Determination of anti-cancer effects of Nigella sativa seed oil on MCF7 breast and AGS gastric cancer cells.

BACKGROUND: This study aimed to investigate the cytotoxic, apoptotic, invasion, metastasis, and heat shock proteins (HSPs) effects of N. sativa oil on breast and gastric cancer cells. METHODS: We assessed the cytotoxic and apoptotic effects of various concentrations of N. sativa oil (10-50-100-200 µg/mL) on MCF7 breast cancer and AGS, an adenocarcinoma of the gastric cell line, at 24, 48 and 72 h using the MTT test. Additionally, the expression of the Caspase-3, BCL2/Bax, MMP2-9 and HSP60-70 gene was examined using RT-PCR in cell lines treating with N. sativa. RESULTS: The MTT experiments demonstrate that N. sativa has a time and dose-dependent inhibitory effect on the proliferation of MCF7 and AGS cancer cells. The vitality rates of MCF7 and AGS cells treated with N. sativa were 77.04-67.50% at 24 h, 65.28-39.14% at 48 h, and 48.95-32.31% at 72 h. The doses of 100 and 200 µg/mL were shown to be the most effective on both cancer cells. RT-PCR analysis revealed that N. sativa oil extract increased caspase-3 levels in both cell lines at higher concentrations and suppressed BCL2/Bax levels. Exposure of MCF7 and AGS cell lines to N. sativa caused a significant decrease in the expression of MMP2-9 and HSP60-70 genes over time, particularly at a dosage of 200 µg/mL compared to the control group (p < 0.05). CONCLUSIONS: Our findings indicate that N. sativa oil has a dose-dependent effect on cytotoxicity and the expression of apoptotic, heat shock proteins, and matrix metalloproteinases genes in breast and gastric cancer.

Allergenicity and structural properties of new Cor a 1 isoallergens from hazel identified in different plant tissues.

The hazel allergen Cor a 1 is a PR-10 protein, closely related to the major birch pollen allergen Bet v 1. Hazel allergies are caused by cross-reactive IgE antibodies originally directed against Bet v 1. Despite the importance of PR-10 proteins in allergy development, their function and localization in the plant remain largely elusive. Therefore, the presence of Cor a 1 mRNA and proteins was investigated in different tissues, i.e., the female flower, immature and mature nuts, catkins, and pollen. Four yet unknown Cor a 1 isoallergens, i.e., Cor a 1.0501-1.0801, and one new Cor a 1.03 variant were discovered and characterized. Depending on the isoallergen, the occurrence and level of mRNA expression varied in different tissues, suggesting different functions. Interestingly, Cor a 1.04 previously thought to be only present in nuts, was also detected in catkins and pollen. The corresponding Cor a 1 genes were expressed in Escherichia coli. The purified proteins were analysed by CD and NMR spectroscopy. Immunoblots and ELISAs to determine their allergenic potential showed that the new proteins reacted positively with sera from patients allergic to birch, hazel and elder pollen and were recognized as novel isoallergens/variants by the WHO/IUIS Allergen Nomenclature Sub-Committee.

Green synthesis of biogenic selenium nanoparticles functionalized with ginger dietary extract targeting virulence factor and biofilm formation in Candida albicans.

To treat the systemic infections caused by Candida albicans (C. albicans), various drugs have been used, however, infections still persisted due to virulence factors and increasing antifungal resistance. As a solution to this problem, we synthesized selenium nanoparticles (SeNPs) by using Bacillus cereus bacteria. This is the first study to report a higher (70 %) reduction of selenite ions into SeNPs in under 6 h. The as-synthesized, biogenic SeNPs were used to deliver bioactive constituents of aqueous extract of ginger for inhibiting the growth and biofilm (virulence factors) in C. albicans. UV-visible spectroscopy revealed a characteristic absorption at 280 nm, and Raman spectroscopy showed a characteristic peak shift at 253 cm-1 for the biogenic SeNPs. The synthesized SeNPs are spherical with 240-250 nm in size as determined by electron microscopy. Fourier transform infrared spectroscopy confirmed the functionalization of antifungal constituents of ginger over the SeNPs (formation of Ginger@SeNPs nanoconjugates). In contrast to biogenic SeNPs, nanoconjugates were active against C. albicans for inhibiting growth and biofilm formation. In order to reveal antifungal mechanism of nanoconjugates', real-time polymerase chain reaction (RT-PCR) analysis was performed, according to RT-PCR analysis, the nanoconjugates target virulence genes involved in C. albicans hyphae and biofilm formation. Nanoconjugates inhibited 25 % growth of human embryonic kidney (HEK) 293 cell line, indicating moderate cytotoxicity of active nanoconjugates in an in-vitro cytotoxicity study. Therefore, biogenic SeNPs conjugated with ginger dietary extract may be a potential antifungal agent and drug carrier for inhibiting C. albicans growth and biofilm formation.

Effect of photobiomodulation at 830 nm on gene expression correlated with JAK/STAT signalling in wounded and diabetic wounded fibroblasts in vitro.

Treatment of chronic diabetic wounds is an ongoing socio-economic challenge. Dysregulated signalling pathways characterise cells from chronic diabetic wounds. Photobiomodulation (PBM) stimulates healing by eliciting photochemical effects that affect gene regulation. JAK/STAT signalling is a primary signal transduction pathway involved in wound healing. This in vitro study aimed to determine if PBM at 830 nm and a fluence of 5 J/cm2 regulates genes related to JAK/STAT signalling in wounded and diabetic wounded fibroblast cells. A continuous wave diode laser (12.53 mW/cm2 ) was used to irradiate cells. Forty-eight hours post-PBM, RT-qPCR was used to analyse 84 genes related to JAK/STAT signalling. Five genes were upregulated and four downregulated in wounded cell models, while six genes were downregulated in diabetic wounded models. The results show drastic gene expression differences between wounded and diabetic wounded cell models in response to PBM using 830 nm.

Diagnostic challenges and forensic implications in a case of infantile fatal myocarditis.

We present the case of a 23-month-old child who died less than 24 h after the onset of cardiac symptoms, despite being admitted to the hospital 72 h earlier. Autopsy revealed no significant macroscopic changes, and histologic examination revealed focal lymphocytic myocarditis with myocyte disruption, diffuse alveolar damage in the exudative phase, and generalized lymphocytic immune activation in other organs. Ante-mortem and post-mortem microbiological exams did not clearly prove a causative role of infectious agents. The peculiarity of this case was characterized by the contrast between the severe clinical features and the mild cardiac histological findings. This discrepancy, coupled with the suspicion of a viral causative role based on both ante-mortem and post-mortem microbiological examinations, presented significant challenges in reaching an etiological diagnosis. This case also confirms that the diagnosis of myocarditis in children cannot be made solely on the basis of histological cut-offs or microbiological results. Using abductive reasoning, various diagnostic hypotheses were formulated and evaluated to arrive at the final diagnosis of fatal myocarditis of viral or post-viral origin. Data from post-mortem examination are often the only source of information that is available to the experts, especially in cases of sudden infant death syndrome. In such cases, the forensic pathologists should accurately evaluate findings that may appear to indicate a different etiology, and, in the absence of clinical or radiological data, interpret post-mortem data in a logically correct manner. The autopsy is the first essential step to evaluate the cause of death and must be integrated with the results of ante- and post-mortem diagnostic tests in a holistic approach, which is crucial to allow forensic pathologists to provide an appropriate and relevant opinion.

Effectiveness of ayurvedic formulation, NAOQ19 along with standard care in the treatment of mild-moderate COVID-19 patients: A double blind, randomized, placebo-controlled, multicentric trial.

BACKGROUND: Medicines in indigenous systems such as Ayurveda have strong antimicrobial activity but double-blind randomized control trials are infrequent in this system of medicine. The efficacy of a new ayurvedic formulation was evaluated during the pandemic. METHODS: 150 mild-moderate COVID-19 patients were enrolled and randomized in 1:1 to NAOQ19 and placebo group. RT-PCR was done on Day 3, 5 and 7. CBC, CRP, LFT, and KFT were assessed at baseline and exit. Duration of hospital stay was noted and clinical assessment was also performed. RESULT: The results demonstrated more people turning RT-PCR negative in the NAOQ19 group compared to the placebo group on day 3 (p-value = 0.033). The mean time duration to turn RT-PCR negative was significantly lower in the NAOQ19 group (4.6 days) compared to placebo group (5.2 days) (p-value = 0.018). There was significant reduction in hospital stay among patients in the NAOQ19 arm who were discharged earlier (5.6 days) compared to placebo group (6.4 days) (p-value = 0.046). Patients in NAOQ19 arm did not show any adverse life-threatening events. CONCLUSION: The ayurvedic preparation given along with standard of care therapy reduced the duration of hospital stay and there was earlier conversion to RT-PCR negative.The integrated approach can help to reduce patient workload in the hospitals as well as limit the transmission of the virus in the community. STUDY REGISTRATION: CTRI/2021/05/033790.

Development and application of reverse transcription-loop mediated isothermal amplification assay for sensitive detection of groundnut bud necrosis virus infecting potato.

Reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detection of groundnut bud necrosis virus (GBNV) causing potato stem necrosis disease. The isothermal temperatures, reaction periods and concentrations of reaction mixture were optimized where, the assay worked well at 65 °C for 50 min, 6 U of WarmStart Bst 2.0 DNA polymerase, 1.4 mM dNTPs and 2.0 mM MgSO4. The optimized assay proved to be specific to GBNV with no cross reactivity to other viruses infecting potato in India. The specificity of RT-LAMP assay was found to be 100 fold more sensitive than that of RT-PCR. The developed assay was applied for the detection of GBNV from 80 potato leaf samples where 24 samples were found infected which was confirmed by RT-PCR. It was concluded that the RT-LAMP assay developed for detection of GBNV was specific, sensitive and suitable for its use in virus indexing under potato seed production programme.

Identification of sex-specific markers based on gene expression in adult Colorado potato beetles.

The Colorado potato beetle, Leptinotarsa decemlineata, (Coleoptera: Chrysomelidae) is an economically important pest insect of potatoes. Understanding how the mechanisms driving its invasiveness vary between sexes will be critical for developing modern control methods. However, the currently available methods for sexing adult Colorado potato beetles are either inefficient or unsuitable for projects that require RNA as an input, like those measuring gene expression. Therefore, the development of simple molecular tools that are tailored to these studies is important. In this study, we used publicly available RNA-seq data to select 5 candidate genes for sex-specific markers in adult Colorado potato beetles. We confirmed that our 5 marker candidates exhibit a sex-specific expression pattern and can be used as PCR markers for sex determination. This method of sex detection will allow researchers to distinguish the sex of the individual with a simple PCR reaction using cDNA as the template and assign sex to RNA-seq samples post hoc.

First report of Watermelon silver mottle orthotospovirus infecting Siraitia grosvenorii in China.

Siraitia grosvenorii, known as "Luohanguo or monk fruit", is a perennial vine belonging to the family Cucurbitaceae. It is cultivated for its fruits, which are used as a Chinese traditional medicine to treat throat, lung and intestine ailments, or as raw material to extract sweet cucurbitane-glycosides as sugar substitute sweeteners (Chen et al., 2007). The production of S. grosvenorii is limited by viral diseases especially cucumber green mottle mosaic virus (CGMMV), papaya ringspot virus (PRSV), watermelon mosaic virus, and zucchini yellow mosaic virus (Liao et al., 2005; Xie et al., 2020). In 2022, virus-like disease consisting of leaf mottling, crinkling, and ringspot was observed on S. grosvenorii plants grown in an insect-proof greenhouse in Guilin City, Guangxi Province, China, with an incidence rate of ~17%. High-throughput sequencing (HTS) was applied to identify potential viruses in the diseased plants. Briefly, total RNA was extracted from a pool of 28 leaf samples (with or without symptoms) of S. grosvenorii using Trizol reagent according to manufacturer's instructions (Invitrogen, U.S.A.). The rRNA was depleted (Epicentre Ribo-zero™ rRNA Removal Kit, Epicentre, U.S.A.), before steps of cDNA library construction (NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®, NEB, U.S.A.), and sequencing (Hiseq 4000 platform, Illumina, U.S.A.). The subsequent bioinformatics analyses were performed according to Liu et al. (2021). HTS of the sample and raw reads processing resulted in 8.4 Gb clean data. The clean reads (150 bp) were de novo assembled into 87,414 contigs (≥200 bp), using CLC Genomics Workbench 21 (Qiagen, Germany). The contigs were annotated by local BLASTX, resulting in matches to CGMMV, PRSV, and watermelon silver mottle virus (WSMoV). Three contigs of 6,557 bp, 4,950 bp, and 3,594 bp were most identical to L (GenBank accession no. JX177647), M (MW051789), and S (KM242056) segments of WSMoV. The complete genome sequences corresponding to the contigs derived from the sample (designated as GL-1 variant of WSMoV, OQ401466-OQ401468) were obtained by reads mapping to segments of these isolates. The reads coverage was ≥99.75% in each RNA segment and the depth of the coverage was in a range of 74-285. To detect the presence of GL-1 in S. grosvenorii plants, three primer pairs D7280F/D7382R (5'-TGATAGCCTGATGAACACCA/5'-TGTCTCTAAACCTTCTACCGC, Tm = 55℃, product size 172 bp), D4512F/D4703R (5'-GCATTGAACTCGCTCACAC/5'-AGTAGACGACCCTGAAGACCT, Tm = 55℃, 192 bp), and D109F/D451R (5'-TTATGGCACAAGAGACAACAGAG/5'-GGGCGTTATGTTCAGTATATTGG, Tm = 56℃, 342 bp) were designed in the L, M, and S segments, respectively. Fresh symptomatic and asymptomatic leaf tissues (n=38) were collected from three fields and their extracted nucleic acids were individually tested with the primers designed by two-steps RT-PCR using TaKaRa RNA PCR kit Ver.3.0 (Takara, Japan). Expected amplicons were obtained in symptomatic samples (n=7) showing mottling, crinkling, and chlorosis. Other samples (n=31) with or without symptoms were negative to WSMoV infection. The amplicons were sequenced, and the sequences obtained shared >99% nt identities with the corresponding GL-1 sequences in GenBank. This is the first report of WSMoV on S. grosvenorii, which provides the basic information for virus disease management.

Identification and Expression of the CorA/MRS2/ALR Type Magnesium Transporters in Tomato.

Magnesium (Mg2+) is the most abundant divalent ion in plants, participating in numerous metabolic processes in growth and development. CorA/MRS2/ALR type Mg2+ transporters are essential for maintaining Mg2+ homeostasis in plants. However, the candidate protein and its potential functions in the tomato plant have not been fully understood. In this study, we identified seven MGT genes (SlMRS2) in tomato based on sequence similarity, domain analysis, conserved motif identification, and structure prediction. Two SlMRS2 genes were analyzed in the bacterial strain MM281, and a functional complementary assay demonstrated their high-affinity transport of Mg2+. Quantitative real-time PCR analysis revealed that the expressions of these Mg2+ transporters were down-regulated in leaves under Mg2+ limitation, with a greater impact on lower and middle leaves compared to young leaves. Conversely, under Mg2+ toxicity, several genes were up-regulated in leaves with a circadian rhythm. Our findings indicate that members of the SlMRS2 family function as Mg2+ transporters and lay the groundwork for further analysis of their distinct functions in tomato.

Investigating the Effects of Alltech Crop Science (ACS) Products on Plant Defence against Root-Knot Nematode Infestation.

Two formulations of Alltech Crop Science products (ACS), a proprietary blend of fermentation products and plant extracts with micronutrients (ACS5075), and a microbial based product (ACS3048), were tested to understand (1) their impact on the tomato plant immune response and (2) whether they are priming a resistance response in plants against root knot nematodes (RKN). Research findings reported previously indicate that tomato plants pre-treated with ACS5075 and ACS3048 were found less sensitive to Meloidogyne javanica infection. In the current study, the expression of six defence-related genes (PR-1, PR-3, PR-5T, ACO, CAT and JERF 3), relative to a housekeeping gene, were monitored via RT-PCR. Results suggest that the treatment with ACS5075 enhanced ACO and PR-1 gene expression levels, both post- treatment and post-infection with M. javanica. Reduced M. javanica infestation that was reported in the previous study could be attributed to the increased expression of these genes in the ACS5075-treated plants. Tomato plants treated with ACS3048, but without RKN infection, also demonstrated higher levels of ACO and PR-1 gene expression. Subsequently, 2D-gel electrophoresis was performed to study the differential protein expression in leaf tissues of treated tomato plants in an effort to elucidate a possible mechanism of action for these products. Protein spot 1 was identified as 'disease resistance protein RPP13-like', protein spot 2 as 'phosphatidylinositol 4-phosphate 5-kinase 2', spot 3 as 'protein SABRE like' and protein spot 4 as 'uncharacterized protein'. Overall research findings indicate that the ACS products could be used as plant immunity-boosting agents, as they play a significant role in the expression of certain genes and proteins associated with plant defence.

Analyses of Elaeocarpus sphaericus Extract for Antioxidant, Antiproliferative and Gene Repression Activities against HIF-1α and VEGF.

The study presents antioxidant, phytochemical, anti-proliferative, and gene repression activities against Hypoxia-inducible factor (HIF-1) alpha and Vascular endothelial growth factor (VEGF) of Elaeocarpus sphaericus extract. Elaeocarpus sphaericus dried and crushed plant leaves were extracted using water and methanol by ASE (Accelerated Solvent Extraction) method. Total phenolic content (TPC) and total flavonoid content (TFC) were used to measure the extracts' phytochemical activity (TFC). Antioxidant potential of the extracts was measured through DPPH, ABTS, FRAP, and TRP. Methanolic extract of the leaves of E. sphaericus has shown a higher amount of TPC (94.666±4.040 mg/gm GAE) and TFC value (172.33±3.21 mg/gm RE). The antioxidant properties of extracts in the yeast model (Drug Rescue assay) showed promising results. Ascorbic acid, gallic acid, hesperidin, and quercetin were found in the aqueous and methanolic extracts of E. sphaericus at varying amounts, according to a densiometric chromatogram generated by HPTLC analysis. Methanolic extract of E. sphaericus (10 mg/ml) has shown good antimicrobial potential against all bacterial strains used in the study except E. coli. The anticancer activity of the extract in HeLa cell lines ranged from 77.94±1.03 % to 66.85±1.95 %, while it ranged from 52.83±2.57 % to 5.44 % in Vero cell lines at varying concentration (1000 µg/ml-31.2 µg/ml). A promising effect of extract was observed on the expression activity of HIF-1 and VEGF gene through RT-PCR assay.

Efficacy of the Unani Regimen as an Add-On to Standard Treatment in Hospitalised RT-PCR-Confirmed Mild to Moderate COVID-19 Patients: An Open-Label Randomized Controlled Trial.

Background The coronavirus disease 2019 (COVID-19) pandemic resulted in mortality and morbidity worldwide. Many treatment modalities have been experimented with limited success. Therefore, the traditional system of medicine needs to be explored. Objective To evaluate the benefits of Unani regimensTiryaq-e-Arba and Unani Joshanda, as adjuvant therapy, were compared to standard treatment alone among reverse transcription polymerase chain reaction (RT-PCR)-confirmed mild to moderate COVID-19 cases. Materials and methods An open-label, double-arm, randomized, controlled interventional clinical study was conducted among 90 RT-PCR-confirmed mild to moderate COVID-19 inpatients admitted to a tertiary care hospital in New Delhi, India. Participants who fulfilled the criteria for inclusion were randomly assigned to two arms, with 43 subjects allocated to the Unani add-on arm and 47 subjects to the control arm receiving standard treatment alone. Results Clinical recovery was achieved in all patients of the Unani arm, while in the control arm, three (6.4%) patients deteriorated and had to be shifted to ICU following admission. In the intervention arm, a shorter duration of hospitalization was observed (mean 5.95 days {SD = 1.99}) than in the control arm (mean 7.62 days {SD, 4.06}); which was a statistically significant difference (p-value 0.017). The majority of the patients recovered within 10 days in the Unani add-on arm. The number of days taken for the reduction of symptoms was significantly less in the intervention arm (mean 5.14 days {SD, 2.39}) as compared with standard treatment (mean 6.53 days {SD, 3.06}) (p < 0.02). Renal and liver safety parameters were within the normal limits in both arms and no serious adverse event was reported. Conclusion Adding Unani formulations to standard treatment significantly reduced the duration of hospital stay and showed early recovery in COVID-19 patients compared with the control arm. It may be concluded that the synergistic effect of the Unani add-on with standard treatment gave more promising results in mild to moderate COVID-19 patients.

Impact of Heartfulness meditation practice on anxiety, perceived stress, well-being, and telomere length.

Objective: Exhaustion, stress, and burnout have all been found to be reduced using techniques like yoga and meditation. This study was carried out to check the effectiveness of Heartfulness practice (a form of meditation) on certain psychological and genetic variables. Methods: A total of 100 healthy individuals (aged 18-24) were recruited and randomized into two groups-Heartfulness intervention and control group. The intervention was carried out for 03 months. Participants from both groups were analysed for their cortisol levels and telomere length before and after the intervention. Psychometric measures of anxiety, perceived stress, well-being and mindfulness were carried out using Beck Anxiety Inventory (BAI), Perceived Stress Scale (PSS), WHO-Well-being Index (WHO-WBI) and Five Facet Mindfulness Questionnaire (FFMQ). Results: The cortisol levels in the meditators group significantly decreased (p < 0.001) after the intervention as compared to the non-meditators group, whereas, the telomere length increased in the mediators group. This increase was not significant (p > 0.05). Anxiety and perceived stress also decreased post intervention, and well-being as well as mindfulness increased, as assessed by the questionnaire tools, although the decrease in perceived stress was statistically insignificant (p > 0.05). A negative correlation was observed between telomere length and cortisol (stress biomarker), whereas a positive correlation was found between telomere length and well-being. Conclusion: Our data provide evidence that Heartfulness meditation practice can improve our mental health. Additionally, telomere length is shown to be affected by cortisol levels, and this meditation practice can also help to increase telomere length, and thereby slow down cellular aging. However, future studies with larger sample size are required to confirm our observations.

First Detection of Potato Spindle Tuber Viroid in Natural Isolates of Potato Blight Agent Phytophthora infestans.

Phytophthora infestans is the oomycete that causes potato blight, an important disease. The potato spindle tuber viroid (PSTVd) is a dangerous pathogen of many plants, including potato. We have previously shown that PSTVd can be transmitted from infected potato plants into the Ph. infestans mycelium, replicated within the mycelium, and then transmitted to other potato plants upon their infection with Ph. infestans in laboratory conditions. The objective of this work was to check the hypothesis that PSTVd transmission, preservation, and replication in Ph. infestans are possible to occur in natural conditions during long-term coevolution of the host and pathogen in the Solanum spp.-Ph. infestans system. A screening test for PSTVd was performed in 111 natural Ph. infestans isolates obtained from potato plants, which represented various cultivars, had signs of potato blight, and were collected from industrial potato fields of the Moscow, Vologda, and Bryansk regions and breeding and variety test plots of the St. Petersburg and Moscow regions in 2020 and 2022. Using RT-PCR with PSTVd-specific primers, 42 Ph. infestans isolates collected in 2020 were tested after five passages and 69 Ph. infestans isolates collected in 2022, after a single passage on rye agar. Diagnostic amplicons were detected in 8 and 50 isolates, respectively. Some of the amplicons were visually assessed as minor amplification products, apparently resulting from nonspecific priming on a host Ph. infestans gene, which codes for a hypothetical protein-coding mRNA in Ph. infestans and other oomycetes. Eight amplicons were sequenced to verify the PSTVd presence in Ph. infestans isolates. Three amplicons corresponded to the complete PSTVd genome and five, to its part (~260 bp). The nucleotide sequences of cloned amplification products were identified to species in the BLAST system and deposited in GenBank. The amplicons obtained with the PSTVd-specific primers were identified as PSTVd sequences in all Ph. infestans isolates examined. The majority of the nucleotide sequences were phylogenetically related to BLAST sequences of PSTVd strains originating from Russia; several strains showed similarity to strains from other countries (France, China, and West African countries). The results demonstrate that PSTVd was for the first time detected in natural (field) Ph. infestans isolates and offer new opportunities for studying the intricate multilevel host-parasite interactions.

Development of Reverse Transcription Recombinase Polymerase Amplification (RT-RPA): A Methodology for Quick Diagnosis of Potato Leafroll Viral Disease in Potato.

Potatoes are developed vegetatively from tubers, and therefore potato virus transmission is always a possibility. The potato leafroll virus (PLRV) is a highly devastating virus of the genus Polerovirus and family Luteoviridae and is regarded as the second-most destructive virus after Potato virus Y. Multiple species of aphids are responsible for the persistent and non-propagating transmission of PLRV. Due to intrinsic tuber damage (net necrosis), the yield and quality are drastically diminished. PLRV is mostly found in phloem cells and in extremely low amounts. Therefore, we have attempted to detect PLRV in both potato tuber and leaves using a highly sensitive, reliable and cheap method of one-step reverse transcription-recombinase polymerase amplification (RT-RPA). In this study, an isothermal amplification and detection approach was used for efficient results. Out of the three tested primer sets, one efficiently amplified a 153-bp product based on the coat protein gene. In the present study, there was no cross-reactivity with other potato viruses and the optimal amplification reaction time was thirty minutes. The products of RT-RPA were amplified at a temperature between 38 and 42 °C using a simple heating block/water bath. The present developed protocol of one-step RT-RPA was reported to be highly sensitive for both leaves and tuber tissues equally in comparison to the conventional reverse transcription-polymerase chain reaction (RT-PCR) method. By using template RNA extracted employing a cellular disc paper-based extraction procedure, the method was not only simplified but it detected the virus as effectively as purified total RNA. The simplified one-step RT-RPA test was proven to be successful by detecting PLRV in 129 samples of various potato cultivars (each consisting of leaves and tubers). According to our knowledge, this is the first report of a one-step RT-RPA performed using simple RNA extracted from cellular disc paper that is equally sensitive and specific for detecting PLRV in potatoes. In terms of versatility, durability and the freedom of a highly purified RNA template, the one-step RT-RPA assay exceeds the RT-PCR assay, making it an effective alternative for the certification of planting materials, breeding for virus resistance and disease monitoring.

Effects of Au@Ag core-shell nanostructure with alginate coating on male reproductive system in mice.

Despite the widespread use of silver nanoparticles (NPs), these NPs can accumulate and have toxic effects on various organs. However, the effects of silver nanostructures (Ag-NS) with alginate coating on the male reproductive system have not been studied. Therefore, this study aimed to investigate the impacts of this NS on sperm function and testicular structure. After the synthesis and characterization of Ag-NS, the animals were divided into five groups (n = 8), including one control group, two sham groups (received 1.5 mg/kg/day alginate solution for 14 and 35 days), and two treatment groups (received Ag-NS at the same dose and time). Following injections, sperm parameters, apoptosis, and autophagy were analyzed by the TUNEL assay and measurement of the mRNA expression of Bax, Bcl-2, caspase-3, LC3, and Beclin-1. Fertilization rate was assessed by in vitro fertilization (IVF), and testicular structure was analyzed using the TUNEL assay and hematoxylin and eosin (H&E) staining. The results showed that the NS was rod-shaped, had a size of about 60 nm, and could reduce sperm function and fertility. Gene expression results demonstrated an increase in the apoptotic markers and a decrease in autophagy markers, indicating apoptotic cell death. Moreover, Ag-NS invaded testicular tissues, especially in the chronic phase (35 days), resulting in tissue alteration and epithelium disintegration. The results suggest that sperm parameters and fertility were affected. In addition, NS has negative influences on testicular tissues, causing infertility in men exposed to these NS.

Clinical efficacy of combination therapy of FuXi-Tiandi-Wuxing Decoction and anti-viral drugs in the treatment of novel coronavirus pneumonia: A prospective interventional study.

Introduction: The National Administration of Traditional Chinese Medicine of the People's Republic of China (NATCM) and the State Administration of Traditional Chinese medicine (TCM) advocated a combination therapy of TCM and anti-viral drugs for novel coronavirus pneumonia (NCP) to improve the efficacy of clinical treatment. Methods: Forty-six patients diagnosed with NCP were sequentially divided into intent-to-treat population: the experimental group (combination of FuXi-Tiandi-Wuxing Decoction and anti-viral drugs; n = 23) and the control group (anti-viral drugs only) (n = 23). The two groups were compared in terms of duration of fever, cough symptom score, fatigue, appetite, dyspnea, out-of-bed activities, chest computer tomography (CT) recovery, virological clearance, average length of hospital stay, and clinical effective rate of drug. After 6 days of observation, patients from the control group were divided into as-treated population: experimental subgroup (n = 14) to obtain clinical benefit and control subgroup (n = 9). Results: There was a significant improvement in the duration of fever (1.087 ± 0.288 vs 4.304 ± 2.490), cough (0.437 ± 0.589 vs 2.435 ± 0.662; P < 0.05), chest CT evaluation (82.6% vs 43.4%; P < 0.05), and virological clearance (60.8% vs 8.7%; P < 0.05) in patients of the experimental group compared with patients in the control group. Further observation in as-treated population reported that cough (0.742 ± 0.463 vs 1.862 ± 0.347; P < 0.05) and fatigue (78.5% vs 33.3%; P < 0.05) were significantly relieved after adding FuXi-Tiandi-Wuxing Decoction to the existing treatment. Conclusion: An early treatment with combination therapy of FuXi-Tiandi-Wuxing Decoction and anti-viral drugs significantly relieves the clinical symptoms of NCP, shows improvement in chest CT scan, improves virological clearance, shortens average length of hospital stay, and reduces the risk of severe illness. The effect of FuXi-Tiandi-Wuxing Decoction in NCP may be clinically important and require further consideration.

Overview of gene expression techniques with an emphasis on vitamin D related studies.

Each cell controls when and how its genes must be expressed for proper function. Every function in a cell is driven by signaling molecules through various regulatory cascades. Different cells in a multicellular organism may express very different sets of genes, even though they contain the same DNA. The set of genes expressed in a cell determines the set of proteins and functional RNAs it contains, giving it its unique properties. Malfunction in gene expression harms the cell and can lead to the development of various disease conditions. The use of rapid high-throughput gene expression profiling unravels the complexity of human disease at various levels. Peripheral blood mononuclear cells (PBMC) have been used frequently to understand gene expression homeostasis in various disease conditions. However, more studies are required to validate whether PBMC gene expression patterns accurately reflect the expression of other cells or tissues. Vitamin D, which is responsible for a multitude of health consequences, is also an immune modulatory hormone with major biological activities in the innate and adaptive immune systems. Vitamin D exerts its diverse biological effects in target tissues by regulating gene expression and its deficiency, is recognized as a public health problem worldwide. Understanding the genetic factors that affect vitamin D has the potential benefit that it will make it easier to identify individuals who require supplementation. Different technological advances in gene expression can be used to identify and assess the severity of disease and aid in the development of novel therapeutic interventions. This review focuses on different gene expression approaches and various clinical studies of vitamin D to investigate the role of gene expression in identifying the molecular signature of the disease.

Gene optimizations are essential in maintaining biological functions. Gene dysregulation results in disease progression. Advanced analytical techniques determine the link between impaired genes and disease conditions. This knowledge can be applied to design clinical trials to aid novel therapeutic interventions and disease prevention.

Modulation of the Functional State of Mouse Neutrophils by Selenium Nanoparticles In Vivo.

This study aimed to discover the immunomodulatory effect of selenium nanoparticles (SeNPs) on the functional state of neutrophils in vivo. Intraperitoneal injections of SeNPs (size 100 nm) 2.5 mg/kg/daily to BALB/c mice for a duration of 7-28 days led to the development of an inflammatory reaction, which was registered by a significant increase in the number of neutrophils released from the peritoneal cavity, as well as their activated state, without additional effects. At the same time, subcutaneous injections of the same SeNPs preparations at concentrations of 0.1, 0.5, and 2.5 mg/kg, on the contrary, modulated the functional state of neutrophils depending on the concentration and duration of SeNPs administration. With the use of fluorescence spectroscopy, chemiluminescence, biochemical methods, and PCR analysis, it was found that subcutaneous administration of SeNPs (0.1, 0.5, and 2.5 mg/kg) to mice for a short period of time (7-14 days) leads to modification of important neutrophil functions (adhesion, the number of migrating cells into the peritoneal cell cavity, ROS production, and NET formation). The obtained results indicated the immunostimulatory and antioxidant effects of SeNPs in vivo during short-term administration, while the most pronounced immunomodulatory effects of SeNPs were observed with the introduction of a low concentration of SeNPs (0.1 mg/kg). Increase in the administration time of SeNPs (0.1 mg/kg or 2.5 mg/kg) up to 28 days led to a decrease in the adhesive abilities of neutrophils and suppression of the expression of mRNA of adhesive molecules, as well as proteins involved in the generation of ROS, with the exception of NOX2; there was a tendency to suppress gene expression pro-inflammatory factors, which indicates the possible manifestation of immunosuppressive and anti-inflammatory effects of SeNPs during their long-term administration. Changes in the expression of selenoproteins also had features depending on the concentration and duration of the administered SeNPs. Selenoprotein P, selenoprotein M, selenoprotein S, selenoprotein K, and selenoprotein T were the most sensitive to the introduction of SeNPs into the mouse organism, which indicates their participation in maintaining the functional status of neutrophils, and possibly mediated the immunomodulatory effect of SeNPs.