Expanding the antiprotozoal activity and the mechanism of action of n-butyl and iso-butyl ester of quinoxaline-1,4-di-N-oxide derivatives against Giardia lamblia, Trichomonas vaginalis, and Entamoeba histolytica. An in vitro and in silico approach.

In this study, n-butyl and iso-butyl quinoxaline-7-carboxylate-1,4-di-N-oxide derivatives were evaluated in vitro against Giardia lamblia (G. lamblia), Trichomonas vaginalis (T. vaginalis), and Entamoeba histolytica (E. histolytica). The potential mechanism of action determination was approached by in silico analysis on G. lamblia and T. vaginalis triosephosphate isomerase (GlTIM and TvTIM, respectively), and on E. histolytica thioredoxin reductase (EhTrxR). Enzyme inactivation assays were performed on recombinant GlTIM and EhTrxR. Compound T-167 showed the best giardicidal activity (IC50 = 25.53 nM) and the highest inactivation efficiency against GlTIM without significantly perturbing its human homolog. Compounds T-142 and T-143 showed the best amoebicidal (IC50 = 9.20 nM) and trichomonacidal (IC50 = 45.20 nM) activity, respectively. Additionally, T-143 had a high activity as giardicial (IC50 = 29.13 nM) and amoebicidal (IC50 = 15.14 nM), proposing it as a broad-spectrum antiparasitic agent. Compounds T-145, and T-161 were the best EhTrxR inhibitors with IC50 of 16 µM, and 18 µM, respectively.

Characterization of the Effect of N-(2-Methoxyphenyl)-1-methyl-1H-benzimidazol-2-amine, Compound 8, against Leishmania mexicana and Its In Vivo Leishmanicidal Activity.

Chemotherapy currently available for leishmaniasis treatment has many adverse side effects and drug resistance. Therefore, the identification of new targets and the development of new drugs are urgently needed. Previously, we reported the synthesis of a N-(2-methoxyphenyl)-1-methyl-1H-benzimidazol-2-amine, named compound 8, with an IC50 value in the micromolar range against L. mexicana, it also inhibited 68.27% the activity of recombinant L. mexicana arginase. Herein, we report studies carried out to characterize the mechanism of action of compound 8, as well as its in vivo leishmanicidal activity. It was shown in our ultrastructural studies that compound 8 induces several changes, such as membrane blebbing, the presence of autophagosomes, membrane detachment and mitochondrial and kinetoplast disorganization, among others. Compound 8 triggers the production of ROS and parasite apoptosis. It reduced 71% of the parasite load of L. mexicana in an experimental model of cutaneous leishmaniasis in comparison with a control. Altogether, the data obtained suggest the potential use of compound 8 in the treatment of cutaneous leishmaniasis.

TATA-Binding Protein-Based Virtual Screening of FDA Drugs Identified New Anti-Giardiasis Agents.

Parasitic diseases, predominantly prevalent in developing countries, are increasingly spreading to high-income nations due to shifting migration patterns. The World Health Organization (WHO) estimates approximately 300 million annual cases of giardiasis. The emergence of drug resistance and associated side effects necessitates urgent research to address this growing health concern. In this study, we evaluated over eleven thousand pharmacological compounds sourced from the FDA database to assess their impact on the TATA-binding protein (TBP) of the early diverging protist Giardia lamblia, which holds medical significance. We identified a selection of potential pharmacological compounds for combating this parasitic disease through in silico analysis, employing molecular modeling techniques such as homology modeling, molecular docking, and molecular dynamics simulations. Notably, our findings highlight compounds DB07352 and DB08399 as promising candidates for inhibiting the TBP of Giardia lamblia. Also, these compounds and DB15584 demonstrated high efficacy against trophozoites in vitro. In summary, this study identifies compounds with the potential to combat giardiasis, offering the prospect of specific therapies and providing a robust foundation for future research.

In vitro, ex vivo and in vivo short-term screening of DHEA nitrate derivatives activity over Trypanosoma cruzi Ninoa and TH strains from Oaxaca State, México.

Chagas disease is a health problem that affects millions of persons, currently Nifurtimox (Nfx) and Benznidazole (Bz) are the unique drugs to treat it. However, these drugs produce adverse effects and high toxicity, which has motivated the search for new candidate drugs. Based on reports about the extensive biological activity of steroidal nitrate esters, in this study three nitrate esters steroids (1b, 2b and 4b) were synthetized and characterized from Dehydroepiandrosterone (DHEA, 1a), 19-hydroxy-DHEA (2a), and Androst-5-en-3ß,17ß-diol (4a), respectively. In addition, compounds 3a and 3b were obtained by introducing an α-ethynyl and a ß-hydroxyl groups at position 17 of 2b and further nitration of the hydroxyl group. The trypanocidal activity of these steroids was evaluated in vitro against the epimastigote stage of two T. cruzi strains, Ninoa and TH, and their cytotoxicity over J774.2 macrophage cell line was assayed. Compounds 3a, 3b, and 4a shown higher trypanocidal activity than Bz and Nfx against epimastigotes of Ninoa strain, whereas DHEA (1a) and its nitrate derivative 1b showed higher activity than the reference drugs against the TH strain epimastigote. None of the compounds showed activity in the ex vivo assays against the blood trypomastigote of both strains. Interestingly, the selectivity index of Androst-5-en-3ß,17ß-diol 4a was almost twice the value of Nfx and 50 times more than Bz, against Ninoa and TH strains, respectively. Therefore, compound 4a could represent a valuable starting point toward the optimization of steroid derivatives as trypanocidal agents.

Novel thiosemicarbazones induce high toxicity in estrogen-receptor-positive breast cancer cells (MCF7) and exacerbate cisplatin effectiveness in triple-negative breast (MDA-MB231) and lung adenocarcinoma (A549) cells.

Cis-diamminedichloroplatinum(II) (CDDP), known as cisplatin, has been extensively used against breast cancer, which is the most frequent cancer among women, and lung cancer, the leading cancer that causes death worldwide. Novel compounds such as thiazole derivatives have exhibited antiproliferative activity, suggesting they could be useful against cancer treatment. Herein, we synthesized two novel thiosemicarbazones and an aldehyde to combine with CDDP to enhance efficacy against ER-positive breast MCF7 cancer cells, triple-negative/basal-B mammary carcinoma cells (MDA-MB231) and lung adenocarcinoma (A549) human cells. We synthesized 2,3,5,6-tetrafluoro-4-(2-mercaptoetanothiolyl)benzaldehyde (ALD), 5-[(2,3,5,6-tetrafluoro-4-(trifluoromethyl)phenyl)thio]-2-furaldehyde thiosemicarbazone (TSC1) and 5-[(4-(trifluoromethyl)phenyl)thio]-2-furaldehyde thiosemicarbazone (TSC2) and used them alone or in combination with subtoxic CDDP concentrations to evaluate cytotoxicity, cytoskeleton integrity and mitochondrial function. We found that none of the synthesized compounds improved CDDP activity against MCF7 cell cultures; however, TSC2 was effective in enhancing the cytotoxicity of CDDP against MDA-MB231 and A549 cancer cell cultures. We demonstrated that the cytotoxic effect is related to the TSC2 capacity to induce disruption in the cytoskeleton network and to decrease mitochondrial function.

Repositioned Drugs for Chagas Disease Unveiled via Structure-Based Drug Repositioning.

Chagas disease, caused by the parasite Trypanosoma cruzi, affects millions of people in South America. The current treatments are limited, have severe side effects, and are only partially effective. Drug repositioning, defined as finding new indications for already approved drugs, has the potential to provide new therapeutic options for Chagas. In this work, we conducted a structure-based drug repositioning approach with over 130,000 3D protein structures to identify drugs that bind therapeutic Chagas targets and thus represent potential new Chagas treatments. The screening yielded over 500 molecules as hits, out of which 38 drugs were prioritized following a rigorous filtering process. About half of the latter were already known to have trypanocidal activity, while the others are novel to Chagas disease. Three of the new drug candidates-ciprofloxacin, naproxen, and folic acid-showed a growth inhibitory activity in the micromolar range when tested ex vivo on T. cruzi trypomastigotes, validating the prediction. We show that our drug repositioning approach is able to pinpoint relevant drug candidates at a fraction of the time and cost of a conventional screening. Furthermore, our results demonstrate the power and potential of structure-based drug repositioning in the context of neglected tropical diseases where the pharmaceutical industry has little financial interest in the development of new drugs.

Application of neural networks for the detection of oral cancer: A systematic review.

One potential application of neural networks (NNs) is the early-stage detection of oral cancer. This systematic review aimed to determine the level of evidence on the sensitivity and specificity of NNs for the detection of oral cancer, following the Preferred Reporting Items for Systematic Reviews and MetaAnalyses (PRISMA) and Cochrane guidelines. Literature sources included PubMed, ClinicalTrials, Scopus, Google Scholar, and Web of Science. In addition, the Quality Assessment of Diagnostic Accuracy Studies 2 (QUADAS-2) tool was used to assess the risk of bias and the quality of the studies. Only 9 studies fully met the eligibility criteria. In most studies, NNs showed accuracy greater than 85%, though 100% of the studies presented a high risk of bias, and 33% showed high applicability concerns. Nonetheless, the included studies demonstrated that NNs were useful in the detection of oral cancer. However, studies of higher quality, with an adequate methodology, a low risk of bias and no applicability concerns are required so that more robust conclusions could be reached.

Structure-based Virtual Screening from Natural Products as Inhibitors of SARS-CoV-2 Spike Protein and ACE2 Receptor Binding and their Biological Evaluation In vitro.

BACKGROUND: In the last years, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused more than 760 million infections and 6.9 million deaths. Currently, remains a public health problem with limited pharmacological treatments. Among the virus drug targets, the SARS-CoV-2 spike protein attracts the development of new anti-SARS-CoV-2 agents. OBJECTIVE: The aim of this work was to identify new compounds derived from natural products (BIOFACQUIM and Selleckchem databases) as potential inhibitors of the spike receptor binding domain (RBD)-ACE2 binding complex. METHODS: Molecular docking, molecular dynamics simulations, and ADME-Tox analysis were performed to screen and select the potential inhibitors. ELISA-based enzyme assay was done to confirm our predictive model. RESULTS: Twenty compounds were identified as potential binders of RBD of the spike protein. In vitro assay showed compound B-8 caused 48% inhibition at 50 µM, and their binding pattern exhibited interactions via hydrogen bonds with the key amino acid residues present on the RBD. CONCLUSION: Compound B-8 can be used as a scaffold to develop new and more efficient antiviral drugs.

In vitro and In vivo Biological Activity of Two Aryloxy-naphthoquinones in Mice Infected with Trypanosoma cruzi Strains.

BACKGROUND: Chagas disease, a condition caused by Trypanosoma cruzi, is an endemic disease in Latin American countries that affects approximately eight million people worldwide. It is a continuing public health problem. As nifurtimox and benznidazole are the two pharmacological treatments currently used to treat it, the present research proposes new therapeutic alternatives. Previous studies conducted on naphthoquinone derivatives have found interesting trypanocidal effects on epimastigotes, with the molecules 2-phenoxy-1,4-naphthoquinone (IC50= 50 nM and SI < 250) and 2-(3-nitrophenoxy)-naphthalene-1,4-dione (IC50= 20 nM and SI=625) presenting the best biological activity.. METHODS: The present study evaluated the efficacy of in vitro, ex vivo and in vivo models of two aryloxyquinones, 2-phenoxy-1,4-naphthoquinone (1) and 2-(3-nitrophenoxy)-naphthalene-1,4- dione (2), against two Mexican T. cruzi strains in both their epimastigote and blood Trypomastigote stage. Both compounds were evaluated against T. cruzi using a mouse model (CD1) infected with Mexican isolates of T. cruzi, nifurtimox and benznidazole used as control drugs. Finally, the cytotoxicity of the two compounds against the J774.2 mouse macrophage cell line was also determined. RESULTS: The in vitro and in vivo results obtained indicated that both quinones were more active than the reference drugs. Compound 1 presents in vivo activity, showing up to 40% parasite reduction after 8 h of administration, a finding which is 1.25 times more effective than the results obtained using nifurtimox. CONCLUSION: These are encouraging results for proposing new naphthoquinone derivatives with potential anti-T. cruzi activity.

Ligand- and Structure-Based Virtual Screening Identifies New Inhibitors of the Interaction of the SARS-CoV-2 Spike Protein with the ACE2 Host Receptor.

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a fast-spreading viral pathogen and poses a serious threat to human health. New SARS-CoV-2 variants have been arising worldwide; therefore, is necessary to explore more therapeutic options. The interaction of the viral spike (S) protein with the angiotensin-converting enzyme 2 (ACE2) host receptor is an attractive drug target to prevent the infection via the inhibition of virus cell entry. In this study, Ligand- and Structure-Based Virtual Screening (LBVS and SBVS) was performed to propose potential inhibitors capable of blocking the S receptor-binding domain (RBD) and ACE2 interaction. The best five lead compounds were confirmed as inhibitors through ELISA-based enzyme assays. The docking studies and molecular dynamic (MD) simulations of the selected compounds maintained the molecular interaction and stability (RMSD fluctuations less than 5 Å) with key residues of the S protein. The compounds DRI-1, DRI-2, DRI-3, DRI-4, and DRI-5 efficiently block the interaction between the SARS-CoV-2 spike protein and receptor ACE2 (from 69.90 to 99.65% of inhibition) at 50 µM. The most potent inhibitors were DRI-2 (IC50 = 8.8 µM) and DRI-3 (IC50 = 2.1 µM) and have an acceptable profile of cytotoxicity (CC50 > 90 µM). Therefore, these compounds could be good candidates for further SARS-CoV-2 preclinical experiments.

The modulation of the hexosamine biosynthetic pathway impacts the localization of CD36 in macrophages.

CD36 is a type 2 cell surface scavenger receptor expressed in various tissues. In macrophages, CD36 recognizes oxidized low-density lipoprotein (ox-LDL), which promotes the formation of foam cells, the first step toward an atherosclerotic arterial lesion. CD36 possesses a variety of posttranslational modifications, among them N-glycosylation and O-GlcNAc modification. Some of the roles of these modifications on CD36 are known, such as N-linked glycosylation, which provides proper folding and trafficking to the plasma membrane in the human embryonic kidney. This study aimed to determine whether variations in the availability of UDP-GlcNAc could impact Rab-5-mediated endocytic trafficking and, therefore, the cellular localization of CD36. These preliminary results suggest that the availability of the substrate UDP-GlcNAc, modulated in response to treatment with Thiamet G (TMG), OSMI-1 (O-GlcNAcylation enzymes modulators) or Azaserine (HBP modulator), influences the localization of CD36 in J774 macrophages, and the endocytic trafficking as evidenced by the regulatory protein Rab-5, between the plasma membrane and the cytoplasm.

Design, Synthesis, and In Vitro and In Silico Evaluation of 1,3,4-Oxadiazoles as Anti-Trypanosoma cruzi and Anti-Leishmania mexicana Agents.

A series of novel 4-acetyl-1,3,4-oxadiazole derivatives was designed and synthesized for their biological evaluation in vitro against Trypanosoma cruzi (T. cruzi) and Leishmania mexicana (L. mexicana). Additionally, all compounds were evaluated by molecular docking on the cruzain of T. cruzi (TcCz) and the cysteine protease B (CPB) of L. mexicana (LmCPB) to know their potential mechanism of binding. Compound OX-12 had better trypanocidal activity against NINOA (IC50=10.5 µM) and A1 (IC50=21.7 µM) T. cruzi strains that reference drug benznidazole (IC50=30.3 µM and 39.8 µM, respectively). Compound OX-2 had the best biological activity against L. mexicana in M379 (IC50=11.9 µM) and FCQEPS (IC50=34.0 µM) strains that the reference drug glucantime (IC50>120 µM). All the compounds showed important interactions with residues on the active site of TcCz (Gly66, Trp26, Leu67, and Ala138) and LmCPB (Gly67, Asn62, Leu68, and Ala140). Finally, the molecular dynamics simulations of the compound OX-12 shown moderate stability from 40-115 ns with an RMSD value of 6.5 Å. Meanwhile, compound OX-2 showed a minor stability in complex with CPB from 25-200 ns of simulation (RMSD<9 Å). These results encourage to develop more potent and efficient trypanocidal and leishmanicidal agents using the 1,3,4-oxadiazole scaffold.

Expanding the chemical space of ester of quinoxaline-7-carboxylate 1,4-di-N-oxide derivatives as potential antitubercular agents.

Tuberculosis is a worldwide health problem that warrants attention given that the current treatment options require a long-term chemotherapeutic period and have reported the development of Mycobacterium tuberculosis (M. tuberculosis) multidrug resistant strains. In this study, n-butyl and isobutyl quinoxaline-7-carboxylate 1,4-di-N-oxide were evaluated against replicating and non-replicating H37Rv M. tuberculosis strains. The results showed that seventeen of the twenty-eight derivatives have minimum inhibitory concentration (MIC) values lower than isoniazid (2.92 µM). The most active antimycobacterial agents were T-148, T-149, T-163, and T-164, which have the lowest MIC values (0.53, 0.57, 0.53, and 0.55 µM respectively). These results confirm the potential of quinoxaline-1,4-di-N-oxide against M. tuberculosis to develop and obtain new and more safety antituberculosis drugs.

Ecoepidemiology of Chagas Disease in a Biological Corridor in Southeastern Mexico: A Promising Approach to Understand the Risk of Chagas Disease.

Ecoepidemiology is an emerging field that attempts to explain how biotic, environmental, and even social factors influence the dynamics of infectious diseases. Particularly in vector-borne diseases, the study under this approach offers us an overview of the pathogens, vectors, and hosts that coexist in a given region and their ecological determinants. As a result of this, risk predictions can be established in a changing environment and how it may impact human populations. This paper is aimed at evaluating some ecoepidemiological characteristics of Chagas disease in a natural reserve in southeastern Mexico that borders human settlements. We carry out a cross-sectional study in 2022 where we search insects manually and with light traps. We set traps for small mammals and bats and conducted interviews with the inhabitants living around the study site. We identified the presence of Triatoma dimidiata and T. huehuetenanguensis species with a percentage of TcI T. cruzi infection of 68.4% (95% CI: 66.9-69.9). Temperature and humidity were not determining factors for the probability of insect capture. Of the 108 wild mammals (Chiroptera, Rodentia, and Didelphimorphia), none was infected with T. cruzi. Knowledge about Chagas disease in nearby inhabitants is poor, and some characteristics were found on the periphery of dwellings that could offer a refuge for insect vectors. With this information, surveillance strategies can be generated in the study area that reduce the risk of transmission of T. cruzi parasite to humans, and it is expected to motivate the use of this field in future research.

What Do You Need to Know before Studying Chagas Disease? A Beginner's Guide.

Chagas disease is one of the most important tropical infections in the world and mainly affects poor people. The causative agent is the hemoflagellate protozoan Trypanosoma cruzi, which circulates among insect vectors and mammals throughout the Americas. A large body of research on Chagas disease has shown the complexity of this zoonosis, and controlling it remains a challenge for public health systems. Although knowledge of Chagas disease has advanced greatly, there are still many gaps, and it is necessary to continue generating basic and applied research to create more effective control strategies. The aim of this review is to provide up-to-date information on the components of Chagas disease and highlight current trends in research. We hope that this review will be a starting point for beginners and facilitate the search for more specific information.

Advances in Protozoan Epigenetic Targets and Their Inhibitors for the Development of New Potential Drugs.

Protozoan parasite diseases cause significant mortality and morbidity worldwide. Factors such as climate change, extreme poverty, migration, and a lack of life opportunities lead to the propagation of diseases classified as tropical or non-endemic. Although there are several drugs to combat parasitic diseases, strains resistant to routinely used drugs have been reported. In addition, many first-line drugs have adverse effects ranging from mild to severe, including potential carcinogenic effects. Therefore, new lead compounds are needed to combat these parasites. Although little has been studied regarding the epigenetic mechanisms in lower eukaryotes, it is believed that epigenetics plays an essential role in vital aspects of the organism, from controlling the life cycle to the expression of genes involved in pathogenicity. Therefore, using epigenetic targets to combat these parasites is foreseen as an area with great potential for development. This review summarizes the main known epigenetic mechanisms and their potential as therapeutics for a group of medically important protozoal parasites. Different epigenetic mechanisms are discussed, highlighting those that can be used for drug repositioning, such as histone post-translational modifications (HPTMs). Exclusive parasite targets are also emphasized, including the base J and DNA 6 mA. These two categories have the greatest potential for developing drugs to treat or eradicate these diseases.

Triose Phosphate Isomerase Structure-Based Virtual Screening and In Vitro Biological Activity of Natural Products as Leishmania mexicana Inhibitors.

Cutaneous leishmaniasis (CL) is a public health problem affecting more than 98 countries worldwide. No vaccine is available to prevent the disease, and available medical treatments cause serious side effects. Additionally, treatment failure and parasite resistance have made the development of new drugs against CL necessary. In this work, a virtual screening of natural products from the BIOFACQUIM and Selleckchem databases was performed using the method of molecular docking at the triosephosphate isomerase (TIM) enzyme interface of Leishmania mexicana (L. mexicana). Finally, the in vitro leishmanicidal activity of selected compounds against two strains of L. mexicana, their cytotoxicity, and selectivity index were determined. The top ten compounds were obtained based on the docking results. Four were selected for further in silico analysis. The ADME-Tox analysis of the selected compounds predicted favorable physicochemical and toxicological properties. Among these four compounds, S-8 (IC50 = 55 µM) demonstrated a two-fold higher activity against the promastigote of both L. mexicana strains than the reference drug glucantime (IC50 = 133 µM). This finding encourages the screening of natural products as new anti-leishmania agents.

Identification of Discrete Typing Units of Trypanosoma cruzi Isolated from Domestic Environments in Southeastern Mexico.

Background: The Trypanosoma cruzi parasite is the causal agent of Chagas disease, recognized by the World Health Organization as a neglected tropical disease. Currently there are seven discrete typing units (DTUs) of T. cruzi distributed in America, but there are still gaps about its distribution in some endemic regions. Materials and Methods: Seventeen units isolated from Chiapas and Oaxaca in Mexico were identified by amplification of the C-5 sterol desaturase gene. Results: Three DTUs of T. cruzi, TcI (6), TcII (10), and TcIV (1) were detected by comparing polymorphic sites in specific regions. Conclusions: New DTUs are reported for both states, where TcII was the most common DTU. The genetic characterization of the isolates can help to understand the epidemiology of Chagas disease.

Improving Adherence to Physical Therapy in the Development of Serious Games: Conceptual Framework Design Study.

BACKGROUND: Insufficient levels of treatment adherence can have adverse effects on the outcomes of physical rehabilitation. To address this issue, alternative approaches to traditional therapies, such as serious games, have been designed to enhance adherence. Nevertheless, there remain gaps in the development of serious games concerning the effective implementation of motivation, engagement, and the enhancement of treatment adherence. OBJECTIVE: This study aims to design a conceptual framework for the development of serious games that incorporate essential adherence factors to enhance patient compliance with physical rehabilitation programs. METHODS: We formulated a conceptual framework using iterative techniques inspired by a conceptual framework analysis. Initially, we conducted a comprehensive literature review, concentrating on the critical adherence factors in physical rehabilitation. Subsequently, we identified, categorized, integrated, and synthesized the concepts derived from the literature review to construct the conceptual framework. RESULTS: The framework resembles a road map, comprising 3 distinct phases. In the initial phase, the patient's characteristics are identified through an initial exploration. The second phase involves the development of a serious game, with a focus on enhancing treatment adherence by integrating the key adherence factors identified. The third phase revolves around the evaluation of the serious game. These phases are underpinned by 2 overarching themes, namely, a user-centered design and the GameFlow model. CONCLUSIONS: The conceptual framework offers a detailed, step-by-step guide for creating serious games that incorporate essential adherence factors, thereby contributing to improved adherence in the physical rehabilitation process. To establish its validity, further evaluations of this framework across various physical rehabilitation programs and user groups are necessary.

Parasitemia and Differential Tissue Tropism in Mice Infected with Trypanosoma cruzi Isolates Obtained from Meccus phyllosoma in the State of Oaxaca, Mexico.

Trypanosoma cruzi is a parasite transmitted by the feces of triatomines. Many triatomine species are found in Mexico, and various T. cruzi variants have been isolated from these species, each showing very different virulence and cell tropism. The isolates were obtained from Meccus phyllosoma specimens in three localities in the state of Oaxaca, Mexico: Tehuantitla, Vixhana, and Guichivere. The virulence of each isolate was assessed by quantifying parasitemia, survival, and histopathologic findings. The lineage of each isolate was identified using the mini-exon gene. The expression of the tssa gene during infection was detected in the heart, esophagus, gastrocnemius, and brain. Our results show that the maximum post-infection parasitemia was higher for the Tehuantitla isolate. On genotyping, all isolates were identified as T. cruzi I. The amastigotes in the heart and gastrocnemius were verified for all isolates, but in the brain only for Tehuantitla and Vixhana. The tssa expression allowed us to detect T. cruzi isolates, for Tehuantitla, predominantly in the heart. For Vixhana, a higher tssa expression was detected in gastrocnemius, and for Guichivere, it was higher in the esophagus. Results show that virulence, tropism, and tssa expression can vary, even when the isolates are derived from the same vector species, in the same region, and at similar altitudes.