Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Biofilm Development by Mycobacterium avium Complex Clinical Isolates. Effect of Clarithromycin in Ultrastructure

Version 1 : Received: 26 February 2024 / Approved: 27 February 2024 / Online: 27 February 2024 (12:20:05 CET)

A peer-reviewed article of this Preprint also exists.

Akir, A.; Senhaji-Kacha, A.; Muñoz-Egea, M.C.; Esteban, J.; Aguilera-Correa, J.J. Biofilm Development by Mycobacterium avium Complex Clinical Isolates: Effect of Clarithromycin in Ultrastructure. Antibiotics 2024, 13, 263. Akir, A.; Senhaji-Kacha, A.; Muñoz-Egea, M.C.; Esteban, J.; Aguilera-Correa, J.J. Biofilm Development by Mycobacterium avium Complex Clinical Isolates: Effect of Clarithromycin in Ultrastructure. Antibiotics 2024, 13, 263.

Abstract

Background: Mycobacterium avium complex includes the commonest nontuberculous mycobacteria associated with human infections. These infections have been associated with the production of biofilms in many cases, but there are only a few studies about biofilms produced by the species included in this group. Methods: Three collection strains (M. avium ATCC25291, M. intracellulare ATCC13950, and M. chimaera DSM756), three clinically significant strains (647, 657, and 655), and three clinically nonsignificant ones (717, 505, and 575) of each species were included. The clinical significance of the clinical isolates was established according to the internationally accepted criteria. Biofilm ultrastructure was studied by Confocal-Laser Scanning Microscopy using Backlight Live dead and Red Nile stains. Viability, covered surface, height and relative autofluorescence were measured in several images/strain. The effect of clarithromycin was studied the technique described by Muñoz-Egea et al with modifications regarding incubation time. The study included clarithromycin in the culture medium at a concentration achievable in the lungs (11.3 mg/L), using one row of wells as the control without antibiotic. Bacterial viability inside the biofilm is expressed as a percentage of viable cells. The differences between the different parameters of biofilm ultrastructure were analysed using Kruskal-Wallis test. The correlation between bacterial viability in the biofilm and treatment time was evaluated using Spearman's rank correlation coefficient (ρ). Results: The strains showed differences between them with all the studied parameters, but neither a species-specific pattern nor a clinical significance-specific patter were detected. For the effect of clarithromycin, the viability of the bacteria contained in the biofilm was inversely proportional to the exposure time of the biofilm (ρ > -0.3; p-value < 0.05), excluding two M. chimaera strains (M. chimaera DSM756 and 575), which showed a weak positive correlation with treatment time (0.2 < ρ < 0.39; p-value < 0.05). Curiously, despite a clarithromycin treatment of 216 hours, the percentage of biofilm viability of the strains evaluated here was not less than 40% at best (M. avium 717). Conclusions: All the M. avium complex studied strains are able to form biofilm in vitro, but ultrastructural characteristics between them suggest that these are strain-specific characteristics unrelated with neither with the species nor with the clinical significance. The clarithromycin effect on MAC species is biofilm age/time of treatment-dependent and appears to be strain-specific while being independent of the clinical significance of the strain.

Keywords

Mycobacterium avium; Mycobacterium intracellulare; Mycobacterium chimaera; biofilm; clarithromycin; treatment; ultrastructure

Subject

Medicine and Pharmacology, Epidemiology and Infectious Diseases

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