Envelope domain III E324, E351, and E380 mutations lever adaptive evolution of DENV-1 genotype I.

Dengue virus (DENV) gains genetic mutations during continuous transmission and evolution, making the virus more adaptive and virulent. The clade of DENV-1 genotype I has expanded and become the predominant genotype in Asia and the Pacific areas, but the underlying mechanisms are unclear. A combined analysis of nonsynonymous mutations in domain III of the envelope protein and their biological effects on virus pathogenesis and transmission was evaluated. Phylogenetic analyses found three nonsynonymous mutations (V324I, V351L, and V380I) in domain III of the envelope protein, which emerged in 1970s-1990s and stably inherited and expanded in contemporary strains after 2000. We generated reverse-mutated viruses (I324V, L351V, and I380V) based on an infectious clone of an epidemic DENV-1 strain (NIID02-20), and the results suggested that the infectivity of the contemporary epidemic virus (wild type, WT) has increased compared to the reverse mutant viruses in mammalian hosts but not mosquito vectors. The WT virus showed a higher binding affinity to host cells and increased virion stability. In addition, weaker immunogenicity and higher resistance to neutralizing antibodies of the WT virus indicated a trend of immune escape. The data suggested that nonsynonymous mutations of the E protein (V324I, V351L, and V380I) promote infectivity and immune evasion of DENV-1 genotype I, which may facilitate its onward transmission on a global scale. IMPORTANCE: We provide evidence that minor sequence variation among dengue virus (DENV) strains can result in increased adaptability and virulence, impacting both the biology of the virus and the antiviral immune response. The genetic mutations of DENV-1 gained during continuous transmission and evolution will offer new clues for the design of novel vaccines against flaviviruses.

Unveiling the seroprevalence of human papillomavirus in Guangdong, China: Implications for vaccination strategies.

Seroepidemiological characteristics of human papillomavirus (HPV) in community residents reflect natural infection and can guide the reform of vaccination programs. A population-based serological survey was conducted in Guangdong Province. Serum anti-HPV IgG antibody levels were determined by an ELISA. Neutralizing antibodies against HPV6, 11, 16, and 18 were detected via a pseudovirus-based neutralization assay (PBNA). A total of 5122 serum samples were collected from community residents, including 1989 males and 3133 females, in three cities of Guangdong Province. The rate of HPV IgG antibody positivity in females was 5.39% (95% CI: 4.6-6.2), which was greater than that in males (2.36%; 95% CI: 1.7-3.1). HPV IgG antibodies were more frequently detected in females aged 51-60 years (11.30%; 95% CI: 7.6-16.0), whereas in males, the detection increased with age and reached 4.94% (95% CI: 2.8-6.9) in the group aged ≥71 years. The seropositivity of neutralizing antibodies against HPV6 and 11 was greater than that against HPV16 and 18. The serum neutralizing antibody titers in individuals who received three doses of a vaccine were 7- to 12-fold greater than those in individuals who did not receive the vaccine. The neutralizing antibody titers slightly decreased within 40 months and ranged from 0.038 to 0.057 log ED50 per month. A moderate consistency between the HPV ELISA and PBNA results was observed (Kappa score = 0.49, r = 0.249, 0.635, 0.382, and 0.466 for HPV6, 11, 16, and 18, respectively). The HPV seropositivity rate among healthy residents of Guangdong Province was found to be low among children and adolescents and to increase with age. The serum neutralizing antibody titers were significantly greater in the vaccine group than that in the control group, and this difference persisted over time, which indicated promising protection against HPV infection.

[Hypoxia promotes lipopolysaccharide-induced CXCL10 expression in microglia].

This study was aimed to investigate the effect of hypoxia on lipopolysaccharide (LPS)-induced CXC-chemokine ligand-10 (CXCL10) expression and the underlying mechanism. C57BL/6J mice were randomly divided into control, hypoxia, LPS, and hypoxia combined with LPS groups. The LPS group was intraperitoneally injected with 0.5 mg/kg LPS, and the hypoxia group was placed in a hypobaric hypoxia chamber (simulated altitude of 6 000 m). The serum and hippocampal tissue samples were collected after 6 h of the treatment. The levels of CXCL10 in the serum and hippocampal tissue of mice were detected by ELISA. The microglia cell line BV2 and primary microglia were stimulated with hypoxia (1% O2) and/or LPS (100 ng/mL) for 6 h. The mRNA expression level of CXCL10 and its content in culture supernatant were detected by real-time quantitative PCR and ELISA, respectively. The phosphorylation levels of nuclear factor κB (NF-κB) signaling pathway-related proteins, p65 and IκBα, were detected by Western blot. Moreover, after NF-κB signaling pathway being blocked with a small molecular compound, PDTC, CXCL10 mRNA expression level was detected in the BV2 cells. The results showed that in the LPS-induced mouse inflammatory model, hypoxia treatment could promote LPS-induced up-regulation of CXCL10 in both serum and hippocampus. Compared with the cells treated with LPS alone, the expression of CXCL10 mRNA and the content of CXCL10 in the culture supernatant of BV2 cells treated with hypoxia combined with LPS were significantly increased. The CXCL10 mRNA level of primary microglial cells treated with hypoxia combined with LPS was significantly up-regulated. Compared with the cells treated with hypoxia or LPS alone, the phosphorylation levels of p65 and IκBα in the BV2 cells treated with hypoxia combined with LPS were significantly increased. PDTC blocked the induction of CXCL10 gene expression by LPS in the BV2 cells. These results suggest that hypoxia promotes LPS-induced expression of CXCL10 in both animal and cell models, and NF-κB signaling pathway plays an important role in this process.

Mass spectrometry imaging: An emerging technology for the analysis of metabolites in insects.

Mass spectrometry imaging (MSI) can visualize the composition, abundance, and spatial distribution of molecules in tissues or cells, which has been widely used in the research of life science. Insects, especially the agricultural pests, have received a great deal of interests from the scientists in biodiversity and food security. This review introduces the major characteristics of MSI, summarizes its application to the investigation of insect endogenous metabolites, exogenous metabolites, and the spatiotemporal changes of metabolites between insects and plants, and discusses its shortfalls and perspectives. The significance of these concerns is beneficial for future insect research such as physiology and metabolism.

Differential Profiles of Gut Microbiota and Metabolites Associated with Host Shift of Plutella xylostella.

Evolutionary and ecological forces are important factors that shape gut microbial profiles in hosts, which can help insects adapt to different environments through modulating their metabolites. However, little is known about how gut microbes and metabolites are altered when lepidopteran pest species switch hosts. In the present study, using 16S-rDNA sequencing and mass spectrometry-based metabolomics, we analyzed the gut microbiota and metabolites of three populations of Plutella xylostella: one feeding on radish (PxR) and two feeding on peas (PxP; with PxP-1 and PxP-17 being the first and 17th generations after host shift from radish to peas, respectively). We found that the diversity of gut microbes in PxP-17 was significantly lower than those in PxR and PxP-1, which indicates a distinct change in gut microbiota after host shift. Kyoto Encyclopedia of Genes and Genomes analysis revealed that the functions of energy metabolism, signal transduction, and xenobiotics biodegradation and metabolism were increased in PxP-17, suggesting their potential roles in host adaptation. Metabolic profiling showed a significant difference in the abundance of gut metabolites between PxR and PxP-17, and significant correlations of gut bacteria with gut metabolites. These findings shed light on the interaction among plants, herbivores, and symbionts, and advance our understanding of host adaptation associated with gut bacteria and metabolic activities in P. xylostella.

Pre-Service Teachers' GenAI Anxiety, Technology Self-Efficacy, and TPACK: Their Structural Relations with Behavioral Intention to Design GenAI-Assisted Teaching.

Generative artificial intelligence (GenAI) has taken educational settings by storm in the past year due to its transformative ability to impact school education. It is crucial to investigate pre-service teachers' viewpoints to effectively incorporate GenAI tools into their instructional practices. Data gathered from 606 pre-service teachers were analyzed to explore the predictors of behavioral intention to design Gen AI-assisted teaching. Based on the Unified Theory of Acceptance and Use of Technology (UTAUT) model, this research integrates multiple variables such as Technological Pedagogical Content Knowledge (TPACK), GenAI anxiety, and technology self-efficacy. Our findings revealed that GenAI anxiety, social influence, and performance expectancy significantly predicted pre-service teachers' behavioral intention to design GenAI-assisted teaching. However, effort expectancy and facilitating conditions were not statistically associated with pre-service teachers' behavioral intentions. These findings offer significant insights into the intricate relationships between predictors that influence pre-service teachers' perspectives and intentions regarding GenAI technology.

Prolyl hydroxylase inhibitor FG-4592 alleviates neuroinflammation via HIF-1/BNIP3 signaling in microglia.

BACKGROUND: Neuroinflammation is responsible for neuropsychiatric dysfunction following acute brain injury and neurodegenerative diseases. This study describes how a hypoxia-inducible factor prolyl hydroxylase (HIF-PHD) inhibitor FG-4592 prevents the lipopolysaccharide (LPS)-induced acute neuroinflammation in microglia. METHODS: The distribution of FG-4592 in mouse brain tissues was determined by collision-induced dissociation tandem mass spectrometry. Microglial activation in the hippocampus was analyzed by immunofluorescence. Moreover, we determined the activation of HIF-1 and nuclear factor-κB (NF-κB) signaling pathways, proinflammatory responses using molecular biological techniques. Transcriptome sequencing and BNIP3 silencing were conducted to explore signaling pathway and molecular mechanisms underlying FG-4592 anti-inflammatory activity. RESULTS: FG-4592 was transported into the brain tissues and LPS increased its transportation. FG-4592 promoted the expression of HIF-1α and induced the downstream gene transcription in the hippocampus. Administration with FG-4592 significantly inhibited microglial hyperactivation and decreased proinflammatory cytokine levels following LPS treatment in the hippocampus. The LPS-induced inflammatory responses and the NF-κB signaling pathway were also downregulated by FG-4592 pretreatment in microglial cells. Mechanistically, Venn diagram analysis of transcriptomic changes of BV2 cells identified that BNIP3 was a shared and common differentially expressed gene among different treatment groups. FG-4592 markedly upregulated the protein levels of BNIP3 in microglia. Importantly, BNIP3 knockdown aggravated the LPS-stimulated inflammatory responses and partially reversed the protection of FG-4592 against microglial inflammatory signaling and microglial activation in the mouse hippocampus. CONCLUSIONS: FG-4592 alleviates neuroinflammation through facilitating microglial HIF-1/BNIP3 signaling pathway in mice. Targeting HIF-PHD/HIF-1/BNIP3 axis is a promising strategy for the development of anti-neuroinflammation drugs.

Enhanced immunity against SARS-CoV-2 in returning Chinese individuals.

Global COVID-19 vaccination programs effectively contained the fast spread of SARS-CoV-2. Characterizing the immunity status of returned populations will favor understanding the achievement of herd immunity and long-term management of COVID-19 in China. Individuals were recruited from 7 quarantine stations in Guangzhou, China. Blood and throat swab specimens were collected from participants, and their immunity status was determined through competitive ELISA, microneutralization assay and enzyme-linked FluoroSpot assay. A total of 272 subjects were involved in the questionnaire survey, of whom 235 (86.4%) were returning Chinese individuals and 37 (13.6%) were foreigners. Blood and throat swab specimens were collected from 108 returning Chinese individuals. Neutralizing antibodies against SARS-CoV-2 were detected in ~90% of returning Chinese individuals, either in the primary or the homologous and heterologous booster vaccination group. The serum NAb titers were significantly decreased against SARS-CoV-2 Omicron BA.5, BF.7, BQ.1 and XBB.1 compared with the prototype virus. However, memory T-cell responses, including specific IFN-γ and IL-2 responses, were not different in either group. Smoking, alcohol consumption, SARS-CoV-2 infection, COVID-19 vaccination, and the time interval between last vaccination and sampling were independent influencing factors for NAb titers against prototype SARS-CoV-2 and variants of concern. The vaccine dose was the unique common influencing factor for Omicron subvariants. Enhanced immunity against SARS-CoV-2 was established in returning Chinese individuals who were exposed to reinfection and vaccination. Domestic residents will benefit from booster homologous or heterologous COVID-19 vaccination after reopening of China, which is also useful against breakthrough infection.

Fungal Spectrum and Susceptibility Against Nine Antifungal Agents in 525 Deep Fungal Infected Cases.

Background: Deep fungal infection has become an important cause of infection and death in hospitalized patients, and this has worsened with increasing antifungal drug resistance. Objective: A 3-year retrospective study was conducted to investigate the clinical characteristics, pathogen spectrum, and drug resistance of deep fungal infection in a regional hospital of Guangzhou, China. Methods: Non-duplicate fungi isolates recovered from blood and other sterile body fluids of in-patients of the clinical department were identified using biochemical tests of pure culture with the API20C AUX and CHROMagar medium. Antifungal susceptibilities were determined by Sensititre YeastOne® panel trays. Results: In this study, 525 patients (283 female, 242 male) with deep fungal infection were included, half of them were elderly patients (≥60 years) (54.67%, n=286). A total of 605 non-repetitive fungi were finally isolated from sterile samples, of which urine specimens accounted for 66.12% (n=400). Surgery, ICU, and internal medicine were the top three departments that fungi were frequently detected. The mainly isolated fungal species were Candida albicans (43.97%, n=266), Candida glabrata (20.00%, n=121), and Candida tropicalis (17.02%, n=103), which contributed to over 80% of fungal infection. The susceptibility of the Candida spp. to echinocandins, 5-fluorocytosine, and amphotericin B remained above 95%, while C. glabrata and C. tropicalis to itraconazole were about 95%, and the dose-dependent susceptibility of C. glabrata to fluconazole was more than 90%. The echinocandins had no antifungal activity against Trichosporon asahi in vitro (MIC90>8 µg/mL), but azole drugs were good, especially voriconazole and itraconazole (MIC90 = 0.25 µg/mL). Conclusion: The main causative agents of fungal infection were still the genus of Candida. Echinocandins were the first choice for clinical therapy of Candida infection, followed with 5-fluorocytosine and amphotericin B. Azole antifungal agents should be used with caution in Candida glabrata and Candida tropicalis infections.

Refocus on Immunogenic Characteristics of Convalescent COVID-19 Challenged by Prototype SARS-CoV-2.

Background: Mass basic and booster immunization programs effectively contained the spread of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) virus, also known as COVID-19. However, the emerging Variants of Concern (VOCs) of COVID-19 evade the immune protection of the vaccine and increase the risk of reinfection. Methods: Serum antibodies of 384 COVID-19 cases recovered from SARS-CoV-2 infection were examined. Correlations between clinical symptoms and antibodies against VOCs were analyzed. Result: All 384 cases (aged 43, range 1−90) were from 15 cities of Guangdong, China. The specific IgA, IgG, and IgM antibodies could be detected within 4−6 weeks after infection. A broad cross-reaction between SARS-CoV-2 and Severe Acute Respiratory Syndrome Coronavirus, but not with Middle East Respiratory Syndrome Coronavirus was found. The titers of neutralization antibodies (NAbs) were significantly correlated with IgG (r = 0.667, p < 0.001), but showed poor neutralizing effects against VOCs. Age, fever, and hormone therapy were independent risk factors for NAbs titers reduction against VOCs. Conclusion: Humoral immunity antibodies from the original strain of COVID-19 showed weak neutralization effects against VOCs, and decreased neutralizing ability was associated with initial age, fever, and hormone therapy, which hindered the effects of the COVID-19 vaccine developed from the SARS-CoV-2 prototype virus.

Vaccination against Varicella Zoster Virus Infection in Less Developed Regions of Guangdong, China: A Cross-Sectional Serosurveillance Study.

Vaccination is the key to prevent varicella zoster virus (VZV) infection in children. Voluntary and self-funded strategies have led to variable vaccination rates against VZV in China. For low-income populations, in particular, the effects of VZV vaccination have been insufficiently estimated. Community-based serosurveillance was conducted in two less developed regions, Zhanjiang and Heyuan, of Guangdong, China. Anti-VZV IgG antibodies in serum were detected by ELISA. The vaccination data were derived from the Guangdong Immune Planning Information System. A total of 4221 participants were involved, of which 3377 were from three counties of Zhanjiang and the other 844 were from one county of Heyuan, Guangdong, China. The total VZV IgG seropositivity rate in vaccinated individuals was 34.30% and 42.76%, while it was 89.61% and 91.62% in non-vaccinated populations of Zhanjiang and Heyuan, respectively. The seropositivity rate increased gradually with age, reaching ~90% in the >20- to 30-year-old group. The VarV vaccination rates of children aged 1-14 years were 60.47% for one dose and 6.20% for two doses in Zhanjiang, and 52.24% for one dose and 4.48% for two doses in Heyuan. Compared with the non-vaccinated group (31.19%) and one-dose group (35.47%), the positivity rate of anti-VZV IgG antibodies was significantly higher in the two-dose group (67.86%). Before the VarV policy was reformed, the anti-VZV IgG positivity rate was 27.85% in the one-dose-vaccinated participants, which increased to 30.43% after October 2017. The high seroprevalence in participants was due to infection of VZV in Zhanjiang and Heyuan, not vaccination against VZV. Children aged 0-5 years are still vulnerable to varicella, so a two-dose vaccination program should be implemented to prevent onward transmission of VZV.

[Effects of blocking lactate production by 2-DG on hypoxic injury of HT22 neurons and its mechanisms].

OBJECTIVE: To investigate the effects of blocking lactate synthesis on the HT22 cell injuries caused by hypoxia. METHODS: 2-deoxy-D-glucose (2-DG) is a non-metabolized glucose analogue that can inhibit lactate synthesis by blocking glycolysis. HT22 cells were divided into 4 groups: Control group, 2-DG group, Hypoxia group and 2-DG+Hypoxia group. The cells in control group and 2-DG treatment group were cultured in a 37℃, 5% CO2 incubator, and thecells in hypoxia group and 2-DG + Hypoxia group were cultured in a hypoxia incubator. The concentrations of 2-DG were 2.5 and 5 mmol/L, the concentration of oxygen was 0.3%, and the treatment time was 24 h. Cell activity was detected by CCK-8 assay, the levels of lactate in cell culture medium were detected by spectrophotometry, cell morphology was observed by fluorescence staining, the level of reactive oxygen species (ROS) was detected by flow cytometry, and the activities of superoxide dismutase (SOD) and catalase (CAT) were determined by enzyme activity kits. The protein expression levels of p-p38, t-p38 and ß-actin were detected by Western blot. RESULTS: Compared with that in control group, the lactate level in culture medium and cell activity were decreased significantly (P＜0.01), the number of adherent cells was decreased, the level of ROS was increased (P＜0.01), and the enzyme activity of CAT was decreased (P＜0.05) in the 2-DG group. In the hypoxia group, the level of lactate in the culture medium was increased significantly (P＜0.01), the cell activity was decreased (P＜0.01), the number of adherent cells was decreased, the ROS levels were increased (P＜0.01), and the enzyme activities of CAT and SOD were decreased (P＜0.01 or P＜0.05). In 2-DG+Hypoxia group, the level of lactate was decreased significantly (P＜0.05), the cell viability was decreased significantly (P＜0.01), the number of cells was decreased significantly, and the ability of adhere to the wall was weakened significantly. The level of ROS was increased significantly (P＜0.01), the enzyme activities of CAT and SOD were decreased significantly (P＜0.01), the protein expression level of p-p38 was increased significantly (P＜0.05), and there was no change in t-p38. Compared with hypoxia groups, in 2-DG combined with hypoxia group, the level of lactate induced by hypoxia, the cell activity, and the enzyme activity level of CAT were decreased significantly (all P＜0.01), while the level of ROS was increased significantly (P＜ 0.01). CONCLUSION: Blocking lactate can reduce the cell activity level under hypoxia and aggravate the oxidative stress injury of HT22 cells. The mechanisms may be related to increasing ROS level and activating p38 signal pathway.

Fitness comparison of Plutella xylostella on original and marginal hosts using age-stage, two-sex life tables.

The diamondback moth, Plutella xylostella, is an important agricultural pest that severely damages cruciferous vegetables. Although previously considered a threat only to Brassica species, P. xylostella has been observed to feed on noncruciferous vegetables. Here, we established a population of P. xylostella on the pea Pisum sativum (PxP population). We compared this PxP population's performance on the pea host plant to a population (PxR) reared on the original host plant radish (Raphanus sativus) for several generations using an age-stage, two-sex life table and analyzed the correlations between different fitness parameters. In the 1st generation of the PxP population, survival rate of immature stage was 17%, while the survival rate of PxR was 68%; the duration of the 4th larval instar (5.30 d) and mortality (25%) of this generation were significantly longer (2.8 d) and higher (1%) than that of PxR, respectively (both p < .001). Upon long-term acclimation, the PxP fitness improved significantly, especially that the survival rate of immature stages increased to approximately 60% in the 15th, 30th, and 45th generations. However, PxP feeding on pea exhibited poorer fitness with longer larval developmental time, shorter total life span, lighter pupa, and lower fecundity in different generations compared with PxP feeding on radish. PxP feeding on pea also showed a significantly lower intrinsic rate of increase (r), net reproduction rate (R 0), finite increase rate (λ), and longer mean generation time (T) than PxP feeding on radish in all generations tested. Significant positive correlations were observed between pupal weight and female fecundity in pea-fed populations, and between female longevity and female fecundity in pea-fed and radish-fed populations. Our findings suggest that P. xylostella adaptation to pea does not improve overall fitness compared with the original host radish, making pea a marginal host for P. xylostella.

A two-step DNA barcoding approach for delimiting moth species: moths of Dongling Mountain (Beijing, China) as a case study.

DNA barcoding, based on a fragment of cytochrome c oxidase I (COI) mtDNA, is as an effective molecular tool for identification, discovery, and biodiversity assessment for most animals. However, multiple gene markers coupled with more sophisticated analytical approaches may be necessary to clarify species boundaries in cases of cryptic diversity or morphological plasticity. Using 339 moths collected from mountains surrounding Beijing, China, we tested a pipeline consisting of two steps: (1) rapid morphospecies sorting and screening of the investigated fauna with standard COI barcoding approaches; (2) additional analyses with multiple molecular markers for those specimens whose morphospecies and COI barcode grouping were incongruent. In step 1, 124 morphospecies were delimited into 116 barcode units, with 90% of the conflicts being associated with specimens identified to the genus Hypena. In step 2, 55 individuals representing all 12 Hypena morphospecies were analysed using COI, COII, 28S, EF-1a, Wgl sequences or their combinations with the BPP (Bayesian Phylogenetics and Phylogeography) multigene species delimitation method. The multigene analyses supported the delimitation of 5 species, consistent with the COI analysis. We conclude that a two-step barcoding analysis pipeline is able to rapidly characterize insect biodiversity and help to elucidate species boundaries for taxonomic complexes without jeopardizing overall project efficiency by substantially increasing analytical costs.