Killing of diverse eye pathogens (Acanthamoeba spp., Fusarium solani, and Chlamydia trachomatis) with alcohols

Abstract

BackgroundBlindness is caused by eye pathogens that include a free-living protist (Acanthamoeba castellanii, A. byersi, and/or other Acanthamoeba spp.), a fungus (Fusarium solani), and a bacterium (Chlamydia trachomatis). Hand-eye contact is likely a contributor to the spread of these pathogens, and so hand washing with soap and water or alcohol–based hand sanitizers (when water is not available) might reduce their transmission. Recently we showed that ethanol and isopropanol in concentrations present in hand sanitizers kill walled cysts of Giardia and Entamoeba, causes of diarrhea and dysentery, respectively. The goal here was to determine whether these alcohols might kill infectious forms of representative eye pathogens (trophozoites and cysts of Acanthamoeba, conidia of F. solani, or elementary bodies of C. trachomatis).Methodology/Principal findingsWe found that treatment with 63% ethanol or 63% isopropanol kills >99% of Acanthamoeba trophozoites after 30 sec exposure, as shown by labeling with propidium iodide (PI) and failure to grow in culture. In contrast, Acanthamoeba cysts, which contain cellulose fibers in their wall, are relatively more resistant to these alcohols, particularly isopropanol. Depending upon the strain tested, 80 to 99% of Acanthamoeba cysts were killed by 63% ethanol after 2 min and 95 to 99% were killed by 80% ethanol after 30 sec, as shown by PI labeling and reduced rates of excystation in vitro. Both ethanol and isopropanol (63% for 30 sec) kill >99% of F. solani conidia, which have a wall of chitin and glucan fibrils, as demonstrated by PI labeling and colony counts on nutrient agar plates. Both ethanol and isopropanol (63% for 60 sec) inactivate 96 to 99% of elementary bodies of C. trachomatis, which have a wall of lipopolysaccharide but lack peptidoglycan, as measured by quantitative cultures to calculate inclusion forming units.Conclusions/SignificanceIn summary, alcohols kill infectious forms of Acanthamoeba, F. solani, and C. trachomatis, although longer times and higher ethanol concentrations are necessary for Acanthamoeba cysts. These results suggest the possibility that expanded use of alcohol-based hand sanitizers in places where water is not easily available might reduce transmission of these important causes of blindness.

The most abundant cyst wall proteins of Acanthamoeba castellanii are lectins that bind cellulose and localize to distinct structures in developing and mature cyst walls

“…Because water for washing hands may be scarce in places where the free-living protist is frequent, we recently showed that alcohols in concentrations present in hand sanitizers kill A . castellanii trophozoites and cysts [15, 16].…”

Effect of ethanol pretreatment in<em> Acanthamoeba</em> keratitis: a long-term follow-up study

“… 15 Furthermore, ethanol has been found to have a cytotoxic effect on some infectious forms of Acanthamoeba . 16 , 17 Therefore, soaking the cornea for 30 seconds in ethanol before debridement not only allows ophthalmologists to easily separate the full thickness of the epithelial layer from the basement membrane in both young and older patients with AK, but also had some parasiticidal effects. Moreover, the larger specimens harvested during debridement ensure not only a higher chance of diagnosing AK but also a better chance of penetration of topical anti-amoebic agents.…”

The most abundant cyst wall proteins ofAcanthamoeba castellaniiare cellulose-binding lectins from three gene families that localize to distinct structures in cyst walls

“…A. castellanii Neff strain trophozoites were purchased from the American Type Culture collection. Trophozoites of A. castellanii MEEI 0184 strain, which was derived from a human corneal infection, were obtained from Dr. Noorjahan Panjwani of Tufts University School of Medicine (9). Trophozoites were grown in T-75 tissue culture flasks at 30°C in 10 ml ATCC medium 712 (PYG plus additives) (Sigma-Aldrich Corporation, St. Louis, MO) (11).…”

“…Acanthamoeba is an emerging pathogen, because 80 to 90% of infections are associated with contact lens use, which is growing worldwide (6, 7). Because water for washing hands may be scarce in places where the free-living protist is frequent, we recently showed that alcohols in concentrations present in hand sanitizers kill A. castellanii trophozoites and cysts (8, 9).…”