Biology subjects

de Miguel, N.

Publications and source records attributed to de Miguel, N..

7 recordsLinked to original sources

bioRxiv2026

Ultrastructural Analysis of Extracellular Vesicles and Surface Protrusions from the Parasite Tritrichomonas foetus

Tritrichomonas foetus is an extracellular flagellated protist responsible for bovine and feline tritrichomonosis, yet several aspects of its cell biology and pathogenicity remain poorly understood. Here, we evaluated the presence of extracellular vesicles (EVs) and membrane protrusions in T. foetus using complementary electron microscopy and immunolabeling approaches. EV-like structures of different sizes were found protruding from the parasite cell body and anterior flagella, and ultrathin sections showed continuity between the membranes of these protrusions and the plasma or flagellar membrane. Multivesicular bodies containing intraluminal vesicles were also observed, and an EV-enriched fraction isolated from conditioned medium contained vesicles with a broad size distribution, consistent with heterogeneous EV populations. Immunofluorescence further showed tetraspanin-1 in intracellular vesicles, flagella, and vesicular structures protruding from the parasite surface. Exposure to bovine preputial epithelial cells resulted in a ninefold increase in the percentage of parasites displaying flagellar cytoneme-like structures, suggesting a possible role for these projections in host-parasite interactions. VPS32, a component of the ESCRT-III machinery, was found in intraluminal vesicles, vesicles protruding from the cell body and flagella, isolated EVs, and flagellar membrane protrusions. In parasites overexpressing VPS32, the number of vesicles per flagellum increased, and the percentage of cells with flagellar cytoneme-like projections was fivefold higher than in control parasites. Pearling-like protrusions containing VPS32 were also observed. Together, these findings provide ultrastructural evidence for heterogeneous EV populations associated with the cell body and flagella of T. foetus and suggest that VPS32 could contribute to flagellar EV and membrane protrusion formation.

cell biology↗

bioRxiv2026

Chemotaxis and selective interactions of Trichomonas vaginalis with the vaginal bacteria

Trichomonas vaginalis is an extracellular parasite that inhabits the human genital tract, yet little is known about how it senses and responds to the complex vaginal microbial ecosystem. Here, we show that T. vaginalis exhibits chemotactic behavior on semisolid surfaces, forming multicellular assemblies that coordinate collective migration. Parasite colonies display both positive and negative chemotactic responses, indicating the ability to detect and react to diffusible signals. Different parasite strains display marked mutual avoidance between neighboring colonies, highlighting specific recognition mechanisms. Furthermore, we show that T. vaginalis is strongly attracted to acidic environments, revealing a niche-adapted pH taxis. Given that vaginal bacteria critically shape local pH, we examined parasite responses to representative members of the vaginal microbiota. T. vaginalis exhibited preferential chemotactic migration toward Lactobacillus gasseri, a hallmark species of eubiotic community state types (CSTs), over Gardnerella vaginalis, which is associated with dysbiotic CST-IV communities, while showing no detectable attraction to Escherichia coli. This selective migration correlated with a robust chemotactic response to lactic acid, a major metabolite produced by lactobacilli. Additionally, when the parasite is co-cultured with the equal number of L. gasseri and G. vaginalis, T. vaginalis exhibits a clear preferential binding to L. gasseri, as demonstrated by flow cytometry and fluorescent microscopy. We show that co-culture of T. vaginalis with either L. gasseri or G. vaginalis results in enhanced parasite growth only in the presence of L. gasseri. Collectively, these findings reveal pH taxis; bacteria-directed migration and preferential association with Lactobacillus as previously underappreciated behavioral traits of T. vaginalis. Such behaviors may destabilize protective microbial communities and drive the transition toward a CST-IV-type dysbiotic state which is frequently associated with trichomoniasis.

microbiology↗

bioRxiv2025

Targeting histone acetylation to overcome drug resistance in the parasite Trichomonas vaginalis

Resistance to antimicrobial drugs is a major global health challenge, yet the regulatory mechanisms underlying resistance phenotypes remain incompletely understood. In particular, whether epigenetic processes contribute to antimicrobial resistance in microbial eukaryotes is largely unexplored. Here, we show that epigenetic regulation plays a central role in modulating metronidazole resistance in Trichomonas vaginalis, the causative agent of the most common non-viral sexually transmitted infection. Pharmacological inhibition of histone deacetylases restores drug sensitivity in a highly resistant clinical isolate, markedly reducing the minimum inhibitory concentration. Integrative transcriptomic reveal widespread transcriptional reprogramming associated with activation of pathways involved in redox metabolism, transcriptional regulation, and stress responses. Chromatin profiling demonstrates that increased H3K9 acetylation is strongly associated with transcriptional activation. Upon TSA treatment, increased H3K9 acetylation is observed at promoters and gene bodies of genes that may be associated with MTZ resistance. By combining epigenetic perturbation with comparative transcriptomics across multiple strains, we identify a set of genes that are epigenetically silenced in resistant parasites that are constitutively expressed in sensitive backgrounds. Functional re-expression of two candidates, thioredoxin reductase and a Myb-like transcription factor (Myb6), significantly reduces resistance, establishing a direct link between epigenetic regulation and drug sensitivity. Together, our findings uncover a reversible, chromatin-based mechanism underlying antimicrobial resistance in a protozoan parasite and highlight epigenetic reprogramming as a potential therapeutic strategy. More broadly, this work suggests that epigenetic regulation may represent a general and previously underappreciated layer of control over drug resistance in microbial eukaryotes.

microbiology↗

bioRxiv2023

Extracellular vesicle-localized miR-203 mediates neural crest-placode communication required for trigeminal ganglia formation

While interactions between neural crest and placode cells are critical for the proper formation of the trigeminal ganglion, the mechanisms underlying this process remain largely uncharacterized. Here, we show that the microRNA-(miR)203, whose epigenetic repression is required for neural crest migration, is reactivated in coalescing and condensing trigeminal ganglion cells. Overexpression of miR-203 induces ectopic coalescence of neural crest cells and increases ganglion size. Reciprocally, loss of miR-203 function in placode, but not neural crest, cells perturbs trigeminal ganglion condensation. Demonstrating intercellular communication, overexpression of miR-203 in the neural crest in vitro or in vivo represses a miR-responsive sensor in placode cells. Moreover, neural crest-secreted extracellular vesicles (EVs), visualized using pHluorin-CD63 vector, become incorporated into the cytoplasm of placode cells. Finally, RT-PCR analysis shows that small EVs isolated from condensing trigeminal ganglia are selectively loaded with miR-203. Together, our findings reveal a critical role in vivo for neural crest-placode communication mediated by sEVs and their selective microRNA cargo for proper trigeminal ganglion formation. SIGNIFICANCE STRATEMENTCellular communication during early development plays a critical role. In this study, we demonstrate a unique role for a microRNA in cell-cell communication between the neural crest (NC) and placode cells (PC) during trigeminal ganglia (TG) formation. By utilizing loss and gain of function experiments in vivo, we demonstrate a requirement for miR-203 during cellular condensation to form the TG. We revealed that NC produces extracellular vesicles, selectively carrying miR-203, which is then taken up by the PC and regulates a sensor vector exclusively expressed in the placode. Taken together, our findings reveal a critical role in TG condensation for miR-203, produced by post-migratory NC and taken up by PC via extracellular vesicles.

developmental biology↗

bioRxiv2023

Role of cytoneme-like structures and extracellular vesicles in Trichomonas vaginalis parasite: parasite communication

Trichomonas vaginalis, the etiologic agent of the most common non-viral sexually transmitted infection worldwide, colonizes the human urogenital tract where it remains extracellular and adheres to epithelial cells. With an estimated prevalence of 276 million new cases annually, mixed infections with different parasite strains are expected. Although it is considered as obvious that parasites interact with their host to enhance their own survival and transmission, evidence of mixed infection call into question the extent to which unicellular parasites communicate with each other. Here, we demonstrated that different T. vaginalis strains are able to communicate through the formation of cytoneme-like membranous cell connections. We showed that T. vaginalis adherent strains form abundant membrane protrusions and cytonemes formation of an adherent parasite strain (CDC1132) is affected in the presence of a different strain (G3 or B7RC2). Using a cell culture inserts assays, we demonstrated that the effect in cytoneme formation is contact independent and that extracellular vesicles (EVs) are responsible, at least in part, of the communication among strains. In this sense, we found that EVs isolated from G3, B7RC2 and CDC1132 strains contain a highly distinct repertoire of proteins, some of them involved in signaling and communication, among other functions. Finally, we showed that parasite adherence to host cells is affected by this communication between strains as binding of adherent T. vaginalis CDC1132 strain to prostate cells is significantly higher in the presence of G3 or B7RC2 strains. Demonstrating that interaction of isolates with distinct phenotypic characteristics may have significant clinical repercussions, we also observed that a poorly adherent parasite strain (G3) adheres more strongly to prostate cells in the presence of an adherent strain. The study of signaling, sensing and cell communication in parasitic organisms will surely enhance our understanding of the basic biological characteristics of parasites that might have important consequences in pathogenesis.

microbiology↗

bioRxiv2021

Ultrastructural and functional analysis of extra-axonemal structures in trichomonads

Trichomonas vaginalis and Tritrichomonas foetus are extracellular flagellated parasites that inhabit humans and other mammals, respectively. In addition to motility, flagella act in a variety of biological processes in different cell types; and extra-axonemal structures (EASs) has been described as fibrillar structures that provide mechanical support and act as metabolic, homeostatic and sensory platforms in many organisms. Here, we identified the presence of EASs forming prominent flagellar swellings in T. vaginalis and T. foetus and we observed that their formation was associated with the parasites adhesion on the host cells, fibronectin, and precationized surfaces; and parasite:parasite interaction. A high number of rosettes, clusters of intramembrane particles that has been proposed as sensorial structures, and microvesicles protruding from the membrane were observed in the EASs. The protein VPS32, a member of the ESCRT-III complex crucial for diverse membrane remodeling events, the pinching off and release of microvesicles, was found in the surface as well as in microvesicles protruding from EASs. Moreover, we demonstrated that overexpression of VPS32 protein induce EAS formation and increase parasite motility in semi-solid medium. These results provide valuable data about the role of the flagellar EASs in the cell-to-cell communication and pathogenesis of these extracellular parasites.

cell biology↗

bioRxiv2019

Exploring the link between adenine DNA methylation and 3D genome organization in the parasite Trichomonas vaginalis

Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogenital tract causing infections that range from asymptomatic to highly inflammatory. Recent works have highlighted the importance of histone modifications in the regulation of transcription and parasite pathogenesis. However, the nature of DNA methylation in the parasite remains unexplored. Using a combination of immunological techniques and UHPLC, we analyzed the abundance of DNA methylation in strains with differential pathogenicity demonstrating that N6-methyladenine (6mA), and not 5-methylcytosine (5mC), is the main DNA methylation mark in T. vaginalis. Genome-wide distribution of 6mA reveals that this mark is enriched at intergenic regions, with a preference for certain super-families of DNA transposable elements. We show that 6mA in T. vaginalis is associated with silencing when present on genes. Interestingly, bioinformatics analysis revealed the presence of transcriptionally active or repressive intervals flanked by 6mA-enriched regions and results from chromatin conformation capture (3C) experiments suggest these 6mA flanked regions are in close spatial proximity. These associations were disrupted when parasites were treated with the demethylation activator ascorbic acid. This finding revealed a new role for 6mA in modulating 3D chromatin structure and gene expression in this divergent member of the Excavata.\n\nSIGNIFICANCE STATEMENTTrichomonas vaginalis causes the most common non-viral sexually transmitted infection yet little is known about the regulation of gene expression in this eukaryotic parasite. We demonstrate that N6-methyladenine (6mA) is the main methylation mark in the T. vaginalis genome. 6mA is widespread in DNA of eubacterial genera, but uncommon in genomes of most eukaryotes. Examination of the genome-wide distribution of 6mA reveals a preference for intergenic regions. Transcriptionally active or repressive intervals are found to be flanked by 6mA-enriched regions and data suggest 6mA flanked regions are in close 3D spatial proximity. Our findings describe for the first time the presence of DNA methylation in T. vaginalis and reveal a new role for 6mA in modulating 3D chromatin architecture.

microbiology↗