A nonstructural protein encoded by a rice reovirus induces an incomplete autophagy to promote viral spread in insect vectors.

Viruses can hijack autophagosomes as the nonlytic release vehicles in cultured host cells. However, how autophagosome-mediated viral spread occurs in infected host tissues or organs in vivo remains poorly understood. Here, we report that an important rice reovirus, rice gall dwarf virus (RGDV) hijacks autophagosomes to traverse multiple insect membrane barriers in the midgut and salivary gland of leafhopper vector to enhance viral spread. Such virus-containing double-membraned autophagosomes are prevented from degradation, resulting in increased viral propagation. Mechanistically, viral nonstructural protein Pns11 induces autophagy and embeds itself in the autophagosome membranes. The autophagy-related protein 5 (ATG5)-ATG12 conjugation is essential for initial autophagosome membrane biogenesis. RGDV Pns11 specifically interacts with ATG5, both in vitro and in vivo. Silencing of ATG5 or Pns11 expression suppresses ATG8 lipidation, autophagosome formation, and efficient viral propagation. Thus, Pns11 could directly recruit ATG5-ATG12 conjugation to induce the formation of autophagosomes, facilitating viral spread within the insect bodies. Furthermore, Pns11 potentially blocks autophagosome degradation by directly targeting and mediating the reduced expression of N-glycosylated Lamp1 on lysosomal membranes. Taken together, these results highlight how RGDV remodels autophagosomes to benefit viral propagation in its insect vector.

Risk assessment of environmental and bystander exposure from agricultural unmanned aerial vehicle sprayers in golden coconut plantations: Effects of droplet size and spray volume.

Unmanned aerial vehicle (UAV) sprayers are widely utilized in commercial aerial application of plant protection products (PPPs) in East Asian countries due to their high flexibility, high efficiency and low cost, but spray drift can lead to low utilization of UAV sprayers application, environmental pollution and bystander exposure risk. Droplet size and spray volume are critical factors affecting spray drift. Currently, the high temperature and humidity environment under the influence of the tropical monsoon climate brings new challenges for UAV sprayers. Therefore, in this study, pesticides were simulated with seduction red solution, and spraying trials were conducted using the DJI commercial T40 UAV sprayers for a typical tropical crop, coconut. In this study, the spray drift distribution of droplets on the ground and in the air, as well as the bystander exposure risk, were comparatively analyzed using droplet size (VF, M, and C) and spray volume (75 L/hm2 and 60 L/hm2) as trial variables. The results indicated that the spray drift characteristics of UAV sprayers were significantly affected by droplet size and spray volume. The spray drift percentage was negatively correlated with the downwind distance and the sampling height. The smaller the droplet size, the farther the buffer zone distance, up to more than 30 m, and the cumulative drift percentage is also greater, resulting in a significant risk of spray drift. The reduction in spray volume helped to reduce the spray drift, and the cumulative drift percentage was reduced by 73.87 % with a droplet size of M. The region of the body where spray drift is deposited the most on bystanders is near chest height. This study provides a reference for the rational and safe use of multirotor UAV sprayers application operations in the tropics and enriches the spray drift database in the tropics.

A plant reovirus hijacks endoplasmic reticulum-associated degradation machinery to promote efficient viral transmission by its planthopper vector under high temperature conditions.

In the field, many insect-borne crop viral diseases are more suitable for maintenance and spread in hot-temperature areas, but the mechanism remains poorly understood. The epidemic of a planthopper (Sogatella furcifera)-transmitted rice reovirus (southern rice black-streaked dwarf virus, SRBSDV) is geographically restricted to southern China and northern Vietnam with year-round hot temperatures. Here, we reported that two factors of endoplasmic reticulum-associated degradation (ERAD) machinery, the heat shock protein DnaJB11 and ER membrane protein BAP31, were activated by viral infection to mediate the adaptation of S. furcifera to high temperatures. Infection and transmission efficiencies of SRBSDV by S. furcifera increased with the elevated temperatures. We observed that high temperature (35°C) was beneficial for the assembly of virus-containing tubular structures formed by nonstructural protein P7-1 of SRBSDV, which facilitates efficient viral transmission by S. furcifera. Both DnaJB11 and BAP31 competed to directly bind to the tubule protein P7-1 of SRBSDV; however, DnaJB11 promoted whereas BAP31 inhibited P7-1 tubule assembly at the ER membrane. Furthermore, the binding affinity of DnaJB11 with P7-1 was stronger than that of BAP31 with P7-1. We also revealed that BAP31 negatively regulated DnaJB11 expression through their direct interaction. High temperatures could significantly upregulate DnaJB11 expression but inhibit BAP31 expression, thereby strongly facilitating the assembly of abundant P7-1 tubules. Taken together, we showed that a new temperature-dependent protein quality control pathway in the ERAD machinery has evolved for strong activation of DnaJB11 for benefiting P7-1 tubules assembly to support efficient transmission of SRBSDV in high temperatures. We thus deduced that ERAD machinery has been hitchhiked by insect-borne crop viruses to enhance their transmission in tropical climates.

The 5-oxoprolinase is required for conidiation, sexual reproduction, virulence and deoxynivalenol production of Fusarium graminearum.

In eukaryotic organisms, the 5-oxoprolinase is one of the six key enzymes in the γ-glutamyl cycle that is involved in the biosynthetic pathway of glutathione (GSH, an antioxidative tripeptide counteracting the oxidative stress). To date, little is known about the biological functions of the 5-oxoprolinase in filamentous phytopathogenic fungi. In this study, we investigated the 5-oxoprolinase in Fusarium graminearum for the first time. In F. graminearum, two paralogous genes (FgOXP1 and FgOXP2) were identified to encode the 5-oxoprolinase while only one homologous gene encoding the 5-oxoprolinase could be found in other filamentous phytopathogenic fungi or Saccharomyces cerevisiae. Deletion of FgOXP1 or FgOXP2 in F. graminearum led to significant defects in its virulence on wheat. This is likely caused by an observed decreased deoxynivalenol (DON, a mycotoxin) production in the gene deletion mutant strains as DON is one of the best characterized virulence factors of F. graminearum. The FgOXP2 deletion mutant strains were also defective in conidiation and sexual reproduction while the FgOXP1 deletion mutant strains were normal for those phenotypes. Double deletion of FgOXP1 and FgOXP2 led to more severe defects in conidiation, DON production and virulence on plants, suggesting that both FgOXP1 and FgOXP2 play a role in fungal development and plant colonization. Although transformation of MoOXP1into ΔFgoxp1 was able to complement ΔFgoxp1, transformation of MoOXP1 into ΔFgoxp2 failed to restore its defects in sexual development, DON production and pathogenicity. Taken together, these results suggest that FgOXP1 and FgOXP2 are likely to have been functionally diversified and play significant roles in fungal development and full virulence in F. graminearum.

The endosomal recycling of FgSnc1 by FgSnx41-FgSnx4 heterodimer is essential for polarized growth and pathogenicity in Fusarium graminearum.

Endosomal sorting machineries regulate the transport of their cargoes among intracellular compartments. However, the molecular nature of such intracellular trafficking processes in pathogenic fungal development and pathogenicity remains unclear. Here, we dissect the roles and molecular mechanisms of two sorting nexin proteins and their cargoes in endosomal recycling in Fusarium graminearum using high-resolution microscopy and high-throughput co-immunoprecipitation strategies. We show that the sorting nexins, FgSnx41 and FgSnx4, interact with each other and assemble into a functionally interdependent heterodimer through their respective BAR domains. Further analyses demonstrate that the dimer localizes to the early endosomal membrane and coordinates endosomal sorting. The small GTPase FgRab5 regulates the correct localization of FgSnx41-FgSnx4 and is consequently required for its trafficking function. The protein FgSnc1 is a cargo of FgSnx41-FgSnx4 and regulates the fusion of secreted vesicles with the fungal growing apex and plasma membrane. In the absence of FgSnx41 or FgSnx4, FgSnc1 is mis-sorted and degraded in the vacuole, and null deletion of either component causes defects in the fungal polarized growth and virulence. Overall, for the first time, our results reveal the mechanism of FgSnc1 endosomal recycling by FgSnx41-FgSnx4 heterodimer which is essential for polarized growth and pathogenicity in F. graminearum.

Comparison of Combined Dissipation Behaviors and Dietary Risk Assessments of Thiamethoxam, Bifenthrin, Dinotefuran, and Their Mixtures in Tea.

In the tea-planting process, insecticides are commonly combined, potentially prolonging the pre-harvest interval and heightening the risk of dietary exposure. This study focused on three frequently used insecticides in tea cultivation: thiamethoxam, bifenthrin, and dinotefuran, aiming to investigate their dissipation behaviors and associated dietary risks upon individual and simultaneous application. The dissipation kinetics of thiamethoxam, bifenthrin, and dinotefuran were successfully characterized by first-order kinetics, yielding respective half-lives of 5.44, 9.81, and 10.16 days. Upon joint application, the dissipation half-lives of thiamethoxam and bifenthrin were notably prolonged compared with their individual applications, resulting in final concentrations after 28 days that were correspondingly elevated by 1.41 and 1.29 times. Assessment of the dietary intake risk revealed that the chronic and acute risk quotients associated with thiamethoxam and bifenthrin escalated by 1.44-1.59 times following their combined application. Although dietary risks associated with Tianmuhu white tea, as determined by the exposure assessment model, were deemed acceptable, the cumulative risks stemming from pesticide mixtures across various dietary sources warrant attention. Molecular docking analyses further unveiled that thiamethoxam and bifenthrin competitively bound to glutathione S-transferase (GST) at amino acid residues, notably at the 76th GLU and the 25th PHE, pivotal in the metabolism and absorption of exogenous substances. Moreover, the interactions between P-glycoprotein and pesticides during transport and absorption were likely to influence dissipation behaviors post-joint application. This research offers valuable insights and data support for optimizing joint pesticide application strategies and assessing risks associated with typical pesticides used in tea cultivation.

Genome-wide mapping of DNase I hypersensitive sites in pineapple leaves.

Pineapple [Ananas comosus (L.) Merr.] is the most economically important crop possessing crassulacean acid metabolism (CAM) photosynthesis which has a higher water use efficiency by control of nocturnal opening and diurnal closure of stomata. To provide novel insights into the diel regulatory landscape in pineapple leaves, we performed genome-wide mapping of DNase I hypersensitive sites (DHSs) in pineapple leaves at day (2a.m.) and night (10a.m.) using a simplified DNase-seq method. As a result, totally 33340 and 28753 DHSs were found in green-tip tissue, and 29597 and 40068 were identified in white-base tissue at 2a.m. and 10a.m., respectively. We observed that majority of the pineapple genes occupied less than two DHSs with length shorter than 1 kb, and the promotor DHSs showed a proximal trend to the transcription start site (>77% promotor DHSs within 1 kb). In addition, more intergenic DHSs were identified around transcription factors or transcription co-regulators (TFs/TCs) than other functional genes, indicating complex regulatory contexts around TFs/TCs. Through combined analysis of tissue preferential DHSs and genes, we respectively found 839 and 888 coordinately changed genes in green-tip at 2a.m. and 10a.m. (AcG2 and AcG10). Furthermore, AcG2-specific, AcG10-specific and common accessible DHSs were dissected from the total photosynthetic preferential DHSs, and the regulatory networks indicated dynamic regulations with multiple cis-regulatory elements occurred to genes preferentially expressed in photosynthetic tissues. Interestingly, binding motifs of several cycling TFs were identified in the DHSs of key CAM genes, revealing a circadian regulation to CAM coordinately diurnal expression. Our results provide a chromatin regulatory landscape in pineapple leaves during the day and night. This will provide important information to assist with deciphering the circadian regulation of CAM photosynthesis.

A plant nonenveloped double-stranded RNA virus activates and co-opts BNIP3-mediated mitophagy to promote persistent infection in its insect vector.

Mitophagy that selectively eliminates damaged mitochondria is an essential mitochondrial quality control mechanism. Recently, mitophagy has been shown to be induced in host cells infected by a few animal viruses. Here, we report that southern rice black-streaked dwarf virus (SRBSDV), a plant nonenveloped double-stranded RNA virus, can also trigger mitophagy in its planthopper vector to prevent mitochondria-dependent apoptosis and promote persistent viral propagation. We find that the fibrillar structures constructed by the nonstructural protein P7-1 of SRBSDV directly target mitochondria via interaction with the mitophagy receptor BNIP3 (BCL2 interacting protein 3), and these mitochondria are then sequestered within autophagosomes to form mitophagosomes. Moreover, SRBSDV infection or P7-1 expression alone can promote BNIP3 dimerization on the mitochondria, and induce autophagy via the P7-1-ATG8 interaction. Furthermore, SRBSDV infection stimulates the phosphorylation of AMP-activated protein kinase (AMPK), resulting in BNIP3 phosphorylation via the AMPKα-BNIP3 interaction. Together, P7-1 induces BNIP3-mediated mitophagy by promoting the formation of phosphorylated BNIP3 dimers on the mitochondria. Silencing of ATG8, BNIP3, or AMPKα significantly reduces virus-induced mitophagy and viral propagation in insect vectors. These data suggest that in planthopper, SRBSDV-induced mitophagosomes are modified to accommodate virions and facilitate persistent viral propagation. In summary, our results demonstrate a previously unappreciated role of a viral protein in the induction of BNIP3-mediated mitophagy by bridging autophagosomes and mitochondria and reveal the functional importance of virus-induced mitophagy in maintaining persistent viral infection in insect vectors.Abbreviations: AMPK: AMP-activated protein kinase; ATG: autophagy related; BNIP3: BCL2 interacting protein 3; CASP3: caspase 3; dsRNA: double strand RNA; ER: endoplasmic reticulum; FITC: fluorescein isothiocyanate; FKBP8: FKBP prolyl isomerase 8; FUNDC1: FUN14 domain containing 1; GFP: green fluorescent protein; GST: glutathione S-transferase; padp: post-first access to diseased plants; Phos-tag: Phosphate-binding tag; PINK1: PTEN induced kinase 1; Sf9: Spodoptera frugiperda; SQSTM1: sequestosome 1; SRBSDV: southern rice black-streaked dwarf virus; STK11/LKB1: serine/threonine kinase 11; TOMM20: translocase of outer mitochondrial membrane 20; RBSDV: rice black-streaked dwarf virus; TUNEL: terminal deoxynucleotidyl dUTP nick end labeling; ULK1: unc-51 like autophagy activating kinase 1; VDAC1: voltage dependent anion channel 1.

Autophagy mediates a direct synergistic interaction during co-transmission of two distinct arboviruses by insect vectors.

Multiple viral infections in insect vectors with synergistic effects are common in nature, but the underlying mechanism remains elusive. Here, we find that rice gall dwarf reovirus (RGDV) facilitates the transmission of rice stripe mosaic rhabdovirus (RSMV) by co-infected leafhopper vectors. RSMV nucleoprotein (N) alone activates complete anti-viral autophagy, while RGDV nonstructural protein Pns11 alone induces pro-viral incomplete autophagy. In co-infected vectors, RSMV exploits Pns11-induced autophagosomes to assemble enveloped virions via N-Pns11-ATG5 interaction. Furthermore, RSMV could effectively propagate in Sf9 cells. Expression of Pns11 in Sf9 cells or leafhopper vectors causes the recruitment of N from the ER to Pns11-induced autophagosomes and inhibits N-induced complete autophagic flux, finally facilitating RSMV propagation. In summary, these results demonstrate a previously unappreciated role of autophagy in the regulation of the direct synergistic interaction during co-transmission of two distinct arboviruses by insect vectors and reveal the functional importance of virus-induced autophagosomes in rhabdovirus assembly.

Arboviruses and symbiotic viruses cooperatively hijack insect sperm-specific proteins for paternal transmission.

Arboviruses and symbiotic viruses can be paternally transmitted by male insects to their offspring for long-term viral persistence in nature, but the mechanism remains largely unknown. Here, we identify the sperm-specific serpin protein HongrES1 of leafhopper Recilia dorsalis as a mediator of paternal transmission of the reovirus Rice gall dwarf virus (RGDV) and a previously undescribed symbiotic virus of the Virgaviridae family, Recilia dorsalis filamentous virus (RdFV). We show that HongrES1 mediates the direct binding of virions to leafhopper sperm surfaces and subsequent paternal transmission via interaction with both viral capsid proteins. Direct interaction of viral capsid proteins mediates simultaneously invasion of two viruses into male reproductive organs. Moreover, arbovirus activates HongrES1 expression to suppress the conversion of prophenoloxidase to active phenoloxidase, potentially producing a mild antiviral melanization defense. Paternal virus transmission scarcely affects offspring fitness. These findings provide insights into how different viruses cooperatively hijack insect sperm-specific proteins for paternal transmission without disturbing sperm functions.

A plant reovirus hijacks the DNAJB12-Hsc70 chaperone complex to promote viral spread in its planthopper vector.

Many viruses usurp the functions of endoplasmic reticulum (ER) for virus-encoded membrane proteins proper functional folding or assembly to promote virus spread. Southern rice black-streaked dwarf virus (SRBSDV), a plant reovirus, exploits virus-containing tubules composed of nonstructural membrane protein P7-1 to spread in its planthopper vector Sogatella furcifera. Here, we report that two factors of the ER-associated degradation (ERAD) machinery, the ER chaperone DNAJB12 and its cytosolic co-chaperone Hsc70, are activated by SRBSDV to facilitate ER-to-cytosol export of P7-1 tubules in S. furcifera. Both P7-1 of SRBSDV and Hsc70 directly bind to the J-domain of DNAJB12. DNAJB12 overexpression induces ER retention of P7-1, but Hsc70 overexpression promotes the transport of P7-1 from the ER to the cytosol to initiate tubule assembly. Thus, P7-1 is initially retained in the ER by interaction with DNAJB12 and then delivered to Hsc70. Furthermore, the inhibitors of the ATPase activity of Hsc70 reduce P7-1 tubule assembly, suggesting that the proper folding and assembly of P7-1 tubules is dependent on the ATPase activity of Hsc70. The DNAJB12-Hsc70 chaperone complex is recruited to P7-1 tubules in virus-infected midgut epithelial cells in S. furcifera. The knockdown of DNAJB12 or Hsc70 strongly inhibits P7-1 tubule assembly in vivo, finally suppressing effective viral spread in S. furcifera. Taken together, our results indicate that the DNAJB12-Hsc70 chaperone complex in the ERAD machinery facilitates the ER-to-cytosol transport of P7-1 for proper assembly of tubules, enabling viral spread in insect vectors in a manner dependent on ATPase activity of Hsc70.

Self- and Conspecific Discrimination Between Unparasitized and Parasitized Green Peach Aphid (Hemiptera: Aphididae), by Aphelinus asychis (Hymenoptera: Aphelinidae).

Aphelinus asychis Walker (Hymenoptera: Aphelinidae) is a solitary koinobiont endoparasitoid, which parasitizes the green peach aphid, Myzus persicae (Sulzer) (Hemiptera: Aphididae). Host discrimination is the ability of a parasitoid to distinguish between unparasitized and parasitized hosts, so parasitoids can avoid wasting eggs and time. We determined the influence of host density (5, 10, 15, 20, and 25 aphids per experienced parasitoid during a 24-h period) on egg distribution, and tested the effects of different time intervals (0, 24, 48, and 72 h) between successive ovipositions on host discrimination between unparasitized and self- or conspecific-parasitized aphids. The results demonstrated that superparasitism decreased with increasing host density. Egg distributions were nonrandom at all host densities, and the observed number of self-superparasitized hosts was lower than expected at all host densities. At 0-h interval, most parasitoids used only antennation to examine and reject parasitized aphids, which might be associated with an external marker. At longer time intervals (≥24 h), A. asychis most often used antennation followed by ovipositor insertion to examine and reject previously parasitized aphids, probably responding to an internal, from chemical substances injected during the first oviposition or changes in host quality associated with parasitism. The results showed that host discrimination in A. asychis was mediated by two consecutive cues, the first external and the second internal, with the relevancy of each depending on the time interval between two ovipositions. Consequently, A. asychis was able to discriminate between unparasitized and self- or conspecific-parasitized M. persicae, but always engaged in some superparasitism.