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Acute hepatopancreatic necrosis disease (AHPND) is a significant deadly infectious disease in the shrimp farming industry, causing serious economic losses globally every year. Because of the rapid progress speed, lack of effective treatment and high mortality rate of AHPND, monitoring with frequent diagnostic tests is vital for a successful prevention. The conventional histopathological diagnosis fell far short of the requirement for efficient monitoring, and the polymerase chain reaction (PCR)-based molecular diagnostic methods that rely on sophisticated thermocycler and trained personnel are hardly applicable in the field. Combining the recombinase polymerase amplification (RPA) and the lateral flow strips (LFSs), a diagnostic method suitable for on-site everyday monitoring of AHPND has been established in this study. This RPA-LFS method targeted the binary toxic photorhabdus insect-related genes PirA and PirB on a virulence plasmid of the AHPND-causative Vibrio parahaemolyticus strains. The diagnostic test was completed within 30 min at 37°C and showed good specificity and good sensitivity of 20 fg DNA of the AHPND shrimp or one colony-forming unit of the causative bacterium per reaction, which was better than the administration-approved standard AP4 assay. Crude templates from sample boiling could be directly used. Tests of clinical samples showed 100% consistency of this method with the standard AP4 assay. This RPA-LFS method can be a good choice for on-site diagnosis of AHPND with quick response time, easy procedure and low demand for resources, and should have significant value for the control of spreading of this dangerous disease in farmed shrimp.

Salmonella, being one of the most widespread foodborne pathogens, is a compulsory test item required by national food safety standard of China and many other countries. More sensitive and specific Salmonella detection method is still needed since traditional methods are time consuming and highly dependent on enormous manpower and material resources. In this research, a bacteria detection method based on CRISPR-Cas13a system (where CRISPR is Clustered Regularly Interspaced Short Palindromic Repeats) was proposed. The target DNA was amplified by PCR and transcribed into RNA by T7 transcriptase, which can activate the RNase activity of the Cas13a protein. The self-folding quenched fluorescent probe can be cleaved by the activated Cas13a protein to generate fluorescent signal. We named this method as PCF detection (PCR-CRISPR-Fluorescence based nucleic acid detection). In this study, PCF detection showed excellent sensitivity, which can detect Salmonella genomic DNA with a minimum of 101 aM or 10° CFU/ml Salmonella bacteria in 2 hr. It also showed good specificity with no cross-reaction with other common foodborne bacteria. PRACTICAL APPLICATION: The PCF detection method proposed in this article can detect Salmonella sensitively and specifically, providing a novel strategy for the detection of foodborne pathogens in food and has great application potential in other microbial detection fields.

Several clinical studies have reported that hearing loss is correlated with autism in children. However, little is known about the underlying mechanism between hearing loss and autism. p21-activated kinases (PAKs) are a family of serine/threonine kinases that can be activated by multiple signaling molecules, particularly the Rho family of small GTPases. Previous studies have shown that Pak1 mutations are associated with autism. In the present study, we take advantage of Pak1 knockout (Pak1-/-) mice to investigate the role of PAK1 in hearing function. We find that PAK1 is highly expressed in the postnatal mouse cochlea and that PAK1 deficiency leads to hair cell (HC) apoptosis and severe hearing loss. Further investigation indicates that PAK1 deficiency downregulates the phosphorylation of cofilin and ezrin-radixin-moesin and the expression of βII-spectrin, which further decreases the HC synapse density in the basal turn of cochlea and disorganized the HC stereocilia in all three turns of cochlea in Pak1-/- mice. Overall, our work demonstrates that the autism-related gene Pak1 plays a crucial role in hearing function. As the first candidate gene linking autism and hearing loss, Pak1 may serve as a potential target for the clinical diagnosis of autism-related hearing loss.

A series of mononuclear lanthanide complexes with the formula [Hex4N][Ln(DBM)4] (1Ln) (Ln = Nd and Tb), [But4N][Dy(DBM)4] (2), [Et4N][Dy(DBM)4] (3), [PhCH2N(CH3)3][Dy(DBM)4] (4) and [Hex4N][Dy(TTA)4] (5) formed by four β-diketonate ligands (i.e. DBM = dibenzoylmethane, TTA = 2-thenoyltrifluoroacetone) and four organic counter ammonium cations (i.e. Hex4N = tetrahexylammonium, But4N = tetrabutylammonium, Et4N = tetraethylammonium and PhCH2N+(CH3)3) were structurally and magnetically characterized. The organic ammonium as contactless cation pairs with [Ln(β-diketonate)4]−1 moieties, the impact of different remote organic cations and the first coordination sphere comprising distinct β-diketonates on their SMM performance were elaborately studied. This is an efficient method by virtue of the interaction of the remote organic ion pair to modulate their single-molecule magnetic (SMM) properties. More importantly, the tetrakis β-diketonate Dy(III) anion can be almost intactly grafted on functional GaN substrate through an anion exchange reaction, resulting in diminishing slow magnetic relaxation behavior, which presents potential application in future SMM devices.

Flavonoids have notable biological activities and have been widely used in the medicinal and chemical industries. However, single-copy integration of heterologous pathway genes limits the production of flavonoids. In this work, we designed and constructed single-step integration of multiple flavonoid (2S)-naringenin biosynthetic pathway genes in S. cerevisiae. The efficiency of the naringenin metabolic pathway gene integration into the rDNA site reached 93.7%. Subsequently, we used a high titer p-coumaric acid strain as a chassis, which eliminated feedback inhibition of tyrosine and downregulated the competitive pathway. The results indicated that increasing the supply of p-coumaric acid was effective for naringenin production. We additionally optimized the amount of donor DNA. The optimum strain produced 149.8 mg/L of (2S)-naringenin. The multi-copy integration of flavonoid pathway genes effectively improved (2S)-naringenin production in S. cerevisiae. We further analyzed the copy numbers and expression levels of essential genes (4CL and CHS) in the (2S)-naringenin metabolic pathway by qPCR. Higher copy numbers of the (2S)-naringenin metabolic pathway genes were associated with greater 4CL and CHS transcription, and the efficiency of naringenin production was higher. Therefore, multi-copy integration of genes in the (2S)-naringenin metabolic pathway was imperative in rewiring p-coumaric acid flux to enhance flavonoid production.

The compound (2S)-eriodictyol is an important flavonoid that can be derived from (2S)-naringenin through flavonoid 3'-hydroxylase (F3'H) catalyzation. F3'H is a cytochrome P450 enzyme that requires a cytochrome P450 reductase (CPR) to function. However, P450s have limited applications in industrial scale biosynthesis, owing to their low activity. Here, an efficient SmF3'H and a matched SmCPR were identified from Silybum marianum. To improve the efficiency of SmF3'H, we established a high-throughput detection method for (2S)-eriodictyol, in which the promoter combination of SmF3'H and SmCPR were optimized in Saccharomyces cerevisiae. The results revealed that SmF3'H/SmCPR should be expressed by using promoters with similar and strong expression levels. Furthermore, directed evolution was applied to further improve the efficiency of SmF3'H/SmCPR. With the optimized promoter and mutated combinations SmF3'HD285N/SmCPRI453V, the (2S)-eriodictyol titer was improved to 3.3 g/L, the highest titer in currently available reports. These results indicated that S. cerevisiae is an ideal platform for functional expression of flavonoid related P450 enzymes.

Severe invasive aspergillosis infection occurs when human immune function is impaired. The interaction between Aspergillus fumigatus (A. fumigatus) conidia and type II lung epithelial cells serves an important role in disease progression. The present study compared the proteomes of A549 human lung epithelial cells with and without A. fumigatus infection. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and protein interaction analyses were performed, and differential protein expression was verified by western blotting and reverse transcription-quantitative PCR (RT-qPCR). In addition, the RNA interference method, an internalization assay and ELISA were performed. Isobaric tags for relative and absolute quantification analysis detected a total of 1,582 proteins, from which 111 proteins with differential expression were obtained (fold change >1.5 or

BACKGROUND: Ovarian cancer is the common tumor in female, the prognostic of which is influenced by a series of factors. In this study, 4 genes relevant to pathological grade in ovarian cancer were screened out by the construction of weighted gene co-expression network analysis. METHODS: GSE9891 with 298 ovarian cancer cases had been used to construct co-expression networks. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses was used to analyze the possible mechanism of genes involved in the malignant process of ovarian cancer. Hub genes were validated in other independent datasets, such as GSE63885, GSE26193 and GSE30161. Survival analysis based on the hub genes was performed by website of Kaplan Meier-plotter. RESULTS: The result based on weighted gene co-expression network analysis indicated that turquoise module has the highest association with pathological grade. Gene Ontology enrichment analysis revealed that the genes in turquoise module main enrichment in inflammatory response and immune response. Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that the genes in turquoise module main enrichment in cytokine-cytokine receptor interaction and chemokine signaling pathway. In turquoise module, a total of 4 hub genes (MS4A4A, CD163, CPR65, MS4A6A) were identified. Then, 4 hub genes were effectively verified in the test datasets (GSE63885, GSE26193 and GSE30161) and tissue samples from Shengjing Hospital of China Medical University. Survival analysis indicated that the 4 hub genes were associated with poor progression-free survival of ovarian cancer. CONCLUSIONS: In conclusion, 4 hub genes (MS4A4A, CD163, CPR65, MS4A6A) were verified associated with pathological grade of ovarian cancer. Moreover, MS4A4A, CD163, MS4A6A may serve as a surface marker for M2 macrophages. Targeting the 4 hub genes may can improve the prognosis of ovarian cancer.

Welsh onions (Allium fistulosum L.) are often affected by stressful environments, such as high light and drought, during summer cultivation, which hinders their growth. We used CO2 assimilation, OJIP transient and MR curves to analyse the photosynthetic characteristics of Welsh onion. The results showed that single high light stress caused a decrease in the net photosynthesis rate through stomatal limitation, while the single drought treatment and the combined stress induced nonstomatal limitation. FO and FJ increased, Fm decreased, and a distinct K-phase was induced. High light and drought stress blocked MR transients, leading to a gradual decrease in VPSI and VPSII-PSI . In general, photosynthesis of Welsh onion was inhibited by high light and drought, which destroyed the receptor and donor side of PSII and reduced electron transport capacity of PSII and PSI.

Non-invasive prenatal testing (NIPT) for common fetal aneuploidies using circulating cell free DNA in maternal plasma has been widely adopted in clinical practice for its sensitivity and accuracy. However, the detection of subchromosomal abnormalities or monogenetic variations showed no cost-effectiveness or satisfactory accuracy. Here we describe the goodness-of-fit and graphical analysis of polymorphic sites based non-invasive prenatal testing (GGAP-NIPT) assay, which was sensitive and accurate for detecting fetal abnormalities simultaneously at the chromosomal, subchromosomal and nucleotide levels. In each sample, fetal fraction was estimated using allelic counts of polymorphic sites from normal reference chromosomal regions and each target amplicon’s genotype was determined with the aid of goodness-of-fit test. Then group of polymorphic amplicons from each target region was analyzed collectively using both statistical and graphical approaches, and fetal genetic abnormality was identified by either overall statistical model fitness or graphical characteristic allelic clusters, whereas regions containing only a single polymorphic site were analyzed using a limited number of replicates. For simulated samples, such a group analytic approach identified all aneuploidies, microdeletions or microduplications and single-gene short variations precisely. As no cross-sample comparison was performed and the relative allelic counts of each target polymorphic amplicon were internally controlled, the GGAP-NIPT assay reported here should be sensitive and accurate inherently. Therefore, the GGAP-NIPT analytical approach has the potential to detect most fetal abnormalities at the chromosomal, subchromosomal and monogenic levels with improved accuracy, facilitating the extension of NIPT to an expanded panel of genetic disorders.

Ulva compressa is one of the causal green macroalgae in many countries. In this study, complete chloroplast genome sequence of U. compressa was reported, and the total length of this species was 94...

Purpose Sporothrix schenckii is a thermally dimorphic fungus. In a saprotrophic environment or culturing at 25 °C, it grows as mycelia, whereas in host tissues or culturing at 37 °C, it undergoes dimorphic transition and division into pathogenic yeast cells. S. schenckii can cause serious disseminated sporotrichosis in immunocompromised hosts and presents an emerging global health problem. The mycelium-to-yeast transition was a consequence of the adaptive process to different environment. Some studies showed that the transition was significantly related to the virulence and pathogenesis of dimorphic fungi. However the genetic mechanisms of this complicated biological process are poorly understood. Method Our study presented a comparative transcriptomic analysis perspective on temperature stress in a visceral isolates of S. schenckii, obtaining more genetic information related to dimorphic transition. Results The 9.38 Gbp dataset was generated and assembled into 14,423 unigenes. Compared with gene and protein databases, 9561 unigenes were annotated. Comparative analysis identified 1259 genes expressed differentially in mycelium and yeast phase, and were categorized into a number of important biological processes, such as synthesis and metabolism, transmembrane transport, biocatalysis, oxidation reduction, and cellular signal transduction. Conclusions The findings suggested that temperature-dependent transition was tightly associated with stress adaptation, growth and development, signal regulation, adhesion, and colonization, which was predicted to be related with virulence and pathogenesis. Collection of these data should offer fine-scale insights into the mechanisms of dimorphism and pathogenesis of S. schenckii, and meanwhile facilitate the evolutionary and function studies of other dimorphic fungi.

The stability and processing of cellular RNA transcripts are efficiently controlled via non-templated addition of single or multiple nucleotides, which is catalyzed by various nucleotidyltransferases including poly(A) polymerases (PAPs). Germline development defective 2 (GLD-2) is among the first reported cytoplasmic non-canonical PAPs that promotes the translation of germline-specific mRNAs by extending their short poly(A) tails in metazoan, such as Caenorhabditis elegans and Xenopus. On the other hand, the function of mammalian GLD-2 seems more diverse, which includes monoadenylation of certain microRNAs. To understand the structural basis that underlies the difference between mammalian and non-mammalian GLD-2 proteins, we determine crystal structures of two rodent GLD-2s. Different from C. elegans GLD-2, mammalian GLD-2 is an intrinsically robust PAP with an extensively positively charged surface. Rodent and C. elegans GLD-2s have a topological difference in the β-sheet region of the central domain. Whereas C. elegans GLD-2 prefers adenosine-rich RNA substrates, mammalian GLD-2 can work on RNA oligos with various sequences. Coincident with its activity on microRNAs, mammalian GLD-2 structurally resembles the mRNA and miRNA processor terminal uridylyltransferase 7 (TUT7). Our study reveals how GLD-2 structurally evolves to a more versatile nucleotidyltransferase, and provides important clues in understanding its biological function in mammals.

Bacillus cereus is an opportunistic pathogen that can cause emetic or diarrheal foodborne illness. Previous studies have identified multiple pathogenic B. cereus strains and characterized a variety of virulence factors. Here, we demonstrate that the virulence and lethality of B. cereus for mammalian cells and host animals involve the interaction of B. cereus flagellin proteins and the host-cell-surface-localized glycosphingolipid Gb3 (CD77, Galα1-4Galβ1-4Glcβ1-Cer). We initially found that B. cereus infection was less lethal for Gb3-deficiencient A4galt−/- mice than for wild-type mice. Subsequent experiments established that some factor other than secreted toxins must account of the observed differential lethality: Gb3-deficiencient A4galt−/- mice were equally susceptible to secreted-virulence-factor-mediated death as WT mice, and we observed no differences in the bacterial loads of spleens or livers of mice treated with B. cereus strain vs. mice infected with a mutant variant of incapable of producing many secreted toxins. A screen for host-interacting B. cereus cell wall components identified the well-known flagellin protein, and both flagellin knockout strain assays and Gb3 inhibitor studies confirmed that flagellin does interact with Gb3 in a manner that affects B. cereus infection of host cells. Finally, we show that treatment with polyclonal antibody against flagellin can protect mice against B. cereus infection. Thus, beyond demonstrating a previously unappreciated interaction between a bacterial motor protein and a mammalian cell wall glycosphingolipid, our study will provide useful information for the development of therapies to treat infection of B. cereus.

Osteosarcoma is a type of aggressive malignant bone tumour that frequently metastasizes to lungs, resulting in poor prognosis. However, the molecular mechanisms of lung metastasis of osteosarcoma remain poorly understood. Here we identify exon-intron fusion genes in osteosarcoma cell lines and tissues. These fusion genes are derived from chromosomal translocations that juxtapose the coding region for amino acids 1-38 of Rab22a (Rab22a1-38) with multiple inverted introns and untranslated regions of chromosome 20. The resulting translation products, designated Rab22a-NeoFs, acquire the ability to drive lung metastasis of osteosarcoma. The Rab22a1-38 moiety governs the function of Rab22a-NeoFs by binding to SmgGDS-607, a GTP-GDP exchange factor of RhoA. This association facilitates the release of GTP-bound RhoA from SmgGDS-607, which induces increased activity of RhoA and promotes metastasis. Disrupting the interaction between Rab22a-NeoF1 and SmgGDS-607 with a synthetic peptide prevents lung metastasis in an orthotopic model of osteosarcoma. Our findings may provide a promising strategy for a subset of osteosarcoma patients with lung metastases.

Pathway optimization plays an important role in fine-tuning metabolic pathways. In most conditions, more than three genes are involved in the biosynthesis pathway of a specific target product. To improve the titer of products, rational regulation of a group of genes by a series of promoters with different strengths is essential. On the basis of a series of RNA-Seq data, a set of 66 native promoters was chosen to fine-tune gene expression in Saccharomyces cerevisiae. Promoter strength was characterized by measuring the fluorescence strength of the enhanced green fluorescent protein through fluorescence-activated cell sorting. The expressions of PTDH1, PPGK1, PINO1, PSED1, and PCCW12 were stronger than that of PTDH3, whereas those of another 15 promoters were stronger than that of PTEF1. Then, 30 promoters were chosen to optimize the biosynthesis pathway of (2S)-naringenin from p-coumaric acid. With a high-throughput screening method, the highest titer of (2S)-naringenin in a 5 L bioreactor reached 1.21 g/L from p-coumaric acid, which is the highest titer according to the currently available reports.

Vibrio parahaemolyticus is an important pathogenic bacterium in both food safety management and mariculture. Rapid and accurate detection technologies are critical for effective control of its outbreak and spreading. Conventional technologies and polymerase chain reaction (PCR)-based approaches have limited usage because of the requirement of laboratory instruments and trained personnel. Using the isothermal recombinase polymerase amplification (RPA) technology, several detection assays have been developed with added convenience. Combining the lateral flow strip (LFS) test with RPA can further simplify the detection. In this study, an improved RPA assay using LFS for visual detection of V. parahaemolyticus was developed. Primers were designed targeting the virulence genes and screened for amplification efficiency, nonspecific amplification, and primer-dimer formation. Probes were designed for the best primer pairs, and the weakness of LFS tests, being easily affected by primer-dependent artifacts, was overcome by sequence modifications on primers and probe. The RPA-LFS assay took 25 min at 35 to 45 °C, and showed excellent specificity. It detected as low as one colony forming unit (CFU) of V. parahaemolyticus per reaction without DNA purification, or 10 CFU/10 g spiked food samples with 2 hr of enrichment. The detection limit was better than the currently available RPA-based detection methods. Application of the RPA-LFS assay for simulated samples or real clinical samples showed accurate and consistent detection results compared to bioassay and quantitative PCR. The RPA-LFS assay provided a rapid, accurate, and convenient V. parahaemolyticus detection method suitable for on-site detection in resource-limited conditions. PRACTICAL APPLICATION: This research developed a rapid and visual detection technology for Vibrio parahaemolyticus that is not dependent on complicated equipment. The detection process takes 25 min and the result is read with the naked eye. A detection kit can be developed based on this technology for on-site detection of V. parahaemolyticus in resource-limited regions for food safety management and mariculture.

Summary Microalgae as a biofuel source are of great interest. Bacterial phycosphere inhabitants of algal cultures are hypothesized to contribute to productivity. In this study, the bacterial composition of the Chlorella sorokiniana phycosphere was determined over several production cycles in different growing seasons by 16S rRNA gene sequencing and identification. The diversity of the phycosphere increased with time during each individual reactor run, based on Faith’s phylogenetic diversity metric versus days post‐inoculation (R = 0.66, P, Microalgae can be efficiently cultivated to produce biomass suitable for conversion to liquid biofuels. Long term experiments conducted over multiple growing seasons in the arid Southwest region of the United States revealed significant associations between algal productivity and the bacterial members of the culture phycospheres. These species offer targets for improving algal biomass yields in mass cultivation.

Oxidative stress is one of the hallmarks of ischemic stroke. Catalase-based (CAT) biomimetic complexes are emerging as promising therapeutic candidates that are expected to act as neuroprotectants for ischemic stroke by decreasing the damaging effects from H2O2. Unfortunately, these molecules result in the unwanted production of the harmful hydroxyl radical, HO•. Here, we report a series of salen-based tri-manganese (Mn(III)) metallocryptands (1-3) that function as catalase biomimetics. These cage-like molecules contain a unique "active site" with three Mn centers in close proximity, an arrangement designed to facilitate metal cooperativity for the effective dismutation of H2O2 with minimal HO• production. In fact, significantly greater oxygen production is seen for 1-3 as compared to the monomeric Mn(Salen) complex, 1c. The most promising system, 1, was studied in further detail and found to confer a greater therapeutic benefit both in vitro and in vivo than the monomeric control system, 1c, as evident from inter alia studies involving a rat model of ischemic stroke damage and supporting histological analyses. We thus believe that metallocryptand 1 and its analogues represent a new and seemingly promising strategy for treating oxidative stress related disorders.

Among the three key active components (KACs) of Magnolia officinalis bark extract (ME), 4-O-methylhonokiol and honokiol showed higher antiproliferation activities than magnolol in the oral squamous cancer cell lines (Cal-27, SCC-9, and SCC-4). Oral bioavailabilities of ME-KACs were poor (

Background Vibrio alginolyticus is an important pathogen that has to be closely monitored and controlled in the mariculture industry because of its strong pathogenicity, quick onset after infection and high mortality rate in aquatic animals. Fast, simple and specific methods are needed for on-site detection to effectively control outbreaks and prevent economic losses. The detection specificity towards the pathogenic strains has to be emphasized to facilitate pointed treatment and prevention. Polymerase chain reaction (PCR)-based molecular approaches have been developed, but their application is limited due to the requirement of complicated thermal cycling machines and trained personnel. Results A fast, simple and highly specific detection method for V. alginolyticus pathogenic strains was established based on isothermal recombinase polymerase amplification (RPA) and lateral flow dipsticks (LFD). The method targeted the virulence gene toxR, which is reported to have good coverage for V. alginolyticus pathogenic strains. To ensure the specificity of the method, the primer-probe set of the RPA system was carefully designed to recognize regions in the toxR gene that diverge in different Vibrio species but are conserved in V. alginolyticus pathogenic strains. The primer-probe set was determined after a systematic screening of amplification performance, primer-dimer formation and false positive signals. The RPA-LFD method was confirmed to have high specificity for V. alginolyticus pathogenic strains without any cross reaction with other Vibrio species or other pathogenic bacteria and was able to detect as little as 1 colony forming unit (CFU) per reaction without DNA purification, or 170 fg of genomic DNA, or 6.25 × 103 CFU/25 g in spiked shrimp without any enrichment. The method finishes detection within 30 min at temperatures between 35 °C and 45 °C, and the visual signal on the dipstick can be directly read by the naked eye. In an application simulation, randomly spiked shrimp homogenate samples were 100% accurately detected. Conclusions The RPA-LFD method developed in this study is fast, simple, highly specific and does not require complicated equipment. This method is applicable for on-site detection of V. alginolyticus pathogenic strains for the mariculture industry.

Human interleukin (IL)-37 is a member of the IL-1 family with potent anti-inflammatory and immunosuppressive properties. Previously, it has been reported that IL-37 suppresses tumor growth and progression. However, the roles of IL-37 in pancreatic cancer development and chemo-resistance remain unknown. Methods: Immunohistochemistry was used to analyze the correlation between IL-37 expression and clinicopathological features of pancreatic ductal adenocarcinoma (PDAC). Western-blot and RT-PCR was used to verify the correlation between IL-37 and hypoxia-inducible factor (HIF)-1α. We performed chromatin immunoprecipitation and luciferase assays to validate HIF-1α suppression of IL-37 expression. Moreover, gain- and loss-of-function studies in vitro and in vivo were used to demonstrate the biological function of IL-37 on PDAC development and chemo-resistance. Results: Our results showed that IL-37 expression was remarkably decreased in PDAC tissues when compared to adjacent normal pancreatic tissues. Reduced IL-37 expression in PDACs was associated with increased PDAC histological grade, tumor size, lymph node metastasis and vessel invasion. IL-37 low patients also have remarkably shorter relapse-free and overall survival. Importantly, IL-37 expression was positively correlated with Gemcitabine efficacy. Mechanistically, HIF-1α attenuated IL-37 transcription by binding to the hypoxia response elements (HREs) in IL-37 promoter. Conversely, IL-37 suppressed HIF-1α expression through STAT3 inhibition. Functionally, downregulation of IL-37 in PDAC cells promoted chemo-resistance, migration and progression in vivo and in vitro. Conclusions: Collectively, our data uncovered IL-37/ STAT3/ HIF-1α negative feedback signaling drives Gemcitabine resistance in PDAC.

Background The Eurasian steppe is an important vegetation type characterized by cold, arid and nitrogen poor conditions. At the Eastern edge, including in the Songnen grassland, the vegetation is dominated by Leymus chinensis (henceforth L. chinensis) and is increasing threatened by elevated anthropogenic nitrogen deposition. L. chinensis is a perennial grass that mainly reproduces vegetatively and its sexual reproduction is limited. However, sexual reproduction plays an important role influencing colonization after large disturbances. To develop an understanding of how elevated nitrogen deposition changes the plant community structure and functioning we need a better understanding how sexual reproduction of L. chinensis changes with nitrogen enrichment. Here we report on a field experiment where we added 10 g N m− 2 yr− 1 and examined changes in seed traits, seed germination and early seedling growth. Results Nitrogen addition increased seed production by 79%, contributing to this seed increases were a 28% increase in flowering plant density, a 40% increase in seed number per plant and a 11% increase in seed weight. Seed size increased with a 42% increase in large seeds and a 49% decrease in the smallest seed size category. Seed germination success improved by 10% for small seeds and 18% for large seeds. Combined, the increased in seed production and improved seed quality doubled the potential seed germination. Subsequent seedling above and below-ground biomass also significantly increased. Conclusions All aspects of L. chinensis sexual reproduction increased with nitrogen addition. Thus, L. chinensis competitive ability may increase when atmospheric nitrogen deposition increases, which may further reduce overall plant diversity in the low diversity Songnen grasslands.

Pseudorabies has caused huge economic losses in China's pig industry and recurred on many large pig farms since late 2011. The disease is caused by highly pathogenic, antigenic variant pseudorabies virus (vPRV) strains. Therefore, the prevention and control of this recurrence of pseudorabies in China has been given priority. In a previous study, we showed that a suitable dose [1 × 106.3 50% tissue culture infectious dose (TCID50) per animal] of commercial Bartha-K61 vaccine protects growing pigs against lethal challenge by the emerging vPRV strain XJ5. In this study, different doses of the Bartha-K61 vaccine and our newly developed rPRV-gI−/gE−/TK− prototype vaccine derived from the vPRV strain XJ5 were used to evaluate immune protection against sublethal challenge by the vPRV strain XJ5. Pigs vaccinated with high doses of the Bartha-K61 vaccine or rPRV-gI−/gE−/TK− prototype vaccine showed no differences in their humoral immune responses, clinical symptoms, body weight gains, viral shedding, or gross and histological lesions after sublethal challenge by the vPRV strain XJ5. Therefore, we concluded that the Bartha-K61 vaccine at a dose of 1 × 105 TCID50 per animal protects pigs against sublethal challenge by the vPRV strain XJ5 and performs equally well as the same dose of the rPRV-gI−/gE−/TK− vaccine, whereas lower doses of the Bartha-K61 vaccine alone do not protect pigs from this challenge. These findings provide useful information for vaccination interventions and the ultimate eradication of pseudorabies caused by vPRV strains emerging in China.

From 2010, porcine epidemic diarrhea virus (PEDV) variants caused sequential outbreaks of disease in Asia and the United States. In this retrospective study, 49 complete spike (S) gene sequences were obtained from PEDV strains collected in China from 2014 to 2016. We observed that variant PEDV strains with novel insertions, deletions, and multiple S gene recombination types were present in China. In addition, mixed infections involving different variant strains were observed in some areas. Based on phylogenetic and recombination analyses, we determined that the newly emerged PEDV variants potentially originated via recombination between the earliest Chinese G1 genogroup strain, JS-2004-2 and earlier Korean pandemic strains. These findings provide important information for understanding ongoing PEDV outbreaks and suggest that novel variants make it more difficult to prevent PEDV infection.

Background Mitochondrial dynamics plays an important role in tumour progression. However, how these dynamics integrate tumour metabolism in hepatocellular carcinoma (HCC) metastasis is still unclear. Methods The mitochondrial fusion protein mitofusin-1 (MFN1) expression and its prognostic value are detected in HCC. The effects and underlying mechanisms of MFN1 on HCC metastasis and metabolic reprogramming are analysed both in vitro and in vivo. Results Mitochondrial dynamics, represented by constant fission and fusion, are found to be associated with HCC metastasis. High metastatic HCC displays excessive mitochondrial fission. Among genes involved in mitochondrial dynamics, MFN1 is identified as a leading downregulated candidate that is closely associated with HCC metastasis and poor prognosis. While promoting mitochondrial fusion, MFN1 inhibits cell proliferation, invasion and migration capacity both in vitro and in vivo. Mechanistically, disruption of mitochondrial dynamics by depletion of MFN1 triggers the epithelial-to-mesenchymal transition (EMT) of HCC. Moreover, MFN1 modulates HCC metastasis by metabolic shift from aerobic glycolysis to oxidative phosphorylation. Treatment with glycolytic inhibitor 2-Deoxy-d-glucose (2-DG) significantly suppresses the effects induced by depletion of MFN1. Conclusions Our results reveal a critical involvement of mitochondrial dynamics in HCC metastasis via modulating glucose metabolic reprogramming. MFN1 may serve as a novel potential therapeutic target for HCC.

BACKGROUND: Human epidermal growth factor receptor 2 (HER2)‐mutant lung cancer remains an orphan of specific targeted therapy. The variable responses to anti‐HER2 therapies in these patients prompt us to examine impact of HER2 variants and co‐mutations on responses to anti‐HER2 treatments in lung cancer. PATIENTS AND METHODS: Patients with stage IV/recurrent HER2‐mutant lung cancers identified through next‐generation sequencings were recruited from seven hospitals. The study comprised a cohort A to establish the patterns of HER2 variants and co‐mutations in lung cancer and a cohort B to assess associations between HER2 variants, co‐mutations, and clinical outcomes. RESULTS: The study included 118 patients (cohort A, n = 86; cohort B, n = 32). Thirty‐one HER2 variants and 35 co‐mutations were detected. Predominant variants were A775_G776insYVMA (49/118, 42%), G778_P780dup (11/118, 9%), and G776delinsVC (9/118, 8%). TP53 was the most common co‐mutation (61/118, 52%). In cohort B, objective response rates with afatinib were 0% (0/14, 95% confidence interval [CI], 0%–26.8%), 40% (4/10, 14.7%–72.6%), and 13% (1/8, 0.7%–53.3%) in group 1 (A775_G776insYVMA, n = 14), group 2 (G778_P780dup, G776delinsVC, n = 10), and group 3 (missense mutation, n = 8), respectively (p = .018). Median progression‐free survival in group 1 (1.2 months; 95% CI, 0–2.4) was shorter than those in group 2 (7.6 months, 4.9–10.4; hazard ratio [HR], 0.009; 95% CI, 0.001–0.079; p < .001) and group 3 (3.6 months, 2.6–4.5; HR, 0.184; 95% CI, 0.062–0.552; p = .003). TP53 co‐mutations (6.317; 95% CI, 2.180–18.302; p = .001) and PI3K/AKT/mTOR pathway activations (19.422; 95% CI, 4.098–92.039; p < .001) conferred additional resistance to afatinib. CONCLUSION: G778_P780dup and G776delinsVC derived the greatest benefits from afatinib among HER2 variants. Co‐mutation patterns were additional response modifiers. Refining patient population based on patterns of HER2 variants and co‐mutations may help improve the efficacy of anti‐HER2 treatment in lung cancer. IMPLICATIONS FOR PRACTICE: Human epidermal growth factor receptor 2 (HER2)‐mutant lung cancers are a group of heterogenous diseases with up to 31 different variants and 35 concomitant genomic aberrations. Different HER2 variants exhibit divergent sensitivities to anti‐HER2 treatments. Certain variants, G778_P780dup and G776delinsVC, derive sustained clinical benefits from afatinib, whereas the predominant variant, A775_G776insYVMA, is resistant to most anti‐HER2 treatments. TP53 is the most common co‐mutation in HER2‐mutant lung cancers. Co‐mutations in TP53 and the PI3K/AKT/mTOR pathway confer additional resistance to anti‐HER2 treatments in lung cancer. The present data suggest that different HER2 mutations in lung cancer, like its sibling epidermal growth factor receptor, should be analyzed independently in future studies.

Dipeptidyl peptidase IV (DPP-IV) inhibitors have become a new choice to treat diabetes. Here, we studied the DPP-IV inhibitory activity of protein hydrolysates from common carp (Cyprinus carpio) roe. The molecular weight distribution and simulated gastrointestinal digestive stability of hydrolysates produced by four enzymes were analyzed. Papain-generated hydrolysate showed the highest DPP-IV inhibition and effective gastrointestinal stability. New effective peptide sequences from papain hydrolysates were identified using liquid chromatography-tandem mass spectrometry. The bioactivity of IPNVAVD, which was identified from papain hydrolysate (IC50 value of 777.35 ± 5.50 μM), was investigated by Caco-2 and HepG2 cell models. We found that the DPP-IV inhibition by papain hydrolysate was not attenuated after simulated gastrointestinal digestion. In addition, IPNVAVD significantly inhibited the DPP-IV secreted by Caco-2 cells with no cytotoxicity. It also promoted glucose uptake in insulin-resistant HepG2 cells. Transport experiments showed that IPNVAVD could be absorbed intactly by the Caco-2 cell monolayer. Protein hydrolysates from common carp roe inhibited DPP-IV effectively, and they may have potential as functional ingredients in food for diabetic patients.

(2S)-Naringenin, a (2S)-flavanone, is widely used in the food, chemical, and pharmaceutical industries because of its diverse physiological activities. The production of (2S)-naringenin in microorganisms provides an ideal source that reduces the cost of the flavonoid. To achieve efficient production of (2S)-naringenin in Saccharomyces cerevisiae (S. cerevisiae), we constructed a biosynthetic pathway from p-coumaric acid, a cost-effective and more efficient precursor. The (2S)-naringenin synthesis pathway genes were integrated into the yeast genome to obtain a (2S)-naringenin production strain. After gene dosage experiments, the genes negatively regulating the shikimate pathway and inefficient chalcone synthase activity were verified as factors limiting (2S)-naringenin biosynthesis. With fed-batch process optimization of the engineered strain, the titer of (2S)-naringenin reached 648.63 mg/L from 2.5 g/L p-coumaric acid. Our results indicate that the constitutive production of (2S)-naringenin from p-coumaric acid in S. cerevisiae is highly promising.

Patients with stage II or III colorectal cancer (CRC) exhibit various clinical outcomes after radical treatments. The 5-year survival rate was between 50 and 87%. However, the underlying mechanisms of the variation remain unclear. Here we show that AMPKα1 is overexpressed in CRC patient specimens and the high expression is correlated with poor patient survival. We further reveal a previously unrecognized function of AMPKα1, which maintains high level of reduced glutathione to keep reduction–oxidation reaction (redox) homeostasis under stress conditions, thus promoting CRC cell survival under metabolic stress in vitro and enhancing tumorigenesis in vivo. Mechanistically, AMPKα1 regulate the glutathione reductase (GSR) phosphorylation possibly through residue Thr507 which enhances its activity. Suppression of AMPKα1 by using nano-sized polymeric vector induces a favorable therapeutic effect, especially when in combination with oxaliplatin. Our study uncovers a novel function of AMPKα1 in redox regulation and identifies a promising therapeutic strategy for treatment of CRC.

Fusarium species exhibit significant intrinsic resistance to most antifungal agents and fungicides, resulting in high mortality rates among immunocompromised patients. Consequently, a thorough characterization of the antifungal resistance mechanism is required for effective treatments and for preventing fungal infections and reducing antifungal resistance. In this study, an isolate of Fusarium oxysporum (wild-type) with broadly resistant to commonly antifungal agents was used to generate 1,450 T-DNA random insertion mutants via Agrobacterium tumefaciens-mediated transformation. Antifungal susceptibility test results revealed one mutant with increased sensitivity to azoles. Compared with the resistant wild-type, the mutant exhibited low MICs to KTZ, ITC, VRC, POS, and PCZ (0.125, 1, 0.06, 0.5, and 0.125μg/ml, respectively). The T-DNA insertion site of this mutant was characterized as involving two adjacent genes, one encoding a hypothetical protein with unknown function and the other encoding the NADPH-cytochrome P450 reductase, referred as CPR1. To confirm the involvement of these genes in the altered azole susceptibility, the independent deletion mutants were generated and the Cpr1 deletion mutant displayed the same phenotypes as the T-DNA random mutant. The deletion of Cpr1 significantly decreased ergosterol levels. Additionally, the expression of the downstream Cyp51 gene was affected, which likely contributed to the observed increased susceptibility to azoles. These findings verified the association between Cpr1 and azole susceptibility in F. oxysporum. Furthermore, this gene may be targeted to improve antifungal treatments.

Rhophilin Rho GTPase binding protein 2 (P76RBE) belongs to rhophilin family of Rho-GTPase-binding proteins and is found to contribute to the development of diverse cancers. Data in Oncomine and Kaplan-Meier Plotter databases showed that P76RBE was upregulated in ovarian cancer tissues compared with normal tissues, and patients with high P76RBE expression had worse overall survival, which indicated P76RBE may be associated with the pathogenesis of ovarian cancer. This study aimed to investigate the role of P76RBE in ovarian cancer and to reveal the possible underlying mechanisms. The results demonstrated that P76RBE was highly expressed in ovarian cancer tissues and ovarian cancer cell lines. Functionally, silencing of P76RBE suppressed the proliferation, induced cell cycle arrest, and inhibited migration and invasion in OVCAR-3 and OV-90 cells, while overexpression of P76RBE showed opposite effects on A2780 cells. Mechanically, P76RBE silencing resulted in downregulation of integrin β1, accompanying the reduced NF-κB p65 phosphorylation and nuclear translocation. Importantly, integrin β1 knockdown effectively rescued the effects of P76RBE overexpression on ovarian cancer cells with suppressed proliferation, migration, and invasion. Additionally, in the xenograft tumors derived from OVCAR-3 and OV-90 cell lines, P76RBE knockdown inhibited tumor growth. Meanwhile, the expression of integrin β1 and NF-κB p65 phosphorylation was decreased. In summary, our findings indicate that P76RBE contributes to the progression of ovarian cancer through regulating the integrin β1/NF-κB signaling, and it may be a promising target for ovarian cancer therapy.

Highlights • FGFR2 rearrangement frequency in Chinese cases is only 5.45%. • The first genomic aberration profiling of FGFRs in Chinese cases is presented. • Cases with FGFRs rearrangement have distinct clinical phenotype. • Patients without FGFR2 rearrangement may harbor other FGFR alterations., Genomic aberrations (GAs) in fibroblast growth factor receptors (FGFRs) are involved in the pathogenesis of intrahepatic cholangiocarcinoma (ICC), and clinical trials have shown efficacy of FGFR inhibitors in treating ICC patients with FGFR GAs such as FGFR2 rearrangement. To clarify the FGFRs GA profile and corresponding clinicopathological features in Chinese patients with ICC, a total of 257 cases were identified. Fourteen cases (5.45%) were positive for FGFR2 rearrangement. Further analysis on the 110 FGFR2 rearrangement negative cases showed that 13 patients present additional FGFRs GAs, including FGFR3 rearrangement (2.73%), and FGFRs mutations. When compared with patients without FGFRs GAs, those with FGFR2 or FGFR3 rearrangement presented more under the age of 58 years, female sex, HBsAb positivity, CD10 expression, and PD-L1 expression. The clinical characteristics between patients with FGFRs mutation and those without FGFRs GAs were similar, with the exception that cases with FGFRs mutation have more hepatolithiasis. We concluded that FGFR rearrangement is associated with unique clinical phenotypes in ICC.

Liquorice is a traditional medicine. Triterpenoids such as glycyrrhizin and glycyrrhetinic acid are the main active constituents of liquorice. Studies have revealed that these compounds exert inhibitory effects on several viruses, including SARS-CoV-2. The main mechanisms of action of these compounds include inhibition of virus replication, direct inactivation of viruses, inhibition of inflammation mediated by HMGB1/TLR4, inhibition of β-chemokines, reduction in the binding of HMGB1 to DNA to weaken the activity of viruses, and inhibition of reactive oxygen species formation. We herein review the research progress on the antiviral effects of glycyrrhizin and its derivatives. In addition, we emphasise the significance of exploring unknown antiviral mechanisms, structural modifications, and drug combinations in future studies.

Shrimp is a globally popular seafood. Shrimp farming has been challenged by various infectious diseases that lead to significant economic losses. The prevention of two important shrimp infectious diseases, the acute hepatopancreatic necrosis disease (AHPND) and the Enterocytozoon hepatopenaei (EHP) infection, is highly dependent on early and accurate diagnostic. On-site monitoring of the two diseases in shrimp farming facilities demands point-of-care-testing (POCT) type of diagnostic assays. This study established a duplex recombinase polymerase amplification (RPA) and lateral flow dipstick (LFD) combined assay that could simultaneously diagnose the two diseases. The optimized RPA-LFD assay could finish the diagnostic in 35 min with good specificity, and the sensitivity reached 101 and 102 gene copies per reaction for EHP and AHPND, respectively, which were at the same level as the currently available molecular diagnostic assays. Test results of clinical samples showed 100% agreement of this assay with the industrial standard nested polymerase chain reaction (PCR) assays, and samples with both diseases were simultaneously identified. Because of the isothermal 37℃ amplification and the visual reading of the signal on dipsticks, the dependence on equipment is minimal. This duplex RPA-LFD assay is well suited for simultaneous POCT diagnostic of the two important shrimp infectious diseases. Moreover, the principle can be applied to multiplex POCT diagnostic of other infectious diseases in aquaculture.

Inappropriate use of antibiotics has accelerated to the emergence of multidrug-resistant bacteria, becoming a major health threat. Moreover, bacterial biofilms contribute to antibiotic resistance and prolonged infections. Bacteriophage (phage) therapy may provide an alternative strategy for controlling multidrug-resistant bacterial infections. In this study, a broad-host-range phage, SHWT1, with lytic activity against multidrug-resistant Salmonella was isolated, characterized and evaluated for the therapeutic efficacy in vitro and in vivo. Phage SHWT1 exhibited specific lytic activity against the prevalent Salmonella serovars, such as Salmonella Pullorum, Salmonella Gallinarum, Salmonella Enteritidis, and Salmonella Typhimurium. Morphological analysis showed that phage SHWT1 was a member of the family Siphoviridae and the order Caudovirales. Phage SHWT1 had a latent period of 5 min and burst size of ~150 plaque-forming units (PFUs)/cell. The phage was stable from pH 3-12 and 4–65°C. Phage SHWT1 also showed capacity to lyse Salmonella planktonic cells and inhibit the biofilm formation at optimal multiplicity of infection (MOI) of 0.001, 0.01, 0.1, and 100, respectively. In addition, phage SHWT1 was able to lyse intracellular Salmonella within macrophages. Genome sequencing and phylogenetic analyses revealed that SHWT1 was a lytic phage without toxin genes, virulence genes, antibiotic resistance genes, or significant genomic rearrangements. We found that phage SHWT1 could successfully protect mice against S. enteritidis and S. typhimurium infection. Elucidation of the characteristics and genome sequence of phage SHWT1 demonstrates that this phage is a potential therapeutic agent against the salmonellosis caused by multidrug-resistant Salmonella.

This study aimed to investigate the effect of myofiber changes and protein oxidation on water holding capacity (WHC) of bighead carp fillets stored at −20 °C. WHC, microstructure, protein oxidation parameters, and specific modifications of oxidized amino acids were analyzed during 9 months of frozen storage. Results indicated that WHC decreased accompanied by myofibers’ structural changes (including the formation of cavities among myofibers, breakage of myofibrils and myofibers, and shortening of sarcomeres) and protein oxidation. SDS-PAGE and carbonyl and sulfhydryl content determination of myofibrillar proteins and exudates gave a detailed description of the protein oxidation. LC-MS/MS analysis demonstrated that oxidation, di-oxidation, and 4-hydroxy-2-nonenal (HNE) and malondialdehyde (MDA) adduction were the top four oxidative modifications of proteins. Oxidative modifications impaired configuration and polarity of proteins, which may further affect WHC. This study provides plausible explanations to support the role of protein oxidation in the decrease of WHC in frozen fillets.

Alveolar echinococcosis (AE) is a malignant and fatal parasitic disease caused by the larvae of Echinococcus multilocularis (E. multilocularis), which inhibits the activity and proliferation of natural killer (NK) cells. In this study, the functional alteration of hepatic NK cells and their related molecules were studied. The AE-infected patient’s tissue was fixed with formalin, embedded in paraffin, and stained with Masson’s trichrome or hematoxylin and eosin (H&E). Single cells from AE-infected patient or E. multilocularis-infected mice were blocked with Fc-receptor (FcR), and stained with monoclonal antibodies, including CD16, CD56, CD3, KIR2DL1, granzyme B, perforin, Interferon gamma (IFN-γ), and tumor necrosis factor-α (TNF-α) or isotype control, to measure molecules and cytokines of NK cells and analyzed by flow cytometry. The Sirius red staining was used to quantitate hepatic fibrosis by calculating quantitative collagen deposition. AE can adjust both the number of hepatic CD56+ NK cells and its KIR2DL1 expression processes. Moreover, the overexpression of KIR2DL1 in NK cells could downregulate the functioning of immune cells in the liver area close to parasitic lesions. The number and dysfunction of NK cells in E. multilocularis infection could be related to the molecule dynamics of cell surface inhibitory receptor Ly49A, leading to hepatic damage and progression of fibrosis. This study illustrated significant increase in hepatic fibrogenesis and apparent upregulation of hepatic CD56+ NK cell population and its KIR2DL1 expression in AE-infected patients. This opposite variation might be related to the impaired NK cells functioning, such as granzyme B, IFN-γ, and TNF-α secretion. In addition, the cell surface inhibitory receptor Ly49A was related to the intracellular cytokine secretion functions of NK cells.

CD73, a cell surface-localized ecto-5'-nucleotidase, is the major enzymatic source of extracellular adenosine. Canonically, it plays multiple roles in cancer-related processes via its metabolite. As a druggable target, clinical trials targeting CD73 in various malignant diseases are currently ongoing. Here, we report the ecto-5'-nucleotidase-independent functions of CD73 in pancreatic ductal adenocarcinoma (PDAC). Our findings support that the elevated expression of CD73 in PDAC cells promotes gemcitabine (GEM) resistance by activating AKT. We discovered that a large amount of intracellular CD73 are localized in the endoplasmic reticulum membrane. Intracellular CD73 physically interacts with major vault protein to activate the SRC-AKT circuit. Troglitazone (TGZ) is a peroxisome proliferator-activated receptor gamma agonist that could inhibit the expression of CD73. The administration of TGZ markedly enhances sensitivity to GEM via downregulating CD73 in PDAC. Our findings support that CD73 could be targeted to overcome chemoresistance in PDAC.

Biodiversity loss, exotic plant invasions and climatic change are currently the three major challenges to our globe and can each affect various ecological processes, including litter composition. To gain a better understanding of global change impacts on ecological processes, these three global change components need to be considered simultaneously. Here we assembled experimental plant communities with species richness levels (1, 2, 4, 8 or 16) and subjected them to drought (no, moderate or intensive drought) and invasion (invasion by the exotic annual plant Symphyotrichum subulatum or not). We collected litter of the native plant communities and let it decompose for nine months within the communities. Drought decreased litter decomposition, while the exotic plant invasion had no impact. Increasing species richness decreased litter decomposition under the mesic condition (no drought), but had little impact under moderate and intensive drought. A structural equation model showed that drought and species richness affected litter decomposition mainly via influencing litter nitrogen concentration, but not via altering the quantity and diversity of soil meso-fauna or soil physio-chemical properties. The negative impact of species diversity on litter decomposition under the mesic condition was mainly ascribed to a sampling effect, i.e. via particularly low litter nitrogen concentrations in the two dominant species. Our results indicate that species richness can interact with drought to affect litter decomposition via effect on litter nitrogen. We conclude that nitrogen-dependent litter decomposition should be a mechanism to predict integrated effects of plant diversity loss, exotic plant invasions and climatic change on litter decomposition.

We investigated the anti-aging effects of velvet antler polypeptide on D-galactose (D-gal)-induced aging mice. D-gal-induced aging mice were established and randomly divided into five groups, the control, model, vitamin E (VE), velvet antler polypeptide low-dose and velvet antler polypeptide high-dose groups. The Morris water maze test was used to evaluate the learning and memory abilities of aging mice. Hippocampal neurons were observed via hematoxylin-eosin staining and transmission electron microscopy. Biochemical methods were used to detect the activities of superoxide dismutase, malonaldehyde and other enzymes and evaluate the influence of velvet antler polypeptide on the antioxidant capacity of aging mice. Using 16S rRNA gene sequencing and meristem technology, we assessed the effect of velvet antler polypeptide on aging mice’s intestinal flora and fatty acid metabolism. The experimental results showed that velvet antler polypeptide could significantly improve aging mice’s learning and cognitive abilities, increase the activities of superoxide dismutase, glutathione peroxidase, and catalase in the serum decrease the malonaldehyde content. Intestinal microecological analysis showed that velvet antler polypeptide could significantly increase the beneficial bacterial genus Lactobacillus abundance. Western blot analysis further demonstrated that velvet antler polypeptide could promote fatty acid metabolism by activating peroxisome proliferator-activated receptor α (PPARα) and upregulating the expression of the downstream enzymes carnitine-palmitoyl transferase-1 A and acyl-CoA oxidase 1 while downregulating that of apolipoprotein E4 (APOE4), thereby reducing fatty acid accumulation and increasing adenosine-triphosphate (ATP) production. Therefore, velvet antler polypeptide improves the intestinal microecology and activates the PPARα/APOE4 pathway to regulate fatty acid metabolism.

The pandemic of COVID-19 caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has led to more than 117 million reported cases and 2.6 million deaths. Accurate diagnosis technologies are vital for controlling this pandemic. Reverse transcription (RT)-based nucleic acid detection assays have been developed, but the strict sample processing requirement of RT has posed obstacles on wider applications. This study established a ligation and recombinase polymerase amplification (L/RPA) combined assay for rapid detection of SARS-CoV−2 on genes N and ORF1ab targeting the specific biomarkers recommended by the China CDC. Ligase-based strategies usually have a low-efficiency problem on RNA templates. This study has addressed this problem by using a high concentration of the T4 DNA ligase and exploiting the high sensitivity of RPA. Through selection of the ligation probes and optimization of the RPA primers, the assay achieved a satisfactory sensitivity of 101 viral RNA copies per reaction, which was comparable to RT-quantitative polymerase chain reaction (RT-qPCR) and other nucleic acid detection assays for SARS-CoV−2. The assay could be finished in less than 30 min with a simple procedure, in which the requirement for sophisticated thermocycling equipment had been avoided. In addition, it avoided the RT procedure and could potentially ease the requirement for sample processing. Once validated with clinical samples, the L/RPA assay would increase the practical testing availability of SARS-CoV-2. Moreover, the principle of L/RPA has an application potential to the identification of concerned mutations of the virus.

MicroRNA-3662 (miR-3662) is minimally expressed in normal human tissues but is highly expressed in all types of cancers, including breast cancer. As determined with The Cancer Genome Atlas dataset, miR-3662 expression is higher in triple-negative breast cancers (TNBCs) and African American breast cancers than in other breast cancer types. However, the functional role of miR-3662 remains a topic of debate. Here, we found that inhibition or knockout of endogenous, mature miR-3662 in TNBC cells suppresses proliferation and migration in vitro and tumor growth and metastasis in vivo. Functional analysis revealed that, for TNBC cells, knockout of miR-3662 reduces the activation of Wnt/β-catenin signaling. Furthermore, using CRISPR-mediated miR-3662 activation and repression, dual-luciferase assays, and miRNA/mRNA immunoprecipitation assays, we established that HMG-box transcription factor 1 (HBP-1), a Wnt/β-catenin signaling inhibitor, is a target of miR-3662 and is most likely responsible for miR-3662-mediated TNBC cell proliferation. Our results suggest that miR-3662 has an oncogenic function in tumor progression and metastasis via an miR-3662-HBP1 axis, regulating the Wnt /β-catenin signaling pathway in TNBC cells. Since miR-3662 expression occurs a tumor-specific manner, it is a promising biomarker and therapeutic target for patients who have TNBCs with dysregulation of miR-3662, especially African Americans.

Avian pathogenic Escherichia coli (APEC), widely spread among poultry, is well-known to cause colibacillosis in chickens, which results in significant losses in poultry industry. The ability to uptake iron in the extra-intestinal environment is prerequisite for APEC survival. For adaptation to the low-iron environments, the bacteria have evolved multiple iron acquisition systems to ensure optimal iron uptake. However, many components of these iron acquisition pathways are still not clearly known. An in silico analysis of the genome of a septicemic APEC O1 strain E516 identified two putative iron transport genes homologous to the c2515 and c2516 genes from uropathogenic E. coli CFT073. In this study, we constructed the single and double gene deletion mutants, and studied their biological characteristic and pathogenic traits through in vitro and in vivo assays. Reverse transcriptase PCR (RT-PCR) analyses demonstrated that the mutations destroying the reading frame of the target genes abolished their transcription. Deletion of the single or double genes of c2515 and c2516 in APEC E516 weakened its ability to produce siderophore. Consistently, the mutants exhibited growth defect under iron-depleted conditions and the intracellular iron levels in the mutants were decreased in comparison with that of the wild-type (WT). Cell infection assays showed that the iron uptake defective mutants were more easily eliminated by the macrophage. Inactivation of the c2515 and c2516 genes affected bacterial colonization of chicken tissues, as well as the 50% lethal dose levels compared with the WT strain. Moreover, the expression levels of several iron uptake-related genes were significantly decreased in the double-deletion mutant. In total, the c2515 and c2516 may involve in siderophore-mediated iron uptake and participate in the pathogenesis of APEC O1 strain E516.

BackgroundThe Glycine N-acyltransferase (GLYAT) gene encodes a protein that catalyzes the transfer of acyl groups from acyl CoA to glycine, resulting in acyl glycine and coenzyme A. Aberrant GLYAT expression is associated with several malignant tumors, but its clinical importance in human breast cancer (BC), has yet to be fully addressed. This study aims to evaluate the clinical function of GLYAT in BC patients.MethodsGLYAT expression was determined by immune blot and immunohistochemistry in three BC cell lines and primary cancer tissues. The MDA-MB 231 cell line was used for GLYAT gene knockdown experiments while the MCF7 cell line for overexpression experiments. Colony formation experiments, soft agar experiments, and transwell assays were utilized for further inspection of cell proliferation and migration capabilities. Immunofluorescence and western blot were used to detect markers of the epithelial-mesenchymal transition (EMT) and changes in the PI3K/AKT/Snail pathway. The role of GLYAT in tumor growth and metastasis was also assessed in nude mice in vivo. Also, a correlation analysis was performed between clinicopathological features and GLYAT expression in BC patients.ResultsGLYAT was decreased in human BC tissues and cell lines. Functional analysis showed that knockdown of GLYAT augmented BC cell proliferation in vitro and in vivo. However, this phenomenon was reversed when GLYAT was overexpressed in the transfected cells. Moreover, downregulation of GLYAT promoted the migratory properties of BC cells, likely through the activation of PI3K/AKT/Snail signaling, which subsequently induced the EMT. IHC analysis indicated that GLYAT was decreased in human BC tissues and lower GLYAT expression was correlated with histological grade, tumor TNM stage, Ki-67 status, and poorer survival in BC patients. Furthermore, lower GLYAT expression seemed as an independent risk factor related to poor prognosis in BC patients based on Cox regression analyses.ConclusionOur findings demonstrate that downregulation of GLYAT expression in human breast cancer is correlated with EMT via the PI3K/AKT/Snail pathway and is also associated with histological grade, tumor TNM stage, Ki-67 status, and poor survival in breast cancer patients.

Ovarian cancer is considered as one of the most fatal gynecologic malignancies. This work aimed to explore the effects and regulatory mechanism of Acyl-CoA medium-chain synthetase-3 (ACSM3, a subunit of CoA ligases) in ovarian cancer progression. As well as employing CCK-8 assay, clone formation assay, and cell cycle analysis were carried out to investigate cell proliferation ability. Wound healing assay and transwell assay were subsequently used to assess cell migration and invasion. Mice xenografts were then conducted to measure the effects of ACSM3 on tumor development in vivo. Our bioinformatics analysis suggested that the expression of ACSM3 was down-regulated in ovarian cancer tissues, and the low expression level of ACSM3 might related with poorer overall survival than high mRNA expression of ACSM3 in ovarian cancer patients. We artificially regulated the expression of ACSM3 to evaluate its effects on ovarian cancer malignant phenotypes. Our data revealed that the overexpression of ACSM3 inhibited cell proliferation, migration, and invasion of ovarian cancer cells. In contrast, the knock-down of ACSM3 received the opposite results. Our western blot results showed that the Integrin β1/AKT signaling pathway was negatively regulated by ACSM3 expression. Moreover, ACSM3 overexpression-induced suppression of cell migration and invasion activities were abolished by the overexpression of ITG β1 (Integrin β1). Additionally, the growth of ovarian cancer xenograft tumors was also repressed by the overexpression of ACSM3. And ACSM3 interference obtained the contrary effects in vivo. In summary, ACSM3 acts as a tumor suppressor gene and may be a potential therapeutic target of ovarian cancer.

The active regulation of the plant growth environment is a common method for optimizing plant yield and quality. In horticulture today, light quality control is carried out using photo-selective nets or membranes to improve the yield and quality of cultivated plants. In the present study, with natural light as the control (CK), we tested different photo-selective nets (white, WN; blue, BN; green, GN; yellow, YN; and red, RN) with 30% shade for characteristics of growth, development, quality, yield, photosynthesis, and chlorophyll fluorescence, considering the antioxidant system, as well as the influence of element absorption and transformation of green onion (Allium fistulosumL.) plants at different growth stages. We found that plants under BN and WN have greater height and fresh weight than those of plants under the other nets. Plants under the BN treatment had the highest quality, yield, photosynthetic pigment content, net photosynthetic rate, transpiration rate, and stomatal conductance, whereas the intercellular CO2concentration was the highest in plants in the YN treatment. The photosynthesis noon break phenomenon was significantly lower in plants with covered photo-selective nets than in CK plants. NPQ was the highest in the YN treatment, and Fv/Fm, ΦPSII, and qP among the plants in the other treatments were different; from highest to lowest, they were as follows: BN > WN > CK > RN > GN > YN. The active oxygen content of green onion leaves in the BN treatment was significantly lower than that in the other treatments, and their key enzyme activity was significantly increased. BN also improved the absorption and transformation of elements in various organs of green onion.

In mammals, sperm-borne regulators can be transferred to oocytes during fertilization and have different effects on the formation of pronuclei, the first cleavage of zygotes, the development of preimplantation embryos and even the metabolism of individuals after birth. The regulatory role of sperm microRNAs (miRNAs) in the development of bovine preimplantation embryos has not been reported in detail. By constructing and screening miRNA expression libraries, we found that miR-202 was highly enriched in bovine sperm. As a target gene of miR-202, co-injection of SEPT7 siRNA can partially reverse the accelerated first cleavage of bovine embryos caused by miR-202 inhibitor. In addition, both a miR-202 mimic and SEPT7 siRNA delayed the first cleavage of somatic cell nuclear transfer (SCNT) embryos, suggesting that miR-202-SEPT7 mediates the delay of first cleavage of bovine embryos. By further exploring the relationship between miR-202/SEPT7, HDAC6 and acetylated α-tubulin during embryonic development, we investigated how sperm-borne miR-202 regulates the first cleavage process of bovine embryos by SEPT7 and demonstrate the potential of sperm-borne miRNAs to improve the efficiency of SCNT.