Heterogeneity in the in vitro susceptibility of Loa loa microfilariae to drugs commonly used in parasitological infections.

Co-infection with loiasis remains a potential problem in control programs targeting filarial infections. The effects of many anti-parasitic drugs often administered to Loa loa infected people are not well documented. This study compared the in vitro activity of several of these drugs on the viability of L. loa microfilariae (mf). Human strain L. loa mf were isolated from baboon blood using iso-osmotic Percoll gradient, and cultured in RPMI 1640/10% FBS with antimalarial drugs (mefloquine, amodiaquine, artesunate, chloroquine and quinine), anthelmintics (ivermectin, praziquantel, flubendazole and its reduced and hydrolyzed metabolites), two potential trypanocidal agents (fexinidazole and Scynexis-7158) and the anticancer drug imatinib. The drug concentrations used varied between 0.156 μg/ml and 10 μg/ml. Mf motility (CR = 50% immotility) and a metabolic viability assay (MTT) were used to assess the effects of these drugs on the parasites. Mf in control cultures showed only a slight reduction in motility after 5 days of culture. Active inhibition of Loa loa motility was seen with mefloquine and amodiaquine (CR values of 3.87 and 4.05 μg/ml, respectively), immobilizing > 90% mf within the first 24 hours: mefloquine killed the mf after 24 hours of culture at concentrations ≥ 5 μg/ml. SCYX-7158 also induced a concentration-dependent reduction in mf motility, with > 50% reduction in mf motility seen after 5 days at 10 μg/ml. The anticancer drug imatinib reduced mf motility at 10 μg/ml from the first day of incubation to 55% by day 5, and the reduction in motility was concentration-dependent. Praziquantel and fexinidazole were inactive, and FLBZ and its metabolites, as well as ivermectin at concentrations > 5 μg/ml, had very minimal effects on mf motility over the first 4 days of culture. The considerable action of the anti-malarial drugs mefloquine and amodiaquine on Loa mf in vitro highlights the possibility of repurposing the existing anti-infectious agents for the development of drugs against loiasis. The heterogeneity in the activity of anti-parasitic agents on Loa loa mf supports the need for further investigation using animal models of loiasis

a. Mean RMS amplitudes for live and dead adult male and female <i>B. pahangi</i>.

<p><b>b</b>. Mean RMS amplitudes for adult females treated with different molar concentrations of imatinib mesylate, (*) significant differences from before drug treatment at 1hpt and on (p<0.0291). <b>c</b>. Mean RMS amplitudes for adult females treated with different molar concentrations of chloroquine significant differences from before drug treatment at 1hpt and on (p<0.0346).</p

Additional file 1: of Heterogeneity in the in vitro susceptibility of Loa loa microfilariae to drugs commonly used in parasitological infections

Table S1. Effects of different drug concentrations on the motility of L. loa mf. (DOCX 33 kb

a. Mean returns (y-axis) for <i>B. pahangi</i> microfilariae from 0–1500 worms (x-axis) per tube.

<p>(*) shows significant difference between 300mf and blank using Tukey’s test (p = 0.0079), <b>b</b>. Mean returns for 5000 live microfilariae, 5000 dead microfilariae, and 5000 live & dead. <b>c</b>. Mean returns for 100 live microfilariae, 100 dead microfilariae, and 100 live & dead. <b>d</b>. 500 live microfilariae, 500 dead microfilariae, and 500 live & dead microfilariae.</p

a. Mean RMS amplitudes of <i>C. elegans</i> L1 from 0 and 100 worms, live and dead.

<p><b>b</b>. FFT of 100 <i>C. elegans</i> L1(blue), overlaid with 100 <i>b. pahangi</i> (black), and blank (red). <b>c</b>. Ratios of selected frequency ranges of <i>C. elegans</i> L1 motion, showing significant differences (*) in RMS amplitudes of degraded worms vs blanks (p<0.0001), healthy worms vs blank (p<0.0001), and healthy worms vs degraded in all but the ratio of middle frequency range to high frequency range (p = 0.3095).</p

a. Mean RMS amplitudes for blank and 100 live and dead <i>B. pahangi</i> L3’s.

<p><b>b</b>. Mean RMS amplitudes over time of L3’s treated at different molar concentrations of imatinib mesylate, (*) significant change from pretreatment earlier than control or other groups (p = 0.0153). <b>c</b>. Mean RMS amplitudes of L3’s treated at different molar concentrations of chloroquine, (*) significant change from pretreatment earlier than control or other groups (p<0.0001). <b>d</b>. Ratios of the maximum amplitudes for the selected frequency ranges of L3 motion, healthy (*) significantly different from blank or degraded (all p<0.0001), blank statistically same as degraded (all p>0.1917)</p