Effect of biotherapy T. cruzi 7x in several therapeutic schemes on experimental infection by Trypanosoma cruzi

Background: Biotherapy is used against infectious diseases treatment and prophylaxis and has been investigated by many researchers [1,2]. Aim: Assess the effect of biotherapy 7x T. cruzi on several treatment schemes, upon experimental infection by T. cruzi. Methodology: A blind, controlled and randomized by drawing experiment was performed. Male Swiss mice, four weeks old were utilized. Groups evaluated: IC – Infection Control (treated with water – 9 animals); TBBA7x3days – Treated with biotherapy 7x 3 days before and 3 days after infection (5 animals); TBB7x3days – Treated with 7x biotherapy 3 days before infection (5 animals); TBBAI7x3days – Treated with 7x biotherapy 3 days before infection and after infection indefinitely (6 animals). Animals were inoculated intraperitoneally with 1400 blood trypomastigotes Y strain. Biotherapy: prepared according to Farmacopéia Homeopática Brasileira [3]. Treatment plan: offered ad libitum, in the water (10µL/mL). Parasitological parameters: parasitemia was assessed according Brener’s technique. [4]. Clinical parameters: body hair aspect, edema, movement, diarrhea, body weight, temperature, food and water intake. Ethics: Registration 030/2008 UEM Ethics Committee for Experiments in Animals. Statistical analysis: was performed using the tests Kruskal Wallis and Mann-Whitney testes, significance 5%. Results: The best effect obtained was with the TBBA7x3days, both for clinical and parasitological parameters. It was expressed by lower parasitemia curve (p=0.04) and decrease of patent period tendency, of total parasitemia, of mortality and survival of the animals increase (Table 1). Evolution of parasitemia was distinct for the several treatment schemes. Survival of at least one mouse by treated groups is an extremely important data, since Y strain causes 100% mortality in Swiss mice. TBBAI7x3days group showed begger tendency in raising total parasitemia compared with IC. Although it might have occurred, this group presented 80% mortality rate compared with other groups. Animals from TBBA7x3days also showed better evolution of weight body, temperature, food (p=0.078-10%) and water intake, body hair aspect and edema development. Diarrhea and paralysis were only observed in IC group mice, highlighting the biotherapy use benefits. Conclusions: Best effect was obtained TBBA7x3days, both for clinical and parasitological parameters. It’s possible to speculate that in this regimen, biotherapy was able to modulate, more effectively, the host’s immune system, decreasing the number of parasites.

Evaluation of biotherapies T.cruzi 15x, 16x, 17x and "potency chords" in experimental infection by Trypanosoma cruzi.

Background: The biotherapies are drugs widely utilized against infectious diseases. Biotherapies’ profylatic and therapeutic action against Chagas Disease is currently being investigated, but it is needed to develop further controlled experiments “in vivo”, which could define more clearly: dilution, dose, time of use and, if possible, the action mechanisms of these ultradiluted medicaments [1,2]. Aim: Evaluate the effect biotherapies T. cruzi 15x, 16x, 17x and “potency chords”, on experimental infection by T. cruzi. Methodology: A blind, controlled and randomized by drawing test was performed. Animals: 29 male Swiss mice, four weeks old were utilized. The animals were kept at Parasitology Vivarium/State University of Maringá (UEM), in ideal conditions of temperature (22±2)ºC and photoperiod (light / dark cycle 12h). Mice have been inoculated intraperitoneally with 1400 blood trypomastigotes Y strain and divided in groups: IC – Infection control (treated with distilled water – 9 animals); TBBA15x3days – Treated with biotherapy 15x 3 days before and 3 days after infection (5 animals); TBBA16x3days – Treated with biotherapy 16x 3 days before and 3 days after infection (5 animals); TBBA17x3days – Treated with biotherapy 17x 3 days before and 3 days after infection (5 animals); TBBAChords3days – Treated with biotherapy 15x, 16x, 17x “potency chords”, 3 days before and 3 days after infection (5 animals). Biotherapies: prepared by a homeopathic pharmacist from UEM, according to Farmacopéia Homeopática Brasileira [3]. Biotherapies treatment schedule: diluted in distilled water (10µL/mL in ambar bottles – renewed each two days) offered ad libitum, 3 days before and 3 days after infection in all groups. Parasitological parameters: parasitemia was assessed from infection until death, according to Brener’s technique [4] with 5µL of blood collected from the tail vein and examined in optical microscope. Pre-patent period, patent period, total parasitemia, survival and morbidity were obtained from the parasitemia curve. Clinical parameters: Visually assessed (presence or absence): body hair aspect (bristling), edema, movement and diarrhea. Measured: body weight, temperature, food and water intake. Ethics: This study has been approved by the UEM Ethics Committee for Experiments in Animals - Registration 030/2008. Statistical analysis: was performed using the tests Kruskal Wallis and Mann-Whitney tests, significance of 5%. Results: There was not statistical difference between total parasitemia of the groups treated with biotherapies and the IC group (p=0.6819). The parasitemia curve of group TBBAChords3days was greater then the IC (p=0.0418). Despite this increase, patent period and mortality both showed a decreasing tendency, while pre patent period and survival time increased (p=0.373). The same tendency results were observed for TBBA17x3days results (Table 1). Survival of at least one mice in groups TBBA17x3days and TBBAChords3days is worthy of discussion, since Y strain causes 100% mortality in these experimental conditions. Groups TBBA17x3days and TBBAChords3days showed better evolution than IC group for body weight, temperature, food and water intake (p=0.05), body hair aspect and edema developing. Diarrhea and hind legs paralysis were only observed in mice belonging to groups IC and TBAA16x3days. Conclusions: Superior effect was obtained with biotherapies 17x and “Potency Chords”, both for clinical and parasitological parameters. “Potency chords” has proper effect which distinguishes it from the individual effects of the dilutions that compound it.

Causticum hahnemanni, Conium maculatum and Lycopodium clavatum highly diluted medications decreases parasitemia in mice infected by Trypanosoma cruzi

Introduction: Benznidazole is the only medicine available in Brazil for Chagas’ disease treatment, however it presents low efficacy in the chronic phase and several adverse effects [1-3]. Aim: Evaluate the effect of Causticum hahnemanni, Conium maculatum and Lycopodium clavatum [4-6] administered to mice infected with T. cruzi. Method: In blind randomized controlled trial 42 male Swiss mice, 8 weeks of age, have been grouped: GCaus –treated with C. hahnemanni 13cH (n=10), GCon –treated with C. maculatum 13 cH (n=11), GLy –treated with L. clavatum13cH (n=10) and CG – control group (n=11) treated with 7% hydro alcoholic solution 13cH. The animals were infected intraperitoneally with 1.400 blood trypomastigotes of T. cruzi - Y strain. Medications were been prepared according to Brazilian Homeopathic Pharmacopoeia [7]. Medication was diluted in water (1mL/100mL) offered ad libitum, from amber recipient during 16 hours administered 48 hours before infection and 48, 96 and 144 hours after inoculation. Parasitological parameters assessed: total parasitemia (TP), maximum peak of parasites (MPP), pre-patent period (PPP) and area under curve (AUC). Parasitemia was evaluated daily counting from the first day of infection. Clinical parameters assessed: weight, temperature, water and food intake and excreta were measured counting from the 5º day before infection until animals’ death or checking negative parasitemia for 3 consecutive days. Mortality was registered for 75 days after infection. Ethics Committee for Experiments in Animals gave approval UEM 054/11. Statistical comparison of data was performed with Kruskal-Wallis test, with 5% significance. Results and Discussion: The diluted medications have been significantly reduced the parasitological parameters: MPP (p

Biotherapy of mice’s serum 13cH modifies parasitological and immunological parameters of Trypanosoma cruzi infection.

Background: T. cruzi biotherapies’ alter the infection course by this protozoan [1,2], fact that encourages the evaluation of other highly diluted medicines which modulates host’s immune system. Aim: Evaluate effect of biotherapy produced from mices’s serum in experimental infection by T. cruzi. Methodology: A blind, randomized and controlled study was performed. Animals: 60 male Swiss mice, four weeks old were inoculated intraperitoneally with 1400 trypomastigotes-Y strain and divided: MNI: mice non-infected by T. cruzi; IC: treated with hydroalcoholic solution 7%; BSI13cH: treated with mice’s serum infected by T. cruzi 13cH. Biotherapies: produced from mice’s serum infected by T. cruzi in 13cH dynamization [3]. Treatment: mice were treated 48 hours before and after infection. Subsequently animals were treated 56/56 hours until 9th day of infection (d.i). The medicine was diluted in water (1/100mL) and offered ad libitum, for 16 hours. Parasitological parameters: were evaluated pre-patent and patent period, parasitemia peak, total parasitemia and mortality [4]. Cytokine dosage: IL-4, IL-17, TNF-α and IFN-γ were measured in serum on 0, 8th and 12th d.i., by enzyme immunoassay. Ethics: study was approved by Ethics Committee for Experiments in Animals/UEM. Statistic: data were compared with Mann Whitney or Student t test, significance 5%. Results: BSI13cH showed tendency to increase total parasitemia (p=0.06) and parasitemia peak (p=0.05), with lower patent period (p=0.03) and higher mortality (p=0.03) compared to IC. In dosage of cytokines BSI13cH group showed on 0 d.i. a decrease in IL-17 (p = 0.02) and increased IL-4 (p = 0.01) compared to MNI (baseline value), probably caused by modulation of medication administration 48 hours before infection. IL-17 concentration didn’t vary throughout the infection to BSI13cH, different IC that tended to decrease concentration of cytokine on 8th d.i. IL-4 increased significantly on 0 d.i. to BSI13cH, with subsequent return to baseline values. IC group didn’t change significantly IL-4 value along the infection. IFN-γ concentration on 12th d.i. to BSI13cH was lower (p = 0.00) than IC, which increased this cytokine on 8th and 12th di. TNF-α concentration of BSI13cH followed the same evolution as IC, with an increase on 8th and 12th d.i. (Figure 1). The medicine seems initially promote Th2 response (IL-4), hindering the development of effective Th1 response (INF-γ), causing an increase of parasitemia and animals' death. Conclusions: BSI13cH demonstrated effect in experimental infection by T. cruzi with increased parasitemia, animals’ premature death and modulated immune response differently of IC.

Zincum metallicum 5cH increases survival and improves clinical mice infected with Trypanosoma cruzi

The Multicenter International Project suggests Zincum Mettalicum high diluted as object of study in different experimental models. Aim: evaluate the effect of substance high diluted Zincum metallicum in murine experimental infection by Trypanosoma cruzi. Metodology: was performed a blind, controlled, randomized, using 60 swiss male mice, 56 days old, divided into groups: CNI - uninfected and untreated animals; CI - infected and untreated animals; infected and treated animals: ZN5cHTA - Zinc 5ch and LAC5cHTA - Lactose 5ch , 48 hours before and after infection, subsequently were treated 56/56 hours until 9th day of infection; ZN5cHTTD - Zinc 5Ch and LAC5cHTTD - Lactose 5cH, everyday from the 4th of infection. Animals were inoculated with 1.400 blood trypomastigotes, strain Y-T. cruzi, intraperitoneally. Medicines were handled, prepared in grain alcohol 70%, and dynamized up to 100 times until 4cH. To obtain the 5cH it was used bi-distilled sterilized water filtered in membrane - 0.22 µm [1], on separate days (first Lactose and then Zinc) and stored in different rooms. Microbiological test in vivo and toxicity were made in accordance with current legislation [2].Test solutions were diluted in water at a concentration of 10% (1mL/100mL) after dilution in water. Clinical (temperature, weight, water/foodintake and excreta)[3] and parasitological parameters (pre-patent and patent period, peak parasitemia, and parasitemia overall survival time)[4] were assessed daily. Data were compared BioEstat 5.0, significance level of 5%. Project was approved by the Ethics Committee on Animal Use in Experimentation of the Universidade Estadual de Maringa by opinion number 025/2014. Results: ZN5cHTA group had a higher survival rate than their control LAC5cHTA (p=0.004). ZN5cHTA shows 55.7% probability of surviving to the 15th day after infection, while LAC5cHTA 29.4%. ZN5cHTA also provides significantly better performance (p= 0.0206) compared to CI, contrary to what occurs with LAC5cHTA x CI (p=0.7410). There is no significant difference in survival between the different treatments schemes TA and TTD, either with ZN5cH (p=0.0754) or LAC5cHTA (p=0.9480), although the best ZN5cHTA present trend toward benefit. Considering parasitological parameters ZN5cHTA group had higher pre-patent period (PPP) meaning benefit to infected animals [5]. Although ZN5cHTA shows greater number of parasites from 6th to 11th day of infection and right shift of parasitemia peak in relation to LAC5cHTA (p=0.020), this group displayed a better performance compared to the other groups as observed in other models [6]. Conclusion: ZN5cHTA group had higher survival, greater pre-patent period and better clinical outcome compared to control LAC5cHTA and to other groups. This result may be related to higher total parasitemia and alterations in the parasite cycle time observed in this group. These findings suggest aggravation with posterior benefit as reported in some cases of homeopathic treatment.

Clinical and parasitological assessment in mice treated with highly diluted Atropa belladonna

Introduction: The infection by Trypanosoma cruzi is a public health problem and there is no effective treatment currently. Immunomodulatory effects of Atropa belladonna may offer benefits1 Objective: To evaluate the effect of A. belladonna in murine infection by T. cruzi. Methodology: The experiment was blind, controlled and randomized by draw. Eighty five Swiss male mice, at 8 weeks of age, were infected with 1400 blood trypomastigotes of T. cruzi Y strain (via IP) and divided into the following groups: without treatment (CI), treated with the mother tincture of A. belladonna (GTM-HN Cristiano), treated with A. belladonna 5cH (G5cH), treated with A. belladonna 6cH (G6cH), treated with A. belladonna 30cH (G30cH). Cereal alcohol 70 ° GL was used for dilutions as well as water in final preparations (Sigma-SP-Brazil). Oral treatment, diluted with water (1mL/100mL water), offered ad libitum 48 hours before infection, available during 16h. After infection, treatment of 56/56h for 16h, until the 9th day of infection2. Parasitological parameters: Curve of parasitemia, total parasitemia (PT), Maximum Peak of Parasites (PMP), Pre-Patent Period (PPP), Patent Period (PP), and Survival. Clinical parameters: water, food, excreta, weight and temperature. Results: G6cH and G30cH groups displayed better survival rates (1.54 and 1.42 times versus IC), and higher curve of parasitemia - G6cH (p = 0.00), G30cH (p = 0.02) – when compared to CI. PMP was lower in GTM (P = 0.01) and G5cH (p = 0.04) groups; PT was lower in GTM (p = 0.01), G5cH (p = 0.05) and G6cH (p = 0.05) groups when compared to CI. There was no difference in PPP and PP parameters in all groups, with a tendency of a higher PPP in G5cH and G6cH groups and lower PPP in G30cHgroup.The mice weight was higher in GTM (

Evaluation of effects of inert vehicles proceeding from homeopathic preparation in mice’s experimental infection by Trypanosoma cruzi

Background: In experiments with homeopathic medicines is important to test the inert vehicle from succussed preparations for the treatment control. Aim: To evaluate the effect of hydro-alcoholic solutions 1cH, 6cH and 30cH in mice’s experimental infection with Trypanosoma cruzi. Methodology: In a blind, randomized, controlled test, two independent experiments with 34 and 51 swiss male mice, 8 weeks old, kept in cages micro acclimated, infected with 1400 blood trypomastigotes of the Y strain of T. cruzi (via IP), were divided: IC-untreated control; G1cH-received hydro-alcoholic solution dynamized 1cH; G6cH-received hydro-alcoholic solution dynamized 6cH and G30cH-received hydro-alcoholic solution dynamized 30cH. The solutions were prepared according to Brazilian Homeophatic Pharmacopoeia1 with alcohol 70 ° GL. Final preparations (1cH, 6cH and 30cH) were manipulated with water (Sigma-SP-Brazil). The treatment was offered diluted with water (1/100mL) ad libitum 48 hours before infection, available for 16h. After infection, the animals were treated 56h/56h for 16h until the 9th day of infection. The parasitological parameters were analyzed: Curve of Parasitemia, Total Parasitemia (TP), Peak Maximum of Parasites (PMP), Pre-Patent Period (PPP), Patent Period (PP) and Survival. The experiment was approved by the UEM’s Ethical Committee. Results: G1cH showed a higher survival (p=0.044) with a life expectancy of 2.58 times larger than the control group (Figure 1.A), as well as lower TP (p=0.002) and PMP (p = 0.018). PPP and PP showed no statistical difference, although in G1cH1 it was observed an increasing trend of PPP (p=0.065). These results are related to host’s benefit. The G6cH group presented a longer survival (p=0,045), with a life expectancy 1.94 times larger than the control group (Figure 1.B). Although no difference to TP, PMP, PP and PPP has been observed, the alcohol 6cH performed protecting animals against infection. The G30cH displayed an increasing trend of PMP (p=0.066) compared to the control group. Effects of inert vehicle succussed have been reported in studies in vitro2. However, no effects had been reported in vivo studies yet2,3. The hydro-alcoholic solution 7% 13CH, tested under the same conditions and animal model, did not change the natural evolution of the infection2. Conclusion: The hydro-alcoholic solutions 1cH and 6cH altered the course of experimental infection by T. cruzi, reducing the parasitemia and/or increasing the survival time, and can not be considered as inert vehicle in the high diluted compounds preparation.

Biotherapic 200dH is harmful to acute murine infection with Trypanosoma cruzi.

Biotherapics employed to treat mice infected by Trypanosoma cruzi were carried out with encouraging results. The aim of this study was evaluated the effect of biotherapic of Trypanosoma cruzi 200dH, using two different schedules of treatment. Swiss male mice, aged 56 days-old were infected intraperitonealy with 1,400 blood trypomastigotes of Trypanosoma cruzi Y strain and were divided into groups: C.I.- infected animals, E.D. – Infected animals treated from the day 1 until the end of the experiment; 200dH S.D. – Infected animals treated on the day 1. Parasitological, clinical and immunological parameters were evaluated. The group of animals that received the medicine in a single dose presented higher value to total parasitaemia and lower value of pre patent period compared to control untreated group (p

Remarks of Eimeria labbeana infection in Egyptian pigeons

This work was conducted to investigate the course of Eimeria labbeana infection in Egyptian pigeons (Columbia Livia). Thirty squabs were experimentally infected orally with 2.5 x104 sporulated Eimeria labbeana oocysts. Daily scarification of three squabs was done from day one post-infection (PI) until day 8 PI for following of the endogenous stages in tissue samples and 6 squabs were kept to follow the patent period and daily oocyst shedding. Paraffin-embedded intestinal samples were sectioned and stained for differentiation of parasitic stages. The infected squabs showed greenish watery diarrhea, weakness, rough feathers, and decrease food intake at day 5 PI. The pre-patent and patent periods were 6 and 14 days PI respectively. The oocyst shedding started at day 6 PI and reached the peak at day 8 PI. Histopathological examination revealed the presence of three schizont stages, macro-and micro-gametes, and oocysts in the duodenal and jejunal parts of the small intestine. In conclusion, it is the first study on Egypt on Eimeria labbeana in Egyptian pigeons and its biology is like recorded before.

Pathogenesis of Isospora amphiboluri in Bearded Dragons (Pogona vitticeps)

Isospora amphiboluri is a common coccidian found in captive bearded dragons (Pogona vitticeps). To minimize the impact of this parasite, it is important to characterize its pathogenesis so that we can develop appropriate methods for diagnosis and treatment. Forty-five juvenile bearded dragons were used for this two-part study. In the first part, ten bearded dragons were infected with 20,000 oocysts per os, while a control group of five animals received only water. Feces were collected over 45 days and screened for oocysts. In the second part, thirty bearded dragons were used to characterize the pathogenesis of I. amphiboluri. Twenty-five bearded dragons were infected as described previously, while five animals served as controls. Five infected bearded dragons and one control were humanely euthanized on days 4, 8, 12, 16, and 20 post-infection for complete necropsies. The pre-patent period for I. amphiboluri was found to be 18.6 ± 1.9 days (range 15–22 days). Histopathology confirmed that I. amphiboluri follows a homoxenous life cycle. Infections begin in the duodenum and progress to the colon over time. The findings of this study can be used to develop better quarantine and treatment protocols for captive bearded dragons.