Biology subjects

Ferrer, L.

Publications and source records attributed to Ferrer, L..

8 recordsLinked to original sources

bioRxiv2026

Direct quantitative PCR detects genetic biomarkers of antileishmanial drug resistance in clinical samples from dogs with leishmaniosis

BackgroundTreatment response in canine leishmaniosis is driven by the dog host, the Leishmania parasite, and pharmacological factors, with drug resistance increasingly undermining the effectiveness of therapy. A direct quantitative PCR test (LeishGenR) was applied to 104 clinical samples from 95 dogs in the Mediterranean area diagnosed with leishmaniosis in veterinary clinical settings and testing positive for Leishmania infantum by PCR. The assay enabled rapid detection of genetic drug-resistance biomarkers for allopurinol (metk), meglumine antimoniate (mrpa), and miltefosine (LdMT), providing a clinically relevant, timely alternative to culture-based approaches by directly analyzing circulating Leishmania infantum amastigotes. ResultsThe assay (LeishGenR) showed high specificity (100%) and sensitivity (>87.5%) for genetic drug-resistance profile assignment and a strong correlation with whole-genome sequencing for gene copy number assessment (metk: r = 0.878; mrpa: r = 0.943 and LdMT = 0.691). Genetic drug-resistance biomarkers were detected in 24.3% of L. infantum DNA from clinical samples analyzed (20/82; 95% CI 16.3-34.6)), most commonly for allopurinol (13.4%; 95% CI 7.6-22.4), then meglumine antimoniate (9.4%; 95% CI 4.6-18.2), and for miltefosine (5.4%; 95% CI 1.8-14.8). Prevalence was higher in dogs previously treated for leishmaniosis. ConclusionThis study demonstrates the ability to detect genetic biomarkers of drug resistance in L. infantum directly from clinical samples of dogs with leishmaniosis. This method enables rapid, precise detection of genomic biomarkers, circumventing delays associated with culture-based methods and supporting more effective clinical management and surveillance. Among dogs with high parasitemia referred to clinics in Mediterranean regions sampled in this study, the findings reveal a significant prevalence of circulating L. infantum strains carrying genomic drug resistance biomarkers to standard treatments for canine leishmaniosis.

genetics↗

bioRxiv2025

Leishmania tarentolae and Leishmania infantum in geckos from Mallorca Island, Spain

Leishmania tarentolae and Leishmania infantum are two sympatric parasites of significant ecological and epidemiological interest in the Mediterranean basin. This study investigated the prevalence of L. tarentolae and L. infantum in two gecko species (Tarentola mauritanica and Hemidactylus turcicus) present on Mallorca Island, Spain, using duplex quantitative PCR. A total of 59 geckos were sampled across the island, including 53 T. mauritanica and six H. turcicus. Tissue and blood samples were screened for both parasites, and generalized linear models were used to assess organ-specific infection patterns. The results revealed the prevalence of Leishmania infection in adult T. mauritanica, with 10/49 (20.41%) testing positive for L. tarentolae and with 1/49 (2.04%) for L. infantum. Coinfection with both parasites was detected in 3/49 geckos (6.12%). No positives were identified in H. turcicus, probably due to small sample size. Regarding positivity by tissues, coleomic organs were more likely to be positive for L. tarentolae in adult T. mauritanica than blood, with a slighter positivity in the liver, spleen and lung. Interestingly, L. infantum was detected more frequently coinfecting with L. tarentolae than as a single parasite. These prevalence rates and the existence of coinfections highlight potential ecological interactions between the two parasites and reptiles. This study provides valuable data on the potential role of geckos in endemic areas like Mallorca. Author SummaryLeishmaniosis is a zoonotic disease caused by parasites that can infect both animals and humans. In Europe, Leishmania infantum is the main species responsible for the disease, with dogs acting as its primary host. However, recent research suggests that other animals, including reptiles, might also play a role in their spread. In this study, centered in wild geckos from Mallorca Island (Spain), we investigated the prevalence of L. tarentolae and L. infantum in two gecko species (Tarentola mauritanica and Hemidactylus turcicus). Using quantitative PCR, it was found that 14/49 (28.57%) of adult Tarentola mauritanica were positive for Leishmania parasites, 13/49 (26.53%) for L. tarentolae and 74/49 (8.16%) for L. infantum. Additionally, coinfection of both parasites was detected in three geckos (3/49, 6.12%). The second gecko species studied, Hemidactylus turcicus, was not positive for Leishmania, possible due to a small sample size. These results reinforce previous studies suggesting that geckos may carry both parasites. Since L. tarentolae has also been detected in dogs, and both Leishmania species are sympatric, it raises questions about the role of reptiles in leishmaniosis cycle and transmission, as well as the reliability of current diagnostic tests in coinfections of closely related parasite species.

microbiology↗

bioRxiv2025

Drug-Resistance Biomarkers in Leishmania infantum through Nanopore-Based Detection of Aneuploidy and Gene Copy Number Variations with LeishGenApp

BackgroundDrug-resistant strains of Leishmania infantum challenge the effectiveness of treatments for clinical leishmaniosis and may lead to more frequent relapses. Copy number variation (CNV) at specific genetic loci is associated with drug resistance and virulence, but information about its prevalence in endemic regions is limited. This study examines the drug resistance and virulence status of Leishmania strains in human and canine isolates from the Mediterranean region. MethodsForty-eight Leishmania infantum isolates were whole-genome sequenced with nanopore long reads, followed by de novo assembly. We analyzed chromosomal aneuploidies and gene copy number variation in loci linked to drug resistance and virulence in Leishmania, alongside the genomic structure and rearrangements responsible for these variations. ResultsComplete genomes were de novo assembled for 35 L. infantum isolates (22 from dogs and 13 from humans), revealing significant chromosomal variability. We assessed copy number variation for 22 potential biomarkers: 15 genes related to drug resistance to first-line drugs (METK for allopurinol; LdSMT for amphotericin B; AQP1 and H-locus for antimonials; LdMT, LdRos3, and MSL for miltefosine; PPM for paramomycin) and seven genes related to virulence (lipophosphoglycan and proteophosphoglycan biosynthesis, and the Lack protein). Drug-resistance biomarkers were identified in 80% of the isolates. Canine strains primarily showed resistance to allopurinol and antimonials, while human isolates exhibited a broader resistance spectrum, especially to antimonials and paromomycin. The co-occurrence of resistance biomarkers was common, especially for allopurinol and antimonial resistance. Distinct mechanisms underlie the observed copy number variations. Virulence-associated genes were less variable among isolates. ConclusionsThe prevalence of drug-resistance biomarkers in Leishmania infantum strains from the Mediterranean region, as revealed by this study, underscores the critical need for routine resistance surveillance in managing clinical leishmaniosis. These findings not only inform current clinical practice but also pave the way for more effective management strategies in the future.

genetics↗

bioRxiv2024

Effects of meglumine antimoniate and allopurinol treatment on the fecal microbiome profile in dogs with leishmaniosis

The combination of meglumine antimoniate and allopurinol is considered one of the most effective treatments for canine leishmaniosis caused by Leishmania infantum. This study investigated the effects of this treatment on the gut microbiome of 10 dogs from Spain, Portugal, and Italy via fecal shotgun metagenomic sequencing over six months. Dogs were sampled at baseline (BL) and at one (M1) and six (M6) months post-treatment. The gut microbiome of Leishmania-infected dogs (BL) is dominated by Prevotella, Collinsella, Bacteroides, and Blautia, with individual variability being the primary determinant of microbiome composition. No significant changes in alpha diversity (Shannon index, gene number) or beta diversity (Bray-Curtis dissimilarity, UniFrac distance) were detected between pre- and post-treatment time points, suggesting that treatment with meglumine antimoniate and allopurinol does not disrupt the gut microbiota. Minor trends in taxonomic shifts were noted, with slight increases in Bifidobacterium pseudocantenulatum, Collinsella tanakaei, and Slackia piriformis after treatment, but these changes were not statistically significant after correction for multiple testing. Linear discriminant analysis and multivariable modeling confirmed that the microbial community structure was resilient to treatment effects. Individual-specific microbiome differences in diversity accounted for 52% of the observed variability, underscoring the personalized nature of the gut microbiota in dogs. Importantly, no adverse microbiome disruptions were detected, even with prolonged allopurinol use. This study highlights the robustness of the canine gut microbiome during antileishmanial therapy and highlights the use of meglumine antimoniate and allopurinol without compromising gut microbial diversity or health. Further studies with larger cohorts are recommended to confirm these findings and explore the functional roles of the gut microbiota in modulating immune responses in Leishmania-infected dogs.

molecular biology↗

bioRxiv2023

Immunogenicity and safety in pigs of PHH-1V, a SARS-CoV-2 RBD fusion heterodimer vaccine candidate

The continuing high global incidence of COVID-19 and the undervaccinated status of billions of persons strongly motivate the development of a new generation of efficacious vaccines. We have developed an adjuvanted vaccine candidate, PHH-1V, based on a protein comprising the receptor binding domain (RBD) of the Beta variant of SARS-CoV-2 fused in tandem with the equivalent domain of the Alpha variant, with its immunogenicity, safety and efficacy previously demonstrated in mouse models. In the present study, we immunized pigs with different doses of PHH-1V in a prime-and-boost scheme showing PHH-1V to exhibit an excellent safety profile in pigs and to produce a solid RBD-specific humoral response with neutralising antibodies to 7 distinct SARS-CoV-2 variants of concern, with the induction of a significant IFN{gamma}+ T-cell response. We conclude that PHH-1V is safe and elicits a robust immune response to SARS-CoV-2 in pigs, a large animal preclinical model.

immunology↗

bioRxiv2022

Preclinical efficacy, safety, and immunogenicity of PHH-1V, a second-generation COVID-19 vaccine, in non-human primates

SARS-CoV-2 emerged in December 2019 and quickly spread worldwide, continuously striking with an unpredictable evolution. Despite the success in vaccine production and mass vaccination programmes, the situation is not still completely controlled, and therefore accessible second-generation vaccines are required to mitigate the pandemic. We previously developed an adjuvanted vaccine candidate coded PHH-1V, based on a heterodimer fusion protein comprising the RBD domain of two SARS-CoV-2 variants. Here, we report data on the efficacy, safety, and immunogenicity of PHH-1V in cynomolgus macaques. PHH-1V prime-boost vaccination induces high levels of RBD-specific IgG binding and neutralising antibodies against several SARS-CoV-2 variants, as well as a balanced Th1/Th2 cellular immune response. Remarkably, PHH-1V vaccination prevents SARS-CoV-2 replication in the lower respiratory tract and significantly reduces viral load in the upper respiratory tract after an experimental infection. These results highlight the potential use of the PHH-1V vaccine in humans, currently undergoing Phase III clinical trials.

immunology↗

bioRxiv2021

Preclinical efficacy, safety, and immunogenicity of PHH-1V, a second-generation COVID-19 vaccine candidate based on a novel recombinant RBD fusion heterodimer of SARS-CoV-2.

Current C0VID-19 vaccines have been associated with a decline in infection rates, prevention of severe disease and a decrease in mortality rates. However, SARS-CoV-2 variants are continuously evolving, and development of new accessible COVID-19 vaccines is essential to mitigate the pandemic. Here, we present data on preclinical studies in mice of a receptor-binding domain (RBD)-based recombinant protein vaccine (PHH-1V) consisting of an RBD fusion heterodimer comprising the B.1.351 and B.1.1.7 SARS-CoV-2 variants formulated in SQBA adjuvant, an oil-in-water emulsion. A prime-boost immunisation with PHH-1V in BALB/c and K18-hACE2 mice induced a CD4+ and CD8+ T cell response and RBD-binding antibodies with neutralising activity against several variants, and also showed a good tolerability profile. Significantly, RBD fusion heterodimer vaccination conferred 100% efficacy, preventing mortality in SARS-CoV-2 infected K18-hACE2 mice, but also reducing Beta, Delta and Omicron infection in lower respiratory airways. These findings demonstrate the feasibility of this recombinant vaccine strategy.

immunology↗

bioRxiv2019

Dysbiosis in a canine model of human fistulizing Crohn’s disease

BackgroundCrohns disease (CD) is a chronic immune-mediated inflammatory condition caused by the loss of mucosal tolerance towards the commensal microbiota. Approximately 70% of CD patients experience perianal complications. Perianal fistulizing is a predictor of poor long-term outcomes. Animal models of gut inflammation have failed to fully recapitulate the human manifestations of fistulizing CD. Here, we evaluated dogs with spontaneous canine anal furunculosis (CAF), a disease with clinical similarities to fistulizing CD, as a surrogate model for human fistulizing CD.\n\nResultsBy comparing the gut microbiomes between dogs suffering from CAF (CAF dogs) and healthy dogs, we show that similar to microbiome population trends in CD humans, CAF dogs microbiomes are either very dissimilar (dysbiotic) or similar, yet unique, to healthy dogs microbiomes. Compared to healthy or healthy-like CAF microbiomes, dysbiotic CAF microbiomes showed an increased abundance of Bacteroides vulgatus and Escherichia coli and a decreased abundance of Megamonas species and Prevotella copri. These same determinant bacteria are associated with human CD.\n\nConclusionsOur results highlight the similarities in microbiome community patterns between CAF dogs and CD humans, including overlapping determinant bacterial taxa, and support the use of CAF dogs as a surrogate model to study human fistulizing CD.

microbiology↗