Abstract
We compared the kill-curve activity of tedizolid and linezolid at clinically relevant (total or free plasma, lung) concentrations against methicillin-resistant Staphylococcus aureus (MRSA) and penicillin-resistant Streptococcus pneumoniae (PRSP) isolated from Chinese patients. Tedizolid had greater in vitro potency than linezolid against staphylococci, streptococci and enterococci species (tedizolid minimum inhibitory concentration (MIC) range: ≤ 0.016–0.5 µg/mL; linezolid MIC range: 0.25–2 µg/mL). In kill-curve experiments, growth of MRSA was inhibited at tedizolid concentration of 0.6 µg/mL (i.e. 4.8 × MIC; MIC = 0.125 µg/mL) and linezolid concentration of 2 µg/mL (2× MIC; MIC = 1 µg/mL). Against PRSP, tedizolid at a concentration of 0.25 µg/mL (representing its MIC) was bacteriostatic, but exerted a bactericidal effect at higher concentrations. Results were similar for linezolid, however, even at 21 µg/mL, a small proportion of organisms survived beyond 24 h. The results demonstrated the potency of tedizolid against clinical strains of Gram-positive pathogens supporting its use as a suitable alternative to linezolid in Chinese patients.
Introduction
The spread and escalation of antimicrobial resistance represent a considerable threat for public health globally,1 including China.2 Among Gram-positive cocci, high prevalence of methicillin-resistant Staphylococcus aureus (MRSA) and/or S. epidermidis (MRSE) was reported among Chinese patients with burn wound infection,3 surgical site infection,4 bloodstream infections,5 and nosocomial pneumonia.6 Furthermore, the rapidly rising rate of penicillin-resistant Streptococcus pneumoniae (PRSP) across the Asia Pacific region,7 and emergence of linezolid-resistant coagulase-negative staphylococci or enterococci, vancomycin-resistant enterococci (VRE) and vancomycin-intermediate S. aureus (VISA) in many Asian countries, including China,8 indicate a serious threat for successful patient care.
Antibiotics demonstrate distinct kinetics and dynamics in their in vitro or in vivo activity against bacterial species.9 The minimum inhibitory concentration (MIC) of antibiotics is a key parameter to characterize the in vitro potency against pathogenic species in local or global surveillance programmes.9–11 When dosing regimen optimization is required for the therapeutic use of an antibiotic, the MIC value is also used to characterize the in vivo pharmacodynamic profile.9,10,11 However, limited information is revealed by the MIC value on the time course of inhibition of bacterial growth and whether any post-antibiotic inhibitory effect continues following cessation of drug exposure.9 Therefore, in vitro time-kill experiments at increasing antibiotic concentrations are required to characterize antibiotics over a time period of 24–72 h.9 These in vitro experiments can demonstrate bacteriostatic and bactericidal activities of antibiotics, which often coexist for the same antibiotic against different species.12
The currently approved oxazolidinones (linezolid, tedizolid) exhibit in vitro bacteriostatic activities against staphylococci and enterococci, and bactericidal activities against streptococcal species.11,12–17 Of note, a greater intracellular accumulation18 and at least 4-times greater in vitro potency of tedizolid compared with linezolid have been demonstrated in previous studies.19,20 Furthermore, tedizolid has been described with a more favourable pharmacokinetic profile than linezolid in both Caucasian21,22 and Asian healthy subjects.23,24 Tedizolid has recently been approved in the USA, Canada, Europe, Japan and other countries for the treatment of acute bacterial skin and skin structure infections (ABSSSIs); its efficacy and safety have also been demonstrated in Chinese patients with ABSSSI.25
The objective of this study was to evaluate 1) the susceptibility of selected clinical Gram-positive pathogens and 2) the in vitro time-kill activity of tedizolid and linezolid against MRSA and PRSP, which were isolated from patients in Chinese hospitals, at concentrations relevant to Chinese subjects based on previously established pharmacokinetic data.
Materials and methods
Determination of antimicrobial susceptibility
Between 2013 and 2016, a total of 135 bacterial strains were isolated from blood, urine, sputum and secretion samples from local sites in China. The MICs of test isolates and quality control strains were measured by broth microdilution method outlined by the Clinical and Laboratory Standards Institute.
The MIC was defined as the lowest concentration of antimicrobial agent that inhibited growth of the organism in the microdilution wells, after confirming aerobic growth in the antibiotic-free broth as a positive growth control. Test results were validated by confirmation of the MIC values determined for quality control strains being within the respective MIC range detailed by CLSI Guideline M100-S26. The quality control strains used were American Type Culture Collection (ATCC) quality control strains S. aureus ATCC29213 and S. pneumoniae ATCC49619.
Assessment of in vitro time-kill activity
Time-kill curves for tedizolid and linezolid were determined for selected clinical strains recently isolated from patients with ABSSSI and respiratory tract infections in China: MRSA 15B183, and PRSP 15L086, using the broth culture and agar plate count method. A starting inoculum (supplemented with 5% horse serum as appropriate) of approximately 1 × 106 colony forming unit (CFU)/mL was used. Tedizolid and linezolid were added to liquid cultures prepared in three series, each containing different clinically relevant drug levels based on those achieved by the respective approved clinical doses (200 mg once daily for tedizolid; 600 mg twice daily for linezolid). Tedizolid concentrations tested were 0.6 µg/mL (estimated free [i.e. unbound] concentration in plasma), 3.2 µg/mL (total concentration in plasma) and 9.05 µg/mL (total concentration in lung epithelial lung fluid [ELF]).23,26 Linezolid concentrations tested were 4.2 µg/mL (minimum free concentration in plasma), 14.7 µg/mL (maximum free concentration in plasma), 6.0 µg/mL (minimum total concentration in plasma) and 21.0 µg/mL (maximum total concentration in plasma and ELF).27–29 In addition, concentrations of 1×, 2× and 4× MIC were also tested for each antibiotic. After inoculation of the antibiotic-containing liquid media, culture tubes were incubated at 35 °C. Duplicate samples were removed immediately after inoculation (baseline) and 1, 3, 6, 8 and 24 h after inoculation, diluted (1/10) in appropriate media, plated on Columbia agar containing 5% sheep blood and incubated at 35 °C for 18 h. Colonies were then counted and mean Δlog10 CFU/mL was calculated.
Results
Antimicrobial susceptibility testing
The MICs for tedizolid against S. aureus were 0.125–0.25 µg/mL irrespective of the test isolate’s sensitivity to methicillin (Table 1). Overall, MIC values for linezolid against S. aureus were around eightfold higher than for tedizolid, with values between 1 and 2 µg/mL. Both antibiotics demonstrated greater activity against S. epidermidis compared with S. aureus, with modal MICs of 0.125 µg/mL for tedizolid and 1 µg/mL for linezolid and similar potency against methicillin-sensitive and -resistant S. epidermis strains.
Table 1. MIC values of test bacterial strains
Tedizolid demonstrated high activity against vancomycin-sensitive Enterococcus spp., particularly E. faecium, with MIC values typically less than 0.016 µg/mL. This is around > 125-fold lower than concentrations required for linezolid to successfully inhibit microbial growth (Table 1). The potency of both tedizolid and linezolid against E. faecium was not reduced by the presence of vancomycin resistance.
The activity of tedizolid against Streptococcus species was comparable to activity against staphylococci, with MICs of 0.25–0.5 µg/mL being demonstrated (Table 1).
Assessment of MIC for strains used for in vitro time-kill curves
The MICs of the two clinical isolates used for time-kill experiments (S. aureus and S. pneumoniae) ranged from 0.125 to 0.25 µg/mL for tedizolid, and from 0.5 to 1 µg/mL for linezolid. The tedizolid MIC was eightfold lower for MRSA 15B183 compared with linezolid (0.125 µg/mL vs 1 µg/mL, respectively) and twofold lower for PRSP 15L086 (0.25 µg/mL vs 0.5 µg/mL). MICs for control ATCC strains were 0.25 µg/mL for tedizolid, and ranged from 1 µg/mL (ATCC49619) to 2 µg/mL (ATCC 29213) for linezolid. For ATCC reference strains, tedizolid MIC was eightfold lower for S. aureus and fourfold lower for S. pneumoniae compared with linezolid.
Table 2. Growth of control and test bacterial strains. Data shown mean log10 CFU/mL at 3 h and 24 h after administration of antibiotics at clinically relevant concentrations
Assessment of in vitro time-kill curves
The growth of reference S. aureus strain ATCC29213 was inhibited at a tedizolid concentration of 0.6 µg/mL (2.4× MIC), but not at 0.25 µg/mL (i.e. the MIC for this strain) (Figure 1(A)). Similarly, linezolid was not effective at 2 µg/mL (i.e. the MIC for this strain), the growth was inhibited only at higher concentrations (Figure 1(C)).
Published online:
19 June 2019Figure 1. Kill curves for tedizolid and linezolid against S. aureus reference strain ATCC29213 and methicillin-resistant S. aureus clinical strain 15B183. CFU: colony-forming unit.
![]({"type":"image","src":"/na101/home/literatum/publisher/tandf/journals/content/yjoc20/2019/yjoc20.v031.i06/1120009x.2019.1623968/20190917/images/medium/yjoc_a_1623968_f0001_b.jpg"})
Figure 1. Kill curves for tedizolid and linezolid against S. aureus reference strain ATCC29213 and methicillin-resistant S. aureus clinical strain 15B183. CFU: colony-forming unit.
For the MRSA clinical isolate 15B183, tedizolid concentrations of 0.6 µg/mL (i.e. 4.8× MIC) were required to inhibit bacterial growth (Figure 1(B)). In contrast, linezolid concentrations of 2 µg/mL (2× MIC) were required to inhibit the growth of the MRSA clinical isolate (Figure 1(D)).
All higher tested concentrations of tedizolid or linezolid exerted bacteriostatic activity during the observational period against either reference or clinical S. aureus strain.
For the reference strain of S. pneumoniae ATCC49619, tedizolid concentrations of 0.25 µg/mL (i.e. the MIC for this strain) displayed bacteriostatic activity (Figure 2(A)). At concentrations of 0.6 µg/mL and higher, tedizolid demonstrated bactericidal activity, reducing the viable bacterial population density from 3.5 × 105 CFU/mL to < 10 CFU/mL over a 24-h period (Figure 2(A); Table 2). The minimum inhibitory concentration of linezolid (i.e. 1 µg/mL) demonstrated bacteriostatic activity, with a horizontal time-kill plot, however, a bactericidal effect on growth was demonstrated at higher concentrations (Figure 2(C)).
Published online:
19 June 2019Figure 2. Kill curves for tedizolid and linezolid against S. pneumoniae reference strain ATCC49619 and penicillin-resistant S. pneumoniae clinical strain 15L086. CFU: colony-forming unit.
![]({"type":"image","src":"/na101/home/literatum/publisher/tandf/journals/content/yjoc20/2019/yjoc20.v031.i06/1120009x.2019.1623968/20190917/images/medium/yjoc_a_1623968_f0002_b.jpg"})
Figure 2. Kill curves for tedizolid and linezolid against S. pneumoniae reference strain ATCC49619 and penicillin-resistant S. pneumoniae clinical strain 15L086. CFU: colony-forming unit.
For the PRSP clinical isolate 15L086, tedizolid concentrations of 0.25 µg/mL (i.e. the MIC for this strain) were bacteriostatic, but exerted a bactericidal effect at higher doses (Figure 2(B)). At concentrations of 12.8× MIC (3.2 µg/mL) tedizolid reduced bacterial density from baseline 7.5 × 104 CFU/mL to below the detection level (< 10 CFU/mL) over a 24-h period (Table 2). Similar results were observed for linezolid, however, even at 21 µg/mL, the highest concentration tested, a small proportion of viable organisms survived beyond 24 h (Figure 2(D)).
Discussion
The results of this study demonstrated the greater in vitro potency as well as the time-kill activity of tedizolid compared with linezolid against clinically relevant drug-resistant strains at concentrations which reflect exposures in patients following once daily dosing of tedizolid phosphate 200 mg or twice daily dosing of linezolid 600 mg.
A number of single-centre or large scale surveillance studies conducted in different regions demonstrated tedizolid to be at least fourfold more potent than linezolid against Gram-positive species isolated from patients with nosocomial pneumonia, skin and soft tissue infections, or bloodstream infections, including multidrug-resistant S. pneumoniae.19,20
The in vitro time-kill experiments for clinical Chinese strains of MRSA and PRSP, which used clinically relevant concentrations of both drugs representing the range of those in plasma and lung tissue,23,26–28 support the results of previous pre-clinical studies. The dosing recommendations were evaluated by Andes et al. (2002) for linezolid (i.e. 600 mg twice daily) and by Louie et al. (2011) for tedizolid (200 mg once daily), and both studies identified the area under the concentration-time curve to MIC (AUC/MIC) ratio as the PD driver for staphylococcal and streptococcal infections.10,11 The previous murine models of thigh infection or pneumonia established that both tedizolid and linezolid exhibit bacteriostatic activities against S. aureus regardless of methicillin resistance status13–15,17 and bactericidal activities against S. pneumoniae, regardless of its penicillin susceptibility.16 Although some discrepancy in the magnitude of the reduction of log CFU exists between studies which may be due to variation in the bacterial and/or mouse strains used or the initial bacterial load.11,13,17 Additionally, a time-dependence in the time-kill activity has been shown in the study by Keel et al. (2012) and by Xiao et al. (2018), which suggest that tedizolid might have a greater anti-staphylococcal activity after exposure at 72 h than at 24 h.13,17
The concentrations applied in this in vitro study represent those achieved with 200 mg once daily administration of tedizolid phosphate taking into account a protein binding of ∼87–90% for tedizolid.26,30 Of note, protein binding is ∼30% for linezolid.29 Bioavailability of tedizolid is high in Asian (i.e. Chinese and Korean) healthy subjects (i.e. 85–95%, respectively), which allows for equivalent dosing via intravenous or oral administration.23,24 The high degree of penetration of tedizolid into skin and soft tissue has previously been established in healthy human subjects.30 Both linezolid and tedizolid has a high accumulation ratio in epithelial lung fluid and alveolar macrophages.26,27
In conclusion, this study demonstrates the increased potency of tedizolid in comparison with linezolid against clinically relevant species of Gram-positive bacteria, including drug-resistant strains. At clinically relevant concentrations, tedizolid exerted a greater bactericidal effect in vitro than linezolid against a Chinese clinical PRSP strain and exerted a bacteriostatic effect against Chinese isolates of MRSA further supporting that tedizolid may be a suitable alternative to linezolid for the treatment of ABSSSI, nosocomial pneumonia and bloodstream infections caused by Gram-positive bacterial pathogens. The efficacy and safety of tedizolid have been established in Chinese subjects with ABSSSI;25 however, further clinical studies are required to prove its efficacy in other indications prior to use in patients.
Notes on contributors
Wang S participates in surveillance studies on clinical isolates.
Li Y has conducted long-term research on various antibacterial agents and participates in Chinese clinical surveillance studies.
Xue F participates in clinical surveillance studies.
Liu J participates in surveillance studies on clinical isolates.
Yang W participates in surveillance studies on clinical isolates.
Zhang J participates in surveillance studies on clinical isolates.
Glenschek-Sieberth M has a role in pharmaceutical pre-clinical development of antibacterial agents.
Lyu Y has conducted long-term research on various antibacterial agents and participates in Chinese clinical surveillance studies.
Data availability
The data that support the findings of this study are available from the corresponding author, [Yuan Lyu], upon reasonable request.
| Bacterial species and strain | Tedizolid (µg/mL) | Linezolid (µg/mL) | ||
|---|---|---|---|---|
| MIC90 | Range | MIC90 | Range | |
| MRSA (n = 10) | 0.25 | 0.125–0.25 | 2 | 1–2 |
| MSSA (n = 10) | 0.5 | 0.125–0.5 | 2 | 1–2 |
| MR S. epidermidis (n = 10) | 0.25 | 0.125–0.25 | 1 | 0.5–1 |
| MS S. epidermidis (n = 10) | 0.25 | 0.125–0.25 | 1 | 0.5–1 |
| S. haemolyticus (n = 10) | 0.125 | 0.125–0.25 | 1 | 1 |
| S. lugdunensis (n = 10) | 0.125 | 0.063–0.25 | 1 | 1 |
| VS E. faecalis (n = 10) | 0.5 | ≤0.016–0.5 | 2 | 0.5–2 |
| VS E. faecium (n = 10) | ≤0.016 | ≤0.016 | 2 | 2 |
| VR E. faecium (n = 10) | ≤0.016 | ≤0.016–0.5 | 2 | 1–2 |
| S. pneumoniae (n = 10) | 0.25 | 0.25 | 0.5 | 0.5 |
| S. pyogenes (n = 10) | 0.5 | 0.25–0.5 | 1 | 1–2 |
| S. agalactiae (n = 10) | 0.5 | 0.5 | 1 | 1 |
| S. anginosus group spp. (n = 15) | 0.25 | 0.031–0.25 | 1 | 0.25–1 |
MIC: minimum inhibitory concentration; MRSA: methicillin-resistant Staphylococcus aureus; MRSE: methicillin-resistant Staphylococcus epidermidis; MSSA: methicillin-sensitive Staphylococcus aureus; MSSE: methicillin-sensitive Staphylococcus epidermidis; VR: vancomycin-resistant; VS: vancomycin-susceptible.
| Bacterial species and strain | Time (h) | Tedizolid | Linezolid | ||
|---|---|---|---|---|---|
| 3.2 (µg/mL) | 9.05 (µg/mL) | 6.0 (µg/mL) | 21.0 (µg/mL) | ||
| S. aureus ATCC29213 | 3 | 5.613 | 5.699 | 5.607 | 5.484 |
| 24 | 4.286 | 4.246 | 4.170 | 4.083 | |
| S. aureus 15B183 (MRSA) | 3 | 5.633 | 5.703 | 5.484 | 5.663 |
| 24 | 4.155 | 4.114 | 4.398 | 4.114 | |
| S. pneumoniae ATCC49619 | 3 | 4.724 | 4.602 | 4.748 | 4.602 |
| 24 | ND | ND | ND | ND | |
| S. pneumoniae 15L086 (PRSP) | 3 | 4.623 | 4.613 | 4.618 | 4.477 |
| 24 | 0.699 | ND | 1.097 | 0.477 | |
CFU: colony-forming unit; MRSA: methicillin-resistant Staphylococcus aureus; ND: not detectable; PRSP: penicillin-resistant Streptococcus pneumoniae.
Disclosure statement
MGS is an employee of Bayer. The institution of SW, Y Li, FX, JL, WY, JZ and Y Lyu received research grant from Bayer AG, Germany. SW, Y Li, FX, JL, WY, JZ have no other conflict of interest. Y Lyu received honoraria from Bayer AG, Germany for participation in advisory board meetings.