Estudio de la actividad antibiótica y desinfectante frente a legionella spp. A nivel planctónico e intracelular

Resumen

Infections caused by Legionella spp. are still a public health problem. There is no standardized method to determine the sensitivity of this bacterium to antimicrobials. Routine methods can provide information about the sensitivity on planktonic growth but no about the intracellular growth of this bacterium, the intracellular activity of the antibiotic, and the ability of the antibiotic to penetrate polymorphonuclear cells; so the results obtained in vitro may not correspond with the in vivo killing effect. On the other hand, the environmental control of Legionella spp. in artificial water systems is commonly performed by chlorination due to its simplicity and cost-benefit. However, chlorine is a very corrosive agent, capable of causing significant damage to the facilities. Therefore, new and less aggressive biocides, such as non-oxidants agents, have been and are being developed. The biocide activity evaluation against Legionella is regulated in Europe by UNE-EN 13623. This regulation does not take into account that Legionella can be present in artificial water systems within amoeba or forming part of biofilms, so the amounts of biocide recommended to achieve a bactericidal effect may not be sufficient in these forms of growth. Rifampicin showed better extracellular activity against Legionella spp. than clarithromycin, azithromycin, telithromycin, levofloxacin, moxifloxacin, tigecycline, doxycycline, trimethoprim-sulfamethoxazole and ampicillin, by both diffusion technique gradient of antibiotic concentration in BCYE-α culture medium (E-test) and broth microdilution in BYE-α culture medium. On the other hand, rifampicin, levofloxacin and moxifloxacin presented better intracellular activity against Legionella spp. than clarithromycin, azithromycin, telithromycin, tigecycline, doxycycline, trimethoprim-sulfamethoxazole and ampicillin. Moreover, the intracellular activity of clarithromycin, levofloxacin and rifampicin determined in J-774 macrophage cell line, can be predicted up to 20.20% by a mathematical relation from the extracellular results obtained by E-test. From the evaluated biocides, calcium hypochlorite was the oxidant biocide with the highest planktonic and intra-amoeba bactericidal activity against L. pneumophila serogroup 1 strains, and 2,2-dibromo-3-nitrilopropionamide was the non-oxidant biocide with the highest activity. Bactericidal concentrations obtained at planktonic level for all the tested compounds (sodium hypochlorite, calcium hypochlorite, bromochloro-5,5-dimethylimidazolidine, trichloroisocyanuric acid, peracetic acid, 2,2-dibromo-3-nitrilopropionamide, didecyldimethylammonium, 5-chloro-2-methyl-4-isothiazolin-3-one/2-methyl-4-isothiazolin-3-one and tetrakis (hydroxymethyl) phosphonium sulfate) were significantly lower than the doses recommended for use. This situation was also observed when analyzed the intra-amoeba bactericidal concentration of sodium hypochlorite, peracetic acid and 2,2-dibromo-3-nitrilopropionamide and tetrakis (hydroxymethyl) phosphonium sulfate, therefore, water facilities may be exposed to higher concentrations than necessary with the increased risk of pipes rupture. Finally, planktonic bactericidal concentrations obtained for calcium hypochlorite, bromochloro-5,5-dione-2,4-dimetilmidazolidina, and trichloroisocyanuric acid are significantly lower than those obtained in the intra-amoeba model, and, hence, evaluation of new biocides based only on extracellular methods, as current regulation UNE-EN 13623 says, may be insufficient to get reliable data of bactericidal effect in water systems.