Leishmaniasis represents a serious health problem worldwide and drug resistance is a growing concern. Leishmania parasites use unusual mechanisms to control their gene expression. In contrast to many other species, they do not have transcriptional regulation. The lack of transcriptional control is mainly compensated by post-transcriptional mechanisms, including tight translational control and regulation of mRNA stability/translatability by RNA-binding proteins. Modulation of translation plays a major role in parasite survival and adaptation to dramatically different environments during change of host; however, our knowledge of fine molecular mechanisms of translation in Leishmania remains limited. Here, we review the current progress in our understanding of how changes in the translational machinery promote parasite differentiation during transmission from a sand fly to a mammalian host, and discuss how translational reprogramming can contribute to the development of drug resistance.

Leishmania is a unicellular protozoan that has a limited transcriptional control and mostly uses post-transcriptional regulation of gene expression, although the molecular mechanisms of the process are still poorly understood. Treatments of leishmaniasis, pathologies associated with Leishmania infections, are limited due to drug resistance. Here, we report dramatic differences in mRNA translation in antimony drug-resistant and sensitive strains at the full translatome level. The major differences (2431 differentially translated transcripts) were demonstrated in the absence of the drug pressure supporting that complex preemptive adaptations are needed to efficiently compensate for the loss of biological fitness once they are exposed to the antimony. In contrast, drug-resistant parasites exposed to antimony activated a highly selective translation of only 156 transcripts. This selective mRNA translation is associated with surface protein rearrangement, optimized energy metabolism, amastins upregulation, and improved antioxidant response. We propose a novel model that establishes translational control as a major driver of antimony-resistant phenotypes in Leishmania.

Leishmania parasites cause leishmaniasis, one of the most epidemiologically important neglected tropical diseases. Leishmania exhibits a high ability of developing drug resistance, and drug resistance is one of the main threats to public health, as it is associated with increased incidence, mortality, and healthcare costs. The antimonial drug is the main historically implemented drug for leishmaniasis. Nevertheless, even though antimony resistance has been widely documented, the mechanisms involved are not completely understood. In this study, we aimed to identify potential metabolite biomarkers of antimony resistance that could improve leishmaniasis treatment. Here, using L. tropica promastigotes as the biological model, we showed that the level of response to antimony can be potentially predicted using ¹H-NMR-based metabolomic profiling. Antimony-resistant parasites exhibited differences in metabolite composition at the intracellular and extracellular levels, suggesting that a metabolic remodeling is required to combat the drug. Simple and time-saving exometabolomic analysis can be efficiently used for the differentiation of sensitive and resistant parasites. Our findings suggest that changes in metabolite composition are associated with an optimized response to the osmotic/oxidative stress and a rearrangement of carbon-energy metabolism. The activation of energy metabolism can be linked to the high energy requirement during the antioxidant stress response. We also found that metabolites such as proline and lactate change linearly with the level of resistance to antimony, showing a close relationship with the parasite’s efficiency of drug resistance. A list of potential metabolite biomarkers is described and discussed.

Leishmania parasites efficiently develop resistance against several types of drugs including antimonials, the primary antileishmanial drug historically implemented. The resistance to antimonials is considered to be a major risk factor for effective leishmaniasis treatment. To detect biomarkers/biopatterns for the differentiation of antimony-resistant Leishmania strains, we employed untargeted global mass spectrometry to identify intracellular lipids present in antimony sensitive and resistant parasites before and after antimony exposure. The lipidomic profiles effectively differentiated the sensitive and resistant phenotypes growing with and without antimony pressure. Resistant phenotypes were characterized by significant downregulation of phosphatidylcholines, sphingolipid decrease, and lysophosphatidylcholine increase, while sensitive phenotypes were characterized by the upregulation of triglycerides with long-chain fatty acids and a tendency toward the phosphatidylethanolamine decrease. Our findings suggest that the changes in lipid composition in antimony-resistant parasites contribute to the physiological response conducted to combat the oxidative stress unbalance caused by the drug. We have identified several lipids as potential biomarkers associated with the drug resistance.

Diagnostic delays prevent most Chagas disease patients from receiving timely therapy during the acute phase when treatment is effective. Quantitative PCR (qPCR)-based diagnostic methods provide high sensitivity during this phase but require specialized equipment and complex protocols. More simple and cost-effective tools are urgently needed to expand early Chagas disease diagnosis in low-income endemic regions. Here, we present a loop-mediated isothermal amplification (LAMP) that targets a highly conserved region in the HSP70 gene of Trypanosoma cruzi , the causative agent of Chagas disease. This assay demonstrates species-specific amplification across multiple parasite genetic lineages while maintaining stability after 1 hour of incubation and at least 8 months of storage at −20°C. Moreover, the assay is at least 12 times less expensive than the TaqMan qPCR that is currently routinely used for acute Chagas diagnostics and is compatible with a low-cost CTAB-based DNA extraction method. Population-based validation in 100 infants born to Chagas-positive mothers in Santa Cruz, Bolivia, yielded a specificity of 100% and sensitivity exceeding 77% when compared to a TaqMan qPCR that targets satellite DNA. This cost-effective DNA extraction method and LAMP assay demonstrate potential for diagnosing Chagas disease in resource-limited endemic regions, particularly where qPCR is unavailable. IMPORTANCE: Chagas disease, caused by the parasite Trypanosoma cruzi , is a life-threatening illness that disproportionately affects resource-limited communities. Congenital Chagas disease, if diagnosed early, presents a unique opportunity for intervention, as treatment in newborns is highly effective with minimal side effects. However, early diagnosis is hindered by the high cost and limited availability of current molecular diagnostic methods in endemic regions. Our study introduces a simple, low-cost, and highly specific LAMP assay targeting the HSP70 gene of T. cruzi . This assay is user-friendly, stable under varying storage and incubation conditions, and designed for accessibility in underserved areas. By providing a detailed, open-access protocol and primers, we aim to facilitate the widespread adoption of this diagnostic assay, enabling earlier detection and treatment. This assay lays the groundwork for a new approach to Chagas disease management, potentially reducing the spread of Chagas disease and improving public health outcomes in vulnerable populations globally.

There is limited information about the innate immunity of triatomines against Trypanosoma rangeli, an infectious, non-pathogenic human parasite, and T. cruzi, the causative agent of Chagas’ disease. This study aimed at addressing this gap by studying the in vitro trypanolytic hemolymph activity from insects not infected by T. rangeli or T. cruzi. Eight triatomine species were examined including Rhodnius prolixus, R. robustus, R. colombiensis, R. pallescens, R. pictipes, Triatoma dimidiata, T. maculata, and Panstrongylus geniculatus. The hemolymph of R. prolixus and R. robustus laboratory colonies demonstrated strong trypanolytic activity during the first 14 hours of parasite incubation with 81% lysis for the T. rangeli C genotype, 90% for the E genotype, 95% for T. cruzi discrete taxonomic unit (DTU) TcII, 94% for TcV, 96% for TcVI, 94% for Tcbat, and 90% for Tcmarinkellei. No lysis activity was detected 14 or 24 hours after parasite incubation with the hemolymph of insects from R. pictipes R. pallescens, P. geniculatus, and T. maculata colonies. Identical results (absence of lysis) were observed using hemolymph from R. colombiensis and T. dimidiata laboratory colonies and specimens captured in sylvatic environments. The hemolymph lytic activity against recently obtained cultures (95% decrease in live parasite count) and long-term T. cruzi TcII ones (96% decrease) was similar. Lytic activity was similar in hemolymph from R. prolixus nymphs, males and females and insects fed on chicken or mouse blood. This is the first comparative study of the trypanolytic activity of hemolymph from different triatomine species against T. rangeli and T. cruzi genotypes. Although the chemical structures of such lysis factors and the mechanisms determining their expression have not been fully determined, their identification furthers our understanding of triatomines’ innate immunity and their role in Trypanosoma transmission.