus influenzae collected in four European countries in 2000–2001

In 2000–2001, 840 clinical isolates of Haemophilus influenzae were collected from laboratories in France, Germany, Italy and Spain (210 isolates/country). β -Lactamase production among the isolates varied considerably by country, ranging from 8.1% in Germany to 34.8% in France. H. influenzae from patients ≤4 years old showed the highest prevalence of β -lactamase production (23.2%), compared with isolates from patients aged 5–17 years (17.8%) and ≥18 years (16.5%). All isolates were susceptible to amoxicillin–clavulanate, ciprofloxacin and levofloxacin; 99.6% and 98.9% of isolates were susceptible to azithromycin and cefuroxime, respectively. Among the macrolides tested, azithromycin (MIC₉₀, 2 mg/L) was eight-fold more potent than clarithromycin (MIC₉₀, 16 mg/L) and roxithromycin (MIC₉₀, 16 mg/L). Despite variations in β -lactamase production between different countries, > 99% of all isolates were susceptible to amoxicillin–clavulanate, ciprofloxacin, levofloxacin, and azithromycin.     

In vitro activity of tigecycline and occurrence of tetracycline resistance determinants in isolates from patients enrolled in phase 3 clinical trials for community-acquired pneumonia

The in vitro activity of tigecycline was evaluated against baseline pathogens isolated from patients enrolled in phase 3 clinical trials for community-acquired pneumonia conducted in 29 countries worldwide. Tigecycline was active against the most prevalent pathogens, including Streptococcus pneumoniae (MIC₉₀ 0.06 mg/L), Staphylococcus aureus (MIC₉₀ 0.25 mg/L), Haemophilus influenzae (MIC₉₀ 0.5 mg/L) and Klebsiella pneumoniae (MIC₉₀ 1 mg/L). Twelve isolates of S. pneumoniae expressing tet (M) and two isolates of K. pneumoniae producing extended-spectrum β-lactamases isolated during the study were susceptible to tigecycline. The excellent in vitro activity of tigecycline against these clinical isolates confirmed its potential utility against pathogens associated with community-acquired pneumonia.     

In vitro activity of quinupristin/dalfopristin in comparison with five antibiotics against worldwide clinical isolates of staphylococci

Objective   To evaluate the in vitro activity of quinupristin/dalfopristin (Q/D), a streptogramin combination, in comparison with five antibiotics against worldwide clinical isolates of staphylococci.
Methods   A multicenter in vitro study was performed using the E test during a period of 3 months (April to June) in 1997 on fresh, clinically significant, non repetitive strains of staphylococci from patients hospitalized in 23 different hospitals in 18 countries tested.
Results   A total of 2132 staphylococcal isolates including methicillin resistant (MR), methicillin susceptible (MS) S. aureus (1003 MS, 462 MR), S. epidermidis (169 MS, 251 MR), S. haemolyticus (28 MS, 46 MR), S. hominis (28 MS, 16 MR), and coagulase negative staphylococci (86 MS, 43 MR) were analyzed.
Q/D was highly active against all species tested. MIC₉₀ (mg/L) ranged from 0.5 to 2 depending on the species. Strains had MIC≤1 mg/L in 97.6%. For S. aureus , S. epidermidis , S. hominis and other coagulase-negative staphylococci no differences in MIC₉₀ were observed for MS or MR. One dilution difference was observed for S. haemolyticus , which overall was the less susceptible species. Erythromycin resistance was observed among 57– 87% of MR-strains and was lower among MS-strains (18–56%). Erythromycin resistance had no or little influence on MIC of Q/D. In comparison to vancomycin, Q/D was two to four times more active.
Conclusions   The streptogramin combination Q/D showed an excellent in vitro activity against all staphylococcal species tested regardless of the resistance pattern to other drug classes, particularly resistance to methicillin. Q/D was two to four times more active than vancomycin and MIC values varied from 0.5–2 according to the species. The synergy of Q/D was well conserved in macrolide-resistant strains.     

Antimicrobial activity of LB10827, a new orally administered cephalosporin, tested against Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae

A new orally administered cephalosporin, LB10827, was compared to 16 other antimicrobial agents tested against Streptococcus pneumoniae (520 strains), Haemophilus influenzae (302 strains) and Moraxella catarrhalis (188 strains) by reference broth microdilution methods. LB10827 (MIC₉₀, 0.12 mg/L; highest MIC, 0.5 mg/L) was 8–16-fold more potent than cefdinir, cefpodoxime or cefuroxime when tested against S. pneumoniae . All Gram-negative strains were inhibited at ≤ 0.5 mg/L LB10827, which is an activity equal or superior to that of currently available and tested oral β-lactams. LB10827 appears to be a promising agent worthy of continued investigations both in vitro and in vivo.     

In vitro activity of levofloxacin against contemporary clinical isolates of Legionella pneumophila, Mycoplasma pneumoniae and Chlamydia pneumoniae from North America and Europe

Objective   To assess the activities of levofloxacin and the comparator agents erythromycin, clarithromycin, azithromycin and doxycycline against atypical respiratory pathogens.
Methods   One hundred and forty-six Legionella pneumophila , 41 Mycoplasma pneumoniae and nine Chlamydia pneumoniae isolates were procured from various culture collections in North America and Europe and tested for susceptibility to the above agents by broth microdilution. The isolates came primarily from clinical sources and were collected from patients between 1995 and 1999.
Results   Against L. pneumophila , levofloxacin was the most active agent, with an MIC₉₀ of 0.03 mg/L, twofold more active than clarithromycin (0.06 mg/L), 16-fold more active than erythromycin and azithromycin (0.5 mg/L) and 64-fold more active than doxycycline. Against M. pneumoniae , azithromycin (MIC₉₀ ≤ 0.0005 mg/L) was the most active agent. However, two isolates of M. pneumoniae , one from the USA and one from Finland, were macrolide resistant (MIC ≥ 4 mg/L), but levofloxacin susceptible (MIC 0.25 mg/L). The geographic origin of L. pneumophila and M. pneumoniae did not affect the MIC range for any antimicrobial agent tested. Against C. pneumoniae , clarithromycin was the most active agent, with an MIC range of ≤0.008–0.03 mg/L.
Conclusions   Levofloxacin had comparable activity to the other agents tested against the atypical respiratory pathogens, confirming its potential as an alternative for empirical therapy of community-acquired pneumonia.     

Antimicrobial Activity of SCH 27899, Oligosaccharide Member of the Everninomycin Class with a Wide Gram-Positive Spectrum

The in vitro antimicrobial activity of SCH 27899 (everninomycin), a novel oligosaccharide compound of the everninomycin class, was compared with vancomycin, chloramphenicol, clinafloxacin, teicoplanin and doxycycline against 428 clinical strains of bacteria. Everninomycin base exhibited the greatest antimicrobial activity compared to other formulations against all strains tested (MIC₉₀: 0.25 μ/ml) followed by clinafloxacin and teicoplanin (MIC₉₀: 0.5 μg/ml), vancomycin (MIC₉₀: 2 μg/ml), and doxycycline (MIC₉₀: 16 μg/ml). Everninomycin demonstrated the best activity against Streptococcus spp. (serogroups A, B, C, F, G) and Streptococcus pneumoniae , and lower activity against coagulase-negative staphylococci (MIC₉₀: 0.5 μg/ml). All enterococci had an everninomycin MIC of 0.5 μg/ml or less. Everninomycin had no measurable antimicrobial activity against gram-negative aerobic organisms except Flavo-bacterium meningosepticum (MIC₅₀: 2 μg/ml). Some everninomycin activity was observed against Clostridium spp., Peptostreptococcus spp., and the Prevotella bivius-disiens group. Everninomycin showed excellent activity (MIC₉₀: 0.25 μg/ml) against the fluoroquinolone-resistant strains and all gram-positive strains resistant to vancomycin (MICs 4 μg/ml). The MBC/MIC ratios and killing curve data suggest that everninomycin is not uniformly or rapidly bactericidal. These in vitro data indicate that everninomycin could be useful against emerging gram-positive strains resistant to other contemporary antimicrobials.     

Antimicrobial activity against strains of Escherichia coli and Klebsiella spp. with resistance phenotypes consistent with an extended-spectrum β-lactamase in Europe

Extended-spectrum β-lactamases (ESBLs) have continued to evolve after their initial detection in Europe nearly two decades ago. The summary results from the MYSTIC Program (31 medical centers) were utilized to assess the extent of ESBL occurrence in Europe from 1997 to 2000. ESBL phenotype rates in Klebsiella spp. (32.8%) and Escherichia coli (14.4%) were generally stable, but extensive hospital-to-hospital and unit-to-unit variations were noted. The highest ESBL rates were found in eastern Europe (including Turkey) and in intensive care unit patient populations. Carbapenems remained active against the ESBL-producing strains (meropenem MIC₉₀, 0.25–1 mg/L), while some other agents, such as aminoglycosides, fluoroquinolones, and piperacillin–tazobactam, were significantly less effective. International surveillance initiatives should be maintained to monitor future progression of this important resistance.     

Highly antibiotic-resistant Acinetobacter baumannii clinical isolates are killed by the green tea polyphenol (–)-epigallocatechin-3-gallate (EGCG)

Acinetobacter baumannii is an increasingly common cause of infection in intensive-care units throughout the world, and the occurrence of multiresistant A. baumannii is increasing. The aim of this study was to determine whether a highly purified polyphenol, (–)-epigallocatechin-3-gallate (EGCG), from green tea ( Camellia sinesis ), had antimicrobial effects against multiresistant clinical isolates of A. baumannii. Standard microplate assays were performed to determine the MIC of EGCG for 21 clinical isolates of A. baumannii. MICs ranged from 0.078 to 0.625 mg/mL, with MIC₅₀ and MIC₉₀ of 0.312 mg/mL and 0.625 mg/mL, respectively. All of the isolates of A. baumannii tested were killed by EGCG. In time-kill assays, EGCG resulted in a 3-log reduction in CFU/mL of A. baumannii after 5 h of incubation with the polyphenol. Synergy between the commonly used topical agent 5% mafenide acetate (Sulfamylon) and EGCG was noted for one clinical isolate, and partial synergy was noted for three other isolates. These findings demonstrate that EGCG is an effective bactericidal agent against antibiotic-resistant A. baumannii clinical strains in laboratory settings. EGCG has previously been shown to be safe, and therefore may be an attractive addition for the treatment of cutaneous A. baumannii infections where high concentrations of the drug can be applied to the wound surface.     

Prevalence of erm methylase genes in clinical isolates of non-pigmented, rapidly growing mycobacteria

The aim of this study was to determine the frequency of erm genes coding for macrolide resistance among clinical isolates of non-pigmented rapidly growing mycobacteria (NPRGM) and to evaluate their importance in phenotypic resistance. Broth microdilution susceptibility testing was performed for all NPRGM tested. A PCR assay with consensus primers was used to evaluate the presence of erm genes among the 167 clinical isolates studied, which belonged to nine species of NPRGM; erm genes were detected in all nine species and 109 strains were erm -positive. The highest percentage of erm -positive isolates was found among Mycobacterium mageritense (100%) and the lowest among Mycobacterium mucogenicum (14%). The MICs of macrolides were found to be lower for erm -negative isolates (MIC₉₀: 2 mg/L) than for erm- positive isolates (MIC₉₀: 16 mg/L), although in some cases high MICs were found for erm -negative isolates. The finding that erm methylases are present in the majority of the species of NPRGM analysed in this study is not in agreement with conventional susceptibility studies. It therefore appears necessary to use a combination therapy to treat infections caused by NPRGM.     

Susceptibility of strains of Streptococcus agalactiae to macrolides and lincosamides, phenotype patterns and resistance genes

The Group B streptococcus ( Streptococcus agalactiae ) is a pathogen of increasing importance in human disease. We therefore studied the susceptibility of clinical isolates of S. agalactiae to penicillin G, erythromycin, azithromycin and clindamycin using National Committee for Clinical Laboratory Standards methodology, and we also determined the phenotypes of macrolide-lincosamide susceptibility and the resistance genes implicated in a group of selected isolates of the different phenotypes. We used 221 isolates collected between 1997 and 1999 in two Health Authority Areas in Móstoles and Granada, Spain. The minimal concentration for 90% inhibition (MIC₉₀) for penicillin G was 0.12 mg/L and all the isolates tested were susceptible. One hundred and eighty-five (83.7%) were susceptible to erythromycin and azithromycin and 191 (86.4%) were susceptible to miocamycin and clindamycin. Twenty-three isolates (10.4%) had a constitutive MLS_B phenotype, seven (3.2%) an inducible phenotype, and six (2.7%) an M phenotype. All except one of the MLS_B phenotype isolates tested ( n  = 23) carried erm genes; in two strains with the mef (A) gene, all the M phenotype ( n  = 6) isolates tested carried mef genes, while erm and mef (A) genes were absent in all the macrolide-lincosamide-susceptible ( n  = 12) isolates tested. In our environment, resistance to macrolide and lincosamide in S. agalactiae was present in 10–16% of the isolates. The majority of resistant strains had the MLS_B phenotype.     

European survey of glycopeptide susceptibility in Staphylococcus spp.

Objectives: To reassess the relative potencies of teicoplanin and vancomycin following several years of clinical usage.
Methods: The glycopeptide susceptibilities of clinical isolates of staphylococci collected from 70 hospitals in 1995 were determined using NCCLS (National Committee for Clinical Laboratory Standards) methods.
Results: In total, 2885 isolates of Staphylococcus aureus and 1480 isolates of coagulase-negative staphylococci were collected. S. aureus was significantly less susceptible to vancomycin (MIC₅₀ 1 mg/L) than teicoplanin (MIC₅₀ 0.5 mg/L), but the reverse was the case for S. haemolyticus and S. epidermidis . No S. aureus isolate was resistant (≥32 mg/L) to either glycopeptide, but nine isolates of coagulase-negative staphylococci had an MIC of teicoplanin of 32 mg/L. Respiratory isolates of S. aureus were less susceptible to glycopeptides than those from other sites. Staphylococci from Belgium and Italy were less susceptible to teicoplanin than isolates from other countries.
Conclusions: This European survey shows that in 10 years of clinical use there have been no major changes in the susceptibility of staphylococci to the glycopeptides.     

Antibiotic susceptibility among Staphylococcus epidermidis isolated from prosthetic joint infections with special focus on rifampicin and variability of the rpoB gene

Staphylococcus epidermidis is the most important pathogen in infections related to implanted foreign materials, especially prosthetic joint infections (PJIs). The aim of this study was to investigate the antimicrobial activities of 16 antibiotics against S. epidermidis isolated from PJIs, with special focus on rifampicin and rpoB variability. Ninety-one per cent of the isolates were multiresistant (i.e. resistant to members of more than three classes of antibiotics). Thirty-nine per cent were resistant to rifampicin, associated with one or two single-nucleotide polymorphisms (SNPs) in rpoB . Using IsoSensitest agar with supplements, 61% were resistant to oxacillin, and using Mueller–Hinton II agar with supplement, 84% were resistant. Using the Etest, 58% were resistant to cefoxitin, and using the disk diffusion test, 91% were resistant. The mecA gene was detected in 85% of the isolates. Regarding recently available antibiotics, all isolates were susceptible to tigecycline and linezolid, and 97% were susceptible to daptomycin. In addition, two novel antibiotics, dalbavancin and ceftobiprole, were tested, although not yet available for routine use. The MIC₅₀ and MIC₉₀ values of these novel antibiotics were 0.032 and 0.047 mg/L and 0.5 and 1.5 mg/L, respectively. Among the other antibiotics, the rates of resistance varied between 0% (vancomycin) and 82% (trimethoprim–sulphamethoxazole). S. epidermidis strains causing PJIs often show multiresistance, including resistance to rifampicin, which is mainly caused by one or two SNPs. Some of the newer antimicrobial agents may provide alternatives for monotherapy or combination therapy with rifampicin. Detection of mecA is necessary before initiating treatment of infections due to S. epidermidis when it displays intermediate susceptibility to cefoxitin.     

Antibacterial effects of Eucalyptus globulus leaf extract on pathogenic bacteria isolated from specimens of patients with respiratory tract disorders

The antibacterial activity of Eucalyptus globulus leaf extract was determined for 56 isolates of Staphylococcus aureus , 25 isolates of Streptococcus pyogenes , 12 isolates of Streptococcus pneumoniae and seven isolates of Haemophilus influenzae obtained from 200 clinical specimens of patients with respiratory tract disorders. MIC₅₀s for these species were 64, 32, 16 and 16 mg/L, respectively; MIC₉₀s were 128, 64, 32 and 32 mg/L, respectively; and MBC s were 512, 128, 64 and 64 mg/L, respectively. These results suggest that further studies to clarify the possible therapeutic role of E. globulus leaf extract in the treatment of respiratory tract infection are warranted.     

Susceptibility testing by E-test and agar dilution of 30 strains of Legionella spp. isolated in Portugal

Objective: To evaluate the activity of erythromycin, roxithromycin, ciprofloxacin, doxycycline, sulfamethoxazole trimethoprim and rifampin against 32 strains of Legionella spp. under different testing conditions.
Method: Minimum inhibitory concentrations (MICs) were determined by the E-test (Ab Biodisk, Solna, Sweden) and agar dilution reference technique (National Committee for Clinical Laboratory Standards (NCCLS), 1990) on two different media, buffered charcoal yeast extract agar (BCYE-α) and buffered yeast extract agar (BYE-α), under 48- and 72-h incubation, without CO₂.
Results: All the antimicrobial agents were inhibited by BCYE-α agar. The MIC₉₀ values on BYE-α were lower than those on BCYE-α but the variation factor was not the same: ciprofloxacin and rifampin, followed by erythromycin, suffered the greatest inhibition by the charcoal in the culture medium. Except for ciprofloxacin and rifampin, the 72-h MIC₉₀ readings were always higher than the 48-h results whenever the agar dilution method was used. The E-test results showed slight variations with some, but not all, antibiotics. The most active agents against the 32 Legionella strains tested were rifampin and ciprofloxacin.
Conclusions: BCYE-α is not suitable for susceptibility testing of Legionella spp. The E-test method on BYE-α agar with 48-h incubation is recommended.