Rapid detection of sacbrood virus (SBV) by one-step reverse transcription loop-mediated isothermal amplification assay

<p>Abstract</p> <p>Background</p> <p><it>Sacbrood virus </it>(SBV) primarily infects honeybee broods, and in order to deal with the problem cost effective detection methods are required.</p> <p>Findings</p> <p>A one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the rapid identification of SBV. The data demonstrated that, in a simple water bath, SBV RNA could be detected as early as 20 min at 65°C, and a positive amplification reaction was visible to the naked eye due to a color change brought on by the addition of nucleic acid stain SYBR Green.</p> <p>Conclusions</p> <p>The current study presents a method for the rapid and simple detection of SBV by RT-LAMP with high sensitivity and analytic specificity.</p

High throughput profiling of the cotton bollworm Helicoverpa armigera immunotranscriptome during the fungal and bacterial infections

Background: Innate immunity is essential in defending against invading pathogens in invertebrates. The cotton bollworm, Helicoverpa armigera (Hubner) is one of the most destructive lepidopteran pests, which causes enormous economic losses in agricultural production worldwide. The components of the immune system are largely unknown in this insect. The application of entomopathogens is considered as an alternative to the chemical insecticides for its control. However, few studies have focused on the molecular mechanisms of host-pathogen interactions between pest insects and their pathogens. Here, we investigated the immunotranscriptome of H. armigera larvae and examined gene expression changes after pathogen infections. This study provided insights into the potential immunity-related genes and pathways in H. armigera larvae.Results: Here, we adopted a high throughput RNA-seq approach to determine the immunotranscriptome of H. armigera larvae injected with buffer, fungal pathogen Beauveria bassiana, or Gram-negative bacterium Enterobacter cloacae. Based on sequence similarity to those homologs known to participate in immune responses in other insects, we identified immunity-related genes encoding pattern recognition receptors, signal modulators, immune effectors, and nearly all members of the Toll, IMD and JAK/STAT pathways. The RNA-seq data indicated that some immunity-related genes were activated in fungus- and bacterium-challenged fat body while others were suppressed in B. bassiana challenged hemocytes, including the putative IMD and JAK-STAT pathway members. Bacterial infection elevated the expression of recognition and modulator genes in the fat body and signal pathway genes in hemocytes. Although fat body and hemocytes both are important organs involved in the immune response, our transcriptome analysis revealed that more immunity-related genes were induced in the fat body than that hemocytes. Furthermore, quantitative real-time PCR analysis confirmed that, consistent with the RNA-seq data, the transcript abundances of putative PGRP-SA1, Serpin1, Toll-14, and Spz2 genes were elevated in fat body upon B. bassiana infection, while the mRNA levels of defensin, moricin1, and gloverin1 were up-regulated in hemocytes.Conclusions: In this study, a global survey of the host defense against fungal and bacterial infection was performed on the non-model lepidopteran pest species. The comprehensive sequence resource and expression profiles of the immunity-related genes in H. armigera are acquired. This study provided valuable information for future functional investigations as well as development of specific and effective agents to control this pest.Peer reviewedEntomology and Plant Patholog

Differential Effects Of Conifer And Broadleaf Litter Inputs On Soil Organic Carbon Chemical Composition Through Altered Soil Microbial Community Composition

A strategic selection of tree species will shift the type and quality of litter input, and subsequently magnitude and composition of the soil organic carbon (SOC) through soil microbial community. We conducted a manipulative experiment in randomized block design with leaf litter inputs of four native subtropical tree species in a Pinus massoniana plantation in southern China and found that the chemical composition of SOC did not differ significantly among treatments until after 28 months of the experiment. Contrasting leaf litter inputs had significant impacts on the amounts of total microbial, Gram-positive bacterial, and actinomycic PLFAs, but not on the amounts of total bacterial, Gramnegative bacterial, and fungal PLFAs. There were significant differences in alkyl/O-alkyl C in soils among the leaf litter input treatments, but no apparent differences in the proportions of chemical compositions (alkyl, O-alkyl, aromatic, and carbonyl C) in SOC. Soil alkyl/O-alkyl C was significantly related to the amounts of total microbial, and Gram-positive bacterial PLFAs, but not to the chemical compositions of leaf litter. Our findings suggest that changes in forest leaf litter inputs could result in changes in chemical stability of SOC through the altered microbial community composition

Mitochondrial Calcium Transporters Mediate Sensitivity to Noise-Induced Losses of Hair Cells and Cochlear Synapses

Mitochondria modulate cellular calcium homeostasis by the combined action of the mitochondrial calcium uniporter (MCU), a selective calcium entry channel, and the sodium calcium exchanger (NCLX), which extrudes calcium from mitochondria. In this study, we investigated MCU and NCLX in noise-induced hearing loss (NIHL) using adult CBA/J mice and noise-induced alterations of inner hair cell (IHC) synapses in MCU knockout mice. Following noise exposure, immunoreactivity of MCU increased in cochlear sensory hair cells of the basal turn, while immunoreactivity of NCLX decreased in a time- and exposure-dependent manner. Inhibition of MCU activity via MCU siRNA pretreatment or the specific pharmacological inhibitor Ru360 attenuated noise-induced loss of sensory hair cells and synaptic ribbons, wave I amplitudes, and NIHL in CBA/J mice. This protection was afforded, at least in part, through reduced cleavage of caspase 9 (CC9). Furthermore, MCU knockout mice on a hybrid genetic CD1 and C57/B6 background showed resistance to noise-induced seizures compared to wild-type littermates. Owing to the CD1 background, MCU knockouts and littermates suffer genetic high frequency hearing loss, but their IHCs remain intact. Noise-induced loss of IHC synaptic connections and reduction of auditory brainstem response (ABR) wave I amplitude were recovered in MCU knockout mice. These results suggest that cellular calcium influx during noise exposure leads to mitochondrial calcium overload via MCU and NCLX. Mitochondrial calcium overload, in turn, initiates cell death pathways and subsequent loss of hair cells and synaptic connections, resulting in NIHL

The Role of Macrolide Antibiotics in Increasing Cardiovascular Risk

AbstractBackgroundLarge cohort studies provide conflicting evidence regarding the potential for oral macrolide antibiotics to increase the risk of serious cardiac events.ObjectivesThis study performed a meta-analysis to examine the link between macrolides and risk of sudden cardiac death (SCD) or ventricular tachyarrhythmias (VTA), cardiovascular death, and death from any cause.MethodsWe performed a search of published reports by using MEDLINE (January 1, 1966, to April 30, 2015) and EMBASE (January 1, 1980, to April 30, 2015) with no restrictions. Studies that reported relative risk (RR) estimates with 95% confidence intervals (CIs) for the associations of interest were included.ResultsThirty-three studies involving 20,779,963 participants were identified. Patients taking macrolides, compared with those who took no macrolides, experienced an increased risk of developing SCD or VTA (RR: 2.42; 95% CI: 1.61 to 3.63), SCD (RR: 2.52; 95% CI: 1.91 to 3.31), and cardiovascular death (RR: 1.31; 95% CI: 1.06 to 1.62). No association was found between macrolides use and all-cause death or any cardiovascular events. The RRs associated with SCD or VTA were 3.40 for azithromycin, 2.16 for clarithromycin, and 3.61 for erythromycin, respectively. RRs for cardiovascular death were 1.54 for azithromycin and 1.48 for clarithromycin. No association was noted between roxithromycin and adverse cardiac outcomes. Treatment with macrolides is associated with an absolute risk increase of 118.1 additional SCDs or VTA, and 38.2 additional cardiovascular deaths per 1 million treatment courses.ConclusionsAdministration of macrolide antibiotics is associated with increased risk for SCD or VTA and cardiovascular death but not increased all-cause mortality

Robust in vitro assay for analyzing the neutralization activity of serum specimens against hepatitis B virus.

Anti-HBs is a well-known marker of protective capability against HBV. However, little is known about the association between the qAnti-HBs determined by immunoassays and the neutralization activity (NAT) derived from functional assays. We developed an in vitro assay for direct measurement of the NAT of human sera. The new assay was highly sensitive, with an analytical sensitivity of 9.6 ± 1.3 mIU/mL for the HBIG standard. For serum detection, the maximum fold dilution required to produce ≥50% inhibition (MDF50) of HBV infection was used as the quantitative index. In vitro NAT evaluations were conducted for a cohort of 164 HBV-free healthy individuals. The results demonstrated that the NAT positively correlated with the qAnti-HBs ( R 2 = 0.473, p < 0.001). ROC analysis indicated that the optimal cutoff value of the qAnti-HBs to discriminate significant NAT (MDF50 ≥ 8) was 62.9 mIU/mL, with an AUROC of 0.920. Additionally, we found that the qAnti-HBc was another independent parameter positively associated with the NAT ( R 2 = 0.300, p < 0.001), which suggested that antibodies against other HBV proteins generated by previous HBV exposure possibly also contribute to the NAT. In summary, the new cell-based assay provides a robust tool to analyse the anti-HBV NAT. Abbreviations: HBV: Hepatitis B virus; HBsAg: Hepatitis B surface antigen; Anti-HBs: Hepatitis B surface antibody; HBeAg: Hepatitis B e antigen; Anti-HBc: Hepatitis B core antibody; qAnti-HBs: quantitative hepatitis B surface antibody; qAnti-HBc: quantitative hepatitis B core antibody; qHBeAg: quantitative hepatitis B e antigen; NAT: neutralization activity; HBIG: hepatitis B immune globulin; NTCP: Na+-taurocholate cotransporting polypeptide; IRES: internal ribosome entry site; ccHBV: cell culture derived hepatitis B virus; GE/cell: genome equivalent per cell; MOI: multiplicity of infection; Dpi: day post infection; HepG2-TetOn: a HepG2-derived cell line that expresses the doxycycline-regulated transactivator; ROC: receiver operating characteristic curve; AUROC: area under receiver operating characteristic curve; LLOQ: the lower limits of quantification; MDF50: the maximum fold dilution required to produce ≥50% inhibition; IC50: half maximal inhibitory concentration

Drought-induced site-specific DNA methylation and its association with drought tolerance in rice (Oryza sativa L.)

An indica pyramiding line, DK151, and its recurrent parent, IR64, were evaluated under drought stress and non-stress conditions for three consecutive seasons. DK151 showed significantly improved tolerance to drought. The DNA methylation changes in DK151 and IR64 under drought stress and subsequent recovery were assessed using methylation-sensitive amplified polymorphism analysis. Our results indicate that drought-induced genome-wide DNA methylation changes accounted for ∼12.1% of the total site-specific methylation differences in the rice genome. This drought-induced DNA methylation pattern showed three interesting properties. The most important one was its genotypic specificity reflected by large differences in the detected DNA methylation/demethylation sites between DK151 and IR64, which result from introgressed genomic fragments in DK151. Second, most drought-induced methylation/demethylation sites were of two major types distinguished by their reversibility, including 70% of the sites at which drought-induced epigenetic changes were reversed to their original status after recovery, and 29% of sites at which the drought-induced DNA demethylation/methylation changes remain even after recovery. Third, the drought-induced DNA methylation alteration showed a significant level of developmental and tissue specificity. Together, these properties are expected to have contributed greatly to rice response and adaptation to drought stress. Thus, induced epigenetic changes in rice genome can be considered as a very important regulatory mechanism for rice plants to adapt to drought and possibly other environmental stresses

Suppress HBV by therapeutic vaccine

乙肝预防性疫苗显著减少了乙肝新发感染，但目前全球仍有约2.5亿慢性乙肝感染者，若未得到有效治疗，可能发展为肝癌、肝硬化等终末期肝病并导致死亡。夏宁邵教授团队研究发展了一种新型的B细胞表位嵌合型类病毒颗粒乙肝治疗性疫苗（治疗性蛋白），在多种模型中证实了其对慢性乙肝感染的治疗潜力，为研发治疗慢性乙肝的原创药物提供了新思路。 我校博士后张天英、博士生郭雪染和博士生巫洋涛为该论文共同第一作者，夏宁邵教授、袁权副教授、张军教授为该论文的共同通讯作者。【Abstract】Objective: This study aimed to develop a novel therapeutic vaccine based on a unique B cell epitope and investigate its therapeutic potential against chronic hepatitis B (CHB) in animal models. Methods: A series of peptides and carrier proteins were evaluated in HBV-tolerant mice to obtain an optimized therapeutic molecule. The immunogenicity,therapeutic efficacy and mechanism of the candidate were investigated systematically. Results: Among the HBsAg-aa119-125-containing peptides evaluated in this study, HBsAg-aa113-135 (SEQ13) exhibited the most striking therapeutic effects. A novel immuno-enhanced virus-like particle carrier (CR-T3) derived from the roundleaf bat HBV core antigen (RBHBcAg) was created and used to display SEQ13, forming candidate molecule CR-T3-SEQ13. Multiple copies of SEQ13 displayed on the surface of this particulate antigen promote the induction of a potent anti-HBs antibody response in mice, rabbits and cynomolgus monkeys. Sera and purified polyclonal IgG from the immunized animals neutralized HBV infection in vitro and mediated efficient HBV/HBsAg clearance in the mice. CR-T3-SEQ13-based vaccination induced long-term suppression of HBsAg and HBV DNA in HBV transgenic mice and eradicated the virus completely in hydrodynamic-based HBV carrier mice. The suppressive effects on HBsAg were strongly correlated with the anti-HBs level after vaccination, suggesting that the main mechanism of CR-T3-SEQ13 vaccination therapy was the induction of a SEQ13-specific antibody response that mediated HBV/HBsAg clearance. Conclusions: The novel particulate protein CR-T3-SEQ13 suppressed HBsAg effectively through induction of a humoral immune response in HBV-tolerant mice. This B cell epitope-based therapeutic vaccine may provide a novel immunotherapeutic agent against chronic HBV infection in humans.This work was supported by the National Scientific and Technological Major project (2017ZX10202203-001), the National Natural Science Foundation of China (31730029, 81672023, 81871316 and 81702006) and the Xiamen University President Fund Project (20720160063). 该研究获得了“艾滋病和病毒性肝炎等重大传染病防治”科技重大专项、国家自然科学基金等资助