Anti-Toxoplasma gondii screening of MMV pandemic response box and evaluation of RWJ-67657 efficacy in chronically infected mice.

Toxoplasmosis is a significant public health concern with limited therapeutic options. The medicines for malaria venture (MMV) developed the pandemic response box (PRB) containing 400 drug-like molecules with broad pathogen activity. The aim of this work is to evaluate PRB compounds for their anti-Toxoplasma gondii activity and identify promising candidates for further evaluation. Screening identified 42 selective compounds with half effective concentration (EC50) ranging from 2.4 to 913.1 nm and half cytotoxic concentration (CC50) ranging from 6 µm to >50 µm. Selectivity index (SI) values (CC50/EC50) ranged from 11 to 17 708. Based on its in silico and in vitro profile and its commercial availability, RWJ-67657 was selected for further studies. Molecular docking analysis showed RWJ-67657 is predicted to bind to T. gondii p38 mitogen-activated protein kinase (TgMAPK). Oral administration of RWJ-67657 (20 mg kg day−1/10 days) significantly reduced parasite burden in chronically infected mice compared to mock-treated group (P < 0.01). These findings highlight the PRB as a promising source for anti-T. gondii compounds, with several showing favourable drug properties, including MMV1634492, MMV002731, MMV1634491, MMV1581551, MMV011565, MMV1581558, MMV1578577, MMV233495 and MMV1580482, firstly described here as anti-T. gondii agents. RWJ-67657 emerges as a valuable drug candidate for experimental chronic cerebral toxoplasmosis therapy.

Chemical Composition, Anthelmintic Activity, and Mechanism of Action of Lippia dominguensis Mold. Essential Oil on Haemonchus contortus.

Gastrointestinal nematode parasitism is a major burden to small ruminant production globally, compounded by increasing anthelmintic resistance. Previous studies have identified essential oils (EOs) from the Lippia genus with antiprotozoal and anthelmintic effects. Lippia dominguensis Moldenke (Ld), an endemic specie from the Dominican Republic, has similar popular uses, however, is chemically and pharmacologically yet uncharacterized. Here, we investigated the in vitro anthelmintic activity of LdEO and its ultrastructural effects on eggs and adult nematodes of Haemonchus contortus multidrug-resistant isolated. The GC/MS analysis showed linalool (33.85 %), 1,8-cineole (30.88 %), and δ-terpineol (10.61 %) as the main EO constituents. The LdEO showed an IC50 =0.523 mg/mL in the egg hatch test, and the motility in the adult worm motility test was 95.8 % at 1 mg/mL. The confocal scanning laser microscopy of eggs indicated permeabilization or disruption of egg cell membranes as the possible mechanism of action of LdEO. The scanning electron microscopy of adult worms showed wrinkling, undulations, and cuticular disruptions. The LdEO displayed significant in vitro anthelmintic activity on eggs and adult worms of H. contortus. Additionally, the LdEO showed low oral toxicity in mice at 2,000 mg/kg. Thus, additional in vivo studies are justified to determine its anthelmintic efficacy in small ruminants.

In vivo evaluation of anti-Leishmania activity of alkyltriazoles and alkylphosphocholines by oral route.

The failures in the treatment of leishmaniasis is an increasing problem around the world, especially related to resistance. Thus, we describe the synthesis and in vivo anti-Leishmania activity of alkylphosphocholine and alkyltriazoles; besides, their likely action mechanisms stem from some eventual inhibition of parasite enzymes using computational tools. These compounds were tested in an in vivo hamster model infected with Leishmania Leishmania infantum chagasi. Fifty days after parasite inoculation, the two compounds 12-azidedodecylphosphocholine (3) and 3-(1-(12-fluorododecyl)-1H-1,2,3-triazol-1-yl)propano-1-ol (9), were separately administered once a day as oral suspensions (25 and 12.5 mg/kg/day, respectively) during ten days, and their efficacy was compared to the reference compound pentavalent antimonial Glucantime (GLU). Compound 3 significantly reduced the number of parasites in the spleen (4.93 × 102 amastigotes/g) and liver (4.52 × 103 amastigotes/g). Compound 9 reduced the number of amastigotes in the spleen to 1.30 × 104 and 1.36 × 103 amastigotes/g in the liver. GLU was the most effective overall treatment (7.50 × 101 and 2.28 × 102 amastigotes/g in the spleen and liver, respectively). The high activity levels of these compounds in vivo may stem from their high in vitro leishmanicidal activity and lipophilicity. The in silico absorption, distribution, metabolism, and excretion studies also showed some anti-Leishmania potential. Compound 9 had more lipophilic characteristics than those of compound 3. In silico studies of the nine enzymes of compounds 3 and 9 showed significant evidence of interactions with nicotimidase and tyrosine aminotransferase, demonstrating possible inhibition enzymes present in L. (L.) infantum chagasi. These compounds could be a promising template for developing a new class of leishmanicidal agents, by oral route, and deserve further investigation to explore different therapeutic regimens.

Amlodipine Increases the Therapeutic Potential of Ravuconazole upon Trypanosoma cruzi Infection.

Mining existing agents that enhance the therapeutic potential of ergosterol biosynthesis inhibitors (EBI) is a promising approach to improve Chagas disease chemotherapy. In this study, we evaluated the effect of ravuconazole, an EBI, combined with amlodipine, a calcium channel blocker, upon Trypanosoma cruzi experimental infection. In vitro assays confirmed the trypanocidal activity of both compounds in monotherapy and demonstrated an additive effect (sum of the fractional inhibitory concentration [ΣFIC] > 0.5) of the combined treatment without additional toxicity to host cells. In vivo experiments, using a murine model of the T. cruzi Y strain in a short-term protocol, demonstrated that amlodipine, although lacking trypanocidal activity, dramatically increased the antiparasitic activity of underdosing ravuconazole regimens. Additional analysis using long-term treatment (20 days) showed that parasitemia relapse until 60 days after treatment was significatively lower in mice treated with the combination (4 out of 14 mice) than ravuconazole monotherapy (10 out of 14 mice), even in the presence of immunosuppressant pressure. Furthermore, the combined therapy was well tolerated and protected the mice from mortality. The treatments also impacted on the cellular and humoral immune response of infected animals, inducing a reduction of serum cytokine levels in all ravuconazole-treated mice. Our findings demonstrate that amlodipine is efficacious in enhancing the antiparasitic activity of ravuconazole in an experimental model of T. cruzi infection and indicates a potential strategy to be explored in Chagas disease treatment.

Investigation of 8-methoxy-3-(4-nitrobenzoyl)-6-propyl-2H-chromen-2-one as a promising coumarin compound for the development of a new and orally effective antileishmanial agent.

Changes in host immunity and parasite resistance to drugs are among the factors that contribute to decreased efficacy of antiparasitic drugs such as the antimonial compounds pentamidine, amphotericin (AMP B) and miltefosine. Bioactive natural products could be alternatives for the development of new drugs to treat neglected human diseases such as leishmaniasis. Natural coumarins and synthetic analogues have shown leishmanicidal activity, mainly in vitro. This study investigated the in vitro and in vivo leishmanicidal activity of synthetic coumarin compounds (C1-C5) in parasites Leishmania (L.) amazonensis and L. (L.) infantum chagasi. The cytotoxicity of these compounds in mammalian cells and their influence on production of reactive oxygen species was also investigated. In vitro assays showed that 8-methoxy-3-(4-nitrobenzoyl)-6-propyl-2H-chromen-2-one (C4) was as active as AMP B mainly in the amastigote form (p < 0.05); C4 presented a selectivity index (65.43) four times higher than C2 (15.4) in L. amazonensis and six times higher (33.94) than C1 (5.46) in L. infantum chagasi. Additionally, coumarin C4 reduced the H2O2 concentration 32.5% more than the control group in L. amazonensis promastigotes during the lag phase of proliferation. No interference of C4 was observed on the mitochondrial membrane potential of the parasites. In vivo, coumarin C4 in corn oil (oral route) led to a reduction in the number of amastigotes from L. infantum chagasi to 1.31 × 106 and 4.09 × 104 in the spleen and liver, respectively (p < 0.05). Thus, C4 represents a candidate for further studies aiming at new treatments of leishmaniasis.

Synthesis and evaluation of the antileishmanial activity of silver compounds containing imidazolidine-2-thione.

A new series of silver compounds could be of interest on designing new drugs for the treatment of leishmaniasis. The compounds [Ag(phen)(imzt)]NO3(1), [Ag(phen)(imzt)]CF3SO3(2), [Ag(phen)2](BF4)·H2O (3), [Ag2(imzt)6](NO3)2(4), and imzt have been synthesized and evaluated in vitro for antileishmanial activity against Leishmania. (L.) amazonensis (La) and L. (L.) chagasi (Lc), and two of them were selected for in vivo studies. In addition to investigating the action on Leishmania, their effects on the hydrogen peroxide production and cysteine protease inhibition have also been investigated. As for antileishmanial activity, compound (4) was the most potent against promastigote and amastigote forms of La (IC50 = 4.67 and 1.88 µM, respectively) and Lc (IC50 = 9.35 and 8.05 µM, respectively); and comparable to that of amphotericin B, reference drug. Beside showing excellent activity, it also showed a low toxicity. In the in vivo context, compound (4) reduced the number of amastigotes in the liver and spleen when compared to the untreated group. In evaluating the effect of the compounds on Leishmania, the level of hydrogen peroxide production was maintained between the lag and log phases; however, in the treatment with compound (4) it was possible to observe a reduction of 25.44 and 49.13%, respectively, in the hydrogen peroxide rates when compared to the lag and log phases. It was noticed that the presence of a nitrate ion and imzt in compound (4) was important for the modulation of the antileishmanial activity. Thus, this compound can represent a potentially new drug for the treatment of leishmaniasis.

Leishmania infantum amastigote nucleoside triphosphate diphosphohydrolase 1 (NTPDase 1): Its inhibition as a new insight into mode of action of pentamidine.

Nucleoside triphosphate diphosphohydrolase (NTPDase) 1 from intracellular amastigotes of Leishmania infantum-infected macrophage was identified by immunocytochemistry and confocal laser scanning microscopy using antibodies that specifically recognize its B-domain. This enzyme was previously characterized in Leishmania promastigote form, and here it is shown to be susceptible to pentamidine isethionate (PEN). In initial assays, this antileishmanial compound (100 µM) reduced 60% phosphohydrolytic activity of promastigotes preparation. An active NTPDase 1 was then isolated by non-denaturing gel electrophoresis, and PEN (10 µM) inhibited 74% and 35% of the ATPase and ADPase activities, respectively, of this pure protein. In addition, PEN 0.1-1 µM inhibited 56% potato apyrase activity, a plant protein that shares high identity with Leishmania NTPDase 1. In contrast, amphotericin B, fluconazole, ketoconazole or allopurinol did not significantly affect phosphohydrolytic activity of either promastigotes preparation or potato apyrase. This work suggests amastigote NTPDase 1 as a new molecular target, and inhibition of its catalytic activity by pentamidine can be part of the mode of action of this drug contributing with the knowledge of its antileishmanial effect.

Comparative analysis of real-time PCR assays in the detection of canine visceral leishmaniasis.

Dogs are important hosts and reservoirs of leishmaniasis, a disease caused by protozoan parasites from the genus Leishmania, affecting ~12 million people worldwide. The detection of visceral leishmaniasis (VL) in dogs by real-time PCR (qPCR) may improve on diagnosis, but the different qPCR methods available for Leishmania DNA detection have not been established as routine in diagnostic tools and/or epidemiologic studies for canine VL. Here, we compared three qPCR assays (DNApol, Linj31, and LDON) in the detection of VL by Leishmania infantum in spleen (n = 48; 7), skin (n = 48; 7), and whole blood (n = 44; 7) samples from serologically positive and negative dogs, respectively. Overall, the DNApol performed better than the Linj31 and LDON assays in the detection of positive samples in all tissues tested, yielding from 66.7 to 100.0% of positivity for both skin and spleen samples. For spleen samples, we observed no statistically significant differences between positive detection by the LDON and DNApol assays. Whole blood samples yielded the lowest rates of positive detection, regardless of the qPCR assay used. In contrast, positive detection of Leishmania DNA was as efficient from skin samples using the DNApol assay as from spleen samples using either the DNApol or the LDON assay. Although qPCR assays from skin samples may not be practical for use in the field, our study suggests that the DNApol and LDON assays from skin samples could be used in future to evaluate canine VL treatment in veterinary clinics.

In vitro evaluation of the schistosomicidal effect of the extracts, fractions and major 3-hydroxy-2,6-dialkyl-substituted piperidine alkaloids from the flowers of Senna spectabilis (Fabaceae).

In this work, we present the in vitro schistosomicidal activity evaluation of the most active dichloromethane fraction (FDm) (ED50=83.5µg/mL) and of a mixture of the major alkaloids ((-)-cassine/(-)-spectaline, C/E) (ED50=37.4µg/mL) from the flowers of Senna spectabilis against adult worms and cercariae. We also demonstrate other toxic effects including paralysis of the adult worms, inhibition of the secretory activity, tegument lesions and cercaricidal activity. In the association test of Praziquantel (PZQ)-C/E, we observed up to 80% mortality of Schistosoma mansoni in comparison to PZQ monotherapy. Due to the diversity of the toxic effects, the schistosomicidal activity of C/E is likely a result of a multitarget mechanism involving the tegument, secretory system and neuromotor action.

Potent Schistosomicidal Constituents from Garcinia brasiliensis.

Praziquantel is the drug of choice for the treatment of schistosomiasis. However, several strains of Schistosoma mansoni are resistant to praziquantel, making it necessary to discover new drugs that might be used for its treatment. With this in mind, the properties of a schistosomicidal ethanolic extract of Garcinia brasiliensis Mart. epicarp, the fractions obtained by partitioning this extract, including the hexane fractions, ethyl acetate fraction, and the aqueous fraction, and the isolated compounds 7-epiclusianone, a major component from these fractions, and fukugetin were tested in vitro on adult worms of S. mansoni. Mortality, damage to membranes, and excretory system activity were observed at 100.0, 50.0, 75.0, and 14.0 µg/mL for the ethanolic extract of G. brasiliensis Mart. epicarp, its hexane fractions, the ethyl acetate fraction, and 7-epiclusianone, respectively. For 7-epiclusianone, these data were confirmed by fluorescent probe Hoechst 33 258 and resorufin. Additionally, the biocidal effect of 7-epiclusianone was even higher than the hexane fractions. Moreover, an inhibitory effect of 7-epiclusianone on the egg laying of female adult S. mansoni worms was observed in cercariae and schistossomula. Thus, 7-epiclusianone is a promising schistosomicidal compound; however, more studies are needed to elucidate its mechanism of toxicity and to evaluate the in vivo activity of this compound.

Shelf life and quality study of minced tilapia with Nori and Hijiki seaweeds as natural additives.

The extraction of mechanically separated meat has emerged as an attractive process. However, it increases the incorporation of oxygen and, consequently, of flavors due to rancidity. Thus, preservatives must be added. The objective of this study was to evaluate the shelf life of minced tilapia to replace synthetic preservatives with Hijiki and Nori seaweeds extracts. The application of the extracts had no effect on the chemical composition of the minced tilapia. The seaweed extracts had inhibitory effect on total volatile base nitrogen. The minced tilapia complied with the microbiological standard set by Brazilin law. The panelists detected no differences in the rancid aroma and only minor differences were detected in the color of the products. It can be concluded that the minced tilapia with added seaweed extracts were within quality standards during frozen storage.

First Description of Marinoquinoline Derivatives' Activity against Toxoplasma gondii.

Toxoplasmosis is a globally prevalent zoonotic disease with significant clinical implications, including neurotoxoplasmosis, a leading cause of cerebral lesions in AIDS patients. The current pharmacological treatments for toxoplasmosis face clinical limitations, necessitating the urgent development of new therapeutics. Natural sources have yielded diverse bioactive compounds, serving as the foundation for clinically used derivatives. The exploration of marine bacteria-derived natural products has led to marinoquinolines, which feature a pyrroloquinoline core and demonstrate in vitro and in vivo anti-Plasmodium activity. This study investigates the in vitro anti-Toxoplasma gondii potential of six marinoquinoline derivatives. Additionally, it conducts absorption, distribution, metabolism, excretion, and toxicity (ADMET) predictions, and evaluates the in vivo efficacy of one selected compound. The compounds displayed half-maximal effective concentration (EC50) values between 1.31 and 3.78 µM and half-maximal cytotoxic concentration (CC50) values ranging from 4.16 to 30.51 µM, resulting in selectivity indices (SI) from 3.18 to 20.85. MQ-1 exhibiting the highest in vitro SI, significantly reduced tachyzoite numbers in the peritoneum of RH-infected Swiss mice when it was orally administered at 12.5 mg/kg/day for eight consecutive days. Also, MQ-1 significantly reduced the cerebral parasite burden in chronically ME49 infected C57BL/6 mice when it was orally administered at 25 mg/kg/day for 10 consecutive days. These findings underscore the promising anti-T. gondii activity of marinoquinolines and their potential as novel therapeutic agents against this disease.

Synthesis and biological evaluation against Leishmania amazonensis of a series of alkyl-substituted benzophenones.

Nine O-alkyl and O-prenyl derivatives were synthesized from commercial 2,4-dihydroxybenzophenone, 4,e4,4'-dihydroxybenzophenone and were evaluated for their leishmanicidal activity against promastigote forms of Leishmania amazonensis, as well their toxicity in murine macrophages. All derivatives exhibited better biological activity than their hydroxylated benzophenones precursors, and new compound LFQM-123 (3c) was 250-fold more active than its precursor 4,4'-dihydroxybenzophenone (3). Moreover, some of the results were comparable to the standard drug Amphotericin B, suggesting that the increase in lipophilicity could facilitate protozoa membrane permeation. In this study we confirmed that benzophenone derivatives exhibit leishmanicidal properties, with relatively low toxicity, and thus could be exploited as promise prototypes for the design and development of new drug for the treatment of leishmaniasis.

Repurposing the Medicines for Malaria Venture's COVID Box to discover potent inhibitors of Toxoplasma gondii, and in vivo efficacy evaluation of almitrine bismesylate (MMV1804175) in chronically infected mice.

Toxoplasmosis, caused by the obligate intracellular parasite Toxoplasma gondii, affects about one-third of the world's population and can cause severe congenital, neurological and ocular issues. Current treatment options are limited, and there are no human vaccines available to prevent transmission. Drug repurposing has been effective in identifying anti-T. gondii drugs. In this study, the screening of the COVID Box, a compilation of 160 compounds provided by the "Medicines for Malaria Venture" organization, was conducted to explore its potential for repurposing drugs to combat toxoplasmosis. The objective of the present work was to evaluate the compounds' ability to inhibit T. gondii tachyzoite growth, assess their cytotoxicity against human cells, examine their absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties, and investigate the potential of one candidate drug through an experimental chronic model of toxoplasmosis. Early screening identified 29 compounds that could inhibit T. gondii survival by over 80% while keeping human cell survival up to 50% at a concentration of 1 µM. The Half Effective Concentrations (EC50) of these compounds ranged from 0.04 to 0.92 µM, while the Half Cytotoxic Concentrations (CC50) ranged from 2.48 to over 50 µM. Almitrine was chosen for further evaluation due to its favorable characteristics, including anti-T. gondii activity at nanomolar concentrations, low cytotoxicity, and ADMET properties. Administering almitrine bismesylate (Vectarion®) orally at dose of 25 mg/kg/day for ten consecutive days resulted in a statistically significant (p < 0.001) reduction in parasite burden in the brains of mice chronically infected with T. gondii (ME49 strain). This was determined by quantifying the RNA of living parasites using real-time PCR. The presented results suggest that almitrine may be a promising drug candidate for additional experimental studies on toxoplasmosis and provide further evidence of the potential of the MMV collections as a valuable source of drugs to be repositioned for infectious diseases.

Case Report: Human Bertiellosis in Brazil.

The Anoplocephalidae family comprises a group of parasites that affect reptiles, birds, and mammals. Humans can be accidentally infected by ingesting contaminated mites. We present a case of human bertiellosis in Brazil. Our report reinforces the importance of correctly identifying the parasite to provide adequate treatment.

Leishmania (Viannia) braziliensis nucleoside triphosphate diphosphohydrolase (NTPDase 1): localization and in vitro inhibition of promastigotes growth by polyclonal antibodies.

Nucleoside triphosphate diphosphohydrolase (NTPDase) activity was recently characterized in Leishmania (Viannia) braziliensis promastigotes (Lb), and an antigenic conserved domain (r82-121) from the specific NTPDase 1 isoform was identified. In this work, mouse polyclonal antibodies produced against two synthetic peptides derived from this domain (LbB1LJ, r82-103; LbB2LJ, r102-121) were used. The anti-LbB1LJ or anti-LbB2LJ antibodies were immobilized on protein A-sepharose and immunoprecipitated the NTPDase 1 of 48 kDa and depleted approximately 40% of the phosphohydrolytic activity from detergent-homogenized Lb preparation. Ultrastructural immunocytochemical microscopy identified the NTPDase 1 on the parasite surface and in its subcellular cytoplasmic vesicles, mitochondria, kinetoplast and nucleus. The ATPase and ADPase activities of detergent-homogenized Lb preparation were partially inhibited by anti-LbB1LJ antibody (43-79%), which was more effective than that inhibition (18-47%) by anti-LbB2LJ antibody. In addition, the immune serum anti-LbB1LJ (67%) or anti-LbB2LJ (33%) was cytotoxic, significantly reducing the promastigotes growth in vitro. The results appoint the conserved domain from the L. braziliensis NTPDase as an important target for inhibitor design and the potential application of these biomolecules in experimental protocols of disease control.

Unbalanced relationships: insights into the interaction between gut microbiota, geohelminths, and schistosomiasis.

Hosts and their microbiota and parasites have co-evolved in an adaptative relationship since ancient times. The interaction between parasites and intestinal bacteria in terms of the hosts' health is currently a subject of great research interest. Therapeutic interventions can include manipulations of the structure of the intestinal microbiota, which have immunological interactions important for modulating the host's immune system and for reducing inflammation. Most helminths are intestinal parasites; the intestinal environment provides complex interactions with other microorganisms in which internal and external factors can influence the composition of the intestinal microbiota. Moreover, helminths and intestinal microorganisms can modulate the host's immune system either beneficially or harmfully. The immune response can be reduced due to co-infection, and bacteria from the intestinal microbiota can translocate to other organs. In this way, the treatment can be compromised, which, together with drug resistance by the parasites makes healing even more difficult. Thus, this work aimed to understand interactions between the microbiota and parasitic diseases caused by the most important geohelminths and schistosomiasis and the consequences of these associations.

In vitro and in silico evaluation of the schistosomicidal activity of eugenol derivatives using biochemical, molecular, and morphological tools.

Background: Eugenol shows both antibacterial and antiparasitic activities, suggesting that it might be evaluated as an option for the treatment of praziquantel-resistant schistosome. Methods: The in vitro activities of three eugenol derivatives (FB1, FB4 and FB9) on adult worms from Schistosoma mansoni were examined by fluorescence and scanning electron microscopy to analyze effects on the excretory system and integument damage, respectively. Biochemical tests with verapamil (a calcium channel antagonist) and ouabain (a Na+/K+-ATPase pump inhibitor) were used to characterize eugenol derivative interactions with calcium channels and the Na+/K+-ATPase, while in silico analysis identified potential Na+/K+-ATPase binding sites. Results: The compounds showed effective doses (ED50) of 0.324 mM (FB1), 0.167 mM (FB4), and 0.340 mM (FB9). In addition, FB4 (0.322 mM), which showed the lowest ED50, ED90 and ED100 (p < 0.05), caused the most damage to the excretory system and integument, according to both fluorescence and scanning electron microscopy analysis. The death of adult worms was delayed by ouabain treatment plus FB1 (192 versus 72 hours) and FB9 (192 versus 168 hours), but the response to FB4 was the same in the presence or absence of ouabain. Besides, no changes were noted when all of the eugenol derivatives were combined with verapamil. Moreover, FB1 and FB9 inhibited Na+/K+-ATPase activity according to in silico analysis but FB4 did not show a time-dependent relationship and may act on targets other than the parasite Na+/K+-ATPase. Conclusion: Eugenol derivatives, mainly FB4 when compared to FB1 and FB9, seem to act more effectively on the integument of adult S. mansoni worms.

Risk of Trypanosoma cruzi transmission in southern Minas Gerais, Brazil - Data from 2014 to 2020.

Trypanosoma cruzi, the etiologic agent of Chagas disease, is widely distributed in the Americas and is transmitted through vectorial, transfusional, and oral routes. This study aimed to evaluate the risk of vectorial transmission of Chagas disease in municipalities located in southern Minas Gerais, Brazil, by analyzing triatomine specimens collected from 2014 to 2020. All 1522 hematophagous triatomines were identified as Panstrongylus megistus, and were subjected to parasitological and molecular examinations. From 2014 to 2016, approximately 10% of insects were positive in the microscopic analysis of intestinal content, and 27% were positive as detected by the quantitative polymerase chain reaction (qPCR) of the same sampling. However, in the last investigated years, an increase in infected triatomines was observed in microscopic analysis (22%) and qPCR methods (41%). This corroborates the findings of acute human Chagas disease cases, which have increased in the study area from a maximum of 2 cases in previous years to 20 cases in 2019, and 17 cases in 2020 through June. Additionally, bloodmeal sources of infected triatomines were investigated; human blood was detected in up to 85.7% of the samples. Moreover, canine blood was also detected in triatomine intestinal content in recent years, reaching 91% of analyzed insects in 2018. Data on bloodmeal sources have demonstrated human-vector contact and have suggested the participation of dogs in the parasite transmission cycle. These results indicate the risk of T. cruzi vectorial transmission in Southern Minas Gerais and São Paulo owing to the boundary between these states. Thus, enhanced surveillance and vector control of Chagas disease are highly recommended in these areas.

Exploring the bioactivity potential of Leonotis nepetifolia: phytochemical composition, antimicrobial and antileishmanial activities of extracts from different anatomical parts.

Leonotis nepetifolia (L.) Br. (Lamiaceae) is an African shrub popularly known as 'cordão-de-frade' in Brazil, traditionally used to treat infectious diseases, among other uses. This study aimed to investigate the phytochemical composition of hydroethanolic extracts from L. nepetifolia prepared from stems, leaves, roots and glomerulus, as well as their cytotoxicity, antileishmanial and antimicrobial activities. The chemical composition of the extracts was assessed by UPLC-ESI-MS/MS, whereas the antileishmanial activity was evaluated against promastigote and amastigote forms of Leishmania amazonensis. Cytotoxicity was tested on murine macrophages and the antimicrobial activity was investigated by a microdilution assay against several strains of fungi, Gram-positive and Gram-negative bacteria. The flavonoids apigenin, cirsiliol apigenin-7-O-glucoside, luteolin, luteolin-4'-O-glucoside, luteolin-4'-O- glucuronide and luteolin-7-O-glucoside were identified in all tested extracts. Extracts from leaves and roots showed more potent antileishmanial activity (IC50 32.90 µg mL-1 and 57.70 µg mL-1, respectively) against amastigotes forms in comparison to the other extracts. The leaf extract inhibited Bacillus cereus and Staphylococcus aureus growth (125 µg mL-1 and 100 µg mL-1, respectively), and also showed anti-Candida activity (10-125 µg mL-1). The biological effect can be related to the identified flavonoids. Our findings disclose the potential of L. nepetifolia as a source of bioactive compounds for the development of new therapeutic options for treating infectious diseases, especially flavonoids.