Optimization of test conditions for development of MTT as in vitro screen.

The quick and easy method of tetrazolium based colorimetric assay with MTT [3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] was used to test the viability of the adult parasites of a rodent filariid Acanthocheilonema viteae in vitro. The ideal conditions required for antifilarial screening were determined by correlating the MTT reduction ability of worms with their size and age in the vertebrate host, also the duration of incubation and temperature of the in vitro culture. It was observed that the worms collected from the host after 90 days of L3 (infective larvae) exposure were not suitable for in vitro screen as they could not reduce MTT to that extent as the worms of early infection. Healthy and full grown worms and also those incubated at 37 degrees C for 16 hr or more caused maximum MTT reduction. Thus, it is recommended to select healthy adult filariids of proper age and size (male > 3.5 cm; female > 7.0 cm). The incubation temperature of the in vitro culture system needs to be adjusted to 37 degrees C and parasites might be exposed to drugs upto 24 hr without much alteration in MTT reduction of untreated controls.

Impact of surface modifications of Acanthocheilonema viteae microfilariae on cell adhesion.

Exposure of A. viteae microfilariae to various lectins reduced their capacity to react with the peritoneal exudate cells of the host, Mastomys natalensis. Sugars corresponding to these lectins with the exception of N-acetyl glucosamine, did not affect the adhesion per se. They however, protected the parasite against the adverse effect of lectins. Neuraminidase and chitinase also suppressed adhesion capacity of the microfilariae. Except sodium dodecylsulphate which enhanced cell attachment, other surfactants inhibited this reaction considerably. The results indicate that antibody dependent adhesion of the microfilariae with the macrophages involves surface moieties of the parasite, where N-acetylglucosamine acts as the principal sugar residue. Participation of -SH groups also is inferred from the observations that p-chloromercuribenzoate and dithiobis-(2-nitrobenzoic acid) inhibited cell attachment and dithiothreitol provided protection against these agents.