A striking molecular portrait of treatment failure in leishmaniasis has emerged from Thailand, where researchers tracked the parasite Leishmania (Mundinia) martiniquensis inside a single patient before therapy and again after clinical relapse following amphotericin B treatment. By comparing these paired clinical isolates at the transcriptomic level, the team uncovered widespread differences in gene expression that extend far beyond the sterol pathways traditionally associated with amphotericin B susceptibility. Alongside these findings, the analysis revealed something unexpected lurking within the parasite: a genetically distinct leishbuvirus, recovered in near-complete form and characterized in detail for the first time in a Southeast Asian isolate of this emerging species. The study, published in Parasites & Vectors, offers a hypothesis-generating framework for understanding why some infections withstand one of the last-line drugs available against visceral leishmaniasis.
L. martiniquensis has attracted growing concern in Southeast Asia as an emerging cause of visceral leishmaniasis, a disease that can be fatal particularly among patients with advanced HIV infection whose immune systems cannot contain the parasite. Amphotericin B remains a mainstay of therapy, yet clinicians in Thailand have documented relapse and treatment failure in patients infected with this species, raising the possibility that some parasite populations carry or acquire reduced susceptibility to the drug. Until now, the molecular machinery underlying such reduced susceptibility in L. martiniquensis has remained largely unexplored, making paired isolates drawn from the same patient at different clinical time points an unusually valuable window into how the parasite changes under drug pressure.
The research team, led by investigators at Chulalongkorn University in Bangkok in collaboration with Hatyai Hospital and King Chulalongkorn Memorial Hospital, obtained one isolate before treatment, designated CULE7.2, and a second, CULE8, after the patient relapsed. They first confirmed the biological difference between the two isolates using in vitro susceptibility assays: CULE8 displayed significantly higher half-maximal and 90 percent inhibitory concentrations of amphotericin B than CULE7.2, with the dose-response curve shifted markedly to the right. Transmission electron microscopy of CULE7.2 promastigotes exposed acutely to the drug revealed the expected cellular damage, including prominent cytoplasmic vacuolization, lipid-like inclusions, mitochondrial swelling, and focal disruption of the nuclear membrane, consistent with amphotericin B’s mechanism of binding membrane sterols and destabilizing cellular membranes.
To explore the transcriptional landscape underlying these phenotypic differences, the researchers profiled both isolates across two developmental stages of the parasite, promastigotes and axenic amastigotes, using RNA sequencing. Importantly, the two isolates were cultured under different conditions: CULE8 was maintained under continuous exposure to a sublethal 0.3 micromolar concentration of amphotericin B, while CULE7.2 was cultured in parallel without the drug. Differential expression analysis, hierarchical clustering, and functional enrichment were performed independently within each stage. This design choice means the observed patterns likely reflect a combination of intrinsic isolate-associated characteristics and the transcriptional response to ongoing drug exposure, an important caveat the authors emphasize in interpreting their results.
In the promastigote stage, the post-relapse isolate CULE8 showed predominant downregulation of genes involved in sterol and lipid metabolism, membrane transport, protein synthesis, motility, and broader metabolic and regulatory processes. Among the affected genes were sterol C24 reductase, an enzyme central to the parasite’s membrane sterol composition and a known determinant of amphotericin B activity, together with upstream enzymes of the mevalonate pathway. At the same time, CULE8 promastigotes displayed increased expression of genes associated with thiol-based redox homeostasis, suggesting a shift toward enhanced management of oxidative stress, a plausible countermeasure to drug-induced membrane and mitochondrial damage.
The axenic amastigote stage, the form that proliferates inside the mammalian host, told a different story. Here, CULE8 showed increased expression of genes linked to lipid metabolism, redox homeostasis, protein synthesis, and proteostasis, alongside broader downregulation of transport genes and genes involved in genome maintenance. Despite the predominantly stage-specific patterns, a small subset of genes showed concordant regulation across both life stages, including genes involved in lipid metabolism and transport. Genes tied to thiol-based redox homeostasis stood out for their elevated expression in both stages, marking this antioxidant capacity as a consistent candidate feature of the relapse-associated parasite and a potential target for functional validation studies.
The virological dimension of the study may prove equally consequential. De novo assembly of RNA-seq reads that failed to map to the parasite genome recovered complete L and S genomic segments of a genetically distinct leishbuvirus from CULE8. This represents, to the authors’ knowledge, the first molecular characterization of a leishbuvirus in a Southeast Asian L. martiniquensis isolate, expanding the known geographic and host range of these viruses. Comparative structural modeling using predicted aligned error matrices supported conservation of the RNA-dependent RNA polymerase core and its Motif C between the CULE8 virus and a reference variant, while negative-stain transmission electron microscopy revealed structures consistent with virus-like particles. Viral RNA abundance was higher in CULE8 than in CULE7.2 across both developmental stages.
The significance of leishbuviruses remains a subject of active debate in the field. Related viruses, including Leishmania RNA virus 1, have been associated in some studies with heightened virulence and treatment failure in other Leishmania species, particularly through modulation of the host inflammatory response, although such associations are not universal and their mechanisms are incompletely understood. The authors are careful to note that the higher leishbuvirus RNA abundance observed in the post-relapse isolate has no established causal relationship with reduced amphotericin B susceptibility or with the patient’s clinical relapse. The finding is best viewed as an observation warranting further investigation in independent isolates rather than as evidence of a viral driver of drug resistance.
Indeed, the authors frame the entire study as hypothesis-generating. Because the paired isolates were examined under different drug-exposure conditions, the transcriptional differences they report cannot be attributed cleanly to either intrinsic isolate biology or adaptive response to amphotericin B, and the findings will require validation in independent isolates under matched experimental conditions. Nevertheless, the breadth of the observed changes, encompassing sterol biosynthesis, lipid handling, membrane transport, redox metabolism, protein homeostasis, and genome maintenance, broadens the molecular landscape associated with reduced amphotericin B susceptibility in this species and identifies multiple candidate pathways for future functional studies, including gene knockout or overexpression experiments that could disentangle correlation from causation.
For a disease that disproportionately strikes immunocompromised patients in resource-limited settings, the stakes of understanding amphotericin B failure are high. Visceral leishmaniasis caused by L. martiniquensis in Thailand is emerging in HIV-coinfected populations, and the therapeutic arsenal is narrow. By pairing rigorous susceptibility testing and ultrastructural imaging with stage-resolved transcriptomics and sensitive viral discovery, this study demonstrates how a single patient’s clinical course can illuminate molecular processes that no laboratory strain could fully recapitulate. The identified candidate pathways, from mevalonate-dependent sterol metabolism to thiol-based antioxidant defenses, now provide concrete starting points for the functional experiments and epidemiological surveillance needed to determine how widespread reduced drug susceptibility might be, and whether hidden viral passengers within these parasites play any role in the clinical outcomes that matter most to patients.
Subject of Research: Transcriptomic analysis of paired Leishmania martiniquensis clinical isolates to identify molecular pathways associated with reduced amphotericin B susceptibility
Article Title: Transcriptomic profiling of paired clinical Leishmania (Mundinia) martiniquensis isolates reveals candidate pathways associated with reduced amphotericin B susceptibility and identifies a novel leishbuvirus variant
Article References: Phadungsaksawasdi, K., Pongpanich, M., Yuanlae, S., Lerona, P. G. E., Sricharoensuk, C., Songumpai, N., Siriyasatien, P., Asawanonda, P., Shotelersuk, V., & Preativatanyou, K. (2026). Transcriptomic profiling of paired clinical Leishmania (Mundinia) martiniquensis isolates reveals candidate pathways associated with reduced amphotericin B susceptibility and identifies a novel leishbuvirus variant. Parasites & Vectors. https://doi.org/10.1186/s13071-026-07628-2
Image Credits: AI Generated
DOI: 10.1186/s13071-026-07628-2
Keywords: Leishmania martiniquensis, Mundinia, amphotericin B, drug susceptibility, transcriptomics, leishbuvirus, visceral leishmaniasis, Thailand, RNA sequencing, parasite virome, sterol metabolism, redox homeostasis
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Juliet Wilcox. (September 12, 2026). Relapse Parasite Genome Study Reveals Drug Resistance Clues and New Leishbuvirus in Thailand. Scienmag. https://scienmag.com/relapse-parasite-genome-study-reveals-drug-resistance-clues-and-new-leishbuvirus-in-thailand/
Juliet Wilcox. “Relapse Parasite Genome Study Reveals Drug Resistance Clues and New Leishbuvirus in Thailand.” Scienmag, 12 September 2026, https://scienmag.com/relapse-parasite-genome-study-reveals-drug-resistance-clues-and-new-leishbuvirus-in-thailand/. Accessed 12 September 2026.
Juliet Wilcox. “Relapse Parasite Genome Study Reveals Drug Resistance Clues and New Leishbuvirus in Thailand.” Scienmag. September 12, 2026. https://scienmag.com/relapse-parasite-genome-study-reveals-drug-resistance-clues-and-new-leishbuvirus-in-thailand/
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Tags: amphotericin BAmphotericin B resistance in leishmaniasisdrug susceptibilitydrug susceptibility in Leishmania martiniquensisemerging Leishmania species in ThailandleishbuvirusLeishbuvirus discovery in Southeast Asialeishbuvirus genome characterizationLeishmania martiniquensisLeishmania parasite drug resistanceLeishmania relapse mechanismsLeishmania transcriptomic analysismolecular basis of leishmaniasis treatment failureMundiniaparasite gene expression differences post-treatmentparasite viromeredox homeostasisRNA sequencingsterol metabolismThailandTranscriptomicsviral co-infection in leishmaniasisvisceral leishmaniasisvisceral leishmaniasis treatment challenges
