Immature salivary gland (SG) infections averaging 103 parasites per fly can apparently develop into mature gland infections averaging 105 parasites per fly in as little as 4 days. Frequently flies which extrude parasites in their saliva prove to have no parasites in the SG, but often show trypanosomes in the esophagus, cibarial pump, and proboscis. In some instances, SG infections have cleared, resulting in a loss of infectivity. Results of studying numbers of parasites regurgitated upon feeding or probing have shown that number to be highly variable and not necessarily related either to previous feeding status or the total number of parasites in the glands. Cloning of metacyclics in mice has been achieved, indicating that the minimum effective dose is one parasite. To date, no infections in mice have resulted from inoculation of extraglandular parasites. Histological and dissection studies support both the classical route and an alternate route of infection development in flies. No SG-infected flies have been found which did not also have proventricular and anterior and posterior midgut (AMG and PMG) infections. Although the AMG is where the heaviest MG infections occur, the PMG seems to support the last survivors in a moribund MG infection. No parasites have been found in the hindgut.