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Resumen Muchos medicamentos y vacunas han tenido implicaciones en el desarrollo de sintomas osteomusculares y articulares. Entre ellos, el uso de retinoides se ha asociado con el desarrollo de sintomas musculoesqueleticos, asi como sintomas axiales sugestivos de espondiloartritis con sacroileitis, y en menor proporcion el desarrollo de sintomas perifericos. Describimos el inicio de sintomas inflamatorios perifericos con el uso de isotretinoina en una paciente previamente sana.

Background Infectious bursal disease (IBD), also known as Gumboro disease, is a viral infection that causes mortality and immunosuppression in chickens (Gallus gallus). VP2 and VP3 are the major structural viral capsid components and are the most immunogenic proteins of IBD virus (IBDV). Reliable diagnostic tests using VP2 and VP3 produced in heterologous systems are important tools to control this infection. One advantage of an IBD diagnostic based on VP3, over those that use VP2, is that VP3 has linear epitopes, enabling its production in bacteria. Results We tested the suitability of recombinant VP3 (rVP3) as a diagnostic reagent in an enzyme-linked immunosorbent assay (ELISA). Compared with a commercial test, rVP3 ELISA showed high sensitivity and specificity as a diagnostic tool for vaccinated animals. In addition, rVP3, but not the commercial ELISA, was able to detect antibodies in nonvaccinated chickens, probably developed against circulating IBDV strains. It was possible the assessment of VP3 regions antigenicity using chicken antisera. Conclusions The full-length recombinant VP3 can be used to assess post vaccination immunological status of chickens and its production is feasible and inexpensive. The evaluation of VP3 regions as candidates for general use in the diagnosis of IBD in chickens should be conducted with caution. Our work was the first to identify several regions of VP3 recognized by chicken antibodies.

A avicultura é um setor de grande importância na economia brasileira tanto pelo aumento do consumo interno quanto pelo crescimento na exportação de carne de frango. A modernização da atividade avícola permitiu a criação adensada de animais, facilitando, no entanto, a rápida disseminação de patógenos que reduzem os índices de produtividade dos plantéis. Nesta revisão, é destacado o reovírus aviário, importante agente de artrite em aves que apresenta distribuição mundial. As aves acometidas apresentam redução no ganho de peso devido à dificuldade de locomoção. Além da artrite, o vírus pode estar relacionado a uma variedade de condições patológicas, como distúrbios entéricos e respiratórios, hepatite e miocardite. A principal forma de prevenção e controle é a vacinação do plantel. No entanto, devido à grande variabilidade genética do reovírus aviário, a vacina utilizada pode não ser eficiente contra estirpes que circulam no campo. O artigo traz uma visão geral sobre a biologia do vírus, sua variabilidade e propostas de classificação dos isolados, patologia da doença e diagnóstico da infecção.

Colorectal cancer (CRC) is one of the most common cancers worldwide. As with other cancers, CRC is a multifactorial disease due to the combined effect of genetic and environmental factors. Most cases are sporadic, but a small proportion is hereditary, estimated at around 5-10%. In both, the tumor interacts with heterogeneous cell populations, such as endothelial, stromal, and immune cells, secreting different signals (cytokines, chemokines or growth factors) to generate a favorable tumor microenvironment for cancer cell invasion and metastasis. There is ample evidence that inflammatory processes have a role in carcinogenesis and tumor progression in CCR. Different profiles of cell activation of the tumor microenvironment can promote pro or anti-tumor pathways; hence they are studied as a key target for the control of cancer progression. Additionally, the intestinal mucosa is in close contact with a microorganism community, including bacteria, bacteriophages, viruses, archaea, and fungi composing the gut microbiota. Aberrant composition of this microbiota, together with alteration in the diet‐derived microbial metabolites content (such as butyrate and polyamines) and environmental compounds has been related to CRC. Some bacteria, such as pks+ Escherichia coli or Fusobacterium nucleatum, are involved in colorectal carcinogenesis through different pathomechanisms including the induction of genetic mutations in epithelial cells and modulation of tumor microenvironment. Epithelial and immune cells from intestinal mucosa have Pattern-recognition receptors and G-protein coupled receptors (receptor of butyrate), suggesting that their activation can be regulated by intestinal microbiota and metabolites. In this review, we discuss how dynamics in the gut microbiota, their metabolites, and tumor microenvironment interplays in sporadic and hereditary CRC, modulating tumor progression.

Additional file 6. Oligonucleotides for PCR assays based on the codon-optimized VP3 sequence. In bold are the targets for restriction endonucleases: Xho I - CTCGAG and EcoR I – GAATTC.

Additional file 1. Recombinant VP3 ELISA optimization. (A) 1:10,000 and (B) 1:40,000 secondary antibodiy dilutions. Negative control: 1; antisera samples: 2 to 8; antisera dilutions: 1:100 (blue bars) and 1:500 (orange bars); antigen concentrations: 0.25, 0.5, 1.0 and 2.0 μg/mL; gray bars: commercial ELISA.

BACKGROUND Trypanosoma cruzi is an important human pathogen in Latin America with nearly seven million people infected. It has a large degree of genetic diversity, classified into six discrete typing units (DTUs), which probably influences its physiological behavior and clinical manifestations. Several genotyping methods are available, with distinct performance on easiness, cost, resolution and applicability; no method excels in all parameters. OBJECTIVES AND METHODS To devise a molecular method for T. cruzi genotyping, based on polymerase chain reaction (PCR) amplification of a single target with multiple copies in the nuclear genome by large scale sequencing. We have applied this method to 29 T. cruzi isolates, comprising all described DTUs. FINDINGS We were able to classify all samples into sub DTU level with high robustness. Evolutionary relationship between DTUs were ascertained, suggesting that TcIII and TcIV DTUs are non-hybrid, and DTU IV is more similar to the common ancestral. CONCLUSION As the TS-LSS method is based on a single PCR reaction, comprising several copies of the target, it is probably useful for clinical samples, when the amount of DNA is a limiting factor. As large scale sequencing systems become more common, the TS-LSS method can be increasingly applied for T. cruzi genotyping.

Additional file 2. Evaluation of the rVP3 ELISA. (A) rVP3 ELISA A/P values of samples considered negative (left) and positive (right) by the commercial test. The horizontal line represents the cut-off 0.0496. (B) Correlation (R2=0.24) between commercial ELISA (y-axis) and rVP3 ELISA (x-axis) O.D. values. A/P = sample O.D. mean – negative control O.D. mean/ positive control O.D. mean – negative control O.D. mean.

The polyphenol nordihydroguaiaretic acid (NDGA) has antineoplastic properties, hence it is critical to understand its action at the molecular level. Here, we establish that NDGA inhibits glucose uptake and cell viability in leukemic HL-60 and U-937 cell lines. We monitored hexose uptake using radio-labeled 2-deoxyglucose (2DG) and found that the inhibition by NDGA followed a noncompetitive mechanism. In addition, NDGA blocked hexose transport in human red blood cells and displaced prebound cytochalasin B from erythrocyte ghosts, suggesting a direct interaction with the glucose transporter GLUT1. We propose a model for the mechanism of action of NDGA on glucose uptake. Our study shows for the first time that NDGA can act as inhibitor of the glucose transporter GLUT1.

Crohn’s disease (CD) is a chronic inflammatory bowel disorder characterized by deregulated inflammation triggered by environmental factors. Notably, adherent-invasive Escherichia coli (AIEC), a bacterium with the ability to survive within macrophages is believed to be one of such factors. Glucocorticoids are the first line treatment for CD and to date, it is unknown how they affect bactericidal and inflammatory properties of macrophages against AIEC. The aim of this study was to evaluate the impact of glucocorticoid treatment on AIEC infected macrophages. First, THP-1 cell-derived macrophages were infected with a CD2-a AIEC strain, in the presence or absence of the glucocorticoid dexamethasone (Dex) and mRNA microarray analysis was performed. Differentially expressed mRNAs were confirmed by TaqMan-qPCR. In addition, an amikacin protection assay was used to evaluate the phagocytic and bactericidal activity of Dex-treated macrophages infected with E. coli strains (CD2-a, HM605, NRG857c, and HB101). Finally, cytokine secretion and the inflammatory phenotype of macrophages were evaluated by ELISA and flow cytometry, respectively. The microarray analysis showed that CD2-a, Dex, and CD2-a + Dex-treated macrophages have differential inflammatory gene profiles. Also, canonical pathway analysis revealed decreased phagocytosis signaling by Dex and anti-inflammatory polarization on CD2-a + Dex macrophages. Moreover, amikacin protection assay showed reduced phagocytosis upon Dex treatment and TaqMan-qPCR confirmed Dex inhibition of three phagocytosis-associated genes. All bacteria strains induced TNF-α, IL-6, IL-23, CD40, and CD80, which was inhibited by Dex. Thus, our data demonstrate that glucocorticoids impair phagocytosis and induce anti-inflammatory polarization after AIEC infection, possibly contributing to the survival of AIEC in infected CD patients.

BACKGROUND Trypanosoma cruzi is an important protozoan parasite and the causative agent of Chagas disease. A critical step in understanding T. cruzi biology is the study of cellular and molecular features exhibited during its growth curve. OBJECTIVES We aimed to acquire a global view of the gene expression profile of T. cruzi during epimastigote growth. METHODS RNA-Seq analysis of total and polysomal/granular RNA fractions was performed along the 10 days T. cruzi epimastigote growth curve in vitro, in addition to cell viability and cell cycle analyses. We also analysed the polysome profile and investigated the presence of granular RNA by FISH and western blotting. FINDINGS We identified 1082 differentially expressed genes (DEGs), of which 220 were modulated in both fractions. According to the modulation pattern, DEGs were grouped into 12 clusters and showed enrichment of important gene ontology (GO) terms. Moreover, we showed that by the sixth day of the growth curve, polysomal content declined greatly and the RNA granules content appeared to increase, suggesting that a portion of mRNAs isolated from the sucrose gradient during late growth stages was associated with RNA granules and not only polyribosomes. Furthermore, we discuss several modulated genes possibly involved in T. cruzi growth, mainly during the stationary phase, such as genes related to cell cycle, pathogenesis, metabolic processes and RNA-binding proteins.

Myosins are motor proteins that comprise a large and diversified family important for a broad range of functions. Two myosin classes, I and XIII, were previously assigned in Trypanosomatids, based mainly on the studies of Trypanosoma cruzi, T. brucei and Leishmania major, and important human pathogenic species; seven orphan myosins were identified in T. cruzi. Our results show that the great variety of T. cruzi myosins is also present in some closely related species and in Bodo saltans, a member of an early divergent branch of Kinetoplastida. Therefore, these myosins should no longer be considered “orphans”. We proposed the classification of a kinetoplastid-specific myosin group into a new class, XXXVI. Moreover, our phylogenetic data suggest that a great repertoire of myosin genes was present in the last common ancestor of trypanosomatids and B. saltans, mainly resulting from several gene duplications. These genes have since been predominantly maintained in synteny in some species, and secondary losses explain the current distribution. We also found two interesting genes that were clearly derived from myosin genes, demonstrating that possible redundant or useless genes, instead of simply being lost, can serve as raw material for the evolution of new genes and functions.

Brain aromatase is a key enzyme exclusively expressed in fish radial glial cells that convert androgens into estrogens, thus controlling neuroendocrine functions and neurogenesis. As an important step in characterizing the neuroendocrine systems of Rhamdia quelen (jundia), a partial cDNA sequence (1045 bp) of brain aromatase (cyp19a1b) was cloned and sequenced. At the nucleotide level the cDNA sequence was found to be 88% identical to cyp19a1b of two species of catfish, Ictalurus punctatus and Silurus meridionalis. The predicted amino acid sequence was between 80 and 91% similar to other teleosts. Real-time RT-qPCR analysis revealed that cyp19a1b was detected in pituitary, hypothalamus, telencephalon, head and posterior kidneys, liver and gonads (testis and ovary) of both males and females. The effects of E2 on cyp19a1b expression are sexually dimorphic in R. quelen. The injection of 17β-estradiol (E2) decreased head kidney mRNA levels of cyp19a1b in males and increased cyp19a1b mRNA levels in the pituitary and head kidney of females. This study demonstrated that the R. quelen cyp19a1b gene is expressed in brain, pituitary and peripheral tissues in both males and females.

The protozoan Trypanosoma cruzi is a parasite exposed to several environmental stressors inside its invertebrate and vertebrate hosts. Although stress conditions are involved in its differentiation processes, little information is available about the stress response proteins engaged in these activities. This work reports the first known association of the stress-inducible protein 1 (STI1) with the cellular differentiation process in a unicellular eukaryote. Albeit STI1 expression is constitutive in epimastigotes and metacyclic trypomastigotes, higher protein levels were observed in late growth phase epimastigotes subjected to nutritional stress. Analysis by indirect immunofluorescence revealed that T. cruzi STI1 (TcSTI1) is located throughout the cell cytoplasm, with some cytoplasmic granules appearing in greater numbers in late growing epimastigotes and late growing epimastigotes subjected to nutritional stress. We observed that part of the fluorescence signal from both TcSTI1 and TcHSP70 colocalized around the nucleus. Gene silencing of sti1 in Trypanosoma brucei did not affect cell growth. Similarly, the growth of T. cruzi mutant parasites with a single allele sti1 gene knockout was not affected. However, the differentiation of epimastigotes in metacyclic trypomastigotes (metacyclogenesis) was compromised. Lower production rates and numbers of metacyclic trypomastigotes were obtained from the mutant parasites compared with the wild-type parasites. These data indicate that reduced levels of TcSTI1 decrease the rate of in vitro metacyclogenesis, suggesting that this protein may participate in the differentiation process of T. cruzi.

BACKGROUND Real-time reverse transcription polymerase chain reaction (RT-PCR) is routinely used to detect viral infections. In Brazil, it is mandatory the use of nucleic acid tests to detect hepatitis C virus (HCV), hepatitis B virus and human immunodeficiency virus in blood banks because of the immunological window. The use of an internal control (IC) is necessary to differentiate the true negative results from those consequent from a failure in some step of the nucleic acid test. OBJECTIVES The aim of this study was the construction of virus-modified particles, based on MS2 bacteriophage, to be used as IC for the diagnosis of RNA viruses. METHODS The MS2 genome was cloned into the pET47b(+) plasmid, generating pET47b(+)-MS2. MS2-like particles were produced through the synthesis of MS2 RNA genome by T7 RNA polymerase. These particles were used as non-competitive IC in assays for RNA virus diagnostics. In addition, a competitive control for HCV diagnosis was developed by cloning a mutated HCV sequence into the MS2 replicase gene of pET47b(+)-MS2, which produces a non-propagating MS2 particle. The utility of MS2-like particles as IC was evaluated in a one-step format multiplex real-time RT-PCR for HCV detection. FINDINGS We demonstrated that both competitive and non-competitive IC could be successfully used to monitor the HCV amplification performance, including the extraction, reverse transcription, amplification and detection steps, without compromising the detection of samples with low target concentrations. In conclusion, MS2-like particles generated by this strategy proved to be useful IC for RNA virus diagnosis, with advantage that they are produced by a low cost protocol. An attractive feature of this system is that it allows the construction of a multicontrol by the insertion of sequences from more than one pathogen, increasing its applicability for diagnosing different RNA viruses.

Trypanosoma cruzi is the etiologic agent of Chagas disease, which is estimated to affect over eight million people around the world. Trypanosoma cruzi has a complex life cycle, involving insect and mammalian hosts and four distinct developmental stages: epimastigotes, metacyclic trypomastigotes, amastigotes, and bloodstream trypomastigotes. Metacyclogenesis is the process by which T. cruzi epimastigotes differentiate into metacyclic trypomastigotes and acquire infectivity, and involves differential gene expression associated with acquisition of virulence. In T. cruzi, gene expression regulation is achieved mainly posttranscriptionally. Therefore, proteomics-based approaches are extremely useful for gaining a better understanding of the changes that occur in the stage-regulated gene expression program of the parasite at the molecular level. Here, we performed an in-depth quantitative MS-based proteomic study of T. cruzi metacyclogenesis and quantified almost 3000 proteins expressed during the process of differentiation. To the best of our knowledge, this work is the most comprehensive quantitative proteomics study of different cell populations of T. cruzi available so far. We identified relevant proteins and pathways involved in the parasite's differentiation and infectivity acquisition, opening new perspectives for further studies that could, ultimately, lead to the identification of new targets for chemotherapy.

CD28 and cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) are two receptors that have complementary functions in control of T-cell activation. Polymorphisms of their genes, CD28 and CTLA4, might confer differential susceptibility to diseases resulting from unbalanced or inefficient immune responses. Thus far, little is known about the CD28 polymorphism in populations and even for CTLA4 just one or two single nucleotide polymorphisms (SNPs) are usually analysed. To assess the allelic and haplotypic diversity and linkage disequilibrium in the Brazilian population, two samples differing according to predominant ancestry - African or European - have been analysed for seven SNPs, CD28 -372(G>A), and int3 17(T>C); CTLA4 -1722(T>C), -1577(G>A) -318(C>T), 49(A>G), 6230(G>A) also named CT60, and three microsatellites, CD28 (CAA)n, CTLA4 (AT)n and D2S72 (CA)n. The two population strata show little differentiation, the only significant differences being the allele frequencies of the CTLA4 -1577(G>A) SNP and the CTLA4 (AT)n microsatellite (P = 0.018 and P = 0.007, respectively). Linkage disequilibrium is high, especially between the CTLA4 polymorphisms. However, low r(2) values indicate that none of the markers is a tag SNP in these populations. These results provide valuable information for optimal selection of markers for use in future association studies. We conclude that disease association studies and functional studies addressing the possible consequences of polymorphisms of the 2q33 genomic region should consider haplotypic data besides analysis of individual polymorphisms.

The strategy used to treat HCV infection depends on the genotype involved. An accurate and reliable genotyping method is therefore of paramount importance. We describe here, for the first time, the use of a liquid microarray for HCV genotyping. This liquid microarray is based on the 5′UTR — the most highly conserved region of HCV — and the variable region NS5B sequence. The simultaneous genotyping of two regions can be used to confirm findings and should detect inter-genotypic recombination. Plasma samples from 78 patients infected with viruses with genotypes and subtypes determined in the Versant™ HCV Genotype Assay LiPA (version I; Siemens Medical Solutions, Diagnostics Division, Fernwald, Germany) were tested with our new liquid microarray method. This method successfully determined the genotypes of 74 of the 78 samples previously genotyped in the Versant™ HCV Genotype Assay LiPA (74/78, 95%). The concordance between the two methods was 100% for genotype determination (74/74). At the subtype level, all 3a and 2b samples gave identical results with both methods (17/17 and 7/7, respectively). Two 2c samples were correctly identified by microarray, but could only be determined to the genotype level with the Versant™ HCV assay. Genotype “1” subtypes (1a and 1b) were correctly identified by the Versant™ HCV assay and the microarray in 68% and 40% of cases, respectively. No genotype discordance was found for any sample. HCV was successfully genotyped with both methods, and this is of prime importance for treatment planning. Liquid microarray assays may therefore be added to the list of methods suitable for HCV genotyping. It provides comparable results and may readily be adapted for the detection of other viruses frequently co-infecting HCV patients. Liquid array technology is thus a reliable and promising platform for HCV genotyping.

The purpose of this study was to analyze the possible influence of the TNF and LTA loci polymorphisms on the susceptibility/resistance to endemic pemphigus foliaceus, also named fogo selvagem (FS), an autoimmune disease characterized by blisters due to acantholysis of the superficial-most epidermal cells. Autoantibodies, mainly of the IgG4 subclass, are directed against a desmosomal glycoprotein known as desmoglein 1. FS shares clinical, histological and immunological features with nonendemic pemphigus foliaceus. Most residents of the endemic regions do not develop the disease, and familial clustering has been documented, suggesting that host factors play a role in susceptibility. In fact, strong positive and negative associations with HLA class II genes have been reported. The TNF and LTA genes are located in the class III region of the Human Major Histocompatibility Complex. Their location, the function of their products, which are cytokines and pluripotent immunomodulators, as well as their genetic variability make them candidate genes for complex diseases with an altered immune response. A total of 162 patients and 191 controls were enrolled in this study. No significant associations were found with any one of the three LTA single nucleotide polymorphisms (SNP) analyzed (at nucleotides 249, 365, 720), nor with the TNF SNP located at positions −863 and −308. The frequency of allele TNF*238A was slightly decreased in patients (OR = 0.45). In conclusion, the results of this study indicate that genetic variability of the TNF and LTA genes does not play a major role in susceptibility/resistance to pemphigus foliaceus.

Endemic pemphigus foliaceus (EPF) is a blistering skin disease characterized by autoantibodies against the desmosomal protein desmoglein 1. Genetic and environmental factors influence its pathogenesis. A total of 128 patients and 402 controls from an ethnically admixed Brazilian population were analyzed for associations by allele and genotype with HLA-DRB1. The alleles DRB1(*)0101, (*)0102, (*)0103, (*)0404, (*)0406, (*)0410, (*)1406 and (*)1601 are significantly more frequent among patients, while DRB1(*)0301, (*)0701, (*)0801, (*)1101, (*)1104 and (*)1402 are negatively associated to EPF. Results of association analysis with protein motifs composed of polymorphic amino-acid residues do not add much to comprehension of the molecular basis of the HLA-DRB1/EPF associations. Interactions between susceptible (SU), protective (PR) and neutral (NE) alleles clearly deviate from the codominant model. Protection is dominant, since the PR/NE and PR/PR genotypes are both equally (P=0.95) and strongly protective (odds ratio OR=0.07 and 0.05, respectively; P

International audience; Endosymbiont-bearing trypanosomatids have been considered excellent models for the study of cell evolution because the host protozoan co-evolves with an intracellular bacterium in a mutualistic relationship. Such protozoa inhabit a single invertebrate host during their entire life cycle and exhibit special characteristics that group them in a particular phylogenetic cluster of the Trypanosomatidae family, thus classified as monoxenics. In an effort to better understand such symbiotic association, we used DNA pyrosequencing and a reference-guided assembly to generate reads that predicted 16,960 and 12,162 open reading frames (ORFs) in two symbiont-bearing trypanosomatids, Angomonas deanei (previously named as Crithidia deanei) and Strigomonas culicis (first known as Blastocrithidia culicis), respectively. Identification of each ORF was based primarily on TriTrypDB using tblastn, and each ORF was confirmed by employing getorf from EMBOSS and Newbler 2.6 when necessary. The monoxenic organisms revealed conserved housekeeping functions when compared to other trypanosomatids, especially compared with Leishmania major. However, major differences were found in ORFs corresponding to the cytoskeleton, the kinetoplast, and the paraflagellar structure. The monoxenic organisms also contain a large number of genes for cytosolic calpain-like and surface gp63 metalloproteases and a reduced number of compartmentalized cysteine proteases in comparison to other TriTryp organisms, reflecting adaptations to the presence of the symbiont. The assembled bacterial endosymbiont sequences exhibit a high A+T content with a total of 787 and 769 ORFs for the Angomonas deanei and Strigomonas culicis endosymbionts, respectively, and indicate that these organisms hold a common ancestor related to the Alcaligenaceae family. Importantly, both symbionts contain enzymes that complement essential host cell biosynthetic pathways, such as those for amino acid, lipid and purine/pyrimidine metabolism. These findings increase our understanding of the intricate symbiotic relationship between the bacterium and the trypanosomatid host and provide clues to better understand eukaryotic cell evolution.

Trypanosoma cruzi is the causative agent of Chagas disease, a neglected disorder that affects millions of people in the Americas. T. cruzi relies mostly upon post-transcriptional regulation to control stage specific gene expression. RNA binding proteins (RBPs) associate with functionally related mRNAs forming ribonucleoprotein complexes that define post-transcriptional operons. The RNA Recognition Motif (RRM) is the most common and ancient family of RBPs. This family of RBPs has been identified in trypanosomatid parasites and only a few of them have been functionally characterized. We describe here the functional characterization of TcRBP40, a T. cruzi specific RBP, and its associated mRNAs. We used a modified version of the recombinant RIP-Chip assay to identify the mRNAs with which it associates and in vivo TAP-tag assays to confirm these results. TcRBP40 binds to an AG-rich sequence in the 3′UTR of the associated mRNAs, which were found to encode mainly putative transmembrane proteins. TcRBP40 is differentially expressed in metacyclogenesis. Surprisingly, in epimastigotes, it is dispersed in the cytoplasm but is concentrated in the reservosomes, a T. cruzi specific organelle, which suggests a putative new function for this parasite organelle.

Background The experimental murine model of leishmaniasis has been widely used to characterize the immune response against Leishmania. CBA mice develop severe lesions, while C57BL/6 present small chronic lesions under L. amazonensis infection. Employing a transcriptomic approach combined with biological network analysis, the gene expression profiles of C57BL/6 and CBA macrophages, before and after L. amazonensis infection in vitro, were compared. These strains were selected due to their different degrees of susceptibility to this parasite. Results The genes expressed by C57BL/6 and CBA macrophages, before and after infection, differ greatly, both with respect to absolute number as well as cell function. Uninfected C57BL/6 macrophages express genes involved in the deactivation pathway of macrophages at lower levels, while genes related to the activation of the host immune inflammatory response, including apoptosis and phagocytosis, have elevated expression levels. Several genes that participate in the apoptosis process were also observed to be up-regulated in C57BL/6 macrophages infected with L. amazonensis, which is very likely related to the capacity of these cells to control parasite infection. By contrast, genes involved in lipid metabolism were found to be up-regulated in CBA macrophages in response to infection, which supports the notion that L. amazonensis probably modulates parasitophorous vacuoles in order to survive and multiply in host cells. Conclusion The transcriptomic profiles of C57BL/6 macrophages, before and after infection, were shown to be involved in the macrophage pathway of activation, which may aid in the control of L. amazonensis infection, in contrast to the profiles of CBA cells.

The life cycle of the protozoan Trypanosoma cruzi exposes it to several environmental stresses in its invertebrate and vertebrate hosts. Stress conditions are involved in parasite differentiation, but little is known about the stress response proteins involved. We report here the first characterization of stress-induced protein-1 (STI-1) in T. cruzi (TcSTI-1). This co-chaperone is produced in response to stress and mediates the formation of a complex between the stress proteins HSP70 and HSP90 in other organisms. Despite the similarity of TcSTI-1 to STI-1 proteins in other organisms, its expression profile in response to various stress conditions, such as heat shock, acidic pH or nutrient starvation, is quite different. Neither polysomal mRNA nor protein levels changed in exponentially growing epimastigotes cultured under any of the stress conditions studied. Increased levels of TcSTI-1 were observed in epimastigotes subjected to nutritional stress in the late growth phase. Co-immunoprecipitation assays revealed an association between TcSTI-1 and TcHSP70 in T. cruzi epimastigotes. Immunolocalization demonstrated that TcSTI-1 was distributed throughout the cytoplasm and there was some colocalization of TcSTI-1 and TcHSP70 around the nucleus. Thus, TcSTI-1 associates with TcHSP70 and TcSTI-1 expression is induced when the parasites are subjected to stress conditions during specific growth phase.

Calpains are calcium-dependent cysteine proteinases found in all living organisms and are involved in diverse cellular processes. Calpain-like proteins have been reported after in silico analysis of the Tritryps genome and are believed to play important roles in cell functions of trypanosomatids. We describe the characterization of a member of this family, which is differentially expressed during the life-cycle of Trypanosoma cruzi.