Progress of the Detection Methods for SARS-CoV-2.

BACKGROUND: The severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) causes a respiratory illness, characterized by symptoms such as fever, dry cough, and drowsiness. This virus is highly contagious, has significant mutation rates, and induces infection despite vaccination. Its widespread prevalence has profoundly impacted global economies, societies, and daily life. In response to these challenges, researchers have committed themselves to advancing rapid and cost-effective diagnostic technologies, holding substantial importance for the rapid evolution of global diagnostic capabilities. Nonetheless, various detection methods diverge in principles, sensitivity, specificity, and other aspects. Additionally, COVID-19 is not an isolated event, but part of a broader history of pandemics in human society. Therefore, this article briefly reviews the existing detection methods of SARS-CoV-2, providing valuable technical insights to diagnose not only SARS-CoV-2 but also other viruses. METHODS: A search was conducted on PubMed by utilizing keywords such as "SARS-CoV-2 detection", "RT-qPCR detection for SARS-CoV-2", "LFA detection for SARS-CoV-2", "Biosensors detection for SARS-CoV-2", and similar terms. The objective was to compile and summarize relevant articles on these topics. RESULTS: Currently, the real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) stands as a widely employed method for detecting SARS-CoV-2, enabling an accurate detection of viral RNA. Furthermore, the lateral flow assay (LFA) assists in detecting viral antigens and antibodies. Gene sequencing technology primarily facilitates the real-time monitoring of mutated SARS-CoV-2 strains, while biosensors could offer a rapid, economical, sensitive, and precise detection of SARS-CoV-2. These methods provide a strong technical support for the early detection and diagnosis of SARS-CoV-2. CONCLUSIONS: This paper offers a concise overview of pathogen detection methods, as molecular biology, and immunological detection techniques, alongside emerging biosensor platforms relevant to SARS-CoV-2, and delineates the strengths and weaknesses of each method.

Electrochemical biosensors for pathogenic microorganisms detection based on recognition elements.

The worldwide spread of pathogenic microorganisms poses a significant risk to human health. Electrochemical biosensors have emerged as dependable analytical tools for the point-of-care detection of pathogens and can effectively compensate for the limitations of conventional techniques. Real-time analysis, high throughput, portability, and rapidity make them pioneering tools for on-site detection of pathogens. Herein, this work comprehensively reviews the recent advances in electrochemical biosensors for pathogen detection, focusing on those based on the classification of recognition elements, and summarizes their principles, current challenges, and prospects. This review was conducted by a systematic search of PubMed and Web of Science databases to obtain relevant literature and construct a basic framework. A total of 171 publications were included after online screening and data extraction to obtain information of the research advances in electrochemical biosensors for pathogen detection. According to the findings, the research of electrochemical biosensors in pathogen detection has been increasing yearly in the past 3 years, which has a broad development prospect, but most of the biosensors have performance or economic limitations and are still in the primary stage. Therefore, significant research and funding are required to fuel the rapid development of electrochemical biosensors. The overview comprehensively evaluates the recent advances in different types of electrochemical biosensors utilized in pathogen detection, with a view to providing insights into future research directions in biosensors.

The Status of Occupational Stress and Its Influence on the Health of Medical Staff in Lanzhou, China.

This study aimed to understand the status quo of occupational stress and its impact on the health of medical staff and provide a theoretical basis for relieving occupational stress and improving the health status of medical staff. The occupational stress and health status of medical staff in 14 hospitals in Lanzhou were studied using a general questionnaire, Effort-Reward Imbalance questionnaire, and Self-Rated Health Measurement Scale. A total of 2169 participants were included in the analysis, and 59.4% of the medical staff experienced occupational stress. The results of the occupational stress survey showed that the prevalence of occupational stress among medical staff aged 40-50, with a master's degree or above, senior professional title, working for 10-20 years, and working more than 48 h per week was higher than in the other groups. The health survey results showed that, compared with other groups, the scores of physical, mental, and social health were lower in medical staff with working years of 10-20 years and working hours of more than 48 h per week. The results show that working years and working hours per week affect not only the level of occupational stress but also physiological, psychological, and social health.

Dietary ß-sitosterol regulates serum lipid level and improves immune function, antioxidant status, and intestinal morphology in broilers.

This research investigated effects of dietary ß-sitosterol addition at different levels on serum lipid levels, immune function, oxidative status, and intestinal morphology in broilers. One-day-old broiler chicks were allocated to 5 groups of 6 replicates. Chickens in the 5 groups were fed a basal diet supplemented with 0 (control group), 40, 60, 80, and 100 mg/kg of ß-sitosterol for 42 D, respectively. ß-Sitosterol linearly decreased (P < 0.05) concentrations of serum total cholesterol, jejunal tumor necrosis factor α (TNF-α), and ileal interleukin 1ß (IL-1ß) and mRNA relative expressions levels of jejunal TLR4 and ileal MyD88, whereas it linearly increased (P < 0.05) contents of jejunal immunoglobulin G (IgG), ileal secreted IgA and glutathione, jejunal catalase activity and Nrf2 mRNA relative expression level, villus height (VH), and VH-to-crypt depth (CD) ratio (VH:CD) in the jejunum and ileum. Linear and quadratic increases (P < 0.05) in absolute and relative spleen weight were observed by dietary ß-sitosterol, whereas malondialdehyde (MDA) concentration in the jejunum and ileum followed the opposite trend (P < 0.05). Compared with the control group, dietary ß-sitosterol at higher than or equal to 60 mg/kg level decreased (P < 0.05) contents of serum total cholesterol, ileal MDA, and jejunal TLR4 mRNA relative expression level, whereas it increased (P < 0.05) absolute spleen weight and ileal glutathione content. Higher than or equal to 80 mg/kg level of ß-sitosterol enhanced (P < 0.05) jejunal IgG concentration, VH, catalase activity, and Nrf2 relative expression level and ileal secreted IgA content, but reduced (P < 0.05) ileal IL-1ß content and MyD88 mRNA relative expression level. ß-Sitosterol addition at 60 and 80 mg/kg levels increased (P < 0.05) relative spleen weight, whereas it decreased (P < 0.05) jejunal MDA accumulation. Moreover, 100 mg/kg level of ß-sitosterol reduced (P < 0.05) jejunal TNF-α level, but it increased (P < 0.05) VH in the jejunum and VH:CD in the jejunum and ileum. Accordingly, dietary ß-sitosterol supplementation could regulate serum cholesterol level, promote immune function, and improve intestinal oxidative status and morphology in broilers.

Effects of Dietary Zeolite Supplementation as an Antibiotic Alternative on Growth Performance, Intestinal Integrity, and Cecal Antibiotic Resistance Genes Abundances of Broilers.

The study investigated the effects of dietary zeolite supplementation as an antibiotic alternative on growth performance, intestinal integrity, and cecal antibiotic resistance genes abundances of broilers. One-day-old chicks were assigned into three groups and fed a basal diet or a basal diet supplemented with antibiotics (50 mg/kg) or zeolite (10 g/kg). Antibiotic or zeolite increased (p < 0.05) average daily gain (ADG) from 1 to 42 days and duodenal villus height to crypt depth ratio (VH:CD) at 21 days. Zeolite increased (p < 0.05) ADG and average daily feed intake from 1 to 21 days, jejunal VH:CD at 21 and 42 days, ileal VH and VH:CD at 42 days, zonula occludens-1 mRNA abundance at 21 days, and duodenal occludin mRNA abundance at 42 days, whereas reduced (p < 0.05) jejunal CD and malondialdehyde levels in ileum at 21 days and duodenum at 42 days, serum D-lactic acid and diamine oxidase levels at 42 days, and plasma lipopolysaccharide content at 21 and 42 days. Antibiotics reduced (p < 0.05) duodenal claudin-2 mRNA abundance at 21 days, whereas increased (p < 0.05) cecal tetB abundance at 42 days. These findings suggested that the beneficial effects of zeolite in broilers were more pronounced than that of antibiotics.

Dietary Chitooligosaccharide Inclusion as an Alternative to Antibiotics Improves Intestinal Morphology, Barrier Function, Antioxidant Capacity, and Immunity of Broilers at Early Age.

This study aimed to investigate the effects of chitooligosaccharide (COS) inclusion as an alternative to antibiotics on growth performance, intestinal morphology, barrier function, antioxidant capacity, and immunity in broilers. In total, 144 one-day-old Arbor Acres broiler chicks were randomly assigned into 3 groups and fed a basal diet free from antibiotics (control group) or the same basal diet further supplemented with either chlortetracycline (antibiotic group) or COS, for 21 days. Compared with the control group, inclusion of COS reduced the feed to gain ratio, the jejunal crypt depth, the plasma diamine oxidase activity, and the endotoxin concentration, as well as jejunal and ileal malondialdehyde contents, whereas increased duodenal villus height, duodenal and jejunal ratio of villus height to crypt depth, intestinal immunoglobulin G, and jejunal immunoglobulin M (IgM) contents were observed, with the values of these parameters being similar or better to that of the antibiotic group. Additionally, supplementation with COS enhanced the superoxide dismutase activity and IgM content of the duodenum and up-regulated the mRNA level of claudin three in the jejunum and ileum, when compared with the control and antibiotic groups. In conclusion, dietary COS inclusion (30 mg/kg), as an alternative to antibiotics, exerts beneficial effects on growth performance, intestinal morphology, barrier function, antioxidant capacity, and immunity in broilers.

Dietary ß-Sitosterol Improves Growth Performance, Meat Quality, Antioxidant Status, and Mitochondrial Biogenesis of Breast Muscle in Broilers.

The present study evaluated effects of ß-sitosterol on growth performance, meat quality, oxidative status, and mitochondrial biogenesis of breast muscle in broilers. One-day-old chicks were allocated to five treatments of six replicates. Broilers were fed a basal diet supplemented either with 0 (control), 40, 60, 80, or 100 mg/kg ß-sitosterol for 42 days. ß-sitosterol linearly and quadratically reduced feed/gain ratio, lightness24h and cooking loss24h in breast muscle, whereas 2, 2-diphenyl-1-picrylhydrazyl scavenging activity of breast muscle followed an opposite trend. ß-sitosterol linearly decreased drip loss24h and malondialdehyde content, whereas linearly increased pH24h, superoxide dismutase activity, and mRNA abundances of peroxisome proliferator-activated receptor γ coactivator 1α (PCG-1α) and mitochondrial transcription factor A (TFAM) in breast muscle. Compared with control, levels of ß-sitosterol higher than 40 mg/kg reduced feed/gain ratio, muscular lightness24h, cooking loss24h, and malondialdehyde level, whereas increased muscular 2, 2-diphenyl-1-picrylhydrazyl scavenging activity, and mRNA abundances (except 60 mg/kg) of PCG-1α and TFAM. Eighty milligram/kilogram ß-sitosterol increased muscular pH24h and superoxide dismutase activity, but decreased its drip loss24h. Therefore, ß-sitosterol could improve growth performance and meat quality, oxidative status, and mitochondrial biogenesis of breast muscle in broilers. Furthermore, supplementation level of 80 mg/kg ß-sitosterol is recommended for broiler diets.

Dietary Tea Tree (Melaleuca alternifolia) Oil Supplementation Improves Growth Performance, Cecal Microflora, Immunity, and Antioxidant Capacity of Partridge Shank Chickens.

The aim of this study was to investigate the effects of tea tree oil (TTO) supplementation on the growth performance, cecal microflora composition, immunity, and antioxidant status of Partridge Shank chickens. A total of 144 one-day-old chicks were allocated into three treatments with six replicates of eight chicks each and fed with a basal diet supplemented with 0 (Control group), 500, and 1000 mg/kg TTO for 50 days. Compared with the control group, the broilers fed with the basal diet supplemented with 1000 mg/kg TTO exhibited an increase in average daily gain from 22 to 50 days (P=0.035) and in both relative thymus weight (P<0.001) and Lactobacillus colonies in the cecal contents (P=0.045) at 50 days of age, but a reduction in the feed/gain ratio during 1 to 50 days (P=0.048). Additionally, dietary TTO supplementation, irrespective of dosage, increased the relative spleen weight (P=0.003) and total antioxidant capacity in the jejunum (P=0.049) and ileum (P=0.001) at 21 days, but decreased the malondialdehyde content in the ileum at both 21 (P=0.003) and 50 days (P<0.001) and in the jejunum at 50 days (P=0.012). The results suggested that TTO supplementation could improve the growth performance, cecal microflora composition, immunity, and antioxidant capacity of Partridge Shank chickens.

LAMP, a new imaging assay of gap junctional communication unveils that Ca2+ influx inhibits cell coupling.

Using a new class of photo-activatible fluorophores, we have developed a new imaging technique for measuring molecular transfer rates across gap junction connexin channels in intact living cells. This technique, named LAMP, involves local activation of a molecular fluorescent probe, NPE-HCCC2/AM, to optically label a cell. Subsequent dye transfer through gap junctions from labeled to unlabeled cells was quantified by fluorescence microscopy. Additional uncagings after prior dye transfers reached equilibrium enabled multiple measurements of dye transfer rates in the same coupled cell pair. Measurements in the same cell pair minimized variation due to differences in cell volume and number of gap junctions, allowing us to track acute changes in gap junction permeability. We applied the technique to study the regulation of gap junction coupling by intracellular Ca(2+) ([Ca(2+)](i)). Although agonist or ionomycin exposure can raise bulk [Ca(2+)](i) to levels higher than those caused by capacitative Ca(2+) influx, the LAMP assay revealed that only Ca(2+) influx through the plasma membrane store-operated Ca(2+) channels strongly reduced gap junction coupling. The noninvasive and quantitative nature of this imaging technique should facilitate future investigations of the dynamic regulation of gap junction communication.

New caged coumarin fluorophores with extraordinary uncaging cross sections suitable for biological imaging applications.

Photocaged fluorescent molecules are important research tools for tracking molecular dynamics with high spatiotemporal resolution in biological systems. We have designed and synthesized a new class of caged coumarin fluorophores. These coumarin cages displayed more than 200-fold fluorescence enhancement after UV photolysis. Remarkably, the uncaging cross section of a 1-(2-nitrophenyl)ethyl (NPE)-caged coumarin is 6600 at wavelength of 365 nm, about 2 orders of magnitude higher than previously described caged fluorophores. Product analysis of the photolytic reaction showed clean conversion of NPE-caged coumarin to 2-nitrosoacetophenone and the parent coumarin, suggesting that the mechanism of the photolysis follows the known photochemical reaction pathway of the 2-nitrobenzyl group. We have also measured the two-photon uncaging cross sections of NPE-caged coumarins 2a and 5 at 740 nm to be near 1 Goeppert-Mayer (GM). The mechanistic study, together with the two-photon uncaging data, suggested that the coumarin moiety serves as an antenna to enhance the light harvesting efficiency of the coumarin cage and that the photonic energy absorbed by coumarin was utilized efficiently to photolyze the NPE group. Future explorations of this type of "substrate-assisted photolysis" may yield other cages of high uncaging cross sections. For cellular imaging applications, we prepared a cell permeable and caged coumarin fluorophore, NPE-HCCC2/AM (10), which can be loaded into fully intact cells to high concentrations. Initial tests of this probe in a number of cultured mammalian cells showed desired properties for the in vivo imaging applications. The combined advantages of robust fluorescence contrast enhancement, remarkably high uncaging cross sections, noninvasive cellular delivery, and flexible chemistry for bioconjugations should generate broad applications of these caged coumarins in biochemical and biological research.