Design of Promising Thiazoloindazole-Based Acetylcholinesterase Inhibitors Guided by Molecular Docking and Experimental Insights.

Alzheimer's disease is characterized by a progressive deterioration of cognitive function and memory loss, and it is closely associated with the dysregulation of cholinergic neurotransmission. Since acetylcholinesterase (AChE) is a critical enzyme in the nervous system, responsible for breaking down the neurotransmitter acetylcholine, its inhibition holds a significant interest in the treatment of various neurological disorders. Therefore, it is crucial to develop efficient AChE inhibitors capable of increasing acetylcholine levels, ultimately leading to improved cholinergic neurotransmission. The results reported here represent a step forward in the development of novel thiazoloindazole-based compounds that have the potential to serve as effective AChE inhibitors. Molecular docking studies revealed that certain of the evaluated nitroindazole-based compounds outperformed donepezil, a well-known AChE inhibitor used in Alzheimer's disease treatment. Sustained by these findings, two series of compounds were synthesized. One series included a triazole moiety (Tl45a-c), while the other incorporated a carbazole moiety (Tl58a-c). These compounds were isolated in yields ranging from 66 to 87% through nucleophilic substitution and Cu(I)-catalyzed azide-alkyne 1,3-dipolar cycloaddition (CuAAC) reactions. Among the synthesized compounds, the thiazoloindazole-based 6b core derivatives emerged as selective AChE inhibitors, exhibiting remarkable IC50 values of less than 1.0 µM. Notably, derivative Tl45b displays superior performance as an AChE inhibitor, boasting the lowest IC50 (0.071 ± 0.014 µM). Structure-activity relationship (SAR) analysis indicated that derivatives containing the bis(trifluoromethyl)phenyl-triazolyl group demonstrated the most promising activity against AChE, when compared to more rigid substituents such as carbazolyl moiety. The combination of molecular docking and experimental synthesis provides a suitable and promising strategy for the development of new efficient thiazoloindazole-based AChE inhibitors.

OsTH1 is a key player in thiamin biosynthesis in rice.

Thiamin is a vital nutrient that acts as a cofactor for several enzymes primarily localized in the mitochondria. These thiamin-dependent enzymes are involved in energy metabolism, nucleic acid biosynthesis, and antioxidant machinery. The enzyme HMP-P kinase/thiamin monophosphate synthase (TH1) holds a key position in thiamin biosynthesis, being responsible for the phosphorylation of HMP-P into HMP-PP and for the condensation of HMP-PP and HET-P to form TMP. Through mathematical kinetic model, we have identified TH1 as a critical player for thiamin biofortification in rice. We further focused on the functional characterization of OsTH1. Sequence and gene expression analysis, along with phylogenetic studies, provided insights into OsTH1 bifunctional features and evolution. The indispensable role of OsTH1 in thiamin biosynthesis was validated by heterologous expression of OsTH1 and successful complementation of yeast knock-out mutants impaired in thiamin production. We also proved that the sole OsTH1 overexpression in rice callus significantly improves B1 concentration, resulting in 50% increase in thiamin accumulation. Our study underscores the critical role of OsTH1 in thiamin biosynthesis, shedding light on its bifunctional nature and evolutionary significance. The significant enhancement of thiamin accumulation in rice callus upon OsTH1 overexpression constitutes evidence of its potential application in biofortification strategies.

Impact of Fe3O4-porphyrin hybrid nanoparticles on wheat: Physiological and metabolic advance.

Iron-magnetic nanoparticles (Fe-NMPs) are widely used in environmental remediation, while porphyrin-based hybrid materials anchored to silica-coated Fe3O4-nanoparticles (Fe3O4-NPs) have been used for water disinfection purposes. To assess their safety on plants, especially concerning potential environmental release, it was investigated for the first time, the impact on plants of a silica-coated Fe3O4-NPs bearing a porphyrinic formulation (FORM) - FORM@NMP. Additionally, FORM alone and the magnetic nanoparticles without FORM anchored (NH2@NMP) were used for comparison. Wheat (Triticum aestivum L.) was chosen as a model species and was subjected to three environmentally relevant doses during germination and tiller development through root application. Morphological, physiological, and metabolic parameters were assessed. Despite a modest biomass decrease and alterations in membrane properties, no major impairments in germination or seedling development were observed. During tiller phase, both Fe3O4-NPs increased leaf length, and photosynthesis exhibited varied impacts: both Fe3O4-NPs and FORM alone increased pigments; only Fe3O4-NPs promoted gas exchange; all treatments improved the photochemical phase. Regarding oxidative stress, lipid peroxidation decreased in FORM and FORM@NMP, yet with increased O2-• in FORM@NMP; total flavonoids decreased in NH2@NMP and antioxidant enzymes declined across all materials. Phenolic profiling revealed a generalized trend towards a decrease in flavones. In conclusion, these nanoparticles can modulate wheat physiology/metabolism without apparently inducing phytotoxicity at low doses and during short-time exposure. ENVIRONMENTAL IMPLICATION: Iron-magnetic nanoparticles are widely used in environmental remediation and fertilization, besides of new applications continuously being developed, making them emerging contaminants. Soil is a major sink for these nanoparticles and their fate and potential environmental risks in ecosystems must be addressed to achieve more sustainable environmental applications. Furthermore, as the reuse of treated wastewater for agricultural irrigation is being claimed, it is of major importance to disclose the impact on crops of the nanoparticles used for wastewater decontamination, such as those proposed in this work.

Mathematical kinetic modelling followed by in vitro and in vivo assays reveal the bifunctional rice GTPCHII/DHBPS enzymes and demonstrate the key roles of OsRibA proteins in the vitamin B2 pathway.

BACKGROUND: Riboflavin is the precursor of several cofactors essential for normal physical and cognitive development, but only plants and some microorganisms can produce it. Humans thus rely on their dietary intake, which at a global level is mainly constituted by cereals (> 50%). Understanding the riboflavin biosynthesis players is key for advancing our knowledge on this essential pathway and can hold promise for biofortification strategies in major crop species. In some bacteria and in Arabidopsis, it is known that RibA1 is a bifunctional protein with distinct GTP cyclohydrolase II (GTPCHII) and 3,4-dihydroxy-2-butanone-4-phosphate synthase (DHBPS) domains. Arabidopsis harbors three RibA isoforms, but only one retained its bifunctionality. In rice, however, the identification and characterization of RibA has not yet been described. RESULTS: Through mathematical kinetic modeling, we identified RibA as the rate-limiting step of riboflavin pathway and by bioinformatic analysis we confirmed that rice RibA proteins carry both domains, DHBPS and GTPCHII. Phylogenetic analysis revealed that OsRibA isoforms 1 and 2 are similar to Arabidopsis bifunctional RibA1. Heterologous expression of OsRibA1 completely restored the growth of the rib3∆ yeast mutant, lacking DHBPS expression, while causing a 60% growth improvement of the rib1∆ mutant, lacking GTPCHII activity. Regarding OsRibA2, its heterologous expression fully complemented GTPCHII activity, and improved rib3∆ growth by 30%. In vitro activity assays confirmed that both OsRibA1 and OsRibA2 proteins carry GTPCHII/DHBPS activities, but that OsRibA1 has higher DHBPS activity. The overexpression of OsRibA1 in rice callus resulted in a 28% increase in riboflavin content. CONCLUSIONS: Our study elucidates the critical role of RibA in rice riboflavin biosynthesis pathway, establishing it as the rate-limiting step in the pathway. By identifying and characterizing OsRibA1 and OsRibA2, showcasing their GTPCHII and DHBPS activities, we have advanced the understanding of riboflavin biosynthesis in this staple crop. We further demonstrated that OsRibA1 overexpression in rice callus increases its riboflavin content, providing supporting information for bioengineering efforts.

Efficient Strategies to Use ß-Cationic Porphyrin-Imidazolium Derivatives in the Photoinactivation of Methicillin-Resistant Staphylococcus aureus.

Bacterial resistance to antibiotics is a critical global health issue and the development of alternatives to conventional antibiotics is of the upmost relevance. Antimicrobial photodynamic therapy (aPDT) is considered a promising and innovative approach for the photoinactivation of microorganisms, particularly in cases where traditional antibiotics may be less effective due to resistance or other limitations. In this study, two ß-modified monocharged porphyrin-imidazolium derivatives were efficiently incorporated into polyvinylpyrrolidone (PVP) formulations and supported into graphitic carbon nitride materials. Both porphyrin-imidazolium derivatives displayed remarkable photostability and the ability to generate cytotoxic singlet oxygen. These properties, which have an important impact on achieving an efficient photodynamic effect, were not compromised after incorporation/immobilization. The prepared PVP-porphyrin formulations and the graphitic carbon nitride-based materials displayed excellent performance as photosensitizers to photoinactivate methicillin-resistant Staphylococcus aureus (MRSA) (99.9999% of bacteria) throughout the antimicrobial photodynamic therapy. In each matrix, the most rapid action against S. aureus was observed when using PS 2. The PVP-2 formulation needed 10 min of exposure to white light at 5.0 µm, while the graphitic carbon nitride hybrid GCNM-2 required 20 min at 25.0 µm to achieve a similar level of response. These findings suggest the potential of graphitic carbon nitride-porphyrinic hybrids to be used in the environmental or clinical fields, avoiding the use of organic solvents, and might allow for their recovery after treatment, improving their applicability for bacteria photoinactivation.

Photomodulation Approaches to Overcome Antimicrobial Resistance.

Photopharmacology is an approach that aims to be an alternative to classical chemotherapy. Herein, the different classes of photoswitches and photocleavage compounds and their biological applications are described. Proteolysis targeting chimeras (PROTACs) containing azobenzene moieties (PHOTACs) and photocleavable protecting groups (photocaged PROTACs) are also mentioned. Furthermore, porphyrins are referenced as successful photoactive compounds in a clinical context, such as in the photodynamic therapy of tumours as well as preventing antimicrobial resistance, namely in bacteria. Porphyrins combining photoswitches and photocleavage systems are highlighted, taking advantage of both photopharmacology and photodynamic action. Finally, porphyrins with antibacterial activity are described, taking advantage of the synergistic effect of photodynamic treatment and antibiotic therapy to overcome bacterial resistance.

Sulfonamide Porphyrins as Potent Photosensitizers against Multidrug-Resistant Staphylococcus aureus (MRSA): The Role of Co-Adjuvants.

Sulfonamides are a conventional class of antibiotics that are well-suited to combat infections. However, their overuse leads to antimicrobial resistance. Porphyrins and analogs have demonstrated excellent photosensitizing properties and have been used as antimicrobial agents to photoinactivate microorganisms, including multiresistant Staphylococcus aureus (MRSA) strains. It is well recognized that the combination of different therapeutic agents might improve the biological outcome. In this present work, a novel meso-arylporphyrin and its Zn(II) complex functionalized with sulfonamide groups were synthesized and characterized and the antibacterial activity towards MRSA with and without the presence of the adjuvant KI was evaluated. For comparison, the studies were also extended to the corresponding sulfonated porphyrin TPP(SO3H)4. Photodynamic studies revealed that all porphyrin derivatives were effective in photoinactivating MRSA (>99.9% of reduction) at a concentration of 5.0 µM upon white light radiation with an irradiance of 25 mW cm-2 and a total light dose of 15 J cm-2. The combination of the porphyrin photosensitizers with the co-adjuvant KI during the photodynamic treatment proved to be very promising allowing a significant reduction in the treatment time and photosensitizer concentration by six times and at least five times, respectively. The combined effect observed for TPP(SO2NHEt)4 and ZnTPP(SO2NHEt)4 with KI seems to be due to the formation of reactive iodine radicals. In the photodynamic studies with TPP(SO3H)4 plus KI, the cooperative action was mainly due to the formation of free iodine (I2).

Porphyrin Photosensitizers Grafted in Cellulose Supports: A Review.

Cellulose is the most abundant natural biopolymer and owing to its compatibility with biological tissues, it is considered a versatile starting material for developing new and sustainable materials from renewable resources. With the advent of drug-resistance among pathogenic microorganisms, recent strategies have focused on the development of novel treatment options and alternative antimicrobial therapies, such as antimicrobial photodynamic therapy (aPDT). This approach encompasses the combination of photoactive dyes and harmless visible light, in the presence of dioxygen, to produce reactive oxygen species that can selectively kill microorganisms. Photosensitizers for aPDT can be adsorbed, entrapped, or linked to cellulose-like supports, providing an increase in the surface area, with improved mechanical strength, barrier, and antimicrobial properties, paving the way to new applications, such as wound disinfection, sterilization of medical materials and surfaces in different contexts (industrial, household and hospital), or prevention of microbial contamination in packaged food. This review will report the development of porphyrinic photosensitizers supported on cellulose/cellulose derivative materials to achieve effective photoinactivation. A brief overview of the efficiency of cellulose based photoactive dyes for cancer, using photodynamic therapy (PDT), will be also discussed. Particular attention will be devoted to the synthetic routes behind the preparation of the photosensitizer-cellulose functional materials.

Photodynamic inactivation of microorganisms in different water matrices: The effect of physicochemical parameters on the treatment outcome.

Wastewater (WW) insufficiently treated for the disinfection of microorganisms, including pathogenic ones, is a source of concern and a possible generator of public health problems. Traditional disinfection methods to reduce pathogens concentration (e.g., chlorination, ozonation, UV) are expensive, unsafe, and/or sometimes ineffective, highlighting the need for new disinfection technologies. The promising results of photodynamic inactivation (PDI) treatment to eradicate microorganisms suggest the efficacy of this treatment to improve WW quality. This work aimed to assess if PDI can be successfully extended to real contexts for the microbial inactivation in WW. For the first time, PDI experiments with 9 different water matrices compositions were performed to inquire about the influence of some of their physicochemical parameters on the effectiveness of microbial inactivation. Bacterial photoinactivation was tested in freshwater, aquaculture water, and seawater samples, as well as in influents and effluents samples from domestic, industrial, and a mixture of industrial and domestic WW receiving wastewater treatment plants (WWTPs). Additionally, PDI assays were performed in phosphate-buffered saline isotonic solution (PBS), used as an aqueous comparative matrix. To relate the PDI disinfection efficiency with the physicochemical compositions of the different used water matrices, a series of statistical analysis were performed, in order to support our main conclusions. Overall, the results showed that PDI is an effective and promising alternative to traditionally used WW disinfection methods, with a bacterial reduction of >3.0 log CFU/mL in all the water matrices within the first hour of PDI treatment, but also that the physicochemical composition of the aqueous matrices to be PDI-disinfected must be taken into account since they seem to influence the PDI efficacy, namely the pH, with acidic pH conditions seeming to be associated to a better PDI performance in general.

Anti-Viral Photodynamic Inactivation of T4-like Bacteriophage as a Mammalian Virus Model in Blood.

The laboratorial available methods applied in plasma disinfection can induce damage in other blood components. Antimicrobial photodynamic therapy (aPDT) represents a promising approach and is approved for plasma and platelet disinfection using non-porphyrinic photosensitizers (PSs), such as methylene blue (MB). In this study, the photodynamic action of three cationic porphyrins (Tri-Py(+)-Me, Tetra-Py(+)-Me and Tetra-S-Py(+)-Me) towards viruses was evaluated under white light irradiation at an irradiance of 25 and 150 mW·cm-2, and the results were compared with the efficacy of the approved MB. None of the PSs caused hemolysis at the isotonic conditions, using a T4-like phage as a model of mammalian viruses. All porphyrins were more effective than MB in the photoinactivation of the T4-like phage in plasma. Moreover, the most efficient PS promoted a moderate inactivation rate of the T4-like phage in whole blood. Nevertheless, these porphyrins, such as MB, can be considered promising and safe PSs to photoinactivate viruses in blood plasma.

New Bis-Cyclometalated Iridium(III) Complexes with ß-Substituted Porphyrin-Arylbipyridine as the Ancillary Ligand: Electrochemical and Photophysical Insights.

An efficient synthetic access to new cationic porphyrin-bipyridine iridium(III) bis-cyclometalated complexes was developed. These porphyrins bearing arylbipyridine moieties at ß-pyrrolic positions coordinated with iridium(III), and the corresponding Zn(II) porphyrin complexes were spectroscopically, electrochemically, and electronically characterized. The features displayed by the new cyclometalated porphyrin-bipyridine iridium(III) complexes, namely photoinduced electron transfer process (PET), and a remarkable efficiency to generate 1O2, allowing us to envisage new challenges and opportunities for their applications in several fields, such as photo(catalysis) and photodynamic therapies.

Can Corrole Dimers Be Good Photosensitizers to Kill Bacteria?

Corroles possess key photophysical and photochemical properties to be exploited as therapeutic agents in antimicrobial photodynamic therapy (aPDT). Herein, we present for the first time the antimicrobial efficiency of three corrole dimers and of the corresponding precursor against the Gram(+) bacterium Staphylococcus aureus. Additionally, to explore future clinical applications, the cytotoxicity of the most promising derivatives towards Vero cells was evaluated. The aPDT assays performed under white light irradiation (50 mW/cm2; light dose 450 J/cm2) and at a corrole concentration of 15 µM showed that some dimers were able to reduce 99.9999% of S. aureus strain (decrease of 5 log10 CFU/mL) and their photodynamic efficiency was dependent on position, type of linkage, and aggregation behavior. Under the same light conditions, the corrole precursor 1 demonstrated notable photodynamic efficiency, achieving total photoinactivation (>8.0 log10 CFU/mL reduction) after the same period of irradiation (light dose 450 J/cm2). No cytotoxicity was observed when Vero cells were exposed to corrole 1 and dimer 3 for 24 h according to ISO guidelines (ISO 10993-5) for in vitro cytotoxicity of medical devices. The results show that corrole dimers, dependent on their structures, can be considered good photosensitizers to kill Staphylococcus aureus.

Photodynamic inactivation of pathogenic Gram-negative and Gram-positive bacteria mediated by Si(IV) phthalocyanines bearing axial ammonium units.

The photodynamic inactivation (PDI) of microorganisms has gained interest as an efficient option for conventional antibiotic treatments. Recently, Si(IV) phthalocyanines (SiPcs) have been highlighted as promising photosensitizers (PSs) to the PDI of microorganisms due to their remarkable absorption and emission features. To increase the potential of cationic SiPcs as PS drugs, one novel (1a) and two previously described (2a and 3a) axially substituted PSs with di-, tetra-, and hexa-ammonium units, respectively, were synthesized and characterized. Their PDI effect was evaluated for the first time against Escherichia coli and Staphylococcus aureus, a Gram-negative and a Gram-positive bacterium, respectively. The photodynamic treatments were conducted with PS concentrations of 3.0 and 6.0 µM under 60 min of white light irradiation (150 mW.cm-2). The biological results show high photodynamic efficiency for di- and tetra-cationic PSs 1a and 2a (6.0 µM), reducing the E. coli viability in 5.2 and 3.9 log, respectively (after 15 min of dark incubation before irradiation). For PS 3a, a similar bacterial reduction (3.6 log) was achieved but only with an extended dark incubation period (30 min). Under the same experimental conditions, the photodynamic effect of cationic PSs 1a-3a on S. aureus was even more promising, with abundance reductions of ca. 8.0 log after 45-60 min of PDI treatment. These results reveal the high PDI efficiency of PSs bearing ammonium groups and suggest their promising application as a broad-spectrum antimicrobial to control infections caused by Gram-negative and Gram-positive bacteria.

Bioluminescent Models to Evaluate the Efficiency of Light-Based Antibacterial Approaches.

The emergence of microbial resistance to antimicrobials among several common pathogenic microbial strains is an increasing problem worldwide. Thus, it is urgent to develop not only new antimicrobial therapeutics to fight microbial infections, but also new effective, rapid, and inexpensive methods to monitor the efficacy of these new therapeutics. Antimicrobial photodynamic therapy (aPDT) and antimicrobial blue light (aBL) therapy are receiving considerable attention for their antimicrobial potential and represent realistic alternatives to antibiotics. To monitor the photoinactivation process provided by aPDT and aBL, faster and more effective methods are required instead of laborious conventional plating and overnight incubation procedures. Bioluminescent microbial models are very interesting in this context. Light emission from bioluminescent microorganisms is a highly sensitive indication of their metabolic activity and can be used to monitor, in real time, the effects of antimicrobial agents and therapeutics. This chapter reviews the efforts of the scientific community concerning the development of in vitro, ex vivo, and in vivo bioluminescent bacterial models and their potential to evaluate the efficiency of aPDT and aBL in the inactivation of bacteria.

The Antimicrobial Photoinactivation Effect on Escherichia coli through the Action of Inverted Cationic Porphyrin-Cyclodextrin Conjugates.

Photodynamic action has been used for diverse biomedical applications, such as treating a broad range of bacterial infections. Based on the combination of light, dioxygen, and photosensitizer (PS), the photodynamic inactivation (PDI) approach led to the formation of reactive oxygen species (ROS) and represented a non-invasive, non-toxic, repeatable procedure for pathogen photoinactivation. To this end, different tetrapyrrolic macrocycles, such as porphyrin (Por) dyes, have been used as PSs for PDI against microorganisms, mainly bacteria. Still, there is significant room for improvement, especially new PS molecules. Herein, unsymmetrical new pyridinone (3−5) and thiopyridyl Pors (7) were prepared with α-, ß-, or γ-cyclodextrin (CD) units, following their quaternization to perform the corresponding free-base Pors (3a−5a and 7a), and were compared with the already-known Pors 6a and 8a, both bearing thiopyridinium and CD units. These water-soluble porphyrins were evaluated as PSs, and their photophysical and photochemical properties and photodynamic effects on E. coli were assessed. The presence of one CD unit and three positive charges on the Por structure (3a−5a and 7a) enhanced their aqueous solubility. The photoactivity of the cationic Pors 3a−5a and 6a−8a ensured their potential against the Gram-negative bacterium E. coli. Within each series of methoxypyridinium vs thiopyridinium dyes, the best PDI efficiency was achieved for 5a with a bacterial viability reduction of 3.5 log10 (50 mW cm−2, 60 min of light irradiation) and for 8a with a total bacterial viability reduction (>8 log10, 25 mW cm−2, 30 min of light irradiation). Here, the presence of the methoxypyridinium units is less effective against E. coli when compared with the thiopyridinium moieties. This study allows for the conclusion that the peripheral charge position, quaternized substituent type/CD unit, and affinity to the outer bacterial structures play an important role in the photoinactivation efficiency of E. coli, evidencing that these features should be further addressed in the pursuit for optimised PS for the antimicrobial PDI of pathogenic microorganisms.

Use of the FLOTAC technique as a new coproparasitological diagnostic method in aquatic mammals and comparison with traditional methods.

The inadequate choice of a diagnostic method or the option for techniques that have low sensitivity and specificity may limit the diagnosis of parasitic agents that affect aquatic mammals. The aim of this study was to evaluate the performance of the FLOTAC technique and compare it with three traditional methods (Willis, sedimentation and centrifugation- flotation) used in the diagnosis of gastrointestinal parasites in aquatic mammals. For this, 129 fecal samples from 12 species were collected. Each sample was submitted to laboratory processing using the Willis, Hoffman techniques, Faust method and FLOTAC. Sensitivity, specificity, real prevalence, estimated prevalence, positive predictive value, negative predictive value, correct classification (accuracy) and incorrect classification were evaluated to compare the different diagnostic methods. The highest frequency of positive samples occurred using FLOTAC (46.51%), compared to Hoffman (23.25%), Faust (10.07%) and Willis techniques (6.97%). In the samples analyzed, the occurrence of Strongylidae eggs and Eimeriidae oocysts was frequently observed. The FLOTAC technique proved to be the most appropriate technique and due to its efficacy, is strongly recommended for coproparasitological evaluations in aquatic mammals.

Photoinactivation of Phage Phi6 as a SARS-CoV-2 Model in Wastewater: Evidence of Efficacy and Safety.

The last two years have been marked by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. This virus is found in the intestinal tract; it reaches wastewater systems and, consequently, the natural receiving water bodies. As such, inefficiently treated wastewater (WW) can be a means of contamination. The currently used methods for the disinfection of WW can lead to the formation of toxic compounds and can be expensive or inefficient. As such, new and alternative approaches must be considered, namely, photodynamic inactivation (PDI). In this work, the bacteriophage φ6 (or, simply, phage φ6), which has been used as a suitable model for enveloped RNA viruses, such as coronaviruses (CoVs), was used as a model of SARS-CoV-2. Firstly, to understand the virus's survival in the environment, phage φ6 was subjected to different laboratory-controlled environmental conditions (temperature, pH, salinity, and solar and UV-B irradiation), and its persistence over time was assessed. Second, to assess the efficiency of PDI towards the virus, assays were performed in both phosphate-buffered saline (PBS), a commonly used aqueous matrix, and a secondarily treated WW (a real WW matrix). Third, as WW is generally discharged into the marine environment after treatment, the safety of PDI-treated WW was assessed through the determination of the viability of native marine water microorganisms after their contact with the PDI-treated effluent. Overall, the results showed that, when used as a surrogate for SARS-CoV-2, phage φ6 remains viable in different environmental conditions for a considerable period. Moreover, PDI proved to be an efficient approach in the inactivation of the viruses, and the PDI-treated effluent showed no toxicity to native aquatic microorganisms under realistic dilution conditions, thus endorsing PDI as an efficient and safe tertiary WW disinfection method. Although all studies were performed with phage φ6, which is considered a suitable model of SARS-CoV-2, further studies using SARS-CoV-2 are necessary; nevertheless, the findings show the potential of PDI for controlling SARS-CoV-2 in WW.

DNA-Based Tools to Certify Authenticity of Rice Varieties-An Overview.

Rice (Oryza sativa L.) is one of the most cultivated and consumed crops worldwide. It is mainly produced in Asia but, due to its large genetic pool, it has expanded to several ecosystems, latitudes and climatic conditions. Europe is a rice producing region, especially in the Mediterranean countries, that grow mostly typical japonica varieties. The European consumer interest in rice has increased over the last decades towards more exotic types, often more expensive (e.g., aromatic rice) and Europe is a net importer of this commodity. This has increased food fraud opportunities in the rice supply chain, which may deliver mixtures with lower quality rice, a problem that is now global. The development of tools to clearly identify undesirable mixtures thus became urgent. Among the various tools available, DNA-based markers are considered particularly reliable and stable for discrimination of rice varieties. This review covers aspects ranging from rice diversity and fraud issues to the DNA-based methods used to distinguish varieties and detect unwanted mixtures. Although not exhaustive, the review covers the diversity of strategies and ongoing improvements already tested, highlighting important advantages and disadvantages in terms of costs, reliability, labor-effort and potential scalability for routine fraud detection.

One Year of COVID-19 in Pregnancy: A National Wide Collaborative Study.

INTRODUCTION: Even though the risk of COVID-19 in pregnancy may be increased, large-scale studies are needed to better understand the impact of the infection in this population. The aim of this study is to describe obstetric complications and the rate of vertical transmission in pregnant women with SARS-CoV-2 infection. MATERIAL AND METHODS: Detected cases of SARS-CoV-2 infection in pregnancy were registered in Portuguese hospitals by obstetricians. Epidemiological, pregnancy and childbirth data were collected. RESULTS: There were 630 positive cases in 23 Portuguese maternity hospitals, most at term (87.9%) and asymptomatic (62.9%). The most frequent maternal comorbidity was obesity. The rates of preterm birth and small-to-gestational-age were 12.1% and 9.9%, respectively. In the third trimester, 2.9% of pregnant women required respiratory support. There were eight cases (1.5%) of fetal death, including two cases of vertical transmission. There were five cases of postpartum respiratory degradation, but no maternal deaths were recorded. The caesarean section rate was higher in the first than in the second wave (68.5% vs 31.5%). RT-PCR SARS-CoV-2 positivity among newborns was 1.3%. CONCLUSION: SARS-Cov-2 infection in pregnancy may carry increased risks for both pregnant women and the fetuses. Individualized surveillance and the prophylaxis of this population with vaccination. is recommended in these cases.

Introdução: Apesar do risco da COVID-19 na gravidez poder ser acrescido, são necessários estudos em larga escala para o melhor conhecimento do impacto desta infeção nesta população. O objetivo deste estudo é descrever as complicações obstétricas e a taxa de transmissão vertical em grávidas com infeção a SARS-CoV-2. Material e Métodos: Os casos conhecidos de infeção por SARS-CoV-2 na gravidez foram registados nos hospitais portugueses por obstetras. Foram recolhidos dados epidemiológicos, da gravidez e do parto. Resultados: Registaram-se 630 casos positivos em 23 maternidades portuguesas, a maioria no termo (87,9%) e assintomática (62,9%). A comorbilidade materna mais frequente foi a obesidade. A taxa de parto pré-termo e de leves para a idade gestacional foi de 12,1% e 9,9%, respectivamente. No terceiro trimestre, 2,9% das grávidas necessitaram de suporte respiratório. Verificou-se uma taxa de 1,5% de morte fetal, incluindo dois casos de transmissão vertical. Houve cinco casos de degradação respiratória no pós-parto, mas sem mortes maternas registadas. A taxa de cesarianas foi mais elevada na primeira do que na segunda vaga (68,5% vs 31,5%). A positividade do RT-PCR SARS-CoV-2 entre os recém-nascidos foi de 1,3%. Conclusão: A infeção pelo SARS-Cov-2 na gravidez pode acarretar riscos aumentados para as grávidas e fetos. Recomenda-se uma vigilância individualizada nestes casos e a profilaxia desta população com a vacinação.

A novel panel of yeast assays for the assessment of thiamin and its biosynthetic intermediates in plant tissues.

Thiamin (or thiamine), known as vitamin B1, represents an indispensable component of human diets, being pivotal in energy metabolism. Thiamin research depends on adequate vitamin quantification in plant tissues. A recently developed quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is able to assess the level of thiamin, its phosphorylated entities and its biosynthetic intermediates in the model plant Arabidopsis thaliana, as well as in rice. However, their implementation requires expensive equipment and substantial technical expertise. Microbiological assays can be useful in deter-mining metabolite levels in plant material and provide an affordable alternative to MS-based analysis. Here, we evaluate, by comparison to the LC-MS/MS reference method, the potential of a carefully chosen panel of yeast assays to estimate levels of total vitamin B1, as well as its biosynthetic intermediates pyrimidine and thiazole in Arabidopsis samples. The examined panel of Saccharomyces cerevisiae mutants was, when implemented in microbiological assays, capable of correctly assigning a series of wild-type and thiamin biofortified Arabidopsis plant samples. The assays provide a readily applicable method allowing rapid screening of vitamin B1 (and its biosynthetic intermediates) content in plant material, which is particularly useful in metabolic engineering approaches and in germplasm screening across or within species.