MIC Values of Iclaprim and Lefamulin against Mycobacterium abscessus Complex

LETTER

Infections with rapid-growing mycobacteria (RGM) are often difficult to treat. The most common RGM identified as a cause of clinical infection are the Mycobacterium abscessus complex (MabsC), which is comprised of three subspecies: Mycobacterium abscessus subsp. abscessus, Mycobacterium abscessus subsp. massiliense, and Mycobacterium abscessus subsp. bolletii. Unlike for Mycobacterium tuberculosis infections and other slow-growing nontuberculous mycobacteria (NTM) infections, there is no standard regimen that is recommended for treatment. The general recommendation is to use a macrolide- and aminoglycoside-based regimen in combination with other antibiotics with demonstrable in vitro activity. However, MabsC are highly resistant against various drug classes (1), resulting in limited antibiotic options. Exploration of activity of newer drugs is useful in identifying potential antibiotic treatment options.

Lefamulin is a pleuromutilin drug for treatment of community-acquired pneumonia (CAP) (2). In vitro activity has been demonstrated against causes of typical and atypical CAP but with limited activity against some Gram-negative organisms, including Enterobacterales. Iclaprim is a dihydrofolate-reductase inhibitor with activity against Gram-positive organisms (3). MIC against Gram-negative organisms was higher than that against Gram-positive organisms, but synergy was seen in combination with sulfamethoxazole (4). MabsC infections, like other mycobacterial infections, are associated with formation of granulomas. They are generally located intracellularly, which influences pharmacokinetic/pharmacodynamic parameters. Iclaprim and lefamulin were selected for testing as both have been reported to have intracellular drug concentrations higher than their serum levels (5, 6).

Broth microdilution was performed as per Clinical and Laboratory Standards Institute (CLSI) standards to test the in vitro activity of iclaprim and lefamulin against clinical isolates of M. abscessus subsp. abscessus (n = 30), M. abscessus subsp. massiliense (n = 30), and M. abscessus subsp. bolletii (n = 3). Iclaprim (HY-101479) and lefamulin (HY-16908A) antibiotic powders were purchased from MedChemExpress (Monmouth Junction, NJ, USA). The testing range was 0.03 to 32 mg/liter for both drugs, with reading performed at day 3. Quality control was performed using Staphylococcus aureus ATCC 29213 (7). The MIC for S. aureus ATCC 29213 was 0.125 mg/liter for both iclaprim and lefamulin. Testing was also performed for M. abscessus ATCC 19977^T. The clinical isolates chosen were selected from a larger collection of isolates which had been whole-genome sequenced on Illumina NovaSeq6000 sequencing platform (Illumina Inc., San Diego, CA, USA) and assembled. An average sequencing depth of 150× was achieved for the genomes. Phylogenetic analysis was used to determine subspecies identity. Multilocus sequence typing (MLST) was performed using the scheme for Mycobacterium abscessus (https://github.com/phac-nml/mab_mabscessus) (8). Isolates with differing sequence types were selected in order to represent genomically diverse isolates (Data Set S1).

The MIC results of the clinical isolates are presented in Table 1. MIC₅₀ and MIC₉₀ were also determined for M. abscessus subsp. abscessus and M. abscessus subsp. massiliense but not for M. abscessus subsp. bolletii due to the limited number of isolates. M. abscessus ATCC 19977^T had an MIC of >32 mg/liter for both lefamulin and iclaprim at day 3 of incubation. Multilocus sequencing data, erm(41) sequevar, and full MIC data are presented in the supplemental material.

TABLE 1.

MIC of lefamulin and iclaprim against members of the Mycobacterium abscessus complex

Organism (n)	MIC (mg/liter)					MIC50	MIC90
	4	8	16	32	>32
Lefamulin
M. abscessus subsp. abscessus (30)	4	3	11	1	11	16	>32
M. abscessus subsp. massiliense (30)	0	4	5	15	6	32	>32
M. abscessus subsp. bolletii (3)	0	0	3	0	0
Iclaprim
M. abscessus subsp. abscessus (30)	0	0	8	7	15	32	>32
M. abscessus subsp. massiliense (30)	0	1	1	6	22	>32	>32
M. abscessus subsp. bolletii (3)	0	1	2	0	0

Although there are currently no interpretive breakpoints available for lefamulin and iclaprim against MabsC, MICs are higher than epidemiological cutoff values (ECOFF) reported in other organisms, suggesting limited in vitro susceptibility when tested singly (2–4, 6, 9, 10). Lefamulin ECOFF of 0.25 mg/liter is reported for Staphylococcus spp. Low MIC₉₀ for S. aureus, methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant Staphylococcus aureus (MRSA), Streptococcus pyogenes, S. agalactiae, S. anginosus, S. constellatus, S. dysgalactiae, and S. intermedius have also been reported (0.12, 0.12, 0.5, 0.03, 0.5, ≤0.004, ≤0.004, 0.12, and 0.008 mg/liter, respectively) (10). However, MabsC infections are usually treated with multiple antibiotics. Combinations of antibiotics may have synergistic activity contributing to their effects. The MIC range of MabsC to iclaprim is similar to that of trimethoprim-sulfamethoxazole (1), which is a combination of a dihydrofolate-reductase inhibitor (trimethoprim) and a sulfonamide (sulfamethoxazole). The activity of trimethoprim-sulfamethoxazole in mycobacteria has been attributed to the sulfamethoxazole component (11). The combination of iclaprim with sulfamethoxazole may result in more favorable synergy, both acting on the metabolic pathway of folic acid. Future studies may consider testing lefamulin and iclaprim in combination with other antibiotics to investigate any potential synergistic effects.