In Vitro Synergism Observed with Azithromycin, Clarithromycin, Minocycline, or Tigecycline in Association with Antifungal Agents against Pythium insidiosum

Abstract

We describe here the in vitro activities of azithromycin, clarithromycin, minocycline, or tigecycline alone and in combination with amphotericin B, itraconazole, terbinafine, voriconazole, anidulafungin, caspofungin, or micafungin against 30 isolates of the oomycete Pythium insidiosum. The assays were based on the CLSI M38-A2 technique and the checkerboard microdilution method. The main synergisms observed were through the combination of minocycline with amphotericin B (73.33%), itraconazole (70%), and micafungin (70%) and of clarithromycin with micafungin (73.33%). Pythiosis is a life-threatening disease caused by the oomycete Pythium insidiosum, which can cause infections in humans and in animals, such as horses, bovines, cats, dogs, and sheep. Clinically, the disease may manifest in cutaneous, gastrointestinal, vascular, and systemic forms and has been described in tropical and subtropical areas (1). The hyphae of P. insidiosum are morphologically similar to those of certain mucoraceous molds, but P. insidiosum is not a true fungus because it does not synthesize ergosterol, which is the target of most antifungal drugs. Despite this challenge, two cases of pythiosis in humans, one case of ocular pythiosis and one case of pleuropericarditis (2, 3), have been successfully treated using combination antifungal therapy. However, combination antifungal therapy has been ineffective in cases of vascular and disseminated human pythiosis (4).Previous studies have shown that the growth of P. insidiosum is inhibited in vitro by the glycylcycline, macrolide, and tetracycline classes of antibacterial drugs (5, 6). However, studies evaluating the antimicrobial combination of antibacterial and antifungal agents against P. insidiosum have not been performed. In this context, this study evaluated the in vitro combination of the antibacterial drugs azithromycin, clarithromycin, minocycline, or tigecycline with the antifungal drugs amphotericin B, itraconazole, voriconazole, terbinafine, anidulafungin, caspofungin, or micafungin against P. insidiosum.Twenty-eight P. insidiosum isolates obtained from Brazilian cases of equine pythiosis and the reference strains ATCC 58.637 and CBS 101555 were evaluated in this study. The identities of the clinical isolates were confirmed using PCR-based assays (7). The antibacterial drugs azithromycin (Pharma Nostra, Rio de Janeiro, Brazil), clarithromycin (Genix, Anápolis, Brazil), minocycline (Pharma Nostra), and tigecycline (Pfizer, New York, NY) and the antifungal drugs amphotericin B (Sigma-Aldrich, St. Louis, MO), itraconazole (Frangon do Brasil Farmacêutica Ltda., São Paulo, Brazil), voriconazole (Pfizer), terbinafine (Pharma Nostra), anidulafungin (Pfizer), caspofungin (Merck, Darmstadt, Germany), and micafungin (Astellas, Chuo, Japan) were obtained commercially and diluted in dimethyl sulfoxide or distilled water, as recommended, to generate stock solutions. The concentrations of the antimicrobial agents tested were 0.03 to 16 g/ml and 1 to 512 g/ml for the antibacterial and antifungal dr...

In Vitro and In Vivo Antimicrobial Activities of Minocycline in Combination with Azithromycin, Clarithromycin, or Tigecycline against Pythium insidiosum

“…The in vitro susceptibility results in our study were consistent with those of previous studies (7)(8)(9)(10). In vivo, the previous studies of induced pythiosis in rabbits demonstrated that caspofungin (1 mg/kg/day) (14), some combinations of terbinafine (125 mg/day) with itraconazole (5 mg/kg/day), fluvastatin (1 mg/kg/day), or caspofungin (1 mg/kg/day) (15), and a vortexed immunotherapy (30 mg at 14-day intervals) (12, 16) presented fungistatic activity, i.e., the progression of subcutaneous lesions was less than that for the untreated group for some therapies, but complete remission of the lesions was not the prevalent observation.…”

“…The susceptibilities of 28 Brazilian P. insidiosum strains isolated from equine pythiosis lesions (11) and two reference strains, P. insidiosum CBS 101555 and ATCC 58.637, were evaluated. A CLSI M38-A2-based in vitro susceptibility test and combination tests were performed as described previously (9). Azithromycin and minocycline (Pharma Nostra, Rio de Janeiro, Brazil), clarithromycin (Genix, Anápolis, Brazil), and tigecycline (Sigma-Aldrich, St. Louis, MO, USA) were purchased as standard antimicrobial powders.…”

“…Despite a few reports of successful therapy with antifungal drugs (2,3), many studies have demonstrated their ineffectiveness (1,(4)(5)(6). We previously described antibacterial compounds belonging to the glycylcycline, macrolide, and tetracycline classes (7)(8)(9)(10) as having potent in vitro activity against P. insidiosum. The present study investigated the in vitro and in vivo interactions among minocycline, azithromycin, clarithromycin, and tigecycline against P. insidiosum.…”

New Insights into the In Vitro Susceptibility of Pythium insidiosum

“…Interestingly, previous studies have shown that P. insidiosum is quite sensitive to antibacterials belonging to the classes of macrolides, tetracyclines, and glycylcyclines (7,8) and that its combination with antifungal drugs can result in synergistic interactions in vitro (9). However, the effects of antibacterial agents that inhibit protein synthesis on P. insidiosum have not been studied extensively.…”

In vitro synergistic combinations of pentamidine, polymyxin B, tigecycline and tobramycin with antifungal agents against Fusarium spp.

“…The antimicrobial agents were selected according to previous reports of synergism when such drugs were combined with antifungal agents against other fungal genera and the oomycete Pythium insidiosum (Afeltra et al, 2002;Chodosh et al, 2000;Jesus et al, 2014;Ku et al, 2010;St-Germain, 1990;Zhai et al, 2010). PNT has been used in prophylaxis of pneumonia caused by Pneumocystis jiroveci in recipients of allogeneic bone marrow transplants and patients with AIDS (Marras et al, 2002).…”