Dating Skin Lesions of Forensic Interest by Immunohistochemistry and Immunofluorescence Techniques: A Scoping Literature Review.

Wound age estimation is a significant issue in forensic pathology. Although various methods have been evaluated, no gold standard system or model has been proposed, and accurate injury time estimation is still challenging. The distinction between vital skin wounds-i.e., ante-mortem lesions-and skin alterations that occur after death is a crucial goal in forensic pathology. Once the vitality of the wound has been confirmed, the assessment of the post-trauma interval (PTI) is also fundamental in establishing the causal relationship between the traumatic event and death. The most frequently used techniques in research studies are biochemistry, molecular biology, and immunohistochemistry (IHC). Biochemical methods take advantage of the chemical and physical techniques. A systematic literature search of studies started on 18 February 2023. The search was conducted in the main databases for biomedical literature, i.e., PubMed and Scopus, for papers published between 1973 and 2022, focusing on different techniques of immunohistochemistry and immunofluorescence (IF) for estimating the PTI of skin wounds. The present study involves a comprehensive and structured analysis of the existing literature to provide a detailed and comprehensive overview of the different IHC techniques used to date skin lesions, synthesize the available evidence, critically evaluate the methodologies, and eventually draw meaningful conclusions about the reliability and effectiveness of the different markers that have been discovered and used in wound age estimation.

Basic Quality Controls Used in Skin Tissue Engineering.

Reconstruction of skin defects is often a challenging effort due to the currently limited reconstructive options. In this sense, tissue engineering has emerged as a possible alternative to replace or repair diseased or damaged tissues from the patient's own cells. A substantial number of tissue-engineered skin substitutes (TESSs) have been conceived and evaluated in vitro and in vivo showing promising results in the preclinical stage. However, only a few constructs have been used in the clinic. The lack of standardization in evaluation methods employed may in part be responsible for this discrepancy. This review covers the most well-known and up-to-date methods for evaluating the optimization of new TESSs and orientative guidelines for the evaluation of TESSs are proposed.

In Vivo and In Vitro Toxicity Testing of Cyanobacterial Toxins: A Mini-Review.

Harmful cyanobacterial blooms are increasing and becoming a worldwide concern as many bloom-forming cyanobacterial species can produce toxic metabolites named cyanotoxins. These include microcystins, saxitoxins, anatoxins, nodularins, and cylindrospermopsins, which can adversely affect humans, animals, and the environment. Different methods to assess these classes of compounds in vitro and in vivo include biological, biochemical, molecular, and physicochemical techniques. Furthermore, toxic effects not attributable to known cyanotoxins can be observed when assessing bloom material. In order to determine exposures to cyanotoxins and to monitor compliance with drinking and bathing water guidelines, it is necessary to have reliable and effective methods for the analysis of these compounds. Many relatively simple low-cost methods can be employed to rapidly evaluate the potential hazard. The main objective of this mini-review is to describe the assessment of toxic cyanobacterial samples using in vitro and in vivo bioassays. Newly emerging cyanotoxins, the toxicity of analogs, or the interaction of cyanobacteria and cyanotoxins with other toxicants, among others, still requires bioassay assessment. This review focuses on some biological and biochemical assays (MTT assay, Immunohistochemistry, Micronucleus Assay, Artemia salina assay, Daphnia magna test, Radionuclide recovery, Neutral red cytotoxicity and Comet assay, Enzyme-Linked Immunosorbent Assay (ELISA), Annexin V-FITC assay and Protein Phosphatase Inhibition Assay (PPIA)) for the detection and measurement of cyanotoxins including microcystins, cylindrospermopsins, anatoxin-a, saxitoxins, and nodularins. Although most bioassay analyses often confirm the presence of cyanotoxins at low concentrations, such bioassays can be used to determine whether some strains or blooms of cyanobacteria may produce other, as yet unknown toxic metabolites. This review also aims to identify research needs and data gaps concerning the toxicity assessment of cyanobacteria.

Hydrothermal Liquefaction of Biomass as One of the Most Promising Alternatives for the Synthesis of Advanced Liquid Biofuels: A Review.

The use of biofuels offers advantages over existing fuels because they come from renewable sources, they are biodegradable, their storage and transport are safer, and their emissions into the atmosphere are lower. Biomass is one of the most promising sustainable energy sources with a wide variety of organic materials as raw material. Chemical, biochemical, and thermochemical methods have been proposed to obtain biofuels from raw materials from biomass. In recent years, a thermochemical method that has generated great interest is hydrothermal liquefaction. In this paper, a brief review of the main sources for liquid biofuels and the synthesis processes is presented, with special emphasis on the production of biofuels using hydrothermal liquefaction by using waste generated by human activity as raw material.

Performance of MALDI-TOF Mass Spectrometry in the Philippines.

Identification of the causative pathogen in infectious diseases is important for surveillance and to guide treatment. In low- and middle-income countries (LMIC), conventional culture and identification methods, including biochemical methods, are reference-standard. Biochemical methods can lack sensitivity and specificity and have slow turnaround times, causing delays in definitive therapy. Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) is a rapid and accurate diagnostic method. Most studies comparing MALDI-TOF MS and biochemical methods are from high-income countries, with few reports from LMIC with tropical climates. The aim of this study was to assess the performance of MALDI-TOF MS compared to conventional methods in the Philippines. Clinical bacterial or fungal isolates were identified by both MALDI-TOF MS and automated (VITEK2) or manual biochemical methods in the San Lazaro Hospital, Metro Manila, the Philippines. The concordance between MALDI-TOF MS and automated (VITEK2) or manual biochemical methods was analyzed at the species and genus levels. In total, 3530 bacterial or fungal isolates were analyzed. The concordance rate between MALDI-TOF MS and biochemical methods was 96.2% at the species level and 99.9% at the genus level. Twenty-three isolates could not be identified by MALDI-TOF MS. In this setting, MALDI-TOF MS was accurate compared with biochemical methods, at both the genus and the species level. Additionally, MALDI-TOF MS improved the turnaround time for results. These advantages could lead to improved infection management and infection control in low- and middle-income countries, even though the initial cost is high.

Valorisation and emerging perspective of biomass based waste-to-energy technologies and their socio-environmental impact: A review.

The economic developments around the globe resulted in the increased demand of energy, which overburdened the supply chain sources of energy. Fossil fuel reserves are exploited to meet the high demand of energy and their combustion is becoming the main source of environmental pollution. So there is dire need to find safe, renewable and sustainable energy resources. Waste to energy (WtE) may be viewed as a possible alternate source of energy, which is economically and environmentally sustainable. Municipal solid waste (MSW) is a major contributor to the development of renewable energy and sustainable environment. At present the scarcity of renewable energy resources and disposal of MSW is a challenging problem for the developing countries, which has generated a wide ranging socioeconomic and environmental problems. This situation stimulates the researchers to develop alternatives for converting WtE under a variety of scenarios. Herein, the present scenario in developing the WtE technologies such as, thermal conversion methods (Incineration, Gasification, Pyrolysis, Torrefaction), Plasma technology, Biochemical methods, Chemical and Mechanical methods, Bio-electrochemical process, Mechanical biological treatment (MBT), Photo-biological processes for efficacious energy recovery and the challenges confronted by developing and developed countries. In this review, a framework for the evaluation of WtE technologies has been presented for the ease of researchers working in the field. Furthermore, this review concluded that WtE is a potential renewable energy source that will partially satisfy the demand for energy and ensure an efficient MSW management to overcome the environmental pollution.

Distance teaching of experimental scientific methodology and scientific thinking.

Objective: The aim of this project was to convert a traditional face-to-face seminar for the teaching of experimental scientific methodology to remote teaching in a timely manner due to the COVID-19 related restrictions to teaching in presence. Methodology: The main focus of the course was on flow cytometry. Basics were developed in a virtual presence phase. Specific teaching contents were taught by an interactive presentation, which came very close to the user experience of a flow cytometer and interactively illustrated the influence of different experimental conditions on the obtained results. Video sequences of authentic sample acquisitions were integrated into Adobe Captivate®. These "virtual acquisitions" were not distinguishable from the original procedure. For interpretation of the resulting diagrams, interactions were inserted, which allowed direct comparison of the obtained results. Implementation: A presentation with interactive elements and video sequences was created and used for the virtual presence phases. After publishing on a web server in HTML 5, contents were made available to the students for post-processing of learning contents by self-paced learning with full (interactive) functionality. Conclusion: Contributions elaborated by the students during the course demonstrate a learning outcome comparable to that archieved in the last years in presence mode. While implementation of this solution represented a highly time-consuming process, narrative feedback was consistently positive. Due to the short time available for implementation, no systematic evaluation could be conducted, which represents a clear limitation of this work.

In Search of Herbal Anti-SARS-Cov2 Compounds.

On March 11, 2020, the World Health Organization (WHO) announced that the spread of the new coronavirus had reached the stage of a pandemic. To date (23.10.2020), there are more than 40 million confirmed cases of the disease in the world, at the same time there is still no effective treatment for the disease. For management and treatment of SARS-Cov2, the development of an antiviral drug is needed. Since the representatives of all human cultures have used medicinal plants to treat viral diseases throughout their history, plants can be considered as sources of new antiviral drug compounds against emerging viruses. The huge metabolic potential of plants allows us to expect discovery of plant compounds for the prevention and treatment of coronavirus infection. This idea is supported by number of papers on the anti-SARS-Cov2 activity of plant extracts and specific compounds in the experiments in silico, in vitro, and in vivo. Here, we summarize information on methods and approaches aimed to search for anti-SARS-Cov2 compounds including cheminformatics, bioinformatics, genetic engineering of viral targets, interacting with drugs, biochemical approaches etc. Our mini-review may be useful for better planning future experiments (including rapid methods for screening compounds for antiviral activity, the initial assessment of the antiviral potential of various plant species in relation to certain pathogens, etc.) and giving a hand to those who are making first steps in this field.

Probing the function of long noncoding RNAs in the nucleus.

The nucleus is a highly organized and dynamic environment where regulation and coordination of processes such as gene expression and DNA replication are paramount. In recent years, noncoding RNAs have emerged as key participants in the regulation of nuclear processes. There are a multitude of functional roles for long noncoding RNA (lncRNA), mediated through their ability to act as molecular scaffolds bridging interactions with proteins, chromatin, and other RNA molecules within the nuclear environment. In this review, we discuss the diversity of techniques that have been developed to probe the function of nuclear lncRNAs, along with the ways in which those techniques have revealed insights into their mechanisms of action. Foundational observations into lncRNA function have been gleaned from molecular cytology-based, single-cell approaches to illuminate both the localization and abundance of lncRNAs in addition to their potential binding partners. Biochemical, extraction-based approaches have revealed the molecular contacts between lncRNAs and other molecules within the nuclear environment and how those interactions may contribute to nuclear organization and regulation. Using examples of well-studied nuclear lncRNAs, we demonstrate that the emerging functions of individual lncRNAs have been most clearly deduced from combined cytology and biochemical approaches tailored to study specific lncRNAs. As more functional nuclear lncRNAs continue to emerge, the development of additional technologies to study their interactions and mechanisms of action promise to continually expand our understanding of nuclear organization, chromosome architecture, genome regulation, and disease states.

Integrated approaches to unravel the impact of protein lipoxidation on macromolecular interactions.

Protein modification by lipid derived reactive species, or lipoxidation, is increased during oxidative stress, a common feature observed in many pathological conditions. Biochemical and functional consequences of lipoxidation include changes in the conformation and assembly of the target proteins, altered recognition of ligands and/or cofactors, changes in the interactions with DNA or in protein-protein interactions, modifications in membrane partitioning and binding and/or subcellular localization. These changes may impact, directly or indirectly, signaling pathways involved in the activation of cell defense mechanisms, but when these are overwhelmed they may lead to pathological outcomes. Mass spectrometry provides state of the art approaches for the identification and characterization of lipoxidized proteins/residues and the modifying species. Nevertheless, understanding the complexity of the functional effects of protein lipoxidation requires the use of additional methodologies. Herein, biochemical and biophysical methods used to detect and measure functional effects of protein lipoxidation at different levels of complexity, from in vitro and reconstituted cell-like systems to cells, are reviewed, focusing especially on macromolecular interactions. Knowledge generated through innovative and complementary technologies will contribute to comprehend the role of lipoxidation in pathophysiology and, ultimately, its potential as target for therapeutic intervention.

A pan-European ring trial to validate an International Standard for detection of Vibrio cholerae, Vibrio parahaemolyticus and Vibrio vulnificus in seafoods.

Globally, vibrios represent an important and well-established group of bacterial foodborne pathogens. The European Commission (EC) mandated the Comite de European Normalisation (CEN) to undertake work to provide validation data for 15 methods in microbiology to support EC legislation. As part of this mandated work programme, merging of ISO/TS 21872-1:2007, which specifies a horizontal method for the detection of V. parahaemolyticus and V. cholerae, and ISO/TS 21872-2:2007, a similar horizontal method for the detection of potentially pathogenic vibrios other than V. cholerae and V. parahaemolyticus was proposed. Both parts of ISO/TS 21872 utilized classical culture-based isolation techniques coupled with biochemical confirmation steps. The work also considered simplification of the biochemical confirmation steps. In addition, because of advances in molecular based methods for identification of human pathogenic Vibrio spp. classical and real-time PCR options were also included within the scope of the validation. These considerations formed the basis of a multi-laboratory validation study with the aim of improving the precision of this ISO technical specification and providing a single ISO standard method to enable detection of these important foodborne Vibrio spp.. To achieve this aim, an international validation study involving 13 laboratories from 9 countries in Europe was conducted in 2013. The results of this validation have enabled integration of the two existing technical specifications targeting the detection of the major foodborne Vibrio spp., simplification of the suite of recommended biochemical identification tests and the introduction of molecular procedures that provide both species level identification and discrimination of putatively pathogenic strains of V. parahaemolyticus by the determination of the presence of theromostable direct and direct related haemolysins. The method performance characteristics generated in this have been included in revised international standard, ISO 21872:2017, published in July 2017.

Insights Into Protein S-Palmitoylation in Synaptic Plasticity and Neurological Disorders: Potential and Limitations of Methods for Detection and Analysis.

S-palmitoylation (S-PALM) is a lipid modification that involves the linkage of a fatty acid chain to cysteine residues of the substrate protein. This common posttranslational modification (PTM) is unique among other lipid modifications because of its reversibility. Hence, like phosphorylation or ubiquitination, it can act as a switch that modulates various important physiological pathways within the cell. Numerous studies revealed that S-PALM plays a crucial role in protein trafficking and function throughout the nervous system. Notably, the dynamic turnover of palmitate on proteins at the synapse may provide a key mechanism for rapidly changing synaptic strength. Indeed, palmitate cycling on postsynaptic density-95 (PSD-95), the major postsynaptic density protein at excitatory synapses, regulates the number of synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) and thus affects synaptic transmission. Accumulating evidence suggests a relationship between impairments in S-PALM and severe neurological disorders. Therefore, determining the precise levels of S-PALM may be essential for understanding the ways in which this PTM is regulated in the brain and controls synaptic dynamics. Protein S-PALM can be characterized using metabolic labeling methods and biochemical tools. Both approaches are discussed herein in the context of specific methods and their advantages and disadvantages. This review clearly shows progress in the field, which has led to the development of new, more sensitive techniques that enable the detection of palmitoylated proteins and allow predictions of potential palmitate binding sites. Unfortunately, one significant limitation of these approaches continues to be the inability to use them in living cells.

Nuclear factor (erythroid-derived 2)-like 2 (NRF2) drug discovery: Biochemical toolbox to develop NRF2 activators by reversible binding of Kelch-like ECH-associated protein 1 (KEAP1).

Mechanisms that activate innate antioxidant responses, as a way to mitigate oxidative stress at the site of action, hold much therapeutic potential in diseases, such as Parkinson's disease, Alzheimer's disease and Huntington's disease, where the use of antioxidants as monotherapy has not yielded positive results. The nuclear factor NRF2 is a transcription factor whose activity upregulates the expression of cell detoxifying enzymes in response to oxidative stress. NRF2 levels are modulated by KEAP1, a sensor of oxidative stress. KEAP1 binds NRF2 and facilitates its ubiquitination and subsequent degradation. Recently, compounds that reversibly disrupt the NRF2-KEAP1 interaction have been described, opening the field to a new era of safer NRF2 activators. This paper describes a set of new, robust and informative biochemical assays that enable the selection and optimization of non-covalent KEAP1 binders. These include a time-resolved fluorescence resonance energy transfer (TR-FRET) primary assay with high modularity and robustness, a surface plasmon resonance (SPR) based KEAP1 direct binding assay that enables the quantification and analysis of full kinetic binding parameters and finally a 1H-15N heteronuclear single quantum coherence (HSQC) NMR assay suited to study the interaction surface of KEAP1 with residue-specific information to validate the interaction of ligands in the KEAP1 binding site.

Current Experimental Methods for Characterizing Protein-Protein Interactions.

Protein molecules often interact with other partner protein molecules in order to execute their vital functions in living organisms. Characterization of protein-protein interactions thus plays a central role in understanding the molecular mechanism of relevant protein molecules, elucidating the cellular processes and pathways relevant to health or disease for drug discovery, and charting large-scale interaction networks in systems biology research. A whole spectrum of methods, based on biophysical, biochemical, or genetic principles, have been developed to detect the time, space, and functional relevance of protein-protein interactions at various degrees of affinity and specificity. This article presents an overview of these experimental methods, outlining the principles, strengths and limitations, and recent developments of each type of method.

Direct Detection of Burkholderia cepacia in Susceptible Pharmaceutical Products Using Semi-Nested PCR.

Burkholderia cepaciahas recently received a considerable attention as one of the major risks in susceptible pharmaceutical products. This microorganism can easily propagate and cause vast and severe contamination, especially to the water supplies for pharmaceutical companies. Moreover, it proliferates within the products and can cause severe infections for humans. Therefore, fast and sensitive detection of these bacteria is of a great demand. The present study introduces improved application of a polymerase chain reaction assay with relatively high sensitivity and specificity for the direct detection ofB. cepaciafrom the aqueous pharmaceutical products. A semi-nested polymerase chain reaction approach using the primer set BCR1/BCR2 followed by BCR1/Mr yielding a 465 bp fragment of the recA gene was applied and tested using both crude lysate from isolated colonies and DNA directly extracted from artificially prepared and spiked reference syrup. The polymerase chain reaction assay showed no interference with other bacterial reference and environmental strains tested, includingStaphylococcus aureusATCC® 6538,Pseudomonas aeruginosaATCC® 9027,Escherichia coliATCC® 8739,Salmonella abonyNCTC® 6017,Bacillus subtilisATCC® 6633,Micrococcus luteus, Staphylococcus warneri, Pseudomonas fluorescens, Pseudomonas putida, andRalstonia pickettii Moreover, this semi-nested assay showed a detection limit of around 10 colony-forming units per sample and could detectB. cepaciastrains isolated from a municipal pre-treated potable water tank. Comparing the results for detection ofB. cepaciain 100 randomly collected commercial syrup preparations using both conventional standard method and polymerase chain reaction assay revealed thatB. cepaciawas detected in two samples using polymerase chain reaction assay while all samples showed negative results by conventional culturing and biochemical methods. These results highlight the advantage of using this polymerase chain reaction assay to detectB. cepaciain contaminated pharmaceutical products and even water for pharmaceutical purposes, without the need of culturing or pre-enrichment, where it may give false-negative results and may be misidentified when biochemically tested.

Métodos para la determinación del estado del magnesio en humanos / Methods of assessment of magnesium status in humans / Métodos para a determinação do estado do magnésio em humanos

En la actualidad, datos epidemiológicos sugieren que, en países occidentales, la ingesta de magnesio no satisface la ingesta recomendada, lo que apoya un riesgo de deficiencia de magnesio latente en estas poblaciones. La evaluación del estado de magnesio sigue siendo un desafío para el laboratorio clínico ya que el magnesio se encuentra distribuido mayoritariamente en el hueso y tejidos blandos. Existe la necesidad de conciliación entre una prueba de fácil acceso, rápida, sensible y representativa del magnesio intracelular. La utilidad de diferentes biomarcadores en sujetos sanos ha sido evaluada; se ha reportado que el magnesio en plasma, eritrocitos y orina parecen ser biomarcadores sensibles a la ingesta dietética y útiles como biomarcadores en la población general. Sin embargo, esto no es concluyente, ya que se resalta que aún se requieren estudios mejor diseñados, que impliquen factores como mayor población empleada, dosis y tiempo de suplementación. El progreso en la genética y la genómica abren perspectivas interesantes en la búsqueda de estos biomarcadores que permitan cuantificar los niveles de magnesio celular así como también las reservas de todo el cuerpo, para poder así establecer recomendaciones dietéticas mejor ajustadas a la población.

Epidemiological studies suggest that dietary magnesium in the Western countries does not meet the recommended intake, supporting a risk of latent magnesium deficiency with Western diet behavior. Assessment of magnesium status remains a major challenge for the clinical laboratory, since, magnesium storage is mostly found in bone and soft tissues. The conciliation between an easy obtained sample, rapid and robust laboratory test, and the parameter representative for intracellular magnesium is extremely difficult to reach. In a current systematic review study, the usefulness of magnesium status biomarkers in healthy subjects has been evaluated. It is proposed that plasma and erythrocyte magnesium, and urinary magnesium excretion which respond to dietary manipulation appear to be useful biomarkers in the general population. However, it is emphasized that well-designed studies of sufficient size with varying doses and duration of magnesium supplementation are still required. The development of specific and sensible biomarkers, making it possible to obtain cell magnesium levels as well as body magnesium pool evaluation, relevant to study individuals, small and large populations, remains a major challenge for the assessment of magnesium status. A progress in genetics and genomics opens new interesting perspectives in the search of these biomarkers.

Na atualidade, dados epidemiológicos sugerem que, nos países ocidentais, a ingestão de magnésio não supre a ingestão recomendada, o que apoia um risco de deficiência de magnésio latente nestas populações. A avaliação do estado do magnésio continua sendo um desafio para o laboratório clínico, visto que o magnésio se encontra distribuído principalmente no osso e nos tecidos moles. Há a necessidade de conciliar evidência facilmente acessível, rápida, sensível e representativa do magnésio intracelular. A utilidade de vários biomarcadores em indivíduos saudáveis foi avaliada, e foi relatado que o magnésio em plasma, eritrócitos e urina parecem ser biomarcadores sensíveis à ingestão dietética e úteis como biomarcadores na população geral. No entanto, esta não é conclusiva, uma vez que se destaca que são requeridos ainda estudos melhor desenhados, envolvendo fatores como utilização de maior população, dosagem e tempo de suplementação. Um avanço na genética e na genômica abre perspectivas interessantes na busca desses biomarcadores para poder quantificar os níveis de magnésio celular bem como as reservas do corpo inteiro, e assim poder estabelecer melhores recomendações na dieta adaptadas à população.

Métodos para la determinación del estado del magnesio en humanos / Methods of assessment of magnesium status in humans / Métodos para a determinaþÒo do estado do magnésio em humanos

En la actualidad, datos epidemiológicos sugieren que, en países occidentales, la ingesta de magnesio no satisface la ingesta recomendada, lo que apoya un riesgo de deficiencia de magnesio latente en estas poblaciones. La evaluación del estado de magnesio sigue siendo un desafío para el laboratorio clínico ya que el magnesio se encuentra distribuido mayoritariamente en el hueso y tejidos blandos. Existe la necesidad de conciliación entre una prueba de fácil acceso, rápida, sensible y representativa del magnesio intracelular. La utilidad de diferentes biomarcadores en sujetos sanos ha sido evaluada; se ha reportado que el magnesio en plasma, eritrocitos y orina parecen ser biomarcadores sensibles a la ingesta dietética y útiles como biomarcadores en la población general. Sin embargo, esto no es concluyente, ya que se resalta que aún se requieren estudios mejor diseñados, que impliquen factores como mayor población empleada, dosis y tiempo de suplementación. El progreso en la genética y la genómica abren perspectivas interesantes en la búsqueda de estos biomarcadores que permitan cuantificar los niveles de magnesio celular así como también las reservas de todo el cuerpo, para poder así establecer recomendaciones dietéticas mejor ajustadas a la población.(AU)

Epidemiological studies suggest that dietary magnesium in the Western countries does not meet the recommended intake, supporting a risk of latent magnesium deficiency with Western diet behavior. Assessment of magnesium status remains a major challenge for the clinical laboratory, since, magnesium storage is mostly found in bone and soft tissues. The conciliation between an easy obtained sample, rapid and robust laboratory test, and the parameter representative for intracellular magnesium is extremely difficult to reach. In a current systematic review study, the usefulness of magnesium status biomarkers in healthy subjects has been evaluated. It is proposed that plasma and erythrocyte magnesium, and urinary magnesium excretion which respond to dietary manipulation appear to be useful biomarkers in the general population. However, it is emphasized that well-designed studies of sufficient size with varying doses and duration of magnesium supplementation are still required. The development of specific and sensible biomarkers, making it possible to obtain cell magnesium levels as well as body magnesium pool evaluation, relevant to study individuals, small and large populations, remains a major challenge for the assessment of magnesium status. A progress in genetics and genomics opens new interesting perspectives in the search of these biomarkers.(AU)

Na atualidade, dados epidemiológicos sugerem que, nos países ocidentais, a ingestÒo de magnésio nÒo supre a ingestÒo recomendada, o que apoia um risco de deficiÛncia de magnésio latente nestas populaþ§es. A avaliaþÒo do estado do magnésio continua sendo um desafio para o laboratório clínico, visto que o magnésio se encontra distribuído principalmente no osso e nos tecidos moles. Há a necessidade de conciliar evidÛncia facilmente acessível, rápida, sensível e representativa do magnésio intracelular. A utilidade de vários biomarcadores em indivíduos saudáveis foi avaliada, e foi relatado que o magnésio em plasma, eritrócitos e urina parecem ser biomarcadores sensíveis O ingestÒo dietética e úteis como biomarcadores na populaþÒo geral. No entanto, esta nÒo é conclusiva, uma vez que se destaca que sÒo requeridos ainda estudos melhor desenhados, envolvendo fatores como utilizaþÒo de maior populaþÒo, dosagem e tempo de suplementaþÒo. Um avanþo na genética e na gen¶mica abre perspectivas interessantes na busca desses biomarcadores para poder quantificar os níveis de magnésio celular bem como as reservas do corpo inteiro, e assim poder estabelecer melhores recomendaþ§es na dieta adaptadas O populaþÒo.(AU)

Diversity in parasitic helminths of Australasian marsupials and monotremes: a molecular perspective.

Marsupials and monotremes are a prominent part of the mammalian fauna in Australia, and harbour an extremely diverse and highly distinctive array of helminth parasites. Their study has been relatively neglected, likely because they have no direct, adverse socioeconomic impact. As the body plans of helminths generally are very simple and morphological characterisation likely underestimates true diversity, molecular tools have been employed to assess genetic diversity. Using biochemical and/or molecular methods, recent studies show extensive diversity in helminths of marsupials, with cryptic species being commonly encountered. The purpose of this article is to review current knowledge about the diversity of parasitic helminths of marsupials and monotremes, to raise questions as to whether current molecular data can be used to estimate diversity, what mechanisms lead to such diversity, to critically appraise the molecular tools that have been employed thus far to explore diversity and to discuss the directions which might be taken in the future employing improved techniques.