Changes in biomarkers of the redox status in whole blood and red blood cell lysates in canine hypothyroidism.

Hypothyroidism is the most commonly diagnosed endocrine disease in dogs. The objective of this study was to evaluate the changes in the redox status in canine hypothyroidism using whole blood (WB) and red blood cell (RBCs) lysates. For this purpose, a panel of five antioxidants and five oxidants biomarkers was measured in WB and RBCs lysates of 30 dogs with hypothyroidism, 26 dogs with non-thyroidal illnesses and 15 healthy dogs. The antioxidants measured were cupric reducing antioxidant capacity (CUPRAC), ferric reducing ability of plasma (FRAP), Trolox equivalent antioxidant capacity (TEAC), thiol and paraoxonase type-1 (PON-1). Oxidants measured include the total oxidant status (TOS), peroxide-activity (POX-Act), reactive oxygen-derived metabolites (d-ROMs), advanced oxidation protein products (AOPP) and thiobarbituric acid reactive substances (TBARS). WB showed a significant decrease of the antioxidants CUPRAC, TEAC and thiol, and also an increase in TBARS and a decrease in AOPP in dogs with hypothyroidism compared to healthy dogs. Meanwhile, RBCs lysates showed a significant increase in FRAP and PON-1 in dogs with hypothyroidism. The changes in the redox biomarkers in this study show that WB in canine hypothyroidism had a higher number of changes in biomarkers of the redox status than RBCs lysates, making it a promising sample type for the evaluation of the redox status in this disease. In addition, WB is easier and simpler to process than RBCs lysates and unlike serum, it does not have any hemolysis interference.

Renin-angiotensin-aldosterone system blockers effect in chronic kidney disease progression in hypertensive elderly patients without proteinuria: PROERCAN trial.

INTRODUCTION: Evidence about nefroprotective effect with RAAS blockers in elderly patients with chronic kidney disease (CKD) without proteinuria is lacking. The primary outcome of our study is to evaluate the impact of RAAS blockers in CKD progression in elderly patients without proteinuria. MATERIALS AND METHODS: Multicenter open-label, randomized controlled clinical trial including patients over 65 year-old with hypertension and CKD stages 3-4 without proteinuria. Patients were randomized in a 1:1 ratio to either receive RAAS blockers or other antihypertensive drugs and were followed up for three years. Primary outcome is estimated glomerular filtration rate (eGFR) decline at 3 years. Secondary outcome measures include BP control, renal and cardiovascular events and mortality. RESULTS: 88 patients were included with a mean age of 77.9±6.1 years and a follow up period of 3 years: 40 were randomized to RAAS group and 48 to standard treatment. Ethiology of CKD was: 53 vascular, 16 interstitial and 19 of unknown ethiology. In the RAAS group eGFR slope during follow up was -4.3±1.1ml/min, whereas in the standard treatment group an increase on eGFR was observed after 3 years (+4.6±0.4ml/min), p=0.024. We found no differences in blood pressure control, number of antihypertensive drugs, albuminuria, potassium serum levels, incidence of cardiovascular events nor mortality during the follow up period. CONCLUSIONS: In elderly patients without diabetes nor cardiopathy and with non proteinuric CKD the use of RAAS blockers does not show a reduction in CKD progression. The PROERCAN (PROgresión de Enfermedad Renal Crónica en ANcianos) trial (trial registration: NCT03195023).

Antimicrobials in Eisenia fetida earthworms: A comprehensive study from method development to the assessment of uptake and degradation.

In this work, an accurate analytical method was developed for the simultaneous analysis of twenty-seven antimicrobials (AMs) in earthworms using liquid chromatography coupled to a triple quadrupole mass spectrometry detector (UHPLC-MS/MS). Adequate apparent recoveries (80-120 %) and limits of quantification (LOQ) (1 µg·kg-1 - 10 µg·kg-1) were obtained, with the exception of norfloxacin (34 µg·kg-1). The method was applied to evaluate the accumulation of sulfamethazine (SMZ) and tetracycline (TC) in earthworms after performing OECD-207 toxicity test, in which Eisenia fetida (E. fetida) organisms were exposed to soils spiked with 10 mg·kg-1, 100 mg·kg-1 or 1000 mg·kg-1 of SMZ and TC, individually. The results confirmed the bioaccumulation of both AMs in the organisms, showing a greater tendency to accumulate SMZ since higher bioconcentration factor values were obtained for this compound at the exposure concentrations tested. In addition, the degradation of both AMs in both matrices, soils and earthworms was studied using liquid chromatography coupled to a q-Orbitrap high resolution mass spectrometry detector. Thirteen transformation products (TPs) were successfully identified, eight of them being identified for the first time in soil/earthworm (such as 4-Amino-3-chloro-n-(4,6-dimethylpyrimidin-2-yl)benzenesulfonamide or 4-(dimethylamino)-1,11,12a-trihydroxy-6,6-dimethyl-3,7,10,12-tetraoxo-3,4,4a,5,5a,6,7,10,12,12a-decahydrotetracene-2-carboxamide, among others) and their formation/degradation trend over time was also studied. Regarding the biological effects, only SMZ caused changes in earthworm growth, evidenced by weight loss in earthworms exposed to concentrations of 100 mg·kg-1 and 1000 mg·kg-1. Riboflavin decreased at all concentrations of SMZ, as well as at the highest concentration of TC. This indicates that these antibiotics can potentially alter the immune system of E. fetida. This research represents a significant advance in improving our knowledge about the contamination of soil by AM over time. It investigates the various ways in which earthworms are exposed to AMs, either by skin contact or ingestion. Furthermore, it explores how these substances accumulate in earthworms, the processes by which earthworms break them down or metabolise them, as well as the resulting TPs. Finally, it examines the potential effects of these substances on the environment.

Multitarget and suspect-screening of antimicrobials in vegetables samples: Uptake experiments and identification of transformation products.

This work provided an accurate analytical method to perform a multitarget analysis of a variety of antimicrobials (AMs) including sulfonamides, tetracyclines, macrolides, fluoroquinolones and quinolones, one imidazole and one nitroimidazole, one triazole, one diaminopyridine and one derivative of Penicillium stoloniferum in vegetables. The analysis is performed using liquid-chromatography coupled to a low-resolution triple quadrupole mass spectrometer (UHPLC-MS/MS) to detect the target analytesor coupled to a high-resolution q-Orbitrap (HRMS) to monitor the formed transformation products (TPs). Both instruments were compared in terms of limits of quantification and matrix effect at the detection. The method was applied to determine the presence of AMs in organic and non-organic vegetables, where sulfadiazine and mycophenolic acid were detected. On the other hand, the transference of four AMs (trimethoprim, sulfamethazine, enrofloxacin, and chlortetracycline) from soils to lettuces was evaluated through controlled uptake experiments. The choice of AMs was based on the classification into different families, and on the fact that those AM families are the most frequently detected in the environment. In this case, each of the AMs with which the soils were contaminated were found in the exposed lettuces. Moreover, in both studies, specific TPs of the AMs were identified, posing the necessity of assessing their effects in relation to food and human safety.

Sample preparation for suspect and non-target screening of xenobiotics in human biofluids by liquid chromatography-High resolution tandem mass spectrometry.

The concept of exposome covers all the exposures an individual suffers from conception to death, which can be partially assessed through the monitoring of human biofluids. In there, target analytical approaches tend to focus on a limited set of xenobiotics, whereas exposomic studies need broad scopes in search of a full understanding. Given the issue, suspect and non-target screening are feasible alternatives. However, adequate sample preparation procedures should minimize interferences without significantly reducing the number of xenobiotics. Within this context, the present article aims to describe comprehensive sample preparation procedures for suspect or non-target screening of organic xenobiotics in several human biofluids, all coupled to unified separation and detection conditions based on ultra-high performance liquid chromatography-high resolution tandem mass spectrometry (UHPLC-HRMS/MS). The referred biofluids consist of human urine, breast milk, saliva and ovarian follicular fluid.•Analytical methods for untargeted analysis of a wide range of xenobiotics in human biofluids are fully described in order to ensure reproducibility.•The sample preparation procedures balance selectivity and sensitivity.•Unified analysis conditions allow simultaneous analysis of diverse biofluids.

Evaluating membrane bioreactor treatment for the elimination of emerging contaminants using different analytical methods.

Since wastewater treatment plants (WWTPs) were not originally designed to eliminate contaminants of emerging concern (CECs), alternative strategies like membrane bioreactor (MBR) technology are gaining importance in achieving effective CEC removal and minimising their environmental impact. In this study, composite wastewater samples were collected from the biggest WWTP in the Basque Country (Galindo, Biscay) and the performance of two secondary treatments (i.e. conventional activated sludge treatment, CAS, and MBR) was assessed. The combination of a suspect screening approach using liquid chromatography tandem high-resolution mass spectrometry (LC-HRMS) and multitarget analysis by gas chromatography-mass spectrometry (GC-MS) allowed the detection of approximately 200 compounds in the WWTP effluents. The estimated removal efficiencies (REs) revealed that only 16 micropollutants exhibited enhanced removal by MBR treatment (RE > 70% or 40 - 60%). The environmental risk posed by the non-eliminated compounds after both treatments remained similar, being anthracene, clarithromycin, bis(2-ethylhexyl) phthalate (DEHP) and dilantin the most concerning pollutants (RQ > 1). The Microtox® bioassay confirmed the MBR's efficiency in removing baseline toxicity, while suggesting a similar performance of CAS treatment. These minimal differences between treatments call into question the worthiness of MBR treatment and emphasise the need to seek more efficient alternative treatment methods.

[Neonatal seizures and progression to epilepsy in a tertiary hospital]. / Crisis neonatales y evolución a epilepsia en un hospital de tercer nivel.

INTRODUCTION: Given the immaturity of the newborn, neonatal seizures are a diagnostic challenge. Most of them are secondary to an acute event. A small percentage constitute the onset of epilepsy. AIMS: The aim was to analyse neonates with a diagnosis of seizures admitted to a tertiary hospital between November 2009 and May 2021, and their subsequent progression to epilepsy. MATERIAL AND METHODS: A retrospective observational study was carried out using the hospital database. Information was collected on neonates with a discharge diagnosis of 'seizures' or 'moderate or severe hypoxic-ischaemic encephalopathy'. Different variables were analysed: aetiology of the seizures, type, persistence over time, treatment and electroclinical correlates. RESULTS: Of 165 patients, 55 presented neonatal seizures. As regards aetiology, 43 patients (78%) had seizures secondary to an acute event, of which 19 (34%) were hypoxic-ischaemic encephalopathies, and 22 (40%) had other acute disorders. Genetic alteration was found in six of them (11%). Thirteen patients (24%) progressed to subsequent epilepsy, of whom seven had symptomatic epilepsy, with a period of latency after the acute event in two patients. Six patients had neonatal epilepsy with unprovoked seizures. Twenty-two (62%) showed electroclinical correlates. All of the confirmed crises (100%) were focal. All the seizures were treated. The drug of choice was phenobarbital. CONCLUSIONS: Diagnosis of neonatal seizures requires high clinical suspicion and electroclinical confirmation. Most of them progress favourably, but a percentage constitute the onset of epilepsy, the identification of which will determine their therapeutic management.

TITLE: Crisis neonatales y evolución a epilepsia en un hospital de tercer nivel.Introducción. Las convulsiones neonatales son un reto diagnóstico, dada la inmadurez del recién nacido. La mayoría son secundarias a un evento agudo. Un pequeño porcentaje constituye el inicio de una epilepsia. Objetivos. Analizar a los neonatos ingresados en un hospital de tercer nivel con diagnóstico de crisis entre noviembre de 2009 y mayo de 2021, y su evolución posterior a epilepsia. Material y métodos. Se ha realizado un estudio observacional retrospectivo utilizando la base de datos del hospital. Se ha recogido la información de los neonatos con diagnóstico en el alta de 'convulsiones' o 'encefalopatía hipóxico-isquémica moderada o grave'. Se analizaron distintas variables: etiología de las crisis, tipo, persistencia temporal, tratamiento y correlato electroclínico. Resultados. De 165 pacientes, 55 presentaron crisis neonatales. En cuanto a la etiología, 43 pacientes (78%) tuvieron crisis secundarias a un evento agudo, de las cuales 19 (34%) fueron encefalopatías hipóxico-isquémicas, y 22 (40%), otras alteraciones agudas. En seis (11%) se encontró alteración genética. Trece pacientes (24%) evolucionaron a una epilepsia posterior, de los cuales siete presentaron una epilepsia sintomática, con un período de latencia tras el evento agudo en dos pacientes. Seis pacientes tuvieron epilepsia neonatal con crisis no provocadas. Veintidós (62%) mostraron correlato electroclínico. El 100% de las crisis confirmadas fueron focales. Todas las crisis se trataron. El fármaco de elección fue el fenobarbital. Conclusiones. El diagnóstico de convulsiones neonatales requiere una alta sospecha clínica y una confirmación electroclínica. La mayoría tiene evolución favorable, pero un porcentaje constituye el inicio de una epilepsia, cuya identificación determinará su manejo terapéutico.

Multitarget and suspect screening of antimicrobials in soil and manure by means of QuEChERS - liquid chromatography tandem mass spectrometry.

The present work aimed to develop an accurate analytical method for the simultaneous analysis of twenty-four antimicrobials in soil:compost and animal manure samples by means of ultra-high performance liquid chromatography coupled to a triple-quadrupole mass spectrometer (UHPLC-QqQ). For this purpose, the effectiveness of two extraction techniques (i.e. focused ultrasound solid-liquid extraction (FUSLE) and QuEChERS (quick, easy, cheap, effective, rugged and safe)) was evaluated, and the clean-up step using solid-phase extraction (SPE) was also thoroughly studied. The method was successfully validated at 10 µg·kg-1, 25 µg·kg-1, and 50 µg·kg-1 showing adequate trueness (70-130%) and repeatability (RSD < 30%), with few exceptions. Procedural limits of quantification (LOQPRO) were determined for soil:compost (0.45 to 7.50 µg·kg-1) and manure (0.31 to 5.53 µg·kg-1) samples. Pefloxacin could not be validated at the lowest level since LOQPRO ≥ 10 µg·kg-1. Sulfamethazine (7.9 ± 0.8 µg·kg-1), danofloxacin (27.1 ± 1.4 µg·kg-1) and trimethoprim (4.9 ± 0.5 µg·kg-1) were detected in soil samples; and tetracycline (56.8 ± 2.8 µg·kg-1), among other antimicrobials, in the plants grown on the surface of the studied soil samples. Similarly, sulfonamides (SAs), tetracyclines (TCs) and fluoroquinolones (FQs) were detected in sheep manure in a range of 1.7 ± 0.3 to 93.3 ± 6.8 µg·kg-1. Soil and manure samples were also analysed through UHPLC coupled to a high-resolution mass-spectrometer (UHPLC-qOrbitrap) in order to extend the multitarget method to suspect screening of more than 22,281 suspects. A specific transformation product (TP) of sulfamethazine (formyl-sulfamethazine) was annotated at 2a level in manure samples, among others. This work contributes to the efforts that have been made in the last decade to develop analytical methods that allow multitarget analysis of a wide variety of antimicrobials, including TPs, which is a complex task due to the diverse physicochemical properties of the antimicrobials.

A novel time-saving multiplex PCR assay for detecting and discriminating the most common canine Babesia species in Europe.

In Europe, most cases of canine babesiosis are caused by Babesia canis, Babesia vogeli (large piroplasms) and Babesia vulpes (small piroplasm). Molecular diagnosis is recommended due to its high sensitivity. Species identification after sequencing allows applying a rapid and efficient treatment, leading to a better prognosis; however, it is expensive and time-consuming. Thus, the objective of the present study was to develop a time-saving multiplex polymerase chain reaction (PCR) for simultaneously detecting and discriminating between large and small forms without sequence analysis. A new multiplex PCR was designed and tested using blood samples from 79 dogs showing clinical signs compatible with babesiosis which were previously analysed using blood smears and molecular methods. Multiplex PCR successfully discriminated between both Babesia groups showing bands of 700 and 890 bp for B. canis/B. vogeli and B. vulpes, respectively. No significant differences in the results of both PCR were detected and a substantial agreement between protocols (κ = 0.64) was found. Our multiplex PCR represents a reliable tool for detecting infections by the major Babesia spp. in dogs from Europe. Since no sequence analysis is required for identifying the species involved, this PCR allows the rapid administration of an appropriate treatment, thus improving the survival rate of the infected animals. In addition, it will represent a helpful tool for unravelling the real prevalence and distribution of B. vulpes and its implication in clinical cases.

Thrombectomy with embed aspiration in acute ischaemic stroke.

BACKGROUND: In addition to stent retrievers, direct aspiration has become a reasonable thrombectomy strategy. OBJECTIVES: We carried out the thrombectomy by guiding the aspiration catheter fully over the clot and performing immediate manual aspiration; we call this procedure "embed aspiration". METHODS: In this prospective, non-randomised, single-centre study, we included all patients treated at a high volume-of-care stroke centre between 2017 and 2018 for the TRIANA (Thrombectomy in Andalusia using Aspiration) registry. Thrombectomy was carried out by embed aspiration. Patients were classified according to the success (eTICI 2b67-2c-3) or failure (eTICI 0-1-2a-2b50) of the procedure. Baseline clinical data and outcomes were compared, and multivariate analysis was performed. RESULTS: The embed aspiration technique was used in 370 patients. Treatment was successful in 90.3% of patients. Mean puncture-to-recanalisation time was 25 minutes. The overall rate of good outcomes (mRS 0-2) at 3 months was 64%. CONCLUSIONS: This study supports real-life evidence that standardised embed aspiration may be an alternative to stent retrievers for thrombectomy.

Tick-borne pathogens in ticks from urban and suburban areas of north-western Spain: Importance of Ixodes frontalis harbouring zoonotic pathogens.

To identify the questing tick populations in urban and suburban areas from the city of Lugo (NW Spain), ticks were collected monthly by flagging. The presence of Borrelia spp., Rickettsia spp. and Anaplasma phagocytophilum also was determined by polymerase chain reaction (PCR) and sequence analysis. Overall, 342 questing ticks were collected; the tick abundance was higher in suburban (95.9%) than in urban areas (4.1%). Ixodes frontalis was the most abundant (86.5%); 88.5% were larvae, 11.1% nymphs and 0.3% adults. All development stages of I. ricinus (7.3%) and adults of Rhipicephalus sanguineus sensu lato (5.8%) and Dermacentor reticulatus (0.3%) were found. Rickettsia spp. (31.9%) was more prevalent than Borrelia spp. (2.7%); no ticks were positive to A. phagocytophilum. Six Rickettsia species were identified (R. slovaca, R. monacensis, R. massiliae, R. raoultii, R. sibirica subsp. mongolitimonae and R. aeschielmanii); Candidatus Rickettsia rioja and two novel Rickettsia species also were detected. In addition, Borrelia turdi (1.8%) and B. valaisiana (0.9%) were identified in Ixodes ticks. This is the first report of R. slovaca in R. sanguineus s.l. and of R. monacensis, R. raoultii, R. slovaca, R. sibirica subsp. mongolitimonae and Ca. R. rioja in I. frontalis. Since most of the pathogens detected are zoonotic, their presence in these areas may have implications for public health.

Comprehensive micropollutant characterization of wastewater during Covid-19 crisis in 2020: Suspect screening and environmental risk prioritization strategy.

Micropollutants monitoring in wastewater can serve as a picture of what is consuming society and how it can impact the aquatic environment. In this work, a suspect screening approach was used to detect the known and unknown contaminants in wastewater samples collected from two wastewater treatment plants (WWTPs) located in the Basque Country (Crispijana in Alava, and Galindo in Vizcaya) during two weekly sampling campaigns, which included the months from April to July 2020, part of the confinement period caused by COVID-19. To that aim, high-resolution mass spectrometry was used to collect full-scan data-dependent tandem mass spectra from the water samples using a suspect database containing >40,000 chemical substances. The presence of > 80 contaminants was confirmed (level 1) and quantified in both WWTP samples, while at least 47 compounds were tentatively identified (2a). Among the contaminants of concern, an increase in the occurrence of some compounds used for COVID-19 disease treatment, such as lopinavir and hydroxychloroquine, was observed during the lockdown. A prioritization strategy for environmental risk assessment was carried out considering only the compounds quantified in the effluents of Crispijana and Galindo WWTPs. The compounds were scored based on the removal efficiency, estimated persistency, bioconcentration factor, mobility, toxicity potential and frequency of detection in the samples. With this approach, 33 compounds (e.g. amantadine, clozapine or lopinavir) were found to be considered key contaminants in the analyzed samples based on their concentration, occurrence and potential toxicity. Additionally, antimicrobial (RQ-AR) and antiviral (EDRP) risk of certain compounds was evaluated, where ciprofloxacin and fluconazole represented medium risk for antibiotic resistance (1 > RQ-AR > 0.1) in the aquatic ecosystems. Regarding mixture toxicity, the computed sum of toxic unit values of the different effluents (> 1) suggest that interactions between the compounds need to be considered for future environmental risk assessments.

Primary Cutaneous Lymphoma Registry of the Spanish Academy of Dermatology and Venereology (AEDV): Data for the First 5 Years. / Registro de linfomas cutáneos primarios (RELCP) de la AEDV: datos tras 5 años de funcionamiento.

BACKGROUND AND OBJECTIVE: Primary cutaneous lymphomas (PCL) are uncommon. Observations based on the first year of data from the Spanish Registry of Primary Cutaneous Lymphomas (RELCP, in its Spanish abbreviation) of the Spanish Academy of Dermatology and Venereology (AEDV) were published in February 2018. This report covers RELCP data for the first 5 years. PATIENTS AND METHODS: RELCP data were collected prospectively and included diagnosis, treatments, tests, and the current status of patients. We compiled descriptive statistics of the data registered during the first 5 years. RESULTS: Information on 2020 patients treated at 33 Spanish hospitals had been included in the RELCP by December 2021. Fifty-nine percent of the patients were men; the mean age was 62.2 years. The lymphomas were grouped into 4 large diagnostic categories: mycosis fungoides/Sézary syndrome, 1112 patients (55%); primary B-cell cutaneous lymphoma, 547 patients (27.1%); primary CD30+lymphoproliferative disorders, 222 patients (11%), and other T-cell lymphomas, 116 patients (5.8%). Nearly 75% of the tumors were registered in stage I. After treatment, 43.5% achieved complete remission and 27% were stable at the time of writing. Treatments prescribed were topical corticosteroids (1369 [67.8%]), phototherapy (890 patients [44.1%]), surgery (412 patients [20.4%]), and radiotherapy (384 patients [19%]). CONCLUSION: The characteristics of cutaneous lymphomas in Spain are similar to those reported for other series. The large size of the RELCP registry at 5 years has allowed us to give more precise descriptive statistics than in the first year. This registry facilitates the clinical research of the AEDV's lymphoma interest group, which has already published articles based on the RELCP data.

Comparison of conventional and dispersive solid phase extraction clean-up approaches for the simultaneous analysis of tetracyclines and sulfonamides in a variety of fresh vegetables.

The extensive use of antibiotics in agriculture has led to the occurrence of residual drugs in different vegetables frequently consumed by humans. This could pose a potential threat to human health, not only because of the possible effects after ingestion but also because the transmission of antibiotic-resistant genes could occur. In this work, two accurate sample preparation procedures were developed and validated for the simultaneous analysis of sulfonamides (SAs) and tetracyclines (TCs) in four of the most widely consumed vegetables (lettuce, onion, tomato, and carrot) in Europe. The evaluated protocols were based on QuECHERS for extraction and subsequent clean-up by SPE (solid phase extraction) or dispersive SPE. Parameters affecting both extraction and clean-up were carefully evaluated and selected for accuracy of results and minimal matrix effect. Overall, apparent recoveries were above 70% for most of the target analytes with both analytical procedures, and adequate precision (RSD<30%) was obtained for all the matrices. The procedural limits of quantification (LOQPRO) values for SPE clean-up remained below 4.4 µg kg-1 for TCs in all vegetables except for chlortetracycline (CTC) in lettuce (11.3 µg kg-1) and 3.0 µg kg-1 for SAs, with the exception of sulfadiazine (SDZ) in onion (3.9 µg kg-1) and sulfathiazole (STZ) in carrot (5.0 µg kg-1). Lower LOQPRO values (0.1-3.7 µg kg-1) were obtained, in general, when dSPE clean-up was employed. Both methods were applied to twenty-five market vegetable samples from ecological and conventional agriculture and only sulfamethazine (SMZ) and sulfapyridine (SPD) were detected in lettuce at 1.2 µg kg-1 and 0.5 µg kg-1, respectively.

Modifying the magnetic response of magnetotactic bacteria: incorporation of Gd and Tb ions into the magnetosome structure.

Magnetotactic bacteria Magnetospirillum gryphiswaldense MSR-1 biosynthesise chains of cube-octahedral magnetosomes, which are 40 nm magnetite high quality (Fe3O4) nanoparticles. The magnetic properties of these crystalline magnetite nanoparticles, which can be modified by the addition of other elements into the magnetosome structure (doping), are of prime interest in a plethora of applications, those related to cancer therapy being some of the most promising ones. Although previous studies have focused on transition metal elements, rare earth (RE) elements are very interesting as doping agents, both from a fundamental point of view (e.g. significant differences in ionic sizes) and for the potential applications, especially in biomedicine (e.g. magnetic resonance imaging and luminescence). In this work, we have investigated the impact of Gd and Tb on the magnetic properties of magnetosomes by using different complementary techniques. X-ray diffraction, transmission electron microscopy, and X-ray absorption near edge spectroscopy analyses have revealed that a small amount of RE ions, ∼3-4%, incorporate into the Fe3O4 structure as Gd3+ and Tb3+ ions. The experimental magnetic characterisation has shown a clear Verwey transition for the RE-doped bacteria, located at T ∼ 100 K, which is slightly below the one corresponding to the undoped ones (106 K). However, we report a decrease in the coercivity and remanence of the RE-doped bacteria. Simulations based on the Stoner-Wohlfarth model have allowed us to associate these changes in the magnetic response with a reduction of the magnetocrystalline (K C) and, especially, the uniaxial (K uni) anisotropies below the Verwey transition. In this way, K uni reaches a value of 23 and 26 kJ m-3 for the Gd- and Tb-doped bacteria, respectively, whilst a value of 37 kJ m-3 is obtained for the undoped bacteria.

Correction: Modifying the magnetic response of magnetotactic bacteria: incorporation of Gd and Tb ions into the magnetosome structure.

[This corrects the article DOI: 10.1039/D2NA00094F.].

Automated assays for trace elements and ferritin measurement in saliva of pigs: Analytical validation and a pilot application to evaluate different iron status.

The aim of this study was to validate automated methods to measure iron (Fe), zinc (Zn), copper (Cu) and ferritin in pig saliva samples. A complete analytical validation was performed of all assays. In addition, these methods were applied to saliva of Fe supplemented (n = 22) and non-supplemented (n = 20) piglets. All assays were able to measure these biomarkers in pig saliva with adequate precision, accuracy and high sensitivity and, in case of trace elements without needing a deproteinization pre-process. The group of piglets supplemented with Fe presented significantly higher levels of ferritin and Zn in saliva. In conclusion, the automated assays evaluated were able to measure Fe, Zn, Cu and ferritin in saliva of pigs, and in case of trace elements, they have the advantage of not needing a deproteinization pre-treatment and thus these analytes can be measured in a simple and fast manner.

Evaluation of acute phase proteins, adiponectin and endothelin-1 to determine vascular damage in dogs with heartworm disease (Dirofilaria immitis), before and after adulticide treatment.

Previous studies have shown the existence of an acute phase response in dogs with heartworm (Dirofilaria immitis), probably caused by the vascular inflammation that occurs during the pathogenesis of this disease. In addition, it has been seen that this acute phase response persists after finishing treatment, especially in dogs with pulmonary hypertension (PH). Furthermore, echocardiographic studies have shown that PH and endarteritis appear to persist for at least 10 months after completion of adulticide treatment, suggesting that the vascular changes in these dogs may not be reversible. Therefore, the objective of this study was to evaluate the serum concentrations of different positive acute phase proteins (APP) [C reactive-protein (CRP), haptoglobin and ferritin] and negative APP (albumin and paraoxonase-1 (PON-1)), and the usefulness of the endothelin-1 (ET-1) and adiponectin, in dogs infected by D. immitis to evaluate their usefulness as diagnostic biomarkers of vascular damage and PH and their progression throughout therapy up to 7 months after the end of adulticide treatment. Twenty-five heartworm-infected dogs received adulticide treatment, and serum measurements were performed on the day of diagnosis (day 0), day of discharge (day 90), and 6 months after discharge (day 270). In addition, presence or absence of PH was also echocardiographically determined using the Right Pulmonary Artery Distensibility Index. PH was present in 44% of the dogs on day 0 and day 90, and in 48% of dogs on day 270. Alterations were observed in the concentrations of all APP throughout the study, persisting the alterations in PON-1 and ferritin on day 270. Depending on the presence or absence of PH, CRP showed significant differences throughout the study, as did ET-1. On the other hand, adiponectin did not show variations throughout the study, so it did not seem a useful marker in this disease. These results could reflect the possible persistence of vascular inflammation up to 7 months after finishing treatment, whether or not there was PH, and consolidate the study of APP as useful markers in heartworm disease. Moreover, persistent PH could be the consequent clinical manifestation in dogs with more severe vascular alterations so the study of APP, especially CRP, and ET-1 could be especially advantageous in these patients in the early evaluation of the disease, as well as for the determination of disease severity, monitoring therapeutic responses, and predicting outcomes.

Differential Expression of Two Copies of the irmA Gene in the Enteroaggregative E. coli Strain 042.

Gene duplications significantly impact the gene repertoires of both eukaryotic and prokaryotic microorganisms. The genomes of pathogenic Escherichia coli strains share a group of duplicated genes whose function is mostly unknown. The irmA gene is one of the duplicates encoded in several pathogenic E. coli strains. The function of its gene product was investigated in the uropathogenic E. coli strain CFT073, which contains a single functional copy. The IrmA protein structure mimics that of human interleukin receptors and likely plays a role during infection. The enteroaggregative E. coli strain 042 contains two functional copies of the irmA gene. In the present work, we investigated their biological roles. The irmA_4509 allele is expressed under several growth conditions. Its expression is modulated by the global regulators OxyR and Hha, with optimal expression at 37°C and under nutritional stress conditions. Expression of the irmA_2244 allele can only be detected when the irmA_4509 allele is knocked out. Differences in the promoter regions of both alleles account for their differential expression. Our results show that under several environmental conditions, the expression of the IrmA protein in strain 042 is dictated by the irmA_4509 allele. The irmA_2244 allele appears to play a backup role to ensure IrmA expression when the irmA_4509 allele loses its function. IMPORTANCE Gene duplications occur in prokaryotic genomes at a detectable frequency. In many instances, the biological function of the duplicates is unknown, and hence, the significance of the presence of multiple copies of these genes remains unclear. In pathogenic E. coli isolates, the irmA gene can be present either as a single copy or in two or more copies. We focused our work on studying why a different pathogenic E. coli strain encodes two functional copies of the irmA gene. We show that under several environmental conditions, one of the alleles dictates IrmA expression, and the second remains silent. The latter allele is only expressed when the former is silenced. The presence of more than one functional copy of the irmA gene in some pathogenic E. coli strains can result in sufficient expression of this virulence factor during the infection process.