Trypanosoma brucei spp. development in the tsetse fly: characterization of the post-mesocyclic stages in the foregut and proboscis.

Post-mesocyclic development of Trypanosoma brucei in the tsetse fly in its migration from midgut to salivary glands, was revisited by sequential microdissection, morphometry and DNA-cytofluorometry. This development started by day 6 after the infective feed, with passage of mesocyclic midgut trypomastigotes through proventriculus and upward migration along foregut and proboscis to the salivary gland ducts. Kinetics of salivary gland infection showed that colonization of the salivary glands by epimastigotes occurred only during the time-limited presence of this developmental phase in the foregut and proboscis. Post-mesocyclic trypanosomes in the foregut and proboscis were pleomorphic, with 4 morphological stages in various constant proportions and present all through from proventriculus up to the salivary gland ducts: 67% long trypomastigotes, 27% long epimastigotes, 4% long epimastigotes undergoing asymmetric cell division and 2% short epimastigotes. Measurements of DNA content demonstrated a predominant tetraploidy for 67% of these trypanosomes, the remainder consisting of the homogeneous diploid short epimastigotes and some long epimastigotes. According to the experimental data, the following sequence of trypanosome differentiation in the foregut and proboscis is proposed as the most obvious hypothesis. Incoming mesocyclic trypomastigotes (2N) from the ectoperitrophic anterior midgut start to replicate DNA to a 4N level, are arrested at this point, and differentiate into the long epimastigote (4N) which give rise, by an asymmetric cell division, to 2 unequal, diploid daughter cells: a long, probably dead-end long epimastigote and a short epimastigote. The latter is responsible for the epimastigote colonization of the salivary glands if launched at the vicinity of the gland epithelium by the asymmetric dividing epimastigote.

Trypanosome-binding proteins of the tsetse flies Glossina palpalis gambiensis and G. morsitans morsitans.

In this paper we describe a new, selective approach to identify protein ligand-receptor interactions between an arthropod vector and the parasite it transmits. Biotinylated vector proteins were incubated with living parasites in physiological conditions. After extensive washing, the parasites were subjected to SDS-PAGE electrophoresis and the polypeptides were electroblotted onto nitrocellulose membrane. Staining with avidin-horseradish peroxidase revealed only biotin-labeled proteins from the vector which were bound to the parasite. A multitude of tissue-specific proteins of Glossina palpalis gambiensis and G. morsitans morsitans proteins, able to bind to cultured procyclic trypanosomes of Trypanosoma brucei spp., has been demonstrated. The relevance of these interactions in relation to the developmental journey of the trypanosome in the tsetse fly is briefly discussed.

Cytogenetic and isozymic comparisons of two laboratory lines of Glossina palpalis gambiensis.

The genetics of two laboratory colonies of Glossina palpalis gambiensis were characterized by C-banding and isoenzyme studies. The colonies, derived from flies collected in the same locality, had different histories in the laboratory and different susceptibilities to trypanosome infection. Although the two lines were also found to differ in the frequencies of chromosome and isozyme variants, the variation was not enough to put their specific status in doubt; it was probably the result of genetic drift since the foundation of the colonies.

Transient adenylate cyclase activation accompanies differentiation of Trypanosoma brucei from bloodstream to procyclic forms.

Pleomorphic bloodstream forms of Trypanosoma brucei differentiate synchronously into procyclic forms when cultivated at 27 degrees C in the presence of citrate/cis-aconitate. The activity of adenylate cyclase was monitored during this process. Two phases of transient stimulation were observed. The first phase occurred 6-10 h after the triggering of differentiation, a period which immediately follows the release of the bulk of the VSG and immediately precedes both the first cell division and the loss of the bloodstream-specific ESAG 4 transmembrane adenylate cyclase. The second phase occurred between 20 and 40 h, when the cells that emerged from the first division began to proliferate. These observations suggest that cAMP may be involved in differentiation/proliferation of the parasite.

Direct isolation in vitro of Trypanosoma brucei from man and other animals, and its potential value for the diagnosis of gambian trypanosomiasis.

A recently described simple kit for isolating African trypanosomes in vitro (KIVI) was tested further with blood samples from man and other animals in Côte d'Ivoire and République du Congo. A high rate of success was achieved, with positive cultures being found 5-36 d after inoculation. The method was also of value in diagnosis. Parasitaemia was initially detected by the haematocrit method; in addition, the mini-anion exchange column was used for human blood and lymph fluid from patients with swollen glands was examined. The card agglutination test (CATT) was applied to the human blood samples. In Côte d'Ivoire, all 5 parasitaemic patients, who were also positive by CATT, yielded positive KIVI cultures. Of 15 animals, 2 parasitaemic and 10 apparently aparasitaemic individuals gave positive cultures. In the Congo, none of the 22 animals was parasitaemic and none gave a positive culture. Of 647 human subjects initially screened, 61, mostly with a positive CATT, were examined by KIVI; 20 gave positive cultures. Seven of these cultures originated from patients in whom no trypanosome had been seen in blood or lymph fluid, although blood from 2 parasitaemic patients failed to yield positive KIVI cultures. Some patients with CATT-negative whole blood and/or serum were positive by KIVI.

Influence of hydroxyethyl starch on coagulation in patients during the perioperative period.

The perioperative use of hydroxyethyl starch (HES) has been implicated as a possible cause of intracranial bleeding. The purpose of this study was to compare the influence on blood coagulation of the isovolemic replacement of 1-L blood loss with either 6% HES (molecular weight [MW] average: 450,000) or 5% human albumin during neurosurgery or lower abdominal surgery. Twenty patients scheduled for brain tumor surgery and 20 patients undergoing transabdominal hysterectomy were studied. The activated partial thromboplastin time, prothrombin time, fibrinogen concentration, factor VIII coagulant, von Willebrand factor antigen, platelet count, and the activated clotting time were compared after induction of anesthesia, after administration of 500 and 1000 mL of colloid solution, and 24 and 48 h postoperatively. All measured coagulation variables remained within physiologic range. Changes in coagulation indices were identical in neurosurgical and hysterectomy patients, except for a larger increase in fibrinogen concentration 24 and 48 h after hysterectomy. The acute phase reaction of factor VIII coagulant and von Willebrand factor, which plays a role in postoperative hypercoagulability, was attenuated by the use of HES. We conclude that isovolemic replacement of 1-L blood loss with either 6% HES (MW average: 450,000) or 5% human albumin does not interfere with normal hemostasis during and after neurosurgery or lower abdominal surgery.

Influence of the salmon mutant of Glossina morsitans morsitans on the susceptibility to infection with Trypanosoma congolense.

Four phenotypes of a sex-linked, maternally influenced semi-lethal eye color mutant of Glossina morsitans morsitans Westwood were fed on Trypanosoma congolense Broden infected guinea pigs. Infection rates were evaluated 25 days later by means of dissection. Procyclic as well as mature infections were significantly more common among females with salmon-colored eyes (sal/sal) than among heterozygous (+/sal, phenotypically wild-type) females. A tendency was found for more mature infections among sal/Y males than among wild-type males. Similarly, females tended to be more infected than males with both procyclic and mature infections. These results indicate that the genotype of the fly, exemplified by the allele salmon, might influence the development of T. congolense in G.m. morsitans. A possible explanation for this phenomenon is discussed.