Tsetse Fly Midgut Research Articles

Procyclic tsetse fly midgut forms and culture forms of African trypanosomes share stage- and species-specific surface antigens identified by monoclonal antibodies.

Procyclic culture form (PCF) trypanosomes were established from a bloodstream form population of cloned Trypanosoma brucei rhodesiense and were used to immunize mice for hybridoma production. Indirect immunofluorescence was used to select 10 hybridomas which secreted antibodies that bound to the surface of homologous living PCF. The antibodies reacted with PCF of several clones of T.b. brucei, T.b. gambiense, and T.b. rhodesiense, but not with PCF of T. congolense or T. vivax, or with promastigotes of several species of Leishmania parasites. The antigens were not detectable in ethanol/acetic acid-fixed bloodstream forms or in lysates of bloodstream forms of any of the T. brucei subspecies, and are thus species-specific and stage-specific markers. Selected monoclonal antibodies bound to procyclic trypanosomes taken directly from the midgut of infected tsetse flies, and to immature epimastigote forms in salivary probes, and may therefore be useful in epidemiologic investigations.

Trypanosoma brucei: Cultivation In vitro of Infective Forms Derived from the Midgut of Glossina morsitans

Infective forms of Trypanosoma brucei derived from the midgut of Glossina morsitans, have been propagated in vitro for 61 days on a bovine embryonic spleen (BESP) feeder layer using RPMI 1640 medium. It was reproducibly shown that only parasites cultured from the midgut of tsetse flies 12-14 hr after feeding on infected animals could be established in vitro. Cultures thus established were infective to rats and tsetse flies. Only midgut vector types of the parasites were identified by light and electron microscopic techniques.

Digestive processes of haematophagous insects. XIV. Haemolytic activity in the midgut of Glossina morsitans morstians Westwood (Diptera: Glossinidae).

A haemolytic agent (haemolysin), which lysed rabbit erythrocytes in an isotonic buffer. occurred in the digestive section of the midgut of adult male and female tsetse flies (Glossina morsitans) but not in the anterior, nondigestive section of the midgut. Haemolysin from females also lysed horse, cow, and pig erythrocytes, and it occurred in the midgut lumen but not in the midgut wall 6, 24. and 48 h after feeding. Little or no haemolysin was present in unfed flies but it was present in the midgut by 3 h after feeding on a rabbit: maximum activity occurred in the midgut 24–48 h after feeding. Flies fed on a meal containing 20 μg or more puromycin/ml produced only small quantities of haemolysin. Flies fed upon erythrocytes, haemoglobin, methaemoglobin, and myoglobin produced large quantities of haemolysin while those fed upon erythrocyte stroma, serum proteins, cytochrome c, or 1 mM ATP in 0.85% NaCl did not.

Quantitative studies on trypanosomes in tsetse tissue culture

Tissues from pupae of Glossina morsitans of various ages were cultured in modified Trager's medium. Cellular outgrowths were produced from explants of proventriculus, brain, and imaginal body wall and large vesicles were extruded from pieces of midgut of young pupae. Complete alimentary tract from older pupae displayed rhythmic contractions for up to 3 weeks. When Trypanosoma brucei and T. congolense in mouse blood were added to hanging drop cultures of tsetse tissues and incubated at 28 C, the organisms multiplied and changed into forms morphologically similar to those found in the tsetse fly midgut. The trypanosomes were maintained for 30 days by serial passage at 5-day intervals. The growth of T. brucei in the presence of different pupal tissues was studied. Of all the tissues tested the complete alimentary tract from pupae older than 21 days gave the best results. Growth also occurred when the trypanosomes were separated from the insect tissue by a semipermeable membrane. The trypanosomes failed to grow in (a) culture medium alone, (b) media containing extracts of alimentary canal and (c) medium in which alimentary tract had been cultured for 3 or 4 days.