Biofilm formation by Rhodococcus equi and putative association with macrolide resistance

Abstract

RESUMO.-[Formação de biofilme por Rhodococcus equi e posspivel associação com resistência a macrolídeos.]Rhodococcus equi é um patógeno intracelular facultativo, o qual causa pneumonia piogranulosa severa em potros e lesões semelhantes à tuberculose em humanos. A sua capacidade de formar biofilme foi descrita em cepas humanas, isoladas a partir de doenças crônicas associadas a falhas de tratamento. Este estudo teve como objetivo verificar a formação de biofilme por 113 cepas de R. equi, isoladas a partir de amostras de equinos (clínicas e fecais), utilizando-se dois diferentes métodos (biofilme em cultura -com e sem adição de glicose -e microscopia de epifluorescência). Além disso, buscou-se determinar a eficácia da azitromicina, claritromicina e eritromicina sobre biofilme consolidado de R. equi. Verificou-se 80,5% (26/41) e 63% dos isolados (58/72) positivos para formação de biofilme, em amostras fecais e clínicas, respectivamente. A adição de glicose amentou a formação de biofilme em amostras fecais, mas não em amostras clínicas. Os antimicrobianos aqui testados não foram capazes de erradicar R. equi em biofilme consolidado, mesmo em concentrações elevadas. Este é o primeiro estudo a demonstrar a formação de biofilme por cepas de R. equi isoladas a partir de amostras de equinos. Os resultados indicam que os isolados de R. equi produtores de biofilme podem ser mais resistentes aos antimicrobianos avaliados. Estudos adicionais são necessários para testar essa hipótese. Brazil. E-mail: sabott20@gmail.com Rhodococcus equi is a facultative intracellular pathogen, which cause severe pyogranulomatous pneumonia in foals and tuberculosis-like lesions in humans. Its ability to form biofilm was described in strains isolated from chronic diseases associated to treatment failures in humans. This study aimed to verify the biofilm formation by 113 R. equi isolated from equine samples (clinical and fecal) using two different methods (biofilm--culturing with and without additional glucose and epifluorescence microscopy). We also aimed to determine the efficacy of azithromycin, clarithromycin and erythromycin on R. equi in established biofilm. We found 80.5% (26/41) and 63% (58/72) biofilm--positive isolates, in fecal and clinical samples, respectively. The additional glucose increased the biofilm formation by R. equi fecal samples, but not by clinical samples. The antimicrobials tested herein were not able to eradicate R. equi in biofilm even at higher concentrations. This is the first study showing the biofilm formation by R. equi isolated from equine samples. Our findings indicate that R. equi biofilm-producers may be more resistant to the antimicrobials evaluated. Further studies are warranted to test this hypothesis. Biofilm formation by

Phenotypic Characterization of Rhodococcus equi Biofilm Grown In Vitro and Inhibiting and Dissolving Activity of Azithromycin/Rifampicin Treatment

“…However, a statistically significant reduction of R. equi sessile aggregates emerged from our data, particularly when treated with AZM and AZM/RIF 2:1 over a prolonged period of time. These results are in good agreement with a previous study [20] which, however, employed different methodological approaches. On the other hand, the role of RIF against R. equi biofilm is controversial.…”

“…To date, poor literature reporting the capacity of R. equi to aggregate in a self-produced matrix has been available. R. equi seems to be able to produce biofilms, although some clinical isolates have been found to be non-biofilm producers at 24 h of incubation [20]. The kinetic crystal violet assay performed in this work demonstrated that 100% (40/40) of the rhodococci evaluated were biofilm producers with a suggested biofilm production time of at least 48 h, while incubation for longer than 72 h allowed more abundant biofilm development.…”

“…Bacteria were grown in CAMHB at 37 • C to reach a spectrophotometric growth absorbance of 0.15 at OD 600 . To assess the growth of rhodococcal biofilms, the microorganisms were diluted 1:50 in CAMHB, and 200 µL of bacterial suspension was seeded in each well of the 96-well plates and then incubated under static conditions at 37 • C for 24, 48, 72, and 96 h. The medium was not supplemented with glucose as it is unlikely to increase the biofilm formation of R. equi clinical isolates [20]. At the end of each incubation time, planktonic bacteria were removed and sessile aggregates were washed with 200 µL of PBS.…”

Moraxella bovis , Moraxella ovis and Moraxella bovoculi : biofilm formation and lysozyme activity

“…Recently, our research group demonstrated the formation of biofilms by Rhodococcus equi from clinical and non‐clinical samples of horses (Gressler et al . ). In addition, Peixoto et al .…”

“…; Gressler et al . ). However, studies involving biofilms in the micro‐organisms of veterinary interest are still scarce, including Moraxella genus.…”