Host cell proteins modulated upon Toxoplasma infection identified using proteomic approaches: a molecular rationale.

Toxoplasma gondii is a pathogenic protozoan parasite belonging to the apicomplexan phylum that infects the nucleated cells of warm-blooded hosts leading to an infectious disease known as toxoplasmosis. Apicomplexan parasites such as T. gondii can display different mechanisms to control or manipulate host cells signaling at different levels altering the host subcellular genome and proteome. Indeed, Toxoplasma is able to modulate host cell responses (especially immune responses) during infection to its advantage through both structural and functional changes in the proteome of different infected cells. Consequently, parasites can transform the invaded cells into a suitable environment for its own replication and the induction of infection. Proteomics as an applicable tool can identify such critical proteins involved in pathogen (Toxoplasma)-host cell interactions and consequently clarify the cellular mechanisms that facilitate the entry of pathogens into host cells, and their replication and transmission, as well as the central mechanisms of host defense against pathogens. Accordingly, the current paper reviews several proteins (identified using proteomic approaches) differentially expressed in the proteome of Toxoplasma-infected host cells (macrophages and human foreskin fibroblasts) and tissues (brain and liver) and highlights their plausible functions in the cellular biology of the infected cells. The identification of such modulated proteins and their related cell impact (cell responses/signaling) can provide further information regarding parasite pathogenesis and biology that might lead to a better understanding of therapeutic strategies and novel drug targets.

Assessment of burden of virus agents in an urban sewage treatment plant in Rio de Janeiro, Brazil.

Sewage discharge is considered to be the main source of virus contamination in aquatic environments. There is no correlation between the presence of viruses and the presence of fecal coliforms in water; therefore virological markers are needed when monitoring contamination. This study investigates DNA and RNA virus concentrations in wastewater and evaluates a potential virus marker of human contamination. Influent and effluent samples were collected twice a month throughout a 1-year period. Viruses were detected using quantitative polymerase chain reaction protocols; nucleotide sequencing was carried out for virus genotyping. Human adenovirus (HAdV) and polyomavirus JC (JCPyV) were the most prevalent viruses found in influent samples (100%) with a virus load that ranged from 10(6) to 10(5) genome copies per liter (gc l(-1)). Norovirus genogroup II (NoV GII) and human astrovirus (HAstV) were less prevalent, and ranged from 10(4) to 10(3)gc l(-1). Quantitative data on virus profiles in wastewaters stress the high level of rotavirus species A environmental dissemination and address the potential of HAdV as a useful virological marker of virus contamination in aquatic environments. This study corroborates other studies performed in developed countries on DNA viruses as good markers of human fecal contamination.

Learning the Biochemical Basis of Axonal Guidance: Using Caenorhabditis elegans as a Model.

AIM: Experimental models are a powerful aid in visualizing molecular phenomena. This work reports how the worm Caenorhabditis elegans (C. elegans) can be effectively explored for students to learn how molecular cues dramatically condition axonal guidance and define nervous system structure and behavior at the organism level. Summary of work: A loosely oriented observational activity preceded detailed discussions on molecules implied in axonal migration. C. elegans mutants were used to introduce second-year medical students to the deleterious effects of gene malfunctioning in neuron response to extracellular biochemical cues and to establish links between molecular function, nervous system structure, and animal behavior. Students observed C. elegans cultures and associated animal behavior alterations with the lack of function of specific axon guidance molecules (the soluble cue netrin/UNC-6 or two receptors, DCC/UNC-40 and UNC-5H). Microscopical observations of these strains, in combination with pan-neuronal GFP expression, allowed optimal visualization of severely affected neurons. Once the list of mutated genes in each strain was displayed, students could also relate abnormal patterns in axon migration/ventral and dorsal nerve cord neuron formation in C. elegans with mutated molecular components homologous to those in humans. SUMMARY OF RESULTS: Students rated the importance and effectiveness of the activity very highly. Ninety-three percent found it helpful to grasp human axonal migration, and all students were surprised with the power of the model in helping to visualize the phenomenon.

Detection of enteric viruses in recreational waters of an urban lagoon in the city of Rio de Janeiro, Brazil.

The aim of this study was to determine the occurrences of the group A rotavirus (RVA), norovirus (NoV) and human adenovirus (HAdV) in the surface waters of an urban lagoon (Rodrigo de Freitas Lagoon) in the city of Rio de Janeiro, Brazil. During one year of surveillance, water samples were obtained from the lagoon and other interconnected ecosystems (river and beach). The samples were concentrated using an adsorption-elution method with a negatively charged membrane and tested by qualitative and quantitative polymerase chain reaction assays. RVA was the most prevalent virus detected (24.3%) with a viral load ranging from 3.0 x 10¹-5.6 x 10(4) genome copies/L, followed by NoV (18.8%) and HAdV (16.7%). Considering water samples suitable for bathing, according to Escherichia coli criterion (< 2,000 most probable number/100 mL), viruses were detected in 50% (57/114) of them. Physicochemical parameters were also measured and showed possible correlations between turbidity and RVA presence and between pH and NoV presence. These data demonstrate the importance of considering viral parameters to ensure water quality and the utilisation of these parameters as additional tools for the characterisation of environmental contamination.

Fast screening of enteropathogens in marine water samples.

This study aimed to fast screen the microbiological contamination of recreational waters using a TaqMan Array Card (TAC), a multiplexed platform designed for the simultaneous detection of 35 enteropathogens. Surface and deep marine water samples were concentrated by skimmed milk flocculation and processed for nucleic acid extraction protocol using QIAamp Fast DNA Stool Mini Kit. Twelve microorganisms and parasites, including bacteria (n = 6), protozoa (4), and viruses (2), were detected in 85.7% (24/28) of samples. Campylobacter (82.1%), Cryptosporidium (39.3%), and adenovirus (14.3%) were the most detected pathogens. Neither fungi nor helminths were detected. A spatial pollution profile of microbiological contamination was observed in the area. Methodologies for simultaneous detection of multiple pathogens, such as TAC, can assist decision-makers by providing a quick assessment of the microbiological water quality in areas used for recreational purposes, which in many cases are in accordance with the bacteriological indicators.

Immunomodulatory Potential of Non-Classical HLA-G in Infections including COVID-19 and Parasitic Diseases.

Human Leukocyte Antigen-G (HLA-G), a polymorphic non-classical HLA (HLA-Ib) with immune-regulatory properties in cancers and infectious diseases, presents both membrane-bound and soluble (sHLA-G) isoforms. Polymorphism has implications in host responses to pathogen infections and in pathogenesis. Differential expression patterns of HLA-G/sHLA-G or its polymorphism seem to be related to different pathological conditions, potentially acting as a disease progression biomarker. Pathogen antigens might be involved in the regulation of both membrane-bound and sHLA-G levels and impact immune responses during co-infections. The upregulation of HLA-G in viral and bacterial infections induce tolerance to infection. Recently, sHLA-G was found useful to identify the prognosis of Coronavirus disease 2019 (COVID-19) among patients and it was observed that the high levels of sHLA-G are associated with worse prognosis. The use of pathogens, such as Plasmodium falciparum, as immune modulators for other infections could be extended for the modulation of membrane-bound HLA-G in COVID-19-infected tissues. Overall, such information might open new avenues concerning the effect of some pathogens such as parasites in decreasing the expression level of HLA-G to restrict pathogenesis in some infections or to influence the immune responses after vaccination among others.

Aripiprazole Offsets Mutant ATXN3-Induced Motor Dysfunction by Targeting Dopamine D2 and Serotonin 1A and 2A Receptors in C. elegans.

The atypical antipsychotic aripiprazole is a Food and Drug Administration-approved drug for the treatment of psychotic, mood, and other psychiatric disorders. Previous drug discovery efforts pinpointed aripiprazole as an effective suppressor of Machado-Joseph disease (MJD) pathogenesis, as its administration resulted in a reduced abundance and aggregation of mutant Ataxin-3 (ATXN3) proteins. Dopamine partial agonism and functional selectivity have been proposed as the main pharmacological mechanism of action of aripiprazole in the treatment of psychosis; however, this mechanism remains to be determined in the context of MJD. Here, we focus on confirming the efficacy of aripiprazole to reduce motor dysfunction in vivo, using a Caenorhabditis elegans (C. elegans) model of MJD, and on unveiling the drug targets required for its positive action against mutant ATXN3 pathogenesis. We employed pharmacogenetics and pharmacological approaches to identify which dopamine and serotonin receptors are critical for aripiprazole-mediated improvements in motor function. We demonstrated that dopamine D2-like and serotonin 5-HT1A and 5-HT2A receptors play important roles in this process. Our findings strengthen the relevance of dopaminergic and serotoninergic signaling modulation against mutant ATXN3-mediated pathogenesis. The identification of aripiprazole's cellular targets, relevant for MJD and perhaps other neurodegenerative diseases, may pave the way for prospective drug discovery and development campaigns aiming to improve the features of this prototypical compound and reduce side effects not negligible in the case of aripiprazole.

WDR60-mediated dynein-2 loading into cilia powers retrograde IFT and transition zone crossing.

The dynein-2 motor complex drives retrograde intraflagellar transport (IFT), playing a pivotal role in the assembly and functions of cilia. However, the mechanisms that regulate dynein-2 motility remain poorly understood. Here, we identify the Caenorhabditis elegans WDR60 homologue, WDR-60, and dissect the roles of this intermediate chain using genome editing and live imaging of endogenous dynein-2/IFT components. We find that loss of WDR-60 impairs dynein-2 recruitment to cilia and its incorporation onto anterograde IFT trains, reducing retrograde motor availability at the ciliary tip. Consistent with this, we show that fewer dynein-2 motors power WDR-60-deficient retrograde IFT trains, which move at reduced velocities and fail to exit cilia, accumulating on the distal side of the transition zone. Remarkably, disrupting the transition zone's NPHP module almost fully restores ciliary exit of underpowered retrograde trains in wdr-60 mutants. This work establishes WDR-60 as a major contributor to IFT, and the NPHP module as a roadblock to dynein-2 passage through the transition zone.

Genetic diversity of noroviruses in Brazil.

Norovirus (NoV) infections are a major cause of acute gastroenteritis outbreaks around the world. In Brazil, the surveillance system for acute diarrhoea does not include the diagnosis of NoV, precluding the ability to assess its impact on public health. The present study assessed the circulation of NoV genotypes in different Brazilian states by partial nucleotide sequencing analysis of the genomic region coding for the major capsid viral protein. NoV genogroup II genotype 4 (GII.4) was the prevalent (78%) followed by GII.6, GII.7, GII.12, GII.16 and GII.17, demonstrating the great diversity of NoV genotypes circulating in Brazil. Thus, this paper highlights the importance of a virological surveillance system to detect and characterize emerging strains of NoV and their spreading potential.

Microbiological assessment of an urban lagoon system in the coastal zone of Rio de Janeiro, Brazil.

This study aims to assess microbiological contamination using a molecular tool for detection of multiple enteropathogens in a coastal ecosystem area in Rio de Janeiro, Brazil. Ten litres of superficial water samples were obtained during the spring ebb tide from sampling sites along the Jacarepaguá watershed. Samples were concentrated using skimmed milk flocculation method for TaqMan array card (TAC), designed to identify 35 enteric pathogens simultaneously, and single TaqMan qPCR analysis for detecting human adenovirus (HAdV) and JC human polyomavirus (JCPyV) as faecal indicator viruses (FIV). TAC results identified 17 enteric pathogens including 4/5 viral species investigated, 8/15 bacteria, 4/6 protozoa and 1/7 helminths. Escherichia coli concentration was also measured as faecal indicator bacteria (FIB) using Colilert Quanti-Tray System with positivity in all samples studied. HAdV and JCPyV qPCR were detected in 8 and 4 samples, respectively, with concentration ranging from 8 × 102 to 2 × 106 genome copies/L. Partial nucleotide sequencing demonstrated the occurrence of species HAdV A, C, D, and F, present in faeces of individuals with enteric and non-enteric infections, and JCPyV type 3 (Af2), prevalent in a high genetically mixed population like the Brazilian. The diversity of enteropathogens detected by TAC emphasizes the utility of this methodology for quick assessment of microbiological contamination of the aquatic ecosystems, speeding up mitigation actions where the risk of the exposed population is detected, as well as pointing out the infrastructure gaps in areas where accelerated urban growth is observed.

Expression of an alternatively spliced variant of SORL1 in neuronal dendrites is decreased in patients with Alzheimer's disease.

SORL1 is strongly associated with both sporadic and familial forms of Alzheimer's disease (AD), but a lack of information about alternatively spliced transcripts currently limits our understanding of the role of SORL1 in AD. Here, we describe a SORL1 transcript (SORL1-38b) characterized by inclusion of a novel exon (E38b) that encodes a truncated protein. We identified E38b-containing transcripts in several brain regions, with the highest expression in the cerebellum and showed that SORL1-38b is largely located in neuronal dendrites, which is in contrast to the somatic distribution of transcripts encoding the full-length SORLA protein (SORL1-fl). SORL1-38b transcript levels were significantly reduced in AD cerebellum in three independent cohorts of postmortem brains, whereas no changes were observed for SORL1-fl. A trend of lower 38b transcript level in cerebellum was found for individuals carrying the risk variant at rs2282649 (known as SNP24), although not reaching statistical significance. These findings suggest synaptic functions for SORL1-38b in the brain, uncovering novel aspects of SORL1 that can be further explored in AD research.

One year monitoring of norovirus in a sewage treatment plant in Rio de Janeiro, Brazil.

Norovirus (NoV) is one of the most important aetiological agents of acute gastroenteritis both in developed and developing countries. NoV is shed in high concentrations by infected persons and contaminates recreational and drinking water through sewage discharge into the environment. The aim of this study was to determine the prevalence, genotypes and removal ratio of NoV by PCR, seminested-PCR and quantitative PCR (qPCR) assays in a sewage treatment plant in Rio de Janeiro city, Brazil, during one year of surveillance. NoV was detected in 7 (15%), 14 (29%) and 28 (58%) samples using PCR, seminested-PCR and qPCR, respectively. The mean removal ratio for the activated sludge process was 0.6 log10 for NoV genogroup I (GI) and 0.32 log10 for NoV genogroup II (GII). The peak NoV concentration was detected in the coldest months, with 53,300 genomic copies/litre. Nucleotide sequencing and phylogenetic analysis revealed that five strains clustered with GI strains and six with GII strains. This study demonstrates that NoV spreads into the environment despite the sewage treatment process and remains a source of waterborne outbreaks of acute gastroenteritis.

Evaluation of an adsorption-elution method for detection of astrovirus and norovirus in environmental waters.

Human astroviruses (HAstV) and noroviruses (NoV) are shed frequently at high concentrations and persist for long periods in the environment, raising a significant health risk of water-related gastroenteritis. The aim of this study was to evaluate an adsorption-elution method with an HA (mixed cellulose esters) negatively charged membrane to determine the best recovery of HAstV and NoV from different environmental waters. As the presence of MgCl(2) affects viral adsorption onto the membrane, three different MgCl(2) concentrations were evaluated. The best recovery of both NoV and HAstV from mineral and river water samples was between 18% and 64%, while recovery from tap water and sea water samples was between 3% and 14%. These results suggest that detection and recovery of each enteric virus with this adsorption-elution method requires a specific MgCl(2) concentration and depends on the source of environmental water tested. The combination of the HA negatively charged membrane to concentrate viruses with quantitative PCR detection allows for the identification of gastroenteritis viruses implicated in acute outbreaks of gastroenteritis.

Major Histocompatibility Complex Class II (DRB3) Genetic Diversity in Spanish Morucha and Colombian Normande Cattle Compared to Taurine and Zebu Populations.

Bovine leukocyte antigens (BoLA) have been used as disease markers and immunological traits in cattle due to their primary role in pathogen recognition by the immune system. A higher MHC allele diversity in a population will allow presenting a broader peptide repertoire. However, loss of overall diversity due to domestication process can decrease a population's peptide repertoire. Within the context of zebu and taurine cattle populations, BoLA-DRB3 genetic diversity in Spanish Morucha and Colombian Normande cattle was analyzed and an approach to estimate functional diversity was performed. Sequence-based typing was used for identifying 29, 23, 27, and 28 alleles in Spanish Morucha, Nariño-, Boyacá-, and Cundinamarca-Normande cattle, respectively. These breeds had remarkably low heterozygosity levels and the Hardy-Weinberg principle revealed significant heterozygote deficiency. FST and DA genetic distance showed that Colombian Normande populations had greater variability than other phenotypically homogeneous breeds, such as Holstein. It was also found that Spanish Morucha cattle were strongly differentiated from other cattle breeds. Spanish Morucha had greater divergence in the peptide-binding region regarding other cattle breeds. However, peptide-binding region covariation indicated that the potential peptide repertoire seemed equivalent among cattle breeds. Despite the genetic divergence observed, the extent of the potential peptide repertoire in the cattle populations studied appears to be similar and thus their pathogen recognition potential should be equivalent, suggesting that functional diversity might persist in the face of bottlenecks imposed by domestication and breeding.

Molecular detection of human astrovirus in an urban sewage treatment plant in Rio de Janeiro, Brazil.

The objective of this study was to evaluate the prevalence and dissemination of human astroviruses (HAstV) in the environment by analyzing urban sewage samples from a wastewater treatment plant in the city of Rio de Janeiro, Brazil. A one-year study was performed with a total of 48 raw and treated sewage composite samples, which were collected biweekly from an activated sludge plant. Virus particles were concentrated by the adsorption-elution method using negatively charged membranes associated to a Centriprep Concentrator 50 (Nihon Millipore). HAstV were detected in 16.7% of the samples in raw and treated sewage by using both qualitative and quantitative reverse transcriptase-polymerase chain reactions (RT-PCR and qPCR, respectively). Positive untreated sewage sample exhibited mean values of 1.1 x 10(4) gEq/mL. The qPCR sensitivity was 18 gEq/reaction. Through utilization of qPCR, a HAstV recovery efficiency of 4.2% and 4.3% was demonstrated for raw and treated sewage samples, respectively. The presence of HAstV in both the raw and treated sewage samples demonstrated the dissemination of these viruses in the environment as well as viral permanence after sewage treatment. There was a reduction in the total and faecal coliform levels, indicating efficiency of the wastewater treatment plant.

[Evaluation and control of missing contacts of leprosy patients]. / Avaliação e controle de contatos faltosos de doentes com hanseníase.

Descriptive study, with qualitative variables, carried on in the Ambulatory of Specialties Region of Taubaté, in order to evaluate the missing contacts of leprosy patients. Thirty-six records of new cases were analyzed, from January 2003 to July 2004. Ninety-two contacts were identified, being 64.1% missing ones, having 25% from 20 to 29 years and 58.5% female. Of these, 61% were submitted to consultation of home nursing. Reported non-membership to the control for forgetfulness (66.6%), and lack of time (11.1%), 4 contacts had symptoms of leprosy, and 1 was confirmed (transmissible form - Dimorfa). Without achieving 100% coverage active search, we suggest, to ensure the goal of eliminating of the disease, partnerships and decentralization of the control activities.

[The basic health and assistance to Hansen's Disease in health care services of a municipality of São Paulo State]. / Atenção básica de saúde e a assistência em Hanseníase em serviços de saúde de um município do Estado de São Paulo.

UNLABELLED: This descriptive study was carried out in a municipality of Sao Paulo State. The objective was to identify and to characterize the Leprosy Control Program in primary care in the city of research. METHODOLOGY: the interviews were conducted with professionals responsable for care in Hansen's Disease and with manager of public health policies. RESULTS: the public health policies prioritize the development of primary care and emphasis on the public health. The control actions are held by trained and experient professionals. It emphasizes absence of active search for cases which undermines the real epidemiological situation, and the important health education to the reduction of stigma and bring the subject of adjustments necessary in new situation of life and overcome limitations.

[Health services organization on the elimination of Leprosy in municipalities of São Paulo State]. / Organização de serviços de saúde na eliminação da Hanseníase em municípios do Estado de São Paulo.

The present study aims to analyze actions to control Hansen's disease developed in nine municipalities of the State of São Paulo and one of the health authorities of the city of São Paulo, selected based on their population, geographic location and prevalence coefficients. The study was approved by the Ethics Committee n masculine 514/05 and instruments were administered by means of interviews to 59 professionals, 17 interlocutors and 9 managers. Data were analyzed using the program SPSS (Statistical Package for Social Scientists). It was verified that prevalence is on the decline and detection remains stable. The physical incapacity prevention actions, health education and active search have not been satisfactorily developed. The most qualified professionals are in the reference units and teamwork is fragile.

Detection and quantification of human adenovirus (HAdV), JC polyomavirus (JCPyV) and hepatitis A virus (HAV) in recreational waters of Niterói, Rio de Janeiro, Brazil.

This study evaluated the impact of sewage discharge in recreational coastal marine environments of Niteroi, Rio de Janeiro, Brazil, over a six-month period by the detection of waterborne enteric viruses. Ten-liter water samples were collected in four beaches from January to July 2017. Viruses were concentrated by an organic flocculation and human adenoviruses (HAdV), polyomavirus (JCPyV), and Hepatitis A virus (HAV) detected by qPCR. Forty-eight water samples were collected, being 43% positive for HAdV and 23% for JCPyV; only one sample was positive for HAV. Viruses were detected in all sampling sites, including in areas suitable for bathing according to the current bacterial standards. The results herein provide an overview of the viral contamination of beaches used for recreational purposes. The viral presence in the sampled areas indicates the need for more rigid effluent discharge controls in these areas, as sewage represents a possible transmission risk for waterborne viral diseases.

Mediterranean spotted fever in Spain, 1997-2014: Epidemiological situation based on hospitalization records.

INTRODUCTION: Mediterranean spotted fever (MSF) is a zoonotic disease caused by Rickettsia conorii. In Spain, deficiencies in the official reporting result in misreporting of this disease. This study aims to describe the clinical and temporal-spatial characteristics of MSF hospitalizations between 1997 and 2014. MATERIALS AND METHODS: We performed a retrospective descriptive study using the Hospitalization Minimum Data Set (CMBD). All CMBD's hospital discharges with ICD-9 CM code 082.1 were analyzed. Hospitalization rates were calculated and clinical characteristics were described. Spatial distribution of cases and their temporal behavior were also assessed. RESULTS: A total of 4,735 hospitalizations with MSF diagnosis were recorded during the study period, out of which 62.2% were male, mean age of 48. Diabetes mellitus, alcohol dependence syndrome, and chronic liver disease occurred in 10.8%, 2.4% and 2.8% hospitalizations, respectively. The median annual hospitalization rate showed a decreasing trend from a maximum of 12.9 in 1997 to a minimum rate of 3.1 in 2014. Most admissions occurred during the summer, showing a significant annual seasonal behavior. Important regional differences were found. DISCUSSION: Although MSF hospitalization rates have decreased considerably, it remains a public health problem due to its severity and economic impact. Therefore, it would be desirable to improve its oversight and surveillance.