Anti-arrhythmia potential of honey-processed licorice in zebrafish model: Antioxidant, histopathological and tissue distribution.

ETHNOPHARMACOLOGICAL RELEVANCE: Honey-processed licorice (HPL) is the roasted product of licorice. It is recorded in the "Shang Han Lun" that licorice has better protection on heart after honey-processed. However, researches regarding its protective effect on the heart and the distribution of HPL in vivo are still limited. AIM OF THE STUDY: To evaluate the cardio-protection of HPL and explore the law of ten main components distribution in vivo under physiological and pathological conditions for an attempt to clarify the pharmacological substance basis of HPL in treating arrhythmia. MATERIALS AND METHODS: The adult zebrafish arrhythmia model was established by doxorubicin (DOX). Electrocardiogram (ECG) was used to detect the heart rate changes of zebrafish. SOD and MDA assays were used to evaluate oxidative stress levels in the myocardium. HE staining was used to observe the morphological change of myocardial tissues after HPL treatment. The UPLC-MS/MS was adapted to detect the content of ten main components of HPL in heart, liver, intestine, and brain under normal and heart injury conditions. RESULTS: Heart rate of zebrafish was decreased, the SOD activity was attenuated and MDA content was increased in myocardium after administration of DOX. Moreover, tissue vacuolation and inflammatory infiltration were detected in zebrafish myocardium induced by DOX. HPL could ameliorate heart injury and bradycardia induced by DOX to a certain extent by increasing SOD activity and reducing MDA content. In addition, the study of tissue distribution revealed that the content of liquiritin, isoliquiritin, and isoliquiritigenin in the heart was higher in the presence of arrhythmias than those in the normal condition. Under pathological conditions, the heart highly exposed to these three components could elicit anti-arrhythmic effects by regulating immunity and oxidation. CONCLUSION: These findings indicate that the HPL is protective against heart injury induced by DOX, and its effect is associated with the alleviation of oxidative stress and tissue injury. And the cardioprotective effect of HPL under pathological conditions may be related to the high distribution of liquiritin, isoliquiritin, and isoliquiritigenin in heart tissue. This study provides an experimental basis for the cardioprotective effects and tissue distribution of HPL.

Deoxynivalenol Biosynthesis in Fusarium pseudograminearum Significantly Repressed by a Megabirnavirus.

Deoxynivalenol (DON) is a mycotoxin widely detected in cereal products contaminated by Fusarium. Fusarium pseudograminearum megabirnavirus 1 (FpgMBV1) is a double-stranded RNA virus infecting Fusarium pseudograminearum. In this study, it was revealed that the amount of DON in F. pseudograminearum was significantly suppressed by FpgMBV1 through a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) assay. A total of 2564 differentially expressed genes were identified by comparative transcriptomic analysis between the FpgMBV1-containing F. pseudograminearum strain FC136-2A and the virus-free strain FC136-2A-V-. Among them, 1585 genes were up-regulated and 979 genes were down-regulated. Particularly, the expression of 12 genes (FpTRI1, FpTRI3, FpTRI4, FpTRI5, FpTRI6, FpTRI8, FpTRI10, FpTRI11, FpTRI12, FpTRI14, FpTRI15, and FpTRI101) in the trichothecene biosynthetic (TRI) gene cluster was significantly down-regulated. Specific metabolic and transport processes and pathways including amino acid and lipid metabolism, ergosterol metabolic and biosynthetic processes, carbohydrate metabolism, and biosynthesis were regulated. These results suggest an unrevealing mechanism underlying the repression of DON and TRI gene expression by the mycovirus FpgMBV1, which would provide new methods in the detoxification of DON and reducing the yield loss in wheat.

The first report of Lasiodiplodia theobromae causing leaf spot on Illicium verum in China.

Star anise (Illicium verum) production is an important industry in parts of southern China (Wang et al. 2011). The production of star anise (cultivar Dahong) was seriously affected by a leaf spot disease during the summer of 2020 in Rong County, Guangxi province, China. Approximately 20% to 30% of the trees (n = 200, 15-16 years old) had obvious leaf spots on more than 80% of the leaves. Local growers had to cut down the seriously diseased trees. Symptomatic samples were collected and sent to us at the end of August 2021. A single brown-edged round gray spot appeared on each leaf. The spot was ~20 mm in diameter. The margin was cut into 5 mm pieces, then disinfested with 3% NaOCl for 30 s, 75% ethanol for 30 s and sterile deionized water for 1 min before dried and placed on potato dextrose agar (PDA) medium at 25°C in the dark. A total of 25 fungal isolates were obtained (isolation rate 82%). Genomic DNAs was extracted from the mycelia of these isolates and three diagnostic regions including ITS1/ITS4 (White et al. 1990), EF1-728F/EF1-986R (Carbone and Kohn 1999) and Bt2a/Bt2b (Glass and Donaldson 1995) were amplified. The colony morphology on PDA and the sequences of the five isolates BJ20-1, BJ20-4, BJ20-5, BJ20-7, BJ20-8 were identical. Fungal colonies had light gray mycelium and black pigment on PDA. The average colony growth rate was 4.25±0.31 cm per day and no spores were produced. Sequences of the representative isolate BJ20-1 were deposited in GenBank (Accession nos. OK483326, OL547596 and OL547597). BLASTn search indicated high identity 99.6%, 98.58% and 100% to ITS (AY640255), EF1-α (AY640258), and ß-tubulin (KU887532) of Lasiodiplodia theobromae, respectively. Combined phylogenetic analysis using MEGAX (Kumar et al. 2018) clustered BJ20-1 and L. theobromae CBS164.96 in one clade. To test pathogenicity, 2-years-old healthy I. verum trees (cultivar Dahong) maintained in a greenhouse were inoculated. Leaves were surface sterilized with 70% ethanol. One PDA plug (5mm in diameter) was placed on each wound acupunctured with a sterile needle pick. Ten PDA plugs with mycelial of BJ20-1 growing 7 days on it were inoculated on five leaves. Four sterile PDA plugs placed on two leaves served as controls. All the plugs were removed from the leaves the day after inoculation. The experiment was repeated three times. At 1 day post-inoculation (DPI), brown expanding lesions were observed on the inoculated leaves. At 7 DPI, a mature ellipse of necrosis formed with 18±4 mm in diameter with black pycnidia in the center. Conidia were observed in the pycnidia. The immature conidia were thick-walled, hyaline, aseptate and ellipsoid, measuring 20 - 25.2 × 11 - 13 µm (n = 25). The mature conidia were dark brown with one central septum, measuring 24.3 - 27 × 13 - 14 µm (n = 25). At 10 DPI, the control leaves remained asymptomatic. Re-isolation was successful from the spot on the inoculated leaves. The colony morphology and molecular identification of the re-isolations were all the same as that of BJ20-1. In conclusion, the morphological and molecular evidence consistently indicated these isolates were L. theobromae. Koch's postulates were fulfilled that L. theobromae was pathogenic on star anise. Although L. theobromae has been reported to cause leaf spot disease on Camellia sinensis (Bao et al, 2021), Kadsura longipedunculata (Fan et al, 2020) and Broussonetia papyrifera (Luo et al, 2020), etc., this is the first report of L. theobromae causing leaf spot on I. verum in China. Due to the leaf spot disease resulting in serious yield reduction on star anise, accurate pathogen identification in this study would significantly improve the control of the leaf spot disease on star anise.

Identification and functional characterization of sex pheromone receptors in mirid bugs (Heteroptera: Miridae).

Mirid bugs are a group of important insect pests that cause large annual losses in agricultural production. Many studies have focused on the isolation and identification of sex pheromones in mirid bugs, and the components and biological activity of the sex pheromones have also been studied as a way to control these pests. However, few studies have focused on the mechanisms of pheromone perception. In this study, we identified the odorant receptor repertoire in three mirid bug species, Apolygus lucorum, Adelphocoris lineolatus, and Adelphocoris suturalis using antennal transcriptome sequencing and bioinformatics analysis. The candidate pheromone receptor (PR) genes were then identified by comparative transcriptomic and expression pattern analysis. Importantly, in vitro functional studies have shown that the candidate PRs have robust responses to the main mirid bug sex pheromone components (E)-2-hexenyl butyrate (E2HB) and hexyl butyrate (HB). Our study uncovered the mechanism of pheromone peripheral coding in these three species and elucidated the mechanism by which mirid bugs can specifically recognize a mate. Moreover, the results of our study will provide a theoretical basis for screening effective sex attractants or mating disturbance agents at the molecular and neural levels for enhanced control of these destructive pests.

The olfactory coreceptor IR8a governs larval feces-mediated competition avoidance in a hawkmoth.

Finding a suitable oviposition site is a challenging task for a gravid female moth. At the same time, it is of paramount importance considering the limited capability of most caterpillars to relocate to alternative host plants. The hawkmoth, Manduca sexta (Sphingidae), oviposits on solanaceous plants. Larvae hatching on a plant that is already attacked by conspecific caterpillars can face food competition, as well as an increased exposure to predators and induced plant defenses. Here, we show that feces from conspecific caterpillars are sufficient to deter a female M. sexta from ovipositing on a plant and that this deterrence is based on the feces-emitted carboxylic acids 3-methylpentanoic acid and hexanoic acid. Using a combination of genome editing (CRISPR-Cas9), electrophysiological recordings, calcium imaging, and behavioral analyses, we demonstrate that ionotropic receptor 8a (IR8a) is essential for acid-mediated feces avoidance in ovipositing hawkmoths.

A Female-Biased Odorant Receptor from Apolygus lucorum (Meyer-Dür) Tuned to Some Plant Odors.

Apolygus lucorum (Meyer-Dür) (Hemiptera: Miridae) is a serious pest of cotton, jujube, grape and many other crops around the world. Understanding how olfactory information directs this insect to its host plants may provide environment-friendly approaches to the control of its population in agriculture. In our study, we cloned an odorant receptor gene, AlucOR46, that was specifically expressed in antennae and female-biased. Functional expression of AlucOR46 in Xenopus oocytes showed that it is tuned to six plant volatiles (S)-(-)-Limonene, (R)-(+)-Limonene, (E)-2-Hexenal, (E)-3-Hexenol, 1-Heptanol and (1R)-(-)-Myrtenol. Electroantennogram (EAG) recordings revealed that all six compounds could elicit electrophysiological responses from the antennae of A. lucorum, higher in females. Our results are in agreement with previous reports showing that (E)-2-Hexenal could attract female A. lucorum in behavior experiments. These results suggest that AlucOR46 might play an important role in locating the host plants of A. lucorum and therefore represents a suitable target for green pest control.

An olfactory receptor from Apolygus lucorum (Meyer-Dur) mainly tuned to volatiles from flowering host plants.

Apolygus lucorum (Meyer-Dür) (Hemiptera: Miridae) is one of the most serious agricultural pests, feeding on a wide range of cultivated plants, including cotton, cereals and vegetables in the north of China. This insect can frequently switch between habitats and host plants over seasons and prefer plants in bloom. A. lucorum relies heavily on olfaction to locate its host plants finely discriminating different plant volatiles in the environment. Despite its economical importance, research on the olfactory system of this species has been so far very limited. In this study, we have identified and characterized an olfactory receptor which is sensitively tuned to (Z)-3-Hexenyl acetate and several flowering compounds. Besides being present in the bouquet of some flowers, these compounds are produced by plants that have suffered attacks and are supposed to act as chemical messengers between plants. This OR may play an important role in the selection of host plants.

Identification and functional characterization of sex pheromone receptors in the common cutworm (Spodoptera litura).

Male moths can finely discriminate the sex pheromone emitted by conspecific females from similar compounds. Pheromone receptors, expressed on the dendritic membrane of sensory neurons housed in the long trichoid sensilla of antennae, are thought to be associated with the pheromone reception. In this study, we identified and functionally characterized 4 pheromone receptors from the antennae of Spodoptera litura (Lepidoptera: Noctuidae). A tissue distribution analysis showed that the expression of the 4 SlituPRs was restricted to antennae. In addition, SlituOR6 and SlituOR13 were specifically expressed in male antennae whereas SlituOR11 and SlituOR16 were male-biased. Functional investigation by heterologous expression in Xenopus oocytes revealed that SlituOR6 was specifically tuned to the second major pheromone component, Z9,E12-14:OAc, SlituOR13 was equally tuned to Z9,E12-14:OAc and Z9-14:OAc, with a small response to the major pheromone component Z9,E11-14:OAc, SlituOR16 significantly responded to the behavioral antagonist Z9-14:OH, whereas SlituOR11 did not show response to any of the pheromone compounds tested in this study. Our results provide molecular data to better understand the mechanisms of sex pheromone detection in the moth S. litura and bring clues to investigate the evolution of the sexual communication channel in closely related species through comparison with previously reported pheromone receptors in other Spodoptera species.