Biology subjects

Valenzuela, J. G.

Publications and source records attributed to Valenzuela, J. G..

10 recordsLinked to original sources

bioRxiv2025

Discovery of an Adaptive Neuroimmune Response Driving Itch and Fast Tick Removal with Implications for Preventing Pathogen Transmission

Acquired tick resistance (ATR) is well characterized in tick-exposed animals, compromising tick fitness through antibody-mediated activation of basophils. Yet, anti-tick vaccines inducing ATR have had limited success. Here, we describe a neuroimmune event preceding ATR that leads to rapid host-mediated tick removal. Tick-sensitized guinea pigs mechanically remove ticks within 3-6 hours via an acquired neuroimmune-induced itch response that correlates with increased dermal expression of itch-associated genes like OSM and skin infiltration by T cells and macrophages, independently of IgG and IgE antibodies. When we expose humans to ticks, a similar immune response is observed. Blocking T cells before tick sensitization prevents immune cell infiltration to bite sites and abrogates scratching and tick removal. This neuroimmune response is independent of Trpv1 as tick-sensitized guinea pigs treated with resiniferatoxin remove ticks effectively. Itch-induced tick removal or IITR offers a novel approach to tick-borne disease prevention through early tick detection and removal.

neuroscience↗

bioRxiv2025

Comprehensive evaluation of antibody responses to mosquitoes and mosquito-borne arboviruses using highly multiplexed serology

Vector-borne diseases (VBDs) are a leading cause of morbidity and mortality worldwide, and their future impact may increase due to climate change. Antigens driving host immune responses to pathogens and vectors can serve as vaccine candidates and biomarkers of previous exposure, but the immunogenicity of the relevant proteomes remains undercharacterized. To comprehensively profile antibody responses to VBDs and to the vectors themselves, we developed a highly multiplexed phage display library (VectorScan) containing over 250,000 peptides derived from diverse arthropod vectors and prevalent vector-borne pathogens. We used phage immunoprecipitation sequencing (PhIP-Seq) to screen VectorScan against blood samples from non-human primates and humans with experimental and natural exposures to arboviruses and mosquitoes (Anopheles gambiae and Aedes aegypti). We analyzed quantitative measurements of peptide seroreactivity to identify epitopes driving viral serotype-level exposure signatures, as well as novel mosquito sialome antigens. Mosquito-directed antibody responses were associated with natural viral exposure, but highly heterogeneous at an individual level. However, recurrent responses to insect-derived cuticular proteins, mucin-like proteins, and fibrinogen-like proteins will inform development of future serosurveillance tools for vector exposure and vector-based vaccines.

immunology↗

bioRxiv2025

Lutzomyia longipalpis salivary proteins elicit human innate and adaptive immune responses detrimental to Leishmania parasites

In Leishmania-endemic areas, humans are constantly exposed to sand fly bites. To explore the immune consequences of this chronic vector exposure, we performed a human challenge study with the sand fly Lutzomyia longipalpis. Peripheral blood mononuclear cells were collected from fifteen healthy volunteers who underwent multiple controlled exposures to sand fly bites. We identified two Lu. longipalpis salivary proteins, LJM19 and LJL143, which elicited type 1 cytokine responses in cells from exposed individuals and which correlated with enhanced killing of Leishmania parasites in co-cultured macrophages. Interestingly, LJM19 also exerted this parasite-killing effect in cells from unexposed individuals, consistent with innate immune activation. In support of this, both LJM19 and LJL143 stimulated the production of the innate cytokines IL-1{beta} and IFN-. Our results demonstrate that repeated exposure to sand fly bites induces innate and adaptive cytokine responses to vector salivary proteins that can be co-opted to protect humans against Leishmania infection.

immunology↗

bioRxiv2022

Centrin-deficient Leishmania mexicana confers protection against Old World visceral leishmaniasis

Leishmaniasis is one of the top neglected tropical diseases with significant morbidity and mortality in low and middle-income countries (LMIC). However, this disease is also spreading in the developed world. Currently, there is a lack of effective strategies to control this disease. Vaccination can be an effective measure to control leishmaniasis and has the potential to achieve disease elimination. Recently, we have generated centrin gene-deleted new world L. mexicana (LmexCen-/-) parasites using CRISPR/Cas9 and showed that they protect mice against a homologous L. mexicana infection that causes cutaneous disease. In this study, we tested whether LmexCen-/- parasites can also protect against visceral leishmaniasis caused by L. donovani in a hamster model. We show that immunization with LmexCen-/- parasites is safe and does not cause lesions. Furthermore, such immunization conferred protection against visceral leishmaniasis caused by a needle-initiated L. donovani challenge, as indicated by a significant reduction in the parasite burdens in the spleen and liver and lack of mortality. Similar control of parasite burden was also observed against a sand fly mediated L. donovani challenge. Importantly, immunization with LmexCen-/- down-regulated the Th2 response as indicated by a significant reduction in the anti-inflammatory cytokines such as IL-10 and IL-4 and increased pro-inflammatory cytokine IFN-{gamma} resulting in higher IFN-{gamma}/IL-10 and IFN-{gamma}/IL4 ratios compared to non-immunized animals. This contrasts with our studies with L. major centrin deletion mutants that showed a dominant Th1 response compared to L. major wild-type infection suggesting the divergent mechanisms of protection in the two mutant parasites. LmexCen-/- immunization resulted in long-lasting protection against L. donovani infection. Further, since the efficacy of LmCen-/- has not been determined against Leishmania strains prevalent in the Americas, LmexCen-/- may be a viable alternative. Taken together, our study demonstrates that immunization with LmexCen-/- parasites is safe and efficacious against old world visceral leishmaniasis.

immunology↗

bioRxiv2022

Genetic exchange in Leishmania is facilitated by IgM natural antibodies

Host factors mediating Leishmania genetic exchange are not well defined. Here, we demonstrate that IgM antibodies, but not IgG or IgA, facilitate parasite genetic hybridization in vitro and in vivo. IgM induces the gradual and transient formation of a structured parasite clump in a process essential for L. major and L. tropica hybridization in vitro. Parasite hybrids and 3-nucleated parasites were observed inside this structure, named the Leishmania mating clump. IgM was also required for or significantly increased Leishmania hybrid formation in vivo. At minimum, we observed a 12-fold increase in the proportion of hybrids recovered from sand flies provided a second blood meal containing IgM compared to controls. Notably, genetic backcross events in sand flies were only observed in the presence of IgM, and were reproducibly recovered, reinforcing the relevance of IgM for Leishmania genetic exchange in vivo. The in vitro and in vivo Leishmania crosses from these studies resulted in full genome hybrids. Leishmania co-option of a host antibody to facilitate mating in the insect vector establishes a new paradigm of parasite-host-vector coevolution that promotes parasite diversity and fitness through genetic exchange.

microbiology↗

bioRxiv2022

First-in-human evaluation of cutaneous innate and adaptive immunomodulation by mosquito bites

Mosquito-borne viruses are a growing global threat. Initial viral inoculation occurs in the skin via the mosquito bite, eliciting immune responses that shape the establishment of infection and pathogenesis. We aimed to cutaneous innate and adaptive immune responses to mosquito bites in individuals from endemic areas. In this single-arm, cross-sectional interventional study, we enrolled 30 healthy adult participants aged 18 to 45 years of age from Cambodia between October 2020 and January 2021. We performed 3-mm skin biopsies at baseline as well as 30 minutes, 4 hours, and 48 hours after a controlled feeding by uninfected Aedes aegypti mosquitos. The primary endpoints were measurement of changes in early and late innate responses in bitten vs unbitten skin by gene expression profiling, immunophenotyping, and cytokine profiling. Results revealed induction of neutrophil degranulation and recruitment of skin-resident dendritic cells and M2-macrophages in bitten skin. As the immune reaction progressed over time, T cell priming and regulatory pathways were upregulated along with a shift to a Th2-driven response and CD8+ T cell activation. In accordance, participants bitten skin cells produced less pro-inflammatory cytokines when stimulated by Ae. aegypti salivary gland extract. No unexpected adverse events occurred, and one patient was lost to follow-up at Day 14. These results identify key immune genes, cell types, and pathways in the human response to mosquito bites that can be leveraged to develop novel therapeutics and vector-targeted vaccine candidates to arboviral diseases. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/489145v2_ufig1.gif" ALT="Figure 1"> View larger version (77K): org.highwire.dtl.DTLVardef@1ba2eaaorg.highwire.dtl.DTLVardef@4d9dd6org.highwire.dtl.DTLVardef@1b54dc8org.highwire.dtl.DTLVardef@5a104_HPS_FORMAT_FIGEXP M_FIG Panel shows evolution of the human skin response to Aedes aegypti bites over time. C_FIG

immunology↗

bioRxiv2020

Preclinical validation of a second generation leishmanization vaccine against vector transmitted fatal visceral leishmaniasis

Visceral Leishmaniasis (VL) is fatal if untreated. There is no licensed vaccine available against human leishmaniasis. We recently demonstrated protection in mice against L. major infection using a CRISPR genome edited attenuated Leishmania major strain (LmCen-/-). Here, as a pre-clinical step, we evaluated the protective efficacy of LmCen-/- against VL induced by sand fly transmitted Leishmania donovani in hamsters. Intradermal immunization of hamsters with LmCen-/- did not develop any lesion; while still priming a pro-inflammatory immune response. When challenged with L. donovani either by intradermal needle injection or by infected sand flies, LmCen-/--immunized hamsters were protected, not showing spleen or liver pathology averting VL fatality compared to control animals. Spleen cells from LmCen-/- immunized and infected sand fly challenged hamsters produced significantly higher Th1-associated cytokines and chemokines including IFN-{gamma} and TNF-, and significantly reduced expression of the anti-inflammatory cytokines IL-10 and IL-21, compared to non-immunized challenged animals. We further developed a GLP-grade LmCen-/- which showed equal protection as laboratory-grade LmCen-/- parasites in hamsters. Importantly, GLP-grade LmCen-/- parasites also induced a proinflammatory immune response in the PBMCs isolated from healthy people living in non-endemic and endemic for VL as well as cured VL people living in endemic region. Together, this study demonstrates that the LmCen-/- parasites are safe and efficacious against VL and it is a strong candidate vaccine to be tested in a human clinical trial.

immunology↗

bioRxiv2020

A Second Generation Leishmanization Vaccine with a Markerless Attenuated Leishmania major Strain using CRISPR gene editing

Leishmaniasis is a debilitating and often fatal neglected tropical disease caused by Leishmania protozoa transmitted by infected sand flies. Vaccination through leishmanization with live Leishmania major has been used successfully but is no longer practiced because it resulted in unacceptable skin lesions. A second generation leishmanization is described here using a CRISPR genome edited L. major strain (LmCen-/-). Notably, LmCen-/- is the first genetically engineered gene deleted Leishmania strain that is antibiotic resistant marker free and does not have any off-target mutations. Mice immunized with LmCen-/- had virtually no visible lesions following challenge with L. major-infected sand flies while non-immunized animals developed large and progressive lesions with a 2-log fold higher parasite burden. LmCen-/- immunization showed protection and an immune response comparable to leishmanization. LmCen-/- is safe since it was unable to cause disease even in immunocompromised mice, induces robust host protection against vector sand fly challenge and because it is marker free, can be advanced to human vaccine trials.

microbiology↗

bioRxiv2019

Leishmania infection induces a limited differential gene expression in the sand fly midgut

BackgroundPhlebotomine sand flies are the vectors of Leishmania worldwide. To develop in the sand fly midgut, Leishmania multiplies and undergoes multiple stage differentiations leading to the infective form, the metacyclic promastigotes. To gain a better understanding of the influence of Leishmania infection in midgut gene expression, we performed RNA-Seq comparing uninfected Lutzomyia longipalpis midguts and Leishmania infantum-infected Lutzomyia longipalpis midguts at seven time points which cover the various developmental Leishmania stages including early time points when blood digestion is taking place and late time points when the parasites are undergoing metacyclogenesis. ResultsOut of over 13,841 transcripts assembled de novo, only 113 sand fly transcripts, about 1%, were differentially expressed. Further, we observed a low overlap of differentially expressed sand fly transcripts across different time points suggesting a specific influence of each Leishmania stage on midgut gene expression. Two main patterns of sand fly gene expression modulation were noticed. At early time points (days 1-4), more transcripts were down-regulated by Leishmania infection at large fold changes (> -32 fold). Among the down-regulated genes, the transcription factor Forkhead/HNF-3 and hormone degradation enzymes were differentially regulated on day 4 and appear to be the upstream regulators of nutrient transport, digestive enzymes, and peritrophic matrix proteins. Conversely, at later time points (days 6 onwards), most of the differentially expressed transcripts were up-regulated by small fold changes (< 32 fold), and the molecular function of such genes are associated with the metabolism of lipids and detoxification of xenobiotics (P450). ConclusionOverall, it appears that Leishmania modulates sand fly gene expression early on in order to overcome the barriers imposed by the midgut, yet it behaves like a commensal at later time points, when modest midgut gene expression changes correlate with a massive amount of parasites in the anterior midgut.

microbiology↗

bioRxiv2019

Distinct gene expression patterns in vector-residing Leishmania infantum identify parasite stage-enriched markers

Promastigotes of Leishmania infantum undergo a series of extracellular developmental stages inside the natural sand fly vector Lutzomyia longipalpis to reach the infectious stage, the metacyclic promastigote. There is limited information regarding the expression profile of L. infantum developmental stages inside the sand fly vector, and molecular markers that can distinguish the different parasite stages are lacking. We performed RNAseq on unaltered midguts of the sand fly Lutzomyia longipalpis after infection with L. infantum parasites. RNAseq was carried out at various time points throughout parasite development. Principal component analysis mapped the sequences corresponding to the procyclic, nectomonad, leptomonad or metacyclic promastigote stage into distinct positions, with the procyclic stage being the most divergent population. Transcriptional levels across genes varied on average between 10- to 100-fold. Comparison between procyclic and nectomonad promastigotes resulted in 836 differentially expressed (DE) genes; between nectomonad and leptomonad promastigotes in 113 DE genes; and between leptomonad and metacyclic promastigotes in 302 DE genes. Most of the DE genes do not overlap across stages, highlighting the uniqueness of each stage. Furthermore, the different stages of Leishmania parasites exhibited specific transcriptional enrichment across chromosomes. Using the transcriptional signatures exhibited by distinct Leishmania stages during their development in the sand fly midgut, we determined the genes predominantly enriched in each stage, identifying multiple stage-specific markers for L. Infantum. Leading stage-specific marker candidates include genes encoding a zinc transporter in procyclics, a beta-fructofuranidase in nectomonads, a surface antigen-like protein in leptomonads, and an amastin-like surface protein in metacyclics. Overall, these findings demonstrate the transcriptional plasticity of the Leishmania parasite inside the sand fly vector and provide a repertoire of stage-specific markers for further development as molecular tools for epidemiological studies.

microbiology↗