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BACKGROUND: Infection with Helicobacter pylori (Hp) mostly occurs during childhood, and persistent infection may lead to severe gastric diseases and even gastric cancer. Currently, the primary method for eradicating Hp is through antibiotic treatment. However, the increasing multidrug resistance in Hp strains has diminished the effectiveness of antibiotic treatments. Vaccination could potentially serve as an effective intervention to resolve this issue. AIMS: Through extensive research and analysis of the vital protein characteristics involved in Hp infection, we aim to provide references for subsequent vaccine antigen selection. Additionally, we summarize the current research and development of Hp vaccines in order to provide assistance for future research. MATERIALS AND METHODS: Utilizing the databases PubMed and the Web of Science, a comprehensive search was conducted to compile articles pertaining to Hp antigens and vaccines. The salient aspects of these articles were then summarized to provide a detailed overview of the current research landscape in this field. RESULTS: Several potential antigens have been identified and introduced through a thorough understanding of the infection process and pathogenic mechanisms of Hp. The conserved and widely distributed candidate antigens in Hp, such as UreB, HpaA, GGT, and NAP, are discussed. Proteins such as CagA and VacA, which have significant virulence effects but relatively poor conservatism, require further evaluation. Emerging antigens like HtrA and dupA have significant research value. In addition, vaccines based on these candidate antigens have been compiled and summarized. CONCLUSIONS: Vaccines are a promising method for preventing and treating Hp. While some Hp vaccines have achieved promising results, mature products are not yet available on the market. Great efforts have been directed toward developing various types of vaccines, underscoring the need for developers to select appropriate antigens and vaccine formulations to improve success rates.
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Antígenos de Bactérias , Vacinas Bacterianas , Infecções por Helicobacter , Helicobacter pylori , Desenvolvimento de Vacinas , Helicobacter pylori/imunologia , Vacinas Bacterianas/imunologia , Infecções por Helicobacter/prevenção & controle , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/microbiologia , Humanos , Antígenos de Bactérias/imunologia , AnimaisRESUMO
Transition-metal dichalcogenides (TMDs) have shown great promise as selective and high-capacity sorbents for Hg(II) removal from water. Yet, their design should consider safe disposal of spent materials, particularly the subsequent formation of methylmercury (MeHg), a highly potent and bioaccumulative neurotoxin. Here, we show that microbial methylation of mercury bound to MoS2 nanosheets (a representative TMD material) is significant under anoxic conditions commonly encountered in landfills. Notably, the methylation potential is highly dependent on the phase compositions of MoS2. MeHg production was higher for 1T MoS2, as mercury bound to this phase primarily exists as surface complexes that are available for ligand exchange. In comparison, mercury on 2H MoS2 occurs largely in the form of precipitates, particularly monovalent mercury minerals (e.g., Hg2MoO4 and Hg2SO4) that are minimally bioavailable. Thus, even though 1T MoS2 is more effective in Hg(II) removal from aqueous solution due to its higher adsorption affinity and reductive ability, it poses a higher risk of MeHg formation after landfill disposal. These findings highlight the critical role of nanoscale surfaces in enriching heavy metals and subsequently regulating their bioavailability and risks and shed light on the safe design of heavy metal sorbent materials through surface structural modulation.
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Mercúrio , Metilação , Adsorção , Nanoestruturas/química , Poluentes Químicos da Água/química , Compostos de Metilmercúrio , Molibdênio/químicaRESUMO
Strain estimation is vital in phase-sensitive optical coherence elastography (PhS-OCE). In this Letter, we introduce a novel, to the best of our knowledge, method to improve strain estimation by using a dual-convolutional neural network (Dual-CNN). This approach requires two sets of PhS-OCE systems: a high-resolution system for high-quality training data and a cost-effective standard-resolution system for practical measurements. During training, high-resolution strain results acquired from the former system and the pre-existing strain estimation CNN serve as label data, while the narrowed light source-acquired standard-resolution phase results act as input data. By training a new network with this data, high-quality strain results can be estimated from standard-resolution PhS-OCE phase results. Comparison experiments show that the proposed Dual-CNN can preserve the strain quality even when the light source bandwidth is reduced by over 80%.
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Nanoplastics are an increasing environmental concern. In aquatic environments, nanoplastics will acquire an eco-corona by interacting with macromolecules (e.g., humic substances and extracellular polymeric substances (EPS)). Here, we show that the properties of the eco-corona and, consequently, its ability to enhance the transport of nanoplastics vary significantly with the surface functionality of nanoplastics and sources of macromolecules. The eco-corona derived from the EPS of Gram-negative Escherichia coli MG1655 enhances the transport of polystyrene (PS) nanospheres in saturated porous media to a much greater extent than the eco-corona derived from soil humic acid and fulvic acid. In comparison, the eco-corona from all three sources significantly enhance the transport of carboxylated PS (HOOC-PS). We show that the eco-corona inhibits the deposition of the two types of nanoplastics to the porous media mainly via steric repulsion. Accordingly, an eco-corona consisting of a higher mass of larger-sized macromolecules is generally more effective in enhancing transport. Notably, HOOC-PS tends to acquire macromolecules of lower hydrophobicity than PS. The more disordered and flexible structures of such macromolecules may result in greater elastic repulsion between the nanoplastics and sand grains and, consequently, greater transport enhancement. The findings of this study highlight the critical role of eco-corona formation in regulating the mobility of nanoplastics, as well as the complexity of this process.
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Microplásticos , Nanosferas , Porosidade , Solo , Poliestirenos , Substâncias Húmicas/análiseRESUMO
Due to the extremely low solubility, mercury sulfide minerals, as the major environmental mercury sinks, are generally considered to be inert mercury species with minimal bioavailability. Here, we demonstrate that extracellular polymeric substances (EPS), continuously secreted and released by anaerobic methylating bacteria, enhance the dissolution processes of cinnabar (α-HgS) minerals. The enhancing effects of EPS occur to a greater extent in the dissolution of nanoparticulate α-HgS compared to the bulk-scale counterpart. The released EPS-Hg(II) species are available for microbial methylation to produce bioaccumulative neurotoxin, methylmercury. This is probably due to the abundant aromatic proteins in EPS that strongly interact with surface Hg(II) via inner-sphere complexation as well as cation-π interaction. Our study discovers the potential environmental risks of "inert" mercury sulfide minerals in natural microbial habitats, particularly benthic biofilms with abundant microbial EPS, transformed to the severely toxic methylmercury. The mechanistic findings will facilitate an accurate understanding of the interactions between soft and transition metals and microorganism-derived organics, which may dictate the environmental fate and impact of these elements.
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Mercúrio , Compostos de Metilmercúrio , Compostos de Metilmercúrio/toxicidade , Matriz Extracelular de Substâncias Poliméricas/metabolismo , Solubilidade , Metilação , Mercúrio/toxicidade , SulfetosRESUMO
The distribution of nanoparticles between aqueous and organic phases is universally considered as the starting point in predicting the fate and bioavailability of engineered nanoparticles in the environment. However, the theoretical basis for determining the distribution of nanoparticles in the immiscible water-oil system remains unclear. Here, for the first time, theoretical calculations were conducted to illustrate the underlying mechanism. It was suggested that the distribution of nanoparticles was largely controlled by the surface charge, particle size and surface hydrophobicity, and the water-oil interface was not the favorable phase for nanoparticles until a size threshold (10 nm) was met and the particle surface became amphiphilic. The theoretical results were verified by the experimental approaches of different nanoparticles distributed in the water-octanol mixture. The neutralization of a charged surface led to enhanced distribution into octanol for hydrophobic nanoparticles (e.g., aqueous C60), yet it had little effect on hydrophilic nanoparticles (e.g., fullerol). More nanoparticles were trapped at the water-oil interface when size grew larger (e.g., Ag-CIT and Au-CIT with citrate) and the surface rendered amphiphilic by polymeric coatings (e.g., Ag-PVP with polyvinylpyrrolidone), though larger hydrophobic nanoparticles like aqu-nC60 tended to stay in the octanol. The surface charge and hydrophobicity may have an important impact on the path-dependent distribution of nanoparticles in water- octanol system. The mechanistic insights based on theoretical calculations and experimental approaches will facilitate the accurate prediction of the distribution of engineered nanoparticles in biological and environmental systems.
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Nanopartículas , Água , Água/química , Interações Hidrofóbicas e Hidrofílicas , Povidona/química , Octanóis , Nanopartículas/químicaRESUMO
An effective strategy to construct low fouling electrochemical biosensors for assaying serum biomarkers was proposed based on specially designed α-aminoisobutyric acid (Aib) incorporated peptides. The Aib-peptides were designed to be of antifouling properties, and at the same to incorporate Aib residues in their interior to enhance the hydrolytic stability. In order to construct the electrochemical biosensor, two kinds of Aib-peptides labelled with biotin were modified on the electrode surface: One with cysteine terminal for easy attachment to the electrode modified with gold nanoparticles, the other with unique terminal peptide sequence for specific binding of immunoglobulin G (IgG), and they were connected through the streptavidin-biotin affinity system. Owing to the interposition of Aib residues, the peptides as well as the constructed biosensors showed excellent antifouling performances and enhanced stability against enzymatic degradation in serum. Furthermore, the IgG biosensor constructed with the Aib-peptides displayed a very low detection limit (29.5 pg mL-1) and a broad linear range (100 pg mL-1 - 10 µg mL-1), and it was able to assay IgG in clinical human sera with decent accuracy and reliability. This strategy provides a new path for the construction of stable antifouling biosensors based on functional peptides for practical biomarker assaying in real clinical samples.
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Técnicas Biossensoriais , Nanopartículas Metálicas , Humanos , Biotina , Ouro , Reprodutibilidade dos Testes , Peptídeos , Imunoglobulina GRESUMO
Despite the extensive study of tetracycline photolysis in aquatic environments, the phototransformation of tetracycline and its metabolites under natural day-night succession has not been examined. In this study, we investigated tetracycline photolysis and associated ecotoxicity in two natural surface waters and one artificial ultrapure water under simulated day/night cycling over two days. Previously unrecognized and highly pH- and temperature-dependent dark interconversions of tetracycline metabolites were observed. The liquid chromatography-mass spectrometry/mass spectrometry analysis identified a range of isomerized, hydroxylated, demethylated, deaminated, and open-ring photoproducts. The hydrolysis of tetracycline, isotetracycline, and several intermediate products was proposed as the major mechanism for the observed dark transformations. Exposure studies employing Escherichia coli indicated that although the tetracycline degradation products had lower bacterial toxicities than the parent compound, increasing toxicity with irradiation time after the near-complete degradation of the parent compound in natural waters implied that product mixtures retain ecotoxicity. The dark transformations also affected the bacterial toxicity and fluorescence properties of irradiated tetracycline solutions. Overall, this study provides new insights into the photochemical behavior of tetracycline and its associated ecological risk in aquatic environments.
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Poluentes Químicos da Água , Fotólise , Poluentes Químicos da Água/química , Tetraciclina/análise , Antibacterianos/química , Escherichia coli , Água , CinéticaRESUMO
Microplastics (MPs) tend to accumulate and undergo a sulfur weathering process that leads to significant surface changes in sulfur-rich anaerobic environments, such as sewage and wastewater treatment plants. Aged MPs can have a profound impact on environmental behaviors of various toxic pollutants, especially heavy metals. Although previous studies have investigated the adsorption characteristics of metal ions on MPs that are aged in aerobic environments, the sorptive interactions of sulfur-aged MPs in anaerobic environments with mercury, i.e., Hg(II), are largely unknown. In this study, laboratory investigations were conducted to study the sorptive behaviors of Hg(II) by six common MPs treated anaerobically in the presence of sulfide. Adsorption isotherms show that the sulfur aging process greatly enhances the MP sorption capacity of Hg(II). The mechanisms including changes in the specific surface area, electrostatic interactions, surface precipitation, and surface functional groups are responsible for the enhanced adsorption capacities of sulfur-aged MPs. The thiol group that forms on the MP surface plays a dominant role in enhancing the MP adsorption capacity of Hg(II), which is determined by the formation of unsaturated bonds in the molecular chains of MPs. Furthermore, the pathways of surface chemical transformation of MPs during sulfur aging have been proposed. This study promotes our understanding of the potential hazard of MPs as well as the fate and transport of heavy metals in the presence of aged MPs.
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Poluentes Ambientais , Mercúrio , Metais Pesados , Adsorção , Poluentes Ambientais/química , Microplásticos , Plásticos/química , Esgotos , Compostos de Sulfidrila , Sulfetos , EnxofreRESUMO
Synthetic genomics will advance our understanding of life and allow us to rebuild the genomes of industrial microorganisms for enhancing performances. Corynebacterium glutamicum, a Gram-positive bacterium, is an important industrial workhorse. However, its genome synthesis is impeded by the low efficiencies in DNA delivery and in genomic recombination/replacement. In the present study, we describe a genomic iterative replacement system based on RecET recombination for C. glutamicum, involving the successive integration of up to 10 kb DNA fragments obtained in vitro, and the transformants are selected by the alternative use of kanR and speR selectable markers. As a proof of concept, we systematically redesigned and replaced a 54.3 kb wild-type sequence of C. glutamicumATCC13032 with its 55.1 kb synthetic counterpart with several novel features, including decoupled genes, the standard PCRTags, and 20 loxPsym sites, which was for the first time incorporated into a bacterial genome. The resulting strain semi-synCG-A1 had a phenotype and fitness similar to the wild-type strain under various stress conditions. The stability of the synthetic genome region faithfully maintained over 100 generations of nonselective growth. Genomic deletions, inversions, and translocations occurred in the synthetic genome region upon induction of synthetic chromosome rearrangement and modification by loxP-mediated evolution (SCRaMbLE), revealing potential genetic flexibility for C. glutamicum. This strategy can be used for the synthesis of a larger region of the genome and facilitate the endeavors for metabolic engineering and synthetic biology of C. glutamicum.
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Corynebacterium glutamicum , Corynebacterium glutamicum/metabolismo , Genoma Bacteriano/genética , Genômica , Engenharia Metabólica/métodos , Biologia SintéticaRESUMO
Growing evidence has suggested that microbial biofilms are potential environmental "hotspots" for the production and accumulation of a bioaccumulative neurotoxin, methylmercury. Here, we demonstrate that extracellular polymeric substances (EPS), the main components of biofilm matrices, significantly interfere with mercury sulfide precipitation and lead to the formation of nanoparticulate metacinnabar available for microbial methylation, a natural process predominantly responsible for the environmental occurrence of methylmercury. EPS derived from mercury methylating bacteria, particularly Desulfovibrio desulfuricans ND132, substantially increase the methylation potential of nanoparticulate mercury. This is likely due to the abundant aromatic biomolecules in EPS that strongly interact with mercury sulfide via inner-sphere complexation and consequently enhance the short-range structural disorder while mitigating the aggregation of nanoparticulate mercury. The EPS-elevated bioavailability of nanoparticulate mercury to D. desulfuricans ND132 is not induced by dissolution of these nanoparticles in aqueous phase, and may be dictated by cell-nanoparticle interfacial reactions. Our discovery is the first step of mechanistically understanding methylmercury production in biofilms. These new mechanistic insights will help incorporate microbial EPS and particulate-phase mercury into mercury methylation models, and may facilitate the assessment of biogeochemical cycling of other nutrient or toxic elements driven by EPS-producing microorganisms that are prevalent in nature.
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Mercúrio , Compostos de Metilmercúrio , Nanopartículas , Matriz Extracelular de Substâncias Poliméricas , Metilação , SulfetosRESUMO
Natural organic matter (NOM) and crystalline metal oxide nanoparticles are both prevalent in natural aquatic environments, and their interactions have important environmental and biogeochemical implications. Here, we show that these interactions are significantly affected by an intrinsic property of metal oxide nanocrystals, the exposed facets. Both anatase (TiO2) and hematite (α-Fe2O3) nanocrystals, representing common engineered and naturally occurring metal oxides, exhibited apparent facet-dependent adsorption of humic acid and fulvic acid. This facet-dependent binding was primarily driven by surface complexation between the NOM carboxyl groups and surficial metal atoms. Thus, the adsorption affinity of different-faceted nanocrystals was determined by the atomic arrangements of crystal facets that controlled the activity of metal atoms and, consequently, the ligand exchange and binding configuration of the carboxyl groups in the first hydration shell of nanocrystals. Distinct facet-dependent fractionation patterns were observed during adsorption of NOM components, particularly the low-molecular-weight and photorefractory constituents. The molecular fractionation of NOM between water and metal oxide nanoparticles was dictated by the combined effects of facet-dependent metal complexation, hydrophobic interaction, and steric hindrance and may significantly influence the NOM-driven processes occurring both in aqueous phases and at water-nanoparticle interfaces.
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Nanopartículas Metálicas , Nanopartículas , Adsorção , Substâncias Húmicas/análise , Compostos Orgânicos , ÓxidosRESUMO
BACKGROUND: Osteoarthritis (OA) is one of the most common rheumatic diseases of which clinical symptoms includes swelling, synovitis and inflammatory pain, affect patients' daily life. It was reported that non-coding RNAs play vital roles in OA. However, the regulation mechanism of ncRNA in OA pathogenesis has not been fully elucidated. METHODS: The expression of SNHG7, miR-34a-5p and SYVN1 was detected using qRT-PCR in tissues, serum and cells. The protein expression of SYVN1, PCNA, cleavage-caspase 3, beclin1 and LC3 were measured using western blot. The RNA immunoprecipitation (RIP), RNA pulldown, and luciferase reporter assays were used to verify the relationship between SNHG7, miR-34a-5p and SYVN1. The MTT and flow cytometry assay was performed to detected cell proliferation and cell apoptosis respectively. RESULTS: In this study, SNHG7 and SYVN1 expression were down-regulated, but miR-34a-5p was up-regulated in OA tissues and IL-1ß treated cells compared with normal tissues and chondrocyte. Functional investigation revealed that up-regulated SNHG7 or down-regulated miR-34a-5p could promote cell proliferation and inhibit cell apoptosis and autophagy in OA cells. More than that, RIP, pulldown and luciferase reporter assay was applied to determine that miR-34a-5p was a target miRNA of SNHG7 and SYVN1 was a target mRNA of miR-34-5p. Rescue experiments showed that overexpression of miR-34a reversed high expression of SNHG7-mediated suppression of apoptosis and autophagy as well as promotion of proliferation, while its knockdown inhibited cell apoptosis and autophagy and promoted cell proliferation which could be impaired by silencing SYVN1. In addition, SNHG7 regulated SYVN1 through sponging miR-34a-5p. CONCLUSION: SNHG7 sponged miR-34a-5p to affect cell proliferation, apoptosis and autophagy through targeting SYVN1 which provides a novel sight into the pathogenesis of OA.
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Apoptose/fisiologia , Autofagia/fisiologia , MicroRNAs/metabolismo , Osteoartrite/metabolismo , RNA Longo não Codificante/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Apoptose/genética , Autofagia/genética , Western Blotting , Proliferação de Células , Regulação para Baixo , Ensaio de Imunoadsorção Enzimática , Humanos , MicroRNAs/genética , Osteoartrite/genética , RNA Longo não Codificante/genética , Reação em Cadeia da Polimerase em Tempo Real , Ubiquitina-Proteína Ligases/genética , Regulação para CimaRESUMO
BACKGROUND: Osteoarthritis (OA) is one of the most common rheumatic diseases of which clinical symptoms includes swelling, synovitis and inflammatory pain, affect patients' daily life. It was reported that non-coding RNAs play vital roles in OA. However, the regulation mechanism of ncRNA in OA pathogenesis has not been fully elucidated. METHODS: The expression of SNHG7, miR-34a-5p and SYVN1 was detected using qRT-PCR in tissues, serum and cells. The protein expression of SYVN1, PCNA, cleavage-caspase 3, beclinl and LC3 were measured using western blot. The RNA immunoprecipitation (RIP), RNA pulldown, and luciferase reporter assays were used to verify the relationship between SNHG7, miR-34a-5p and SYVN1. The MTT and flow cytometry assay was performed to detected cell proliferation and cell apoptosis respectively. RESULTS: In this study, SNHG7 and SYVN1 expression were down-regulated, but miR-34a-5p was up-regulated in OA tissues and IL-1P treated cells compared with normal tissues and chondrocyte. Functional investigation revealed that up-regulated SNHG7 or down-regulated miR-34a-5p could promote cell proliferation and inhibit cell apoptosis and autophagy in OA cells. More than that, RIP, pulldown and luciferase reporter assay was applied to determine that miR-34a-5p was a target miRNA of SNHG7 and SYVN1 was a target mRNA of miR-34-5p. Rescue experiments showed that overexpression of miR-34a reversed high expression of SNHG7-mediated suppression of apoptosis and autophagy as well as promotion of proliferation, while its knockdown inhibited cell apoptosis and autophagy and promoted cell proliferation which could be impaired by silencing SYVN1. In addition, SNHG7 regulated SYVN1 through sponging miR-34a-5p. CONCLUSION: SNHG7 sponged miR-34a-5p to affect cell proliferation, apoptosis and autophagy through targeting SYVN1 which provides a novel sight into the pathogenesis of OA.
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Humanos , Osteoartrite/metabolismo , Autofagia/fisiologia , Apoptose/fisiologia , MicroRNAs/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , RNA Longo não Codificante/metabolismo , Osteoartrite/genética , Autofagia/genética , Ensaio de Imunoadsorção Enzimática , Regulação para Baixo , Regulação para Cima , Western Blotting , Apoptose/genética , MicroRNAs/genética , Ubiquitina-Proteína Ligases/genética , Proliferação de Células , Reação em Cadeia da Polimerase em Tempo Real , RNA Longo não Codificante/genéticaRESUMO
BACKGROUND: Osteoarthritis (OA) is a common chronic degenerative disease, and the chondrocyte is reported to be a key player in OA progression. Increasing evidence has verified the regulatory role of miRNAs in OA. However, the function and underlying mechanism of miR-107 in cartilage function are still not clarified. METHODS: Abundance of miR-107 and PTEN mRNA were detected by qRT-PCR. Relative protein levels of PTEN, Bcl-2, Bax, Caspase-3, aggrecan, collagen II, MMP-13, and MMP-9 were measured by western blotting (WB). A biological web server Targetscan was used to predict the putative binding sites between miR-107 and PTEN, and luciferase reporter assay was employed to further verify the true interplay between them. Cell proliferative or apoptotic activity was assessed by MTT or flow cytometry (FCM) analysis. RESULTS: miR-107 was downregulated and PTEN was upregulated in OA tissues. PTEN could be negatively regulated by miR-107 by targeted interaction. Interference of PTEN induced proliferation of C28/I2 cells, but inhibited cell apoptosis. Restoration of PTEN reversed miR-107-stimulated cell proliferation and miR-107-inhibited apoptosis in C28/I2 cells. Furthermore, enforced abundance of miR-107 promoted the expressions of aggrecan and collagen II protein, while it attenuated MMP-13 and MMP-9 expression in C28/I2 cells, which was overturned by PTEN restoration. CONCLUSION: miR-107 induced chondrocyte growth and ameliorated cartilage degradation by targeting to PTEN, providing a potential therapeutic target for OA.
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OBJECTIVE: To improve the accuracy of preoperative evaluation of cervical lymph node metastasis in thoracic esophageal squamous carcinoma. METHODS: Forty-two patients with thoracic esophageal squamous carcinoma underwent neck ultrasonography. Enlarged lymph nodes with their long axis greater than 10 mm and a short-to-long axis ratio greater than 0.5 were considered as metastatic. RESULTS: Preoperative neck ultrasonography revealed the enlarged lymph nodes in 16 patients, but only in 5 (31%) cases the nodes were palpable. Among them 9 were classified as metastatic (cM(1-LN)), including 4 patients with palpable nodes. In 5 cM(1-LN) patients surgical intervention was canceled and the remaining 37 patients underwent trans-thoracic esophagectomy. Cervical node metastasis (pM(1-LN)) was confirmed pathologically in 6 surgical patients, 4 with tumors invading the adventitia (pT3) and the other 2 into the surrounding structure (pT(4)) (pT(1), pT(2) vs. pT(3), pT(4), P = 0.020). All 6 pM(1-LN) patients had concomitant mediastinal node metastasis and 4 of them had upper abdominal node metastasis. Statistically significant relationship was detected between cervical and abdominal nodal status (r = 0.536, P = 0.007). In comparison with the results of pathological examination and treatment response, the accuracy and sensitivity were 81% and 95% (P = 0.043), 36% and 82% (P = 0.081), respectively, for palpation and ultrasonography. Five out of 39 (13%) patients had their therapy changed due to ultrasonographic findings. CONCLUSIONS: Neck ultrasonography for cervical lymphadenopathy is of high sensitivity and accuracy, which plays an important role in the preoperative evaluation and therapeutic decision-making.