Trypanosoma vivax Infections: Pushing Ahead with Mouse Models for the Study of Nagana. II. Immunobiological Dysfunctions

Trypanosoma vivax is the main species involved in trypanosomosis, but very little is known about the immunobiology of the infective process caused by this parasite. Recently we undertook to further characterize the main parasitological, haematological and pathological characteristics of mouse models of T. vivax infection and noted severe anemia and thrombocytopenia coincident with rising parasitemia. To gain more insight into the organism's immunobiology, we studied lymphocyte populations in central (bone marrow) and peripherical (spleen and blood) tissues following mouse infection with T. vivax and showed that the immune system apparatus is affected both quantitatively and qualitatively. More precisely, after an initial increase that primarily involves CD4+ T cells and macrophages, the number of splenic B cells decreases in a step-wise manner. Our results show that while infection triggers the activation and proliferation of Hematopoietic Stem Cells, Granulocyte-Monocyte, Common Myeloid and Megacaryocyte Erythrocyte progenitors decrease in number in the course of the infection. An in-depth analysis of B-cell progenitors also indicated that maturation of pro-B into pre-B precursors seems to be compromised. This interferes with the mature B cell dynamics and renewal in the periphery. Altogether, our results show that T. vivax induces profound immunological alterations in myeloid and lymphoid progenitors which may prevent adequate control of T. vivax trypanosomosis

Trypanosoma vivax Infections: Pushing Ahead with Mouse Models for the Study of Nagana. I. Parasitological, Hematological and Pathological Parameters

African trypanosomiasis is a severe parasitic disease that affects both humans and livestock. Several different species may cause animal trypanosomosis and although Trypanosoma vivax (sub-genus Duttonella) is currently responsible for the vast majority of debilitating cases causing great economic hardship in West Africa and South America, little is known about its biology and interaction with its hosts. Relatively speaking, T. vivax has been more than neglected despite an urgent need to develop efficient control strategies. Some pioneering rodent models were developed to circumvent the difficulties of working with livestock, but disappointedly were for the most part discontinued decades ago. To gain more insight into the biology of T. vivax, its interactions with the host and consequently its pathogenesis, we have developed a number of reproducible murine models using a parasite isolate that is infectious for rodents. Firstly, we analyzed the parasitical characteristics of the infection using inbred and outbred mouse strains to compare the impact of host genetic background on the infection and on survival rates. Hematological studies showed that the infection gave rise to severe anemia, and histopathological investigations in various organs showed multifocal inflammatory infiltrates associated with extramedullary hematopoiesis in the liver, and cerebral edema. The models developed are consistent with field observations and pave the way for subsequent in-depth studies into the pathogenesis of T. vivax - trypanosomosis

Histopathological study of mice infected with <i>Trypanosoma vivax</i>.

<p>8-week-old Outbred mice were injected i.p. with 10² bloodstream forms of <i>T. vivax</i> and different lymphoid and non lymphoid organs were harvested for histopathological examination 20 days post-infection. Spleen (A–D): (A) Diffuse lesions characterized by large necrotic foci in the red pulp (black star), associated with lymphoid tissue disorganization in the white pulp (white star). (B) Infiltration of a necrotic focus by activated macrophages (top of the Fig., arrows) and trypanosomes (arrowhead shows very small basophilic points in the inset depicting a higher magnification). (C) Presence of lower density hematopoiesis compared to non-infected mice. (D) Infiltration of the white pulp by activated macrophages and presence of a Mott cell (arrow). Liver (E–G): (E) Multifocal inflammatory lesions centered on portal tracts/centrilobular veins (arrows), and focal necrotic focus (star). (F) Peri-venous inflammatory infiltrate composed of plasma cells (mostly), but also lymphocytes and macrophages. In the inset depicting a higher magnification, arrowhead points to trypanosomes in the vascular spaces. (G) Foci of extramedullary hematopoiesis. Kidney (H–J): (H) Interstitial inflammatory infiltrates (I) mostly composed of plasma cells. (J) Trypanosomes in an arcuate artery (star); in the inset depicting a higher magnification, arrowhead points to trypanosomes in the vascular spaces. Cerebellum (K–M): (K) Multifocal lesions centered on blood vessels (arrows). (L) Blood vessel lumen filled by trypanosomes, proteins and erythrocytes (star), with perivascular edema (arrow) and ischemic neurons (arrowheads). (M) Trypanosomes in a meningeal blood vessel. Hematoxylin-eosin staining, scale bars are indicated at the bottom of each photograph. In the inset depicting a higher magnification, the arrowhead points to trypanosomes.</p

<i>T. vivax</i> induces major perturbations in hematological parameters and causes severe thrombocytopenia.

<p>8-week-old Outbred (white symbols) or C57BL/6 (black symbols) mice were injected i.p. with 1×10² bloodstream forms of <i>T. vivax</i>. Blood samples were collected individually every 2–3 days and red blood cell counts (RBC, A), hematocrit (HCT, B), hemoglobin concentrations (HGB, C), leukocytes (WBC, D) and platelets (PLA, E), were determined. Results are given for days 5, 10, 15 and 20 as arithmetic means ± standard deviations of at least three different experiments with 3–5 mice per time point/experimental group. *** p<0.001, ** p<0.01, * p<0.05, when compared with samples from day 0. $ = not determined on day 10 for C57BL/6 mice.</p

Molecular identity of <i>T. vivax</i> IL 1392.

<p>Full lengh of <i>ILDat1.2 VSG</i> gene (A). Initiation and stop codons are underlined. The specific <i>T. vivax</i> 20-amino acids sequence at the N-terminal end of the gene is depicted in red (positions 64 to 123). Forward and reverse primers used in this experiment are in italics. DNA was extracted from <i>T. vivax</i> bloodstream forms and amplified by PCR using <i>VSG-1.2</i>F and <i>VSG-1.2</i>R primers. A fragment of 148 bp was obtained and the resulting sequence aligned with the Y486 reference strain (B). Two point mutations are squared. Blood smears of a mouse infected with <i>T. vivax</i> were fixed and stained with Giemsa (C and D); k = kinetoplast, f = flagellum. The high number of circulating parasites at the peak of parasitemia can be evaluated in the picture (E).</p

Effect of host background on parasite load and fate.

<p>BALB/c, C57BL/6 or Outbred mice were injected i.p. with 1×10² bloodstream forms of <i>T. vivax</i> and the mean parasitemia recorded individually during infection (A, B, C). Mean mortality (D) is depicted compared to BALB/c mice. Results are given as arithmetic means ± standard deviations of at least three independent experiments. Cumulative mortality was recorded over time for all groups and Kaplan-Meir survival curves plotted for the three mouse strains (D); Comparison between survival curves was performed using Log-rank Mantel-Cox test: * p<0.028, ** p<0.0018, when compared with BALB/c survival.</p