In Vitro Effects of Amphotericin B on Growth and Ultrastructure of the Amoeboflagellates Naegleria gruberi and Naegleria fowleri

Abstract

In vitro effects of the polvene antibiotic amphotericin B (AmB) on growth, viability, and ultrastructure of amoeboflagellates of the genus Naegleria were examined. The strains studied were the nonpathogenic Naegleria gruberi EGB and the Carter and TY strains of the pathogenic Naegleria fowleri. AmB was amoebicidal at all concentrations used (0.25, 0.50, and 1.0 ,ug/ml) when the drug was added to cultures in lag phase, as determined by viability testing, but was mainly inhibitory when added to log-phase cultures. The drug produced ultrastructural modifications at all concentrations (0.05 to 1.0 gg/ml). These changes included distortion of nuclear shape, increase in cytoplasmic membranes (both rough and smooth endoplasmic reticulum), decrease in number of food vacuoles, absence of pseudopod formation, mitochondrial abnormalities, increase in autophagic vacuoles, and blebbing of the plasma membrane. These alterations of amoebic ultrastructure became more pronounced with increased time in AmB and with increase in AmB concentration in the growth medium.

In Vitro Susceptibilities of Naegleria fowleri Strain HB-1 to Selected Antimicrobial Agents, Singly and in Combination

“…Antiamoebic effects of drugs that have been previously noted include inhibition of growth (5,8,9), inhibition of amoebic motility (5,8,9), and cytotoxic changes observed by light or electron microscopy (17). Definitions of the MIC of a drug have varied in previous studies (5,8,9).…”

“…There was no indication ofsynergy when AMB was combined with miconazole or when the methyl ester of AMB was combined with either minocycline, tetracycline, or miconazole. DISCUSSION AMB (5,8,9,17), tetracycline (19), rifampin (19), clotrimazole (9,12), and miconazole (9,21) have all shown activity against Naegleria spp. The majority of the assays of these reagents have been carried out either on agar plates or in axenic media, both with and without bacteria as an added nutrient.…”

“…Many different antimicrobial agents (and treatment regimens) have been used to treat PAM, but the most promising appears to be the antifungal polyene, AMB. Both in vitro amoebicidal activity (5,8,9,17) …”

In Vitro Susceptibility of Pathogenic Naegleria and Acanthamoeba Species to a Variety of Therapeutic Agents

“…Schuster and Rechthand have recently shown that amphotericin B is amebicidal when used during the lag phase of amoebic growth but mainly inhibitory when used during the log phase of growth (24). Ultrastructural changes that they observed were distortions of nuclear shape, increase in cytoplasmic membranes (both rough and smooth endoplasmic reticulum), decrease in food vacuoles, absence of pseudopods, abnormal mitochondria, increase in autophagic vacuoles, and blebbing of the plasma membrane.…”

“…The MMIC was defined as the lowest concentration of drug at which no active pseudopodial activity was seen as compared with controls. After reincubation of the chamber slide for 24 h and again for a total of 48 h, the minimal inhibitory concentration (MIC) of growth was determined. The MIC was defined as the lowest concentration of drug at which growth was 50% less than that of the control, as determined visually on the inverted microscope (Fig.…”