Combined effect of corn in the barrier crop and plant extracts against Cowpea mild mottle virus infecting soybean [Glycine max (L. ) Merr. ] in the field

Cowpea mild mottle virus (CpMMV) is one of the problems that can decrease soybean production. The research was conducted on the combined effects of corn in the barrier crop with plant extracts against CpMMV infecting soybean in the field. The field data was conducted using a Completely Randomized Design. The mean of disease incidence and disease severity is measured from total plants in each replicate plot on each treatment. Planting one or two of corn lines were grown at the edge four weeks before planting soybeans. Cashew nut shell (CNS), pagoda leaf, and rhizome of ginger extracts were applied using the sprayer and applied at 24 h before virus acquisition and transmission by whiteflies. The result showed that the virus incubation period ranged from 9−38 days after transmission longer than the untreated control. Planting two corn lines at the edge with CNS extract as bioactivator on soybean was the most extended incubation period of the virus and the lowest absorbance value DAS-ELISA of 0.20. There was a 73.11 % increase in the relative inhibition level of the virus. Planting corn at the edge with CNS extract proved to be more effective than soybean monoculture with CNS extract. However, soybean monoculture with CNS extract provides a better relative inhibition level of the virus (64.32 %) than planting two rows of corn on the edge combined with ginger of rhizome extract and planting two rows of corn on the edge with pagoda leaf extract as bioactivator on a soybean plant.

Assessment of Two Novel Host-Derived Beauveria bassiana (Hypocreales: Cordycipitaceae) Isolates Against the Citrus Pest, Diaphorina citri (Hemiptera: Liviidae).

The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Liviidae), vectors 'Candidatus Liberibacter spp.', the causative agent of Citrus Greening Disease (CGD) or Huanglongbing (HLB). Managing populations of psyllids in the Lower Rio Grande Valley (LRGV), TX, United States is imperative given a continuous increase in HLB-positive trees. A component of integrated pest management (IPM) program is the use of strains of entomopathogenic fungi for the biological control of D. citri. In an attempt to find endemic strains of entomopathogenic fungi that grow favorably under LRGV environmental conditions and naturally infect D. citri, psyllids were collected from local residential areas, surface sterilized, and plated on a semi-selective agar medium. Collection of over 9,300 samples from 278 sites throughout the LRGV led to the positive identification of two Beauveria bassiana (Balsamo-Crivellii) Vuillemin (Hypocreales: Cordycipitaceae) isolates, ACP18001 and ACP18002. Chi-square analysis of primary and secondary acquisition bioassays revealed that both field isolated strains outperformed Cordyceps (Isaria) fumosorosea (Wize) (Hypocreales: Cordycipitaceae) Apopka97 under both primary (direct spray) and secondary acquisition (adult exposure to sprayed foliage) bioassays with ACP18002 marginally outperforming ACP18001 under secondary acquisition. Slopes of the dose response regression lines for the three fungi were not significantly different. In addition, the thermal profiles for vegetative growth of each isolate indicated that the field isolates grew at higher rates than the standard at higher temperatures. The new isolates may prove to be good candidates for the management of D. citri populations in the LRGV.

Overexpression of an Osa-miR162a Derivative in Rice Confers Cross-Kingdom RNA Interference-Mediated Brown Planthopper Resistance without Perturbing Host Development.

Rice is a main food crop for more than half of the global population. The brown planthopper (BPH, Nilaparvata lugens) is one of the most destructive insect pests of rice. Currently, repeated overuse of chemical insecticides represents a common practice in agriculture for BPH control, which can induce insect tolerance and provoke environmental concerns. This situation calls for innovative and widely applicable strategies for rice protection against BPH. Here we report that the rice osa-miR162a can mediate cross-kingdom RNA interference (RNAi) by targeting the NlTOR (Target of rapamycin) gene of BPH that regulates the reproduction process. Through artificial diet or injection, osa-miR162a mimics repressed the NlTOR expression and impaired the oviposition of BPH adults. Consistently, overproduced osa-miR162a in transgenic rice plants compromised the fecundity of BPH adults fed with these plants, but meanwhile perturbed root and grain development. To circumvent this issue, we generated osa-miR162a-m1, a sequence-optimized osa-miR162a, by decreasing base complementarity to rice endogenous target genes while increasing base complementarity to NlTOR. Transgenic overexpression of osa-miR162a-m1 conferred rice resistance to BPH without detectable developmental penalty. This work reveals the first cross-kingdom RNAi mechanism in rice-BPH interactions and inspires a potentially useful approach for improving rice resistance to BPH. We also introduce an effective strategy to uncouple unwanted host developmental perturbation from desirable cross-kingdom RNAi benefits for overexpressed plant miRNAs.

Transmission of Grapevine leafroll-associated virus-1 (Ampelovirus) and Grapevine virus A (Vitivirus) by the Cottony Grape Scale, Pulvinaria vitis (Hemiptera: Coccidae).

The cottony grape scale Pulvinaria vitis is a scale insect colonizing grapevine; however, its capacity as a vector of grapevine viruses is poorly known in comparison to other scale species that are vectors of viral species in the genera Ampelovirus and Vitivirus. The ability of P. vitis to transmit the ampeloviruses Grapevine leafroll-associated viruses [GLRaV]-1, -3, and -4, and the vitivirus Grapevine virus A (GVA), to healthy vine cuttings was assessed. The scale insects used originated from commercial vine plots located in Alsace, Eastern France. When nymphs sampled from leafroll-infected vineyard plants were transferred onto healthy cuttings, only one event of transmission was obtained. However, when laboratory-reared, non-viruliferous nymphs were allowed to acquire viruses under controlled conditions, both first and second instar nymphs derived from two vineyards were able to transmit GLRaV-1 and GVA. This is the first report of GLRaV-1 and GVA transmission from grapevine to grapevine by this species.

RNA-Seq based global transcriptome analysis of rice unravels the key players associated with brown planthopper resistance.

Rice production is adversely affected by biotic and abiotic stresses. Among the biotic stresses, brown planthopper (BPH) majorly affects the rice yield. Comprehending the genome and candidate players is essential for the resistance to BPH. This holistic study aimed to dissect the complex BPH resistance mechanism of the host against pathogen. Transcriptome analysis of six samples comprising of two-resistant (PTB33, BM71) and one-sensitive (TN1) genotypes under control and stress conditions was carried-out. A total of 148 million filtered reads were generated after quality check. Among these, 127 million filtered reads were aligned to the rice genome. These aligned reads were taken for further analysis. A total of 14,358 DEGs across the genotypes under stress were identified. Of which, 4820 DEGs were functionally annotated from 9266 uniquely mapped DEGs. Fifty-five potential BPH stress players were selected from the in-silico analysis of DEGs. qRT-PCR results revealed key players were differentially regulated in both resistant and sensitive genotypes. Spatio-temporal and hormone level expression signature of 55 BPH associated players were analyzed and noted their differential expression in tissues and hormones, respectively. This study inferred the significant differences in gene expression signatures may contribute to the process of BPH resistance mechanism in rice.

A rapid multiplex PCR assay for species identification of Asian rice planthoppers (Hemiptera: Delphacidae) and its application to early-instar nymphs in paddy fields.

Rice (Oryza sativa L.) is the main cereal crop in many Asian countries. The Asian rice planthoppers, Nilaparvata lugens (Stål) (brown planthopper), Sogatella furcifera (Horváth) (white-backed planthopper), and Laodelphax striatellus (Fallén) (small brown planthopper) (Hemiptera: Delphacidae), are the most economically important pests of rice. These three rice planthopper species often co-occur in the same paddy field. Traditionally, species identification of individuals of the three rice planthopper species has relied on morphological characters, but accurate discrimination of early-instar nymphs is very difficult, even for expert researchers. In this study, we developed a rapid one-step multiplex PCR assay using conserved and species-specific 5.8S-ITS2 rDNA gene primers for simultaneous identification of individuals of the three rice planthopper species. The multiplex PCR results showed that the three rice planthopper species could be identified accurately based on the length of the resultant amplicon, regardless of the individual developmental stage. Furthermore, we applied this assay for the first accurate quantification of early-instar nymphs of each rice planthopper species in paddy fields. Notably, we found that the species composition of early-instar nymphs cannot be extrapolated from that of adults. Thus, the multiplex PCR assay developed here facilitates detection of each rice planthopper species at the beginning of outbreaks in paddy fields.

Mitogenomic analysis of diversity of key whitefly pests in Kenya and its implication to their sustainable management.

Whiteflies (Hemiptera: Aleyrodidae) are devastating agricultural pests of economic importance vectoring pathogenic plant viruses. Knowledge on their diversity and distribution in Kenya is scanty, limiting development of effective sustainable management strategies. The present study is aimed at identifying whitefly pest species present in Kenya across different agroecological zones and establish predictive models for the most abundant species in Africa. Whiteflies were sampled in Kenya from key crops known to be severely infested and identified using 16S rRNA markers and complete mitochondrial genomes. Four whitefly species were identified: Aleyrodes proletella, Aleurodicus dispersus, Bemisia afer and Trialeurodes vaporariorum, the latter being the most dominant species across all the agroecology. The assembly of complete mitogenomes and comparative analysis of all 13 protein coding genes confirmed the identities of the four species. Furthermore, prediction spatial models indicated high climatic suitability of T. vaporariorum in Africa, Europe, Central America, parts of Southern America, parts of Australia, New Zealand and Asia. Consequently, our findings provide information to guide biosecurity agencies on protocols to be adopted for precise identification of pest whitefly species in Kenya to serve as an early warning tool against T. vaporariorum invasion into unaffected areas and guide appropriate decision-making on their management.

Identification and analysis of miRNAs in IR56 rice in response to BPH infestations of different virulence levels.

Rice production and sustainability are challenged by its most dreadful pest, the brown planthopper (Nilaparvata lugens Stål, BPH). Therefore, the studies on rice-BPH interactions and their underlying mechanisms are of high interest. The rice ontogenetic defense, such as the role of microRNAs (miRNAs) has mostly been investigated against the pathogens, with only a few reports existing against the insect infestations. Thus, revealing the involvement of rice miRNAs in response to BPH infestations will be beneficial in understanding these complex interactions. In this study, the small RNA profiling of the IR56 rice in response to separate BPH infestations of varied virulence levels identified the BPH-responsive miRNAs and revealed the differential transcript abundance of several miRNAs during a compatible and incompatible rice-BPH interaction. The miRNA sequence analysis identified 218 known and 28 novel miRNAs distributed in 54 miRNA families. Additionally, 138 and 140 numbers of differentially expressed (DE) miRNAs were identified during the compatible and incompatible interaction, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed the target gene candidates of DE miRNAs (including osa-miR2871a-3p, osa-miR172a, osa-miR166a-5p, osa-miR2120, and osa-miR1859) that might be involved in the IR56 rice defense responses against BPH infestation. Conversely, osa-miR530-5p, osa-miR812s, osa-miR2118g, osa-miR156l-5p, osa-miR435 and two of the novel miRNAs, including novel_16 and novel_52 might negatively modulate the IR56 rice defense. The expressional validation of the selected miRNAs and their targets further supported the IR56 rice defense regulatory network. Based on our results, we have proposed a conceptual model depicting the miRNA defense regulatory network in the IR56 rice against BPH infestation. The findings from the study add further insights into the molecular mechanisms of rice-BPH interactions and will be helpful for the future researches.

Cry1C rice doesn't affect the ecological fitness of rice brown planthopper, Nilaparvata lugens either under RDV stress or not.

The potential risks of Bt rice on non-target arthropods (NTAs) should be evaluated and defined before commercial production. Recently, effects of Bt rice on NTAs under abiotic and biotic stress conditions attracted much attention. Here we reported the effects of Bt rice T1C-19 (Cry1C rice) on the non-target herbivore, Nilaparvata lugens (rice brown planthopper, BPH) with or without RDV (rice dwarf virus) infection conditions. BPH showed no feeding and oviposition preference between Bt rice T1C-19 and its non-Bt parental rice Minghui 63 (MH63), as well as between RDV-infected and RDV-free rice plants. Meanwhile, rice type, RDV infection status, and their interaction had little impacts on the survival, development and fecundity of BPH. By comparison with non-Bt control, Bt rice T1C-19 with or without RDV infection had no significant effects on the life-table parameters of BPH including rm, R0, T, DT and λ. Thus, it could be concluded that Bt rice T1C-19 doesn't affect the ecological fitness of BPH either under RDV stress or not.

Comparative transcriptome analysis of defense response of rice to Nilaparvata lugens and Chilo suppressalis infestation.

The brown planthopper (BPH, Nilaparvata lugens) and striped stem borer (SSB, Chilo suppressalis) are two of the most devastating insect pests in rice, causing significant losses of rice yield. Plants evolve multiple defense responses in the process of coexisting with pests. According to different pest infestation, the plants selectively activate related pathways and downstream gene expression. However, there are very few reports of differences in defense signaling pathways after rice was attacked by BPH or SSB. We determined the transcriptional responses of rice infested with BPH and SSB for 3 and 6 h using Illumina sequencing. By comparing the difference in gene changes caused by BPH and SSB infestation in rice, multiple signal pathways and gene expression patterns, including phytohormones, secondary metabolites, plant-pathogen interaction, reactive oxygen species, defense response, transcription factors, protease inhibitor and chitinase were found significantly different. Our results provide a basis for further exploring the molecular mechanism of rice defense response caused by BPH and SSB infestation, which will add to further understanding the interactions between plants and insects, and could provide valuable resources that could be applied in insect-resistant crop breeding.

Changes in reflectance of rice seedlings during planthopper feeding as detected by digital camera: Potential applications for high-throughput phenotyping.

Damage to grasses and cereals by phloem-feeding herbivores is manifest as nutrient and chlorophyll loss, desiccation, and a gradual decline in host vigour. Chlorophyll loss in particular leads to a succession of colour changes before eventual host death. Depending on the attacking herbivore species, colour changes can be difficult to detect with the human eye. This study used digital images to examine colour changes of rice seedlings during feeding by the brown planthopper, Nilaparvata lugens (Stål) and whitebacked planthopper, Sogatella furcifera (Horváth). Values for red (580 nm), green (540 nm) and blue (550 nm) reflectance for 39 rice varieties during seedling seed-box tests were derived from images captured with a digital camera. Red and blue reflectance gradually increased as herbivore damage progressed until final plant death. Red reflectance was greater from plants attacked by the brown planthopper than plants attacked by the whitebacked planthopper, which had proportionately more green and blue reflectance, indicating distinct impacts by the two planthoppers on their hosts. Analysis of digital images was used to discriminate variety responses to the two planthoppers. Ordination methods based on red-green-blue reflectance and vegetation indices such as the Green Leaf Index (GLI) that included blue reflectance were more successful than two-colour indices or indices based on hue, saturation and brightness in discriminating between damage responses among varieties. We make recommendations to advance seed-box screening methods for cereal resistance to phloem feeders and demonstrate how images from digital cameras can be used to improve the quality of data captured during high-throughput phenotyping.

A plant DNA virus replicates in the salivary glands of its insect vector via recruitment of host DNA synthesis machinery.

Whereas most of the arthropod-borne animal viruses replicate in their vectors, this is less common for plant viruses. So far, only some plant RNA viruses have been demonstrated to replicate in insect vectors and plant hosts. How plant viruses evolved to replicate in the animal kingdom remains largely unknown. Geminiviruses comprise a large family of plant-infecting, single-stranded DNA viruses that cause serious crop losses worldwide. Here, we report evidence and insight into the replication of the geminivirus tomato yellow leaf curl virus (TYLCV) in the whitefly (Bemisia tabaci) vector and that replication is mainly in the salivary glands. We found that TYLCV induces DNA synthesis machinery, proliferating cell nuclear antigen (PCNA) and DNA polymerase δ (Polδ), to establish a replication-competent environment in whiteflies. TYLCV replication-associated protein (Rep) interacts with whitefly PCNA, which recruits DNA Polδ for virus replication. In contrast, another geminivirus, papaya leaf curl China virus (PaLCuCNV), does not replicate in the whitefly vector. PaLCuCNV does not induce DNA-synthesis machinery, and the Rep does not interact with whitefly PCNA. Our findings reveal important mechanisms by which a plant DNA virus replicates across the kingdom barrier in an insect and may help to explain the global spread of this devastating pathogen.

Transcriptomic analysis of Dubas bug (Ommatissus lybicus Bergevin) infestation to Date Palm.

Date palm (Phoenix dactylifera L.) and its fruit possess sociocultural, health and economic importance in Middle East. The date palm plantations are prone to Dubas bug (DB; Ommatissus lybicus DeBergevin; Homoptera: Tropiduchidae) attacks that severely damages the tree's growth and reduces fruit production. However, the transcriptome related datasets are not known to understand how DB activates physiological and gene regulatory mechanisms during infestation. Hence, we performed RNA-Seq of leaf infected with or without DB to understand the molecular responses of date palm seedlings. Before doing that, we noticed that DB infestation significantly increase superoxide anion and malondialdehyde production to two-folds as compared to healthy control. Stress-responsive genes such as proline transporter 2, NADP-dependent glyceraldehyde and superoxide dismutase were found significantly upregulated in infected seedlings. The infection repercussions were also revealed by significantly higher contents of endogenous phytohormonal signaling of jasmonic acid (JA) and salicylic acid (SA) compared with control. These findings persuaded to dig out intrinsic mechanisms and gene regulatory networks behind DB infestation to date palm by RNA-Seq analysis. Transcriptome analysis revealed upregulation of 6,919 genes and down-regulation of 2,695 genes in leaf during the infection process. The differentially expressed genes were mostly belongs to cellular functions (calcium and MAPK), phytohormones (auxin, gibberellins, abscisic acid, JA and SA), and secondary metabolites (especially coumarinates and gossypol). The data showed that defense responses were aggravated by gene networks involved in hypersensitive responses (PAR1, RIN4, PBS1 etc.). In conclusion, the results revealed that date palm's leaf up-regulates both cellular and phytohormonal determinants, followed by intrinsic hypersensitive responses to counter infestation process by Dubas bug.

Constitutive expression of Asparaginase in Gossypium hirsutum triggers insecticidal activity against Bemisia tabaci.

Whitefly infestation of cotton crop imparts enormous damage to cotton yield by severely affecting plant health, vigour and transmitting Cotton Leaf Curl Virus (CLCuV). Genetic modification of cotton helps to overcome both the direct whitefly infestation as well as CLCuV based cotton yield losses. We have constitutively overexpressed asparaginase (ZmASN) gene in Gossypium hirsutum to overcome the cotton yield losses imparted by whitefly infestation. We achieved 2.54% transformation efficiency in CIM-482 by Agrobacterium-mediated shoot apex transformation method. The relative qRT-PCR revealed 40-fold higher transcripts of asparaginase in transgenic cotton line vs. non-transgenic cotton lines. Metabolic analysis showed higher contents of aspartic acid and glutamic acid in seeds and phloem sap of the transgenic cotton lines. Phenotypically, the transgenic cotton lines showed vigorous growth and height, greater number of bolls, and yield. Among six representative transgenic cotton lines, line 14 had higher photosynthetic rate, stomatal conductance, smooth fiber surface, increased fiber convolutions (SEM analysis) and 95% whitefly mortality as compared to non-transgenic cotton line. The gene integration analysis by fluorescence in situ hybridization showed single copy gene integration at chromosome number 1. Collectively, asparaginase gene demonstrated potential to control whitefly infestation, post-infestation damages and improve cotton plant health and yield: a pre-requisite for farmer's community.

Relationship between the honeydew of mealy bugs and the growth of Phlebopus portentosus.

BACKGROUND: Phlebopus portentosus and mealy bugs form a fungus-insect gall on the roots of host plants. The fungus and mealy bugs benefit mutually through the gall, which is the key link in the nutritional mechanism of P. portentosus. The cavity of the fungus-insect gall provides an ideal shelter for mealy bugs survival and reproduction, but how does P. portentosus benefit from this symbiotic relationship? METHODOLOGY AND RESULTS: Anatomical examination of fungus-insect galls revealed that one or more mealy bugs of different generations were living inside the galls. The mealy bug's mouthpart could penetrate through the mycelium layer of the inside of the gall and suck plant juice from the host plant root. Mealy bugs excreted honeydew inside or outside the galls. The results of both honeydew agar medium and quartz tests showed that the honeydew can attract and promote the mycelial growth of P. portentosus. A test of the relationship between the honeydew and the formation of the fungus-insect gall showed that honeydew promoted gall formation. CONCLUSIONS: All experimental results in this study show that the honeydew secreted by mealy bugs can attract and promote the mycelial growth of P. portentosus, forming a fungus-insect gall, because mealy bugs' honeydew is rich in amino acids and sugars.

Cassava whitefly species in eastern Nigeria and the threat of vector-borne pandemics from East and Central Africa.

Bemisia tabaci (sensu latu) is a group of >40 highly cryptic whitefly species that are of global agricultural importance, both as crop pests and plant-virus vectors. Two devastating cassava diseases in East and Central Africa are spread by abundant populations of one of these species termed Sub-Saharan Africa 1 (SSA1). There is a substantive risk that these whitefly-borne pandemics will continue to spread westwards and disrupt cassava production for millions of smallholder farmers in West Africa. We report here, therefore, the first comprehensive survey of cassava B. tabaci in eastern Nigeria, a West African region likely to be the first affected by the arrival of these whitefly-borne pandemics. We found one haplotype comprising 32 individuals with 100% identical mtCO1 sequence to the East African SSA1 populations (previously termed SSA1-SG1) and 19 mtCO1 haplotypes of Sub-Saharan Africa 3 (SSA3), the latter being the most prevalent and widely distributed B. tabaci species in eastern Nigeria. A more divergent SSA1 mtCO1 sequence (previously termed SSA1-SG5) was also identified in the region, as were mtCO1 sequences identifying the presence of the MED ASL B. tabaci species and Bemisia afer. Although B. tabaci SSA1 was found in eastern Nigeria, they were not present in the high abundances associated with the cassava mosaic (CMD) and cassava brown streak disease (CBSD) pandemics of East and Central Africa. Also, no severe CMD or any CBSD symptoms were found in the region.

A biological control model to manage the vector and the infection of Xylella fastidiosa on olive trees.

Xylella fastidiosa pauca ST53 is the bacterium responsible for the Olive Quick Decline Syndrome that has killed millions of olive trees in Southern Italy. A recent work demonstrates that a rational integration of vector and transmission control measures, into a strategy based on chemical and physical control means, can manage Xylella fastidiosa invasion and impact below an acceptable economic threshold. In the present study, we propose a biological alternative to the chemical control action, which involves the predetermined use of an available natural enemy of Philaenus spumarius, i.e., Zelus renardii, for adult vector population and infection biocontrol. The paper combines two different approaches: a laboratory experiment to test the predation dynamics of Zelus renardii on Philaenus spumarius and its attitude as candidate for an inundation strategy; a simulated experiment of inundation, to preliminary test the efficacy of such strategy, before eventually proceeding to an in-field experimentation. With this double-fold approach we show that an inundation strategy with Zelus renardii has the potential to furnish an efficient and "green" solution to Xylella fastidiosa invasion, with a reduction of the pathogen incidence below 10%. The biocontrol model presented here could be promising for containing the impact and spread of Xylella fastidiosa, after an in-field validation of the inundation technique. Saving the fruit orchard, the production and the industry in susceptible areas could thus become an attainable goal, within comfortable parameters for sustainability, environmental safety, and effective plant health protection in organic orchard management.

Transcriptome profiling and dimorphic expression of sex-related genes in fifth-instar nymphs of Sogatella furcifera, an important rice pest.

Sogatella furcifera is an important rice pest. In order to understand the molecular basis of the sex determination in this pest, we performed de novo transcriptome sequencing of six cDNA libraries (three biological replicates) of female and male fifth-instar nymphs. Total 65,199 unigenes were obtained, with an average length of 971.5 bp and N50 length of 1708 bp. 20,287 open reading frames (ORFs) were predicted and annotated. Total 1019 differentially expressed genes with 873 upregulated and 146 downregulated were found in male compared to female. Total 164 sex-determining genes were identified, including the key sex-determining genes in fruit flies, such as Sxl, tra, dsx, etc. It implied that the sex determination mechanisms of S. furcifera may be the same as that of fruit flies. This study provided transcriptome resource as a fundamental support for future functional studies to elucidate the sex determination regulatory networks governing sexual dimorphism of S. furcifera.

Detection of a secreted protein biomarker for citrus Huanglongbing using a single-walled carbon nanotubes-based chemiresistive biosensor.

Citrus greening, or Huanglongbing (HLB), is currently the most devasting disease of citrus, creating unprecedented crisis for the multibillion-dollar global citrus industry. To-date, there is no effective cure and disease management relies on early detection and removal of infected trees. Thus, it is imperative that accurate, timely, and robust disease detection and diagnosis technologies are available to minimize the spread of disease. This study reports a sensitive and selective label-free biosensor that combines the physical and chemical advantages of carbon nanomaterials like single-walled carbon nanotubes (SWNTs) in a field-effect transistor (FET)/chemiresistor architecture with selective antibodies against Sec-delivered effector 1 (SDE1), a secreted protein biomarker, for the detection of HLB. The biosensor detected SDE1 biomarkers for citrus greening in plant tissue extracts with the dynamic range over three orders of magnitude in the low nanomolar to micromolar concentration range and limit of detection of 5 nM. The study also demonstrated the use of the standard additions assay method with the biosensor to attain a 90-percent signal recovery in concentrated plant tissue extract, allowing for quantitative detection without an external calibration. Adopting the novel detection strategy targeting the secreted protein biomarker, SDE1, addresses some of the challenges faced by current methods of nucleic acid-based assays and symptom-based diagnosis, which have been found prone to false negatives and misdiagnoses, respectively.

Characterization and application of a gall midge resistance gene (Gm6) from Oryza sativa 'Kangwenqingzhan'.

KEY MESSAGE: The resistance gene Gm6 was mapped and characterized using near-isogenic and pyramided lines, followed by marker-assisted selection to develop lines with resistance to both gall midge and brown planthopper. The Asian rice gall midge (GM; Orseolia oryzae; Diptera: Cecidomyiidae) is a major destructive pest affecting rice cultivation regions. The characterization of GM-resistance genes and the breeding of resistant varieties are together considered the most efficient strategy for managing this insect. Here, the Gm6 resistance gene derived from the Kangwenqingzhan (KW) variety was found to be located on the long arm of chromosome 4 using the F2 population of 9311/KW. The region was narrowed to a 90-kb segment flanked by the markers YW91 and YW3-4 using backcrossing populations. Based on no-choice feeding and host choice tests, GM development and growth in near-isogenic lines (NILs) were severely restricted compared to that in the 9311 control. On day 8, the average GM body length was 0.69 mm and 0.56 mm on NILs and 9311, respectively, and the differences were more significant at later time points. However, GM insects exhibited no host preference between NILs and 9311, and there was normal egg hatching on the resistant plants. We developed pyramided lines carrying BPH27, BPH36, and Gm6 by crossing and backcrossing with marker-assisted selection. These lines were similar to the KW parent in terms of agronomic traits while also exhibiting high resistance to brown planthopper (BPH) and GM. The present mapping and characterization of Gm6 will facilitate map-based cloning of this important resistance gene and its application in the breeding of insect-resistant rice varieties.