Peak Parasitemia Research Articles

Efficacy of diminazene diaceturate and isometamidium chloride hydrochloride for the treatment of Trypanosoma evansi in mice model.

Diminazene diaceturate (DIM) and isometamidium chloride hydrochloride (ISMM) have been widely used for the treatment of animal trypanosomosis. We evaluated the efficacy of standard doses of DIM and ISMM followed by their double doses for the treatment of Trypanosoma evansi in experimentally infected mice. A T. evansi strain obtained from a naturally infected camel in Afar was used. 25 swiss white mice randomly divided in to five groups were inoculated with 0.2mL of blood containing 103 trypanosomes. At the peak of parasitemia (≈ 2weeks post infection), groups A and B were treated with the standard dose (3.5mg/kg body weight [BWT]) of DIM; groups C and D were treated with the standard dose (0.5mg/kg BWT) of ISMM; and group E served as infected control. In the DIM standard dose groups, relapses and peak parasitemia were observed 20- and 25-days post treatment respectively. Similarly, relapses and peak parasitemia were observed 21- and 27-days post treatment in the ISMM standard dose groups. All mice in the control group died within two weeks post infection. Following relapses, mice were treated with the double doses of DIM (7mg/kg BWT) or ISMM (1mg/kg BWT). Parasitemia was not detected for 3months following the double dose treatments. Following dexamethasone administration for 7days, all but one mouse in the DIM group remained negative for another month. In general, although the T. evansi strain was resistant to the standard doses of DIM and ISMM their double doses completely cleared the infection.

Dynamics of blood stage and sporozoite-induced malarial infections in experimentally infected passerines

Complex experimental studies of vertebrate host, vector, and parasite interactions are essential in understanding virulence, but are difficult or impossible to conduct if vector species are unknown. Subinoculation of erythrocytic meronts of avian malarial parasites into susceptible hosts can avoid this problem, but this approach omits early exoerythrocytic stages, e.g. cryptozoites and metacryptozoites, that normally develop from sporozoites. A fundamental question that has remained unanswered is whether blood stage and sporozoite-induced malarial infections lead to differences in the dynamics of parasitemia in acute infections, patterns of parasite development, and host mortality. Here we demonstrate in a Carduelis spinus – Plasmodium relictum (genetic lineage pSGS1) system that experimental infections using inoculation of infected blood and using mosquito bite show similar peaks of parasitemias, but some measures of parasite development in the vertebrate host differ. Infected birds from all groups show decreased activity during the peak of parasitemia. There is no doubt that experimental infections using vectors provide the most precise information about the development of a parasite and its virulence in the host, but experimental infections using blood stages demonstrate similar parasitemias and effects on the host. These results are important for further experimental research of malarial parasites, especially studying avian Plasmodium parasites with unknown vectors.

In vitro and in vivo activity of voriconazole and benznidazole combination on trypanosoma cruzi infection models

Combination therapy has been proposed as an ideal strategy to reduce drug toxicity and improve treatment efficacy in Chagas disease. Previously, we demonstrated potent in vivo anti-Trypanosoma cruzi activity of voriconazole. In this work, we aimed to study the synergistic effect of voriconazole (VCZ) and benznidazole (BZ) both in vitro and in vivo models of T. cruzi infection using the Tulahuen strain. Combining VCZ and BZ at fixed concentrations, the inhibitory concentration 50% (IC50) on amastigotes was lower than the obtained IC50 for BZ alone and the Fractional Inhibitory Concentration Index (∑FIC) suggested an in vitro additive effect on T. cruzi amastigotes inhibition at concentrations devoid of cytotoxic effects. Treatment response in the in vivo model was evaluated by comparing behavior and physical aspects, parasitemia and mortality of mice infected with Tulahuen strain. VCZ and BZ treatments alone or in combination were well tolerated. All treated animals displayed significantly lower mean peak parasitemia and mortality compared to infected non-treated controls (p< 0.05). However, VCZ + BZ combination elicited no additional benefits over BZ monotherapy. VCZ efficacy was not enhanced by combination therapy with BZ at the doses studied, requiring further and astringent non-clinical studies to establish the VCZ efficacy and eventually moving forward to clinical trials.

A Critical Blimp-1-Dependent IL-10 Regulatory Pathway in T Cells Protects From a Lethal Pro-inflammatory Cytokine Storm During Acute Experimental Trypanosoma brucei Infection.

In many infectious diseases, the immune response operates as a double-edged sword. While required for protective immunity, infection-induced inflammation can be detrimental if it is not properly controlled, causing collateral body damage and potentially leading to death. It is in this context that the potent anti-inflammatory cytokine interleukin-10 (IL-10) is required to dampen the pro-inflammatory immune response that hallmarks trypanosomosis. Effective control of this infection requires not just the action of antibodies specific for the parasite's variable surface glycoprotein (VSG) coat antigens, but also a pro-inflammatory immune response mediated mainly by IFNγ, TNF, and NO. However, strict control of inflammation is mandatory, as IL-10-deficient mice succumb from an unrestrained cytokine storm within 10 days of a Trypanosome brucei infection. The relevant cellular source of IL-10 and the associated molecular mechanisms implicated in its trypanosomosis associated production are poorly understood. Using an IL-10 reporter mouse strain (Vert-X), we demonstrate here that NK cells, CD8+ T cells and CD4+ T cells as well as B cells and plasma cells constitute potential cellular sources of IL-10 within the spleen and liver during acute infection. The IL-10 wave follows peak pro-inflammatory cytokine production, which accompanied the control of peak parasitemia. Similar results were observed following conventional experimental needle infection and physiological infections via T. brucei-infected tsetse flies. Our results show that conditional T cell-specific ablation of the IL-10 regulating Prdm1 gene (encoding for the Blimp-1 transcription factor), leads to an uncontrolled trypanosome-induced pro-inflammatory syndrome like the one observed in infected IL-10-deficient mice. This result indicates that the biological role of IL-10-derived from non-T cells, including NK cells, is of minor importance when considering host survival. The cytokine IL-27 that is also considered to be an IL-10 regulator, did not affect IL-10 production during infection. Together, these data suggest that T. brucei activates a Blimp-1-dependent IL-10 regulatory pathway in T cells that acts as a critical anti-inflammatory rheostat, mandatory for host survival during the acute phase of parasitemia.

The many faces of transferrin: Does genotype modulate immune response?

In this study, the expression of pro-inflammatory and iron metabolism genes were analysed under Trypanoplasma borreli (T. borreli) challenge in common carp. Three transferrin (Tf) genotypic groups: two homozygous - DD, GG, and heterozygous DG were intraperitoneally infected with a dose of 2.16 × 105/100 μL parasites. Organ and blood samples were collected at weekly intervals. During the infection period, mortality and parasitaemia were assessed along with measurements of blood iron concentrations and antibody levels. Expression of Tf, Fer, IRP1 and 2, TfR 1a and 1b, Hep, TNF α1 and α2, and IL-1 β was measured in the peak of parasitaemia and the week preceding the peak.Study revealed, that changes in iron blood level induced by parasite were not correlated with the activities of iron homeostasis genes. Neither iron content nor the specific antibody response correlated with survival. We demonstrate that challenged carp, display three distinct, Tf genotype dependent activity patterns of iron homeostasis genes expression. The expected, “classical” way of up-regulation represented homozygous DD individuals. In contrast, GG individuals demonstrated downward trend, while gene expressions of heterozygous DG carp could be defined as an intermediate. We speculate, whether this phenomenon is related to the transferrin molecule itself or to Tf-genotypes being markers of other factors, that influence the iron homeostasis genes activities. We discussed the role of alarmins in triggering the immune response. Distinct genes activating patterns of homozygous genotypes DD and GG had no consequences in terms of mortality rates caused by T.borreli. The highest mortality was observed in the heterozygous group DG.In conclusion, this study suggest that transferrin variant, but not iron blood concentration, has a significant impact on carp immune response to blood parasite infection. This research sheds a new light on the inflammation process and interaction between a host and invaders.

Role of Dual Oxidase 1 in severe malaria

Abstract Despite important advances in malaria control, the disease is still a major public health issue in sub-Saharan Africa. Recent reports indicate over 400,000 deaths annually. Parasite resistance to known antimalarial drugs delays control and elimination. Dual oxidase 1 (Duox1), a member of the NADPH oxidase enzyme family, generates hydrogen peroxide that is used by its partner enzyme lactoperoxidase to generate antimicrobial hypothiocyanite. Malaria parasites are susceptible to hypothiocyanite in vitro. Duox1 is required for the generation of hypothiocyanite in vivo. We hypothesized that malaria parasites are susceptible to the Duox1-mediated antimicrobial system in vivo. Duox1-deficient and wild-type, Duox1-expressing mice were infected with Plasmodium berghei by either intravenous injection of sporozoites or exposure to infected mosquitos. The results indicate that Duox1-deficient mice either infected with intravenous injection of P. berghei sporozoites (p&amp;lt;0.004) or by exposure to P. berghei-infected mosquito bites (p&amp;lt;0.002), have significantly higher blood parasite density at peak parasitemia compared to wild-type mice. Survival data indicated that Duox1-deficient mice are more susceptible to P. berghei infection than control animals (p&amp;lt;0.03 for intravenous infection, p&amp;lt;0.02 for exposure to mosquito bites). Histopathological analysis of brain tissues revealed a slightly higher number of indices of cerebral pathology including cerebrovascular infected red blood cells and hemozoin accumulation, hemostasis, and hemorrhage. Overall, these data propose a novel role of Duox1 in delaying the onset of parasitemia and clinical outcomes in malaria infection.

Accounting for red blood cell accessibility reveals distinct invasion strategies in Plasmodium falciparum strains.

The growth of the malaria parasite Plasmodium falciparum in human blood causes all the symptoms of malaria. To proliferate, non-motile parasites must have access to susceptible red blood cells, which they invade using pairs of parasite ligands and host receptors that define invasion pathways. Parasites can switch invasion pathways, and while this flexibility is thought to facilitate immune evasion, it may also reflect the heterogeneity of red blood cell surfaces within and between hosts. Host genetic background affects red blood cell structure, for example, and red blood cells also undergo dramatic changes in morphology and receptor density as they age. The in vivo consequences of both the accessibility of susceptible cells, and their heterogeneous susceptibility, remain unclear. Here, we measured invasion of laboratory strains of P. falciparum relying on distinct invasion pathways into red blood cells of different ages. We estimated invasion efficiency while accounting for red blood cell accessibility to parasites. This approach revealed different tradeoffs made by parasite strains between the fraction of cells they can invade and their invasion rate into them, and we distinguish “specialist” strains from “generalist” strains in this context. We developed a mathematical model to show that generalist strains would lead to higher peak parasitemias in vivo compared to specialist strains with similar overall proliferation rates. Thus, the ecology of red blood cells may play a key role in determining the rate of P. falciparum parasite proliferation and malaria virulence.

Vaccination accelerates hepatic erythroblastosis induced by blood-stage malaria

BackgroundVaccination induces survival of otherwise lethal blood-stage infections of the experimental malaria Plasmodium chabaudi. Blood-stage malaria induces extramedullary erythropoiesis in the liver. This study investigates how vaccination affects the course of malaria-induced expression of erythrocytic genes in the liver.MethodsFemale Balb/c mice were vaccinated at week 3 and week 1 before challenging with 106P. chabaudi-parasitized erythrocytes. The non-infectious vaccine consisted of erythrocyte ghosts isolated from P. chabaudi-infected erythrocytes. Gene expression microarrays and quantitative real-time PCR were used to compare mRNA expression of different erythrocytic genes in the liver of vaccination-protected and non-protected mice during infections on days 0, 1, 4, 8, and 11 p.i.ResultsGlobal transcriptomics analyses reveal vaccination-induced modifications of malaria-induced increases in hepatic gene expression on days 4 and 11 p.i. On these days, vaccination also alters hepatic expression of the erythropoiesis-involved genes Ermap, Kel, Rhd, Rhag, Slc4a1, Gypa, Add2, Ank1, Epb4.1, Epb4.2, Epb4.9, Spta1, Sptb, Tmod1, Ahsp, Acyp1, Gata1, Gfi1b, Tal1, Klf1, Epor, and Cldn13. In vaccination-protected mice, expression of these genes, except Epb4.1, is significantly higher on day 4 p.i. than in un-protected non-vaccinated mice, reaches maximal expression at peak parasitaemia on day 8 p.i., and is slowed down or even decreased towards the end of crisis phase on day 11 p.i.. After day 1 p.i., Epor expression takes about the same course as that of the other erythroid genes. Hepatic expression of Epo, however, is delayed in both vaccinated and non-vaccinated mice for the first 4 days p.i. and is maximal at significantly higher levels in vaccinated mice on day 8 p.i., before declining towards the end of crisis phase on day 11 p.i.ConclusionThe present data indicate that vaccination accelerates malaria-induced erythroblastosis in the liver for 1–2 days. This may contribute to earlier replenishment of peripheral red blood cells by liver-derived reticulocytes, which may favour final survival of otherwise lethal blood-stage malaria, since reticulocytes are not preferred as host cells by P. chabaudi.

A comparative study on susceptibility, course of parasitemia, and pattern of infection with Trypanosoma evansi between different laboratory animals

This study investigated the susceptibility of eight different animal species (rat, mice, guinea pigs, hamsters, rabbits, sheep, goats, and chicken) experimentally infected with Trypanosoma evansi isolated from camels. In all laboratory animals, the number of trypanosomes was standardized according to the weight of the animal, and daily examination of the blood was conducted to assess the presence of trypanosomes to determine the prepatent period and the peak of parasitemia. Results suggested that mice and rats were the most susceptible laboratory animals to infection, whereas hamsters and guinea pigs displayed a certain degree of tolerance to infection. Rabbits exhibited a chronic course of infection, but the level of parasitemia was very low. In sheep and goats, trypanosomes were detectable only by subpassage to rats and mice, whereas all the chickens remained uninfected even with increasing doses of infection.

Using two phases of the CD4 T cell response to blood-stage murine malaria to understand regulation of systemic immunity and placental pathology in Plasmodium falciparum infection.

Plasmodium falciparum infection and malaria remain a risk for millions of children and pregnant women. Here, we seek to integrate knowledge of mouse and human T helper cell (Th) responses to blood-stage Plasmodium infection to understand their contribution to protection and pathology. Although there is no complete Th subset differentiation, the adaptive response occurs in two phases in non-lethal rodent Plasmodium infection, coordinated by Th cells. In short, cellular immune responses limit the peak of parasitemia during the first phase; in the second phase, humoral immunity from T cell-dependent germinal centers is critical for complete clearance of rapidly changing parasite. A strong IFN-γ response kills parasite, but an excess of TNF compared with regulatory cytokines (IL-10, TGF-β) can cause immunopathology. This common pathway for pathology is associated with anemia, cerebral malaria, and placental malaria. These two phases can be used to both understand how the host responds to rapidly growing parasite and how it attempts to control immunopathology and variation. This dual nature of T cell immunity to Plasmodium is discussed, with particular reference to the protective nature of the continuous generation of effector T cells, and the unique contribution of effector memory T cells.

COMPARISON OF PATHOGENESIS OF P. BERGHEI INFECTION IN MOUSE AND RAT MODELS

Background: The cytokine cascade in the immunopathogenesis of malaria infection had been widely studied. However, their specific association with survival and severe infection remained obscure. Methods: The study investigated the cytokine profiles and histopathological features of malaria in the severe infection and survival models by using male ICR mice and male Sprague Dawley rats respectively. Results: The severe model, the infected ICR mice, exhibited a high parasitemia with 100% mortality after peak parasitemia at day 5 post-infection. The survival model, the infected Sprague Dawley rats, showed mild parasitemia with full recovery by day 14 of infection. Both severe and survival models showed similar histopathological severity during peak parasitemia. The severe model produced highly elevated levels of proinflammatory cytokines, TNF-I± and IL-1I±, and low levels of the anti-inflammatory cytokine, IL-4; while the survival model showed low levels of TNF-I± and IL-1I± with high levels of IL-4. Conclusion: There were differences in the pathogenesis of the severe and survival models of malaria infection. These could be a basis for immunotherapy of malaria in the future.

Antimalarial Activity of Human Group IIA Secreted Phospholipase A2 in Relation to Enzymatic Hydrolysis of Oxidized Lipoproteins.

The level of human group IIA secreted phospholipase A2 (hGIIA sPLA2) is increased in the plasma of malaria patients, but its role is unknown. In parasite culture with normal plasma, hGIIA is inactive against Plasmodium falciparum, contrasting with hGIIF, hGV, and hGX sPLA2s, which readily hydrolyze plasma lipoproteins, release nonesterified fatty acids (NEFAs), and inhibit parasite growth. Here, we revisited the anti-Plasmodium activity of hGIIA under conditions closer to those of malaria physiopathology where lipoproteins are oxidized. In parasite culture containing oxidized lipoproteins, hGIIA sPLA2 was inhibitory, with a 50% inhibitory concentration value of 150.0 ± 40.8 nM, in accordance with its capacity to release NEFAs from oxidized particles. With oxidized lipoproteins, hGIIF, hGV, and hGX sPLA2s were also more potent, by 4.6-, 2.1-, and 1.9-fold, respectively. Using specific immunoassays, we found that hGIIA sPLA2 is increased in plasma from 41 patients with malaria over levels for healthy donors (median [interquartile range], 1.6 [0.7 to 3.4] nM versus 0.0 [0.0 to 0.1] nM, respectively; P < 0.0001). Other sPLA2s were not detected. Malaria plasma, but not normal plasma, contains oxidized lipoproteins and was inhibitory to P. falciparum when spiked with hGIIA sPLA2 Injection of recombinant hGIIA into mice infected with P. chabaudi reduced the peak of parasitemia, and this was effective only when the level of plasma peroxidation was increased during infection. In conclusion, we propose that malaria-induced oxidation of lipoproteins converts these into a preferential substrate for hGIIA sPLA2, promoting its parasite-killing effect. This mechanism may contribute to host defense against P. falciparum in malaria where high levels of hGIIA are observed.

Experimental characterization of the complete life cycle of Haemoproteus columbae, with a description of a natural host-parasite system used to study this infection

Characterization of complete life cycles of haemoparasites requires the maintenance of suitable susceptible vertebrate hosts and vectors for long periods in captivity, in order to follow the complete parasitic cycle in definitive and intermediate hosts. Currently, there are few host-parasite models established in avian haemosporidian research, and those have been developed mainly for species of Passeriformes and their parasites. This study aimed to develop an experimental methodology to access the complete life cycle of Haemoproteus columbae (cytb lineage HAECOL1), which parasitizes the Rock Pigeon (Columba livia) and louse fly (Pseudolynchia canariensis). A colony of louse flies, which are the natural vectors of this parasite, was established. Thirty newly emerged insects were exposed to H. columbae infection and used to infect naïve Rock Pigeons. The peak of parasitaemia (acute stage) was seen between 27 and 32 days p.i. when up to 70.8% of red blood cells were infected. The crisis occurred approximately 1 week after the peak, and the long-lasting chronic parasitaemia stage followed. Exo-erythrocytic meronts were seen mainly in the lungs where extensive tissue damage was reported, but also in the kidneys and spleen. In the vector, the sporogonic cycle of H. columbae was completed between 13 and 16 days p.i., at an average temperature ranging between 12 and 15 °C. This host-parasite model is tractable for maintenance in captivity. It is recommended for use in studies aiming for detailed characterization of host-parasite relationships in areas such as physiology, pathology, immunobiology, genetics, as well as for evaluative treatments and to follow the infection in any stage of parasite development both in the vertebrate or invertebrate host.

CASE SERIES OF BABESIOSIS AT A SINGLE CENTER

SESSION TITLE: Wednesday Abstract Posters SESSION TYPE: Original Investigation Posters PRESENTED ON: 10/23/2019 09:45 AM - 10:45 AM PURPOSE: The purpose of this study was to review trends in the management of cases of babesiosis in a single tertiary care hospital. We also try to elucidate clinical factors that could predict outcomes METHODS: Billing ICD-9 (088.82) & 10(B60.0) codes for babesia from 2011 to 2017 were used to select patients. Only inpatient confirmed cases per CDC Babesiosis 2011 case definition criteria (presence of parasitemia and the appropriate objective & subjective clinical criteria) were included. The electronic medical record was accessed for demographics, relevant labs, interventions & outcome data. The statistical analysis was done for subgroup of patients requiring intensive care RESULTS: 85 patients had a confirmed diagnosis of babesiosis. Of these, 39 required hospitalization. These patients were predominantly male (71% n=27) & white (89.5% n=34), (remaining were African American & Latinos). The median age was 70 years. The most common comorbid conditions were hypertension (66% n=25) & heart disease (i.e heart failure, coronary artery disease & chronic arrhythmias) (58% n=22). Malignancy (n=8), COPD (n=8), CKD(n=4) & Blood dyscrasias (n=6) were less common.Co-infections were present in 42%, (n=16) namely Lyme(31%, n=12), Ehrilica(n=2), Anaplasma(n=2) & Powassan virus(n=1). 44% (n=17) presented with fever, the median temperature was 37.85°C. Median peak parasitemia among all patients was 3.85% (Range 0.05-13%), & median parasitemia among patients admitted to ICU was 8% (Range: 3%-23%). This difference in means between ICU & non-ICU patients using the unpaired t-test was significant (p=0.0122). The median age of non-ICU admission was lower compared to the ICU median age of 68 yers vs 84 years. This was also very significant (p=0.0091). Using the unpaired t-test the following lab parameters were significantly different between ICU & non-ICU patients: LDH (p=0.0103), AST (p=0.0002) and TBilli (p=0.0039) while Hemoglobin (p=0.2350 ), Platelets (p= 0.3973 ), ALT (p=0.149) were not. The median hospital stay was 4.5 days with ICU patients staying statistically longer (Median 7 vs 4 days, p=0.022) Of all patients, 29% (n=10) required intensive care. 11 required exchange transfusion, 8 of which were in ICU (p=0.001, Fisher Exact test). One patient died from multiorgan failure & septic shock, representing a mortality rate of 2.6%. Most patients received atovaquone (n=37) usually with Azithromycin (n=33). Other agents used in combination, were Clindamycin(n=16),Doxycycline(n=25) & Quinine(n=16) CONCLUSIONS: Babesiosis is a geographically limited rare disease which can result in serious illness requiring intensive care. Our study suggests that age, AST & LDH on presentation and peak parasitemia are factors that may indicate need for ICU admission CLINICAL IMPLICATIONS: Early recognition of clinical risk factors of severe disease can help effectively triage and treat patients with Babesiosis DISCLOSURES: No relevant relationships by Scott Beegle, source=Web Response No relevant relationships by Abdul Moiz Khan, source=Web Response No relevant relationships by Muhammad Salick, source=Web Response No relevant relationships by Ali Wazir, source=Web Response No relevant relationships by Robert Wilcott, source=Web Response

Fluctuations of Spleen Cytokine and Blood Lactate, Importance of Cellular Immunity in Host Defense Against Blood Stage Malaria Plasmodium yoelii.

Our previous studies of protective immunity and pathology against blood stage malaria parasites have shown that not only CD4+ T cells, but also CD8+ T cells and macrophages, are important for host defense against blood stage malaria infection. Furthermore, we found that Plasmodium yoelii 17XNL (PyNL) parasitizes erythroblasts, the red blood cell (RBC) precursor cells, which then express MHC class I molecules. In the present study, we analyzed spleen cytokine production. In CD8+ T cell-depleted mice, IL-10 production in early stage infection was increased over two-fold relative to infected control animals and IL-10+ CD3− cells were increased, whereas IFN-γ production in the late stage of infection was decreased. At day 16 after PyNL infection, CD8+ T cells produced more IFN-γ than CD4+ T cells. We evaluated the involvement of the immunoproteasome in induction of immune CD8+ T cells, and the role of Fas in protection against PyNL both of which are downstream of IFN-γ. In cell transfer experiments, at least the single molecules LMP7, LMP2, and PA28 are not essential for CD8+ T cell induction. The Fas mutant LPR mouse was weaker in resistance to PyNL infection than WT mice, and 20% of the animals died. LPR-derived parasitized erythroid cells exhibited less externalization of phosphatidylserine (PS), and phagocytosis by macrophages was impaired. Furthermore, we tried to identify the cause of death in malaria infection. Blood lactate concentration was increased in the CD8+ T cell-depleted PyNL-infected group at day 19 (around peak parasitemia) to similar levels as day 7 after infection with a lethal strain of Py. When we injected mice with lactate at day 4 and 6 of PyNL infection, all mice died at day 8 despite demonstrating low parasitemia, suggesting that hyperlactatemia is one of the causes of death in CD8+ T cell-depleted PyNL-infected mice. We conclude that CD8+ T cells might control cytokine production to some extent and regulate hyperparasitemia and hyperlactatemia in protection against blood stage malaria parasites.

Trypanosoma brucei brucei causes a rapid and persistent influx of neutrophils in the spleen of infected mice.

Trypanosomosis is a chronic parasitic infection, affecting both humans and livestock. A common hallmark of experimental murine infections is the occurrence of inflammation and the associated remodelling of the spleen compartment. The latter involves the depletion of several lymphocyte populations, the induction of T‐cell‐mediated immune suppression, and the activation of monocyte/macrophage cell populations. Here, we show that in experimental T b brucei infections in mice, these changes are accompanied by the alteration of the spleen neutrophil compartment. Indeed, mature neutrophils are rapidly recruited to the spleen, and cell numbers remain elevated during the entire infection. Following the second peak of parasitemia, the neutrophil cell influx coincides with the rapid reduction of splenic marginal zone (MZ)B and follicular (Fo)B cells, as well as CD8+ T and NK1.1+ cells, the latter encompassing both natural killer (NK) and natural killer T (NKT) cells. This report is the first to show a comprehensive overview of all alterations in spleen cell populations, measured with short intervals throughout the entire course of an experimental T b brucei infection. These data provide new insights into the dynamic interlinked changes in spleen cell numbers associated with trypanosomosis‐associated immunopathology.

A non-lethal malarial infection results in reduced drug metabolizing enzyme expression and drug clearance in mice

BackgroundGiven the central importance of anti-malarial drugs in the treatment of malaria, there is a need to understand the effect of Plasmodium infection on the broad spectrum of drug metabolizing enzymes. Previous studies have shown reduced clearance of quinine, a treatment for Plasmodium infection, in individuals with malaria.MethodsThe hepatic expression of a large panel of drug metabolizing enzymes was studied in the livers of mice infected with the AS strain of Plasmodium chabaudi chabaudi, a nonlethal parasite in most strains of mice with several features that model human Plasmodium infections. C57BL/6J mice were infected with P. chabaudi by intraperitoneal injection of infected erythrocytes and sacrificed at different times after infection. Relative hepatic mRNA levels of various drug metabolizing enzymes, cytokines and acute phase proteins were measured by reverse transcriptase-real time PCR. Relative levels of cytochrome P450 proteins were measured by Western blotting with IR-dye labelled antibodies. Pharmacokinetics of 5 prototypic cytochrome P450 substrate drugs were measured by cassette dosing and high-resolution liquid chromatography-mass spectrometry. The results were analysed by MANOVA and post hoc univariate analysis of variance.ResultsThe great majority of enzyme mRNAs were down-regulated, with the greatest effects occurring at the peak of parasitaemia 8 days post infection. Protein levels of cytochrome P450 enzymes in the Cyp 2b, 2c, 2d, 2e, 3a and 4a subfamilies were also down-regulated. Several distinct groups differing in their temporal patterns of regulation were identified. The cassette dosing study revealed that at the peak of parasitaemia, the clearances of caffeine, bupropion, tolbutamide and midazolam were markedly reduced by 60–70%.ConclusionsThese findings in a model of uncomplicated human malaria suggest that changes in drug clearance in this condition may be of sufficient magnitude to cause significant alterations in exposure and response of anti-malarial drugs and co-medications.

Evaluation of the inhibitory effect of ivermectin on the growth of Babesia and Theileria parasites in vitro and in vivo

BackgroundTreatment is the principle way to control and eliminate piroplasmosis. The search for new chemotherapy against Babesia and Theileria has become increasingly urgent due to parasite resistance to current drugs. Ivermectin (IVM) was the world’s first endectocide, capable of killing a wide variety of parasites and vectors, both inside and outside the body. It is currently authorized to treat onchocerciasis, lymphatic filariasis, strongyloidiasis, and scabies. The current study documented the efficacy of IVM on the growth of Babesia and Theileria in vitro and in vivo.MethodsThe fluorescence-based assay was used for evaluating the inhibitory effect of IVM on four Babesia species, including B. bovis, B. bigemina, B. divergens, B. caballi, and Theileria equi, the combination with diminazene aceturate (DA), clofazimine (CF), and atovaquone (AQ) on in vitro cultures, and on the multiplication of a B. microti-infected mouse model. The cytotoxicity of compounds was tested on Madin–Darby bovine kidney (MDBK), mouse embryonic fibroblast (NIH/3 T3), and human foreskin fibroblast (HFF) cell lines.ResultsThe half-maximal inhibitory concentration (IC50) values determined for IVM against B. bovis, B. bigemina, B. divergens, B. caballi, and T. equi were 53.3 ± 4.8, 98.6 ± 5.7, 30.1 ± 2.2, 43.7 ± 3.7, and 90.1 ± 8.1 μM, respectively. Toxicity assays on MDBK, NIH/3 T3, and HFF cell lines showed that IVM affected the viability of cells with a half-maximal effective concentration (EC50) of 138.9 ± 4.9, 283.8 ± 3.6, and 287.5 ± 7.6 μM, respectively. In the in vivo experiment, IVM, when administered intraperitoneally at 4 mg/kg, significantly (p < 0.05) inhibited the growth of B. microti in mice by 63%. Furthermore, combination therapies of IVM–DA, IVM–AQ, and IVM–CF at a half dose reduced the peak parasitemia of B. microti by 83.7%, 76.5%, and 74.4%, respectively. Moreover, this study confirmed the absence of B. microti DNA in groups treated with combination chemotherapy of IVM + DA and IVM + AQ 49 days after infection.ConclusionsThese findings suggest that IVM has the potential to be an alternative remedy for treating piroplasmosis.

Route of inoculation influences Trypanosoma congolense and Trypanosoma brucei brucei virulence in Swiss white mice.

Experiments on infections caused by trypanosomes are widely performed in Swiss white mice through various inoculation routes. To better understand the effect of route of trypanosome inoculation on disease outcomes in this model, we characterised the virulence of two isolates, Trypanosoma brucei KETRI 2710 and T. congolense KETRI 2765 in Swiss white mice. For each of the isolates, five routes of parasite inoculation, namely intraperitoneal (IP), subcutaneous (SC), intramuscular (IM) intradermal (ID) and intravenous (IV) were compared using groups (n = 6) of mice, with each mouse receiving 1x104 trypanosomes. We subsequently assessed impact of the routes on disease indices that included pre-patent period (PP), parasitaemia levels, Packed Cell Volume (PCV), bodyweight changes and survival time. Pre-patent period for IP inoculated mice was a mean ± SE of 3.8 ± 0.2 and 6.5 ± 0.0 for the T brucei and T. congolense isolates respectively; the PP for mice groups inoculated using other routes were not significantly different(p> 0.05) irrespective of route of inoculation and species of trypanosomes. With ID and IP routes, parasitaemia was significantly higher in T. brucei and significantly lower in T. congolense infected mice and the progression to peak parasitaemia routes showed no significant different between the routes of either species of trypanosome. The IM and ID routes in T. congolense inoculations, and IP and IV in T. b. brucei induced the fastest and slowest parasitaemia progressions respectively. There were significant differences in rates of reduction of PCV with time post infection in mice infected by the two species and which was more pronounced in sc and ip injected mice. No significant differences in mice body weight changes and survivorship was observed between the routes of inoculation. Inoculation route therefore appears to be a critical determinant of pathogenicity of Trypanosoma congolense and Trypanosoma brucei brucei in murine mouse model of African trypanosomiasis.

Impaired epididymal function of albino rats infected and treated with Trypanosoma brucei brucei and Trypanosoma congolense

The effect of trypanosomosis (Trypanosoma brucei brucei and Trypanosoma congolense infection) and diminazene aceturate treatment on the epididymal function of male albino rats was investigated. Twenty-five adult male albino rats were used for this study. They were randomly assigned into 5 groups of 5 rats each. Group 1 was the uninfected untreated group while groups 2 and 3 were infected with 5 × 105T. congolense. Groups 4 and 5 were infected with 5 × 105T. brucei brucei. Rats in groups 2 and 4 were humanely sacrificed at the peak of parasitemia, i.e., on days 7 and 14 PI respectively and their epididymides dissected out for the enumeration of epididymal sperm cells. Groups 3 and 5 rats were treated with 3.5 mg/kg and 7 mg/kg diminazene aceturate (Tryponil®) respectively and later sacrificed at week 7 of the experiment. The epididymides of the rats were also dissected out and epididymal sperm reserves (ESR) of all the rats enumerated. Hyperacute and acute trypanosomosis were observed following T. congolense and T. brucei brucei infections respectively. The uninfected control group had significantly higher (P ˂ 0.05) ESR compared with the infected groups with the group 3 rats having the least ESR, thus implying impaired epididymal function. There was an improvement (P ˂ 0.05) in the ESR of group 5 rats compared with the group 4 rats while treatment was of no effect in the T. congolense–treated group. It was therefore concluded that trypanosomal infection adversely impaired epididymal function. Resolution of epididymal function in T. brucei brucei–infected rats may be possible following treatment with diminazene aceturate.