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1. Wide reference databases for typing Trypanosoma cruzi based on amplicon sequencing of the minicircle hypervariable region.

Author

Fanny Rusman, Anahí G Díaz, Tatiana Ponce, Noelia Floridia-Yapur, Christian Barnabé, Patricio Diosque, and Nicolás Tomasini

Subjects
Arctic medicine. Tropical medicine, RC955-962, Public aspects of medicine, RA1-1270
Abstract

BackgroundTrypanosoma cruzi, the etiological agent of Chagas Disease, exhibits remarkable genetic diversity and is classified into different Discrete Typing Units (DTUs). Strain typing techniques are crucial for studying T. cruzi, because their DTUs have significant biological differences from one another. However, there is currently no methodological strategy for the direct typing of biological materials that has sufficient sensitivity, specificity, and reproducibility. The high diversity and copy number of the minicircle hypervariable regions (mHVRs) makes it a viable target for typing.Methodology/principal findingsApproximately 24 million reads obtained by amplicon sequencing of the mHVR were analyzed for 62 strains belonging to the six main T. cruzi DTUs. To build reference databases of mHVR diversity for each DTU and to evaluate this target as a typing tool. Strains of the same DTU shared more mHVR clusters than strains of different DTUs, and clustered together. Different identity thresholds were used to build the reference sets of the mHVR sequences (85% and 95%, respectively). The 95% set had a higher specificity and was more suited for detecting co-infections, whereas the 85% set was excellent for identifying the primary DTU of a sample. The workflow's capacity for typing samples obtained from cultures, a set of whole-genome data, under various simulated PCR settings, in the presence of co-infecting lineages and for blood samples was also assessed.Conclusions/significanceWe present reference databases of mHVR sequences and an optimized typing workflow for T. cruzi including a simple online tool for deep amplicon sequencing analysis (https://ntomasini.github.io/cruzityping/). The results show that the workflow displays an equivalent resolution to that of the other typing methods. Owing to its specificity, sensitivity, relatively low cost, and simplicity, the proposed workflow could be an alternative for screening different types of samples.

Published
2023
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2. Guide RNA Repertoires in the Main Lineages of Trypanosoma cruzi: High Diversity and Variable Redundancy Among Strains

Author

Fanny Rusman, Noelia Floridia-Yapur, Nicolás Tomasini, and Patricio Diosque

Subjects
Chagas disease, kinetoplastids, minicircles, RNA editing, DTU, Microbiology, QR1-502
Abstract

Trypanosoma cruzi, as other kinetoplastids, has a complex mechanism of editing of mitochondrial mRNAs that requires guide RNAs (gRNAs) coded in DNA minicircles in the kinetoplast. There are many variations on this mechanism among species. mRNA editing and gRNA repertoires are almost unknown in T. cruzi. Here, gRNAs were inferred based on deep-sequenced minicircle hypervariable regions (mHVRs) and editing cascades were rebuilt in strains belonging to the six main T. cruzi lineages. Inferred gRNAs were clustered according to their sequence similarity to constitute gRNA classes. Extreme diversity of gRNA classes was observed, which implied highly divergent gRNA repertoires among different lineages, even within some lineages. In addition, a variable gRNA class redundancy (i.e., different gRNA classes editing the same mRNA region) was detected among strains. Some strains had upon four times more gRNA classes than others. Such variations in redundancy affected gRNA classes of all mRNAs in a concerted way, i.e., there are correlated variations in the number of gRNAs classes editing each mRNA. Interestingly, cascades were incomplete for components of the respiratory complex I in several strains. Finally, gRNA classes of different strains may potentially edit mitochondrial mRNAs from other lineages in the same way as they edit their own mitochondrial mRNAs, which is a prerequisite for biparental inheritance of minicircle in hybrids. We propose that genetic exchange and biparental inheritance of minicircles combined with minicircle drift due to (partial) random segregation of minicircles during kDNA replication is a suitable hypothesis to explain the divergences among strains and the high levels of gRNA redundancy in some strains. In addition, our results support that the complex I may not be required in some stages in the life cycle as previously shown and that linkage (in the same minicircle) of gRNAs that edit different mRNAs may prevent gRNA class lost in such stage.

Published
2021
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3. Evidence of hybridization, mitochondrial introgression and biparental inheritance of the kDNA minicircles in Trypanosoma cruzi I.

Author

Fanny Rusman, Noelia Floridia-Yapur, Paula G Ragone, Patricio Diosque, and Nicolás Tomasini

Subjects
Arctic medicine. Tropical medicine, RC955-962, Public aspects of medicine, RA1-1270
Abstract

BACKGROUND:Genetic exchange in Trypanosoma cruzi is controversial not only in relation to its frequency, but also to its mechanism. Parasexual genetic exchange has been proposed based on laboratory hybrids, but population genomics strongly suggests meiosis in T. cruzi. In addition, mitochondrial introgression has been reported several times in natural isolates although its mechanism is not fully understood yet. Moreover, hybrid T. cruzi DTUs (TcV and TcVI) have inherited at least part of the kinetoplastic DNA (kDNA = mitochondrial DNA) from both parents. METHODOLOGY/PRINCIPAL FINDINGS:In order to address such topics, we sequenced and analyzed fourteen nuclear DNA fragments and three kDNA maxicircle genes in three TcI stocks which are natural clones potentially involved in events of genetic exchange. We also deep-sequenced (a total of 6,146,686 paired-end reads) the minicircle hypervariable region (mHVR) of the kDNA in such three strains. In addition, we analyzed the DNA content by flow cytometry to address cell ploidy. We observed that most polymorphic sites in nuclear loci showed a hybrid pattern in one cloned strain and the other two cloned strains were compatible as parental strains (or nearly related to the true parents). The three clones had almost the same ploidy and the DNA content was similar to the reference strain Sylvio (a nearly diploid strain). Despite maxicircle genes evolve faster than nuclear housekeeping ones, we detected no polymorphisms in the sequence of three maxicircle genes showing mito-nuclear discordance. Lastly, the hybrid stock shared 66% of its mHVR clusters with one putative parent and 47% with the other one; in contrast, the putative parental stocks shared less than 30% of the mHVR clusters between them. CONCLUSIONS/SIGNIFICANCE:The results suggest a reductive division, a natural hybridization, biparental inheritance of the minicircles in the hybrid and maxicircle introgression. The models including such phenomena and explaining the relationships between these three clones are discussed.

Published
2020
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4. Elucidating diversity in the class composition of the minicircle hypervariable region of Trypanosoma cruzi: New perspectives on typing and kDNA inheritance.

Author

Fanny Rusman, Nicolás Tomasini, Noelia-Floridia Yapur, Andrea F Puebla, Paula G Ragone, and Patricio Diosque

Subjects
Arctic medicine. Tropical medicine, RC955-962, Public aspects of medicine, RA1-1270
Abstract

BACKGROUND:Trypanosoma cruzi, the protozoan causative of Chagas disease, is classified into six main Discrete Typing Units (DTUs): TcI-TcVI. This parasite has around 105 copies of the minicircle hypervariable region (mHVR) in their kinetoplastic DNA (kDNA). The genetic diversity of the mHVR is virtually unknown. However, cross-hybridization assays using mHVRs showed hybridization only between isolates belonging to the same genetic group. Nowadays there is no methodologic approach with a good sensibility, specificity and reproducibility for direct typing on biological samples. Due to its high copy number and apparently high diversity, mHVR becomes a good target for typing. METHODOLOGY/PRINCIPAL FINDINGS:Around 22 million reads, obtained by amplicon sequencing of the mHVR, were analyzed for nine strains belonging to six T. cruzi DTUs. The number and diversity of mHVR clusters was variable among DTUs and even within a DTU. However, strains of the same DTU shared more mHVR clusters than strains of different DTUs and clustered together. In addition, hybrid DTUs (TcV and TcVI) shared similar percentages (1.9-3.4%) of mHVR clusters with their parentals (TcII and TcIII). Conversely, just 0.2% of clusters were shared between TcII and TcIII suggesting biparental inheritance of the kDNA in hybrids. Sequencing at low depth (20,000-40,000 reads) also revealed 95% of the mHVR clusters for each of the analyzed strains. Finally, the method revealed good correlation in cluster identity and abundance between different replications of the experiment (r = 0.999). CONCLUSIONS/SIGNIFICANCE:Our work sheds light on the sequence diversity of mHVRs at intra and inter-DTU level. The mHVR amplicon sequencing workflow described here is a reproducible technique, that allows multiplexed analysis of hundreds of strains and results promissory for direct typing on biological samples in a future. In addition, such approach may help to gain knowledge on the mechanisms of the minicircle evolution and phylogenetic relationships among strains.

Published
2019
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5. A Novel Genotype and First Record of Trypanosoma lainsoni in Argentina

Author

Anahí G. Díaz, Paula G. Ragone, Fanny Rusman, Noelia Floridia-Yapur, Rubén M. Barquez, M. Mónica Díaz, Nicolás Tomasini, and Patricio Diosque

Subjects
Trypanosoma lainsoni, Leopardus geoffroyi, Calomys spp., 18S rDNA genes, gGAPDH genes, morphology, Medicine
Abstract

Trypanosomes are a group of parasitic flagellates with medical and veterinary importance. Despite many species having been described in this genus, little is known about many of them. Here, we report a genetic and morphological characterization of trypanosomatids isolated from wild mammals from the Argentine Chaco region. Parasites were morphologically and ultrastructurally characterized by light microscopy and transmission electron microscopy. Additionally, 18s rRNA and gGAPDH genes were sequenced and analyzed using maximum likelihood and Bayesian inference. Morphological characterization showed clear characteristics associated with the Trypanosoma genus. The genetic characterization demonstrates that the studied isolates have identical sequences and a pairwise identity of 99% with Trypanosoma lainsoni, which belongs to the clade of lizards and snakes/rodents and marsupials. To date, this species had only been found in the Amazon region. Our finding represents the second report of T. lainsoni and the first record for the Chaco region. Furthermore, we ultrastructurally described for the first time the species. Finally, the host range of T. lainsoni was expanded (Leopardus geoffroyi, Carenivora, Felidae; and Calomys sp., Rodentia, Cricetidae), showing a wide host range for this species.

Published
2020
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6. Evolution of Trypanosoma cruzi: clarifying hybridisations, mitochondrial introgressions and phylogenetic relationships between major lineages

Author

Nicolás Tomasini and Patricio Diosque

Subjects
Chagas disease, parasite, phylogeny, Trypanosomatidae, discrete typing units, Microbiology, QR1-502, Infectious and parasitic diseases, RC109-216
Abstract

Several different models of Trypanosoma cruzi evolution have been proposed. These models suggest that scarce events of genetic exchange occurred during the evolutionary history of this parasite. In addition, the debate has focused on the existence of one or two hybridisation events during the evolution of T. cruzi lineages. Here, we reviewed the literature and analysed available sequence data to clarify the phylogenetic relationships among these different lineages. We observed that TcI, TcIII and TcIV form a monophyletic group and that TcIII and TcIV are not, as previously suggested, TcI-TcII hybrids. Particularly, TcI and TcIII are sister groups that diverged around the same time that a widely distributed TcIV split into two clades (TcIVS and TcIVN). In addition, we collected evidence that TcIII received TcIVS kDNA by introgression on several occasions. Different demographic hypotheses (surfing and asymmetrical introgression) may explain the origin and expansion of the TcIII group. Considering these hypotheses, genetic exchange should have been relatively frequent between TcIII and TcIVS in the geographic area in which their distributions overlapped. In addition, our results support the hypothesis that two independent hybridisation events gave rise to TcV and TcVI. Consequently, TcIVS kDNA was first transferred to TcIII and later to TcV and TcVI in TcII/TcIII hybridisation events.

Published
2015
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7. Epidemiological modeling of Trypanosoma cruzi: Low stercorarian transmission and failure of host adaptive immunity explain the frequency of mixed infections in humans.

Author

Nicolás Tomasini, Paula Gabriela Ragone, Sébastien Gourbière, Juan Pablo Aparicio, and Patricio Diosque

Subjects
Biology (General), QH301-705.5
Abstract

People living in areas with active vector-borne transmission of Chagas disease have multiple contacts with its causative agent, Trypanosoma cruzi. Reinfections by T. cruzi are possible at least in animal models leading to lower or even hardly detectable parasitaemia. In humans, although reinfections are thought to have major public health implications by increasing the risk of chronic manifestations of the disease, there is little quantitative knowledge about their frequency and the timing of parasite re-inoculation in the course of the disease. Here, we implemented stochastic agent-based models i) to estimate the rate of re-inoculation in humans and ii) to assess how frequent are reinfections during the acute and chronic stages of the disease according to alternative hypotheses on the adaptive immune response following a primary infection. By using a hybrid genetic algorithm, the models were fitted to epidemiological data of Argentinean rural villages where mixed infections by different genotypes of T. cruzi reach 56% in humans. To explain this percentage, the best model predicted 0.032 (0.008-0.042) annual reinfections per individual with 98.4% of them occurring in the chronic phase. In addition, the parasite escapes to the adaptive immune response mounted after the primary infection in at least 20% of the events of re-inoculation. With these low annual rates, the risks of reinfection during the typically long chronic stage of the disease stand around 14% (4%-18%) and 60% (21%-70%) after 5 and 30 years, with most individuals being re-infected 1-3 times overall. These low rates are better explained by the weak efficiency of the stercorarian mode of transmission than a highly efficient adaptive immune response. Those estimates are of particular interest for vaccine development and for our understanding of the higher risk of chronic disease manifestations suffered by infected people living in endemic areas.

Published
2017
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8. Experimental evidence of biological interactions among different isolates of Trypanosoma cruzi from the Chaco Region.

Author

Paula G Ragone, Cecilia Pérez Brandán, Mercedes Monje Rumi, Nicolás Tomasini, Juan J Lauthier, Rubén O Cimino, Alejandro Uncos, Federico Ramos, Anahí M Alberti D'Amato, Miguel A Basombrío, and Patricio Diosque

Subjects
Medicine, Science
Abstract

Many infectious diseases arise from co-infections or re-infections with more than one genotype of the same pathogen. These mixed infections could alter host fitness, the severity of symptoms, success in pathogen transmission and the epidemiology of the disease. Trypanosoma cruzi, the etiological agent of Chagas disease, exhibits a high biological variability often correlated with its genetic diversity. Here, we developed an experimental approach in order to evaluate biological interaction between three T. cruzi isolates belonging to different Discrete Typing Units (DTUs TcIII, TcV and TcVI). These isolates were obtained from a restricted geographical area in the Chaco Region. Different mixed infections involving combinations of two isolates (TcIII + TcV, TcIII + TcVI and TcV + TcVI) were studied in a mouse model. The parameters evaluated were number of parasites circulating in peripheral blood, histopathology and genetic characterization of each DTU in different tissues by DNA hybridization probes. We found a predominance of TcVI isolate in blood and tissues respect to TcIII and TcV; and a decrease of the inflammatory response in heart when the damage of mice infected with TcVI and TcIII + TcVI mixture were compared. In addition, simultaneous presence of two isolates in the same tissue was not detected. Our results show that biological interactions between isolates with different biological behaviors lead to changes in their biological properties. The occurrence of interactions among different genotypes of T. cruzi observed in our mouse model suggests that these phenomena could also occur in natural cycles in the Chaco Region.

Published
2015
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9. Optimized multilocus sequence typing (MLST) scheme for Trypanosoma cruzi.

Author

Patricio Diosque, Nicolás Tomasini, Juan José Lauthier, Louisa Alexandra Messenger, María Mercedes Monje Rumi, Paula Gabriela Ragone, Anahí Maitén Alberti-D'Amato, Cecilia Pérez Brandán, Christian Barnabé, Michel Tibayrenc, Michael David Lewis, Martin Stephen Llewellyn, Michael Alexander Miles, and Matthew Yeo

Subjects
Arctic medicine. Tropical medicine, RC955-962, Public aspects of medicine, RA1-1270
Abstract

Trypanosoma cruzi, the aetiological agent of Chagas disease possess extensive genetic diversity. This has led to the development of a plethora of molecular typing methods for the identification of both the known major genetic lineages and for more fine scale characterization of different multilocus genotypes within these major lineages. Whole genome sequencing applied to large sample sizes is not currently viable and multilocus enzyme electrophoresis, the previous gold standard for T. cruzi typing, is laborious and time consuming. In the present work, we present an optimized Multilocus Sequence Typing (MLST) scheme, based on the combined analysis of two recently proposed MLST approaches. Here, thirteen concatenated gene fragments were applied to a panel of T. cruzi reference strains encompassing all known genetic lineages. Concatenation of 13 fragments allowed assignment of all strains to the predicted Discrete Typing Units (DTUs), or near-clades, with the exception of one strain that was an outlier for TcV, due to apparent loss of heterozygosity in one fragment. Monophyly for all DTUs, along with robust bootstrap support, was restored when this fragment was subsequently excluded from the analysis. All possible combinations of loci were assessed against predefined criteria with the objective of selecting the most appropriate combination of between two and twelve fragments, for an optimized MLST scheme. The optimum combination consisted of 7 loci and discriminated between all reference strains in the panel, with the majority supported by robust bootstrap values. Additionally, a reduced panel of just 4 gene fragments displayed high bootstrap values for DTU assignment and discriminated 21 out of 25 genotypes. We propose that the seven-fragment MLST scheme could be used as a gold standard for T. cruzi typing, against which other typing approaches, particularly single locus approaches or systematic PCR assays based on amplicon size, could be compared.

Published
2014
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10. How often do they have sex? A comparative analysis of the population structure of seven eukaryotic microbial pathogens.

Author

Nicolás Tomasini, Juan José Lauthier, Francisco José Ayala, Michel Tibayrenc, and Patricio Diosque

Subjects
Medicine, Science
Abstract

The model of predominant clonal evolution (PCE) proposed for micropathogens does not state that genetic exchange is totally absent, but rather, that it is too rare to break the prevalent PCE pattern. However, the actual impact of this "residual" genetic exchange should be evaluated. Multilocus Sequence Typing (MLST) is an excellent tool to explore the problem. Here, we compared online available MLST datasets for seven eukaryotic microbial pathogens: Trypanosoma cruzi, the Fusarium solani complex, Aspergillus fumigatus, Blastocystis subtype 3, the Leishmania donovani complex, Candida albicans and Candida glabrata. We first analyzed phylogenetic relationships among genotypes within each dataset. Then, we examined different measures of branch support and incongruence among loci as signs of genetic structure and levels of past recombination. The analyses allow us to identify three types of genetic structure. The first was characterized by trees with well-supported branches and low levels of incongruence suggesting well-structured populations and PCE. This was the case for the T. cruzi and F. solani datasets. The second genetic structure, represented by Blastocystis spp., A. fumigatus and the L. donovani complex datasets, showed trees with weakly-supported branches but low levels of incongruence among loci, whereby genetic structuration was not clearly defined by MLST. Finally, trees showing weakly-supported branches and high levels of incongruence among loci were observed for Candida species, suggesting that genetic exchange has a higher evolutionary impact in these mainly clonal yeast species. Furthermore, simulations showed that MLST may fail to show right clustering in population datasets even in the absence of genetic exchange. In conclusion, these results make it possible to infer variable impacts of genetic exchange in populations of predominantly clonal micro-pathogens. Moreover, our results reveal different problems of MLST to determine the genetic structure in these organisms that should be considered.

Published
2014
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11. Growth of peripheral and central nervous system tumors is supported by cytoplasmic c-Fos in humans and mice.

Author

David C Silvestre, Germán A Gil, Nicolás Tomasini, Daniela F Bussolino, and Beatriz L Caputto

Subjects
Medicine, Science
Abstract

BACKGROUND: We have previously shown that the transcription factor c-Fos is also capable of associating to endoplasmic reticulum membranes (ER) and activating phospholipid synthesis. Herein we examined phospholipid synthesis status in brain tumors from human patients and from NPcis mice, an animal model of the human disease Neurofibromatosis Type 1 (NF1). PRINCIPAL FINDINGS: In human samples, c-Fos expression was at the limit of detection in non-pathological specimens, but was abundantly expressed associated to ER membranes in tumor cells. This was also observed in CNS of adult tumor-bearing NPcis mice but not in NPcis fos(-/-) KO mice. A glioblastoma multiforme and a malignant PNS tumor from a NF1 patient (MPNST) showed a 2- and 4- fold c-Fos-dependent phospholipid synthesis activation, respectively. MPNST samples also showed increased cell proliferation rates and abundant c-Fos expression. CONCLUSIONS: Results highlight a role of cytoplasmic c-Fos as an activator of phospholipid synthesis in events demanding high rates of membrane biogenesis as occurs for the exacerbated growth of tumors cells. They also disclose this protein as a potential target for controlling tumor growth in the nervous system.

Published
2010
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12. Hydrophobicity-Driven Increases in Editing in Mitochondrial mRNAs during the Evolution of Kinetoplastids

Author

Fanny Rusman, Noelia Floridia-Yapur, Anahí G Díaz, Tatiana Ponce, Patricio Diosque, and Nicolás Tomasini

Subjects
Genetics, Molecular Biology, Ecology, Evolution, Behavior and Systematics
Abstract

Kinetoplastids are a diverse group of flagellates which exhibit editing by insertion/deletion of Us in the mitochondrial mRNAs. Some mRNAs require editing to build most of their coding sequences, a process known as pan-editing. Evidence suggests that pan-editing is an ancestral feature in kinetoplastids. Here, we investigate how the transition from nonedited to pan-edited states occurred. The mitochondrial mRNAs and protein sequences from nine kinetoplastids and related groups (diplonemids, euglenids, and jakobids) were analyzed. RNA editing increased protein hydrophobicity to extreme values by introducing Us in the second codon position, despite the absence of editing preferences related to codon position. In addition, hydrophobicity was maintained by purifying selection in species that lost editing by retroposition of the fully edited mRNA. Only a few hydrophobic to hydrophilic amino acid changes were inferred for such species. In the protein secondary structure, these changes occurred spatially close to other hydrophilic residues. The analysis of coevolving sites showed that multiple changes are required together for hydrophobicity to be lost, which suggest the proteins are locked into extended hydrophobicity. Finally, an analysis of the NAD7 protein–protein interactions showed they can also influence hydrophobicity increase in the protein and where editing can occur in the mRNA. In conclusion, our results suggest that protein hydrophobicity has influenced editing site selection and how editing expanded in mRNAs. In effect, the hydrophobicity increase was entrenched by a neutral ratchet moved by a mutational pressure to introduce Us, thus helping to explain both RNA editing increase and, possibly, persistence.

Published
2023
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13. TcTASV Antigens of Trypanosoma cruzi: Utility for Diagnosis and High Accuracy as Biomarkers of Treatment Efficacy in Pediatric Patients

Author

Rubén O. Cimino, Mercedes Monje-Rumi, Anahí A. Alberti D’Amato, Daniel O. Sánchez, José Fernando Gil, Valeria Tekiel, Nicolás Tomasini, Noelia Floridia-Yapur, Patricio Diosque, Julio R. Nasser, Juan José Lauthier, and Paula G. Ragone

Subjects
030231 tropical medicine, CHAGAS, Diagnostic accuracy, Antibody level, Strongyloides stercoralis, Serology, purl.org/becyt/ford/3.3 [https], 03 medical and health sciences, SEROLOGIA, FAMILAI MULTIGENICA, 0302 clinical medicine, Antigen, Virology, Parasite hosting, Medicine, Trypanosoma cruzi, TcTASV, biology, business.industry, ANTIGENOS SUPERFICIE, biology.organism_classification, Treatment efficacy, Infectious Diseases, Immunology, DIAGNOSTICO, SEGUIMIENTO, ELISA, purl.org/becyt/ford/3 [https], Parasitology, business
Abstract

The discovery and characterization of novel parasite antigens to improve the diagnosis of Trypanosoma cruzi by serological methods and for accurate and rapid follow-up of treatment efficiency are still needed. TcTASV is a T. cruzi-specific multigene family, whose products are expressed on the parasite stages present in the vertebrate host. In a previous work, a mix of antigens from subfamilies TcTASV-A and TcTASV-C (Mix A + C) was sensitive and specific to identify dogs with active infection of high epidemiological relevance. Here, TcTASV-A and TcTASV-C were assayed separately as well as together (Mix A + C) in an ELISA format on human samples. The Mix A + C presented moderate sensitivity (78%) but high diagnostic accuracy with a 100% of specificity, evaluated on healthy, leishmaniasic, and Strongyloides stercoralis infected patients. Moreover, antibody levels of pediatric patients showed-2 years posttreatment-diminished reactivity against the Mix A + C (P < 0.0001), pointing TcTASV antigens as promising tools for treatment follow-up. Fil: Floridia Yapur, Noelia Aldana del Rosario. Universidad Nacional de Salta. Facultad de Ciencias Naturales; Argentina. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; Argentina Fil: Monje Rumi, Maria Mercedes. Universidad Nacional de Salta. Facultad de Ciencias Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: D'Amato, Anahí Alberti. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Cimino, Rubén Oscar. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; Argentina Fil: Gil, José Fernando. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Investigaciones en Energía no Convencional. Universidad Nacional de Salta. Facultad de Ciencias Exactas. Departamento de Física. Instituto de Investigaciones en Energía no Convencional; Argentina Fil: Sanchez, Daniel Oscar. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina Fil: Nasser, Julio Rubén. Universidad Nacional de Salta. Facultad de Ciencias Naturales; Argentina Fil: Tekiel, Valeria Sonia. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina

Published
2019
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14. Guide RNA Repertoires in the Main Lineages of Trypanosoma cruzi: High Diversity and Variable Redundancy Among Strains

Author

Noelia Floridia-Yapur, Nicolás Tomasini, Fanny Rusman, and Patricio Diosque

Subjects
0301 basic medicine, Microbiology (medical), Chagas disease, RNA editing, Trypanosoma cruzi, Trypanosoma brucei brucei, 030231 tropical medicine, Immunology, KINETOPLASTIDS, Biology, Minicircle, Microbiology, purl.org/becyt/ford/3.3 [https], 03 medical and health sciences, chemistry.chemical_compound, Cellular and Infection Microbiology, 0302 clinical medicine, MINICIRCLES, RNA EDITING, Humans, Guide RNA, Original Research, minicircles, Genetics, Base Sequence, DNA, Kinetoplast, CHAGAS DISEASE, Inheritance (genetic algorithm), biology.organism_classification, QR1-502, Hypervariable region, kinetoplastids, 030104 developmental biology, Infectious Diseases, chemistry, Kinetoplast, DTU, purl.org/becyt/ford/3 [https], DNA, RNA, Guide, Kinetoplastida
Abstract

Trypanosoma cruzi, as other kinetoplastids, has a complex mechanism of editing of mitochondrial mRNAs that requires guide RNAs (gRNAs) coded in DNA minicircles in the kinetoplast. There are many variations on this mechanism among species. mRNA editing and gRNA repertoires are almost unknown in T. cruzi. Here, gRNAs were inferred based on deep-sequenced minicircle hypervariable regions (mHVRs) and editing cascades were rebuilt in strains belonging to the six main T. cruzi lineages. Inferred gRNAs were clustered according to their sequence similarity to constitute gRNA classes. Extreme diversity of gRNA classes was observed, which implied highly divergent gRNA repertoires among different lineages, even within some lineages. In addition, a variable gRNA class redundancy (i.e., different gRNA classes editing the same mRNA region) was detected among strains. Some strains had upon four times more gRNA classes than others. Such variations in redundancy affected gRNA classes of all mRNAs in a concerted way, i.e., there are correlated variations in the number of gRNAs classes editing each mRNA. Interestingly, cascades were incomplete for components of the respiratory complex I in several strains. Finally, gRNA classes of different strains may potentially edit mitochondrial mRNAs from other lineages in the same way as they edit their own mitochondrial mRNAs, which is a prerequisite for biparental inheritance of minicircle in hybrids. We propose that genetic exchange and biparental inheritance of minicircles combined with minicircle drift due to (partial) random segregation of minicircles during kDNA replication is a suitable hypothesis to explain the divergences among strains and the high levels of gRNA redundancy in some strains. In addition, our results support that the complex I may not be required in some stages in the life cycle as previously shown and that linkage (in the same minicircle) of gRNAs that edit different mRNAs may prevent gRNA class lost in such stage. Fil: Rusman, Fanny. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Floridia Yapur, Noelia Aldana del Rosario. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina

Published
2021
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15. Evidence of hybridization, mitochondrial introgression and biparental inheritance of the kDNA minicircles in Trypanosoma cruzi I

Author

Paula G. Ragone, Noelia Floridia-Yapur, Fanny Rusman, Patricio Diosque, and Nicolás Tomasini

Subjects
0301 basic medicine, Introgression, Genes, Protozoan, RC955-962, Biochemistry, purl.org/becyt/ford/1 [https], Spectrum Analysis Techniques, 0302 clinical medicine, Arctic medicine. Tropical medicine, Genotype, Cell Cycle and Cell Division, Energy-Producing Organelles, Protozoans, Genetics, KDNA, education.field_of_study, Chromosome Biology, DNA, Kinetoplast, Database and informatics methods, Sequence analysis, High-Throughput Nucleotide Sequencing, Eukaryota, Bioquímica y Biología Molecular, Flow Cytometry, Mitochondria, Nuclear DNA, Meiosis, Infectious Diseases, Cell Processes, Spectrophotometry, TCI, Cytophotometry, Cellular Structures and Organelles, Public aspects of medicine, RA1-1270, CIENCIAS NATURALES Y EXACTAS, HYBRIDIZATION, Research Article, Trypanosoma, Mitochondrial DNA, Evolutionary Processes, Bioinformatics, Trypanosoma cruzi, 030231 tropical medicine, Population, Genomics, Bioenergetics, Biology, Minicircle, DNA sequencing, Ciencias Biológicas, 03 medical and health sciences, BIPARENTAL INHERITANCE, education, purl.org/becyt/ford/1.6 [https], Gene, DNA sequence analysis, Evolutionary Biology, Ploidies, Organisms, Public Health, Environmental and Occupational Health, Biology and Life Sciences, Sequence Analysis, DNA, Cell Biology, TRYPANOSOMA CRUZI, DNA, Protozoan, Parasitic Protozoans, MITOCHONDRIAL INTROGRESSION, Hypervariable region, Research and analysis methods, Kinetoplasts, 030104 developmental biology, Genetic Loci, Hybridization, Genetic
Abstract

Background Genetic exchange in Trypanosoma cruzi is controversial not only in relation to its frequency, but also to its mechanism. Parasexual genetic exchange has been proposed based on laboratory hybrids, but population genomics strongly suggests meiosis in T. cruzi. In addition, mitochondrial introgression has been reported several times in natural isolates although its mechanism is not fully understood yet. Moreover, hybrid T. cruzi DTUs (TcV and TcVI) have inherited at least part of the kinetoplastic DNA (kDNA = mitochondrial DNA) from both parents. Methodology/Principal findings In order to address such topics, we sequenced and analyzed fourteen nuclear DNA fragments and three kDNA maxicircle genes in three TcI stocks which are natural clones potentially involved in events of genetic exchange. We also deep-sequenced (a total of 6,146,686 paired-end reads) the minicircle hypervariable region (mHVR) of the kDNA in such three strains. In addition, we analyzed the DNA content by flow cytometry to address cell ploidy. We observed that most polymorphic sites in nuclear loci showed a hybrid pattern in one cloned strain and the other two cloned strains were compatible as parental strains (or nearly related to the true parents). The three clones had almost the same ploidy and the DNA content was similar to the reference strain Sylvio (a nearly diploid strain). Despite maxicircle genes evolve faster than nuclear housekeeping ones, we detected no polymorphisms in the sequence of three maxicircle genes showing mito-nuclear discordance. Lastly, the hybrid stock shared 66% of its mHVR clusters with one putative parent and 47% with the other one; in contrast, the putative parental stocks shared less than 30% of the mHVR clusters between them. Conclusions/significance The results suggest a reductive division, a natural hybridization, biparental inheritance of the minicircles in the hybrid and maxicircle introgression. The models including such phenomena and explaining the relationships between these three clones are discussed., Author summary Chagas disease, an important public health problem in Latin America, is caused by the parasite Trypanosoma cruzi. Despite being a widely studied parasite, several questions on the biology of genetic exchange remain unanswered. Population genomic studies have inferred meiosis in T. cruzi, but this cellular division mechanism has not been observed in laboratory yet. In addition, previous results suggest that mitochondrial DNA (called kDNA) may be inherited from both parents in hybrids. Here, we analyzed a hybrid strain and its potential parents to address the mechanisms of genetic exchange at nuclear and mitochondrial levels. We observed that the hybrid strain had heterozygous patterns and DNA content compatible with a meiosis event. Also, we observed that the evolutionary histories of nuclear DNA and kDNA maxicircles were discordant and that the three strains shared identical DNA sequences. Mitochondrial introgression of maxicircle DNA from one genotype to another may explain this observation. In addition, we demonstrated that the hybrid strain shared kDNA minicircles with both parental strains. Our results suggest that hybridization implied meiosis and biparental inheritance of the kDNA. Further research is required to address such phenomena in detail.

Published
2020

16. Potential association of Trypanosoma cruzi DTUs TcV and TcVI with the digestive form of Chagas disease

Author

S. Nuñez, C.M. Pérez Brandán, María M. Monje-Rumi, Patricio Diosque, Paula G. Ragone, Nicolás Tomasini, María Paola Zago, Noelia Floridia-Yapur, and N. Barrientos

Subjects
0301 basic medicine, Microbiology (medical), Chagas disease, Adult, Male, Adolescent, DTUS, Trypanosoma cruzi, 030106 microbiology, Argentina, Biology, Microbiology, Asymptomatic, law.invention, 03 medical and health sciences, purl.org/becyt/ford/3.3 [https], Young Adult, law, parasitic diseases, Genetics, medicine, Prevalence, Humans, Blood culture, Chagas Disease, Typing, Molecular Biology, Ecology, Evolution, Behavior and Systematics, Polymerase chain reaction, Aged, Aged, 80 and over, ARGENTINA, medicine.diagnostic_test, CHAGAS DISEASE, DNA, Protozoan, Middle Aged, medicine.disease, biology.organism_classification, Virology, 030104 developmental biology, Infectious Diseases, DNA BLOT, CLINICAL PICTURES, purl.org/becyt/ford/3 [https], Female, medicine.symptom, Mixed infection
Abstract

The relationship among genetic diversity of Trypanosoma cruzi and clinical forms of Chagas disease remain elusive. In order to assess the possible association between different T. cruzi Discrete Typing Units (DTUs) and the clinical pictures of the disease, 205 chronic patients from Salta province, Argentina, were analysed. One hundred and twenty-two of these patients were clinically categorized as: cardiac 38.5% (47/122), digestive 15% (18/122), cardio-digestive 16% (20/122) and asymptomatic 30% (37/122). From each patient, blood samples were taken for both, Polymerase Chain Reaction (PCR) targeting kDNA and blood culture analyses. The presence of T. cruzi kDNA was detected in 43% (88/205) of the patients. T. cruzi DTUs were identified in 74% (65/88) of the kDNA positive patients by PCR-hybridization using specific probes. We detected the presence of DTUs TcI, TcII, TcV and TcVI. Single infections (i.e. presence of only one DTU in the sample) were detected in 38.64% of the samples (34/88), while mixed infections were 35.23% (31/88). TcV was the most prevalent DTU (60.3%- 53/88). The association analyses showed, for the first time to the best of our knowledge, that TcV and TcVI were associated with the digestive form of Chagas Disease (Fisher p = .0001). Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Floridia Yapur, Noelia Aldana del Rosario. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Zago, María Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Pérez Brandán, Jimena María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Nuñez, Sonia. Gobierno de la Provincia de Salta. Hospital San Bernardo.; Argentina Fil: Barrientos, Natalia. Gobierno de la Provincia de Salta. Hospital San Bernardo.; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina

Published
2019

17. TcTASV Antigens of

Author

Noelia, Floridia-Yapur, Mercedes, Monje-Rumi, Paula, Ragone, Juan J, Lauthier, Nicolás, Tomasini, Anahí, Alberti D'Amato, Patricio, Diosque, Rubén, Cimino, José F, Gil, Daniel O, Sanchez, Julio R, Nasser, and Valeria, Tekiel

Subjects
Trypanosoma cruzi, Antiprotozoal Agents, Antibodies, Protozoan, Humans, Antigens, Protozoan, Chagas Disease, Enzyme-Linked Immunosorbent Assay, Articles, Child, Sensitivity and Specificity, Biomarkers
Abstract

The discovery and characterization of novel parasite antigens to improve the diagnosis of Trypanosoma cruzi by serological methods and for accurate and rapid follow-up of treatment efficiency are still needed. TcTASV is a T. cruzi–specific multigene family, whose products are expressed on the parasite stages present in the vertebrate host. In a previous work, a mix of antigens from subfamilies TcTASV-A and TcTASV-C (Mix A + C) was sensitive and specific to identify dogs with active infection of high epidemiological relevance. Here, TcTASV-A and TcTASV-C were assayed separately as well as together (Mix A + C) in an ELISA format on human samples. The Mix A + C presented moderate sensitivity (78%) but high diagnostic accuracy with a 100% of specificity, evaluated on healthy, leishmaniasic, and Strongyloides stercoralis infected patients. Moreover, antibody levels of pediatric patients showed—2 years posttreatment—diminished reactivity against the Mix A + C (P < 0.0001), pointing TcTASV antigens as promising tools for treatment follow-up.

Published
2019

18. The TcTASV proteins are novel promising antigens to detect activeTrypanosoma cruziinfection in dogs

Author

Juan José Lauthier, Paula G. Ragone, Jorge Diego Marco, Noelia Floridia-Yapur, Rubén O. Cimino, A. M. Alberti d'Amato, Daniel O. Sánchez, M. Monje Rumi, Nicolás Tomasini, Valeria Tekiel, Paola Andrea Barroso, Julio R. Nasser, and Patricio Diosque

Subjects
0301 basic medicine, Chagas disease, Subfamily, Protein family, Otras Ciencias Biológicas, Trypanosoma cruzi, 030106 microbiology, 030231 tropical medicine, Antibodies, Protozoan, Antigens, Protozoan, Enzyme-Linked Immunosorbent Assay, Sensitivity and Specificity, law.invention, Ciencias Biológicas, Mice, 03 medical and health sciences, Dogs, 0302 clinical medicine, Antigen, law, medicine, Animals, Parasite hosting, Chagas Disease, Dog Diseases, Gene, IMMUNODIAGNOSIS, TCTASV ANTIGENS, biology, TRYPANOSOMA CRUZI, biology.organism_classification, medicine.disease, Virology, DOGS, Infectious Diseases, Recombinant DNA, Animal Science and Zoology, Parasitology, CIENCIAS NATURALES Y EXACTAS
Abstract

In regions where Chagas disease is endemic, canine Trypanosoma cruzi infection is highly correlated with the risk of transmission of the parasite to humans. Herein we evaluated the novel TcTASV protein family (subfamilies A, B, C), differentially expressed in bloodstream trypomastigotes, for the detection of naturally infected dogs. A gene of each TcTASV subfamily was cloned and expressed. Indirect enzyme-linked immunosorbent assays (ELISA) were developed using recombinant antigens individually or mixed together. Our results showed that dogs with active T. cruzi infection differentially reacted against the TcTASV-C subfamily. The use of both TcTASV-C plus TcTASV-A proteins (Mix A+C-ELISA) enhanced the reactivity of sera from dogs with active infection, detecting 94% of the evaluated samples. These findings agree with our previous observations, where the infected animals exhibited a quick anti-TcTASV-C antibody response, coincident with the beginning of parasitaemia, in a murine model of the disease. Results obtained in the present work prove that the Mix A+C-ELISA is a specific, simple and cheap technique to be applied in endemic areas in screening studies. The Mix A+C-ELISA could help to differentially detect canine hosts with active infection and therefore with high impact in the risk of transmission to humans. Fil: Floridia Yapur, Noelia Aldana del Rosario. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biología. Cátedra de Química Biologica; Argentina. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Alberti D'amato, Anahí Maitén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Cimino, R.. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Marco, Jorge Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Barroso, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Sanchez, Daniel Oscar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina Fil: Nasser, Julio Rubén. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Tekiel, Valeria Sonia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina

Published
2016
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19. A Novel Genotype and First Record of Trypanosoma lainsoni in Argentina

Author

Fanny Rusman, M. Mónica Díaz, Rubén M. Barquez, Noelia Floridia-Yapur, Anahí G Díaz, Paula G. Ragone, Nicolás Tomasini, and Patricio Diosque

Subjects
Microbiology (medical), 18S RDNA GENES, 18S rDNA genes, lcsh:Medicine, Leopardus geoffroyi, Article, 18S ribosomal RNA, purl.org/becyt/ford/1 [https], gGAPDH genes, Genus, Calomys spp, morphology, transmission electron microscopy, parasitic diseases, Genotype, GGAPDH GENES, Immunology and Allergy, purl.org/becyt/ford/1.6 [https], Clade, Molecular Biology, Gene, Trypanosoma lainsoni, General Immunology and Microbiology, biology, lcsh:R, biology.organism_classification, TRANSMISSION ELECTRON MICROSCOPY, Infectious Diseases, Evolutionary biology, CALOMYS SPP, TRYPANOSOMA LAINSONI, Trypanosoma, MORPHOLOGY, LEOPARDUS GEOFFROYI, Cricetidae
Abstract

Trypanosomes are a group of parasitic flagellates with medical and veterinary importance. Despite many species having been described in this genus, little is known about many of them. Here, we report a genetic and morphological characterization of trypanosomatids isolated from wild mammals from the Argentine Chaco region. Parasites were morphologically and ultrastructurally characterized by light microscopy and transmission electron microscopy. Additionally, 18s rRNA and gGAPDH genes were sequenced and analyzed using maximum likelihood and Bayesian inference. Morphological characterization showed clear characteristics associated with the Trypanosoma genus. The genetic characterization demonstrates that the studied isolates have identical sequences and a pairwise identity of 99% with Trypanosoma lainsoni, which belongs to the clade of lizards and snakes/rodents and marsupials. To date, this species had only been found in the Amazon region. Our finding represents the second report of T. lainsoni and the first record for the Chaco region. Furthermore, we ultrastructurally described for the first time the species. Finally, the host range of T. lainsoni was expanded (Leopardus geoffroyi, Carenivora, Felidae, and Calomys sp., Rodentia, Cricetidae), showing a wide host range for this species.

Published
2020
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20. MLST Reveals a Separate and Novel Clonal Group for Acidovorax avenae Strains Causing Red Stripe in Sugarcane from Argentina

Author

Cecilia Alejandra Fontana, Paola Daniela Fontana, Nicolás Tomasini, Sergio Miguel Salazar, Valentina Di Pauli, Pier Sandro Cocconcelli, and Graciela Vignolo

Subjects
Otras Ciencias Biológicas, Genotypes, Genotipos, Argentina, Plant Science, Biology, SUGARCANE, purl.org/becyt/ford/1 [https], Saccharum, Ciencias Biológicas, Comamonadaceae, Phylogenetics, Genotype, RED STRIPE, Enfermedades de las Plantas, Genetics, Estría Roja de la Caña de Azúcar, purl.org/becyt/ford/1.6 [https], Phylogeny, Plant Diseases, Genetic diversity, Phylogenetic tree, Acidovorax, Nucleotide Sequence, Nucleic acid sequence, food and beverages, biology.organism_classification, Caña de Azúcar, Genética, Secuencia Nucleotídica, Multilocus sequence typing, Agronomy and Crop Science, ACIDOVORAX AVENAE, CIENCIAS NATURALES Y EXACTAS, MLST, Multilocus Sequence Typing
Abstract

Acidovorax spp. cause a wide range of economically important diseases in monocotyledonous and dicotyledonous plants, including sugarcane, corn, rice, oat, millet, foxtail watermelon, and orchid. In Argentina, the red stripe disease of sugarcane caused by Acidovorax avenae affects 30% of the milling stems with important economic losses. To explore the genetic diversity of this bacterium associated with red stripe in Argentina, multilocus sequence typing (MLST) was applied. This study included 15 local strains isolated from four different sugarcane planting regions and selected after random amplified polymorphic DNA analysis and reference strains of A. citrulli, A. avenae, and A. oryzae to investigate their phylogenetic relationships. MLST analysis resulted in five sequence types among the sugarcane A. avenae strains which constitute a clonal complex, meaning a common and close origin. Sugarcane strains were related to A. avenae from other hosts and distant to A. citrulli. Signals of frequent recombination in several lineages of A. avenae was detected and we observed that A. oryzae is closely related to A. avenae strains. This study provides valuable data in the field of epidemiological and evolutionary investigations of novel clone of A. avenae strains causing sugarcane red stripe. The knowledge of the genetic diversity and strain-host specificity are important to select the genotypes with the best response to the red stripe disease. Fil: Fontana, Paola. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Tucuman-Santiago del Estero. Estación Experimental Agropecuaria Famaillá; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Fontana, Cecilia Alejandra. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Tucuman-Santiago del Estero. Estación Experimental Agropecuaria Famaillá; Argentina Fil: Di Pauli, Valentina. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Tucuman-Santiago del Estero. Estación Experimental Agropecuaria Famaillá; Argentina Fil: Cocconcelli, Pier Sandro. Università Cattolica del Sacro Cuore; Italia Fil: Vignolo, Graciela Margarita. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina Fil: Salazar, Sergio Miguel. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Tucuman-Santiago del Estero. Estación Experimental Agropecuaria Famaillá; Argentina

Published
2018

21. Introgression of the Kinetoplast DNA: An Unusual Evolutionary Journey in Trypanosoma cruzi

Author

Nicolás Tomasini

Subjects
0301 basic medicine, Chagas disease, CIENCIAS MÉDICAS Y DE LA SALUD, PHYLOGENY, 030231 tropical medicine, Ciencias de la Salud, Introgression, Context (language use), 03 medical and health sciences, 0302 clinical medicine, Phylogenetics, parasitic diseases, Genetics, medicine, Trypanosoma cruzi, Genetics (clinical), Salud Ocupacional, biology, Phylogenetic tree, TRYPANOSOMA CRUZI, biology.organism_classification, medicine.disease, EVOLUTION, MITOCHONDRIAL INTROGRESSION, Sexual reproduction, 030104 developmental biology, Evolutionary biology, Kinetoplast, DTU, KINETOPLAST, HYBRIDIZATION
Abstract

Introduction: Phylogenetic relationships between different lineages of Trypanosoma cruzi, the agent of Chagas disease, have been controversial for several years. However, recent phylogenetic and phylogenomic analyses clarified the nuclear relationships among such lineages. However, incongruence between nuclear and kinetoplast DNA phylogenies has emerged as a new challenge. This incongruence implies several events of mitochondrial introgression at evolutionary level. However, the mechanism that gave origin to introgressed lineages is unknown. Here, I will review and discuss how maxicircles of the kinetoplast were horizontally and vertically transferred between different lineages of T. cruzi. Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina

Published
2018
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22. Trypanosoma cruzi diversity in the Gran Chaco: Mixed infections and differential host distribution of TcV and TcVI

Author

Patricio Diosque, Juan José Lauthier, Paula G. Ragone, Nicolás Tomasini, Cecilia Pérez Brandán, María M. Monje-Rumi, Viviana Orellana, Miguel A. Basombrío, Rubén O. Cimino, and Anahí M. Alberti D’Amato

Subjects
Adult, Male, Rural Population, Microbiology (medical), CIENCIAS MÉDICAS Y DE LA SALUD, Adolescent, Genotype, Trypanosoma cruzi, Argentina, Ciencias de la Salud, Biology, Microbiology, Young Adult, Dogs, parasitic diseases, Genetics, Epidemiología, Animals, Humans, Parasite hosting, Parasitología, Chagas Disease, Dog Diseases, Typing, Child, Molecular Biology, Ecology, Evolution, Behavior and Systematics, DOMESTIC CYCLE, ARGENTINA, Coinfection, Host (biology), Hybridization probe, DNA, Protozoan, Middle Aged, biology.organism_classification, Virology, PREVALENCE, PCR, Cross-Sectional Studies, Infectious Diseases, HYBRIDIZATION, Mixed infection
Abstract

The transmission cycles of Trypanosoma cruzi in the Gran Chaco are complex networks involving domestic and wild components, whose interrelationships are not well understood. Knowing the circuit of transmission of the different Discrete Typing Units (DTUs) of T. cruzi in the complex environment of the Chaco region is relevant to understanding how the different components (reservoirs, vectors, ecotopes) interact. In the present study we identified the DTUs infecting humans and dogs in two rural areas of the Gran Chaco in Argentina, using molecular methods which avoid parasite culture. Blood samples of humans and dogs were typified by PCR-DNA blotting and hybridization assays with five specific DNA probes (TcI, TcII, TcIII, TcV and TcVI). PCR analyses were performed on seropositive human and dog samples and showed the presence of T. cruzi DNA in 41.7% (98/235) and 53% (35/66) samples, respectively. The identification of infective DTUs was determined in 83.6% (82/98) and 91.4% (32/35) in human and dog samples, respectively. Single infections (36.7% – 36/98) and a previously not detected high proportion of mixed infections (47.9% – 47/98) were found. In a 15.3% (15/98) of samples the infecting DTU was not identified. Among the single infections TcV was the most prevalent DTU (30.6% – 30/98) in human samples; while TcVI (42.8% – 15/35) showed the highest prevalence in dog samples. TcV/TcVI was the most prevalent mixed infection in humans (32.6% – 32/98); and TcI/TcVI (14.3% – 5/35) in dogs. Significant associations between TcV with humans and TcVI with dogs were detected. For the first time, the presence of TcIII was detected in humans from this region. The occurrence of one human infected whit TcIII (a principally wild DTU) could be suggested the emergence of this, in domestic cycles in the Gran Chaco. Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Perez Brandan, Cecilia Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Alberti D'Amato, Anahi M.. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Cimino, Ruben O.. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biologia. Catedra de Quimica Biologica; Argentina. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Orellana, Viviana. Universidad Nacional de Salta. Facultad de Ciencias de la Salud; Argentina Fil: Basombrio, Miguel Angel Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina

Published
2015
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23. Phylogenomics of Trypanosoma cruzi: Few evidence of TcI/TcII mosaicism in TcIII challenges the hypothesis of an ancient TcI/TcII hybridization

Author

Nicolás Tomasini and Patricio Diosque

Subjects
0301 basic medicine, Microbiology (medical), CIENCIAS MÉDICAS Y DE LA SALUD, Genotype, Genetic Speciation, Trypanosoma cruzi, 030231 tropical medicine, Zoology, Ciencias de la Salud, Biology, PHYLOGENOMICS, Microbiology, 03 medical and health sciences, 0302 clinical medicine, Phylogenetics, Phylogenomics, Genetics, Molecular Biology, Phylogeny, Ecology, Evolution, Behavior and Systematics, Salud Ocupacional, Phylogenetic tree, Mosaicism, Genetic Variation, TRYPANOSOMA CRUZI, biology.organism_classification, 030104 developmental biology, Infectious Diseases, Evolutionary biology, Hybridization, Genetic, Genome, Protozoan, HYBRIDIZATION
Abstract

Phylogenetic relationships among major lineages of Trypanosoma cruzi are still debatable. Particularly, it is controversial the origin of two main lineages: TcIII and TcIV. Some authors proposed that these lineages have been the result of an ancient hybridization between TcI and TcII, and this was one of the most accepted evolutionary models in the scientific community for several years. In the present paper we analyse several genomes of T. cruzi in order to examine if there is evidence supporting that TcIII is an ancient TcI/TcII hybrid. Our results show that TcIII is mainly related to TcI and not to TcII and there is few evidence of mosaicism for TcIII. Our results challenge the hypothesis of the ancient TcI/TcII hybridization. Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina

Published
2017

24. Epidemiological modeling of Trypanosoma cruzi: Low stercorarian transmission and failure of host adaptive immunity explain the frequency of mixed infections in humans

Author

Juan P. Aparicio, Paula G. Ragone, Sébastien Gourbière, Nicolás Tomasini, and Patricio Diosque

Subjects
0301 basic medicine, Mixed infections, Disease, Pathogenesis, Adaptive Immunity, Pathology and Laboratory Medicine, Systems Science, purl.org/becyt/ford/1 [https], 0302 clinical medicine, Agent-Based Modeling, Epidemiology, Genotype, Medicine and Health Sciences, Immune Response, lcsh:QH301-705.5, Protozoans, Mammals, Ecology, Transmission (medicine), Coinfection, Simulation and Modeling, purl.org/becyt/ford/1.2 [https], Acquired immune system, Infectious Diseases, Computational Theory and Mathematics, Modeling and Simulation, Host-Pathogen Interactions, Physical Sciences, Vertebrates, Algorithms, CIENCIAS NATURALES Y EXACTAS, Research Article, Neglected Tropical Diseases, Chagas disease, medicine.medical_specialty, Trypanosoma, Computer and Information Sciences, Trypanosoma cruzi, 030231 tropical medicine, Immunology, Humans transmission, Biology, Research and Analysis Methods, Models, Biological, Epidemiological modeling, 03 medical and health sciences, Cellular and Molecular Neuroscience, Immune system, Dogs, Genetics, medicine, Parasitic Diseases, Animals, Humans, Chagas Disease, Molecular Biology, Ecology, Evolution, Behavior and Systematics, Protozoan Infections, Organisms, Computational Biology, Biology and Life Sciences, biology.organism_classification, medicine.disease, Tropical Diseases, Virology, Parasitic Protozoans, Insect Vectors, Ciencias de la Computación, Vector-Borne Diseases, 030104 developmental biology, lcsh:Biology (General), Ciencias de la Computación e Información, Amniotes, Mathematics
Abstract

People living in areas with active vector-borne transmission of Chagas disease have multiple contacts with its causative agent, Trypanosoma cruzi. Reinfections by T. cruzi are possible at least in animal models leading to lower or even hardly detectable parasitaemia. In humans, although reinfections are thought to have major public health implications by increasing the risk of chronic manifestations of the disease, there is little quantitative knowledge about their frequency and the timing of parasite re-inoculation in the course of the disease. Here, we implemented stochastic agent-based models i) to estimate the rate of re-inoculation in humans and ii) to assess how frequent are reinfections during the acute and chronic stages of the disease according to alternative hypotheses on the adaptive immune response following a primary infection. By using a hybrid genetic algorithm, the models were fitted to epidemiological data of Argentinean rural villages where mixed infections by different genotypes of T. cruzi reach 56% in humans. To explain this percentage, the best model predicted 0.032 (0.008–0.042) annual reinfections per individual with 98.4% of them occurring in the chronic phase. In addition, the parasite escapes to the adaptive immune response mounted after the primary infection in at least 20% of the events of re-inoculation. With these low annual rates, the risks of reinfection during the typically long chronic stage of the disease stand around 14% (4%-18%) and 60% (21%-70%) after 5 and 30 years, with most individuals being re-infected 1–3 times overall. These low rates are better explained by the weak efficiency of the stercorarian mode of transmission than a highly efficient adaptive immune response. Those estimates are of particular interest for vaccine development and for our understanding of the higher risk of chronic disease manifestations suffered by infected people living in endemic areas., Author summary Chagas disease is caused by a unicellular parasite mainly transmitted by blood-feeding bugs. The disease starts with an acute phase when the parasite multiplies in tissues. The immune system is then activated and almost clears the parasites before the host enters the chronic phase of the disease. As hosts remain exposed to the vectors, secondary contacts may result in re-inoculation with the parasite. The re-inoculated parasites then have to face an adaptive immune response that may prevent reinfection. There is little quantitative knowledge about the frequency of such re-inoculation and the efficiency of the immune system to prevent them. Since reinfections can lead to people carrying two or more strains of the parasite, we can potentially infer on the frequency of reinfection from the detection of such mixed infections. We fitted agent-based models to field data to estimate the frequency of reinfection in humans and its timing after primary infection. Our models predicted that most reinfections occur during the chronic phase as the adaptive immune response fails to prevent them in 20% of re-inoculations. Such a rate of failure is particularly important as reinfections are thought to increase the risk of heart damage in the chronic phase of the disease.

Published
2017

25. Introgression of the Kinetoplast DNA: An Unusual Evolutionary Journey in

Author

Nicolás, Tomasini

Subjects
Kinetoplast, Evolution, Trypanosoma cruzi, parasitic diseases, Mitochondrial introgression, DTU, Hybridization, Article, Phylogeny
Abstract

Introduction: Phylogenetic relationships between different lineages of Trypanosoma cruzi, the agent of Chagas disease, have been controversial for several years. However, recent phylogenetic and phylogenomic analyses clarified the nuclear relationships among such lineages. However, incongruence between nuclear and kinetoplast DNA phylogenies has emerged as a new challenge. This incongruence implies several events of mitochondrial introgression at evolutionary level. However, the mechanism that gave origin to introgressed lineages is unknown. Here, I will review and discuss how maxicircles of the kinetoplast were horizontally and vertically transferred between different lineages of T. cruzi. Conclusion: Finally, I will discuss what we know - and what we don't - about the kDNA transference and inheritance in the context of sexual reproduction in this parasite.

Published
2017

26. Biological behavior of different Trypanosoma cruzi isolates circulating in an endemic area for Chagas disease in the Gran Chaco region of Argentina

Author

Angel M. Padilla, Nélida Marcela Romero, Cecilia Pérez Brandán, Mercedes Monje Rumi, Miguel A. Basombrío, Patricio Diosque, Nicolás Tomasini, Anahí M. Alberti D’Amato, Marcela Portelli, Rubén O. Cimino, Juan José Lauthier, Julio R. Nasser, and Paula G. Ragone

Subjects
Chagas disease, Male, Biological Properties, CIENCIAS MÉDICAS Y DE LA SALUD, Endemic Diseases, Veterinary (miscellaneous), Trypanosoma cruzi, 030231 tropical medicine, Argentina, Antibodies, Protozoan, Biology, Natural Isolates, Serology, 03 medical and health sciences, Mice, 0302 clinical medicine, Genetic variation, medicine, Parasite hosting, Animals, Humans, Chagas Disease, Typing, Muscle, Skeletal, 030304 developmental biology, Trypanosoma Cruzi, 0303 health sciences, Myocardium, Genetic Variation, Heart, DNA, Protozoan, Bioquímica y Biología Molecular, medicine.disease, biology.organism_classification, Virology, DNA Fingerprinting, 3. Good health, Enzymes, Mice, Inbred C57BL, Disease Models, Animal, Medicina Básica, Infectious Diseases, Blood, DNA profiling, Insect Science, Multilocus sequence typing, Parasitology
Abstract

The biological behavior of the different . Trypanosoma cruzi strains is still unclear and the importance of exploring the relevance of these differences in natural isolates is of great significance. Herein we describe the biological behavior of four . T. cruzi isolates circulating sympatrically in a restricted geographic area in Argentina endemic for Chagas Disease. These isolates were characterized as belonging to the Discrete Typing Units (DTUs) TcI, TcIII, TcV and TcVI as shown by Multilocus Enzyme Electrophoresis and Multilocus Sequence Typing. In order to study the natural behavior of the different isolates and to preserve their natural properties, we developed a vector transmission model that allows their maintenance in the laboratory. The model consisted of serial passages of these parasites between insect vectors and mice. Vector-derived parasite forms were then inoculated in C57BL/6J mice and number of parasite in peripheral blood, serological response and histological damage in acute and chronic phases of the infection were measured. Parasites from DTUs TcI, TcIII and TcVI were detected by direct fresh blood examination, while TcV parasites could only be detected by Polimerase Chain Reaction. No significant difference in the anti-. T. cruzi antibody response was found during the chronic phase of infection, except for mice infected with TcV parasites where no antibodies could be detected. Histological sections showed that TcI isolate produced more damage in skeletal muscle while TcVI induced more inflammation in the heart. This work shows differential biological behavior among different parasite isolates obtained from the same cycle of transmission, permitting the opportunity to formulate future hypotheses of clinical and epidemiological importance. Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Pérez Brandan, Cecilia María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Padilla, Angel M.. University of Georgia; Estados Unidos Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Alberti D'amato, Anahí Maitén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Cimino, Rubén Oscar. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biología; Argentina Fil: Romero, Nélida M.. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biología; Argentina Fil: Portelli, Marcela. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biología; Argentina Fil: Nasser, Julio Rubén. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biología; Argentina Fil: Basombrío, Miguel Ángel Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina

Published
2012
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27. Interest and limitations of Spliced Leader Intergenic Region sequences for analyzing Trypanosoma cruzi I phylogenetic diversity in the Argentinean Chaco

Author

Maria Mercedes Monje Rumi, Nicolás Tomasini, Alejandra Falla, Cecilia Pérez Brandán, Anahí A. Alberti D’Amato, Alejandro G. Schijman, Felipe Guhl, Paula G. Ragone, Christian Barnabé, Juan José Lauthier, Michel Tibayrenc, Patricio Diosque, Carolina Cura, Claudia Herrera, and Miguel A. Basombrío

Subjects
RNA, Spliced Leader, Microbiology (medical), Genotype, Sequence analysis, Trypanosoma cruzi, Molecular Sequence Data, 030231 tropical medicine, Argentina, Biology, Polymerase Chain Reaction, Microbiology, 03 medical and health sciences, 0302 clinical medicine, Intergenic region, Phylogenetics, Genetics, Humans, Chagas Disease, Gene conversion, Molecular Biology, Phylogeny, Ecology, Evolution, Behavior and Systematics, 030304 developmental biology, 0303 health sciences, Geography, Molecular epidemiology, Haplotype, Genetic Variation, Bayes Theorem, Sequence Analysis, DNA, DNA, Protozoan, Phylogenetic diversity, Infectious Diseases, Haplotypes, Microsatellite, DNA, Intergenic, Microsatellite Repeats, Multilocus Sequence Typing
Abstract

Internal and geographical clustering within Trypanosoma cruzi I (TcI) has been recently revealed by using Multilocus Microsatellite Typing and sequencing of the Spliced-Leader Intergenic Region (SL-IR). In the present work, 14 isolates and 11 laboratory-cloned stocks obtained from a geographically restricted area in Chaco Province, Argentina, were analyzed by PCR and sequencing of SL-IR. We were able to differentiate 8 different genotypes that clustered into 4 groups. One of these groups was classified within the formerly described haplotype A and another one within the recently described SL-IR group E. Both were phylogenetically well-supported. In contrast, none of the stocks from the Chaco province were grouped within the cluster previously named haplotype D despite the fact that they shared a similar microsatellite motif in the SL-IR. No evidence of recombination or gene conversion within these stocks was found. On the other hand, multiple ambiguous alignments in the microsatellite region of SL-IR, affecting the tree topology and relationships among groups were detected. Finally, since there are multiple copies of the SL-IR, and they are arranged in tandem, we discuss how molecular processes affecting this kind of sequences could mislead phylogenetic inference.

Published
2011
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28. Evolution of Trypanosoma cruzi: clarifying hybridisations, mitochondrial introgressions and phylogenetic relationships between major lineages

Author

Patricio Diosque and Nicolás Tomasini

Subjects
Microbiology (medical), Chagas disease, lcsh:Arctic medicine. Tropical medicine, discrete typing units, Genotype, lcsh:RC955-962, PHYLOGENY, Trypanosoma cruzi, lcsh:QR1-502, Introgression, Biology, phylogeny, lcsh:Microbiology, purl.org/becyt/ford/1 [https], Ciencias Biológicas, Monophyly, Biología Celular, Microbiología, Phylogenetics, Genetic variation, Clade, purl.org/becyt/ford/1.6 [https], Genetics, Trypanosomatidae, Phylogenetic tree, DISCRETE TYPING UNITS, Genetic Variation, HYBRIDISATION, Sequence Analysis, DNA, Articles, TRYPANOSOMA CRUZI, DNA, Protozoan, Biological Evolution, EVOLUTION, Mitochondria, Sister group, parasite, Hybridization, Genetic, CIENCIAS NATURALES Y EXACTAS
Abstract

Different models of evolution of Trypanosoma cruzi have been proposed. These models suggest scarce events of genetic exchange in the evolutionary history of this parasite. In addition, the debate focused on the existence of one or two hybridization events during the evolution of T. cruzi lineages. Here we reviewed literature and analyzed available sequence data in order to clarify the phylogenetic relationships among lineages. We observed that TcI, TcIII and TcIV form a monophyletic group and consequently TcIII and TcIV are not, as previously suggested, TcI-TcII hybrids. Particularly, TcI and TcIII are sister groups that diverged around the same time that a widely distributed TcIV was splitted into two clades (TcIVS and TcIVN). In addition, we collected evidence that TcIII received the TcIVS kDNA by introgression at several opportunities. Different demographic hypotheses (surfing and asymmetrical introgression) may explain the origin and expansion of the TcIII group. Considering these hypotheses, genetic exchange should have been relatively frequent between TcIII and TcIVS in the geographic area in which their distributions were superimposed. In addition, our results support the hypothesis that two independent hybridization events gave origin to TcV and TcVI. Consequently, TcIVS kDNA was first transferred to TcIII and later to TcV and TcVI in TcII/TcIII hybridization events. Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina

Published
2015

29. Multilocus sequence typing approach for a broader range of species of Leishmania genus: describing parasite diversity in Argentina

Author

Carlos Lorenzo Hoyos, Masataka Korenaga, S. Pamela Cajal, Nicolás Tomasini, Juan José Lauthier, Fabricio M. Locatelli, Marisa Juarez, Paola Andrea Barroso, Paula Ruybal, Jorge Diego Marco, J. Octavio Estevez, Yoshihisa Hashiguchi, Julio R. Nasser, and Cecilia Nevot

Subjects
Microbiology (medical), Population, Argentina, Biology, Microbiology, Ciencias Biológicas, Genética y Herencia, Dogs, Cutaneous leishmaniasis, parasitic diseases, Genetics, medicine, Animals, Humans, Genetic variability, education, Molecular Biology, Leishmaniasis, Ecology, Evolution, Behavior and Systematics, Phylogeny, Leishmania, education.field_of_study, Genetic diversity, ARGENTINA, Ecology, DNA, Protozoan, medicine.disease, biology.organism_classification, Infectious Diseases, Visceral leishmaniasis, Haplotypes, Evolutionary biology, Multilocus sequence typing, LEISHMANIA, CIENCIAS NATURALES Y EXACTAS, MLST, LEISHMANIASIS, Multilocus Sequence Typing
Abstract

Leishmaniasis is a vector-borne protozoan infection affecting over 350 million people around the world. In Argentina cutaneous leishmaniasis is endemic in nine provinces and visceral leishmaniasis is spreading from autochthonous transmission foci in seven provinces. However, there is limited information about the diversity of the parasite in this country. Implementation of molecular strategies for parasite typing, particularly multilocus sequence typing (MLST), represents an improved approach for genetic variability and population dynamics analyses. We selected six loci as candidates implemented in reference strains and Argentinean isolates. Phylogenetic analysis showed high correlation with taxonomic classification of the parasite. Autochthonous Leishmania (Viannia) braziliensis showed higher genetic diversity than L. (Leishmania) infantum but low support was obtained for intra-L. braziliensis complex variants suggesting the need of new loci that contribute to phylogenetic resolution for an improved MLST or nested-MLST scheme. This study represents the first characterization of genetic variability of Leishmania spp. in Argentina. Fil: Marco, Jorge Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Barroso, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Locatelli, Fabricio M.. Kochi University. Kochi Medical School. Departament of Parasitology; Japón Fil: Cajal, S. Pamela. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Hoyos, Carlos Lorenzo. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Nevot, Cecilia. Kochi University. Kochi Medical School. Departament of Parasitology; Japón Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Juarez, Marisa. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Estevez, J. Octavio. Veterinaria Del Oeste; Argentina Fil: Korenaga, Masataka. Kochi University. Kochi Medical School. Departament of Parasitology; Japón Fil: Nasser, Julio. Universidad Nacional de Salta. Sede Regional Oran. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Hashiguchi, Yoshihisa. Kochi University. Kochi Medical School. Dept.of Parasitology; Japón. Universidad Central del Ecuador y Proyecto Prometeo. Centro de Biomedicina; Ecuador Fil: Ruybal, Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones En Microbiología y Parasitología Médica; Argentina

Published
2014

30. Evaluation of recombinant antigens of Trypanosoma cruzi to diagnose infection in naturally infected dogs from Chaco region, Argentina

Author

Noelia Floridia-Yapur, M. Monje Rumi, Julio R. Nasser, Nicolás Tomasini, I. Lopez-Quiroga, Paula G. Ragone, Iván Sergio Marcipar, A. M. Alberti d'Amato, Patricio Diosque, A. F. Vega-Benedetti, Rubén O. Cimino, and Juan José Lauthier

Subjects
dogs, Trypanosoma cruzi, Immunology, Argentina, Positive control, Antigens, Protozoan, Enzyme-Linked Immunosorbent Assay, Biology, Sensitivity and Specificity, law.invention, Kappa index, Dogs, Antigen, law, parasitic diseases, Animals, Chagas Disease, Dog Diseases, Antigens, Area under the curve, recombinant antigens, purl.org/becyt/ford/3.1 [https], biology.organism_classification, Virology, Recombinant Proteins, Recombinant DNA, Parasitology, ELISA, purl.org/becyt/ford/3 [https], Xenodiagnosis
Abstract

Dogs are considered the main mammal reservoir of Trypanosoma cruzi in domiciliary environments. Consequently, accurate detection of T. cruzi infection in canine populations is epidemiologically relevant. Here we analyzed the utility of the T. cruzi recombinant antigens FRA, SAPA, CP1, Ag1 and a SAPA/TSSA VI mixture, in an ELISA format. We used a positive control group of sera obtained from 38 dogs from the Chaco region in Argentina with positive Homogenate-ELISA reaction, all of them also positive by xenodiagnosis and/or PCR. The negative group included 19 dogs from a non-endemic area. Sensitivity, specificity, Area Under the Curve (AUC) of the Receiver-Operating Charactheristic (ROC) curve and Kappa index were obtained in order to compare the diagnostic efficiency of the tests. The SAPA/TSSA VI had the highest performance, with a sensitivity of 94.7% and an AUC ROC of 0.99 that indicates high accuracy. Among individual antigens, SAPA-ELISA yielded the highest sensitivity (86.8%) and AUC ROC (0.96), whereas FRA-ELISA was the least efficient test (sensitivity=36.8%; AUC ROC=0.53). Our results showed that the use of SAPA/TSSA VI in ELISA assays could be a useful tool to study dogs naturally infected with T. cruzi in endemic areas. Fil: Floridia Yapur, Noelia Aldana del Rosario. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Universidad Nacional de Salta. Facultad de Ciencias Naturales. Escuela de Biología. Cátedra de Química Biológica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; Argentina Fil: Vega Benedetti, Ana Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Alberti D'amato, Anahí Maitén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: López Quiroga, Inés Raquel. Universidad Nacional de Salta. Facultad de Ciencias Naturales. Escuela de Biología. Cátedra de Química Biológica; Argentina. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina Fil: Marcipar, Iván Sergio. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Tecnología Inmunológica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; Argentina Fil: Nasser, Julio Rubén. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Universidad Nacional de Salta. Facultad de Ciencias Naturales. Escuela de Biología. Cátedra de Química Biológica; Argentina Fil: Cimino, Rubén Oscar. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; Argentina. Universidad Nacional de Salta. Facultad de Ciencias Naturales. Escuela de Biología. Cátedra de Química Biológica; Argentina

Published
2014
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31. Optimized multilocus sequence typing (MLST) scheme for Trypanosoma cruzi

Author

Nicolás Tomasini, Michel Tibayrenc, Paula G. Ragone, Cecilia Pérez Brandán, Maria Mercedes Monje Rumi, Juan José Lauthier, Christian Barnabé, Martin S. Llewellyn, Patricio Diosque, Anahí Maitén Alberti-D'Amato, Michael A. Miles, Louisa A. Messenger, Matthew Yeo, and Michael D. Lewis

Subjects
CIENCIAS MÉDICAS Y DE LA SALUD, lcsh:Arctic medicine. Tropical medicine, Genotype, Sequence analysis, lcsh:RC955-962, Trypanosoma cruzi, 030231 tropical medicine, SCHEME OF OPTIMIZATION, Ciencias de la Salud, Biology, Microbiology, 03 medical and health sciences, purl.org/becyt/ford/3.3 [https], 0302 clinical medicine, Medicine and Health Sciences, Parasitic Diseases, Chagas Disease, Typing, Parasite Evolution, 030304 developmental biology, Genetics, Whole genome sequencing, 0303 health sciences, Genetic diversity, Protozoan Infections, Strain (biology), lcsh:Public aspects of medicine, Public Health, Environmental and Occupational Health, Biology and Life Sciences, lcsh:RA1-1270, TRYPANOSOMA CRUZI, Amplicon Size, 3. Good health, Enfermedades Infecciosas, Infectious Diseases, Multilocus sequence typing, GENETIC DIVERSITY, purl.org/becyt/ford/3 [https], Parasitology, MULTILOCUS SEQUENCE TYPING, Research Article, Multilocus Sequence Typing
Abstract

Trypanosoma cruzi, the aetiological agent of Chagas disease possess extensive genetic diversity. This has led to the development of a plethora of molecular typing methods for the identification of both the known major genetic lineages and for more fine scale characterization of different multilocus genotypes within these major lineages. Whole genome sequencing applied to large sample sizes is not currently viable and multilocus enzyme electrophoresis, the previous gold standard for T. cruzi typing, is laborious and time consuming. In the present work, we present an optimized Multilocus Sequence Typing (MLST) scheme, based on the combined analysis of two recently proposed MLST approaches. Here, thirteen concatenated gene fragments were applied to a panel of T. cruzi reference strains encompassing all known genetic lineages. Concatenation of 13 fragments allowed assignment of all strains to the predicted Discrete Typing Units (DTUs), or near-clades, with the exception of one strain that was an outlier for TcV, due to apparent loss of heterozygosity in one fragment. Monophyly for all DTUs, along with robust bootstrap support, was restored when this fragment was subsequently excluded from the analysis. All possible combinations of loci were assessed against predefined criteria with the objective of selecting the most appropriate combination of between two and twelve fragments, for an optimized MLST scheme. The optimum combination consisted of 7 loci and discriminated between all reference strains in the panel, with the majority supported by robust bootstrap values. Additionally, a reduced panel of just 4 gene fragments displayed high bootstrap values for DTU assignment and discriminated 21 out of 25 genotypes. We propose that the seven-fragment MLST scheme could be used as a gold standard for T. cruzi typing, against which other typing approaches, particularly single locus approaches or systematic PCR assays based on amplicon size, could be compared., Author Summary The single-celled parasite Trypanosoma cruzi occurs in mammals and insect vectors in the Americas. When transmitted to humans it causes Chagas disease (American trypanosomiasis) a major public health problem. T. cruzi is genetically diverse and currently split into six groups, known as TcI to TcVI. Multilocus sequence typing (MLST) is a method used for studying the population structure and diversity of pathogens and involves sequencing DNA of several different genes and comparing the sequences between isolates. Here, we assess 13 T. cruzi genes and select the best combination for diversity studies. Outputs reveal that a combination of 7 genes can be used for both lineage assignment and high resolution studies of genetic diversity, and a reduced combination of four loci for lineage assignment. Application of MLST for assigning field isolates of T. cruzi to genetic groups and for detailed investigation of diversity provides a valuable approach to understanding the taxonomy, population structure, genetics, ecology and epidemiology of this important human pathogen.

Published
2014

32. Preponderant clonal evolution of Trypanosoma cruzi I from Argentinean Chaco revealed by Multilocus Sequence Typing (MLST)

Author

Paula G. Ragone, Nicolás Tomasini, Juan José Lauthier, Patricio Diosque, Anahí M. Alberti D’Amato, Miguel A. Basombrío, Maria Mercedes Monje Rumi, and Cecilia Pérez Brandán

Subjects
RNA, Spliced Leader, Microbiology (medical), Lineage (genetic), CIENCIAS MÉDICAS Y DE LA SALUD, Genotype, Trypanosoma cruzi, Molecular Sequence Data, Argentina, Ciencias de la Salud, Microbiology, Linkage Disequilibrium, Evolution, Molecular, Intergenic region, Genetics, Humans, Chagas Disease, Molecular Biology, Alleles, Phylogeny, Ecology, Evolution, Behavior and Systematics, Polymorphism, Genetic, biology, Genetic Variation, biology.organism_classification, INTERCAMBIO GENÉTICO, RAPD, Enfermedades Infecciosas, Infectious Diseases, Genetic Loci, Genetic marker, TCI, Genetic structure, Multilocus sequence typing, DNA, Intergenic, ENFERMEDAD DE CHAGAS, Multilocus Sequence Typing, MLST
Abstract

Trypanosoma cruzi has been historically classified as a species with preponderant clonal evolution (PCE). However, with the advent of highly polymorphic markers and studies at geographically reduced scales,the PCE in T. cruzi was challenged. In fact, some studies have suggested that recombination in T. cruzi lineage I (TcI) is much more frequent than previously believed. Further analyses of TcI populations from different geographical regions of Latin America are needed to examine this hypothesis. In the present study,we contribute to this topic by analyzing the population structure of TcI from a restricted geographical area in the Chaco region, Argentina. We analyzed TcI isolates from different hosts and vectors using a Multilocus Sequence Typing (MLST) approach. These isolates were previously characterized by sequencing the spliced leader intergenic region (SL-IR). Low levels of incongruence and well-supported clusters for MLST dataset were obtained from the analyses. Moreover, high linkage disequilibrium was found and five repeated and overrepresented genotypes were detected. In addition, a good correspondence between SL-IR and MLST was observed which is expected under PCE. However, recombination is not ruled out because five out of 28 pairs of loci were incompatible with strict clonality and one possible genetic exchange event was detected. Overall, our results represent evidence of PCE in TcI from the study area.Finally, considering our findings we discuss the scenario for the genetic structure of TcI. Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Alberti D'amato, Anahi M.. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Perez Brandan, Cecilia Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Basombrio, Miguel Angel Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina

Published
2014

33. Reassessment of MLST schemes for Leptospira spp. typing worldwide

Author

Juan José Lauthier, Bibiana Brihuega, Nicolás Tomasini, Paula Ruybal, Vanina Delia Varni, Karina Cynthia Caimi, and Ariel Koval

Subjects
DNA, Bacterial, Microbiology (medical), CIENCIAS MÉDICAS Y DE LA SALUD, Ciencias de la Salud, Locus (genetics), Biology, Microbiology, Ciencias Biológicas, Biología Celular, Microbiología, Leptospira, Zoonoses, Genotype, Genetics, medicine, Animals, Humans, Epidemiología, Leptospirosis, Reassessment, Typing, Molecular Biology, Ecology, Evolution, Behavior and Systematics, Phylogenetic tree, Zoonosis, typing, Neglected Diseases, bacterial infections and mycoses, medicine.disease, biology.organism_classification, Enfermedades Infecciosas, Infectious Diseases, Multilocus sequence typing, CIENCIAS NATURALES Y EXACTAS, Multilocus Sequence Typing, MLST
Abstract

Leptospirosis is a neglected zoonosis of global importance. Several multilocus sequence typing (MLST) methods have been developed for Leptospira spp., the causative agent of leptospirosis. In this study we reassessed the most commonly used MLST schemes in a set of worldwide isolates, in order to select the loci that achieve the maximum power of discrimination for typing Leptospira spp. Global eBURST algorithm was used to detect clonal complexes among STs and phylogenetic relationships among concatenated and individual sequences were inferred through maximum likelihood (ML) analysis. The evaluation of 12 loci combined to type a subset of strains rendered 57 different STs. Seven of these loci were selected into a final scheme upon studying the number of alleles and polymorphisms, the typing efficiency, the discriminatory power and the ratio dN/dS per nucleotide site for each locus. This new 7-locus scheme was applied to a wider collection of worldwide strains. The ML tree constructed from concatenated sequences of the 7 loci identified 6 major clusters corresponding to 6 Leptospira species. Global eBURST established 8 CCs, which showed that genotypes were clearly related by geographic origin and host. ST52 and ST47, represented mostly by Argentinian isolates, grouped the higher number of isolates. These isolates were serotyped as serogroups Pomona and Icterohaemorrhagiae, showing a unidirectional correlation in which the isolates with the same ST belong to the same serogroup. In summary, this scheme combines the best loci from the most widely used MLST schemes for Leptospira spp. and supports worldwide strains classification. The Argentinian isolates exhibited congruence between allelic profile and serogroup, providing an alternative to serological methods. Fil: Varni, Vanina Delia. Instituto Nacional de Tecnología Agropecuaria; Argentina Fil: Ruybal, Paula. Instituto Nacional de Tecnología Agropecuaria; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Brihuega, Bibiana. Instituto Nacional de Tecnología Agropecuaria; Argentina Fil: Koval, Ariel. Biogenesis Bago S.a; Argentina Fil: Caimi, Karina Cynthia. Instituto Nacional de Tecnología Agropecuaria; Argentina

Published
2014

34. MLSTest: novel software for multi-locus sequence data analysis in eukaryotic organisms

Author

Nicolás Tomasini, Martin S. Llewellyn, Patricio Diosque, and Juan José Lauthier

Subjects
Microbiology (medical), CIENCIAS MÉDICAS Y DE LA SALUD, Trypanosoma cruzi, MLSTEST, Ciencias de la Salud, Locus (genetics), Candida glabrata, Computational biology, SOFTWARE, Biology, Neisseria meningitidis, Microbiology, DNA sequencing, INCONGRUENCE, Software, Genetics, Epidemiología, Typing, Cluster analysis, Molecular Biology, Genotyping, Ecology, Evolution, Behavior and Systematics, business.industry, Aspergillus fumigatus, CONCATENATION, Eukaryota, Genomics, Diploidy, Haemophilus influenzae, Infectious Diseases, Multilocus sequence typing, MULTILOCUS SEQUENCE TYPING, business, SCHEME OPTIMIZATION, Algorithms, Type I and type II errors, MLST, Multilocus Sequence Typing
Abstract

Multi-locus sequence typing (MLST) is a frequently used genotyping method whose goal is the unambiguous assignment of microorganisms to genetic clusters. MLST typically involves analysis of DNA sequence results generated from several house-keeping gene loci. MLST remains the gold standard for molecular typing of many bacterial pathogens. Eukaryotic pathogens have also been the subject of MLST, however, few tools are available to deal with diploid sequence data. Here we present novel software for MLST data analysis tailored towards diploid Eukaryotes: MLSTest. This software meets various methods used in MLST and introduces some novel methodologies for the evaluation of the data set. In addition to construction of allelic profiles and basic clustering analysis, the MLSTest looks for network structures that suggest genetic exchange in BURST graphs. Additionally, it uses several simple methods for tree construction with the advantage of managing heterozygous or three-state sites. Additionally, the software analyses whether concatenation of fragments from different genes is suitable for the data set using different tests (bionj-incongruence length difference test, Templeton test). It evaluates how the incongruence is distributed across the tree using a variation of the localized incongruence length difference test based on a modified neighbour joining algorithm. We tested the last method in simulated datasets. We showed that is conservative (adequate type I error rate) and moderately to highly powerful as well as useful to localize incongruences in two bacterial and two eukaryotic MLST datasets. MLSTest was also designed for developing MLST schemes. It thus has tools to optimize locus combinations and to reduce the number of targets required for typing. MLSTest also analyses whether the discriminatory power of the typing scheme is increased by including more loci. We evaluated the software over simulated and real datasets from bacterial and eukaryotic microorganisms. The software is freely available at http://www.ipe.unsa.edu.ar/software. Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Llewellyn, Martin S.. University of Wales. School of Biological Sciences; Reino Unido Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina

Published
2013

35. Benznidazole treatment in chronic children infected with Trypanosoma cruzi: serological and molecular follow-up of patients and identification of Discrete Typing Units

Author

Julio R. Nasser, Rubén O. Cimino, Viviana Orellana, Nicolás Tomasini, Miguel A. Basombrío, Juan José Lauthier, José Fernando Gil, C.D. Lacunza, Patricio Diosque, A. M. Alberti d'Amato, C.M. Pérez Brandán, M. Monje Rumi, and Paula G. Ragone

Subjects
Chagas disease, Male, Rural Population, Adolescent, Genotype, Veterinary (miscellaneous), Trypanosoma cruzi, Antiprotozoal Agents, Argentina, Antibodies, Protozoan, Serology, medicine, Humans, Chagas Disease, Typing, Seroconversion, Child, biology, DNA, Kinetoplast, Nucleic Acid Hybridization, DNA, Protozoan, biology.organism_classification, medicine.disease, Virology, Hypervariable region, Infectious Diseases, Treatment Outcome, Benznidazole, Nitroimidazoles, Insect Science, Child, Preschool, Chronic Disease, Parasitology, Female, medicine.drug, Follow-Up Studies
Abstract

A total of 221 children from two rural settlements in Northeast Argentina were examined for T. cruzi infection. Blood samples were taken for serology tests and PCR assays. In addition, T. cruzi Discrete Typing Units (DTUs) were determined by hybridization with specific DNA probes of the minicircle hypervariable regions (mHVR). Serological results indicated that 26% (57/215) were reactive against T. cruzi antigens. PCR analyses were performed on seropositive samples showing presence of parasite DNA in 31 out of 53 samples (58.5%). All seropositive children underwent specific chemotherapy with Benznidazole (5mg/kg/day) for a period of two months and were monitored two and five years after treatment. Overall the treatment was well tolerated and low side effects were observed. Serological conversion was observed at two years post -treatment in one child form Pampa Avila and at five years in two children from Tres Estacas. However, at the end of the follow-up period, T. cruzi DNA could not be detected by PCR in samples from treated children, except in two cases. In addition, the results of hybridizations with specific DNA probes showed that DTU TcV was detected in 68% (21/31), TcVI in 7% (2/31) and TcV/VI in 3% (1/31) of the samples. Altogether, results of the follow-up of treated children showed a low rate of seroconversion; however trend toward seroconversion was evident at five years post-treatment. On the other hand, detection of T. cruzi DNA by PCR significantly decreased after Benznidazole treatment. The existence of data regarding serological and molecular follow-ups from controlled studies in the Chaco Region will be important for future treatment efforts against T. cruzi infection in this region. The results obtained in the present study represent a contribution in this regard.

Published
2012

36. Controlling cytoplasmic c-Fos controls tumor growth in the peripheral and central nervous system

Author

Nicolás Tomasini, Beatriz L. Caputto, David C. Silvestre, Germán A. Gil, and D. F. Bussolino

Subjects
Central Nervous System, Central nervous system, Cell, Endoplasmic Reticulum, Biochemistry, c-Fos, PC12 Cells, Central Nervous System Neoplasms, Cellular and Molecular Neuroscience, Mice, Peripheral Nervous System Neoplasms, Cell Line, Tumor, medicine, Animals, Humans, Transcription factor, Phospholipids, Cell Proliferation, biology, Endoplasmic reticulum, General Medicine, Oligonucleotides, Antisense, Cell biology, Rats, medicine.anatomical_structure, Cytoplasm, Tumor progression, Membrane biogenesis, biology.protein, Proto-Oncogene Proteins c-fos
Abstract

Some 20 years ago c-Fos was identified as a member of the AP-1 family of inducible transcription factors (Angel and Karin in Biochim Biophys Acta 1072:129–157, 1991). More recently, an additional activity was described for this protein: it associates to the endoplasmic reticulum and activates the biosynthesis of phospholipids (Bussolino et al. in FASEB J 15:556–558, 2001), (Gil et al. in Mol Biol Cell 15:1881–1894, 2004), the quantitatively most important components of cellular membranes. This latter activity of c-Fos determines the rate of membrane genesis and consequently of growth in differentiating PC12 cells (Gil et al. in Mol Biol Cell 15:1881–1894, 2004). In addition, it has been shown that c-Fos is over-expressed both in PNS and CNS tumors (Silvestre et al. in PLoS One 5(3):e9544, 2010). Herein, it is shown that c-Fos-activated phospholipid synthesis is required to support membrane genesis during the exacerbated growth characteristic of brain tumor cells. Specifically blocking c-Fos-activated phospholipid synthesis significantly reduces proliferation of tumor cells in culture. Blocking c-Fos expression also prevents tumor progression in mice intra-cranially xeno-grafted human brain tumor cells. In NPcis mice, an animal model of the human disease Neurofibromatosis Type I (Cichowski and Jacks in Cell 104:593–604, 2001), animals spontaneously develop tumors of the PNS and the CNS, provided they express c-Fos (Silvestre et al. in PLoS One 5(3):e9544, 2010). Treatment of PNS tumors with an antisense oligonucleotide that specifically blocks c-Fos expression also blocks tumor growth in vivo. These results disclose cytoplasmic c-Fos as a new target for effectively controlling brain tumor growth.

Published
2011

37. Both JNK and apoptosis pathways regulate growth and terminalia rotation during Drosophila genital disc development

Author

Nicolás Tomasini, Ana Macías, Sergio Gonzalo Benitez, and Claudia Marcela Sosa

Subjects
Male, Embryology, Programmed cell death, Apoptosis, Growth, Genitalia, Male, Ciencias Biológicas, purl.org/becyt/ford/1 [https], Animals, Humans, purl.org/becyt/ford/1.6 [https], Caspase, Cell Proliferation, biology, Cell Death, Cell growth, Kinase, Reaper, Biología del Desarrollo, JNK Mitogen-Activated Protein Kinases, Genitalia, Female, Head involution, engrailed, Cell biology, APOPTOSIS, DROSOPHILA, GENITAL DISC, Caspases, biology.protein, REAPER, Drosophila, Female, JNK, CIENCIAS NATURALES Y EXACTAS, Developmental Biology
Abstract

Apoptosis is necessary for the regulation of growth during development, but the precise details of this regulation have not been completely characterized. In this study, we used the Drosophila genital disc as a model to analyze the contribution of apoptosis to growth regulation. We studied the expression or activity of several elements of the apoptotic death pathway such as Drosophila inhibitor of apoptosis1, caspases, the apoptotic genes reaper (rpr) and head involution defective, as well as elements of the Jun-NH2-terminal kinase (JNK) pathway. We found that the JNK pathway is active in a dynamic, asymmetric and genitalia-specific manner. Apoptosis, as measured in terms of the expression of a variety of apoptotic molecules, occurs in a JNK-dependent and -independent manner and was detected among engrailed (en) expressing cells. JNK regulation of apoptotic genes is necessary to control growth in both sexes and rotation in males; this regulatory role is necessary to execute en(+) cell death and to activate expression of rpr in these cells. rpr is up-regulated at antero-posterior borders, and this expression appears to be of particular importance in the control of growth, since the balance between cell proliferation and death in those regions appears to depend on the equilibrium between pro- and anti-apoptotic factors at a cellular level. Fil: Benitez, Sergio Gonzalo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Departamento de Fisiología. Laboratorio de Genética del Desarrollo; Argentina Fil: Sosa, Claudia Marcela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Departamento de Fisiología. Laboratorio de Genética del Desarrollo; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Departamento de Fisiología. Laboratorio de Genética del Desarrollo; Argentina Fil: Macías, Ana. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Departamento de Fisiología. Laboratorio de Genética del Desarrollo; Argentina

Published
2010

38. Trypanosoma cruzi i genotypes in different geographical regions and transmission cycles based on a microsatellite motif of the intergenic spacer of spliced-leader genes

Author

Carolina Cura, Teresa Abate, Rodrigo Gurgel-Gonçalves, Aldo Solari, Marta Victoria Cardinal, Christine Aznar, Juan M. Burgos, Denis Blanchet, Ricardo E. Gürtler, Luis Miguel De Pablos, Patricio Diosque, Alexandre J. da Silva, Ana María Mejía-Jaramillo, Marcela Silvina Rodriguero, Nicolás Tomasini, César Augusto Cuba Cuba, Tomás Duffy, Antonio Osuna, Alejandro G. Schijman, Graciela Russomando, Sonia A. Kjos, Omar Triana-Chávez, and Mariano J. Levin

Subjects
RNA, Spliced Leader, Chagas disease, Disease reservoir, Genotype, Trypanosoma cruzi, Molecular Sequence Data, Article, Intergenic region, parasitic diseases, Triatoma infestans, medicine, Animals, Humans, Chagas Disease, Triatominae, Phylogeny, Disease Reservoirs, Genetics, Base Sequence, Geography, biology, Haplotype, Triatoma gerstaeckeri, DNA, Protozoan, medicine.disease, biology.organism_classification, Virology, Insect Vectors, Infectious Diseases, DNA, Intergenic, Parasitology, Sequence Alignment, Microsatellite Repeats
Abstract

The intergenic region of spliced-leader (SL-IR) genes from 105 Trypanosoma cruzi I (Tc I) infected biological samples, culture isolates and stocks from 11 endemic countries, from Argentina to the USA were characterised, allowing identification of 76 genotypes with 54 polymorphic sites from 123 aligned sequences. On the basis of the microsatellite motif proposed by Herrera et al. (2007) to define four haplotypes in Colombia, we could classify these genotypes into four distinct Tc I SL-IR groups, three corresponding to the former haplotypes Ia (11 genotypes), Ib (11 genotypes) and Id (35 genotypes); and one novel group, Ie (19 genotypes). Genotypes harbouring the Tc Ic motif were not detected in our study. Tc Ia was associated with domestic cycles in southern and northern South America and sylvatic cycles in Central and North America. Tc Ib was found in all transmission cycles from Colombia. Tc Id was identified in all transmission cycles from Argentina and Colombia, including Chagas cardiomyopathy patients, sylvatic Brazilian samples and human cases from French Guiana, Panama and Venezuela. Tc Ie gathered five samples from domestic Triatoma infestans from northern Argentina, nine samples from wild Mepraia spinolai and Mepraia gajardoi and two chagasic patients from Chile and one from a Bolivian patient with chagasic reactivation. Mixed infections by Tc Ia + Tc Id, Tc Ia + Tc Ie and Tc Id + Tc Ie were detected in vector faeces and isolates from human and vector samples. In addition, Tc Ia and Tc Id were identified in different tissues from a heart transplanted Chagas cardiomyopathy patient with reactivation, denoting histotropism. Trypanosoma cruzi I SL-IR genotypes from parasites infecting Triatoma gerstaeckeri and Didelphis virginiana from USA, T. infestans from Paraguay, Rhodnius nasutus and Rhodnius neglectus from Brazil and M. spinolai and M. gajardoi from Chile are to our knowledge described for the first time.

Published
2010

39. Experimental Evidence of Biological Interactions among Different Isolates of Trypanosoma cruzi from the Chaco Region

Author

Alejandro Uncos, Paula G. Ragone, Federico Ramos, Nicolás Tomasini, Anahí M. Alberti D´Amato, Miguel A. Basombrío, Rubén O. Cimino, Juan José Lauthier, Mercedes Monje Rumi, Cecilia Pérez Brandán, and Patricio Diosque

Subjects
Chagas disease, CIENCIAS MÉDICAS Y DE LA SALUD, Genotype, BIOLOGICAL INTERACTIONS, Trypanosoma cruzi, Science, Ciencias de la Salud, Parasitemia, Biology, Mice, purl.org/becyt/ford/3.3 [https], medicine, Animals, Humans, Chagas Disease, ISOLATES, Typing, Pathogen, Inflammation, Genetic diversity, Multidisciplinary, Genetic Variation, Heart, TRYPANOSOMA CRUZI, medicine.disease, biology.organism_classification, Virology, Biological interaction, Enfermedades Infecciosas, Immunology, Medicine, purl.org/becyt/ford/3 [https], Research Article
Abstract

Many infectious diseases arise from co-infections or re-infections with more than one genotype of the same pathogen. These mixed infections could alter host fitness, the severity of symptoms, success in pathogen transmission and the epidemiology of the disease. Trypanosoma cruzi, the etiological agent of Chagas disease, exhibits a high biological variability often correlated with its genetic diversity. Here, we developed an experimental approach in order to evaluate biological interaction between three T. cruzi isolates belonging to different Discrete Typing Units (DTUs TcIII, TcV and TcVI). These isolates were obtained from a restricted geographical area in the Chaco Region. Different mixed infections involving combinations of two isolates (TcIII + TcV, TcIII + TcVI and TcV + TcVI) were studied in a mouse model. The parameters evaluated were number of parasites circulating in peripheral blood, histopathology and genetic characterization of each DTU in different tissues by DNA hybridization probes. We found a predominance of TcVI isolate in blood and tissues respect to TcIII and TcV; and a decrease of the inflammatory response in heart when the damage of mice infected with TcVI and TcIII + TcVI mixture were compared. In addition, simultaneous presence of two isolates in the same tissue was not detected. Our results show that biological interactions between isolates with different biological behaviors lead to changes in their biological properties. The occurrence of interactions among different genotypes of T. cruzi observed in our mouse model suggests that these phenomena could also occur in natural cycles in the Chaco Region. Fil: Ragone, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Perez Brandan, Cecilia Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Monje Rumi, Maria Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Tomasini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Lauthier, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Cimino, Ruben O.. Universidad Nacional de Salta. Facultad de Cs.naturales. Escuela de Biologia. Catedra de Quimica Biologica; Argentina Fil: Uncos, Delfor Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Ramos, Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Alberti D'amato, Anahi M.. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina Fil: Basombrio, Miguel Angel Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina Fil: Diosque, Patricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Salta. Instituto de Patología Experimental; Argentina. Universidad Nacional de Salta; Argentina

Published
2015
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40. Growth of Peripheral and Central Nervous System Tumors Is Supported by Cytoplasmic c-Fos in Humans and Mice

Author

Nicolás Tomasini, Beatriz L. Caputto, Germán A. Gil, David C. Silvestre, and D. F. Bussolino

Subjects
Male, Nervous system, Cytoplasm, Pathology, medicine.medical_specialty, Neurofibromatosis 1, Genotype, Central nervous system, Cell Biology/Cell Growth and Division, lcsh:Medicine, Biology, Endoplasmic Reticulum, c-Fos, Neurological Disorders/Neuro-Oncology, Central Nervous System Neoplasms, Mice, medicine, Animals, Humans, Phosphorylation, lcsh:Science, Molecular Biology, Transcription factor, Oncology/Neuro-Oncology, Cell Proliferation, Mice, Knockout, Multidisciplinary, Cell growth, Activator (genetics), Endoplasmic reticulum, lcsh:R, Brain, Mice, Inbred C57BL, Disease Models, Animal, medicine.anatomical_structure, Membrane biogenesis, Cancer research, biology.protein, lcsh:Q, Proto-Oncogene Proteins c-fos, Research Article
Abstract

Background We have previously shown that the transcription factor c-Fos is also capable of associating to endoplasmic reticulum membranes (ER) and activating phospholipid synthesis. Herein we examined phospholipid synthesis status in brain tumors from human patients and from NPcis mice, an animal model of the human disease Neurofibromatosis Type 1 (NF1). Principal Findings In human samples, c-Fos expression was at the limit of detection in non-pathological specimens, but was abundantly expressed associated to ER membranes in tumor cells. This was also observed in CNS of adult tumor-bearing NPcis mice but not in NPcis fos(−/−) KO mice. A glioblastoma multiforme and a malignant PNS tumor from a NF1 patient (MPNST) showed a 2- and 4- fold c-Fos-dependent phospholipid synthesis activation, respectively. MPNST samples also showed increased cell proliferation rates and abundant c-Fos expression. Conclusions Results highlight a role of cytoplasmic c-Fos as an activator of phospholipid synthesis in events demanding high rates of membrane biogenesis as occurs for the exacerbated growth of tumors cells. They also disclose this protein as a potential target for controlling tumor growth in the nervous system.

Published
2010
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