Quality, supply chain, and use of Trypanocidal drugs among camel keepers in north-eastern Kenya: A cross-sectional study.

The non-cyclic trypanosomiasis (surra), caused by Trypanosoma evansi, and mechanically transmitted by biting flies, hinders camel productivity in Kenya. Trypanocides are the most commonly used drugs to control surra. However, emergence of drug resistance by the parasites is a major limitation to control efforts. There is limited information on the quality of trypanocides, the supply chain and drug-use practices among camel keepers potentially contributing to development of drug resistance in Kenya. We sought to fill this gap by conducting a cross-sectional study among camel keepers in Isiolo and Marsabit counties, Kenya. We mapped the trypanocide drugs supply chain through quantitative and qualitative surveys. We administered a semi-structured questionnaire to camel keepers to generate data on trypanocides-use practices, including the types, sources, person who administers treatment, reconstitution, dosage, route and frequency of administration, among others. Additionally, we tested the quality of trypanocidal drugs retailed in the region. We mapped a total of 55 and 49 agro-veterinary outlets and general (ordinary) shops retailing veterinary drugs in the two counties, respectively. These comprised of 29 and 26 agro-veterinary outlets, as well as 24 and 25 general shops in Isiolo and Marsabit counties, respectively. Overall, the respondents experienced 283 surra cases in the three-month recall period, which were treated with trypanocides. The majority of these cases were diagnosed by camel owners (71.7%) and herders (24.1%). A significant proportion of the cases were treated by camel owners (54.8%), herders (35.3%), the owner's son (3.2%) and veterinary personnel (1.1%) (χ2 = 24.99, p = 0.000). Most of the households sourced the drugs from agro-veterinary outlets (59.0%), followed by general shops (19.8%), veterinary personnel (2.1%), and open-air markets (0.4%) (χ2 = 319.24, p = 0.000). Quinapyramine was the most (56.9%) predominantly used trypanocide in treatment of surra, followed by homidium (19.8%), isometamidium (15.9%), diminazene aceturate (6.7%), and ethidium (0.7%) (χ2 = 340.75, p < 0.000). Only a meager proportion of respondents (15.2%) used the drugs correctly as instructed by the manufacturers. We recorded an association between correct drug usage, with the person who administers the treatment (χ2 = 17.7, p = 0.003), and the type of trypanocide used (χ2 = 19.4, p < 0.001). All the drug samples tested had correct concentrations of active ingredient (100.0%), and therefore of good quality. We have demonstrated that whereas the trypanocides retailed in the region by authorized vendors are of good quality, there is widespread incorrect handling and use of the drugs by unqualified individuals, which may contribute to treatment failure and emergence of trypanocide resistance.

Molecular characterization of trypanocide-resistant strains derived from a single field isolate of Trypanosoma evansi.

Four strains (SB-PR, SB-RS, SB-RD, and SB-RM) of Trypanosoma evansi (T. evansi) were used in this study. SB-PR is known to be trypanocide-sensitive, while the others are trypanocide-resistant to suramin, diminazene diaceturate, and melarsomine hydrochloride, respectively. SB-RS, SB-RD, and SB-RM are derivatives of a single field isolate of SB-PR. Trypanocide resistance will not only increase costs and decrease production efficiency but will also affect effective treatment strategies. Therefore, studies on this topic are important to avoid inefficient production and ineffective treatment. This paper aims to presents a comparative molecular characterization of the trypanocide-resistant strains compared to the parent population. Comparative molecular characterization of these strains based on a protein profile analysis performed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), DNA fingerprinting of random amplified polymorphic DNA (RAPD), and the molecular characterization of expression-site-associated 6 (ESAG6), variant surface glycoprotein (VSG), and T. evansi adenosine transporter-1 (TevAT1) gene sequences. The results show three derived strains (SB-RS, SB-RD, and SB-RM) exhibit different banding patterns than SB-PR. According to the RAPD results, SB-RS and SB-RD are different strains with DNA fingerprint similarities of about 77.8 %, while the DNA fingerprint of SB-RM has a similarity of 44.4 % to SB-RS and SB-RD. No differences in VSG were found among the four strains; however, ESAG6 showed differences in both nucleotide and amino acid sequences, as well as in its secondary and 3D structure. In conclusion, all molecular analyses of the ESAG6 gene showed that SB-PR, SB-RS, SB-RD, and SB-RM are different strains. Furthermore, SB-PR, SB-RS, SB-RD, and SB-RM did not exhibit the TevAT1 gene, so the resistance mechanism was determined to be unrelated to that gene.

Drug resistance in animal trypanosomiases: Epidemiology, mechanisms and control strategies.

Animal trypanosomiasis (AT) is a complex of veterinary diseases known under various names such as nagana, surra, dourine and mal de caderas, depending on the country, the infecting trypanosome species and the host. AT is caused by parasites of the genus Trypanosoma, and the main species infecting domesticated animals are T. brucei brucei, T. b. rhodesiense, T. congolense, T. simiae, T. vivax, T. evansi and T. equiperdum. AT transmission, again depending on species, is through tsetse flies or common Stomoxys and tabanid flies or through copulation. Therefore, the geographical spread of all forms of AT together is not restricted to the habitat of a single vector like the tsetse fly and currently includes almost all of Africa, and most of South America and Asia. The disease is a threat to millions of companion and farm animals in these regions, creating a financial burden in the billions of dollars to developing economies as well as serious impacts on livestock rearing and food production. Despite the scale of these impacts, control of AT is neglected and under-resourced, with diagnosis and treatments being woefully inadequate and not improving for decades. As a result, neither the incidence of the disease, nor the effectiveness of treatment is documented in most endemic countries, although it is clear that there are serious issues of resistance to the few old drugs that are available. In this review we particularly look at the drugs, their application to the various forms of AT, and their mechanisms of action and resistance. We also discuss the spread of veterinary trypanocide resistance and its drivers, and highlight current and future strategies to combat it.

Gallium maltolate, a promising low toxicity drug with curative effect on mice chronically infected with Trypanosoma evansi.

Trypanosoma evansi is a flagellate protozoan that infects a wide range of hosts, especially horses. Clinically, the infection is characterized by rapid weight loss, anemia and mobility disorders. This study evaluated the efficacy of treatment gallium maltolate (GaM) in rats infected with T. evansi in the acute and chronic phases of the disease and its influence on the enzyme and blood parameters. 48 animals (Rattus norvegicus) were divided into 8 groups (A-H) of 6 animals each, namely: A: (negative control) uninfected; B: acutely infected positive control; C: chronically infected positive control; D: acutely infected, treated with GaM for 7 days post infection (p.i.); E: acutely infected treated with GaM for 3 days before infection (b.i) and 7 days p.i.; F: chronically infected, treated with GaM for 7 days p.i.; G: chronically infected, treated with GaM for 3 days b.i. and 7 days p.i.; and H: uninfected treated with GaM for 10 days. Acute infected animals (B, D and E) had a progressive increase in parasitemia and were died or euthanized before completing treatment days (5th days p.i.) as they had high parasitemia (over 100 field trypanosomes in the blood smear). Thus, it can be concluded that GaM was not effective against an acute infection. In untreated chronically infected animals (C) the parasitemia also increased progressively and they were euthanized on the 7th day p.i.. The chronically infected and treated animals (F and G) showed low parasitemia and after treatment became negative, showing no trypanosomes in the bloodstream until the 50th day of the experiment. Thus, we conclude that GaM was effective against chronic infections. In uninfected and treated animals (H) hematological, biochemical and enzymatic parameters had no significant changes when compared to the negative control group (A) demonstrating the low toxicity of GaM.

Parasitological, Serological and Molecular Prevalence of Trypanosoma evansi among Arabian Camels (Camelus dromedaries) with Evaluation of Antitrypanosomal Drugs.

PURPOSE: This study was carried out to assess the prevalence of Trypanosoma evansi infection in naturally diseased Dromedary camels in Dammam, Eastern region of Saudi Arabia. The detection of Trypanosoma evansi was performed using the parasitological, serological, and molecular diagnosis and a comparison between such methods were analyzed. In addition, evaluation of therapeutic efficacy of selected antitrypanosomal drugs, cymelarsan and quinapyrmine (aquin-1.5), was trialed for treatment of diagnosed infected cases. METHODS: A total 350 randomly selected camels were evaluated using thin blood smear (TBS), RoTat1.2 PCR and CATT/T. evansi techniques. RESULTS: The total prevalence was 6.9%, 7.7%, and 32.8% by TBS, RoTat1.2 PCR and CATT/T. evansi techniques, respectively. Although PCR detect T. evansi in more samples than TBS, the agreement was good (K = 0.9). Among the CATT/T. evansi results, PCR detect T. evansi in 12 and 15 CATT positive and negative camels, respectively, with low agreement (Kappa = 0.1). The use of cymelarsan and quinapyramine sulfate in the treatment of naturally infected cases demonstrated a very efficient therapeutic response. CONCLUSION: It was found that 1. Comparing the CATT/T. evansi and PCR results, the positivity of CATT was higher than PCR detection, while the agreement was poor (K = 0.1). 2. Cymelarsan and aquin-1.5 proved to be effective in the treatment of naturally infected camels, but cymelarsan presented with higher effectiveness (100%) than aquin-treated camels (83.3%). a 3. The use of cymelarsan and CATT is recommended for disease treatment and control.

In vitro trypanocidal potency and in vivo treatment efficacy of oligomeric ethylene glycol-tethered nitrofurantoin derivatives.

African trypanosomiasis is a significant vector-borne disease of humans and animals in the tsetse fly belt of Africa, particularly affecting production animals such as cattle, and thus, hindering food security. Trypanosoma congolense (T. congolense), the causative agent of nagana, is livestock's most virulent trypanosome species. There is currently no vaccine against trypanosomiasis; its treatment relies solely on chemotherapy. However, pathogenic resistance has been established against trypanocidal agents in clinical use. This underscores the need to develop new therapeutics to curb trypanosomiasis. Many nitroheterocyclic drugs or compounds, including nitrofurantoin, possess antiparasitic activities in addition to their clinical use as antibiotics. The current study evaluated the in vitro trypanocidal potency and in vivo treatment efficacy of previously synthesized antileishmanial active oligomeric ethylene glycol derivatives of nitrofurantoin. The trypanocidal potency of analogues 2a-o varied among the trypanosome species; however, T. congolense strain IL3000 was more susceptible to these drug candidates than the other human and animal trypanosomes. The arylated analogues 2k (IC50 0.04 µM; SI >6365) and 2l (IC50 0.06 µM; SI 4133) featuring 4-chlorophenoxy and 4-nitrophenoxy moieties, respectively, were revealed as the most promising antitrypanosomal agents of all analogues against T. congolense strain IL3000 trypomastigotes with nanomolar activities. In a preliminary in vivo study involving T. congolense strain IL3000 infected BALB/c mice, the oral administration of 100 mg/kg/day of 2k caused prolonged survival up to 18 days post-infection relative to the infected but untreated control mice which survived 9 days post-infection. However, no cure was achieved due to its poor solubility in the in vivo testing medium, assumably leading to low oral bioavailability. These results confirm the importance of the physicochemical properties lipophilicity and water solubility in attaining not only in vitro trypanocidal potency but also in vivo treatment efficacy. Future work will focus on the chemical optimization of 2k through the investigation of analogues containing solubilizing groups at certain positions on the core structure to improve solubility in the in vivo testing medium which, in the current investigation, is the biggest stumbling block in successfully treating either animal or human Trypanosoma infections.

State-of-the-art Review on the Antiparasitic Activity of Benzimidazolebased Derivatives: Facing Malaria, Leishmaniasis, and Trypanosomiasis.

Protozoan parasites represent a significant risk for public health worldwide, afflicting particularly people in more vulnerable categories and cause large morbidity and heavy economic impact. Traditional drugs are limited by their toxicity, low efficacy, route of administration, and cost, reflecting their low priority in global health management. Moreover, the drug resistance phenomenon threatens the positive therapy outcome. This scenario claims the need of addressing more adequate therapies. Among the diverse strategies implemented, the medicinal chemistry efforts have also focused their attention on the benzimidazole nucleus as a promising pharmacophore for the generation of new drug candidates. Hence, the present review provides a global insight into recent progress in benzimidazole-based derivatives drug discovery against important protozoan diseases, such as malaria, leishmaniasis and trypanosomiasis. The more relevant chemical features and structure-activity relationship studies of these molecules are discussed for the purpose of paving the way towards the development of more viable drugs for the treatment of these parasitic infections.

Exploring microalgal and cyanobacterial metabolites with antiprotozoal activity against Leishmania and Trypanosoma parasites.

Neglected tropical diseases (NTD) like Leishmaniasis and trypanosomiasis affect millions of people annually, while currently used antiprotozoal drugs have serious side effects. Drug research based on natural products has shown that microalgae and cyanobacteria are a promising platform of biochemically active compounds with antiprotozoal activity. These unicellular photosynthetic organisms are rich in polyunsaturated fatty acids, pigments including phycocyanin, chlorophylls and carotenoids, polyphenols, bioactive peptides, terpenes, alkaloids, which have proven antioxidant, antimicrobial, antiviral, antiplasmodial and antiprotozoal properties. This review provides up-to-date information regarding ongoing studies on substances synthesized by microalgae and cyanobacteria with notable activity against Leishmania spp., Trypanosoma cruzi, and Trypanosoma brucei, the causative agents of Leishmaniasis, Chagas disease, and human African trypanosomiasis, respectively. Extracts of several freshwater or marine microalgae have been tested on different strains of Leishmania and Trypanosoma parasites. For instance, ethanolic extract of Chlamydomonas reinhardtii and Tetraselmis suecica have biological activity against T. cruzi, due to their high content of carotenoids, chlorophylls, phenolic compounds and flavonoids that are associated with trypanocidal activity. Halophilic Dunaliella salina showed moderate antileishmanial activity that may be attributed to the high ß-carotene content in this microalga. Peptides such as almiramides, dragonamides, and herbamide that are biosynthesized by marine cyanobacteria Lyngbya majuscula were found to have increased activity in micromolar scale IC50 against L. donovani, T. Cruzi, and T. brucei parasites. The cyanobacterial peptides symplocamide and venturamide isolated from Symploca and Oscillatoria species, respectively, and the alkaloid nostocarbonile isolated from Nostoc have shown promising antiprotozoal properties and are being explored for pharmaceutical and medicinal purposes. The discovery of new molecules from microalgae and cyanobacteria with therapeutic potential against Leishmaniasis and trypanosomiasis may address an urgent medical need: effective and safe treatments of NTDs.

Bis-6-amidino-benzothiazole Derivative that Cures Experimental Stage 1 African Trypanosomiasis with a Single Dose.

We designed and synthesized a series of symmetric bis-6-amidino-benzothiazole derivatives with aliphatic central units and evaluated their efficacy against bloodstream forms of the African trypanosome Trypanosoma brucei. Of these, a dicationic benzothiazole compound (9a) exhibited sub-nanomolar in vitro potency with remarkable selectivity over mammalian cells (>26,000-fold). Unsubstituted 5-amidine groups and a cyclohexyl spacer were the crucial determinants of trypanocidal activity. In all cases, mice treated with a single dose of 20 mg kg-1 were cured of stage 1 trypanosomiasis. The compound displayed a favorable in vitro ADME profile, with the exception of low membrane permeability. However, we found evidence that uptake by T. brucei is mediated by endocytosis, a process that results in lysosomal sequestration. The compound was also active in low nanomolar concentrations against cultured asexual forms of the malaria parasite Plasmodium falciparum. Therefore, 9a has exquisite cross-species efficacy and represents a lead compound with considerable therapeutic potential.

Intracellular ROS production and apoptotic effect of quinoline and isoquinoline alkaloids on the growth of Trypanosoma evansi.

Trypanosoma evansi, a hemoflagellate poses huge economic threat to the livestock industry of several countries of Asia, Africa, South America and Europe continents of the world. Limited number of available chemical drugs, incidents of growing drug resistance, and related side effects encouraged the use of herbal substitutes. In the present investigation, the impact of six alkaloids of quinoline and isoquinoline group was evaluated on the growth and multiplication of Trypanosoma evansi and their cytotoxic effect was examined on horse peripheral blood mononuclear cells in an in vitro system. Quinine, quinindine, cinchonine, cinchonidine, berbamine and emetine showed potent trypanocidal activities with IC50/24 h values 6.631 ± 0.244, 8.718 ± 0.081, 16.96 ± 0.816, 33.38 ± 0.653, 2.85 ± 0.065, and 3.12 ± 0.367 µM, respectively, which was comparable to the standard anti-trypanosomal drug, quinapyramine sulfate (20 µM). However, in the cytotoxicity assay, all the drugs showed dose dependent cytotoxic effect and quinine, berbamine and emetine showed selectivity index more than 5, based of ration of CC50 to IC50. Among the selected alkaloids, quinidine, berbamine and emetine exhibited higher apoptotic effects in T. evansi. Likewise, drug treated parasites showed a dose-dependent and time-dependent increase in reactive oxygen species (ROS) production. Therefore, increased apoptosis in combination with ROS generation could be responsible for the observed trypanocidal effect which could be further evaluated in T. evansi-infected mice model.

Effects of Synthetic Ligustrazine-Based Chalcone Derivatives on Trypanosoma brucei brucei and Leishmania spp. Promastigotes.

Current medication therapy for leishmaniasis and trypanosomiasis remains a major challenge due to its limited efficacy, significant adverse effects, and inaccessibility. Consequently, locating affordable and effective medications is a pressing concern. Because of their easy-to-understand structure and high functionalization potential, chalcones are promising candidates for use as bioactive agents. Thirteen synthetic ligustrazine-containing chalcones were evaluated for their ability to inhibit the growth of leishmaniasis and trypanosomiasis in etiologic agents. The tetramethylpyrazine (TMP) analogue ligustrazine was chosen as the central moiety for the synthesis of these chalcone compounds. The most effective compound (EC50 = 2.59 µM) was the chalcone derivative 2c, which featured a pyrazin-2-yl amino on the ketone ring and a methyl substitution. Multiple actions were observed for certain derivatives, including 1c, 2a-c, 4b, and 5b, against all strains tested. Eflornithine served as a positive control, and three ligustrazine-based chalcone derivatives, including 1c, 2c, and 4b, had a higher relative potency. Compounds 1c and 2c are particularly efficacious; even more potent than the positive control, they are therefore promising candidates for the treatment of trypanosomiasis and leishmaniasis.

Antimicrobial Peptides (AMPs): Potential Therapeutic Strategy against Trypanosomiases?

Trypanosomiases are a group of tropical diseases that have devastating health and socio-economic effects worldwide. In humans, these diseases are caused by the pathogenic kinetoplastids Trypanosoma brucei, causing African trypanosomiasis or sleeping sickness, and Trypanosoma cruzi, causing American trypanosomiasis or Chagas disease. Currently, these diseases lack effective treatment. This is attributed to the high toxicity and limited trypanocidal activity of registered drugs, as well as resistance development and difficulties in their administration. All this has prompted the search for new compounds that can serve as the basis for the development of treatment of these diseases. Antimicrobial peptides (AMPs) are small peptides synthesized by both prokaryotes and (unicellular and multicellular) eukaryotes, where they fulfill functions related to competition strategy with other organisms and immune defense. These AMPs can bind and induce perturbation in cell membranes, leading to permeation of molecules, alteration of morphology, disruption of cellular homeostasis, and activation of cell death. These peptides have activity against various pathogenic microorganisms, including parasitic protists. Therefore, they are being considered for new therapeutic strategies to treat some parasitic diseases. In this review, we analyze AMPs as therapeutic alternatives for the treatment of trypanosomiases, emphasizing their possible application as possible candidates for the development of future natural anti-trypanosome drugs.

In vitro and in vivo evaluation of efficacy of berberine chloride: Phyto-alternative approach against Trypanosoma evansi infection.

Current chemotherapy against the Surra organism, Trypanosoma evansi has several limitations in terms of efficacy, toxicity, availability and emerging resistance. These reasons make the search of new chemo-preventive and chemo-therapeutic agent with high potency and low toxicity. Alkaloid phyto-molecules, berberine has shown promising anti-kinetoplastid activity against T. cruzi, T. congolense, T. brucei, Leishmania donovani and L. tropica. However, till date, there is no investigation of therapeutic efficacy of berberine chloride (BC) against T. evansi. The IC50 value of BC for growth inhibition of T. evansi at 24 h of culture was calculated as 12.15 µM. The specific selectivity index (SSI) of BC was calculated as 19.01 and 10.43 against Vero cell line and Equine PBMC's, respectively. Thirteen drug target genes affecting various metabolic pathways were studied to investigate the mode of trypanocidal action of BC. In transcript analysis, the mRNA expression of arginine kinase 1 remained refractory to exposure with BC, which provides metabolic plasticity in adverse environmental conditions. In contrary, rest all the drug target gene were down-regulated, which indicates that drug severely affect DNA replication, cell proliferation, energy homeostasis, redox homeostasis and calcium homeostasis of T. evansi, leading to the death of parasite in low concentrations. It is the first attempt to investigate in vitro anti-trypanosomal activity of BC against T. evansi. These data imply that phytochemicals as alternative strategies can be explored in the future as an alternative treatment for Surra in animal.

An atypical Trypanosoma lewisi infection in a 22-day-old neonate from India: An emergent zoonosis.

Reports on atypical human trypanosomiasis, caused by Trypanosoma lewisi, are rare and so far a total of 19 reports on human infection with animal trypanosomes, which includes nine cases from Trypanosoma lewisi exist. Trypanosoma lewisi, a Stercorarian trypanosoma of rats, is transmitted by the fecal contamination of the wound or the bite caused by rat flea Ceratophyllus fasciatus. We report here an atypical neonatal infection of T. lewisi in a 22-day-old infant from Agra. The infant presented with a history of high fever, poor appetite, and lethargy for 3 days. The hematological parameters were normal except for a low platelet count. A high C-reactive protein (CRP) concentration of 70.49 mg/L indicated marked inflammation. The Leishman-stained thin blood smears were microscopically positive for the hemoflagellate. Based on the morphological features and further confirmed by polymerase chain reaction (PCR) assay, the hemoflagellate was identified as T. lewisi. Symptomatic treatment and antibiotic therapy helped in an uneventful recovery of the patient.

Comparative pathogenicity of drug-resistant and drug-sensitive Trypanosoma brucei and Trypanosoma congolense infections in Nigerian local dogs.

Animal trypanosomosis is an important endemic and wasting disease in sub-Saharan Africa. Its control relies on chemotherapy, and resistance to trypanocides has been widely reported. The pathogenicity of drug-resistant canine trypanosomes is not clear with scanty information available. Thus, this study assessed the comparative pathogenicity of drug-resistant and drug-sensitive Trypanosoma brucei and Trypanosoma congolense infections in dogs. Twenty Nigerian local dogs were used and were randomly assigned into five groups (A-E) of four dogs each. Group A served as the uninfected-control group, while groups B and C were infected with 106 drug-sensitive T. congolense and T. brucei. Groups D and E were infected with 106 multidrug-resistant T. congolense and T. brucei, respectively. The pre-patent period (PPP), clinical signs, level of parasitaemia (LOP), rectal temperature, body weight, packed cell volume (PCV), red blood cell count (RBC), haemoglobin concentration (HbC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), total leucocyte count (TLC) and survivability were assessed. Groups D and E had longer (p < 0.05) mean PPP than groups B and C. Also, group E dogs had lower (p < 0.05) mean LOP, longer (p < 0.05) mean survivability, and higher (p < 0.05) mean body weight, PCV, HbC and RBC than group C dogs. The clinical signs were very severe in group C dogs, compared to group E dogs. However, these parameters did not differ statistically between groups B and D. Thus, multidrug-resistant T. brucei was of lower pathogenicity than drug-sensitive T. brucei, while multidrug-resistant and drug-sensitive T. congolense had comparable pathogenicity following infection in dogs.

Gene expression study to elucidate the anti-trypanosomal activity of quinapyramine methyl sulphate (QPS).

The kinetoplastid protozoan parasite, Trypanosoma evansi causes a fatal disease condition known as Surra in equines throughout the globe. Disease condition being acute in nature, entrust a huge economic and health impact on the equine industry. Till date, quinapyramine methyl sulphate (QPS) is the first line of treatment and a panacea for the T. evansi infection in equines. Still after the >70 years of its discovery, there is no clue about the mode of action of QPS in T. evansi. The establishment of in vitro cultivation of T. evansi in HMI-9 media has provided opportunity to study the alteration in mRNA expression of parasite on exposure to the drug. With this research gap, the present study aimed to investigate the relative mRNA expression of 13 important drug target genes to elucidate the anti-trypanosomal activity of QPS against T. evansi. The IC50 of QPS against a pony isolate of T. evansi was determined as 276.4 nM(147.21 ng/ mL) in the growth inhibitory assay. The in vitro cultured T. evansi population were further exposed to IC50 of QPS and their relative mRNA expression was studied at 12 h, 24 h and 48 h interval.The mRNA expression of several genes such as hexokinase, trypanothione reductase, aurora kinase, oligopeptidase B and ribonucleotide reductase II were found refractory (non-significant, p > 0.1234) to the exposure of QPS. Significant up-regulation of trans-sialidase (p < 0.0001), ESAG8 (p < 0.0021), ribonucleotide reductase I (p < 0.0001), ornithine decarboxylase (p < 0.0001), topoisomerase II (p < 0.0021) and casein kinase I (p < 0.0021) were recorded after exposure with QPS. The arginine kinase 1 and calcium ATPase I showed highly significant (p < 0.0001) down-regulation in the drug kinetics. Therefore, the arginine kinase 1 and calcium ATPase I can be explored further to elucidate the trypanocidal activity of QPS. The preliminary data generated provide the potential of arginine kinase 1 and calcium ATPase I mRNA mediated pathway of trypanocidal action of QPS. Further, transcriptomics approach is required to investigate the possible mechanism of action of drugs at molecular level against the targeted organism.

In vitro anti-trypanosomal effect of ivermectin on Trypanosoma evansi by targeting multiple metabolic pathways.

High cytotoxicity and increasing resistance reports of existing chemotherapeutic agents against T. evansi have raised the demand for novel, potent, and high therapeutic index molecules for the treatment of surra in animals. In this regard, repurposing approach of drug discovery has provided an opportunity to explore the therapeutic potential of existing drugs against new organism. With this objective, the macrocyclic lactone representative, ivermectin, has been investigated for the efficacy against T. evansi in the axenic culture medium. To elucidate the potential target of ivermectin in T. evansi, mRNA expression profile of 13 important drug target genes has been studied at 12, 24, and 48 h interval. In the in vitro growth inhibition assay, ivermectin inhibited T. evansi growth and multiplication significantly (p < 0.001) with IC50 values of 13.82 µM, indicating potent trypanocidal activity. Cytotoxicity assays on equine peripheral blood mononuclear cells (PBMCs) and Vero cell line showed that ivermectin affected the viability of cells with a half-maximal cytotoxic concentration (CC50) at 17.48 and 22.05 µM, respectively. Data generated showed there was significant down-regulation of hexokinase (p < 0.001), ESAG8 (p < 0.001), aurora kinase (p < 0.001), casein kinase 1 (p < 0.001), topoisomerase II (p < 0.001), calcium ATPase 1 (p < 0.001), ribonucleotide reductase I (p < 0.05), and ornithine decarboxylase (p < 0.01). The mRNA expression of oligopeptidase B remains refractory to the exposure of the ivermectin. The arginine kinase 1 and ribonucleotide reductase II showed up-regulation on treatment with ivermectin. The ivermectin was found to affect glycolytic pathways, ATP-dependent calcium ATPase, cellular kinases, and other pathway involved in proliferation and maintenance of internal homeostasis of T. evansi. These data imply that intervention with alternate strategies like nano-formulation, nano-carriers, and nano-delivery or identification of ivermectin homologs with low cytotoxicity and high bioavailability can be explored in the future as an alternate treatment for surra in animals.

Nucleoside analogues for the treatment of animal trypanosomiasis.

Animal trypanosomiasis (AT) is a parasitic disease with high socio-economic impact. Given the limited therapeutic options and problems of toxicity and drug resistance, this study assessed redirecting our previously identified antitrypanosomal nucleosides for the treatment of AT. Promising hits were identified with excellent in vitro activity across all important animal trypanosome species. Compound 7, an inosine analogue, and our previously described lead compound, 3'-deoxytubercidin (8), showed broad spectrum anti-AT activity, metabolic stability in the target host species and absence of toxicity, but with variable efficacy ranging from limited activity to full cure in mouse models of Trypanosoma congolense and T. vivax infection. Several compounds show promise against T. evansi (surra) and T. equiperdum (dourine). Given the preferred target product profile for a broad-spectrum compound against AT, this study emphasizes the need to include T. vivax in the screening cascade given its divergent susceptibility profile and provides a basis for lead optimization towards such broad spectrum anti-AT compound.

Ruthenium Metallotherapeutics: Novel Approaches to Combatting Parasitic Infections.

Human parasitic infections cause a combined global mortality rate of over one million people per annum and represent some of the most challenging diseases for medical intervention. Current chemotherapeutic strategies often require prolonged treatment, coupled with subsequent drug-induced cytotoxic morbidity to the host, while resistance generation is also a major concern. Metals have been used extensively throughout the history of medicine, with more recent applications as anticancer and antimicrobial agents. Ruthenium metallotherapeutic antiparasitic agents are highly effective at targeting a range of key parasites, including the causative agents of malaria, trypanosomiasis, leishmaniasis, amoebiasis, toxoplasmosis and other orphan diseases, while demonstrating lower cytotoxicity profiles than current treatment strategies. Generally, such compounds also demonstrate activity against multiple cellular target sites within parasites, including inhibition of enzyme function, cell membrane perturbation, and alterations to metabolic pathways, therefore reducing the opportunity for resistance generation. This review provides a comprehensive and subjective analysis of the rapidly developing area of ruthenium metal- based antiparasitic chemotherapeutics, in the context of rational drug design and potential clinical approaches to combatting human parasitic infections.