Dose Of Diminazene Aceturate Research Articles

Evaluation of the Inhibitory Effect of Moringa oleifera Leaves Methanolic Extract against In Vitro Growth of Several Babesia Species and Theileria equi and the In Vivo Growth of Babesia microti.

The current study evaluated the inhibitory effect of Moringa oleifera leaves methanolic extract (MOL) against the in vitro growth of Babesia bovis (B. bovis), B. caballi, B. bigemina, and Theileria equi (T. equi), as well as in vivo growth of B. microti in mice. Active principles of MOL extract were determined using liquid chromatography mass spectrometry (LC-MS). MOL's anti-piroplasm efficacy was assessed both in vitro and in vivo using the SYBR Green I fluorescence assay. Every 96 hours, the hematological parameters, including red blood cell count (RBCs; 104/UL), hemoglobin content (HGB; g/dl), and hematocrit percent (HCT; %), in the treated mice were monitored using a Celltac MEK6450 automated hematological analyzer. LC-MS of MOL revealed that the most abundant polyphenolic catechism found in the MOL extract was isoquercetin and rutin. MOL inhibited B. bovis, B. caballi, B. bigemina, and T. equi in vitro growth in a dose-dependent way, with IC50 values of 45.29 ± 6.14, 19.16 ± 0.45, 137.49 ± 16.07, and 9.29 ± 0.014 μg/ml, respectively. MOL's in vitro antibabesial activity was enhanced when administrated simultaneously with either diminazene aceturate (DA) or MMV665875 compound from malaria box. In mice infected by B. microti, a combination of MOL and a low dose of DA (12.5 mg·kg-1) resulted in a significant (P < 0.05) reduction in B. microti growth. These findings suggest that MOL is an effective herbal anti-piroplasm therapy, especially when combined with a low dosage of either DA or MMV665875.

Diminazene aceturate and imidocarb dipropionate-based combination therapy for babesiosis – A new paradigm

In the present study, the effect of a combination therapy consisting of diminazene aceturate (DA) and imidocarb dipropionate (ID) on the in vitro growth of several parasitic piroplasmids, and on Babesia microti in BALB/c mice was evaluated using a fluorescence-based SYBR Green I test. We evaluated the structural similarities between the regularly used antibabesial medications, DA and ID, and the recently found antibabesial drugs, pyronaridine tetraphosphate, atovaquone, and clofazimine, using atom pair fingerprints (APfp). The Chou–Talalay approach was used to determine the interactions between the two drugs. A Celltac MEK-6450 computerized hematology analyzer was used to detect hemolytic anemia every 96 hours in mice infected with B. microti and in those treated with either mono- or combination therapy. According to the APfp results, DA and ID have the most structural similarities (MSS). DA and ID had synergistic and additive interactions against the in vitro growth of Babesia bigemina and Babesia bovis, respectively. Low dosages of DA (6.25 mg kg−1) and ID (8.5 mg kg−1) in conjunction with each other inhibited B. microti growth by 16.5 %, 32 %, and 4.5 % more than 25 mg kg−1 DA, 6.25 mg kg−1 DA, and 8.5 mg kg−1 ID monotherapies, respectively. In the blood, kidney, heart, and lung tissues of mice treated with DA/ID, the B. microti small subunit rRNA gene was not detected. The obtained findings suggest that DA/ID could be a promising combination therapy for treating bovine babesiosis. Also, such combination may overcome the potential problems of Babesia resistance and host toxicity induced by utilizing full doses of DA and ID.

Diminazene aceturate and imidocarb dipropionate-based combination therapy for babesiosis - a new paradigm.

In the present study, the effect of a combination therapy consisting of diminazene aceturate (DA) and imidocarb dipropionate (ID) on the in vitro growth of several parasitic piroplasmids, and on Babesia microti in BALB/c mice was evaluated using a fluorescence-based SYBR Green I test. We evaluated the structural similarities between the regularly used antibabesial medications, DA and ID, and the recently found antibabesial drugs, pyronaridine tetraphosphate, atovaquone, and clofazimine, using atom pair fingerprints (APfp). The Chou-Talalay approach was used to determine the interactions between the two drugs. A Celltac MEK-6450 computerized hematology analyzer was used to detect hemolytic anemia every 96h in mice infected with B. microti and in those treated with either mono- or combination therapy. According to the APfp results, DA and ID have the most structural similarities (MSS). DA and ID had synergistic and additive interactions against the in vitro growth of Babesia bigemina and Babesia bovis, respectively. Low dosages of DA (6.25mg kg-1) and ID (8.5mg kg-1) in conjunction with each other inhibited B. microti growth by 16.5, 32, and 4.5% more than 25mg kg-1 DA, 6.25mg kg-1 DA, and 8.5mg kg-1 ID monotherapies, respectively. In the blood, kidney, heart, and lung tissues of mice treated with DA/ID, the B. microti small subunit rRNA gene was not detected. The obtained findings suggest that DA/ID could be a promising combination therapy for treating bovine babesiosis. Also, such combination may overcome the potential problems of Babesiaresistance and host toxicity induced by utilizing full doses of DA and ID.

Pomegranate (Punica granatum) Peel Inhibits the In Vitro and In Vivo Growth of Piroplasm Parasites.

Pomegranate (Punica granatum) peel has seen a rapid surge in attention as a medical and nutritional product over the last decade. The impact of pomegranate peel methanolic extract monotherapy and combination therapy on the in vitro growth of Babesia (B.) bovis, B. bigemina, B. divergens, B. caballi, and Theileria (T.) equi, as well as B. microti in mice, was investigated in this work. Fluorescence-based SYBR green I assay was used for evaluating the inhibitory antibabesial efficacy of pomegranate (Punica granatum) peel against the growth of several piroplasm parasites in vitro and in vivo. Celltac α MEK-6450 computerized haematology analyzer was used for monitoring the haematological parameters of treated mice every 4 days. Pomegranate peel inhibited the in vitro growth of B. bovis, B. bigemina, B. divergens, T. equi, and B. caballi in a dose-dependent manner, with IC50 values of 154.45 ± 23.11, 40.90 ± 9.35, 72.71 ± 14.77, 100 ± 16.20, and 77.27 ± 16.94 μg/ml, respectively. On a B. bovis culture, the in vitro inhibitory effect of pomegranate peel was amplified when it was combined with diminazene aceturate (DA). Combination therapy of pomegranate peel and a low dose of DA (15 mg kg−1) inhibited B. microti growth significantly (P < 0.05) higher than the treatment with the full dose of DA (25 mg kg−1) in B. microti-infected mice. These findings suggest that pomegranate peel might be a potential medicinal plant for babesiosis treatment, especially when combined with a low dosage of DA.

Effect of methanolic extract from Capsicum annuum against the multiplication of several Babesia species and Theileria equi on in vitro cultures, and Babesia microti in mice

Background and Aim:Piroplasmosis is a serious disease that infects animals, inflicting significant economic losses in the livestock industry and animal trade worldwide. Anti-piroplasm drugs now on the market have demonstrated host toxicity and parasite resistance. As a result, developing more effective and safer anti-piroplasm drugs becomes an urgent issue. This study aimed to evaluate the inhibitory effect of Capsicum annuum methanolic extract (CA) against the growth of Babesia bovis, Babesia divergens, Babesia caballi, and Theileria equi in vitro and against B. microti in mice.Materials and Methods:Fluorescence-based SYBR Green I assay was used to evaluate CA’s inhibitory effect in vitro and in vivo when used either as a monotherapy or combined with diminazene aceturate (DA). The hematological parameters (HCT, hemoglobin, and red blood cells counts) were determined in the blood of mice every 96 h using Celltac a MEK-6450 electronic hematology analyzer.Results:The in vitro growth of B. bovis, B. divergens, T. equi, and B. caballi was inhibited by CA in a dose-dependent manner with IC50 values of 4.87±1.23, 44.11±8.03, 8.23±2.54, and 1.26±0.50 mg/mL, respectively. In B. microti-infected mice, a combination therapy consisting of CA and a low dose of DA showed a significant (p<0.05) inhibition of B. microti growth nearly similar to those obtained by treatment with the full dose of DA.Conclusion:The obtained results indicate that CA might be a promising medicinal plant for treating babesiosis, especially when used with a low dose of DA.

Trypanocidal effect of Diminazene aceturate by intranasal administration. Comparison among formulations

Background: The standard treatment for nagana and surra, vector–born parasite diseases, is a single intramuscular (i.m.) dose of diminazene aceturate (Berenil), an aromatic diamidine. Due to discontinuation of public provision of veterinary services or to lack of access to remote areas, low income farmers inject livestock and other domestic animals by themselves. We tested a not explored administration route for drugs against parasitological infections: the intranasal one. Berenil alone dissolved in water did not reach an effective drug concentration. Methods: Mice were infected with Trypanosoma brucei brucei or Trypanosoma evansi and treated with different formulations of Berenil. Survival, parasitemia, body weight and behavior were recorded. Results: Berenil formulated with chitosan reached a lethal concentration for bloodstream parasites. Residual parasites were absent, as demonstrated by immunosuppression. Conclusion: The intranasal route is an effective, safer, and easier way to perform antiparasitic treatments in animals.

Evaluation of the inhibitory effect of Zingiber officinale rhizome on Babesia and Theileria parasites.

The effect of Zingiber officinale rhizome methanolic extract (ZOR) on the in vitro growth of bovine Babesia (B. bovis, B. bigemina, and B. divergens) and equine piroplasm (B. caballi, and Theileria equi) parasites and on the growth of B. microti in mice was evaluated in this study. The possible in vitro synergistic interaction between ZOR and either diminazene aceturate (DA) or potent Medicines for Malaria Venture (MMV) hits from the malaria box was also investigated. In vitro, ZOR reduced the growth of B. bovis, B. bigemina, T. equi, and B. caballi in a dose-dependent manner. B. divergens was the most susceptible parasite to the in vitro inhibitory effect of ZOR. DA and MMV compounds enhanced the in vitro inhibitory antibabesial activity of ZOR. 12.5mg/kg DA when administrated in combination with ZOR in mice exhibited a significant inhibition (P<0.05) in B. microti growth better than those observed after treatment with 25mg/kg DA monotherapy. These findings suggest that ZOR could be a viable medicinal plant for babesiosis treatment, particularly when combined with a modest dose of either DA or powerful anti-B. bigemina MMV hits.

Evaluation of the Efficacy of a Combination of Artemether and ‎Diminazene Aceturate Therapy in Experimental Trypanosoma brucei ‎Infection in Rats

This study evaluated the efficacy of a combination of artemether and diminazene aceturate therapy in experimental Trypanosoma brucei infection in rats. Thirty five male albino rats used for the study were randomly assigned to seven groups of five rats each as follows: Group A- infected and treated with diminazene aceturate (DA) at 7.0 mg/kg body weight (bw), intramuscular (IM) once on day 7 post-infection (pi); Group B – infected and treated with artemether (ART) at 3.2 mg/kg bw IM on day 7pi and 1.6mg/kg bw IM on days 8,9,10 and 11pi; Groups C,D and E, – infected and treated with DA at 7.0 mg/kg bw IM, 3.5 mg/kg bw IM, and 1.75 mg/kg bw IM respectively, once on day 7pi and ART at 3.2 mg/kg bw IM on day 7pi plus 1.6mg/kg bw on days 8,9,10 and 11pi; Group F –infected, untreated and Group G – uninfected, untreated. Onset of parasitaemia (OP), level of parasitaemia (LOP), clearance of parasites post treatment, mortality post infection, relapse of parasitaemia post clearance, rectal temperature, and body weight, were determined at specified intervals during the 70-day experimental period. Results showed that there were no significant (p&gt;0.05) variations in the OP and LOP between the infected groups. Trypanosomes were cleared from the blood of rats in group A, C, D, and E after treatment. All the rats in groups B and F were dead by day 14pi. The infection relapsed in groups C and E. It was concluded that a combination of DA (3.5mg/kg bw once) and ART (3.2 mg/kg bw on day 1 of treatment and 1.6 mg/kg bw for 4 consecutive days) exhibited efficacy comparable to the standard dose of DA at 7mg/kg in the treatment of trypanosome brucei in rats and could thus possibly constitute an effective treatment regimen to reduce the dose of DA and avoid toxicity.&#x0D;

Myrrh Oil in Vitro Inhibitory Growth on Bovine and Equine Piroplasm Parasites and Babesia microti of Mice.

The present experimental study was conducted for the assessment of the efficacy of in vitro inhibition of myrrh oil on the propagation of Babesia bovis, B. divergens, B. bigemina, Theileria equi, and B. caballi and in vivo efficacy on B. microti in mice through fluorescence assay based on SYBR green I. The culture of B. divergens B. bovis and was used to evaluate the in vitro possible interaction between myrrh oil and other commercial compound, such as pyronaridine tetraphosphate (PYR), diminazene aceturate (DA), or luteolin. Nested-polymerase chain reaction protocol using primers of the small-subunit rRNA of B. microti was employed to detect any remnants of DNA for studied parasitic species either in blood or tissues. Results elucidated that; Myrrh oil significantly inhibit the growth at 1% of parasitic blood level for all bovine and equine piroplasm under the study. Parasitic regrowth was inhibited subsequently by viability test at 2 µg/mL for B. bigemina and B. bovis, and there was a significant improvement in the in vitro growth inhibition by myrrh oil when combined with DA, PYR, and luteolin. At the same time; mice treated with a combination of myrrh oil/DA showed a higher inhibition in emitted fluorescence signals than the group that challenged with 25 mg/kg of diminazene aceturate at 10 and 12 days post-infection. In conclusion, this study has recommended the myrrh oil to treat animal piroplasmosis, especially in combination with low doses of DA.

Development of unstable resistance to diminazene aceturate in Babesia bovis

Diminazene aceturate (DA) is commonly used in the treatment of bovine babesiosis caused by Babesia bovis. In this study, we attempted to develop resistance in B. bovis in vitro to DA and clofazimine (CF, a novel antibabesial agent) using short- and long-term drug pressures. In the short term, we found that 6.7 ± 2 (0.54 ± 0.16 μM)-, 12.9 ± 8.6 (1.05 ± 0.7 μM)-, and 14 ± 5.9 (1.14 ± 0.48 μM)-fold increases in the half-maximal inhibitory concentration (IC50) of DA were demonstrated on B. bovis cultivated with 0.04 μM of DA pressure for 4, 8, and 12 days, respectively, as compared to that on parental culture (0.08 ± 0.0065 μM) before drug pressure was initiated. However, in B. bovis cultivated with 0.04 μM of DA pressure after 16 days, the parasites could not tolerate 0.8 μM of DA. In the long term, 7.6 ± 3.5-, 20.5 ± 0.1-, and 26.8 ± 5.5-fold increases in the IC50 of DA were demonstrated on parasites from subcultures at days 8, 3, and 5 post-cultivation, respectively, in a drug-free medium, where these subcultures were obtained from B. bovis cultivated with DA pressure with changing doses for 30, 60, and 90 days, respectively. However, the second and third times, no increase was demonstrated on B. bovis from these subcultures at days 15 and 30 post-cultivation in a drug-free medium. In addition, in B. bovis cultivated with drug pressure after 90 days, the parasites tolerate up to 0.64 μM DA. All findings demonstrated that DA resistance in B. bovis is unstable and lost within 15 days of drug withdrawal. However, treatment with subtherapeutic doses of DA in cattle might result in the development of resistance in B. bovis, which may not even respond to subsequent treatments with high doses of DA. Thus, if the bovine babesiosis caused by B. bovis is unresponsive to DA, treatment with other antibabesial agents might be recommended.

Evaluation of efficacy and effects of daily dosing of diminazene aceturate in dogs experimentally infected with &lt;i&gt;Trypanosoma congolense&lt;/i&gt;

Resistance to trypanocidal drugs or relapse of infection poses a big challenge to the effective treatment of trypanosomosis. This study investigated the efficacy and effect of multiple daily doses of diminazene aceturate (DA) in Nigerian indigenous dogs (NID) experimentally infected with Trypanosoma congolense (2.5 x 105). Fifteen NID of both sexes and mean weight of 9.4 0.98 kg were randomly divided into 3 equal groups. Group A dogs were infected but not treated. Groups B and C were infected with T. congolense, and treated with single and four multiple daily divided doses of 3.5 mg/kg of DA, respectively. Parasitaemia was detected in the infected dogs on day 10 post infection (pi) and treatment was instituted in Groups B and C dogs on day 17 pi. Parasitaemia was cleared on days 10 and 8 post-treatment in Groups B and C respectively. There was an increase in urea, creatinine, aminotransferase and aspartate aminotransferase levels in all the groups while, alkaline phosphatase, Packed Cell Volume and Red Blood Cells count decreased. There was relapse of infection in treated groups on day 25 post-treatment. We concluded that four multiple daily divided doses of DA (3.5 mg/kg) were not effective in preventing relapse of T. congolense infection in NID.Keywords: Diminazene aceturate multiple doses; trypanosomosis; biochemical changes; dogs

Prevalence of bovine trypanosomosis and assessment of trypanocidal drug resistance in tsetse infested and non-tsetse infested areas of Northwest Ethiopia

The Northwestern region of Ethiopia is affected by both tsetse and non-tsetse transmitted trypanosomosis with a significant impact on livestock productivity. The control of trypanosomosis in Ethiopia relies on either curative or prophylactic treatment of animals with diminazene aceturate (DA) or isometamidium chloride (ISM). In the present work; questionnaire survey, cross-sectional and experimental studies were carried out to; a) assess the utilization of trypanocidal drugs; b) determine the prevalence of bovine trypanosomosis and; c) assess the drug resistant problems respectively in Tsetse and non-tsetse infested areas on NW Ethiopia. A total of 100 respondents were included for the survey and the questionnaires focused on the drug utilization practices for the control of Trypanosomosis. Blood from cattle 640 (324 cattle tested in 2011, 316 cattle tested in 2012) and 795 (390 cattle tested in 2011, 405 cattle tested in 2012) were examined from tsetse infested and non-tsetse infested areas respectively using the buffy coat technique and thin blood smear for the detection of trypanosomes and measurement of packed cell volume (PCV). For the assessment of trypanocidal drug resistance three isolates, one from tsetse (TT) and two from non-tsetse (NT) areas were used on thirty six trypanosome naïve calves. The experimental animals were divided randomly into six groups of six animals (TT-ETBS2-DA, TT-ETBS2-ISM, NT-ETBD2-DA, NT-ETBD2-ISM, NT-ETBD3-DA and NT-ETBD3-ISM), which were infected with T. vivax isolated from a tsetse-infested or non-tsetse infested area with 2×106 trypanosomes from donor animals, and in each case treated with higher dose of DA or ISM. The results of the questionnaire survey showed trypanosomosis was a significant animal health constraint for 84% and 100% of the farmers questioned in non-tsetse and tsetse infested areas of Northwest Ethiopia respectively. Responses on trypanocidal drug utilization practices indicated that risk factors for the development of drug resistance are common and treatment failures are frequently seen. Accordingly, the majority of farmers in tsetse infested area get trypanocides from drug stores and unauthorized sources whereas those from non-tsetse area get from veterinary clinics. Moreover, treatment administration is mainly by animal health personnel and treatment frequency is a maximum of three times/year/animal in non-tsetse area whereas it is administered mainly by the farmers more than seven times/year/animal in tsetse infested area. The prevalence of trypanosomosis varied from 17.59% in 2011 to 25.0% in 2012 in tsetse infested areas with a significant (P=0.023) difference. Similarly, in non-tsetse infested area the prevalence was varied from 3.85% in 2011 to 5.93% in 2012 without significant rise. Trypanosoma congolense (75%) was the most prevalent followed by T. vivax (20.58%) and mixed infections (4.41%) in tsetse infested area while in non-tsetse infested area only T. vivax was detected. The overall mean PCV in parasitaemic animals (20±2.3 SD) was significantly (P<0.001) lower than that of aparasitaemic animals (27±4.3 SD). The assessment of trypanocidal drug resistance tests revealed one isolate of non-tsetse infested area against DA in group NT-ETBD2-DA is resistant to the higher dose used with 3 relapsing animals (50% relapses) in the group. Another two relapses were detected one against ISM for the isolate from tsetse infested area (TT-ETBS2-ISM) and one against DA for another isolate (NT-ETBD3-DA) from the non-tsetse area. In conclusion, trypanosomosis is widely prevalent in both study areas causing significant reduction in the mean PCV values. Farmers' trypanocidal utilization practices appear to pose risks of drug resistance problems. The in vivo drug resistance tests indicated the presence of resistant parasites with the higher dose against DA for NT-ETBD2 isolate and suspected resistance problems were detected against ISM and DA for TT-ETBS2 and NT-ETBD3 isolates respectively. Therefore, trypanosomosis is a major constraint in Northwest Ethiopia and drug resistance is a threat in the control of trypanosomosis in both study areas.

Efficacy of repeated doses of diminazene aceturate (Dinazene®) in the treatment of experimental Trypanosoma brucei infection of Albino rats.

The efficacy of repeated doses of Dinazene® in Albino rats experimentally infected with Trypanosoma brucei (Gboko strain) was investigated. A total of 30 adult female Albino rats weighing 130-190 g were used for the study. They were assigned to six groups (groups A-F) of five rats each. Groups A-D were infected intraperitoneally with 1.0 × 106 trypanosomes in 400 µL of PBS diluted blood while groups E (uninfected treated) and F (uninfected untreated) served as controls. The rats in the groups A-D as well as those in group E were treated with 7.0 mg/kg body weight at day 11 post infection. Groups B, C and D however received two, three and four repeated doses of the drug at weekly intervals following initial treatment. There was complete clearance of the parasite within 120 h post treatment. Parasitaemia, packed cell volume (PCV), total red blood cell (RBC) and white blood cell (WBC) counts, haemoglobin concentration (Hb), rectal temperature, and body weight were used to assay the efficacy of treatment. Following treatment and parasite clearance from the blood, there was improvement (P<0.05) in the values of parameters measured when compared to the uninfected controls. However, relapse infection was observed in the rats of group A, B and C, with a resultant decline in clinical condition and values of parameters used to assess efficacy. We concluded that four consecutive treatments using same dose at weekly intervals proved efficacious in the experimental management of T. brucei infection in rats.

In vivo experimental drug resistance study in Trypanosoma vivax isolates from tsetse infested and non-tsetse infested areas of Northwest Ethiopia

Ethiopia, particularly in the Northwest region, is affected by both tsetse fly and non-tsetse fly transmitted trypanosomosis with a significant impact on livestock productivity. The control of trypanosomosis in Ethiopia relies on either curative or prophylactic treatment of animals with diminazene aceturate (DA) or isometamidium chloride (ISM), respectively. However, since these two trypanocides have been on the market for more than 40 years, this may have resulted in drug-resistance. Therefore, in vivo drug resistance tests on two Ethiopian isolates of Trypanosoma vivax were completed, one from an area where tsetse flies are present and one from an area where tsetse flies are not present. Twenty four cattle (Bos indicus) aged between 6 and 12 months, purchased from a trypanosome-free area (Debre Brehan: Northcentral Ethiopia) and confirmed to be trypanosome-negative, were randomly assigned into four groups of six animals, which were infected with T. vivax isolated from a tsetse-infested or non-tsetse infested area, and in each case treated with curative doses of DA or ISM. Each animal were inoculated intravenously 3×106 trypanosomes from donor animals. Parasitaemia became patent earlier in infections with non-tsetse T. vivax (∼7 days post-infection) than tsetse (∼14 days post-infection). Both groups were treated at the highest peak parasitaemia with DA or ISM and nine cattle, four with non-tsetse T. vivax (two ISM- and two DA-treated) and five with tsetse T. vivax (three ISM- and two DA-treated) showed relapses of parasitaemia. Moreover, treatment did not improve diagnostic host markers of trypanosome infections in these animals. In conclusion, in vivo drug tests indicated the presence of resistant parasites (>20% of treated animals in each group relapsed) against recommended doses of both available trypanocidal drugs.

Abstract W P196: Post-Stroke Activation of Angiotensin Converting Enzyme 2 is Neuroprotective

Background: Recent discoveries point to the angiotensin converting enzyme 2/angiotensin-(1-7)/mas (ACE2-Ang-(1-7)-Mas) axis as a potential target for neuroprotective stroke therapies. Central pre-stroke activation of this axis by infusion of diminazene aceturate (DIZE), an activator of ACE2, improves stroke outcomes in rats. This led us to hypothesize that intraperitoneal (IP) post-stroke injections of DIZE may exert similar Mas-mediated neuroprotection and thus offer additional clinically-relevant evidence to support this approach. We also assessed the direct effects of DIZE on ACE2 activity in brain tissue. Methods: Male SD rats underwent ischemic stroke by endothelin-1-induced middle cerebral artery occlusion and were randomly divided into groups (n=~12/group) that received IP injections of dH2O or DIZE (0.75, 2.5, or 7.5 mg/kg) at 4, 24, and 48 h after stroke, as well as blinded neurological assessments at 4, 24, and 72 h after stroke. Additional groups received pre- and post-stroke intracerebroventricular (ICV) infusion of sterile 0.9% saline or Mas receptor antagonist A-779 in addition to post-stroke control or drug therapy. Immediately after the 72 h tests, animals were euthanized, cerebral infarct size assessed by TTC staining, and tissue harvested for ACE2 activity assay. Data are expressed as mean ± SEM with significance inferred at p&lt;0.05. Results: Mean infarct sizes (%) were significantly decreased by post-stroke IP injections of all doses of DIZE (0.75 mg/kg: 29.9±5.0; 2.5 mg/kg: 27.9±4.2; 7.5 mg/kg: 31.36±4.1) vs. dH2O (41.9±3.8). At 24 h post-stroke, neurologic deficits (Garcia and Bederson scales) were significantly improved in DIZE-treated rats (7.5 mg/kg: 16.7±0.2 and 0.5±0.1) vs. controls (15.5±0.4 and 1.1±0.2). ICV infusion of A-779 in DIZE-treated rats abolished the improvement in infarct size observed in saline infused DIZE-treated rats (A-779: 43.8%±7.5; saline: 25.8±6.7). Addition of DIZE (4 nmol/μg protein) to brain homogenate increased ACE2 activity by more than 3-fold. Conclusions: This suggests that targeting the ACE2/Ang-(1-7)/Mas axis by post-stroke DIZE injections exerts neuroprotection in a Mas-dependent fashion.

Evaluation of In vivo Antitrypanosomal Activity of Aqueous and Methanol Leaf Extracts of Clutia abyssinica (Euphorbiaceae) against Trypanosoma congolense Field Isolate

Aqueous and methanol leaf extracts of C. abyssinica were investigated for their in vivo activity against Trypanosoma congolence, the main causative agent of African animal trypanosomosis in Sub-Saharan Africa and Ethiopia. The in vivo antitrypanosomal efficacy of the aqueous and methanol extracts was evaluated in Swiss albino mice infected with T. congolense field isolate. The leaf extracts were administered 12 days post-infection at peak parasitaemia level of ~108 trypanosomes/ml at doses of 100, 200 and 400 mg/kg by intraperitoneal injection once daily for 7 days. Parasitaemia, packed cell volume (PCV), mean survival time and change in body weight were used as indices for monitoring the efficacy of the extracts by comparing with the positive control: 28 mg/kg dose of diminazene aceturate and negative control: 2% tween 80 treated groups. The extracts had a lethal dose greater than 2000 mg/kg and there were no evidences of acute toxicity at the doses tested. Highly significant (p<0.001) reduction in pre-treatment parasitaemia by 3.91% (7.38 ± 0.18), increase in PCV by 1.12% (48.66 ± 0.20), body weight improvement by 1.36% (22.34 ± 0.27) and mean survival time of 39.20 ± 0.37 days was observed in the group treated by 400 mg/kg methanol leaf extract of C. abyssinica. In general, the results obtained suggest ethno-pharmacological usefulness of the plant and necessitate further studies to be carried on isolated active substances from the plant.

In vivo trypanocidal effect of aqueous root extracts of securidaca longepedunculata and its phytochemical analysis

The aqueous root extracts of Securidaca longepedunculata (Fresen, polygalacaea) were used to treat trypanosomiasis in this experiment. 25 Wister albino rats were inoculated with Trypanosoma brucei. Its trypanocidal activity was assessed through daily examination of blood samples, clinical and haematological changes at intervals, and possible deaths were among the parameters which were carefully monitored. The treatment involved a therapeutic dose of diminazene aceturate (3.5 mg/kg), a combination of sub-therapeutic dose of diminazene (1.75 mg/kg) and sub-therapeutic dose of the extract, oral infusion of 200 and 100 mg/kg of the extract, respectively for 7 days. In all rats treated with diminazene and the extract, there was a significant decrease (p < 0.05) in parasitemia even though those that received the extract alone relapsed. And there was a significant increase (p < 0.05) in haematological values as well. Hence, these findings provide a possible, cheap and available alternative to the existing but costly trypanocides additionally, due to phytochemical data revealed. Key words: Securidaca longepedunculata, trypanosomiasis, diminazene aceturate, parasitemia, Trypanosoma brucei, Human African trypanosomiasis.

Blocking &lt;i&gt;Babesia bovis&lt;/i&gt; vaccine reactions of dairy cattle in milk

The use of 1.16 mg/kg (one third) of the recommended dose of diminazene aceturate, administered indiscriminately to cattle on day seven of the unfrozen Babesia bovis and Babesia bigemina bivalent live blood vaccine reaction, was an infection and block treatment method of immunisation used successfully with no known adverse effect on the parasites or the development of protective immunity. Continuing with this practice after replacement of the unfrozen vaccine with deep-frozen monovalent B. bovis and B. bigemina live blood vaccines resulted in reports of vaccine failure. Laboratory investigation indicated the harmful effect of block treatment in preventing the development of durable immunity against B. bigemina as opposed to the much lesser effect it had on B. bovis. Consequently the practice was no longer recommended. A B. bovis vaccination attempt aimed at controlling the disease of dairy cows in milk (n = 30) resulted in 20% fatalities during the expected vaccine reaction period. The practice of block treating B. bovis was therefore reinvestigated, this time in a field trial using dairy cattle in milk (n = 11). Using 0.88 mg/kg (one quarter) of the recommended dose of diminazene administered on day 12 of the B. bovis vaccine reaction resulted in only two animals (n = 5) testing ≥ 1/80 positive with the indirect fluorescent antibody test (IFAT) although parasites could be demonstrated in three. In the untreated control group, by contrast, five of the vaccinated animals (n = 6) tested ≥ 1/80 positive with IFAT and parasites could be demonstrated in all. The unsatisfactory outcome obtained in this study, combined with that of the earlier investigation, indicated that there are more factors that influence successful vaccination than previously considered. It is therefore concluded that block treatment of the live frozen South African cattle babesiosis vaccines reactions is not recommended.

Occurrence of Trypanosoma evansi in Horses in the State of Minas Gerais, Brazil

Ttrypanosomosis caused by Trypanosoma evansi is a condition that causes significant losses to farmers in endemic areas. This study aimed to report one case of trypanosomiasis in the municipality of Itabira, state of Minas Gerais, Brazil. In November 2010, on a farm with 16 horses, three horses had clinical signs of anemia, limb edema, weight loss, lameness, muscle atrophy, and incoordination of the hind limbs. Trypanosomosis was suspected, and the animals with clinical signs were treated with two doses of diminazene aceturate (7 mg/kg, intramuscular) every 7 days; the treated animals recovered from the clinical symptoms. Blood samples were collected from six horses on the farm, including the three treated animals, to perform polymerase chain reaction specific for T evansi. Four samples were polymerase chain reaction negative, including those collected from three treated horses. However, two other asymptomatic horses were positive for the parasite based on the molecular testing. Based on the results, we concluded that the initial clinical suspect of trypanosomosis was correct, and the treatment used was effective. Probably the disease was introduced to Minas Gerais State through a stallion, which acquired the infection in an endemic area in southern Brazil.

A Preliminary Trypanocidal Study of Natural and Synthetic Supplementation of Zinc and Magnesium in Combination with Diminazene Aceturate in Wistar Rats

Eighty healthy Wistar albino rats were used to investigate the trypanocidal effect of natural and synthetic supplements of zinc and magnesium in combination with diminazene aceturate in Trypanosoma congolense-inoculated rats. The rats were randomly divided into eight groups in study ‘1’ (involving the natural supplements of zinc and magnesium) and eight groups in study ‘2’ (the synthetic supplements ZnCl2 and MgCl2), each group of both studies having five rats. Hence, studies 1 and 2 had groups A - H each. Parasitaemia of trypanosome parasite was assessed using ‘wet mount method’ after inoculation of all test groups except group ‘G’ of both studies (the normal, not infected, not treated control). Group ‘E’ of both studies in which combinations of natural source of zinc ion (maize bran) and magnesium ion (wheat bran) were combined with subtherapeutic dose of diminazene aceturate at 1.75 mg/kg cleared the trypanosome parasites with no relapse. The salts supplements of ZnCl2 and MgCl2 with the same subtherapeutic dose of diminazene aceturate also elicited an effect similar to the group ‘E’ in natural supplementation study. There was thus, no significant difference (p>0.05) between the PCV and RBC of normal group ‘G’ and the natural and synthetic supplemented groups with the trypanocide. The groups B, C, and D of both studies however, only prolonged the live of the animals (though with relapse occurring) there was still the death of the animals before the end of the experiment. The improvement of PCV and RBC values of treated groups ‘E’ in the two studies towards the normal gave credence to the fact that the supplements of Zn2+ and Mg2+ with the subtherapeutic dose of 1.75 mg/kg had a better trypanocidal and rejuvenating tendency and could be used for treatment.