Mast cells, histamine and the pathogenesis of intestinal damage in experimental Trypanosoma brucei brucei infections

Intestinal damage with increased permeability is a prominent feature of experimental African trypanosomiasis. The possible involvement of mast cells and histamine in the altered gut integrity was investigated, at the level of the jejunum, in BALB/c mice infected with Trypanosoma brucei brucei. Mast cells were studied by selective staining of granule content with Alcian Blue/Safranin and quantitative histology, and histamine concentrations were determined by a fluorimetric method. Mast-cell activation, shown by a marked reduction in the numbers of positive-staining cells seen per villous section, was prominent on days 7 and 14 post-infection (there was, for example, a reduction to 36% of the control value by day 14; P=0.0001). By day 21, however, there were 131% more staining cells per villous section in the infected mice than in the uninfected controls (P=0.003). Histamine levels in homogenates of the jejunal mucosae of the infected mice were found to be significantly elevated at each time-point. The maximum increase was observed on day 14, when the numbers of granulated mast cells were at their lowest, with mean (S.E.) concentrations of 6.744 (0.890) ng/mg tissue for the infected mice and 2.813 (0.321) ng/mg for the uninfected controls (P=0.0008). The jejunal mucosa suffered progressive morphological damage during the infection, with oedema of the lamina propria and villi and disruption of the endothelium. These results indicate that mast cells are involved with the intestinal pathology that develops during experimental African trypanosomiasis.     

Physiologic interactions between L-proline and D-glucose in Leishmania tarentolae, L. donovani and Trypanosoma scelopori culture forms.

L-proline is more completely catabolized by Leishmania tarentolae and Trypanosoma scelopori than by L. donovani. In the two former organisms the activity of 14CO2 derived from L-proline-14C was as high or higher than that derived from D-glucose-14C; in L. donovani more 14CO2 is derived from D-glucose-14C than from L-proline-14C. Addition of unlabeled D-glucose to cells incubated with L-proline-14C resulted in increased 14C-activity of TCA-soluble fractions of L. tarentolae and T. scelopori; 14C-activity of 5% TCA insoluble fractions of these cells remained unchanged. Some of the increased 14C-label found in the soluble fraction was in glutamate and aspartate. This increased activity in glutamate and aspartate was accompanied by a decrease in alanine-14C in the free amino acid pool. These results suggest that there are complex physiological interactions between glucose and proline in the metabolism of L. tarentolae, L. donovani and T. scelopori. In addition, proline appears to have a more central role in the amino acid metabolism of L. tarentolae and T. scelopori whereas glucose may be more significant in L. donovani.     

Comparison of maxicircle DNAs of Leishmania tarentolae and Trypanosoma brucei.

The conserved portions of the maxicircle DNAs of Leishmania tarentolae and Trypanosoma brucei are organized in a basically colinear manner over a 15- to 17-kilobase region that is interrupted by two small less-homologous sequences. The most highly conserved regions are those encoding the 9S and 12S genes. An approximately 12-kilobase region directly upstream of the 12S gene in the L. tarentolae maxicircle showed no sequence homology with the T. brucei maxicircle and also was not transcribed. An approximately 6-kilobase region in the T. brucei maxicircle in the same relative location also showed no sequence homology with the L. tarentolae maxicircle. We propose that evolution of maxicircle DNA occurs mainly within this "divergent region."     

Parasite-driven pathogenesis in Trypanosoma brucei infections

Trypanosomes are protozoan parasites of medical and veterinary importance. It is well established that different species, subspecies and strains of trypanosome can cause very different disease in the mammalian host, exemplified by the two human-infective subspecies of Trypanosoma brucei that cause either acute or chronic disease. We are beginning to understand how the host response shapes the course of the disease and how genetic variation in the host can be a factor in disease severity, particularly in the mouse model, but until recently the role of parasite genetic variation that determines differential disease outcome has been a neglected area. This review will discuss the recent advances in this field, covering both our current knowledge of the T.?brucei genes involved and the approaches that are leading towards the identification of T.?brucei virulence genes. Finally, the potential for using parasite genotype variation to examine the evolutionary context of virulence will be discussed.     

Cerebral trypanosomiasis in cattle with mixed Trypanosoma congolense and T. brucei brucei infections

Six Boran steers were infected simultaneously with Trypanosoma congolense and T. brucei brucei while another group of 3 was inoculated with T. b. brucei one year after infection with T. congolense. Three further steers were infected with T. b. brucei alone. Whereas, the six animals which received simultaneous infections developed clinical signs of cerebral trypanosomiasis as evidenced by depression, ataxia and occasional circling, those infected with T. b. brucei alone did not. At necropsy, 4 out of the 6 simultaneously infected animals had a mild to severe disseminated non-suppurative meningoencephalitis. Trypanosoma b. brucei was isolated from the cerebrospinal fluid (CSF) of three out of the four animals with histological lesions. Two of the cattle superinfected with T. b. brucei one year after infection with T. congolense also developed both clinical and histological evidence of cerebral trypanosomiasis. Trypanosoma congolense was isolated from the CSF of one of these 2 animals. Specific antibodies to the variable surface glycoproteins (VSGs) of the infecting T. b. brucei and T. congolense clones were found in the CSF of the 8 animals that developed cerebral trypanosomiasis. In these animals however, there was neither temporal nor quantitative correlation between VSG-specific antibodies in serum and in CSF, implying a de novo synthesis of antibodies to the infecting trypanosomes in the CSF.     

The relapse of Trypanosoma brucei brucei infections after chemotherapy in rabbits

Infections of T. brucei in the rabbit were found to relapse after chemotherapy. The results indicated that 25 mg/kg diminazene aceturate given 3 days after infection resulted in a complete cure but if given 7 days after infection relapses frequently occurred. However, treatment was apparently successful if delayed until 14 or 21 days. Six of the rabbits originally treated with diminazene aceturate on day 7 were treated with suramin 21 days later; in 3 rabbits the infections relapsed. In all rabbits in which drug treatment was not curative, the clinical condition nevertheless improved. An attempt to locate a cryptic focus of infection in rabbits was unsuccessful.     

Specificity in signal transduction among glycosylphosphatidylinositols of Plasmodium falciparum, Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp

Glycosylphosphatidylinositols (GPIs) and related glycoconjugates of parasite origin have been shown to regulate both the innate and acquired immune systems of the host. This is achieved through the activation of novel GPI-dependent signalling pathways in macrophages, lymphocytes and other cell types. Parasite GPIs impart at least two distinct signals to host cells through the structurally distinct inositolphosphoglycan (IPG) and fatty acid domains. Binding of IPG to as yet uncharacterized cell surface receptor(s) leads to activation of src-family protein tyrosine kinases: depending upon structure, GPI-derived fatty acids can either activate or antagonize protein kinase C, and may enter the sphingomyelinase pathway. The degree of fatty acid saturation may also contribute to signalling activity. Thus, variation in structure of parasite GPIs imparts different properties of signal transduction upon this class of glycolipid. The divergent activities of GPIs from various protozoal taxa reflect global aspects of the host/parasite relationship, suggesting that GPI signalling is a central determinant of disease in malaria, leishmaniasis and both American and African trypanosomiases.     

New fluorescence markers to distinguish co-infecting Trypanosoma brucei strains in experimental multiple infections

Multiple-genotype infections are increasingly recognized as important factors in disease evolution, parasite transmission dynamics, and the evolution of drug resistance. However, the distinction of co-infecting parasite genotypes and the tracking of their dynamics have been difficult with traditional methods based on various genotyping techniques, leaving most questions unaddressed. Here we report new fluorescence markers of various colours that are inserted into the genome of Trypanosoma brucei to phenotypically label live parasites of all life cycle stages. If different parasite strains are labelled with different colours they can be easily distinguished from each other in experimental studies. A total of 10 T. brucei strains were successfully transfected with different fluorescence markers and were monitored in culture, tsetse flies and mice, to demonstrate stability of marker expression. The use of fluorescence activated cell sorting (FACS) allowed rapid and accurate identification of parasite strains labelled with different markers. Cell counts by FACS were virtually identical to counts by traditional microscopy (n=75, Spearman's rho: 0.91, p<0.0001) but were considerably faster and had a significantly lower sampling error (66% lower, d.f.=73, t=-17.1, p<0.0001). Co-infecting strains transfected with fluorescence genes of different colour were easily distinguished by eye and their relative and absolute densities were reliably counted by FACS in experimental multiple infections in mice. Since the FACS can simultaneously determine the population sizes of differently labelled T. brucei strains or subspecies it allows detailed and efficient tracking of multiple-genotype infections within a single host or vector individual, enabling more powerful studies on parasite dynamics. In addition, it also provides a simple way to separate genotypes after experimental mixed infections, to measure responses of the single strains to an applied treatment, thus eliminating the need for laborious cloning steps. The markers presented broaden the spectrum of tools available for experimental studies on multiple-genotype infections. They are fundamentally different from isoenzyme analysis and other genotyping approaches in that they allow the distinction of parasite genotypes based on an easily recognizable phenotypic trait. They will be of specific interest to researches addressing ecological, evolutionary and epidemiological questions using trypanosomes as an experimental system.     

Metabonomic investigation of single and multiple strain Trypanosoma brucei brucei infections

Although co-infections are common and can have important epidemiologic and evolutionary consequences, studies exploring biochemical effects of multiple-strain infections remain scarce. We studied metabolic responses of NMRI mice to Trypanosoma brucei brucei single (STIB777AE-Green1 or STIB246BA-Red1) and co-infections using a (1)H nuclear magnetic resonance (NMR) spectroscopy-based metabolic profiling strategy. All T. b. brucei infections caused an alteration in urinary biochemical composition by day 4 postinfection, characterized by increased concentrations of 2-oxoisocaproate, D-3-hydroxybutyrate, lactate, 4-hydroxyphenylacetate, phenylpyruvate, and 4-hydroxyphenylpyruvate, and decreased levels of hippurate. Although there were no marked differences in metabolic signatures observed in the mouse infected with a single or dual strain of T. b. brucei, there was a slower metabolic response in mice infected with T. b. brucei green strain compared with mice infected with either the red strain or both strains concurrently. Pyruvate, phenylpyruvate, and hippurate were correlated with parasitemia, which might be useful in monitoring responses to therapeutic interventions.     

Antitumor phthalanilides active in acute and chronic Trypanosoma brucei brucei murine infections

A series of phthalanilides and related compounds were tested on a short-term, fulminating, mouse infection of Trypanosoma brucei brucei (EATRO 110 isolate). The most effective compound was [4,4'-bis (4-methylimidazolin-2-yl)-terephthalanilide] which had a cure rate of 75% at 0.1 mg/kg body weight and 100% at 0.5 mg/kg when administered as three single daily intraperitoneal injections starting 24 hours post-infection. Several related phthalanilides and similarly substituted ureas showed definite but lower activity. In tests with a chronic neurotropic T. b. brucei isolate (TREU 667), cure rates greater than 90% were obtained with 10 or 25 mg/kg [4,4'-bis(4-methylimidazolin-2-yl)-terephthalanilide]. Cured animals survived for at least 200 days after infection with no evidence of recrudescence of parasitemia or of toxicity; blood or brain homogenates of cured animals were non-infective to immunosuppressed animals. These studies indicate that this series of compounds, previously studied as antitumor agents, should be re-examined as potential trypanocides.     

Multiple-strain infections of Trypanosoma brucei across Africa

It is becoming increasingly clear that parasitic infections frequently contain multiple strains of the same parasite species. This may have important consequences for the parasite dynamics in the host and thus alter disease and transmission dynamics. In Trypanosoma brucei, the causal agent of human African trypanosomiasis (sleeping sickness), multiple-strain infections have previously been demonstrated to occur. Here, we analyzed field isolates of T. b. gambiense, T. b. rhodesiense, and T. b. brucei, isolated throughout Africa to assess the commonness of multiple-strain infections across the natural range of this parasite. Using eight highly variable microsatellite loci, we found multiple strains in 8.8% of our isolates. Due to the technical challenges of detecting multiple infections this number represents a minimum estimate and the true frequency of multiple-strain infections is likely to be higher. Multiple-strain infections occurred across the entire East-West range of the parasite. Together with previous results, these findings strongly suggest that multiple-strain infections are common for this parasite and that their consequences for epidemiology and parasite evolution should be investigated in detail.     

Leishmania mexicana complex: human infections in the Republic of Panama

Parasites of the genus Leishmania responsible for human cutaneous leishmaniasis in the New World form 2 major taxonomic divisions: the Leishmania braziliensis and the L. mexicana complexes. We report the isolation and characterization of the L. mexicana complex among humans in the Republic of Panama. Characterization was based on parasite morphology, pathogenesis in infected golden hamsters, cellulose acetate isoenzyme electrophoretic mobilities, and membrane-specific monoclonal antibodies using the radioimmune binding assay technique.     

Leishmaniasis in Brazil: XII. Observations on cross-immunity in monkeys and man infected with Leishmania mexicana mexicana, L. m. amazonensis, L. braziliensis braziliensis, L. b. guyanensis and L. b. panamensis.

Cross-immunity trials in monkeys (Cebus apella apella) and observations on experimental and natural infections in man confirm the separate identity of L. mexicana mexicana, L. m. amazonensis, L. b. braziliensis, L. b. guyanensis and L. b. panamensis. Neither L. m. mexicana nor L. m. amazonensis infections gave protection against subsequent challenge with parasites of the L. braziliensis complex; but recovery from infection with subspecies of L. braziliensis in most cases gave firm resistance to infection with the mexicana parasites. The failure of certain New World leishmanias to immunize against each other has an important bearing on taxonomy, future attempts to prepare vaccines against Leishmania, epidemiology and diagnosis.