Activity of nadifloxacin and three other antimicrobial agents against Cutibacterium acnes isolated from patients with acne vulgaris

Conflict of interest

C Neumeister, M R Götz and U Schwantes are employees of Dr. Pfleger Arzneimittel GmbH. P Nenoff, D Koch and C Krüger are employees of the laboratory for medical microbiology, which was commissioned by Dr. Pfleger Arzneimittel GmbH with the isolation of C. acnes and susceptibility testing. R‐H Bödeker and C. Borelli have received honoraria for their advisory activities from Dr. Pfleger Arzneimittel GmbH.

Funding

This study was funded by Dr. Pfleger Arzneimittel GmbH, Bamberg, Germany.

Dear Sir,

Nadifloxacin^* (OPC‐7251) represents an established antimicrobial agent belonging to the quinolone group and was developed exclusively for topical administration ⁴ . Nadifloxacin is effective in treating a variety of bacterial skin infections and acne. ² , ³ , ⁵ , ⁷ , ⁸ , ⁹ Here, the in vitro activity of nadifloxacin was assessed and compared with those of erythromycin, clindamycin and tetracycline against Cutibacterium (C.) acnes, formerly Propionibacterium acnes, to gain a picture of the resistance situation in Germany. The study was approved by the Ethics Committee of the Bayerische Landesärztekammer (Munich, Germany; EC‐No.17088) and registered at Deutsches Register Klinische Studien (ID: DRKS00014231). Samples were collected between March and May 2018 at 45 sites. Each sample was obtained from a pustule of the facial area using a sterile sample collection swab and directly inserted in the transport tube medium (Amies medium, Sarstedt AG & Co. KG, Nümbrecht, Germany, order number 80.1361.500). Further information was acquired to characterize the patient population, acne score (Leeds revised Acne Grading System ⁶ ) and prior antibiotic treatment of acne. The VITEK 2 ANC Card System for Identification of Clinical Isolates of Anaerobic Bacteria (bioMérieux sa, Marcy‐l´Etoile, France, distributed by bioMérieux Deutschland GmbH, Nürtingen, Germany) was used. Reference standards of antimicrobial agents were purchased by Sigma‐Aldrich Chemie GmbH (Taufkirchen, Germany): nadifloxacin (Order no. SMB00375‐1G), clindamycin phosphate (Order no. PHR1021‐1G), erythromycin (Order no. E5389‐1G) and tetracycline hydrochloride (Order no. T7660‐5G). Nadifloxacin was dissolved in 0.1 N NaOH, erythromycin in ethanol 96% (Dr. K. Hollborn & Söhne GmbH & Co. KG, Leipzig, Germany), clindamycin and tetracycline in sterile distilled water.

Before susceptibility testing, the isolates were transferred on Columbia 5% SB (sheep blood) agar (BD, Heidelberg, Germany) to ensure purity and good growth. The antimicrobial susceptibility against the different drugs was determined by an agar dilution method using Gifu Anaerobic Medium agar modified ‘Nissui’ (Nissui Pharmaceutical Inc. Ltd., Tokyo, Japan; pH 7.3, distributed by HyServe GmbH & Co. KG, Uffing, Germany) and a bacterial cell inoculum of 10⁶ cfu per ml as it was previously described ⁵ . The plates were incubated at 37°C in anaerobic atmosphere using the Anaerocult system (Merck, Darmstadt, Germany) and analysed after 48 h incubation for anaerobic C. acnes.

Minimum inhibitory concentration for clindamycin was determined by E‐test (Liofilchem, Roseto degli Abruzzi, Italy) on Mueller‐Hinton agar plates (BD, Heidelberg, Germany), which covered a continuous concentration range from 0.016 to 256 µg/mL.

Two control organisms (C. acnes DSM 1897 and 108415, Braunschweig, Germany) were tested simultaneously (MIC [µg/mL]: nadifloxacin: 0.195; erythromycin: 0.049; tetracycline: 0.78; clindamycin: 0.016; [n = 2]).

A total of 73 strains of C. acnes from 115 samples were isolated. Of these 73 samples, 23 (31.5%) came from patients who had been pretreated with antibiotics. More than half of the patients (51.4%) were ≤21 years old (Table 1). Most of the patients suffered from mild to moderate acne, represented by a Leeds score of 6 for the 3rd quartile (75%) and 4 for the median (Table 1). ⁶

Table 1.

Patient age [years] and severity of acne at time of sample collection

Characteristics	N	Missing	Minimum	1st quartile	Median	3rd quartile	Maximum
Age [years]	72	1	10.0	18.0	21.0	26.0	64.0
Severity grade of acne*	73	0	1.0	3.0	4.0	6.0	11.0

^*According to Leeds revised acne grading system (O’Brien et al. ⁶ ).

None of the 73 strains revealed elevated minimum inhibitory concentration (MIC) levels against nadifloxacin or tetracycline. Table 2 shows the number of samples in which resistance was detected according to the applied clinical breakpoints. The highest percentage (15.07%, 11 strains) was shown for erythromycin, followed by clindamycin (4.11%, 3 strains).

Table 2.

Minimal inhibitory concentration (MIC) range, MIC₅₀, MIC₉₀ of antimicrobial agents against C. acnes; breakpoint resistance, percentage and no. of resistant strains

Antimicrobial agent	Test	MIC range (min to max) [µg/mL]	MIC50 [µg/mL]	MIC90 [µg/mL]	Breakpoint resistance [µg/mL]	% resistance (no. of strains)
Nadifloxacin	Agar dilution	0.097 to 0.390	0.195	0.195	≥4†	0.00 (0)
Clindamycin	E‐Test	0.023 to >256	0.047	0.750	≥8†, ‡, §, ¶	4.11 (3)
Erythromycin	Agar dilution	<0.024 to >200	0.024	200	≥2‡ ≥8†	For both breakpoints: 15.07 (11)
Tetracycline	Agar dilution	0.390 to 3.125	0.390	0.780	≥16‡, §	0.00 (0)

^†Alba et al. ¹ .

^‡Sardana et al. ⁸ .

^§Clinical and Laboratory Standards Institute (CLSI) Performance Standards for Antimicrobial Susceptibility Testing, 29th Edition, CLSI Supplement M100, Wayne PA: Pennsylvania; 2019.

^¶The European Committee on Antimicrobial Susceptibility Testing. Breakpoint tables for interpretation of MICs and zone diameters. Version 9.0, 2019. http://www.eucast.org.

As for erythromycin a significant number of resistances was found (n = 11), an investigation into possible influencing factors on occurrence of resistant strains was carried out by means of contingency tables. There is no evidence of an influence of age, severity of acne or antibiotic pretreatment on the occurrence of resistant strains.

In summary, nadifloxacin was found to be highly active against C. acnes isolated from patients with acne vulgaris. However, a considerable number of C. acnes strains showed in vitro resistance against erythromycin and clindamycin.