The involvement of cyclotides in mutual interactions of violets and the two-spotted spider mite.

Plants employ different chemicals to protect themselves from herbivory. These defenses may be constitutive or triggered by stress. The chemicals can be toxic, act as repellents, phagosuppressants and/or phago-deterrents. The two-spotted spider mite (Tetranychus urticae) is a generalist arthropod herbivorous pest and its feeding causes extensive damage both to crops and wild plants. Cyclotides are cyclic peptides involved in host-plant defenses. A single Viola sp. can produce more than a hundred cyclotides with different biological activities and roles. The organ and tissue specific cyclotide patterns change over the seasons and/or with environment, but the role of biotic/abiotic stress in shaping them remains unclear. Here, we demonstrate the involvement of cyclotides in mutual interactions between violets and mites. We used immunohistochemistry and mass spectrometry imaging to show the ingested cyclotides in T. urticae and assess the Viola odorata response to mite feeding. Moreover, to assess how mites are affected by feeding on violets, acceptance and reproductive performance was compared between Viola uliginosa, V. odorata and Phaseolus vulgaris. We demonstrate that cyclotides had been taken in by mites feeding on the violets. The ingested peptides were found in contact with epithelial cells of the mite digestive system, in the fecal matter, feces, ovary and eggs. Mites preferred common bean plants (P. vulgaris) to any of the violet species; the latter affected their reproductive performance. The production of particular cyclotides in V. odorata (denoted by molecular weights: 2979, 3001, 3017, 3068, 3084, 3123) was activated by mite feeding and their levels were significantly elevated compared to the control after 5 and 21 days of infestation. Specific cyclotides may affect mites by being indigestible or through direct interaction with cells in the mite digestive tract and reproductive organs. A group of particular peptides in V. odorata appears to be involved in defense response against herbivores.

Insecticides action of neonicotinoids group in fall armyworm (Spodoptera frugiperda) control on bean leaves (Phaseolus vulgaris)

Among several factors that impact the bean culture productivity there are pest insects, which affect the plant since seeding until postharvest, causing loss in the culture yield. The fall armyworm (Spodoptera frugiperda) stands out among the main pests of common bean (Phaseolus vulgaris). The aim of this work was to evaluate the effectiveness of insecticides of the neonicotinoid group in the control of the fall armyworm in the bean crop, comparing dinotefuran, acetamiprid, thiamethoxam and imidacloprid performance. Experimental design was composed of blocks entirely randomized, with 9 treatments and 10 replicates. Three caterpillars, at the stage of second instar, were used in each experiment, focusing on the ingestion of leaves containing the treatments. Evaluations were realized in the intervals of 1, 3, 6 and 8 days after every application, counting the number of caterpillars alive in the Petri's dish, attributing visual notes on bean leaves, according to the foliar area affected (consumed). The dinotefuran treatment with the highest dose presented superior efficiency at 80% in the first evaluation. The thiamethoxam treatment with the lowest dose, in the latest analyses, showed efficient superior at 90%. The major doses of all treatments presented efficiency higher than 80% in the last evaluations, being efficient in the S. frugiperda control.

Molecular insight into the endophytic growth of Beauveria bassiana within Phaseolus vulgaris in the presence or absence of Tetranychus urticae.

Entomopathogenic fungi are an important factor in the natural regulation of arthropod populations. Moreover, some can exist as an endophyte in many plant species and establish a mutualistic relationship. In this study, we have investigated the endophytic growth of Beauveria bassiana within different tissues of Phaseolus vulgaris in the presence and absence of Tetranuychus urticae. After the colonization of the B. bassiana within the internal tissues of P. vulgaris. The susceptibility of T. urticae appeared to depend on the life stage where high, moderate, and low mortalities were recorded among adults, nymphs, and eggs, respectively. In addition, this study provided, for the first time, molecular insight into the endophytic growth of B. bassiana by analyzing the expression of several genes involved in the development of the entomopathogenic fungi at 0-, 2-, and 7- days post-inoculation. B. bassiana displayed preferential tissue colonization within P. vulgaris that can be put into the following order based on the detection rate: leaf > stem > root. After analyzing the development-implicated genes (degrading enzymes, sugar transporter, hydrophobins, cell wall synthesis, secondary metabolites, stress management), the most remarkable finding is the detection of behavioral change between parasitic and endophytic Beauveria during post-penetration events. This study elucidates the tri-trophic interaction between fungus-plant-herbivore.

Comparative transcriptome provides molecular insight into defense-associated mechanisms against spider mite in resistant and susceptible common bean cultivars.

Common bean (Phaseolus vulgaris L.) is a major source of proteins and one of the most important edible foods for more than three hundred million people in the world. The common bean plants are frequently attacked by spider mite (Tetranychus urticae Koch), leading to a significant decrease in plant growth and economic performance. The use of resistant cultivars and the identification of the genes involved in plant-mite resistance are practical solutions to this problem. Hence, a comprehensive study of the molecular interactions between resistant and susceptible common bean cultivars and spider mite can shed light into the understanding of mechanisms and biological pathways of resistance. In this study, one resistant (Naz) and one susceptible (Akhtar) cultivars were selected for a transcriptome comparison at different time points (0, 1 and 5 days) after spider mite feeding. The comparison of cultivars in different time points revealed several key genes, which showed a change increase in transcript abundance via spider mite infestation. These included genes involved in flavonoid biosynthesis process; a conserved MYB-bHLH-WD40 (MBW) regulatory complex; transcription factors (TFs) TT2, TT8, TCP, Cys2/His2-type and C2H2-type zinc finger proteins; the ethylene response factors (ERFs) ERF1 and ERF9; genes related to metabolism of auxin and jasmonic acid (JA); pathogenesis-related (PR) proteins and heat shock proteins.

Enemigos naturales nativos de mosca blanca (Hemiptera: Aleyrodidae) en el cultivo de ejote francés en Chimaltenango, Guatemala / tive natural enemies of whitefly (Hemiptera: Aleyrodidae) in French bean crop in Chimaltenango, Guatemala

El complejo de mosca blanca (Hemiptera: Aleyrodidae) incluye algunas de las principales plagas del ejote francés (Phaseolus vulgaris L.). Dentro de las cuales, Bemisia tabaci es vector del virus del mosaico dorado que afecta la calidad y rendimiento del cultivo, con pérdidas hasta del 100% y un control difícil debido a la resistencia adquirida por las plagas hacia algunos agroquímicos. El ejote francés ocupa el segundo lugar entre de los productos no tradicionales de exportación de Guatemala. Su manejo agronómico ha sido principalmente a través del control químico, el cual afecta insectos y otros organismos que no son el objetivo del control, tales como: polinizadores, insectos benéficos, humanos y fauna silvestre. Los objetivos del estudio fueron: determinar la presencia de enemigos naturales nativos de la mosca blanca e identificar las especies de mosca blanca presentes en el cultivo del ejote francés en Chimaltenango. Para el estudio se establecieron cuatro parcelas de 300 m², se realizaron muestreos semanales durante dos ciclos del cultivo. En cada parcela se muestrearon cinco sitios y en cada sitio cinco plantas. Las especies de parasitoides nativos encontrados fueron: Encarsia Formosa Gahan, Eretmocerus eremicus Rose y Zolnerowuch y Amitus fuscipennis MacGown y Nebeker, la especie más abundante fue A. fuscipennis. Los depredadores identificados fueron Chrysoperla carnea (Stephens) e Hippodamia convergens Guerin-Meneville. La especie más abundante fue H. convergens. Estas especies podrían ser herramientas valiosas para ser empleadas en programas de control biológico, producciones orgánicas o en programas de manejo integrado de plagas.

The whitefly complex (Hemiptera: Aleyrodidae) includes some of the main pests of the French green bean (Phaseolus vulgaris L.). Among which, Bemisia tabaci is a vector of the golden mosaic virus that affects the quality and yield of the crop, with losses up to 100% and difficult control due to the resistance acquired by pests towards some agrochemicals. The French green bean ranks second among the non-traditional export products of Guatemala. Its agronomic management has been mainly through chemical control, which affects insects and other organisms that are not the objective of the control, such as: pollinators, beneficial insects, humans and wildlife. The objectives of the study were: to determine the presence of natural enemies native to the whitefly and identify the species of whitefly present in the cultivation of the French bean in Chimaltenango. For the study, four 300 m² plots were established, weekly sampling was carried out during two crop cycles. Five sites were sampled on each plot and five plants on each site. The native parasitoid species found were: Encarsia Formosa Gahan, Eretmocerus eremicus Rose and Zolnerowuch and Amitus fuscipennis MacGown and Nebeker, the most abundant species was A. fuscipennis. The predators identified were Chrysoperla carnea (Stephens) and Hippodamia convergens Guerin-Meneville. The most abundant species was H. convergens. These species could be valuable tools to be used in biological control programs, organic productions or in integrated pest management programs.

Construction of a high density linkage map and genome dissection of bruchid resistance in zombi pea (Vigna vexillata (L.) A. Rich).

Zombi pea (Vigna vexillata) is a legume crop that is resistant to several biotic and abiotic stresses. Callosobruchus maculatus and Callosobruchus chinensis are serious stored-insect pests of legume crops. We constructed a high-density linkage map and performed quantitative trait loci (QTLs) mapping for resistance to these insect species in zombi pea. An F2 population of 198 individuals from a cross between 'TVNu 240' (resistant) and 'TVNu 1623' (susceptible) varieties was used to construct a linkage map of 6,529 single nucleotide polymorphism markers generated from sequencing amplified fragments of specific loci. The map comprised 11 linkage groups, spanning 1,740.9 cM, with an average of 593.5 markers per linkage group and an average distance of 0.27 cM between markers. High levels of micro-synteny between V. vexillata and cowpea (Vigna unguiculata), mungbean (Vigna radiata), azuki bean (Vigna angularis) and common bean (Phaseolus vulgaris) were found. One major and three minor QTLs for C. chinensis resistance and one major and one minor QTLs for C. maculatus resistance were identified. The major QTLs for resistance to C. chinensis and C. maculatus appeared to be the same locus. The linkage map developed in this study will facilitate the identification of useful genes/QTLs in zombi pea.

Role of cyanogenic glycosides in the seeds of wild lima bean, Phaseolus lunatus: defense, plant nutrition or both?

MAIN CONCLUSION: Cyanogenic glycosides present in the seeds of wild lima bean plants are associated with seedling defense but do not affect seed germination and seedling growth. Wild lima bean plants contain cyanogenic glycosides (CNGs) that are known to defend the plant against leaf herbivores. However, seed feeders appear to be unaffected despite the high levels of CNGs in the seeds. We investigated a possible role of CNGs in seeds as nitrogen storage compounds that influence plant growth, as well as seedling resistance to herbivores. Using seeds from four different wild lima bean natural populations that are known to vary in CNG levels, we tested two non-mutually exclusive hypotheses: (1) seeds with higher levels of CNGs produce seedlings that are more resistant against generalist herbivores and, (2) seeds with higher levels of CNGs germinate faster and produce plants that exhibit better growth. Levels of CNGs in the seeds were negatively correlated with germination rates and not correlated with seedling growth. However, levels of CNGs increased significantly soon after germination and seeds with the highest CNG levels produced seedlings with higher CNG levels in cotyledons. Moreover, the growth rate of the generalist herbivore Spodoptera littoralis was lower in cotyledons with high-CNG levels. We conclude that CNGs in lima bean seeds do not play a role in seed germination and seedling growth, but are associated with seedling defense. Our results provide insight into the potential dual function of plant secondary metabolites as defense compounds and storage molecules for growth and development.

Vines avoid coiling around neighbouring plants infested by polyphagous mites.

Vines that coil around plants heavily infested with ambulate polyphagous mites can be heavily damaged by the mites. To explore whether vines avoid mite-infested plants, we observed the coiling responses of morning glory (Ipomoea nil var. Heavenly Blue) vines and bush killer (Cayratia japonica (Thunb) Gagnep) tendrils around nearby kidney bean (Phaseolus vulgaris L.) plants that were either uninfested or heavily infested with the two-spotted spider mite (Tetranychus urticae Koch). The proportions of I. nil vines that coiled around spider mite-infested and uninfested bean plants did not differ significantly; however, no C. japonica tendril coiled around spider mite-infested plants. The proportion of such tendrils was thus significantly lower than that around uninfested plants. The ability of C. japonica tendrils to avoid spider mite-infested plants would prevent serious "contact infections" by mites. We further found that tendril avoidance seemed to be attributable to the mite webs that covered infested plants; neither spider mite-induced bean volatiles nor spider mite intrusion onto tendrils seemed to explain the avoidance.

Tetranins: new putative spider mite elicitors of host plant defense.

The two-spotted spider mite (Tetranychus urticae) is a plant-sucking arthropod herbivore that feeds on a wide array of cultivated plants. In contrast to the well-characterized classical chewing herbivore salivary elicitors that promote plant defense responses, little is known about sucking herbivores' elicitors. To characterize the sucking herbivore elicitors, we explored putative salivary gland proteins of spider mites by using an Agrobacterium-mediated transient expression system or protein infiltration in damaged bean leaves. Two candidate elicitors (designated as tetranin1 (Tet1) and tetranin2 (Tet2)) triggered early leaf responses (cytosolic calcium influx and membrane depolarization) and increased the transcript abundances of defense genes in the leaves, eventually resulting in reduced survivability of T. urticae on the host leaves as well as induction of indirect plant defenses by attracting predatory mites. Tet1 and/or Tet2 also induced jasmonate, salicylate and abscisic acid biosynthesis. Notably, Tet2-induced signaling cascades were also activated via the generation of reactive oxygen species. The signaling cascades of these two structurally dissimilar elicitors are mostly overlapping but partially distinct and thus they would coordinate the direct and indirect defense responses in host plants under spider mite attack in both shared and distinct manners.

Endophytic entomopathogenic fungi enhance the growth of Phaseolus vulgaris L. (Fabaceae) and negatively affect the development and reproduction of Tetranychus urticae Koch (Acari: Tetranychidae).

Entomopathogenic fungi (EPF) have primarily been applied as an inundative approach to manage pests. However, in recent decade multifunctional role of EPF have been documented which provide multiple benefits to host plants when colonized as an endophyte. In this study five fungal isolates from the genus Beauveria (three), Isaria (one) and Lecanicillium (one) were evaluated for their ability to colonize common bean, Phaseolus vulgaris and to assess their effects in planta on plant growth promotion and possible negative effects on the two-spotted spider mites, Tetranychus urticae. All the tested isolates in this study were able to endophytically colonize root, stem and even leaves of inoculated plants examined at 7 and 14 days post inoculation, indicating the systemic colonization of EPF. Colonized plants showed increased plant heights, fresh shoot and root weights compared to plants without inoculation. Survivorship of T. urticae significantly differed among the treatments with higher survival probability in control plants. Significant reduction in larval development, adult longevity and female fecundity of spider mites were observed when fed on treated plants compared to control plants. The negative effects were found to be carried over the second generation fed on fresh plants. Overall, our results show (i) the positive effects of fungal endophytes on plant growth, (ii) reduction in population growth rate and (iii) negative effects of endophytes on growth and reproduction of spider mites in successive generations. The study presents reports on the endophytic management of plant-feeding mites and highlights the possibility of utilizing entomopathogenic fungal endophytes in the integrated pest management program.

Zein Nanoparticles as Eco-Friendly Carrier Systems for Botanical Repellents Aiming Sustainable Agriculture.

Botanical repellents represent one of the main ways of reducing the use of synthetic pesticides and the contamination of soil and hydric resources. However, the poor stability and rapid degradation of these compounds in the environment hinder their effective application in the field. Zein nanoparticles can be used as eco-friendly carrier systems to protect these substances against premature degradation, provide desirable release characteristics, and reduce toxicity in the environment and to humans. In this study, we describe the preparation and characterization of zein nanoparticles loaded with the main constituents of the essential oil of citronella (geraniol and R-citronellal). The phytotoxicity, cytotoxicity, and insect activity of the nanoparticles toward target and nontarget organisms were also evaluated. The botanical formulations showed high encapsulation efficiency (>90%) in the nanoparticles, good physicochemical stability, and effective protection of the repellents against UV degradation. Cytotoxicity and phytotoxicity assays showed that encapsulation of the botanical repellents decreased their toxicity. Repellent activity tests showed that nanoparticles containing the botanical repellents were highly repellent against the Tetranychus urticae Koch mite. This nanotechnological formulation offers a new option for the effective use of botanical repellents in agriculture, reducing toxicity, protecting against premature degradation, and providing effective pest control.

Viral metagenomics of aphids present in bean and maize plots on mixed-use farms in Kenya reveals the presence of three dicistroviruses including a novel Big Sioux River virus-like dicistrovirus.

BACKGROUND: Aphids are major vectors of plant viruses. Common bean (Phaseolus vulgaris L.) and maize (Zea mays L.) are important crops that are vulnerable to aphid herbivory and aphid-transmitted viruses. In East and Central Africa, common bean is frequently intercropped by smallholder farmers to provide fixed nitrogen for cultivation of starch crops such as maize. We used a PCR-based technique to identify aphids prevalent in smallholder bean farms and next generation sequencing shotgun metagenomics to examine the diversity of viruses present in aphids and in maize leaf samples. Samples were collected from farms in Kenya in a range of agro-ecological zones. RESULTS: Cytochrome oxidase 1 (CO1) gene sequencing showed that Aphis fabae was the sole aphid species present in bean plots in the farms visited. Sequencing of total RNA from aphids using the Illumina platform detected three dicistroviruses. Maize leaf RNA was also analysed. Identification of Aphid lethal paralysis virus (ALPV), Rhopalosiphum padi virus (RhPV), and a novel Big Sioux River virus (BSRV)-like dicistrovirus in aphid and maize samples was confirmed using reverse transcription-polymerase chain reactions and sequencing of amplified DNA products. Phylogenetic, nucleotide and protein sequence analyses of eight ALPV genomes revealed evidence of intra-species recombination, with the data suggesting there may be two ALPV lineages. Analysis of BSRV-like virus genomic RNA sequences revealed features that are consistent with other dicistroviruses and that it is phylogenetically closely related to dicistroviruses of the genus Cripavirus. CONCLUSIONS: The discovery of ALPV and RhPV in aphids and maize further demonstrates the broad occurrence of these dicistroviruses. Dicistroviruses are remarkable in that they use plants as reservoirs that facilitate infection of their insect replicative hosts, such as aphids. This is the first report of these viruses being isolated from either organism. The BSRV-like sequences represent a potentially novel dicistrovirus infecting A. fabae.

Multivariate statistics applied to the reaction of common bean plants to parasitism by Meloidogyne javanica.

The availability of common bean cultivars tolerant to Meloidogyne javanica is limited in Brazil. Thus, the present study aimed to evaluate the reactions of 33 common bean genotypes (23 landrace, 8 commercial, 1 susceptible standard and 1 resistant standard) to M. javanica, employing multivariate statistics to discriminate the reaction of the genotypes. The experiment was conducted in a greenhouse using a completely randomized design with seven replicates. The seeds were sown in 1-L pots containing autoclaved soil and sand in a 1:1 ratio (v:v). On day 19, after emergence of the seedlings, the plants were treated with inoculum containing 4000 eggs + second-stage juveniles (J2). At 60 days after inoculation, the seedlings were evaluated based on biometric and parasitism-related traits, such as number of galls, final nematode population per root system, reproduction factor, and percent reduction in the reproduction factor of the nematode (%RRF). The data were subjected to analysis of variance using the F-test. The Mahalanobis generalized distance was used to obtain the dissimilarity matrix, and the average linkage between groups was used for clustering. The use of multivariate statistics allowed groups to be separated according to the resistance levels of genotypes, as observed in the %RRF. The landrace genotypes FORT-09, FORT-17, FORT-31, FORT-32, FORT-34 and FORT-36 presented resistance to M. javanica; thus, these genotypes can be considered potential sources of resistance.

Double strand RNA delivery system for plant-sap-feeding insects.

Double-stranded RNA (dsRNA)-mediated gene silencing, also known as RNA interference (RNAi), has been a breakthrough technology for functional genomic studies and represents a potential tool for the management of insect pests. Since the inception of RNAi numerous studies documented successful introduction of exogenously synthesized dsRNA or siRNA into an organism triggering highly efficient gene silencing through the degradation of endogenous RNA homologous to the presented siRNA. Managing hemipteran insect pests, especially Halyomorpha halys (Stål) (Heteroptera: Pentatomidae), the brown marmorated stink bug (BMSB), is critical to food productivity. BMSB was recently introduced into North America where it is both an invasive agricultural pest of high value specialty, row, and staple crops, as well as an indoor nuisance pest. RNAi technology may serve as a viable tool to manage this voracious pest, but delivery of dsRNA to piercing-sucking insects has posed a tremendous challenge. Effective and practical use of RNAi as molecular biopesticides for biocontrol of insects like BMSB in the environment requires that dsRNAs be delivered in vivo through ingestion. Therefore, the key challenge for molecular biologists in developing insect-specific molecular biopesticides is to find effective and reliable methods for practical delivery of stable dsRNAs such as through oral ingestion. Here demonstrated is a reliable delivery system of effective insect-specific dsRNAs through oral feeding through a new delivery system to induce a significant decrease in expression of targeted genes such as JHAMT and Vg. This state-of-the-art delivery method overcomes environmental delivery challenges so that RNAi is induced through insect-specific dsRNAs orally delivered to hemipteran and other insect pests.

Intraspecific variation among Tetranychid mites for ability to detoxify and to induce plant defenses.

Two genotypes coexist among Kanzawa spider mites, one of which causes red scars and the other of which causes white scars on leaves, and they elicit different defense responses in host plants. Based on RNA-Seq analysis, we revealed here that the expression levels of genes involved in the detoxification system were higher in Red strains than White strains. The corresponding enzyme activities as well as performances for acaricide resistance and host adaptation toward Laminaceae were also higher in Red strains than White strains, indicating that Red strains were superior in trait(s) of the detox system. In subsequent generations of strains that had survived exposure to fenpyroximate, both strains showed similar resistance to this acaricide, as well as similar detoxification activities. The endogenous levels of salicylic acid and jasmonic acid were increased similarly in bean leaves damaged by original Red strains and their subsequent generations that inherited high detox activity. Jasmonic acid levels were increased in leaves damaged by original White strains, but not by their subsequent generations that inherited high detox activity. Together, these data suggest the existence of intraspecific variation - at least within White strains - with respect to their capacity to withstand acaricides and host plant defenses.

Isolated and Community Contexts Produce Distinct Responses by Host Plants to the Presence of Ant-Aphid Interaction: Plant Productivity and Seed Viability.

Ant-aphid interactions may affect host plants in several ways, however, most studies measure only the amount of fruit and seed produced, and do not test seed viability. Therefore, the aim of this study was to assess the effects of the presence of ant-aphid interactions upon host plant productivity and seed viability in two different contexts: isolated and within an arthropod community. For this purpose we tested the hypothesis that in both isolated and community contexts, the presence of an ant-aphid interaction will have a positive effect on fruit and seed production, seed biomass and rate of seed germination, and a negative effect on abnormal seedling rates, in comparison to plants without ants. We performed a field mesocosm experiment containing five treatments: Ant-aphid, Aphid, Community, Ant-free community and Control. We counted fruits and seeds produced by each treatment, and conducted experiments for seed biomass and germinability. We found that in the community context the presence of an ant-aphid interaction negatively affected fruit and seed production. We think this may be because aphid attendance by tending-ants promotes aphid damage to the host plant, but without an affect on seed weight and viability. On the other hand, when isolated, the presence of an ant-aphid interaction positively affected fruit and seed production. These positive effects are related to the cleaning services offered to aphids by tending-ants, which prevent the development of saprophytic fungi on the surface of leaves, which would cause a decrease in photosynthetic rates. Our study is important because we evaluated some parameters of plant fitness that have not been addressed very well by other studies involving the effects of ant-aphid interactions mainly on plants with short life cycles. Lastly, our context dependent approach sheds new light on how ecological interactions can vary among different methods of crop management.

Pathogenicity and Virulence of Soilborne Oomycetes on Phaseolus vulgaris.

Dry bean (Phaseolus vulgaris L.) is a globally important leguminous food crop. Yields can be reduced by high incidence of soilborne oomycetes that cause seedling disease. Breeders have attempted to develop Pythium root rot-resistant bean varieties; however, relationships between dry bean and most soilborne oomycete species remain uncharacterized. Oomycete species (n = 28), including Pythium spp. and Phytopythium spp., were tested in a growth chamber seedling assay at 20°C and an in vitro seed assay at 20°C and 26°C to evaluate their pathogenicity and virulence on 'Red Hawk' dark red kidney bean and 'Zorro' black bean. Root size or disease severity was significantly impacted by 14 oomycete species, though results varied by bean variety, temperature, and assay. Of these 14 pathogenic oomycete species, 11 species exhibited significant differences in DSI due to temperature on at least one bean variety. Pythium aphanidermatum, P. myriotylum, P. ultimum, P. ultimum var. sporangiiferium, and P. ultimum var. ultimum were the most virulent species in both assays, causing seed rot and pre-emergence damping-off of dry bean. Oomycete species were clustered into three groups based on symptom development: seed rot pathogens, root rot pathogens, or nonpathogens. Intraspecific variability in virulence was observed for eight of the 14 pathogenic oomycete species. Improved understanding of Pythium and Phytopythium interactions with dry bean may enable breeders and pathologists to more effectively evaluate strategies for oomycete seedling disease management.

Genotyping-by-Sequencing to Predict Resistance to Lima Bean Downy Mildew in a Diversity Panel.

Lima bean is affected by severe downy mildew epidemics caused by the oomycete Phytophthora phaseoli. There are six documented races of P. phaseoli (A to F). Race F is currently predominant in the mid-Atlantic region, creating the need for resistant lima bean cultivars with desirable agronomic characteristics. In order to develop markers for detecting race F resistance, bulked segregant analysis (BSA) using genotyping-by-sequencing (GBS) was used on a biparental F2 population comprised of 216 lima bean progeny segregating for a dominant race F resistance phenotype. Data were analyzed using a custom bioinformatic analysis pipeline (redrep). Kompetitive allele-specific polymerase chain reaction assays were developed using 12 GBS markers associated with the race F resistance phenotype. Using these assays, the F2 population was used to map the race F resistance locus. Seven markers were in linkage and significantly associated with race F resistance that mapped between two markers located approximately 4.88 centimorgan (cM) apart. These assays were successfully used to genotype a newly acquired lima bean diversity panel consisting of 256 landraces, cultivars, and wild germplasm, and a haplotype consisting of two of the seven linked markers was demonstrated to accurately predict race F resistance. This confirmed the ability of our customized methods to accurately predict phenotypes in diverse lines of lima bean.

Comparative Transcriptome Analysis of Resistant and Susceptible Common Bean Genotypes in Response to Soybean Cyst Nematode Infection.

Soybean cyst nematode (SCN; Heterodera glycines Ichinohe) reproduces on the roots of common bean (Phaseolus vulgaris L.) and can cause reductions in plant growth and seed yield. The molecular changes in common bean roots caused by SCN infection are unknown. Identification of genetic factors associated with SCN resistance could help in development of improved bean varieties with high SCN resistance. Gene expression profiling was conducted on common bean roots infected by SCN HG type 0 using next generation RNA sequencing technology. Two pinto bean genotypes, PI533561 and GTS-900, resistant and susceptible to SCN infection, respectively, were used as RNA sources eight days post inoculation. Total reads generated ranged between ~ 3.2 and 5.7 million per library and were mapped to the common bean reference genome. Approximately 70-90% of filtered RNA-seq reads uniquely mapped to the reference genome. In the inoculated roots of resistant genotype PI533561, a total of 353 genes were differentially expressed with 154 up-regulated genes and 199 down-regulated genes when compared to the transcriptome of non- inoculated roots. On the other hand, 990 genes were differentially expressed in SCN-inoculated roots of susceptible genotype GTS-900 with 406 up-regulated and 584 down-regulated genes when compared to non-inoculated roots. Genes encoding nucleotide-binding site leucine-rich repeat resistance (NLR) proteins, WRKY transcription factors, pathogenesis-related (PR) proteins and heat shock proteins involved in diverse biological processes were differentially expressed in both resistant and susceptible genotypes. Overall, suppression of the photosystem was observed in both the responses. Furthermore, RNA-seq results were validated through quantitative real time PCR. This is the first report describing genes/transcripts involved in SCN-common bean interaction and the results will have important implications for further characterization of SCN resistance genes in common bean.

Is protection against florivory consistent with the optimal defense hypothesis?

BACKGROUND: Plant defense traits require resources and energy that plants may otherwise use for growth and reproduction. In order to most efficiently protect plant tissues from herbivory, one widely accepted assumption of the optimal defense hypothesis states that plants protect tissues most relevant to fitness. Reproductive organs directly determining plant fitness, including flowers and immature fruit, as well as young, productive leaf tissue thus should be particularly well-defended. To test this hypothesis, we quantified the cyanogenic potential (HCNp)-a direct, chemical defense-systemically expressed in vegetative and reproductive organs in lima bean (Phaseolus lunatus), and we tested susceptibility of these organs in bioassays with a generalist insect herbivore, the Large Yellow Underwing (Noctuidae: Noctua pronuba). To determine the actual impact of either florivory (herbivory on flowers) or folivory on seed production as a measure of maternal fitness, we removed varying percentages of total flowers or young leaf tissue and quantified developing fruit, seeds, and seed viability. RESULTS: We found extremely low HCNp in flowers (8.66 ± 2.19 µmol CN(-) g(-1) FW in young, white flowers, 6.23 ± 1.25 µmol CN(-) g(-1) FW in mature, yellow flowers) and in pods (ranging from 32.05 ± 7.08 to 0.09 ± 0.08 µmol CN(-) g(-1) FW in young to mature pods, respectively) whereas young leaves showed high levels of defense (67.35 ± 3.15 µmol CN(-) g(-1) FW). Correspondingly, herbivores consumed more flowers than any other tissue, which, when taken alone, appears to contradict the optimal defense hypothesis. However, experimentally removing flowers did not significantly impact fitness, while leaf tissue removal significantly reduced production of viable seeds. CONCLUSIONS: Even though flowers were the least defended and most consumed, our results support the optimal defense hypothesis due to i) the lack of flower removal effects on fitness and ii) the high defense investment in young leaves, which have high consequences for fitness. These data highlight the importance of considering plant defense interactions from multiple angles; interpreting where empirical data fit within any plant defense hypothesis requires understanding the fitness consequences associated with the observed defense pattern.