Search

Asthma is a chronic respiratory condition predominantly driven by a type 2 immune response. Epithelial-derived alarmins such as thymic stromal lymphopoietin (TSLP), interleukin (IL)-33, and IL-25 orchestrate the activation of downstream Th2 cells and group 2 innate lymphoid cells (ILC2s), along with other immune effector cells. While these alarmins are produced in response to inhaled triggers, such as allergens, respiratory pathogens or particulate matter, disproportionate alarmin production by airway epithelial cells can lead to asthma exacerbations. With alarmins produced upstream of the type 2 inflammatory cascade, understanding the pathways by which these alarmins are regulated and expressed is critical to further explore new therapeutics for the treatment of asthmatic patients. This review emphasizes the critical role of airway epithelium and epithelial-derived alarmins in asthma pathogenesis and highlights the potential of targeting alarmins as a promising therapeutic to improve outcomes for asthma patients. [ABSTRACT FROM AUTHOR]

Disruption of the airway epithelium triggers a defensive immune response that begins with the production and release of alarmin cytokines. These epithelial-derived alarmin cytokines, including thymic stromal lymphopoietin (TSLP), are produced in response to aeroallergens, viruses, and toxic inhalants. An alarmin response disproportionate to the inhaled trigger can exacerbate airway diseases such as asthma. Allergens inhaled into previously sensitized airways are known to drive a T2 inflammatory response through the polarization of T cells by dendritic cells mediated by TSLP. Harmful compounds found within air pollution, microbes, and viruses are also triggers causing airway epithelial cell release of TSLP in asthmatic airways. The release of TSLP leads to the development of inflammation which, when unchecked, can result in asthma exacerbations. Genetic and inheritable factors can contribute to the variable expression of TSLP and the risk and severity of asthma. This paper will review the various triggers and consequences of TSLP release in asthmatic airways. [ABSTRACT FROM AUTHOR]

The genetically modified (GM) maize DBN9936 with a biosafety certificate will soon undergo commercial application. To monitor the safety of DBN9936 maize, three genomic DNA (gDNA) reference materials (RMs) (DBN9936a, DBN9936b, and DBN9936c) were prepared with nominal copy number ratios of 100%, 3%, and 1% for the DBN9936 event, respectively. DBN9936a was prepared from the leaf tissue gDNA of DBN9936 homozygotes, while DBN9936b and DBN9936c were prepared by the quantitative mixing of gDNA from the leaf tissues of DBN9936 homozygotes and non-GM counterparts. Validated DBN9936/zSSIIb duplex droplet digital PCR was demonstrated to be an accurate reference method for conducting homogeneity study, stability study, and collaborative characterization. The minimum intake for one measurement was determined to be 2 μL, and the gDNA RMs were stable during transport at 37 °C for 14 days and storage at −20 °C for 18 months. Each gDNA RM was certified for three property values: DBN9936 event copy number concentration, zSSIIb reference gene copy number concentration, and DBN9936/zSSIIb copy number ratio. The measurement uncertainty of the certified values took the uncertainty components related to possible inhomogeneity, instability, and characterization into account. This batch of gDNA RMs can be used for calibration and quality control when quantifying DBN9936 events. [ABSTRACT FROM AUTHOR]

Among the cucurbit-infecting viruses, watermelon mosaic virus (WMV) and zucchini yellow mosaic virus (ZYMV) (Potyvirus: Potyviridae) are responsible for severe symptoms on cucumber, melon, watermelon, and zucchini cultivations worldwide. In this study, reverse transcription real-time PCR (real-time RT-PCR) and droplet-digital PCR (RT-ddPCR) assays targeting the coat protein (CP) genes of WMV and ZYMV were developed and validated according to the international standards of plant pest diagnosis (EPPO PM 7/98 (5)). First, the diagnostic performance of WMV-CP and ZYMV-CP real-time RT-PCRs was evaluated, and the assays displayed an analytical sensitivity of 10−5 and 10−3, respectively. The tests also showed an optimal repeatability, reproducibility and analytical specificity, and were reliable for the virus detection in naturally infected samples and across a wide range of cucurbit hosts. Based on these results, the real-time RT-PCR reactions were adapted to set up RT-ddPCR assays. These were the first RT-ddPCR assays aiming at the detection and quantification of WMV and ZYMV and showed a high sensitivity, being able to detect until 9 and 8 copies/µL of WMV or ZYMV, respectively. The RT-ddPCRs allowed the direct estimation of the virus concentrations and opened to a broad range of applications in disease management, such as the evaluation of partial resistance in breeding processes, identification of antagonistic/synergistic events, and studies on the implementation of natural compounds in the integrated management strategies. [ABSTRACT FROM AUTHOR]

Wastewater-based surveillance can be used as a complementary method to other SARS-CoV-2 surveillance systems. It allows the emergence and spread of infections and SARS-CoV-2 variants to be monitored in time and place. This study presents an RT-ddPCR method that targets the T19I amino acid mutation in the spike protein of the SARS-CoV-2 genomes, which is specific to the BA.2 variant (omicron). The T19I assay was evaluated both in silico and in vitro for its inclusivity, sensitivity, and specificity. Moreover, wastewater samples were used as a proof of concept to monitor and quantify the emergence of the BA.2 variant from January until May 2022 in the Brussels-Capital Region which covers a population of more than 1.2 million inhabitants. The in silico analysis showed that more than 99% of the BA.2 genomes could be characterized using the T19I assay. Subsequently, the sensitivity and specificity of the T19I assay were successfully experimentally evaluated. Thanks to our specific method design, the positive signal from the mutant probe and wild-type probe of the T19I assay was measured and the proportion of genomes with the T19I mutation, characteristic of the BA.2 mutant, compared to the entire SARS-CoV-2 population was calculated. The applicability of the proposed RT-ddPCR method was evaluated to monitor and quantify the emergence of the BA.2 variant over time. To validate this assay as a proof of concept, the measurement of the proportion of a specific circulating variant with genomes containing the T19I mutation in comparison to the total viral population was carried out in wastewater samples from wastewater treatment plants in the Brussels-Capital Region in the winter and spring of 2022. This emergence and proportional increase in BA.2 genomes correspond to what was observed in the surveillance using respiratory samples; however, the emergence was observed slightly earlier, which suggests that wastewater sampling could be an early warning system and could be an interesting alternative to extensive human testing. [ABSTRACT FROM AUTHOR]

Bangladesh's aquaculture sector has contributed progressively to the nation's economy over the years, but the COVID-19 pandemic has impeded fish farmers' access to markets, reduced their production and sales capacity, resulted in lower income, and increased food security vulnerability. This study assesses how COVID-19 affects smallholder fish farmers and their response strategies by employing data collected from 250 fish farmers and traders from intensive fish-growing areas of Bangladesh. The results reveal that most farmers experienced difficulty obtaining inputs, and the price of those inputs skyrocketed during the COVID-19 period, resulting in several months of decreased production and operations. As a result of COVID-19, farm gate prices for silver carp, ruhu, common carp, grass carp, and tilapia fish dropped by 25%, 23%, 23%, 22%, 23%, and 40%, respectively. On the other hand, fish feed prices were found to increase significantly. Reduced income from fish farming and other sources has triggered a significant drop in capital for farming operations and production capacity improvement, leading to food insecurity. The most common coping strategies include reduced buying from the market (vegetables, fruits, meat, milk, etc.), relying on less expensive or less preferred food, purchasing food on credit, and selling assets. Notably, due to COVID-19, a new mode of marketing has evolved as an adaptation strategy in the fish marketing system, such as the use of the mobile phone (18%) and Facebook/internet to sell fish directly to the customer (16%). The sector requires short-term financial assistance to assist fish actors with production and marketing challenges. [ABSTRACT FROM AUTHOR]

Cytokine receptor-like factor 2 B-cell acute lymphoblastic leukemia (CRLF2 B-ALL) is a high-risk subtype characterized by CRLF2 overexpression with poor survival rates in children and adults. CRLF2 and interleukin-7 receptor alpha (IL-7Rα) form a receptor for the cytokine thymic stromal lymphopoietin (TSLP), which induces JAK/STAT and PI3K/AKT/mTOR pathway signals. Previous studies from our group showed that low TSLP doses increased STAT5, AKT, and S6 phosphorylation and contributed to CRLF2 B-ALL cell survival. Here we investigated the role of TSLP in the survival and proliferation of CRLF2 B-ALL cells in vitro and in vivo. We hypothesized that high doses of TSLP increase CRLF2 signals and contribute to increased proliferation of CRLF2 B-ALL cells in vitro and in vivo. Interestingly, we observed the opposite effect. Specifically, high doses of TSLP induced apoptosis in human CRLF2 B-ALL cell lines in vitro, prevented engraftment of CRLF2 B-ALL cells, and prolonged the survival of +TSLP patient-derived-xenograft mice. Mechanistically, we showed that high doses of TSLP induced loss of its receptor and loss of CRLF2 signals in vitro. These results suggest that high doses of TSLP could be further investigated as a potential therapy for the treatment of CRLF2 B-ALL. [ABSTRACT FROM AUTHOR]

Salmonella is a crucial food‐borne pathogen causing food poisoning, leading to severe public health events. Here, we developed a technique by integrating recombinase polymerase amplification with CRISPR‐LbCas12a and employing two targets with engineered crRNA for detection of Salmonella (RPA‐LbCas12a‐TTECDS). Our findings revealed that this novel method rapidly detects trace Salmonella in food through fluorescence intensity and provides a template for other food‐borne pathogen detection methods. Further, crRNA was optimized to increase detection sensitivity. Double targets were used to enhance the detection accuracy, reaching the level of qPCR, which was superior to fluorescent RPA. The RPA‐LbCas12a‐TTECDS system specifically detected Salmonella levels as low as 50 CFU per ml at 37°C in 1 h. In summary, a simple, rapid, sensitive and high accuracy detection technique based on CRISPR‐Cas12a was created for Salmonella detection without complicated equipment. Significance and Impact of the Study: The emergence of CRISPR‐Cas12a provides a new technique for detection methods. In this study, we developed a rapid, sensitive, and high accuracy RPA‐LbCas12a‐TTECDS based toolbox for Salmonella detection. This method is easy to operate with a completion span of 50 min by a fluorescence detector. The simple and quick‐to‐read method is potentially useful as point‐of‐care testing for Salmonella. [ABSTRACT FROM AUTHOR]

mRNA decay is an important process in post-transcriptional regulation; in addition, it plays a crucial role in plant development and response to stress. The development of new tools to quantify mRNA decay intermediates is thus important to better characterize the dynamic of mRNA decay in various conditions. Here, we applied droplet digital PCR (ddPCR), a recent and precise PCR technology, to determine mRNA half-life in Arabidopsis seedlings. We demonstrated that ddPCR can correctly assess mRNA half-life from a wide variety of transcripts in a reproducible manner. We also demonstrated that thanks to multiplexing mRNA, the half-life of multiple transcripts can be followed in the same reaction. As ddPCR allows precise quantification, we proposed that this approach is highly suitable when a low amount of RNA is available; for the detection of many targets or for the analysis of lowly expressed transcripts. [ABSTRACT FROM AUTHOR]

Droplet digital polymerase chain reaction (ddPCR) is one of the newest and most promising tools providing absolute quantification of target DNA molecules. Despite its emerging applications in microorganisms, few studies reported its use for detecting lactic acid bacteria. This study evaluated the applicability of a ddPCR assay targeting molecular genes obtained from in silico analysis for detecting Lactiplantibacillus plantarum subsp. plantarum, a bacterium mainly used as a starter or responsible for fermentation in food. The performance characteristics of a ddPCR were compared to those of a quantitative real-time PCR (qPCR). To compare the linearity and sensitivity of a qPCR and ddPCR, the calibration curve for a qPCR and the regression curve for a ddPCR were obtained using genomic DNA [102–108 colony-forming units (CFU)/mL] extracted from a pure culture and spiked food sample. Both the qPCR and ddPCR assays exhibited good linearity with a high coefficient of determination in the pure culture and spiked food sample (R2 ≥ 0.996). The ddPCR showed a 10-fold lower limit of detection, suggesting that a ddPCR is more sensitive than a qPCR. However, a ddPCR has limitations in the absolute quantitation of high bacterial concentrations (>106 CFU/mL). In conclusion, a ddPCR can be a reliable method for detecting and quantifying lactic acid bacteria in food. [ABSTRACT FROM AUTHOR]

The shrimp industry in the Philippines plays a vital role in the local and national economy through exports to markets abroad such as South Korea, Japan, the USA, and others. In this study, we aimed to describe the various cultural and operational characteristics of smallholder and commercial shrimp (P. vannamei) farms in the Davao region. We also evaluated the current risks and challenges faced by the shrimp farmers. A semi-structured questionnaire that focused on shrimp farmers and operators in the region was used to collect data from N = 41 farmers and operators. The results showed that respondents who were engaged in smallholder farming activities had an average yield of 10 tons/ha. The commercial farms that operate intensively had an average yield of 24 tons/ha. Most smallholder operators used electric generator machines to conduct aeration in their farms using paddlewheels and blowers. More paddlewheels and blowers were employed per pond in the commercial farms compared to smallholder farms. Generally, the income of a farm was related to their yield or the number of fries rather than social factors or their size. In terms of input costs, feeds were found to have the highest cost, followed by the fry, fuel, labor, and others (fertilizers and water treatment chemicals). Most of the farmers mentioned that their shrimp are affected by diseases such as white spot syndrome (60%), black gills (35%), and red tail (5%). They perceived that the main contamination comes from the water source (31%). The main threats mentioned were declining shrimp prices in the market, source of fry, water disposal, overstocking, and water quality. This study shows that small-holding fish farmers should be supported by the government so that they can make use of the more advanced technology employed by commercial shrimp farmers in order to increase their economic productivity and lower their environmental footprint. [ABSTRACT FROM AUTHOR]

The impacts of climate change on shrimp aquaculture can vary widely and can have environmental and socioeconomic consequences. This study assessed the vulnerability to climate change impacts of selected small-scale shrimp farms of Penaeus vannamei and shrimpfish market vendors in the Davao region, the Philippines, using a modified Fisheries Vulnerability Assessment Tool (FishVool). Shrimp farmers and vendors were interviewed using two separate semi-structured questionnaires. A total of thirty-nine (n = 39) shrimp farmers and forty-eight (n = 48) market vendors from various market areas within the region were interviewed. Data regarding exposure (E), sensitivity (S), and adaptive capacity (AC) were collected following the FishVool parameters with modifications. Results revealed that the overall climate change vulnerability of the shrimp farmers was medium (M), where both exposure and adaptive capacity were low (L) while sensitivity was medium (M). In addition, the shrimp market vulnerability of the various sites examined revealed medium (M) scores for markets in Pantukan, Mabini, Tagum, Maco, Lupon, Davao City, and Digos, and high (H) vulnerability scores for the markets in Panabo and Sta Cruz. Overall, the study provided a better understanding of shrimp farming in relation to climate change impacts and vulnerability and provided information for future shrimp farm management, marketing, and climate change adaptation in the region. [ABSTRACT FROM AUTHOR]

The aim of this study was to develop homogeneous and stable plasmid DNA reference materials for detecting the mechanisms of resistance to quinolones and fluoroquinolones in foodborne pathogens. The DNA fragments of 11 target genes associated with quinolone and fluoroquinolone resistance were artificially synthesized, inserted into plasmid vectors, and transferred into recipient cells. PCR and sequencing of DNA were performed to assess the genetic stability of the target DNA in recombinant Escherichia coli DH5α cells during subculturing for 15 generations. The limit of detection (LOD) of the target DNA was determined using PCR and real-time qualitative PCR (qPCR). The homogeneity and storage stability of plasmid DNA reference materials were evaluated in terms of plasmid DNA quantity, PCR-measured gene expression, and qPCR threshold cycle. All 11 target DNAs were successfully synthesized and inserted into vectors to obtain recombinant plasmids. No nucleotide mutations were identified in the target DNA being stably inherited and detectable in the corresponding plasmids during subculturing of recombinant strains. When the target DNA was assessed using PCR and qPCR, the LOD was ≤1.77 × 105 and 3.26 × 104 copies/μL, respectively. Further, when the reference materials were stored at 37 °C for 13 days, 4 °C for 90 days, and −20 °C for 300 days, each target DNA was detectable by PCR, and no mutations were found. Although the threshold cycle values of qPCR varied with storage time, they were above the LOD, and no significant differences were found in the quantity of each plasmid DNA at different timepoints. Further, the homogeneity and stability of the materials were highly consistent with the requirements of standard reference materials. To summarize, considering that our plasmid DNA reference materials conformed to standard requirements, they can be used to detect the mechanisms of quinolone and fluoroquinolone resistance in foodborne pathogens. [ABSTRACT FROM AUTHOR]

Cancer is a major health problem that poses a great challenge to health care systems worldwide. Tools for cancer treatment have rapidly advanced in recent years, resulting in therapeutic strategies which are alternative and complementary to conventional treatment. To identify the cell surface receptors used by a tumor cell-adapted rotavirus and the cell death markers induced by its infection, we use Wt1-5, a rotavirus isolate recently adapted to tumor cells, to infect the human acute lymphoblastic leukemia cell line, Reh. The expression of cell surface receptors used by Wt1-5 was determined using flow cytometry and an antibody blocking assay to test for their implication in virus infection. Viral antigens and cell death markers induced by rotavirus infection were followed by flow cytometric analysis. The present study showed that rotavirus Wt1-5 was able to use cell surface proteins such as heat shock proteins (HSPs) 90, 70, 60 and 40, Hsc70, PDI and integrin β3. Rotavirus Wt1-5 induced cytotoxic effects including changes in cell membrane permeability, alteration of mitochondrial membrane potential, DNA fragmentation and activation of cell death signaling. Wt1-5 deserves to be further studied as a candidate oncolytic agent due to its ability to induce apoptosis in lymphoblastic leukemia-derived cells. [ABSTRACT FROM AUTHOR]