Lysis of Trypanosoma cruzi by Pseudomonas fluorescens

Abstract

Trypomastigotes of Trypanosoma cruzi isolated from the blood of infected mice were lysed within 24 h by an extracellular substance produced by Pseudomonas fluorescens. Isolation of the anti-trypanosomal factor (ATF) was accomplished by growth of the organisms in a defined medium, extracellular secretion by the sedimented cells, sterilization by filtration, lyophilization, dialysis, and gel filtration. Chromatographic separation with Sephadex G-25 and G-200 disclosed the occurrence of three active fractions. ATF-I(1) exhibited a molecular weight higher than 440,000. ATF-II and ATF-III were considerably smaller (molecular weights approximately 1,355 and 1,060, respectively). The lytic substance contained protein and lipopolysaccharide, was resistant to heat and freezing, was not proteolytic or hemolytic, and was not inhibited by trypsin but was suppressed by pronase.During studies of Trypanosoma cruzi, we observed that the number of trypomastigotes isolated from the blood of infected mice markedly diminished when maintained in vitro at 4°C for 3 days. Microscopic examination revealed that the parasite samples had become contaminated by a small, motile, rod-shaped microorganism that was attracted to the parasites, particularly to the posterior end. As a result of the interaction, the trypomastigotes became immobilized, and most were lysed within 24 h. A gram-negative bacterium isolated in blood agar and nutrient broth cultures was identified as Pseudomonas fluorescens by the Clinical Pathology Department, National Institutes of Health. We considered these observations to be significant not only in terms of cellular biology and physiology, but also because of their potential significance in the chemotherapy of Chagas' disease; further studies were therefore conducted to analyze the parasite-bacterium interaction.MATERIALS AND METHODS Parasites. The Tulahuen strain of T. cruzi is routinely maintained in this laboratory by blood passaging every 6 days in NIH-Swiss 16-to 20-g male mice; 4 x 106 to 5 x 106 trypomastigotes are usually inoculated intraperitoneally (14). The infection becomes acute after 6 days, producing marked pathology of liver and spleen, and blood counts of 108 parasites per ml are often obtained. The animals are not immunosuppressed. Blood samples are obtained under ether anesthesia directly from the heart after exposure of the cardiac surface. Parasites were isolated from blood by a two-step procedure (15) involving separation of the erythrocytes with Lymphoprep (9.6% sodium-Nmethyl-3,5-diacetamido-2,4,6-triiodobenzoate and 5.5% Ficoll; specific gravity, 1.077; Nyegaard and Co., Oslo, Norway) and elution through a DEAEcellulose column to remove lymphocytes and platelets. The eluted flagellates exhibited the usual lanceolate shape (6), were actively motile and elicited typical infections. Two myotropic strains of T. cruzi, the House 510 and the House 11, of Costa Rican and Nicaraguan origin, respectively, trypomastigotes of T. rhodesiense 1886, T. equiperdum, and promastigotes of Leishmania brasilien...

Genome‐based discovery, structure prediction and functional analysis of cyclic lipopeptide antibiotics in Pseudomonas species

“…For the beneficial Pseudomonas species, a range of functions and activities have been reported (Raaijmakers et al ., 2006). For example, viscosin was shown to have antiviral activity (Groupe et al ., 1951) and to cause lysis of trypomastigotes of the human pathogen Trypanosoma cruzi , the causal agent of Chagas' disease (Mercado and Colon‐Whitt, 1982). For strain P. fluorescens SBW25, we focused on the role of CLP synthesis in surface motility, biofilm formation and activity against zoospores of P. infestans , the causal agent of late blight of tomato and potato.…”

“…2000). Also for the human pathogen T. cruzi , similar microscopic observations have been made (Mercado and Colon‐Whitt, 1982). In their study, the viscosin‐producing P. fluorescens isolate approached a single flagellate of T. cruzi at the posterior end, the site of the flagellar origin, resulting in immobilization.…”

Field-collected Triatoma sordida from central Brazil display high microbiota diversity that varies with regard to developmental stage and intestinal segmentation

“…Thus, Pseudomonas putida decreases Plasmodium falciparum levels in Anopheles gambiae mosquitoes [ 23 ], and Pseudomonas fluorescens causes T . cruzi lysis in vitro [ 55 ]. It is possible that some of the Pseudomonas species present in all T .…”

Glossina spp. gut bacterial flora and their putative role in fly-hosted trypanosome development

“…In sandflies ( Phlebotomus papatasi ), microbial infections significantly reduced the rates of infection with Leishmania major (Schlein et al, 1985). In addition, strains of Pseudomonas fluorescens (Mercado and Colon-Whitt, 1982), as well as of S. marcescens (which was isolated from Rhodnius prolixus ) (Azambuja et al, 2004) have been reported to be able to lyse Trypanosoma cruzi in vitro . All these examples show potential implication of bacteria isolated from insects in their vector competence.…”

“…(Thomson et al, 2000); hemagglutinins (Gilboa-Garber, 1972) and siderophore by P. aeruginosa (Schalk et al, 2002). An antitrypanosomal factor has been shown to be produced by P. fluorescens (Mercado and Colon-Whitt, 1982). Pigments such as prodigiosin are produced by the Gram-negative bacteria such as Serratia spp.…”