Biology subjects

Meneses, C.

Publications and source records attributed to Meneses, C..

At least 19 recordsLinked to original sources

bioRxiv2026

The PEARL toolkit: Using sand flies to identify leishmaniasis animal reservoirs

Animal reservoirs remain unknown in emerging and most endemic leishmaniasis foci hindering control efforts. Here, we developed a field-applicable PERL (Phlebotomines Establish Reservoirs of Leishmania) toolkit using individual blood fed sand flies (IBF) to identify leishmaniasis reservoirs. Using IBF, we optimized DNA and RNA co-extraction and parasite detection of [≥]1 parasite/s by kDNA qPCR and ssu rRNA RT-qPCR. We then screened IBF for expression of two parasite genes, sherp and HPB, identified by RNAseq as having low-to-absent expression in IBF given a first Leishmania donovani-infected blood meal (IBF-iBM1) and high expression in specimens provided subsequent uninfected blood meals (IBF-BMS+). Linear discriminant analysis of target gene expression classified iBM1 parasites with a predictive accuracy of ~87% and ~82% in membrane- or naturally-fed on hamsters sand flies, respectively. By determining the blood source in specimens determined as IBF-iBM1, the PERL toolkit provides an innovative and practical approach to identification of leishmaniasis reservoirs.

Molecular Biology↗

bioRxiv2026

Cryptic Sex in Leishmania Depends on SPO11 Paralogs

Genetic exchange in Leishmania is established, yet the molecular mechanisms enabling hybrid formation in sand flies remain poorly defined. In Leishmania, as in plants and several protists, two paralogs of the conserved meiotic endonuclease SPO11 are present, but their contribution to hybridization is unknown. Here, we dissect the roles of SPO11-1 and SPO11-2 during in vivo sand fly infections using targeted gene deletions, catalytically-dead mutants, expression analyses, and genome-wide characterization of hybrid progeny. We show that both SPO11 paralogs are essential for efficient hybridization: deletion of either paralog in both parents abolishes hybrid recovery, and catalytic inactivation fails to rescue mating. When only one parent lacks SPO11-1 or SPO11-2, hybrid formation is reduced in a strain-dependent manner, revealing asymmetric requirements for each paralog. Genomic analysis of hybrids from SPO11-deficient crosses reveals polyploidy and altered parental genome contributions, including unbalanced and near-balanced triploid configurations, indicating disrupted reductional processes. Together, these results establish SPO11-dependent DNA break formation as a core requirement for Leishmania hybridization and define distinct, strain-specific roles for the two SPO11 paralogs. Highlights{square} Leishmania encodes two non-redundant SPO11 paralogs that are differentially required in each mating partner for efficient hybridization. {square}The catalytic activity of SPO11 is essential, indicating conservation of its ancestral mode of action. {square}Loss of either paralog disrupts reductional division and prevents balanced genome segregation. {square}Cell fusion proceeds without SPO11, revealing a separation between fusion and meiosis-like functions. {square}SPO11-deficient hybrids show widespread aneuploidy and highly distorted parental genome contributions.

evolutionary biology↗

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↗

bioRxiv2024

Photosynthetic and Genetic Adaptations Underpinning the Resilience of Cistanthe longiscapa in the Atacama Desert

O_LIThe Atacama Desert is one of the most hostile environments for life. However, the plant species Cistanthe longiscapa (C. longiscapa) completes its life cycle in the Atacama Desert after sporadic rainfall. C_LIO_LIPhysiological analyses under controlled environmental conditions revealed superior photosynthetic performance, better light acclimation mechanisms, and larger accumulation of photosystem II in C. longiscapa compared to its mesophilic sister species. C_LIO_LIC. longiscapa shows evolutionary expansions in gene families related to DNA repair, photosynthesis, and protein homeostasis. In addition, we observed substantial gene duplication and polymorphic variations between coastal and inland populations in the Atacama Desert. Finally, our assembled mitochondrial genome provides genetic information for all DNA-containing compartments of C. longiscapa. C_LIO_LIDiurnal oscillations of malic acid and time-resolved transcriptome analyses of plants harvested in the Atacama Desert indicate that C. longiscapa engages in CAM metabolism. We observed significant differences in transcripts encoding plastid-localized proteins, including those involved in carbon metabolism, light harvesting, and photoprotection, highlighting the critical role of chloroplasts in the adaptation of C. longiscapa to the Atacama Desert. C_LIO_LIOur study provides physiological and genetic evidence for the adaptations of C. longiscapa and advances our understanding of how plants can cope with extreme environmental conditions. C_LI

plant biology↗

bioRxiv2024

Enhanced auxin signaling hub triggers root hair growth at moderate low temperature in Arabidopsis thaliana

Root hairs (RH) as a mixed tip- and non-tip growing protrusions that develop from root epidermal cells are important for nutrient and water uptake, root anchoring, and interaction with soil microorganisms. Although nutrient availability and temperature are critical interlinked factors for sustained plant growth, the molecular mechanisms underlying their sensing and downstream signaling pathways remain unclear. Here, we identified a moderate low temperature (10{degrees}C) condition that triggers a strong RH elongation response involving several molecular components of the auxin pathway. Then, we have determined that auxin biosynthesis carried out by YUCCAs/TAA1, the auxin transport conducted by PIN2/PIN4 and AUX1/PGP4, and the auxin signaling controlled by TIR1/AFB2 coupled to four specific ARFs (ARF6/ARF8 and ARF7/ARF19), are all crucial for the RH response at moderate low temperature. These results uncover the auxin pathway as one central hub under moderate low temperature in the roots to trigger RH growth. Our work highlights the importance of moderate low temperature stimulus as a complex nutritional signal from the media soil into the roots that may be fine-tuned for future biotechnological applications to enhance nutrient uptake.

plant biology↗

bioRxiv2024

Sup4h5-L19 activation tagging line partially restores root hair growth in p4h5 mutant by introducing small transcriptomic changes in Arabidopsis thaliana

An specific group of 2-oxoglutarate (2OG) dioxygenases named as Prolyl 4-Hydroxylases (P4H) produce trans-4-hydroxyproline (Hyp/O) from peptidyl-proline, catalyzing proline hydroxylation of cell wall glycoproteins EXT, AGPs, and HRGPs in plant cells, a crucial modification for O-glycosylation. Out of the Arabidopsis thaliana 13 P4Hs, P4H5 regulate root hair cell elongation and T-DNA insertional p4h5 mutant has arrested cell elongation and shortened root hairs. P4H5 selectively hydroxylates EXT proline units indicating that EXT proline hydroxylation as an essential modification for root hair growth. In this work, we isolate an activation-tagging line called Sup4h5-L19/p4h5 (p4h5-L19) that partially suppressed root hair phenotype in the p4h5 mutant background. The T-DNA insertion site was mapped by Thermal Asymmetric Interlaced PCR (TAIL-PCR) followed by PCR product sequencing and the T-DNA is inserted at the beginning of the sixth exon of the AT3G17750 gene, an uncharacterized cytosolic kinase. By analyzing expression changes and mutants analysis in this loci, no clear direct effect was detected. By RNA-seq analysis, it become clear that p4h5-L19 may largely reverse the genetic alterations caused by the p4h5 mutant in Wt Col-0, particularly at 10{degrees}C where there is an increase in root hair growth, with a total of 14 genes that have been activated and 83 genes that have been suppressed due to the enhancer of the activation tagging L19 in p4h5 L19 compared to p4h5. Among these genes, 3 of them, Tonoplast Intrinsic Proteins (TIPs), were identified to be root hair specific (TIP1;1, TIP2;2, and TIP2;3) and the corresponding mutants for two of them (TIP1;1 and TIP2;3) showed reduced root hair growth response at low temperature. This study unmasked new components of the root hair growth response at low temperature that works independently of the O-glycosyated EXTs in the cell walls.

plant biology↗

bioRxiv2024

RSL4 regulatory network acts as an early driver of root hair growth at low temperature in Arabidopsis thaliana

The root hair (RH) cells can elongate to several hundred times their initial size, and are an ideal model system for investigating cell size control. Their development is influenced by both endogenous and external signals, which are combined to form an integrative response. Surprisingly, a low temperature condition of 10{degrees}C causes an increased RH growth in Arabidopsis and in several monocots, even when the development of the rest of the plant are halted. Previously, we demonstrated a strong correlation between the RH growth response and a significant decrease in nutrient availability in the medium under low temperature conditions. However, the molecular basis responsible for receiving and transmitting signals related to the availability of nutrients in the soil, and their relation to plant development, remain largely unknown. In this study, we have discovered two antagonic gene regulatory networks (GRNs) controlling RH early transcriptome responses to low temperature. One GNR enhances RH growth and it is commanded by the transcription factors (FTs) ROOT HAIR DEFECTIVE 6 (RHD6), HAIR DEFECTIVE 6-LIKE 2 and 4 (RSL2-RSL4) and a member of the homeodomain leucine zipper (HD-Zip I) group I 16 (AtHB16). On the other hand, a second GRN was identified as a negative regulator of RH growth at low temperature and it is composed by the trihelix TF GT2-LIKE1 (GTL1) and the associated DF1, a previously unidentified MYB-like TF (AT2G01060) and several members of HD-Zip I group (AtHB3, AtHB13, AtHB20, AtHB23). Functional analysis of both GRNs highlights a complex regulation of RH growth response to low temperature, and more importantly, these discoveries enhance our comprehension of how plants synchronize the RH growth in response to variations in temperature at the cellular level. Significance StatementRoot hair (RH) cells may expand hundreds of times, making them a useful cell size model. Integrated endogenous and exogenous cues affect their development. Even when plant development ceases, Arabidopsis and other monocots grow RH at 10{degrees}C. We previously established a strong correlation between growth response and a substantial medium nutrition reduction at low temperature. Receiving and transmitting soil nutrient signals and their impact on plant development are unknown molecularly. Our study identified two opposing gene regulatory networks (GRN) that govern early transcriptome responses linked to RH growth at low temperature. Functional analysis shows a complex regulation of the transcriptional cascade to influence the low-temperature RH growth. These results explain how cells coordinate RH formation in response to temperature.

plant biology↗

bioRxiv2022

Transcriptomic reprogramming in a susceptible Phaseolus vulgaris L. variety during Pseudomonas syringae attack: The key role of homogalacturonan methylation

The susceptibility of common bean varieties to Pseudomonas syringae pv. phaseolicola (Pph) has been well-documented. However, the molecular mechanism that drives this susceptibility has not been clarified yet. In an attempt to understand this process, 15-day-old common bean plants, variety rinon, were infected with Pph to analyze the transcriptomic changes during the first steps of the infection (at 2 and 9 h). RNA-seq analysis showed an upregulation of defense-and signaling-related genes at 2h, most of them being downregulated at 9h, suggesting that Pph would inhibit the transcriptomic reprogramming of the plant. This trend was also observed in the modulation of 101 cell wall (CW) related genes, suggesting that Pph could produce/induce changes in the CW. However, the changes in CW composition at early stages of Pph infection were related to homogalacturonan (HG) methylation and the formation of HG egg boxes. From all HG-related genes modulated by the infection, a common bean pectin methylesterase inhibitor 3 (PvPMEI3) gene - closely related to AtPMEI3 -- was detected. In addition, PMEI3 protein was located in the apoplast and its PME inhibitory activity was demonstrated. Therefore, PvPMEI3 seems to be a good candidate to play a key role in Pph infection. This premise was supported by the analysis of Arabidopsis pmei3 mutant, which showed susceptibility to Pph, in contrast to resistant Col-0 control plants. All these changes could be an attempt to reinforce the CW structure and thus, hinder the attack of the bacterium. However, these transcriptional and CW-remodeling processes are neither maintained during the necessary time, nor are deep enough to block the action of the pathogen, facilitating the well-known susceptibility of rinon variety to Pph.

plant biology↗

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

Transcriptomic analysis of the C3-CAM transition in Cistanthe longiscapa, a drought tolerant plant in the Atacama Desert

One of the most outstanding plant species during the blooming of the Atacama Desert is the annual plant Cistanthe longiscapa. This plant can perform CAM photosynthesis, but the ecophysiological and molecular mechanisms that this plant uses to withstand the extreme conditions it inhabits in the field are unknown. Morphological and ecophysiological traits were studied and leaf samples at dawn/dusk times were collected from three sites distributed across an increasing south to north arid gradient, to evaluate CAM expression and transcriptomic differences, and search for links between photosynthetic path and abiotic response. Plants from the different sites presented significant differences in nocturnal leaf acid accumulation, isotopic carbon ratio ({delta}13C), succulence and other four traits that clearly indicated a spectrum of CAM photosynthesis intensity that correlated with aridity intensity. The differential gene expression analysis among Dawn vs Dusk between sampling sites showed higher gene expression in the arid northern site (3991 v/s 2293) with activation of regulatory processes associated with abscisic acid and circadian rhythm. The analysis highlights clear ecophysiological differences and the requirement of a strong rewiring of the gene expression to allow a transition from a weak into a strong CAM in C. longiscapa.

plant biology↗

bioRxiv2021

Apoplastic class III peroxidases PRX62 and PRX69 regulate ROS-homeostasis and cell wall associated extensins linked to root hair growth at low-temperature in Arabidopsis thaliana

Root Hairs (RHs) growth is highly influenced by endogenous as well as by external environmental signals that coordinately regulate its final cell size. RHs actively expand the root surface responsible for nutrient uptake and water absorption. We have recently determined that RH growth was unexpectedly boosted when Arabidopsis thaliana seedlings are cultivated at low temperatures. It was proposed that RH growth plasticity in response to low temperature was linked to a reduced nutrient availability in the media. Here, we explored the molecular basis of this strong RH growth response by using the Genome Wide Association Studies (GWAS) approach on Arabidopsis thaliana natural accessions. We identified the poorly characterized PEROXIDASE 62 (PRX62) as a key protein triggering this conditional growth under a moderate low-temperature stress. In addition, we identified the related protein PRX69 as an important factor in this developmental process. The prx62 prx69 double mutant and the PRX62 and PRX69 over-expressing lines showed contrasting RH phenotypes, peroxidase activities and cyt/apoReactive Oxygen Species (ROS) levels. Strikingly, a cell wall protein extensin (EXT) reporter revealed the effect of peroxidase activity on the EXT cell wall association at 10{degrees}C in the RH apical zone. EXT cell wall insolubilization was enhanced at 10{degrees}C, which was completely abolished under the PRX inhibitor salicylhydroxamic acid (SHAM) treatment. Finally, we demonstrated that the Root Hair defective 6-like 4 (RSL4) bHLH family transcription factor directly controls the expression of PRX69. Collectively, our results indicate that both PRX62 and PRX69 are key apoplastic PRXs that modulate ROS-homeostasis and cell wall EXT-insolubilization linked to RH elongation at low-temperature.

plant biology↗

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

Binding of Leishmania infantum LPG to the midgut is not sufficient to define vector competence in Lutzomyia longipalpis sand flies

The major surface lipophosphoglycan (LPG) of Leishmania parasites is critical to vector competence in restrictive sand fly vectors by mediating Leishmania attachment to the midgut epithelium, considered essential to parasite survival and development. However, the relevance of LPG for sand flies that harbor multiple species of Leishmania remains elusive. We tested binding of Leishmania infantum wild type (WT), LPG-defective ({Delta}lpg1 mutants) and add-back lines ({Delta}lpg1 + LPG1) to sand fly midguts in vitro and their survival in Lutzomyia longipalpis sand flies in vivo. Le. infantum WT parasites attached to the Lu. longipalpis midgut in vitro with late-stage parasites binding to midguts in significantly higher numbers compared to early-stage stage promastigotes. {Delta}lpg1 mutants did not bind to Lu. longipalpis midguts, and this was rescued in the {Delta}lpg1 + LPG1 lines, indicating that midgut binding is mediated by LPG. When Lu. longipalpis sand flies were infected with either Le. infantum WT, {Delta}lpg1, or {Delta}lpg1 + LPG1 of the BH46 or BA262 strains, the BH46 {Delta}lpg1 mutant, but not the BA262 {Delta}lpg1 mutant, survived and grew to similar numbers as the WT and {Delta}lpg1 + LPG1 lines. Exposure of BH46 and BA262 {Delta}lpg1 mutants to blood engorged midgut extracts led to the mortality of the BA262 {Delta}lpg1 but not the BH46 {Delta}lpg1 parasites. These findings suggest that Le. infantum LPG protects parasites on a strain-specific basis early in infection, likely against toxic components of blood digestion, however, it is not necessary to prevent Le. infantum evacuation along with the feces in the permissive vector Lu. longipalpis. IMPORTANCEIt is well established that LPG is sufficient to define the vector competence of restrictive sand fly vectors to Leishmania parasites. However, the permissiveness of other sand flies to multiple Leishmania species suggests that other factors might define vector competence for these vectors. In this study, we investigated the underpinnings of Leishmania infantum survival and development in its natural vector Lutzomyia longipalpis. We found out that LPG-mediated midgut binding persists in late-stage parasites. This observation is paradigm-changing and suggests that only a subset of infective metacyclics lose their ability to attach to the midgut with implications for parasite transmission dynamics. However, our data also demonstrate that LPG is not a determining factor in Leishmania infantum retention in the midgut of Lutzomyia longipalpis, a permissive vector. Rather, LPG appears to be more important in protecting some parasite strains from the toxic environment generated during blood meal digestion in the insect gut. Thus, the relevance of LPG in parasite development in permissive vectors appears to be complex and should be investigated on a strain-specific basis.

microbiology↗

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

Analysis of co-regulated abundance of genes associated with arsenic and phosphate metabolism in Andean Microbial Ecosystems

Phosphate and arsenate are very similar compounds, and there is great interest in studying their relationship and their interaction with biological systems. Despite having no apparent biological function, specific genes regulate arsenic interaction with cells and can be located in regions of the genome called arsenic islands, where phosphate metabolism genes are also present. Although they are neighboring genes, the nature of their relationship and how they have been selected is still unknown. In this work, we analyzed the metagenomes of the four microbial ecosystems inhabiting hypersaline lakes of the Argentine Puna and the Atacama salt flat in Chile and have evaluated the presence and abundance of both arsenic and phosphate metabolism genes. The samples analyzed included microbialites, biofilms and microbial mats; all of them established under high arsenic concentrations, high UV radiation and high temperature fluctuation, among others. The results show great differences in the dispersion and abundance of genes related to both phosphate and arsenic metabolism in the analyzed samples. The main difference is given in the Diamante Lake, located in the crater of the Galan volcano characterized by being one of the lakes with the highest arsenic concentration (2.34 mM). Correlating genes abundance with the physicochemical parameters of the lakes studied, our results suggest that arsenic and phosphate metabolism are intricately co-regulated in environmental conditions.

genomics↗

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↗