Selective and non-selective evolutionary signatures found in the simplest replicative biological entities.

Mitoviruses, which are considered evolutionary relics of extinct alpha-proteobacteria RNA phages, represent one of the simplest self-replicating biological systems. This study aims to quantitatively describe genomes and identify potential genomic signatures that support the protein phylogenetic-based classification criterion. Genomic variables, such as mononucleotide and dinucleotide composition, codon usage bias, and minimal free energy derived from optimized predicted RNA secondary structure, were analyzed. From the values obtained, the main evolutionary pressures were discussed, indicating that natural selection plays a significant role in shaping mitovirus genomes. However, neutral evolution also makes a significant contribution. This study reveals a significant discovery of structural divergence in Kvaramitovirus. The energy minimization approach employed to study 2D folding in this study reveals a distinct spatial organization of their genomes, providing evidence for the hypothesis of a single evolutionary event of circularization in the most recent common ancestor of the lineage. This hypothesis was discussed in light of recent discoveries by other researchers that partially support the existence of mitoviruses with circular genomes. Finally, this study represents a significant advancement in the understanding of mitoviruses, as it quantitatively describes the nucleotide sequence at the family and genus taxonomic levels. Additionally, we provide hypotheses that can be experimentally validated to inspire new research and address the gaps in knowledge of this fascinating, basally divergent RNA virus lineage.

An in silico analysis revealed a novel evolutionary lineage of putative mitoviruses.

Mitoviruses (family Mitoviridae) are small capsid-less RNA viruses that replicate in the mitochondria of fungi and plants. However, to date, the only authentic animal mitovirus infecting an insect was identified as Lutzomyia longipalpis mitovirus 1 (LulMV1). Public databases of transcriptomic studies from several animals may be a good source for identifying the often missed mitoviruses. Consequently, a search of mitovirus-like transcripts at the NCBI transcriptome shotgun assembly (TSA) library, and a search for the mitoviruses previously recorded at the NCBI non-redundant (nr) protein sequences library, were performed in order to identify new mitovirus-like sequences associated with animals. In total, 10 new putative mitoviruses were identified in the TSA database and 8 putative mitoviruses in the nr protein database. To our knowledge, these results represent the first evidence of putative mitoviruses associated with poriferan, cnidarians, echinoderms, crustaceans, myriapods and arachnids. According to different phylogenetic inferences using the maximum likelihood method, these 18 putative mitoviruses form a robust monophyletic lineage with LulMV1, the only known animal-infecting mitovirus. These findings based on in silico procedures provide strong evidence for the existence of a clade of putative mitoviruses associated with animals, which has been provisionally named 'kvinmitovirus'.

Microscopic and PCR-based detection of microsporidia spores in human stool samples.

Microsporidia are obligate intracellular fungi with a remarkable ability to infect a wide range of invertebrate and vertebrate hosts. Namely, Enterocytozoon bieneusi is the most frequently microsporidia reported worldwide, and mainly associated with chronic diarrhea and wasting syndrome in AIDS patients. Microscopy and PCR-based detection techniques are effective for diagnosis and identification of species and genotypes; however, these methods should be standardized in each laboratory. In this study, we performed microscopy and nested PCR techniques with PCR product sequencing to detect E. bieneusi in human stool samples. These techniques, if applied together, might prove useful for diagnosis and future epidemiological studies of intestinal microsporidiosis in Argentina.

Antioxidant activity of chia flour as a food supplement in a cellular model: Repercussions of processing and in vitro digestion.

Food processing and digestion can alter bioactive compound composition of food, affecting their potential biological activity. In this study, we evaluated the direct and protective antioxidant effects of polyphenols extracted from defatted chia flour (DCF) (salviaflaside, rosmarinic and fertaric acid as major compounds), sweet cookies supplemented with DCF (CFC) (same major compounds), and their digested fractions (rosmarinic acid, salviaflaside, fertaric and salvianolic E/B/L acid as major compounds) in HepG2 cells in basal and in oxidative stress conditions. DCF showed protective antioxidant effects by decreasing reactive oxygen species (ROS) and protein oxidation products (POP) while increasing reduced glutathione (GSH). Additionally, CFC revealed similar protective effects and even showed enhanced modulation of the antioxidant system due to the activation of antioxidant enzymes. However, the digested fractions only decreased ROS, indicating continued antioxidant effects. This study underscores the importance of evaluating manufacturing and digestion effects to confirm a food's antioxidant properties.

Involvement of aryl hydrocarbon receptor in the aflatoxin B1 and fumonisin B1 effects on in vitro differentiation of murine regulatory-T and Th17 cells.

Aflatoxin B1 (AFB1) and fumonisin B1 (FB1) are mycotoxins widely found as cereal contaminants, and their co-consumption is associated with liver cancer. Both are immunotoxic, but their interactions have been little studied. This work was aimed to evaluate in mouse spleen mononuclear cells (SMC) the effects of the exposure to AFB1 (5-50 µM), FB1 (25-250 µM), and AFB1-FB1 mixtures (MIX) on the in vitro differentiation of regulatory T cells (Treg and Tr1-like) and Th17 cells, as well as elucidate the contribution of aryl hydrocarbon receptor (Ahr) in such effects. AFB1 and mainly MIX induced cytotoxicity in activated CD4 cells via Ahr signaling. AFB1 (5 µM) increased the Treg cell differentiation, but its combination with FB1 (25 µM) also reduced Th17 cell expansion by Ahr-dependent mechanisms. Therefore, this mixture could enhance the Treg/Th17 cell ratio and favor immunosuppression and escape from tumor immunosurveillance to a greater extent than individual mycotoxins. Whereas, AFB1-FB1 mixtures at medium-high doses inhibited the Tr1-like cell expansion induced by the individual mycotoxins and affected Treg and Th17 cell differentiation in Ahr-independent and dependent manners, respectively, which could alter anti-inflammatory and Th17 immune responses. Moreover, individual FB1 altered regulatory T and Th17 cell development independently of Ahr. In conclusion, AFB1 and FB1 interact by modifying Ahr signaling, which is involved in the immunotoxicity as well as in the alteration of the differentiation of Treg, Tr1-like, and Th17 cells induced by AFB1-FB1 mixtures. Therefore, Ahr is implicated in the regulation of the anti- and pro-inflammatory responses caused by the combination of AFB1 and FB1.

Putative Mitoviruses without In-Frame UGA(W) Codons: Evolutionary Implications.

Mitoviruses are small vertically transmitted RNA viruses found in fungi, plants and animals. Taxonomically, a total of 105 species and 4 genera have been formally recognized by ICTV, and recently, 18 new putative species have been included in a new proposed genus. Transcriptomic and metatranscriptomic studies are a major source of countless new virus-like sequences that are continually being added to open databases and these may be good sources for identifying new putative mitoviruses. The search for mitovirus-like sequences in the NCBI databases resulted in the discovery of more than one hundred new putative mitoviruses, with important implications for taxonomy and also for the evolutionary scenario. Here, we propose the inclusion of four new putative members to the genus Kvaramitovirus, and the existence of a new large basally divergent lineage composed of 144 members that lack internal UGA codons (subfamily "Arkeomitovirinae"), a feature not shared by the vast majority of mitoviruses. Finally, a taxonomic categorization proposal and a detailed description of the evolutionary history of mitoviruses were carried out. This in silico study supports the hypothesis of the existence of a basally divergent lineage that could have had an impact on the early evolutionary history of mitoviruses.

Effects of chlorine and peroxyacetic acid wash treatments on growth kinetics of Salmonella in fresh-cut lettuce.

Fresh-cut produces are often consumed uncooked, thus proper sanitation is essential for preventing cross contamination. The reduction and subsequent growth of Salmonella enterica sv Thompson were studied in pre-cut iceberg lettuce washed with simulated wash water (SWW), sodium hypochlorite (SH, free chlorine 25 mg/L), and peroxyacetic acid (PAA, 80 mg/L) and stored for 9 days under modified atmosphere at 9, 13, and 18 °C. Differences in reduction between SH and PAA were non-existent. Overall, visual quality, dehydration, leaf edge and superficial browning and aroma during storage at 9 °C were similar among treatments, but negative effects increased with temperature. These results demonstrated that PAA can be used as an effective alternative to chlorine for the disinfection of Salmonella spp. in fresh-cut lettuce. The growth of Salmonella enterica sv Thompson was successfully described with the Baranyi and Roberts growth model in the studied storage temperature range, and after treatment with SWW, chlorine, and PAA. Subsequently, predictive secondary models were used to describe the relationship between growth rates and temperature based on the models' family described by Belehrádek. Interestingly, the exposure to disinfectants biased growth kinetics of Salmonella during storage. Below 12 °C, growth rates in lettuce treated with disinfectant (0.010-0.011 log CFU/h at 9 °C) were lower than those in lettuce washed with water (0.016 log CFU/h at 9 °C); whereas at higher temperatures, the effect was the opposite. Thus, in this case, the growth rate values registered at 18 °C for lettuce treated with disinfectant were 0.048-0.054 log CFU/h compared to a value of 0.038 log CFU/h for lettuce treated with only water. The data and models developed in this study will be crucial to describing the wash-related dynamics of Salmonella in a risk assessment framework applied to fresh-cut produce, providing more complete and accurate risk estimates.

Unveiling Fresh-Cut Lettuce Processing in Argentine Industries: Evaluating Salmonella Levels Using Predictive Microbiology Models.

A survey was performed to gather information on the processing steps, conditions, and practices employed by industries processing ready-to-eat (RTE) leafy vegetables in Argentina. A total of seven industries participated in the survey. A cluster analysis of the data obtained was performed to identify homogeneous groups among the participating industries. The data collected were used as inputs of two predictive microbiology models to estimate Salmonella concentrations after chlorine washing, during storage and distribution of final products, and to rank the different practices according to the final estimated Salmonella levels. Six different clusters were identified by evaluating the parameters, methods, and controls applied in each processing step, evidencing a great variability among industries. The disinfectant agent applied by all participating industries was sodium hypochlorite, though concentrations and application times differed among industries from 50 to 200 ppm for 30 to 110 s. Simulations using predictive models indicated that the reductions in Salmonella in RTE leafy vegetables would vary in the range of 1.70-2.95 log CFU/g during chlorine-washing depending on chlorine concentrations applied, washing times, and vegetable cutting size, which varied from 9 to 16 cm2 among industries. Moreover, Salmonella would be able to grow in RTE leafy vegetables during storage and distribution, achieving levels of up to 2 log CFU/g, considering the storage and transportation temperatures and times reported by the industries, which vary from 4 to 14 °C and from 18 to 30 h. These results could be used to prioritize risk-based sampling programs by Food Official Control or determine more adequate process parameters to mitigate Salmonella in RTE leafy vegetables. Additionally, the information gathered in this study is useful for microbiological risk assessments.

Association between altered tryptophan metabolism, plasma aryl hydrocarbon receptor agonists, and inflammatory Chagas disease.

Introduction: Chagas disease causes a cardiac illness characterized by immunoinflammatory reactions leading to myocardial fibrosis and remodeling. The development of Chronic Chagas Cardiomyopathy (CCC) in some patients while others remain asymptomatic is not fully understood, but dysregulated inflammatory responses are implicated. The Aryl hydrocarbon receptor (AhR) plays a crucial role in regulating inflammation. Certain tryptophan (Trp) metabolites have been identified as AhR ligands with regulatory functions. Methods results and discussion: We investigated AhR expression, agonist response, ligand production, and AhR-dependent responses, such as IDO activation and regulatory T (Treg) cells induction, in two T. cruzi-infected mouse strains (B6 and Balb/c) showing different polymorphisms in AhR. Furthermore, we assessed the metabolic profile of Trp catabolites and AhR agonistic activity levels in plasma samples from patients with chronic Chagas disease (CCD) and healthy donors (HD) using a luciferase reporter assay and liquid chromatography-mass spectrophotometry (LC-MS) analysis. T. cruzi-infected B6 mice showed impaired AhR-dependent responses compared to Balb/c mice, including reduced IDO activity, kynurenine levels, Treg cell induction, CYP1A1 up-regulation, and AhR expression following agonist activation. Additionally, B6 mice exhibited no detectable AhR agonist activity in plasma and displayed lower CYP1A1 up-regulation and AhR expression upon agonist activation. Similarly, CCC patients had decreased AhR agonistic activity in plasma compared to HD patients and exhibited dysregulation in Trp metabolic pathways, resulting in altered plasma metabolite profiles. Notably, patients with severe CCC specifically showed increased N-acetylserotonin levels in their plasma. The methods and findings presented here contribute to a better understanding of CCC development mechanisms and may identify potential specific biomarkers for T. cruzi infection and the severity of associated heart disease. These insights could be valuable in designing new therapeutic strategies. Ultimately, this research aims to establish the AhR agonistic activity and Trp metabolic profile in plasma as an innovative, non-invasive predictor of prognosis for chronic Chagas disease.

Indoleamine 2,3-dioxigenase (IDO) is critical for host resistance against Trypanosoma cruzi.

Indoleamine 2,3-dioxygenase (IDO) is an inflammatory cytokine-inducible rate-limiting enzyme of the tryptophan (Trp) catabolism, which is involved in the inhibition of intracellular pathogen replication as well as in immunomodulation. Here we demonstrated the effect of IDO-dependent Trp catabolism on Trypanosoma cruzi resistance to acute infection. Infection with T. cruzi resulted in the systemic activation of IDO. The blocking of IDO activity in vivo impaired resistance to the infection and exacerbated the parasite load and infection-associated pathology. In addition, IDO activity was critical to controlling the parasite's replication in macrophages (Mos), despite the high production of nitric oxide produced by IDO-blocked T. cruzi-infected Mos. Analysis of the mechanisms by which IDO controls the parasite replication revealed that T. cruzi amastigotes were sensitive to L-kynurenine downstream metabolites 3-hydroxykynurenine (3-HK) and 3-hydroxyanthranilic acid, while 3-HK also affected the trypomastigote stage. Finally, 3-HK treatment of mice acutely infected with T. cruzi was able to control the parasite and to improve the survival of lethally infected mice. During infection, IDO played a critical role in host defense against T. cruzi; therefore, the intervention of IDO pathway could be useful as a novel antitrypanosomatid therapeutic strategy.

A Survey of Mycoviral Infection in Fusarium spp. Isolated from Maize and Sorghum in Argentina Identifies the First Mycovirus from Fusarium verticillioides.

Mycoviruses appear to be widespread in Fusarium species worldwide. The aim of this work was to identify mycoviral infections in Fusarium spp., isolated from maize and sorghum grown in Argentina, and to estimate their potential effects on the pathogenicity and toxigenesis of the host fungus towards maize. Mycoviruses were identified in 2 out of 105 isolates analyzed; Fusarium verticillioides strain Sec505 and Fusarium andiyazi strain 162. They were characterized as members of the genus Mitovirus by high-throughput sequencing and sequence analysis. The F. verticillioides mitovirus was a novel mycovirus whereas the F. andiyazi mitovirus was found to be a new strain of a previously identified mitovirus. We have named these mitoviruses, Fusarium verticillioides mitovirus 1 (FvMV1) and Fusarium andiyazi mitovirus 1 strain 162 (FaMV1-162). To our knowledge, FvMV1 is the first mycovirus reported as naturally infecting F. verticillioides, the major causal agent of ear rot and fumonisin producer in corn. Both mitoviruses exhibited 100% vertical transmission rate to microconidia. The Fa162 strain infected with FaMV1-162 did not show phenotypic alterations. In contract, F. verticillioides Sec505 infected with FvMV1 showed increased virulence as well as microconidia and fumonisin-B1 production, compared with two uninfected strains. These results suggest that FvMV1 could have a role in modulating F. verticillioides pathogenicity and toxin production worth further exploring.

Deficiency of CD73 activity promotes protective cardiac immunity against Trypanosoma cruzi infection but permissive environment in visceral adipose tissue.

Damaged cells release the pro-inflammatory signal ATP, which is degraded by the ectonucleotidases CD39 and CD73 to the anti-inflammatory mediator adenosine (ADO). The balance between ATP/ADO is known to determine the outcome of inflammation/infection. However, modulation of the local immune response in different tissues due to changes in the balance of purinergic metabolites has yet to be investigated. Here, we explored the contribution of CD73-derived ADO on the acute immune response against Trypanosoma cruzi parasite, which invades and proliferates within different target tissues. Deficiency of CD73 activity led to an enhanced cardiac microbicidal immune response with an augmented frequency of macrophages with inflammatory phenotype and increased CD8+ T cell effector functions. The increment of local inducible nitric oxide (NO) synthase (iNOS)+ macrophages and the consequent rise of myocardial NO production in association with reduced ADO levels induced protection against T. cruzi infection as observed by the diminished cardiac parasite burden compared to their wild-type (WT) counterpart. Unexpectedly, parasitemia was substantially raised in CD73KO mice in comparison with WT mice, suggesting the existence of tissue reservoir/s outside myocardium. Indeed, CD73KO liver and visceral adipose tissue (VAT) showed increased parasite burden associated with a reduced ATP/ADO ratio and the lack of substantial microbicidal immune response. These data reveal that the purinergic system has a tissue-dependent impact on the host immune response against T. cruzi infection.

Helminth Infections: Recognition and Modulation of the Immune Response by Innate Immune Cells.

The survival of helminths in the host over long periods of time is the result of a process of adaptation or dynamic co-evolution between the host and the parasite. However, infection with helminth parasites causes damage to the host tissues producing the release of danger signals that induce the recruitment of various cells, including innate immune cells such as macrophages (Mo), dendritic cells (DCs), eosinophils, basophils, and mast cells. In this scenario, these cells are able to secrete soluble factors, which orchestrate immune effector mechanisms that depend on the different niches these parasites inhabit. Here, we focus on recent advances in the knowledge of excretory-secretory products (ESP), resulting from helminth recognition by DCs and Mo. Phagocytes and other cells types such as innate lymphocyte T cells 2 (ILC2), when activated by ESP, participate in an intricate cytokine network to generate innate and adaptive Th2 responses. In this review, we also discuss the mechanisms of innate immune cell-induced parasite killing and the tissue repair necessary to assure helminth survival over long periods of time.

Microscopio and PCR-based detection of microsporidia spores in human stool samples

Abstract Microsporidia are obligate intracellular fungi with a remarkable ability to infect a wide range of invertebrate and vertebrate hosts. Namely, Enterocytozoon bieneusi is the most frequently microsporidia reported worldwide, and mainly associated with chronic diarrea and wasting syndrome in AIDS patients. Microscopy and PCR-based detection techniques are effective for diagnosis and identification of species and genotypes; however, these methods should be standardized in each laboratory. In this study, we performed microscopy and nested PCR techniques with PCR product sequencing to detect E. bieneusi in human stool samples. These techniques, if applied together, might prove useful for diagnosis and future epidemiological studies of intestinal microsporidiosis in Argentina.

Resumen Los microsporidios son hongos intracelulares obligados con una notable capacidad para infectar una amplia gama de hospedadores invertebrados y vertebrados. Enterocytozoon bieneusi es el microsporidio más frecuentemente reportado en todo el mundo, principalmente tricrómicaasociado con diarrea crónica y síndrome debilitante en pacientes con sida. Las técnicas dedetección basadas en microscopía y PCR son útiles para el diagnóstico y la identificación deespecies y genotipos, pero estos métodos deben estar estandarizados en cada laboratorio.En este estudio evaluamos técnicas de microscopía y PCR anidada, con secuenciación de losproductos, para detectar E. bieneusi en muestras de heces humanas. Estas técnicas, usadas con-juntamente, podrían ser útiles para su aplicación en el diagnóstico de microsporidiosis intestinaly para realizar estudios epidemiológicos de esta afección en Argentina.