Flexible Label-Free Platinum and Bio-PET-Based Immunosensor for the Detection of SARS-CoV-2.

The demand for new devices that enable the detection of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) at a relatively low cost and that are fast and feasible to be used as point-of-care is required overtime on a large scale. In this sense, the use of sustainable materials, for example, the bio-based poly (ethylene terephthalate) (Bio-PET) can be an alternative to current standard diagnostics. In this work, we present a flexible disposable printed electrode based on a platinum thin film on Bio-PET as a substrate for the development of a sensor and immunosensor for the monitoring of COVID-19 biomarkers, by the detection of L-cysteine and the SARS-CoV-2 spike protein, respectively. The electrode was applied in conjunction with 3D printing technology to generate a portable and easy-to-analyze device with a low sample volume. For the L-cysteine determination, chronoamperometry was used, which achieved two linear dynamic ranges (LDR) of 3.98-39.0 µmol L-1 and 39.0-145 µmol L-1, and a limit of detection (LOD) of 0.70 µmol L-1. The detection of the SARS-CoV-2 spike protein was achieved by both square wave voltammetry (SWV) and electrochemical impedance spectroscopy (EIS) by a label-free immunosensor, using potassium ferro-ferricyanide solution as the electrochemical probe. An LDR of 0.70-7.0 and 1.0-30 pmol L-1, with an LOD of 0.70 and 1.0 pmol L-1 were obtained by SWV and EIS, respectively. As a proof of concept, the immunosensor was successfully applied for the detection of the SARS-CoV-2 spike protein in enriched synthetic saliva samples, which demonstrates the potential of using the proposed sensor as an alternative platform for the diagnosis of COVID-19 in the future.

Novel and highly stable strategy for the development of microfluidic enzymatic assays based on the immobilization of horseradish peroxidase (HRP) into cotton threads.

The use of biological components in the development of new methods of analysis and point-of-care (POC) devices is an ever-expanding theme in analytical chemistry research, due to the immense potential for early diagnosis of diseases and monitoring of biomarkers. In the present work, the evaluation of an electrochemical microfluidic device based on the immobilization of horseradish peroxidase (HRP) enzyme into chemically treated cotton threads is described. This bioreactor was used as a channel for the build of the microfluidic device, which has allowed to use of a non-modified screen-printed electrode (SPE) as an amperometric detector. Cotton threads were treated using citric acid, and the immobilization of HRP has been performed by EDC/NHS crosslinking, connecting amine groups of the enzymes to carboxylic acids in the cellulosic structure. For the analytical evaluation, an amperometric assay for hydrogen peroxide detection was performed after the injection of H2O2 and hydroquinone (HQN) concomitantly. The enzymatic reaction consumes H2O2 leading to the formation of O-quinone, which is readily reducible at non-modified SPE. Several experimental parameters related to enzyme immobilization have been investigated and under the best set of conditions, a good analytical performance was obtained. In addition, the threads were freezer-stored and, after 12 weeks, 84% of hydrogen peroxide sensitivity was maintained, which is very reasonable for enzyme-based systems and still offers good analytical precision. Therefore, a simple and inexpensive microfluidic system was reported by crosslinking carboxylic groups to amine-containing macromolecules, suggesting a new platform for many other protein-based assays.

Influence of filament aging and conductive additive in 3D printed sensors.

3D printing technology combined with electrochemical techniques have allowed the development of versatile and low-cost devices. However, some aspects need to be considered for the good quality and useful life of the sensors. In this work, we have demonstrated herein that the filament aging, the conductive material, and the activation processes (post-treatments) can influence the surface characteristics and the electrochemical performance of the 3D printed sensors. Commercial filaments and 3D printed sensors were morphologically, thermally, and electrochemically analyzed. The activated graphene-based (Black Magic®) sensor showed the best electrochemical response, compared to the carbon black-filament (Proto-Pasta®). In addition, we have proven that filament aging harms the performance of the sensors since the electrodes produced with three years old filament had a considerably lower intra-days reproducibility. Finally, the activated graphene-based sensor has shown the best performance for the electrochemical detection of bisphenol A, demonstrating the importance of evaluating and control the characteristics and quality of filaments to improve the mechanical, conductive, and electrochemical performance of 3D printed sensors.

Biosensing strategies for the electrochemical detection of viruses and viral diseases - A review.

Viruses are the causing agents for many relevant diseases, including influenza, Ebola, HIV/AIDS, and COVID-19. Its rapid replication and high transmissibility can lead to serious consequences not only to the individual but also to collective health, causing deep economic impacts. In this scenario, diagnosis tools are of significant importance, allowing the rapid, precise, and low-cost testing of a substantial number of individuals. Currently, PCR-based techniques are the gold standard for the diagnosis of viral diseases. Although these allow the diagnosis of different illnesses with high precision, they still present significant drawbacks. Their main disadvantages include long periods for obtaining results and the need for specialized professionals and equipment, requiring the tests to be performed in research centers. In this scenario, biosensors have been presented as promising alternatives for the rapid, precise, low-cost, and on-site diagnosis of viral diseases. This critical review article describes the advancements achieved in the last five years regarding electrochemical biosensors for the diagnosis of viral infections. First, genosensors and aptasensors for the detection of virus and the diagnosis of viral diseases are presented in detail regarding probe immobilization approaches, detection methods (label-free and sandwich), and amplification strategies. Following, immunosensors are highlighted, including many different construction strategies such as label-free, sandwich, competitive, and lateral-flow assays. Then, biosensors for the detection of viral-diseases-related biomarkers are presented and discussed, as well as point of care systems and their advantages when compared to traditional techniques. Last, the difficulties of commercializing electrochemical devices are critically discussed in conjunction with future trends such as lab-on-a-chip and flexible sensors.

A Reliable Stem Cell Carrier: An Experimental Study in Wistar Rats.

INTRODUCTION: Treatments based on cell biology need reliable and precise carriers for reaching the desired targets. For that reason, a PDO-based cell carrier was idealized, with the purpose of carrying stem cells to distant sites at room temperature. MATERIALS AND METHODS: Three modalities of the same carrier were evaluated: one containing undifferentiated human dental pulp stem cells (DPSCs); one loaded with stem cells induced to neurogenic differentiation (DPSCNs); and one without cells (Blank). The carriers were implanted in sciatic nerve gaps in 48 Wistar rats that were divided in three groups. Two other rats were included in a SHAM control group. Immunohistochemical, histological and clinical analyses were performed in two, four, six and eight weeks of time. RESULTS: Efficacy of human stem cell transportation at room temperature to rats was attested. Moreover, it was possible to confirm that those cells show tropism for inflamed environments and are also prone to induction of neurogenesis in the first two weeks, vanishing after that period. CONCLUSION: Clinical evaluation of the animals' gait recovery shows a promising perspective of success with the inclusion of stem cell-loaded PDO tubes in nerve gaps, which may be positively compared to previously published studies. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors - www.springer.com/00266.

Palonosetron versus ondansetron for prevention of nausea and vomiting after total abdominal hysterectomy under spinal anesthesia with intrathecal morphine: a double-blind, randomized controlled trial.

BACKGROUND: Hysterectomy is a widely performed surgery and neuraxial anesthesia with intrathecal morphine provides superior quality of recovery. Postoperative nausea and vomiting (PONV) is a frequent problem with intrathecal morphine use. Although palonosetron is effective for prevention of PONV after general anesthesia, its efficacy after neuraxial anesthesia has not been established. This study was conducted to compare the use of palonosetron with ondansetron for PONV prophylaxis in patients at a high risk of PONV during total abdominal hysterectomy (TAH) under spinal anesthesia with intrathecal morphine. METHODS: This prospective, randomized double-blind study conducted at São Rafael Hospital involved 140 American Society of Anesthesiologists physical status I or II women who underwent TAH under spinal anesthesia with intrathecal morphine and who had at least 3 risk factors for PONV based on Apfel's simplified score. The patients were randomized into two groups: one received palonosetron whereas the other received ondansetron. All patients received spinal anesthesia with intrathecal morphine, as well as dexamethasone plus palonosetron or ondansetron for PONV prophylaxis. The overall incidence of PONV, incidence of early- and late-onset nausea and vomiting, severity of nausea, and use of rescue antiemetics were recorded. RESULTS: The overall incidence of PONV was 42.9% in the palonosetron group and 52.9% in the ondansetron group (p > 0.05). No significant differences existed in the incidence of early- and late-onset nausea or early-onset vomiting between the two groups. The incidence of late-onset vomiting was significantly lower in the palonosetron group. CONCLUSIONS: Palonosetron exhibited efficacy similar to that of ondansetron for reducing the overall incidence of PONV after TAH under spinal anesthesia with intrathecal morphine; however, palonosetron reduced the incidence of late-onset vomiting significantly better than ondansetron. TRIAL REGISTRATION: RBR-4gnm8n ( ensaiosclinicos.gov.br ), date of registration: August 18, 2014.

Epidemiology of Megninia spp. in laying flocks from the State of Minas Gerais, Brazil.

Aspects related to the epidemiology of Megninia ginglymura were studied, in order to determine predisposing factors for the occurrence in layer chicken houses of commercial farms in the State of Minas Gerais, Brazil. An observational, cross-sectional, analytical survey was conducted for identifying major risks or protection factors for infestations. Four hundred thirty-one houses located in forty-three farms were visited in 2012. Regarding the farms, 17 (39.53%) had chickens infested by M. ginglymura and 1 (2.32%) presented Megninia cubitalis. The epidemiological aspects of the occurrence of M. ginglymura were investigated using multiple logistic regression models. The chicken house organization was an important protection factor involved in the occurrence of M. ginglymura (odds ratio [OR] = 0.85). The presence of subsistence breedings of poultry within the premises or at the proximity of a farm was characterized as a risk factor for the occurrence of infestations (OR = 3.92). The use of mineral oil (OR = 0.16) was considered protective against new infestations. Some regions of the state were characterized by a higher risk for mite infestation than others.

An electroanalytical approach for evaluation of biochar adsorption characteristics and its application for lead and cadmium determination.

This work describes for first time the use of electroanalytical techniques for evaluation of adsorptive proprieties of biochar using it as electrode modifier and its application for preconcentration and determination of Lead(II) and Cadmium(II) under differential pulse adsorptive voltammetric conditions (DPAdSV). Samples of biochars were obtained from castor oil cake using a predefined set of experimental conditions varying the heating rate (V), final temperature (T) and warm-up period (P) and subsequently used for carbon paste modified electrode (CPME) preparation. The proposed method was applied for Lead(II) and Cadmium(II) determination in spiked simulated industrial effluents and the limit of detection obtained for both metals were adequated for determination of these evaluated ions taking into account the limits established by Brazilian legislation. For all samples analyzed, recoveries ranged from 95% to 104% were obtained and no significative interferences were observed for common cations in water samples.

Principal component analysis on the perceptions of milk producers about Rhipicephalus (Boophilus) microplus control in Minas Gerais.

Milk producers in Lavras, Passos and Divinópolis, Minas Gerais, were interviewed with the aim of evaluating their perceptions and attitudes regarding control over Rhipicephalus (Boophilus) microplus. Multivariate correlation between the variables was done by means of principal component analysis. The producers' perceptions and attitudes regarding R. (B.) microplus were similar: most of them did not have any basic knowledge of tick biology or control, and they applied acaricide products through backpack spraying, without any defined technical criteria. Some of the results obtained were: I. a negative correlation between schooling level and the frequency of spraying cattle with acaricides; II. a positive correlation between milk production, quality of equipment for acaricide application and technological level of the farm; III. farm properties with greater production and technification tended to keep the intervals between acaricide applications constant over the course of the year. After applying principal component analysis, a positive correlation was observed between schooling level, technological level of the farm and perceptions regarding R. (B.) microplus, but without any correlation with attitudes towards controlling this tick. It was concluded that higher technological level and schooling level improved the producers' perceptions relating to the biology of the tick R. (B.) microplus, but did not achieve effectiveness with regard to using controls more rationally.

Quill mites in Brazilian psittacine birds (Aves: Psittaciformes).

The primary and secondary feathers of 170 Brazilian psittacine birds (Aves: Psittaciformes) were examined in order to identify feather quill mite fauna. Birds were held captive in two locations in the state of Minas Gerais (MG), and two in the state of Espirito Santo (ES). The quills were cut longitudinally and were examined under optical microscopy. The genus of quill mites most frequently found was Paralgopsis (Astigmata: Pyrogliphidae), followed by Cystoidosoma (Astigmata: Syringobiidae). Astigmata: Syringophilidae mites were sporadically observed. After analyzing the data using logistic regression models, it was determined that there was higher infestation risk for psittacines in ES state, as compared with those in MG, and a significant increase in risk depending on the psittacine host species. However, the location of captivity did not have a significant effect. Lesions were observed in infested feathers. Cystoidosoma sp. and Paralgopsis sp. were always observed together, with parts of Paralgopsis found inside Cystoidosoma sp., suggesting thanatochresis or predation.

Hepatozoon spp. (Apicomplexa: Hepatozoidae) infection and selected hematological values of the neotropical rattlesnake, Crotalus durissus collilineatus (Linnaeus, 1758) (Serpentes: Viperidae), from Brazil.

This study aims to establish the hematological values of Crotalus durissus collilineatus snakes captured in Brazil as well as to verify the effects of hematozoan infection on these snakes. Eighty-three blood samples were drawn from C. d. collilineatus specimens for analysis. The sample set was composed of 30 males and 30 females, recently caught from the wild, and 11 males and 12 females bred in captivity. Blood samples were used to determine red blood cell counts, white blood cell counts, thrombocyte counts, hematocrit values, hemoglobin concentration, and total plasma protein. Blood smears were used to diagnose Hepatozoon spp. infection and to calculate the parasitic load in the sample as well as the percentage of immature red cells. Results obtained for the wild-caught animals, with and without parasites, were compared among themselves and with the values obtained for the captive-bred animals. Hematological values for C. durissus were established. Wild-caught snakes had an infection rate of 38.3%, while no Hepatozoon sp. infection was detected in the captive-bred animals. The snakes which were not infected by the Hepatozoon sp. exhibited average weight, length, and weight-length ratios higher than those of the infected animals. An increase in immature red cells was noted in the Hepatozoon-infected snakes.

Amblyomma rotundatum (Koch, 1844) (Acari: Ixodidae) two-host life-cycle on Viperidae snakes.

Amblyomma rotundatum is an ixodid tick that infests ectothermic animals and reproduces exclusively by parthenogenesis. This tick has been frequently reported to infest reptiles and amphibians, under natural conditions and sometimes in captivity. It was described in Brazil and several other countries of South, Central and North America. Although many studies have reported aspects of its biology, none of them has used regularly either ophidian as hosts, or controlled temperature, humidity and luminosity for parasitic stages. The objective of this experiment was to study the life cycle of A. rotundatum feeding on Viperidae snakes under room controlled conditions at 27 ± 1 °C temperature, 85 ± 10% relative humidity and 12:12 hours photoperiod for parasitic stages, and under B.O.D incubator conditions at 27 ± 1 °C temperature, 85 ± 10% relative humidity and scotophase for non-parasitic stages. The total duration of the life cycle ranged from 56 to 163 days (mean of 105 days). Two-host life cycle was observed for most of the ixodid population studied.

Salivary immunoglobulin a response to a match in top-level brazilian soccer players.

It has been suggested that several parameters of mucosal immunity, including salivary immunoglobulin A (s-IgA), are affected by heavy exercise either in field sports or in the laboratory environment. Few observations have been made during a true sporting environment, particularly in professional soccer. We tested the hypothesis that salivary IgA levels will be decreased after a 70-minute regulation in a top-level professional soccer friendly match. Saliva samples from 24 male professional soccer players collected before and after the match were analyzed. Salivary immunoglobulin A concentration was measured by enzyme-linked immunosorbent assay and expressed as the absolute concentration (s-IgAabs), s-IgA relative to total protein concentration (IgA-Pro), and the secretion rate of IgA (s-IgArate). Rate of perceived exertion (RPE) was used to monitor the exercise intensity. The paired t-test showed no significant changes in s-IgAabs and s-IgArate (p > 0.05) from PRE to POST match. However, a significant (p < 0.05) increase in total protein concentration (1.46 +/- 0.4 to 2.00 +/- 07) and a decrease in IgA-Pro were observed. The best and most significant correlation was obtained with the RPE and changes in IgA-Pro (rs = -0.43) and could indicate that this expression may be an interesting marker of intensity in a soccer match. However, further investigation regarding exercise intensity, protein concentration, and immune suppression, particularly in team sports, is warranted. From a practical application, the variability of the responses among the players leads us to suggest that there is a need to individually analyze the results with team sports. Some athletes showed a decrease in s-IgA expressions, suggesting the need for taking protective actions to minimize contact with cold viruses or even reducing the training load.

The impact of a 17-day training period for an international championship on mucosal immune parameters in top-level basketball players and staff members.

This investigation examined the impact of a 17-d training period (that included basketball-specific training, sprints, intermittent running exercises, and weight training, prior to an international championship competition) on salivary immunoglobulin A (SIgA) levels in 10 subjects (athletes and staff members) from a national basketball team, as a biomarker for mucosal immune defence. Unstimulated saliva samples were collected at rest at the beginning of the preparation for the Pan American Games and 1 d before the first game. The recovery interval from the last bout of exercise was 4 h. The SIgA level was measured using enzyme-linked immunosorbent assay and expressed as absolute concentrations, secretion rate, and SIgA level relative to total protein. The decrease in SIgA levels following training was greater in athletes than in support staff; however, no significant differences between the two groups were detected. A decrease in SIgA level, regardless of the method used to express IgA results, was verified for athletes. Only one episode of upper respiratory tract illness symptoms was reported, and it was not associated with changes in SIgA levels. In summary, a situation of combined stress for an important championship was found to decrease the level of SIgA-mediated immune protection at the mucosal surface in team members, with greater changes observed in the athletes.

[Water quality evaluation in rural areas of Lavras, Minas Gerais, Brazil, 1999-2000]. / Avaliação da qualidade da água e percepção higiênico-sanitária na área rural de Lavras, Minas Gerais, Brasil, 1999-2000.

In addition to personal interviews, laboratory analyses were performed using 80 water samples from 45 rural areas that are crossed by the Agua Limpa and Santa Cruz streams close to the city of Lavras, southern Minas Gerais State. The results allowed comparing the quality of water used for agriculture and the identification of determinant factors. The Agua Limpa stream mostly crosses an area used primarily for housing and characterized by low schooling. Many houses are supplied by shallow water wells and have ordinary cesspits for human waste disposal. All springs are polluted. The Santa Cruz stream displays a different scenario. The land is used mostly for agricultural purposes. Most owners live in town, with widely varied levels of school, from none to university. The houses are supplied by surface water. Most of the springs are polluted. The perception by both home and land owners concerning quality of the drinking water is determined solely by the water's physical and organoleptic characteristics. Sanitary parameters are not taken into account. Moreover, there is no relationship between fecal contamination and the type of spring. Land use and anthropic activity are far more important than the type of spring for water quality.

Biology of Amblyomma aureolatum (Pallas, 1772) (Acari: Ixodidae) on some laboratory hosts in Brazil.

The ixodid Amblyomma aureolatum is suspected to play a role in the epidemiology of wild life-cycle hemoparasites, which frequently infect dogs in rural and hunting areas in Brazil. Little is known about its bionomics. The objective of the present study was to evaluate some bionomic aspects of A. aureolatum ticks in Brazil. One engorged female, collected from a dog (Canis familiaris) in São Sebastião das Aguas Claras, State of Minas Gerais, was used to establish a colony in the laboratory. Subsequently its parasitic stage progeny were fed on domestic dogs and laboratory animals. The free-living stages were incubated at 27 degrees C +/- 2 degrees C and minimum 70% relative humidity in a BOD incubator. The egg incubation period ranged from 31 to 34 days; the parasitic period of larvae ranged from 4 to 6 days and ecdysis to nymphs occurred from day 19 up to day 22. The parasitic period of nymphs ranged from 5 to 8 days and the period of ecdysis to adults from 31 to 33 days. The parasitic period of adults ranged from 11 to 15 days, the pre-oviposition period from 6 to 12 days, and the oviposition period from 9 to 38 days. The total duration of the life cycle ranged from 116 to 168 days.