Nano copper in the diet of laying quails: productive performance, metabolism, and tissue concentration.

The study evaluated the use of nano copper in semi-purified diets for laying quails and its effect on performance, metabolic state, and bioavailability. A total of 160 (180-days-old) quails were distributed in a completely randomized design, in a 3x3+1 factorial. The copper sources used were copper sulfate, copper oxide, and nano copper oxide, at levels of 200, 400, and 800 ppm each, totaling nine treatments plus a negative control (with no copper inclusion). The following variables were determined: weight gain, feed intake, egg production, egg weight, hemoglobin, hematocrit, Cu in the tissues and Cu bioavailability. Data were subjected to analysis of variance at 5% probability. The effect of sources and levels, as well as the interaction between the factors were evaluated. When interaction was observed, the effect of sources was evaluated separately by the Tukey's test and the effect of levels by regression, both at 5% probability. Copper nano oxide can be used at up to 800 ppm in the diet of laying quails without altering the productive performance, and with higher bioavailability than conventional copper oxide. Hemoglobin increases with the inclusion of 200 and 400 ppm of nano copper oxide and the hematocrit with 400 ppm.

Mitochondrion genomes of seven species of the endangered genus Sporophila (Passeriformes: Thraupidae).

We announce the mitochondrial genomes of seven species of the genus Sporophila (S. bouvreuil, S. iberaensis, S. melanogaster, S. minuta, S. nigrorufa, S. pileata, and S. ruficollis) which were validated by comparative genomic and phylogenetic analysis with related species. The mitochondrial genomes of seven passerines of the genus Sporophila were assembled (three complete and four nearly complete genomes) and were validated by reconstructing phylogenetic relations within Thraupidae. The complete mitogenomes ranged from 16,781 bp in S. ruficollis to 16,791 bp in S. minuta. We identified a conserved genome composition within all mitogenomes with 13 protein-coding genes, 22 tRNAs and two rRNAs. We observed a bias in the nucleotide composition and six mutational hotspots in Sporophila mitogenomes. Our mitogenome-based phylogenetic tree has S. minuta, S. maximiliani and S. nigricollis as sister species of the remaining species in the genus. We present new mitogenome sequences for seven Sporophila species, providing new genomic resources that may be useful for research on the evolution, comparative genetics, and conservation of this threatened group.

Chloroplast genome assembly of Serjania erecta Raldk: comparative analysis reveals gene number variation and selection in protein-coding plastid genes of Sapindaceae.

Serjania erecta Raldk is an essential genetic resource due to its anti-inflammatory, gastric protection, and anti-Alzheimer properties. However, the genetic and evolutionary aspects of the species remain poorly known. Here, we sequenced and assembled the complete chloroplast genome of S. erecta and used it in a comparative analysis within the Sapindaceae family. S. erecta has a chloroplast genome (cpDNA) of 159,297 bp, divided into a Large Single Copy region (LSC) of 84,556 bp and a Small Single Copy region (SSC) of 18,057 bp that are surrounded by two Inverted Repeat regions (IRa and IRb) of 28,342 bp. Among the 12 species used in the comparative analysis, S. erecta has the fewest long and microsatellite repeats. The genome structure of Sapindaceae species is relatively conserved; the number of genes varies from 128 to 132 genes, and this variation is associated with three main factors: (1) Expansion and retraction events in the size of the IRs, resulting in variations in the number of rpl22, rps19, and rps3 genes; (2) Pseudogenization of the rps2 gene; and (3) Loss or duplication of genes encoding tRNAs, associated with the duplication of trnH-GUG in X. sorbifolium and the absence of trnT-CGU in the Dodonaeoideae subfamily. We identified 10 and 11 mutational hotspots for Sapindaceae and Sapindoideae, respectively, and identified six highly diverse regions (tRNA-Lys - rps16, ndhC - tRNA-Val, petA - psbJ, ndhF, rpl32 - ccsA, and ycf1) are found in both groups, which show potential for the development of DNA barcode markers for molecular taxonomic identification of Serjania. We identified that the psaI gene evolves under neutrality in Sapindaceae, while all other chloroplast genes are under strong negative selection. However, local positive selection exists in the ndhF, rpoC2, ycf1, and ycf2 genes. The genes ndhF and ycf1 also present high nucleotide diversity and local positive selection, demonstrating significant potential as markers. Our findings include providing the first chloroplast genome of a member of the Paullinieae tribe. Furthermore, we identified patterns in variations in the number of genes and selection in genes possibly associated with the family's evolutionary history.

Histochemistry and transcriptomics of mucins and peritrophic membrane (PM) proteins along the midgut of a beetle with incomplete PM and their complementary function.

The midgut of Zabrotes subfasciatus (Coleoptera) and other insects may have regions lacking a peritrophic membrane (matrix, PM) and covered with a jelly-like material known as peritrophic gel. This work was undertaken to test the hypothesis that the peritrophic gel is a vertebrate-like mucus. By histochemistry we identified mucins along the whole midgut, which contrasts with the known occurrence of PM only at the posterior midgut. We also analyzed the expression of the genes coding for mucus-forming mucins (Mf-mucins), peritrophins, chitin synthases and chitin deacetylases along the midgut and carcass (insect without midgut) by RNA-seq. Mf-mucins were identified as proteins with high O-glycosylation and multiple tandem repeats of Pro/Thr/Ser residues. Peritrophins were separated into PM proteins, cuticular proteins analogous to peritrophins (CPAPs) and ubiquitous-chitin-binding domain-(CBD)-containing proteins (UCBPs). PM proteins have at least 3, CPAP one or 3, and UCBPs have a varied number of CBDs. PM proteins are more expressed at midgut, CPAP at the carcass, and UCBP at both. The results showed that most PM proteins are mainly expressed at the posterior midgut, together with midgut chitin synthase and chitin deacetylase, and in agreement with the presence of PM only at the posterior midgut by visual inspection. The excretion of most midgut chitinase is avoided, suggesting that the shortened PM is functional. Mf-mucins are expressed along the whole midgut, probably forming the extracellular mucus layer observed by histochemistry. Thus, the lack of PM at anterior and middle midgut causes the exposure of a mucus, which may correspond to the previously described peritrophic gel. The putative functional interplay of mucus and PM is discussed. The major role of mucus is proposed to be tissue protection and of PM to enhancing digestive efficiency by allowing enzyme recycling.

The complete chloroplast genome sequence of Eugenia klotzschiana O. Berg unveils the evolutionary dynamics in plastomes of Myrteae DC. Tribe (Myrtaceae).

Myrteae is the most diversified tribe in the Myrtaceae family and has great ecological and economic importance. Here, we performed the assembly and annotation of the chloroplast genome of Eugenia klotzschiana O. Berg and used this in a comparative analysis with other 13 species from the Myrteae tribe. The E. klotzschiana plastome exhibited a length of 158,977 bp and a very conserved structure and gene composition when compared with other Myrteae genomes. We identified 34 large repetitive sequences and 94 SSR repeats in E. klotzschiana plastome. The trnT-trnL, rpl32-trnL, ndhF-rpl32, psbE-petL, and ycf1 regions were identified as mutational hotspots. A negative selection signal was detected in 74 protein-coding genes while neutral evolution was detected in two genes (rps12 and psaI). Furthermore, 222 RNA editing sites were identified in the E. klotzschiana plastome. We also obtained a plastome-based Myrtales phylogenetic tree, including E. klotzschiana for the first time in a molecular phylogeny, recovering its sister relationship for all other Eugenia species. Our results illuminate how evolution shaped the chloroplast genome structure and composition in the Myrteae tribe, especially in the E. klotzschiana plastome.

Spiders' digestive system as a source of trypsin inhibitors: functional activity of a member of atracotoxin structural family.

Spiders are important predators of insects and their venoms play an essential role in prey capture. Spider venoms have several potential applications as pharmaceutical compounds and insecticides. However, transcriptomic and proteomic analyses of the digestive system (DS) of spiders show that DS is also a rich source of new peptidase inhibitor molecules. Biochemical, transcriptomic and proteomic data of crude DS extracts show the presence of molecules with peptidase inhibitor potential in the spider Nephilingis cruentata. Therefore, the aims of this work were to isolate and characterize molecules with trypsin inhibitory activity. The DS of fasting adult females was homogenized under acidic conditions and subjected to heat treatment. After that, samples were submitted to ion exchange batch and high-performance reverse-phase chromatography. The fractions with trypsin inhibitory activity were confirmed by mass spectrometry, identifying six molecules with inhibitory potential. The inhibitor NcTI (Nephilingis cruentata trypsin inhibitor) was kinetically characterized, showing a KD value of 30.25 nM ± 8.13. Analysis of the tertiary structure by molecular modeling using Alpha-Fold2 indicates that the inhibitor NcTI structurally belongs to the MIT1-like atracotoxin family. This is the first time that a serine peptidase inhibitory function is attributed to this structural family and the inhibitor reactive site residue is identified. Sequence analysis indicates that these molecules may be present in the DS of other spiders and could be associated to the inactivation of prey trypsin (serine peptidase) ingested by the spiders.

Spiders digestive system as a source of trypsin inhibitors: functional activity of a member of atracotoxin structural family

Spiders are important predators of insects and their venoms play an essential role in prey capture. Spider venoms have several potential applications as pharmaceutical compounds and insecticides. However, transcriptomic and proteomic analyses of the digestive system (DS) of spiders show that DS is also a rich source of new peptidase inhibitor molecules. Biochemical, transcriptomic and proteomic data of crude DS extracts show the presence of molecules with peptidase inhibitor potential in the spider Nephilingis cruentata. Therefore, the aims of this work were to isolate and characterize molecules with trypsin inhibitory activity. The DS of fasting adult females was homogenized under acidic conditions and subjected to heat treatment. After that, samples were submitted to ion exchange batch and high-performance reverse-phase chromatography. The fractions with trypsin inhibitory activity were confirmed by mass spectrometry, identifying six molecules with inhibitory potential. The inhibitor NcTI (Nephilingis cruentata trypsin inhibitor) was kinetically characterized, showing a KD value of 30.25 nM ± 8.13. Analysis of the tertiary structure by molecular modeling using Alpha-Fold2 indicates that the inhibitor NcTI structurally belongs to the MIT1-like atracotoxin family. This is the first time that a serine peptidase inhibitory function is attributed to this structural family and the inhibitor reactive site residue is identified. Sequence analysis indicates that these molecules may be present in the DS of other spiders and could be associated to the inactivation of prey trypsin (serine peptidase) ingested by the spiders.

Adaptation of Helicoverpa armigera to Soybean Peptidase Inhibitors Is Associated with the Transgenerational Upregulation of Serine Peptidases.

Molecular phenotypes induced by environmental stimuli can be transmitted to offspring through epigenetic inheritance. Using transcriptome profiling, we show that the adaptation of Helicoverpa armigera larvae to soybean peptidase inhibitors (SPIs) is associated with large-scale gene expression changes including the upregulation of genes encoding serine peptidases in the digestive system. Furthermore, approximately 60% of the gene expression changes induced by SPIs persisted in the next generation of larvae fed on SPI-free diets including genes encoding regulatory, oxidoreductase, and protease functions. To investigate the role of epigenetic mechanisms in regulating SPI adaptation, the methylome of the digestive system of first-generation larvae (fed on a diet with and without SPIs) and of the progeny of larvae exposed to SPIs were characterized. A comparative analysis between RNA-seq and Methyl-seq data did not show a direct relationship between differentially methylated and differentially expressed genes, while trypsin and chymotrypsin genes were unmethylated in all treatments. Rather, DNA methylation potential epialleles were associated with transcriptional and translational controls; these may play a regulatory role in the adaptation of H. armigera to SPIs. Altogether, our findings provided insight into the mechanisms of insect adaptation to plant antiherbivore defense proteins and illustrated how large-scale transcriptional reprograming of insect genes can be transmitted across generations.

Immunologic Diagnosis of Endemic Mycoses.

The endemic mycoses blastomycosis, coccidioidomycosis, histoplasmosis, paracoccidioidomycosis, cryptococcosis, sporotrichosis, talaromycosis, adiaspiromycosis, and emergomycosis are mostly caused by geographically limited thermally dimorphic fungi (except for cryptococcosis), and their diagnoses can be challenging. Usual laboratory methods involved in endemic mycoses diagnosis include microscopic examination and culture of biological samples; however, serologic, histopathologic, and molecular techniques have been implemented in the last few years for the diagnosis of these mycoses since the recovery and identification of their etiologic agents is time-consuming and lacks in sensitivity. In this review, we focus on the immunologic diagnostic methods related to antibody and antigen detection since their evidence is presumptive diagnosis, and in some mycoses, such as cryptococcosis, it is definitive diagnosis.

The Genome of Rhyzopertha dominica (Fab.) (Coleoptera: Bostrichidae): Adaptation for Success.

The lesser grain borer, Rhyzopertha dominica (F.) (Coleoptera: Bostrichidae), is a major global pest of cereal grains. Infestations are difficult to control as larvae feed inside grain kernels, and many populations are resistant to both contact insecticides and fumigants. We sequenced the genome of R. dominica to identify genes responsible for important biological functions and develop more targeted and efficacious management strategies. The genome was assembled from long read sequencing and long-range scaffolding technologies. The genome assembly is 479.1 Mb, close to the predicted genome size of 480.4 Mb by flow cytometry. This assembly is among the most contiguous beetle assemblies published to date, with 139 scaffolds, an N50 of 53.6 Mb, and L50 of 4, indicating chromosome-scale scaffolds. Predicted genes from biologically relevant groups were manually annotated using transcriptome data from adults and different larval tissues to guide annotation. The expansion of carbohydrase and serine peptidase genes suggest that they combine to enable efficient digestion of cereal proteins. A reduction in the copy number of several detoxification gene families relative to other coleopterans may reflect the low selective pressure on these genes in an insect that spends most of its life feeding internally. Chemoreceptor genes contain elevated numbers of pseudogenes for odorant receptors that also may be related to the recent ontogenetic shift of R. dominica to a diet consisting primarily of stored grains. Analysis of repetitive sequences will further define the evolution of bostrichid beetles compared to other species. The data overall contribute significantly to coleopteran genetic research.

Diagnosis of MALT Lymphoma from Surveillance Endoscopy of a Patient with a CDH1 Gene Germline Mutation.

Carriers of the mutated CDH1 gene have an increased risk of developing early-onset signet-ring cell (diffuse) gastric cancer. We present a case of a young patient with a confirmed mutation of the CDH1 gene, who was diagnosed with a gastric marginal zone B-cell lymphoma (MZL) of mucosa-associated lymphoid tissue (MALT lymphoma) from surveillance endoscopy. He underwent Helicobacter pylori eradication treatment and was subsequently submitted to a total prophylactic gastrectomy. The surgical specimen only revealed foci of signet-ring cell carcinoma (SRCC) in situ without lymphoma signs. We highlight here the occurrence of other pathology in high-risk patients as well as its possible influence on the decision to perform gastrectomy.

A mutação do gene CDH1 determina um risco aumentado de desenvolvimento precoce de cancro gástrico de células em anel de sinete (tipo difuso). Apresentamos um caso de um doente jovem portador de uma mutação no gene CDH1 que foi diagnosticado com linfoma de MALT gástrico numa endoscopia de vigilância. O doente foi submetido a terapêutica de erradicação da Helicobacter pylori e subsequentemente realizou uma gastrectomia total profilática. A avaliação histológica da peça cirúrgica identificou focos de carcinoma in situ de células de anel em sinete, sem evidência de linfoma. O nosso objetivo é salientar a ocorrência de outras patologias em doentes de alto risco assim como a sua possível influência na decisão cirúrgica.

Recruitment of lysosomal cathepsins B, L and D as digestive enzymes in Coleoptera.

The recruitment of the lysosomal cathepsins B (CAB), L (CAL) and D (CAD) as luminal digestive enzymes was investigated in 3 species of beetles. Gene expression was determined by RNA-seq in different regions of the midgut and in the carcasses from the transcriptomes of Dermestes maculatus, Tenebrio molitor and Zabrotes subfasciatus. These data together with phylogenetic analyses, allowed us to identify the sequences of the gene coding for digestive and lysosomal CABs, CADs and CALs in T. molitor and Z. subfasciatus and observe the absence of digestive cathepsins in D. maculatus. Comparisons of structures based on the overall similarity of modelled structures were performed and subsite residues in the lysosomal and digestive CALs were identified by molecular docking. The data showed that S2 subsites are very variable, probably as an adaption to a luminal digestive role. The survey of sequences of the gene coding for cathepsins in the genomes of 13 beetle species from different phylogenetic groups showed that expansion of CAL and CAB genes occurred only in the Cucujiformia clade. Several digestive CABs have a reduced occluding loop, probably to act as digestive enzymes. Pollen-feeding was proposed to be the selective pressure to recruit cathepsins as digestive enzymes in Cucujiformia beetles.

Structural and Evolutionary Analyses of PR-4 SUGARWINs Points to a Different Pattern of Protein Function.

SUGARWINs are PR-4 proteins associated with sugarcane defense against phytopathogens. Their expression is induced in response to damage by Diatraea saccharalis larvae. These proteins play an important role in plant defense, in particular against fungal pathogens, such as Colletothricum falcatum (Went) and Fusarium verticillioides. The pathogenesis-related protein-4 (PR-4) family is a group of proteins equipped with a BARWIN domain, which may be associated with a chitin-binding domain also known as the hevein-like domain. Several PR-4 proteins exhibit both chitinase and RNase activity, with the latter being associated with the presence of two histidine residues H11 and H113 (BARWIN) [H44 and H146, SUGARWINs] in the BARWIN-like domain. In sugarcane, similar to other PR-4 proteins, SUGARWIN1 exhibits ribonuclease, chitosanase and chitinase activities, whereas SUGARWIN2 only exhibits chitosanase activity. In order to decipher the structural determinants involved in this diverse range of enzyme specificities, we determined the 3-D structure of SUGARWIN2, at 1.55Å by X-ray diffraction. This is the first structure of a PR-4 protein where the first histidine has been replaced by asparagine and was subsequently used to build a homology model for SUGARWIN1. Molecular dynamics simulations of both proteins revealed the presence of a flexible loop only in SUGARWIN1 and we postulate that this, together with the presence of the catalytic histidine at position 42, renders it competent as a ribonuclease. The more electropositive surface potential of SUGARWIN1 would also be expected to favor complex formation with RNA. A phylogenetic analysis of PR-4 proteins obtained from 106 Embryophyta genomes showed that both catalytic histidines are widespread among them with few replacements in these amino acid positions during the gene family evolutionary history. We observe that the H11 replacement by N11 is also present in two other sugarcane PR-4 proteins: SUGARWIN3 and SUGARWIN4. We propose that RNase activity was present in the first Embryophyta PR-4 proteins but was recently lost in members of this family during the course of evolution.

High-Intensity Interval Training upon Cognitive and Psychological Outcomes in Youth: A Systematic Review.

Development of innovative and time-efficient strategies to involve youth in physical activity is pivotal in the actual inactivity pandemic. Moreover, physical activity may improve academic performance, of great interest for educators. This present systematic review aimed to analyze the effects of high-intensity interval training (HIIT) on cognitive performance and psychological outcomes in youth. A database search (Web of Science, PubMed, Scopus, and PsycINFO) for original research articles was performed. A total of eight articles met the inclusion criteria, and the Cochrane risk of bias tool was used. The studies' results were recalculated to determine effect sizes using Cohen's d. Different HIIT interventions reported improvements on cognitive performance at executive function (d = 0.75, +78.56%), linguistic reasoning (d = 0.25, +7.66%), concentration (d = 0.71, +61.10%), selective attention (d = 0.81, +60.73%), non-verbal and verbal abilities (d = 0.88, +47.50%; d = 1.58, +22.61%, respectively), abstract reasoning (d = 0.75, +44.50%), spatial and numerical abilities (d = 37.19, +22.85%; d = 1.20, +8.28%, respectively), and verbal reasoning (d = 1.00, +15.71%) in youth. Regarding psychological outcomes, HIIT showed higher self-concept (d = 0.28, +8.71%) and psychological well-being in boys and girls (d = 0.73, +32.43%, d = 0.39, +11.58%, respectively). To sum up, HIIT interventions between 4-16 weeks, for 8-30 min/session, at ≥85% maximal heart rate, would provide positive effects on cognitive performance and psychological outcomes in youth.

New germline GATA1 variant in females with anemia and thrombocytopenia.

Familial forms of bone marrow defects are rare disorders and description of new cases are valuable opportunities to clarify the molecular machinery that triggers hematopoiesis and blood formation, as well as risk to malignant transformation. We investigated the genetic scenario and possible patterns of transmission in a rare case of familial myeloid disorder with a history of exposure to pesticides. Blood counts of two proband sisters, age 41 and 42, revealed mild anemia, neutrophilia and thrombocytopenia with bone marrow finding mimicking primary myelofibrosis in the cellular phase. We analyzed the coding regions of 78 myeloid neoplasms-related genes and 16 encoding xenobiotic metabolizing genes using Next-Generation Sequencing. The GATA1 variant c.788C > T, p.T263M, located in the C-terminal zinc finger domain of GATA1, was detected in the DNA of the two sisters. The screening of the other kindreds also revealed the p.T263M variant in the mother and two daughters with the same bone marrow disorder. This is the first report of an alteration in the GATA1 CF domain causing anemia, thrombocytopenia and megakaryocyte proliferation with mild myelofibrosis, correlating a new GATA1 germline variant with myeloid disorder.

Cathepsins L and B in Dysdercus peruvianus, Rhodnius prolixus, and Mahanarva fimbriolata. Looking for enzyme adaptations to digestion.

Cysteine peptidases (CP) play a role as digestive enzymes in hemipterans similar to serine peptidases in most other insects. There are two major CPs: cathepsin L (CAL), which is an endopeptidase and cathepsin B (CAB) that is both an exopeptidase and a minor endopeptidase. There are thirteen putative CALs in Dysdercus peruvianus, which in some cases were confirmed by cloning their encoding genes. RNA-seq data showed that DpCAL5 is mainly expressed in the anterior midgut (AM), DpCAL10 in carcass (whole body less midgut), suggesting it is a lysosomal enzyme, and the other DpCALs are expressed in middle (MM) and posterior (PM) midgut. The expression data were confirmed by qPCR and enzyme secretion to midgut lumen by a proteomic approach. Two CAL activities were isolated by chromatography from midgut samples with similar kinetic properties toward small substrates. Docking analysis of a long peptide with several DpCALs modeled with digestive Tenebrio molitor CAL (TmCAL3) as template showed that on adapting to luminal digestion DpCALs (chiefly DpCAL5) changed in relation to their ancestral lysosomal enzyme (DpCAL10) mainly at its S2 subsite. A similar conclusion arrived from structure alignment-based clustering of DpCALs based on structural similarity of the modeled structures. Changes mostly on S2 subsite could mean the enzymes turn out less peptide-bond selective, as described in TmCALs. R. prolixus CALs changed on adapting to luminal digestion, although less than DpCALs. Both D. peruvianus and R. prolixus have two digestive CABs which are expressed in the same extension as CALs, in the first digestive section of the midgut, but less than in the other midgut sections. Mahanarva fimbriolata does not seem to have digestive CALs and their digestive CABs are mainly expressed in the first digestive section of the midgut and do not diverge much from their lysosomal counterparts. The data suggest that CABs are necessary at the initial stage of digestion in CP-dependent Hemipterans, which action is completed by CALs with low peptide-bond selectivity in Heteroptera species. In M. fimbriolata protein digestion is supposed to be associated with the inactivation of sap noxious proteins, making CAB sufficient as digestive CP. Hemipteran genomes and transcriptome data showed that CALs have been recruited as digestive enzymes only in heteropterans, whereas digestive CABs occur in all hemipterans.

Performance, metabolic efficiency and egg quality in Japanese quails fed with acidulated soybean soapstock and lecithin for a prolonged period.

This study aimed to evaluate the use of acidulated soybean soapstock in association with lecithin on productive performance, metabolic efficiency in the utilization of nutrientes, and the egg quality of Japanese quails. A total of 192 quails were used, distributed randomly in a 2×2×2 factorial scheme that included two types of oils, two levels of supplementation (4% and 8%) and two levels of lecithin (1% or 0%). At the end of the six-month experimental period, some double significant interactions were shown between the level of oil and lecithin for the performance variables (egg weight p=0.04, feed intake p<0.01 and feed conversion rate p=0.04). The feed conversion rate also was influenced by a double significant interaction between the type of oil and the level of oil (p<0.01). The nutrient digestibility showed that different interactions affected the results. The evaluation of egg quality, was verified that the use of acidulated soybean soapstock did not affect most variables of internal quality. The results showed that it is possible to use 8% acidulated soybean soapstock in combination with 1% lecithin in the diets of Japanese quails for a period of up to six months without a reduction in performance.

Autochthonous canine visceral leishmaniasis cases occur in Paraná state since 2012: isolation and identification of Leishmania infantum / Leishmaniose visceral canina autóctone ocorre no estado do Paraná desde 2012: isolamento e identificação de Leishmania infantum

Abstract The aim of this study was to confirm the emergence of canine visceral leishmaniasis among dogs in Foz do Iguaçu. The disease was diagnosed through the isolation and molecular identification of Leishmania infantum. In the first sample collection stage (2012), three lymph node aspirates and 46 buffy coat samples were obtained mostly from the dogs that were seroreagents for leishmaniasis. In the second sample collection stage (2013), the buffy coat samples were collected from 376 dogs located close to Paraguay, Paraná river, center and peripheral parts of the city. The DNA from the six isolates, four from the first sampling stage (4/49) and two from the second sampling stage (2/376), was subjected to polymerase chain reaction using the K26F/R primers. The isolate was confirmed as L. infantum by sequencing. As none of the dogs had ever left the city, the isolates were confirmed as autochthonous. Further, the study confirmed the emergence of canine visceral leishmaniasis in Paraná through the identification of L. infantum among dogs in Foz do Iguaçu city. Hence, collaborative control measures should be designed and implemented by the public agencies and research institutions of Brazil, Argentina, and Paraguay to control the spread of visceral leishmaniasis.

Resumo O objetivo deste estudo foi confirmar a emergência da leishmaniose visceral canina em Foz do Iguaçu próximo à fronteira com a Argentina e ao Paraguai, por meio do isolamento e identificação molecular de Leishmania infantum. Em um primeiro estágio de coleta de animais (2012), três amostras de aspirados de linfonodos e 46 camadas leucocitárias foram obtidas de cães soropositivos para leishmaniose. Em um segundo estágio de coleta (2013), foram coletadas amostras de camada leucocitária de 376 cães de 20 localidades próximas à fronteira com o Paraguai, rio Paraná, centro e periferia da cidade. Seis isolados foram obtidos, quatro da primeira etapa (4/49) e dois da segunda etapa (2/376); estes isolados foram submetidos à amplificação com iniciadores K26F/R, e a análise de sua sequência confirmou a espécie como L. infantum. A autoctonia dos casos foi confirmada, pois 100% dos cães nunca haviam saído da cidade. O estudo confirma a emergência de leishmaniose visceral canina no Paraná com identificação de L. infantum em cães da cidade de Foz do Iguaçu. Assim, medidas de controle devem ser elaboradas e implementadas por órgãos públicos e instituições de pesquisa do Brasil, Argentina e Paraguai em parceria com o objetivo de controlar a disseminação de zoonoses e os casos humanos de LV.

PpCRN7 and PpCRN20 of Phythophthora parasitica regulate plant cell death leading to enhancement of host susceptibility.

BACKGROUND: Phytophthora species secrete cytoplasmic effectors from a family named Crinkler (CRN), which are characterised by the presence of conserved specific domains in the N- and C-terminal regions. P. parasitica causes disease in a wide range of host plants, however the role of CRN effectors in these interactions remains unclear. Here, we aimed to: (i) identify candidate CRN encoding genes in P. parasitica genomes; (ii) evaluate the transcriptional expression of PpCRN (Phytophthora parasitica Crinkler candidate) during the P. parasitica interaction with Citrus sunki (high susceptible) and Poncirus trifoliata (resistant); and (iii) functionally characterize two PpCRNs in the model plant Nicotiana benthamiana. RESULTS: Our in silico analyses identified 80 putative PpCRN effectors in the genome of P. parasitica isolate 'IAC 01/95.1'. Transcriptional analysis revealed differential gene expression of 20 PpCRN candidates during the interaction with the susceptible Citrus sunki and the resistant Poncirus trifoliata. We have also found that P. parasitica is able to recognize different citrus hosts and accordingly modulates PpCRNs expression. Additionally, two PpCRN effectors, namely PpCRN7 and PpCRN20, were further characterized via transient gene expression in N. benthamiana leaves. The elicitin INF-1-induced Hypersensitivity Response (HR) was increased by an additive effect driven by PpCRN7 expression, whereas PpCRN20 expression suppressed HR response in N. benthamiana leaves. Despite contrasting functions related to HR, both effectors increased the susceptibility of plants to P. parasitica. CONCLUSIONS: PpCRN7 and PpCRN20 have the ability to increase P. parasitica pathogenicity and may play important roles at different stages of infection. These PpCRN-associated mechanisms are now targets of biotechnological studies aiming to break pathogen's virulence and to promote plant resistance.

Autochthonous canine visceral leishmaniasis cases occur in Paraná state since 2012: isolation and identification of Leishmania infantum.

The aim of this study was to confirm the emergence of canine visceral leishmaniasis among dogs in Foz do Iguaçu. The disease was diagnosed through the isolation and molecular identification of Leishmania infantum. In the first sample collection stage (2012), three lymph node aspirates and 46 buffy coat samples were obtained mostly from the dogs that were seroreagents for leishmaniasis. In the second sample collection stage (2013), the buffy coat samples were collected from 376 dogs located close to Paraguay, Paraná river, center and peripheral parts of the city. The DNA from the six isolates, four from the first sampling stage (4/49) and two from the second sampling stage (2/376), was subjected to polymerase chain reaction using the K26F/R primers. The isolate was confirmed as L. infantum by sequencing. As none of the dogs had ever left the city, the isolates were confirmed as autochthonous. Further, the study confirmed the emergence of canine visceral leishmaniasis in Paraná through the identification of L. infantum among dogs in Foz do Iguaçu city. Hence, collaborative control measures should be designed and implemented by the public agencies and research institutions of Brazil, Argentina, and Paraguay to control the spread of visceral leishmaniasis.