Maternal distress, DNA methylation, and fetal programing of stress physiology in Brazilian mother-infant pairs.

Maternal prenatal psychosocial stress is associated with adverse hypothalamic-pituitary-adrenal axis (HPAA) function among infants. Although the biological mechanisms influencing this process remain unknown, altered DNA methylation is considered to be one potential mechanism. We investigated associations between maternal prenatal psychological distress, infant salivary DNA methylation, and stress physiology at 12 months. Mother's distress was measured via depression and anxiety in early and late pregnancy in a cohort of 80 pregnant adolescents. Maternal hair cortisol was collected during pregnancy. Saliva samples were collected from infants at 12 months to quantify DNA methylation of three stress-related genes (FKBP5, NR3C1, OXTR) (n = 62) and diurnal cortisol (n = 29). Multivariable linear regression was used to test for associations between prenatal psychological distress, and infant DNA methylation and cortisol. Hair cortisol concentrations in late pregnancy were negatively associated with two sites of FKBP5 (site 1: B = -22.33, p = .003; site 2: B = -15.60, p = .012). Infants of mothers with elevated anxiety symptoms in late pregnancy had lower levels of OXTR2 CpG2 methylation (B = -2.17, p = .03) and higher evening salivary cortisol (B = 0.41, p = .03). Furthermore, OXTR2 methylation was inversely associated with evening cortisol (B = -0.14, p-value ≤ .001). Our results are, to our knowledge, the first evidence that the methylation of the oxytocin receptor may contribute to the regulation of HPAA during infancy.

Impact of polymorphisms in blaZ, blaR1 and blaI genes and their relationship with ß-lactam resistance in S. aureus strains isolated from bovine mastitis.

There is not a consensus between the presence of the genotypic resistance marker gene and the phenotypic resistance to ß-lactams in Staphylococcus aureus, which means, positive S. aureus blaZ isolates demonstrating sensitivity to ß-lactams. The present study aimed to characterize the blaZ, blaR1 and blaI genes, identify and evaluate single nucleotide polymorphisms (SNPs) and their relationship with ß-lactam resistance in samples of Staphylococcus aureus obtained from cases of bovine mastitis. Five isolates (two resistant and three sensitive to oxacillin) of Staphylococcus aureus with detected production of beta-lactamase, previously evaluated as containing the blaZ gene and negative for the mecA and mecC genes, had the bla operon completely sequenced. Impacts on the protein sequence due to the detected polymorphisms were evaluated by modeling the proteins encoded by the blaZ, blaR1 and blaI genes using a three-dimensional model structure obtained from the Protein Data Bank (PDB) database. Fifteen SNPs were detected in the blaZ gene, 30 in the blaR1 gene and three in the blaI gene. These SNPs caused alterations in amino acid sites. Deleterious mutations were detected in the blaZ gene (E146G, P218S, Y221C) and the blaR1 gene (K481E). Molecular docking analysis revealed that polymorphisms in the blaZ gene may explain the phenotypic sensitivity in isolates that contain the resistance marker gene. Although sensitive and resistant isolates encode beta-lactamase, these proteins are functionally altered due to a change in the binding site with the antibiotic.

Characterization of Multidrug-Resistant Staphylococcus aureus Isolates and Comparison of Methods of Susceptibility to Vancomycin.

S. aureus are among the main bacteria causing problems related to multidrug resistance in nosocomial infections. Therefore, it is necessary to carry out a reliable and rapid diagnosis for the identification of the bacteria and characterization of its susceptibility profile, especially vancomycin, which is an alternative treatment against multidrug-resistant (MDR) S. aureus. Thus, the goal of this study was to characterize isolates of S. aureus regarding the resistance and virulence and to check the susceptibility to vancomycin, through different methods, for comparative purposes. Seventeen antimicrobials were tested to assess the susceptibility profile. It was evaluated the presence of identification (nuc), resistance (mecA and blaZ), biofilm (icaA and icaD) and siderophore (sfaD and sbnD) genes. The susceptibility to vancomycin was evaluated by Minimum Inhibitory Concentration (MIC) by broth microdilution (BMD), E-test, commercial panel (Kit), and Phoenix equipment. Most S. aureus (93,33%) was classified as MDR. These isolates were 100% positive for nuc, mecA, icaA, icaD, and sfaD genes; 96.67% for sbnD and 33.33% for blaZ. In relation to BMD, all methods correctly classified the susceptibility of the isolates; however, regarding the exact MIC value for vancomycin, Phoenix showed agreement of 63.33%, E-test (33.33%) and Kit (26.66%). In conclusion, most of S. aureus was considered MDR. Also, they presented resistance, biofilm production, and siderophores genes, showing the pathogenic potential of these bacteria. Besides, the Phoenix test was considered the most effective, as it presents advantages, such as identification of the microorganism and a greater number of antimicrobials tested at a time.

Overexpression of OCT-1 gene is a biomarker of adverse prognosis for diffuse large B-cell lymphoma (DLBCL): data from a retrospective cohort of 77 Brazilian patients.

BACKGROUND: OCT-1 gene is a member of the POU-homeodomain family of transcriptional regulators of B-lymphocyte differentiation by controlling expression of B-cell specific genes. BCL-2 gene is a potent inhibitor of apoptosis and it is essential during B-cell differentiation into germinal center. These genes may be expressed in diffuse large B-cell lymphoma (DLBCL), but the role of BCL-2 in its prognosis has been contradictory, and OCT-1 has yet to be tested. METHODS: In this study, we aimed to investigate the prognostic impact of OCT-1 and BCL-2 expression in DLBCL treated in the real world with immunochemotherapy in a single center. BCL-2 and OCT-1 genes were available in 78.5% (77/98) DLBCL patients, and the RNA for quantitative real-time PCR was isolated from formalin-fixed paraffin-embedded samples. The values obtained for gene expression were transformed in categorical variable according to their median. RESULTS: Cohort median age was 54.5 years (15-84), 49 (50%) were male, 38/77 (49.4%) and 40/77 (51.9%) presented OCT-1 and BCL-2 expression ≥ median, respectively. The overall response rate (ORR) in all patients was 68.4% (67/98), 65,3% (64/98) of patients acquired complete response, and 3.1% (3/98) partial response, while 6.1% (6/98) were primary refractory. The median follow-up was 3.77 years (95% CI: 3.2-4.1), with 5.43 (95% CI: 2.2-NR) of overall survival (OS) and 5.15 years (95% CI: 2.9-NA) of progression free survival (PFS). OCT-1 ≥ median was associated with shorter OS at univariate analysis (p = 0.013; [HR] 2.450, 95% CI: 1.21-4.96) and PFS (p = 0.019; [HR] 2.270, 95%CI: 1.14-4.51) and BCL-2 gene overexpression presented worse PFS (p = 0.043, [HR] 2.008, 95% CI: 1.02-3.95). At multivariate analysis, OCT-1 overexpression was associated with poor PFS (p = 0.035, [HR] 2.22, 95% CI: 1.06-4.67). CONCLUSION: In this study, we showed that overexpression of OCT1 gene was an independent prognostic factor of adverse outcomes in DLBCL.

Nutritional interference for phenotypic biofilm quantification in Aeromonas spp. isolates containing the fla gene.

The Aeromonas genus has several virulence factors associated with the development of diseases in aquatic organisms, leading to losses in aquaculture. One of these factors is the flagella's formation which allows the biofilm's formation that provides the microorganisms a greater pathogenicity, greater protection to certain substances such as antibiotics. The aim of the study was to verify the presence of the fla gene, related to biofilm production in isolates of Aeromonas spp. from fishes and also to determine the best quantification condition of phenotypic biofilm production in vitro. Polymerase Chain Reactions were performed to obtain the amplification of the region comprising the fla gene. To determine the best condition for the production biofilm, the microplate adhesion test was carried out under different concentrations of TSB broth and it combined with glucose. Of the 43 isolates of Aeromonas spp. analyzed, 28 were positive for the fla gene and, in the quantification of the biofilm, all these were able to form biofilm in the TSB broth without dilution and without addition of glucose, being this the best condition tested. It was observed that the isolates of Aeromonas spp. analyzed have potential for biofilm formation, and hence potential for virulence.

Resistance to antimicrobials and biofilm formation in Staphylococcus spp. isolated from bovine mastitis in the Northeast of Brazil.

Mastitis is the principal disease affecting dairy herds worldwide. The aim of the present study was to characterize phenotypic and genotypic features associated with resistance to antimicrobials in Staphylococcus spp. isolated from 2064 milk samples of 525 lactating cows in the Northeast of Brazil. Of the 218 isolates analyzed, 57.8% were characterized as Staphylococcus aureus, 28% as coagulase-positive staphylococci other than S. aureus (oCPS), and 14.2% as coagulase-negative staphylococci (CNS). The test for susceptibility to antimicrobials showed amoxicillin (32.6%) to be the less effective drug in vitro, and the multi-drug resistance (MDR) rate for beta-lactams varied from 0 to 0.75. The genotypic characterization showed that 93.1% of the samples were tested positive for the blaZ gene, while none amplified mecA. The antibiotic efflux mechanism was observed in 0.9% of isolates. The biofilm formation was found in 3.7 and 96.3% of samples, respectively, on Congo red agar and on the microplate adhesion test, while the icaD gene was present in 92.2% of Staphylococcus spp. The high frequency of blaZ gene observed in this study was associated with the resistance of most Staphylococcus spp. to one or more of the beta-lactams tested, which are routinely used in Brazilian herds for mastitis treatment. The biofilm formation was also detected in the isolates analyzed being an important characteristic for pathogenicity and antimicrobial resistance of bacteria.

Resistance and virulence in Staphylococcus aureus by whole-genome sequencing: a comparative approach in blaZ-positive isolates.

Mastitis caused by Staphylococcus aureus is a worldwide problem in dairy farms, in part because of the pathogenicity of the bacteria, biofilm formation, and mechanisms of antimicrobial resistance that make the disease difficult to diagnose and treat, which is typically done with the use of beta-lactam antibiotics. The aim of the present study was to determine the virulence and resistance factors of S. aureus isolates from subclinical mastitis, blaZ + /mecA - /mecC - , resistant and sensitive to oxacillin. All isolates were classified as CC97 by MLST analysis, a clonal complex well adapted to the mammary gland and although STAU23 and STAU73 were resistant to oxacillin while STAU32 and STAU78 were sensitive, the genomic analysis identified only the blaZ operon corresponding to resistance to beta-lactams. However, the presence of the sdrC gene was revealed exclusively in resistant isolates, an important adhesin in the colonization process that potentiates pathogenicity in S. aureus. In addition, resistance islands (REIs) were identified in these isolates, suggesting more conserved REIs. In the analysis of SNPs throughout the genome, mutations were found in the trmB and smpB genes of the resistant isolates and in the murD and rimM genes of the sensitive isolates. This study highlights the potential benefit of genome-wide characterization tools to identify molecular mechanisms of S. aureus in bovine mastitis.

Distribution of PLD and FagA, B, C and D genes in Corynebacterium pseudotuberculosis isolates from sheep and goats with caseus lymphadenitis.

Caseous lymphadenits (CL) is a chronic and subclinical disease that affects goats and sheep and, consequently, causes economic losses, especially to small producers. The purpose of this study, through use of Polymerase Chain Reaction (PCR), was to verify the presence of virulence genes of phospholipase D (PLD), integral membrane protein (FagA), iron enterobactin transporter (FagB), ATP binding cytoplasmic membrane protein (FagC) and iron siderophore binding protein (FagD) in 168 isolates of C. pseudotuberculosis obtained from cases of caseous lymphadenitis in goats and sheep. FagA, FagB and PLD genes were detected in all 145 strains isolated from abscesses in superficial lymph nodes and in 23 strains isolated from viscera. The FagC gene was positive in 167 (99.40%) isolates. The FagD gene was detected in 160 (95.23%) isolates. All virulence factors analyzed were found more frequently among isolates collected in the viscera of animals with CL, indicating a multifactorial nature, as well as variations, in the invasive potential of C. pseudotuberculosis strains.

Action of crude ethanol extract of Hymenaea martiana leaf, gallic acid, and polypyrrole (PPy) against Aeromonas hydrophila.

Pisciculture represents one of the industries with the fastest growth rates worldwide. However, it presents obstacles to its development, such as bacteriosis, which is conventionally treated with antibiotics. The indiscriminate and inappropriate use of antibiotics can lead to bacterial resistance, thus alternatives to the use of antibiotics have been researched. The study aimed to analyze the potential of crude ethanol extract (CEE) from Hymenaea martiana leaf, gallic acid (GA), and polypyrrole (PPy) against Aeromonas hydrophila. Tests were performed to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the compounds individually and in synergy (checkerboard) against A. hydrophila and in silico tests between the compounds evaluated. The CEE of H. martiana leaf and PPy were effective against A. hydrophila with MBC results of 3125 µg/mL for the CEE of H. martiana and 125 µg/mL for PPy. Evaluating the GA, a MIC and MBC of 125 µg/mL was obtained. In the interaction tests (checkerboard, using PPy/CEE and PPy/GA), there was a significant reduction in individual introductions. Thus, for the PPy/CEE tests, we had a reduction of MIC/MBC to 1.95 and 781.25 µg/mL, and for the synergy tests between PPy/GA to 7.8125 and 31.125 µg/mL, respectively. The synergy tests are encouraging, and it is possible to verify a decrease of up to 98% in the introduction of PPy, 75% in CEE for H. martiana and 75.1% for GA, when compared to their individual tests. The tests with GA are encouraging due to GA's effectiveness as an antimicrobial agent and high synergy with polypyrrole, both in vitro results and molecular docking experiments showed the actions at the same activation site in A. hydrophila. In vivo tests evaluating isolated components of CEE from H. martiana in synergy with PPy should be performed, to verify the quality of the interactions and the improvement of the immune responses of the animals. It was evidenced that gallic acid, a substance isolated from the extract, tends to have more promising results. This is relevant since the industry has been developing these compounds for different uses, thus providing easier access to the product. Thus, the present study indicates an efficient alternative in the use of bioactive compounds as substitutes for conventional antimicrobials.

Blood parasite load by qPCR as therapeutic monitoring in visceral leishmaniasis patients in Brazil: a case series study.

BACKGROUND: This study aimed to describe the kinetics of Leishmania parasite load determined using kinetoplast DNA (kDNA)-based quantitative polymerase chain reaction (qPCR) in visceral leishmaniasis (VL) patients. METHODS: Parasite load in blood was assessed by qPCR at five time points, up to 12 months post-diagnosis. Sixteen patients were followed up. RESULTS: A significant reduction in the parasite load was observed after treatment (P < 0.0001). One patient had an increased parasite load 3 months post-treatment and relapsed clinically at month six. CONCLUSIONS: We have described the use of kDNA-based qPCR in the post-treatment follow-up of VL cases.

Genomic Characterization of Aeromonas veronii Provides Insights into Taxonomic Assignment and Reveals Widespread Virulence and Resistance Genes throughout the World.

Aeromonas veronii is a Gram-negative bacterial species that causes disease in fish and is nowadays increasingly recurrent in enteric infections of humans. This study was performed to characterize newly sequenced isolates by comparing them with complete genomes deposited at the NCBI (National Center for Biotechnology Information). Nine isolates from fish, environments, and humans from the São Francisco Valley (Petrolina, Pernambuco, Brazil) were sequenced and compared with complete genomes available in public databases to gain insight into taxonomic assignment and to better understand virulence and resistance profiles of this species within the One Health context. One local genome and four NCBI genomes were misidentified as A. veronii. A total of 239 virulence genes were identified in the local genomes, with most encoding adhesion, motility, and secretion systems. In total, 60 genes involved with resistance to 22 classes of antibiotics were identified in the genomes, including mcr-7 and cphA. The results suggest that the use of methods such as ANI is essential to avoid misclassification of the genomes. The virulence content of A. veronii from local isolates is similar to those complete genomes deposited at the NCBI. Genes encoding colistin resistance are widespread in the species, requiring greater attention for surveillance systems.

Pain paths among post-COVID-19 condition subjects: A prospective cross-sectional study with in-person evaluation.

BACKGROUND: New-onset chronic pain has been acknowledged as part of the post-COVID-19 condition. However, available fine-grained data about its clinical phenotype, trajectories and main associated characteristics remain scarce. We described the distinct temporal evolutions of post-COVID-19 pain and their epidemiological and phenotypical features. METHODS: A prospective cross-sectional study enrolled post-COVID-19 condition patients (i.e. who had persisting COVID-19-related symptoms over 30 days since their first positive laboratory test), whose COVID-19 diagnosis had been supported by RT-PCR of oral/nasopharyngeal swab or serology. They underwent in-person evaluations with a structured interview, pain and quality-of-life-related questionnaires and thorough physical examination. Chronic pain (CP) and probable neuropathic pain (NP) were defined according to IASP criteria. RESULTS: The present study included 226 individuals, 177 (78.3%) of whom presented over 3 months since their first COVID-19 symptom. New-onset pain occurred in 170 (75.2%) participants and was chronic in 116 (68.2%). A chronic course was associated with COVID-19-related hospitalization, new-onset fatigue, lower cognitive performance, motor and thermal sensory deficits, mood and sleep impairments and overall lower quality-of-life levels. Probable NP occurred in only 7.6% new-onset pain patients, and was associated with pain chronification, new-onset fatigue, motor and thermal sensory deficits, mechanical allodynia and lower rates of SARS-CoV-2 vaccination. Previous CP was reported by 86 (38.1%) individuals and had aggravated after the infection in 66 (76.7%) of them, which was associated with orthostatic hypotension. CONCLUSIONS: Post-COVID pain phenomena follow different paths, which are associated with specific clinical and epidemiological features, and possibly distinct underlying mechanisms, prognostic and therapeutic implications. SIGNIFICANCE: COVID-19-related pain usually follows a chronic course and is non-neuropathic. Its possible courses and phenotypes are associated with distinct clinical and epidemiological features. This suggests differing underlying mechanisms, which may have significant prognostic and therapeutic implications.

Gestational age acceleration is associated with epigenetic biomarkers of prenatal physiologic stress exposure.

BACKGROUND: Physiological maternal stress response, such as imbalance in the glucocorticoid pathway and immune system seems to be mediated by DNA methylation (DNAm) and might translate intrauterine stress exposures into phenotypic changes in a sex-specific manner. DNAm in specific sites can also predict newborn gestational age and gestational age acceleration (GAA). GAA occurs when the predicted biological age is higher than the chronological age. In adults, poor health outcomes related to this deviance are well documented and raise questions for the interpretation and prediction in early stages of life. Boys seem to be more vulnerable to intrauterine stress exposure than girls; however, the mechanisms of adaptive sex-specific responses are still unclear. We hypothesize that intrauterine stress exposure is associated with GAA and could be different in boys and girls if inflammatory or glucocorticoid pathways exposure is considered. RESULTS: Using the Western Region Birth Cohort (ROC-São Paulo, Brazil) (n = 83), we calculated DNAm age and GAA from cord blood samples. Two epigenetic risk scores were calculated as an indirect proxy for low-grade inflammation (i-ePGS) and for glucocorticoid exposure (GES). Multivariate linear regression models were applied to investigate associations of GAA with prenatal exposures. The i-ePGS and GES were included in different models with the same co-variates considering sex interactions. The first multivariate model investigating inflammatory exposure (adj. R2 = 0.31, p = < 0.001) showed that GAA was positively associated with i-ePGS (CI, 0.26-113.87, p = 0.049) and negative pregnancy-related feelings (CI, 0.04-0.48 p = 0.019). No sex interaction was observed. The second model investigating glucocorticoid exposure (adj. R2 = 0.32, p = < 0.001) showed that the higher was the GAA was associated with a lower the lower was the GES in girls (CI, 0.04-2.55, p = 0.044). In both models, maternal self-reported mental disorder was negatively associated with GAA. CONCLUSION: Prenatal epigenetic score of exposure to low-grade inflammatory was a predictor of GAA for both sexes. Glucocorticoid epigenetic score seems to be more important to GAA in girls. This study supports the evidence of sex-specificity in stress response, suggesting the glucocorticoid as a possible pathway adopted by girls to accelerate the maturation in an adverse condition.

DNA methylation mediates a randomized controlled trial home-visiting intervention during pregnancy and the Bayley infant's cognitive scores at 12 months of age.

The crosstalk between maternal stress exposure and fetal development may be mediated by epigenetic mechanisms, including DNA methylation (DNAm). To address this matter, we collect 32 cord blood samples from low-income Brazilian pregnant adolescents participants of a pilot randomized clinical intervention study (ClinicalTrials.gov, Identifier: NCT02807818). We hypothesized that the association between the intervention and infant neurodevelopmental outcomes at 12 months of age would be mediated by DNAm. First, we searched genome methylation differences between cases and controls using different approaches, as well as differences in age acceleration (AA), represented by the difference of methylation age and birth age. According to an adjusted p-value ≤ 0.05 we identified 3090 differentially methylated positions- CpG sites (DMPs), 21 differentially methylated regions (DMRs) and one comethylated module weakly preserved between groups. The intervention group presented a smaller AA compared to the control group (p = 0.025). A logistic regression controlled by sex and with gestational age indicated a coefficient of -0.35 towards intervention group (p = 0.016) considering AA. A higher cognitive domain score from Bayley III scale was observed in the intervention group at 12 months of age. Then, we performed a potential causal mediation analysis selecting only DMPs highly associated with the cognitive domain (adj. R2 > 0.4), DMRs and CpGs of hub genes from the weakly preserved comethylated module and epigenetic clock as raw values. DMPs in STXBP6, and PF4 DMR, mediated the association between the maternal intervention and the cognitive domain at 12 months of age. In conclusion, DNAm in different sites and regions mediated the association between intervention and cognitive outcome.

Tannin as a modulator of rumen microbial profile, apparent digestibility and ingestive behavior of lactating goats: A preliminary metagenomic view of goats adaptability to tannin.

The aim with this study was to use interdisciplinary techniques and visions in order to identify the modulating effect of tannins on adaptive factors in lactating goats with different genetic patterns, through the assessment of digestibility and nutrient intake, ingestive behavior and rumen metagenome. We used in this study 8 of the Repartida ecotype and 6 of the Canindé breed goats, on average five years old, distributed in a completely randomized design, in a 2 × 2 factorial design, with two genetic groups and two diets. The applied diets were: basal diet and inclusion of 5% commercial tannin extract in basal diet. Intake and apparent digestibility of nutrients were evaluated, based on the quantification of the offered feed and refusals, and bromatological evaluation of samples of the offered feed, refusals and feces. Behavioral data were collected in 24-h continuous visual observations. The ruminal fluid was collected and DNA extraction, sequencing, and evaluation of relative abundance of the rumen microbiome were performed. The data obtained were analyzed statistically, through analysis of variance with 5% significance and, when necessary, a comparison of means test was applied. In this preliminary findings was observed that the genetic group factor caused changes in the number of chews and the relative abundance of microorganisms (P = 0.0290 and P = 0.0051). The diet factor influenced digestibility, which better values were observed for the tannin diet (P = 0.0049), in addition, it promoted changes in the rumen microbiota, with a beneficial modulatory characteristic. The inclusion of 5% tannin extract from Acacia mearnsii modulates the rumen microbiome, improving the apparent digestibility of nutrients without affecting the feed intake of goats from the Repartida and Canindé genetic groups.

Molecular screening for the mutation associated with canine degenerative myelopathy (SOD1:c.118G > A) in German Shepherd dogs in Brazil.

Canine Degenerative Myelopathy is a late onset recessive autosomal disease characterized by a progressive ascending degeneration of the spinal cord. Two causal mutations are associated with this disease: a transition (c.118G>A) in exon 2 of the SOD1 that was described in several breeds and a transversion (c.52A>T) in exon 1 of the same gene described in Bernese Mountain dogs. The aim of this study was to understand the impact of the SOD1:c.118G > A mutation by genotyping a population of German Shepherd dogs in Brazil. A PCR-RFLP approach was used to genotype 97 healthy individuals belonging from the Northeast (Bahia and Pernambuco states) and South (Santa Catarina state) regions of Brazil. A total of 95 individuals were successfully genotyped resulting in an observed genotype frequency (with 95% confidence interval) of: 0.758 (0.672-0.844), 0.242 (0.156-0.328) and 0.000 (0.000-0.000) for "GG", "AG" and "AA" genotypes, respectively. To our knowledge, this is the first attempt to describe the presence of the "A" allele associated with CDM (SOD1:c.118G > A) in German Shepherd dogs in Brazil and, as such, these results contribute toward important epidemiological data in this country and to the knowledge of the distribution of the aforementioned mutation worldwide.

Genome-wide DNA methylation profile in the peripheral blood of cocaine and crack dependents.

OBJECTIVE: Cocaine use disorders (CUDs) represent a major public health problem in many countries. To better understand the interaction between the environmental modulations and phenotype, the aim of the present study was to investigate the DNA methylation pattern of CUD patients, who had concomitant cocaine and crack dependence, and healthy controls. METHODS: We studied DNA methylation profiles in the peripheral blood of 23 CUD patients and 24 healthy control subjects using the Illumina Infinium HumanMethylation450 BeadChip arrays. RESULTS: Comparison between CUD patients and controls revealed 186 differentially methylated positions (DMPs; adjusted p-value [adjP] < 10-5) related to 152 genes, with a subset of CpGs confirmed by pyrosequencing. DNA methylation patterns discriminated CUD patients and control groups. A gene network approach showed that the EHMT1, EHMT2, MAPK1, MAPK3, MAP2K1, and HDAC5 genes, which are involved in transcription and chromatin regulation cellular signaling pathways, were also associated with cocaine dependence. CONCLUSION: The investigation of DNA methylation patterns may contribute to a better understanding of the biological mechanisms involved in CUD.

Molecular characterization of viruses associated with encephalitis in São Paulo, Brazil.

The objective of this study was to characterize the prevalence of viral encephalitis due to arbovirus infection of the Togaviridae and Flaviviridae families in São Paulo, Brazil. A total of 500 cerebrospinal fluid (CSF) samples collected between August 2012 and January 2013, from patients with symptoms of acute encephalitis were analyzed. Findings suggestive of viral encephalitis-elevations in cell concentration, glucose and total protein-were observed in 234 (46.8%) samples, designated as Group 1. The remaining 266 samples comprised Group 2. All samples were tested for Flaviviruses (dengue virus 1, 2, 3 and 4, yellow fever virus and West Nile virus), Alphavirus (NS5 region) and enterovirus by RT- PCR and for herpesviruses and enteroviruses using CLART-Entherpex. A presumptive viral etiological agent was detected in 26 samples (5.2%), 18 (8.0%) in Group 1 and 8 (3.0%) in Group 2. In Group 1 human herpesviruses were detected in 9 cases, enteroviruses in 7 cases, dengue viruses (DENV) in 2 CSFs and St. Louis encephalitis virus (SLEV) in one case. In Group 2 there were 3 CSFs positive for human herpesviruses, 2 for enteroviruses, 2 for DENV and 1 for SLEV. Detection of arboviruses, even though present in a minority of infected patients, identifies these viruses as a probable etiological agent of encephalitis. This is of special concern in regions where this class of viruses is endemic and has been linked to other recent epidemics.

Initial findings of striatum tripartite model in OCD brain samples based on transcriptome analysis.

Obsessive-compulsive disorder (OCD) is a psychiatric disorder characterized by obsessions and/or compulsions. Different striatal subregions belonging to the cortico-striato-thalamic circuitry (CSTC) play an important role in the pathophysiology of OCD. The transcriptomes of 3 separate striatal areas (putamen (PT), caudate nucleus (CN) and accumbens nucleus (NAC)) from postmortem brain tissue were compared between 6 OCD and 8 control cases. In addition to network connectivity deregulation, different biological processes are specific to each striatum region according to the tripartite model of the striatum and contribute in various ways to OCD pathophysiology. Specifically, regulation of neurotransmitter levels and presynaptic processes involved in chemical synaptic transmission were shared between NAC and PT. The Gene Ontology terms cellular response to chemical stimulus, response to external stimulus, response to organic substance, regulation of synaptic plasticity, and modulation of synaptic transmission were shared between CN and PT. Most genes harboring common and/or rare variants previously associated with OCD that were differentially expressed or part of a least preserved coexpression module in our study also suggest striatum subregion specificity. At the transcriptional level, our study supports differences in the 3 circuit CSTC model associated with OCD.

Blood parasite load by qPCR as therapeutic monitoring in visceral leishmaniasis patients in Brazil: a case series study

ABSTRACT Background: This study aimed to describe the kinetics of Leishmania parasite load determined using kinetoplast DNA (kDNA)-based quantitative polymerase chain reaction (qPCR) in visceral leishmaniasis (VL) patients. Methods: Parasite load in blood was assessed by qPCR at five time points, up to 12 months post-diagnosis. Sixteen patients were followed up. Results: A significant reduction in the parasite load was observed after treatment (P < 0.0001). One patient had an increased parasite load 3 months post-treatment and relapsed clinically at month six. Conclusions: We have described the use of kDNA-based qPCR in the post-treatment follow-up of VL cases.