Search

Transmission of insect-borne pathogens is mediated by interactions between insects and plants across variable environments. Water stress, for example, affects the physiology, defense, chemistry, and nutritional balance of plants in ways that alter their tolerance to herbivores and pathogens. However, few studies have explored interactions between water stress and insect-borne pathogens as well as the molecular mechanisms mediating these interactions. Here, we address these knowledge gaps by assessing effects of plant water stress on the transmission of a bacterial pathogen, Candidatus Liberibacter solanacearum (CLs), by the vector Bactericera cockerelli Šulc (potato psyllid). We hypothesized that plant water stress would promote pathogen transmission by inducing plant gene transcripts and phytohormones involved in defense. Our results showed water stress was associated with decreased CLs titer with two psyllid haplotypes. Our analysis of plant gene transcripts suggested water stress affected phytohormone pathways in ways that altered plant tolerance to the CLs pathogen. Our study shows that abiotic stressors like drought may mediate the spread of plant pathogens by altering plant signaling pathways in ways that affect pathogen transmission.

Mixed viral infections are common in fields and frequently exacerbate disease severity via synergistic interactions among individual viral genomic components leading to major crop loss. Two predominant species of tomato-infecting begomoviruses, Tomato leaf curl New Delhi virus (ToLCNDV) and Tomato leaf curl Gujarat virus (ToLCGuV), are known to cause severe leaf curl disease of tomato in India. Previously, we have demonstrated asymmetric synergism between these two distinct begomovirus species during mixed infection in solanaceous hosts. In the present study, we have identified the underlying proteins that positively regulate asymmetric synergism and their effect on plant defense machinery. During co-infection, the AC2 and AV2 of ToLCGuV enhanced ToLCNDV DNA accumulation in Nicotiana benthamiana as well as in their natural host, tomato. Furthermore, we found that AC2 and AV2 of ToLCNDV and AV2 of ToLCGuV play a critical role in suppression of post transcriptional gene silencing (PTGS) machinery. Taken together, AC2 and AV2 encoded proteins of ToLCGuV are the crucial viral factors promoting asymmetric synergism with ToLCNDV. KEY POINTS: • Begomoviral suppressors play vital roles in viral synergism. • AC2 and AV2 of ToLCGuV asymmetrically enhance ToLCNDV accumulation. • AC2 and AV2 of ToLCNDV and ToLCGuV AV2 are major PTGS suppressors.

Soil bacteria that form mutualisms with plants, such as rhizobia, affects susceptibility of plants to herbivores and pathogens. Soil rhizobia also promote nitrogen fixation, which mediates host nutrient levels and defenses. However, whether aboveground herbivores affect the function of soil rhizobia remains poorly understood. We assessed reciprocal interactions between Sitona lineatus, a chewing herbivore, and pea (Pisum sativum) plants grown with or without rhizobia (Rhizobium leguminosarum biovar viciae). We also examined the underlying plant-defense and nutritional mechanisms of these interactions. In our experiments, soil rhizobia influenced feeding and herbivory by chewing herbivores. Leaf defoliation by S. lineatus was lower on plants treated with rhizobia, but these insects had similar amino acid levels compared to those on un-inoculated plants. Plants grown with soil rhizobia had increased expression of gene transcripts associated with phytohormone-mediated defense, which may explain decreased susceptibility to S. lineatus. Rhizobia also induced expression of gene transcripts associated with physical and antioxidant-related defense pathways in P. sativum. Conversely, S. lineatus feeding reduced the number of root nodules and nodule biomass, suggesting a disruption of the symbiosis between plants and rhizobia. Our study shows that aboveground herbivores can engage in mutually antagonistic interactions with soil microbes mediated through a multitude of plant-mediated pathways.

The abundance and diversity of pollinator populations are in global decline. Managed pollinator species, like honey bees, and wild species are key ecosystem service providers in both natural and managed agroecosystems. However, relatively few studies have exhaustively characterized pollinator populations in diverse agroecosystems over multiple years, while also thoroughly documenting plant-pollinator interactions. Yet, such studies are needed to fulfill the national pollinator protection plans that have been released by the United States and other nations. Our research is among the first studies to respond to these directives by systematically documenting bee and plant biodiversity, bee-plant interactions, and bee-mediated pollen movement in farming systems of the Pacific Northwest, USA. Our data provides insight into the processes mediating pollinator and plant community assembly, persistence, and resilience across landscapes with variable crop and landscape diversity and agroecosystem management practices. These data will also contribute to the development of a United States pollinator database, supporting the United States' plan to promote pollinators. With few publicly available data sets that systematically take account of agroecosystem practices, plant populations, and pollinators, our research will provide future users the means to conduct synesthetic studies of pollinators and ecosystem function in a period of rapid and global pollinator declines. There are no copyright or proprietary restrictions for research or teaching purposes. Usage of the data set must be cited.

Crop-associated microbiota are key factors affecting host health and productivity. Most crops are grown within heterogeneous landscapes, and interactions between management practices and landscape context often affect plant and animal biodiversity in agroecosystems. However, whether these same factors typically affect crop-associated microbiota is less clear. Here, we assessed whether orchard management strategies and landscape context affected bacterial and fungal communities in pear (Pyrus communis) flowers. We found that bacteria and fungi responded differently to management schemes. Organically-certified orchards had higher fungal diversity in flowers than conventional or bio-based integrated pest management (IPM) orchards, but organic orchards had the lowest bacterial diversity. Orchard management scheme also best predicted the distribution of several important bacterial and fungal genera that either cause or suppress disease, with organic and bio-based IPM best explaining the distributions of bacterial and fungal genera, respectively. Moreover, patterns of bacterial and fungal diversity were affected by interactions between management, landscape context, and climate. When examining the similarity of bacterial and fungal communities across sites, both abundance- and taxa-related turnover were mediated primarily by orchard management scheme and landscape context, and specifically the amount of land in cultivation. Our study reveals local- and landscape-level drivers of floral microbiome structure in a major fruit crop, providing insights that can inform microbiome management to promote host health and high-yielding quality fruit.IMPORTANCE.In tree fruits, proper crop management during bloom is essential for producing disease-free fruit. Tree fruits are often grown in heterogeneous landscapes; however, few studies have assessed whether landscape context and crop management affect the floral microbiome, which plays a critical role in shaping plant health and disease tolerance. Such work is key for identification of tactics and/or contexts where beneficial microbes proliferate, and pathogenic microbes are limited. Here, we characterize the floral microbiome of pear crops in Washington State, USA, where major production occurs in inter-mountain valleys and basins with variable elevation and microclimates. Our results show that both local- (crop management) and landscape-level (habitat types and climate) factors affect floral microbiota, but in disparate ways for each kingdom. More broadly, these findings can potentially inform microbiome management in orchards for promotion of host health and high-quality yields.

Plants are often attacked by multiple antagonists, and traits of the attacking organisms, and their order of arrival onto hosts, may affect plant defenses. However, few studies have assessed how multiple antagonists, and varying attack order, affect plant defense or nutrition. To address this, we assessed defensive and nutritional responses of Pisum sativum plants after attack by a vector herbivore (Acrythosiphon pisum), a non-vector herbivore (Sitona lineatus), and a pathogen (Pea enation mosaic virus, PEMV). We show PEMV-infectious A. pisum induced several pathogen-specific plant defense signals, but these defenses were inhibited when S. lineatus was present in peas infected with PEMV. In contrast, feeding by S. lineatus induced anti-herbivore defense signals, but these defenses were enhanced by PEMV. Sitonalineatus also increased abundance of plant amino acids, but only when they attacked after PEMV-infectious A. pisum. Our results suggest that diverse communities of biotic antagonists alter defense and nutritional traits of plants through complex pathways that depend on the identity of attackers and their order of arrival onto hosts. Moreover, we show interactions among a group of biotic stressors can vary along a spectrum from antagonism to enhancement/synergism based on the identity and order of attackers, and these interactions are mediated by a multitude of phytohormone pathways.

Arthropods that vector plant pathogens often interact with predators within food webs. Predators affect vectors by eating them (consumptive effects) and by inducing antipredator behaviors (non-consumptive effects), and these interactions may affect transmission of vector-borne pathogens. However, it has proven difficult to experimentally tease apart the effects of predators on vector fitness and behavior as they are often correlated. We addressed this problem by assessing how both aphids and an aphid-borne pathogen were affected by variable predation risk. Specifically, we experimentally manipulated ladybeetle predators’ mouthparts to isolate consumptive, and non-consumptive, effects of predators on aphid fitness, movement, and virus transmission. We show that although lethal predators decreased aphid vector abundance, they increased pathogen transmission by increasing aphid movement among hosts. Moreover, aphids responded to risk of predation by moving to younger plant tissue that was more susceptible to the pathogen. Aphids also responded to predator risk through compensatory reproduction, which offset direct consumptive effects. Our results support predictions of disease models showing alterations of vector movement due to predators can have greater effects on transmission of pathogens than vector consumption. Broadly, our study shows isolating direct and indirect predation effects can reveal novel pathways by which predators affect vector-borne pathogens.

Crop-associated microbiota are key factors affecting host health and productivity. Most crops are grown within heterogeneous landscapes, and interactions between management practices and landscape context often affect plant and animal biodiversity in agroecosystems. However, whether these same factors typically affect crop-associated microbiota is less clear. Here, we assessed whether orchard management strategies and landscape context affected bacterial and fungal communities in pear (Pyrus communis) flowers. We found that bacteria and fungi responded differently to management schemes. Organically-certified orchards had higher fungal diversity in flowers than conventional or bio-based integrated pest management (IPM) orchards, but organic orchards had the lowest bacterial diversity. Orchard management scheme also best predicted the distribution of several important bacterial and fungal genera that either cause or suppress disease, with organic and bio-based IPM best explaining the distributions of bacterial and fungal genera, respectively. Moreover, patterns of bacterial and fungal diversity were affected by interactions between management, landscape context, and climate. When examining the similarity of bacterial and fungal communities across sites, both abundance- and taxa-related turnover were mediated primarily by orchard management scheme and landscape context, and specifically the amount of land in cultivation. Our study reveals local- and landscape-level drivers of floral microbiome structure in a major fruit crop, providing insights that can inform microbiome management to promote host health and high-yielding quality fruit.IMPORTANCEIn tree fruits, proper crop management during bloom is essential for producing disease-free fruit. Tree fruits are often grown in heterogeneous landscapes; however, few studies have assessed whether landscape context and crop management affect the floral microbiome, which plays a critical role in shaping plant health and disease tolerance. Such work is key for identification of tactics and/or contexts where beneficial microbes proliferate, and pathogenic microbes are limited. Here, we characterize the floral microbiome of pear crops in Washington State, USA, where major production occurs in inter-mountain valleys and basins with variable elevation and microclimates. Our results show that both local (crop management) and landscape (habitat types and climate) level factors affect floral microbiota, but in disparate ways for each kingdom, suggesting a need for unique management strategies for each group. More broadly, these findings can potentially inform microbiome management in orchards for promotion of host health and high-quality yields.

Mixed viral infections are common in fields and frequently exacerbate disease severity via synergistic interactions among individual viral genomic components leading to major crop loss. Two predominant species of tomato-infecting begomoviruses, Tomato leaf curl New Delhi virus (ToLCNDV) and Tomato leaf curl Gujarat virus (ToLCGuV), are known to cause severe leaf curl disease of tomato in India. Previously, we have demonstrated asymmetric synergism between these two distinct begomovirus species during mixed infection in solanaceous hosts. In the present study, we have identified the underlying proteins that positively regulate asymmetric synergism and their effect on plant defense machinery. During co-infection, the AC2 and AV2 of ToLCGuV enhanced ToLCNDV DNA accumulation in Nicotiana benthamiana as well as in their natural host, tomato. Furthermore, we found that AC2 and AV2 of ToLCNDV and AV2 of ToLCGuV play a critical role in suppression of post transcriptional gene silencing (PTGS) machinery. Taken together, AC2 and AV2 encoded proteins of ToLCGuV are the crucial viral factors promoting asymmetric synergism with ToLCNDV. KEY POINTS: • Begomoviral suppressors play vital roles in viral synergism. • AC2 and AV2 of ToLCGuV asymmetrically enhance ToLCNDV accumulation. • AC2 and AV2 of ToLCNDV and ToLCGuV AV2 are major PTGS suppressors.

ToLCNDV-AV2 is essential for repression of - antiviral silencing mediated by RNA-dependent RNA polymerase 1 and is responsible for contrasting symptom development in Nicotiana species infected with different geminiviruses., RNA silencing is an integral part of the cellular defense mechanisms in plants that act against virus infection. However, the specific role of RNA silencing and the interplay between host and virus components during recovery from geminivirus infection remains unknown. Hence, in this study we aimed to examine the mechanism behind the host-specific recovery of Nicotiana tabacum infected with Tomato leaf curl Gujarat virus (ToLCGV). Unlike Tomato leaf curl New Delhi virus (ToLCNDV), ToLCGV infection resulted in symptom remission in N. tabacum, and we found that this was mainly due to cross-talk between the pre-coat protein (encoded by the AV2 ORF) of the virus and the host RNA-silencing component RNA-dependent RNA polymerase 1 (encoded by NtRDR1) of N. tabacum. Moreover, apart from the AV2 mutant, other mutants of ToLCNDV developed severe symptoms on a transgenic NtRDR1-overexpression line of N. benthamiana. In contrast, inoculation with ToLCGV resulted in symptom remission, which was due to enhanced methylation of the ToLCGV promoter. Our study reveals a novel ‘arms race’ in which the pre-coat protein of ToLCNDV selectively blocks the recovery process through inhibiting host-specific RDR1-mediated antiviral silencing in tobacco.

Herbivores that transmit plant pathogens often share hosts with non-vector herbivores. These co-occurring herbivores can affect vector fitness and behaviour through competition and by altering host plant quality. However, few studies have examined how such interactions may both directly and indirectly influence the spread of a plant pathogen. Here, we conducted field and greenhouse trials to assess whether a defoliating herbivore ( Sitona lineatus ) mediated the spread of a plant pathogen, Pea enation mosaic virus (PEMV), by affecting the fitness and behaviour of Acrythosiphon pisum , the PEMV vector. We observed higher rates of PEMV spread when infectious A. pisum individuals shared hosts with S. lineatus individuals. Using structural equation models, we showed that herbivory from S. lineatus increased A. pisum fitness, which stimulated vector movement and PEMV spread. Moreover, plant susceptibility to PEMV was indirectly enhanced by S. lineatus , which displaced A. pisum individuals to the most susceptible parts of the plant. Subsequent analyses of plant defence genes revealed considerable differences in plant phytohormones associated with anti-herbivore and anti-pathogen defence when S. lineatus was present. Given that vectors interact with non-vector herbivores in natural and managed ecosystems, characterizing how such interactions affect pathogens would greatly enhance our understanding of disease ecology.

Quantitative reverse transcription PCR (RT-qPCR) is one of the most efficient, reliable and widely used techniques to quantify gene expression. In this study, we evaluated the performance of six southern corn rootworm, Diabrotica undecimpunctata howardi (Barber), housekeeping genes (HKG), β-actin (Actin), β-tubulin (Tubulin), elongation factor 1 alpha (EF1α), glyceraldehyde-3 phosphate dehydrogenase (GAPDH), 40 S ribosomal protein S9 (RpS9) and ubiquitin-conjugating protein (Ubi), under different experimental conditions such as developmental stage, exposure of neonate and adults to dsRNA, exposure of adults to different temperatures, different 3rd instar larva tissues, and neonate starvation. The HKGs were analyzed with four algorithms, including geNorm, NormFinder, BestKeeper, and delta-CT. Although the six HKGs showed a relatively stable expression pattern among different treatments, some variability was observed. Among the six genes, EF1α exhibited the lowest Ct values for all treatments while Ubi exhibited the highest. Among life stages and across treatments, Ubi exhibited the least stable expression pattern. GAPDH, Actin, and EF1α were among the most stable HKGs in the majority of the treatments. This research provides HKG for accurate normalization of RT-qPCR data in the southern corn rootworm. Furthermore, this information can contribute to future genomic and functional genomic research in Diabrotica species.

Many insect herbivores are vectors that transmit plant pathogens as they forage. Within food webs, vectors interact with a range of host plants, other herbivores, and predators. Yet, few studies have examined how tri-trophic interactions involving vectors affect the spread of pathogens. Here we assessed effects of food web structure on aphid vectors and the prevalence of an aphid-borne persistent pathogen (Pea enation mosaic virus, PEMV) in pea plants. We experimentally manipulated ladybird predators, alternative host plants, and non-vector herbivores and assessed responses of pea aphids and PEMV using structural equation models. We show that variation in bottom-up, top-down, and horizontal interactions mediated PEMV prevalence. Predators reduced PEMV prevalence by consuming aphids, but an alternative host plant (vetch) had the opposite effect by promoting aphid movement and abundance. Non-vector herbivores (pea leaf weevil) increased PEMV susceptibility in peas. In turn, weevils offset the positive effects of predators on PEMV, but increased the negative effects of vetch. Our results show that tri-trophic interactions within insect and plant food webs can mediate vector biology with synergistic and opposing effects on pathogens. Continuing to assess how community-wide interactions affect vectors will aid in our understanding of vector-borne pathosystems.

Supporting Figures and Tables

Chilli, which encompasses several species in the genus Capsicum, is widely consumed throughout the world. In the Indian subcontinent, production of chilli is constrained due to chilli leaf curl disease (ChiLCD) caused by begomoviruses. Despite the considerable economic consequences of ChiLCD on chilli cultivation in India, there have been scant studies of the genetic diversity and structure of the begomoviruses that cause this disease. Here we report on a comprehensive survey across major chilli-growing regions in India. Analysis of samples collected in the survey indicates that ChiLCD-infected plants are associated with a complex of begomoviruses (including one previously unreported species) with a diverse group of betasatellites found in crops and weeds. The associated betasatellites neither enhanced the accumulation of the begomovirus components nor reduced the incubation period in Nicotiana benthamiana. The ChiLCD-associated begomoviruses induced mild symptoms on Capsicum spp., but both the level of helper virus that accumulated and the severity of symptoms were increased in the presence of cognate betasatellites. Interestingly, most of the begomoviruses were found to be intra-species recombinants. The betasatellites possess high nucleotide variability, and recombination among them was also evident. The nucleotide substitution rates were determined for the AV1 gene of begomoviruses (2.60 × 10- 3 substitutions site- 1 year- 1) and the βC1 gene of betasatellites [chilli leaf curl betasatellite (ChiLCB), 2.57 × 10- 4 substitution site- 1 year- 1; tomato leaf curl Bangladesh betasatellite (ToLCBDB), 5.22 × 10- 4 substitution site- 1 year- 1]. This study underscores the current understanding of Indian ChiLCD-associated begomoviruses and also demonstrates the crucial role of betasatellites in severe disease development in Capsicum spp.

Signaling networks among multiple phytohormones fine-tune plant defense responses to insect herbivore attack. Previously, it was reported that the synergistic combination of ethylene (ET) and jasmonic acid (JA) was required for accumulation of the maize insect resistance1 (mir1) gene product, a cysteine (Cys) proteinase that is a key defensive protein against chewing insect pests in maize (Zea mays). However, this study suggests that mir1-mediated resistance to corn leaf aphid (CLA; Rhopalosiphum maidis), a phloem sap-sucking insect pest, is independent of JA but regulated by the ET-signaling pathway. Feeding by CLA triggers the rapid accumulation of mir1 transcripts in the resistant maize genotype, Mp708. Furthermore, Mp708 provided elevated levels of antibiosis (limits aphid population)- and antixenosis (deters aphid settling)-mediated resistance to CLA compared with B73 and Tx601 maize susceptible inbred lines. Synthetic diet aphid feeding trial bioassays with recombinant Mir1-Cys Protease demonstrates that Mir1-Cys Protease provides direct toxicity to CLA. Furthermore, foliar feeding by CLA rapidly sends defensive signal(s) to the roots that trigger belowground accumulation of the mir1, signifying a potential role of long-distance signaling in maize defense against the phloem-feeding insects. Collectively, our data indicate that ET-regulated mir1 transcript accumulation, uncoupled from JA, contributed to heightened resistance to CLA in maize. In addition, our results underscore the significance of ET acting as a central node in regulating mir1 expression to different feeding guilds of insect herbivores.

Chewing herbivores, such as caterpillars and beetles, while feeding on the host plant, cause extensive tissue damage and release a wide array of cues to alter plant defenses. Consequently, the cues can have both beneficial and detrimental impacts on the chewing herbivores. Herbivore-associated molecular patterns (HAMPs) are molecules produced by herbivorous insects that aid them to elicit plant defenses leading to impairment of insect growth, while effectors suppress plant defenses and contribute to increased susceptibility to subsequent feeding by chewing herbivores. Besides secretions that originate from glands (e.g., saliva) and fore- and midgut regions (e.g., oral secretions) of chewing herbivores, recent studies have shown that insect frass and herbivore-associated endosymbionts also play a critical role in modulating plant defenses. In this review, we provide an update on a growing body of literature that discusses the chewing insect HAMPs and effectors and the mechanisms by which they modulate host defenses. Novel “omic” approaches and availability of new tools will help researchers to move forward this discipline by identifying and characterizing novel insect HAMPs and effectors and how these herbivore-associated cues are perceived by host plant receptors.

The vasculature of plants act as a channel for transport of signal(s) that facilitate long-distance intraplant communication. In maize, Maize insect resistance1-Cysteine Protease (Mir1-CP), which has homology to papain-like proteases, provides defense to different feeding guilds of insect pests. Furthermore, accumulation of Mir1-CP in the vasculature suggests that Mir1-CP can potentially function as a phloem-mobile protein. In a recent study, we provided evidence that Mir1-CP can curtail the growth of phloem-sap sucking insect, corn leaf aphid (CLA; Rhopalosiphum maidis). Our current study further examined whether aboveground feeding by CLA can induce resistance to subsequent herbivory by belowground feeding western corn rootworm (WCR; Diabrotica virgifera virgifera). Aboveground feeding by CLA systemically induced the accumulation of Mir1-CP in the roots. Furthermore, foliage feeding by CLA provided enhanced resistance to subsequent herbivory by belowground feeding of WCR. Taken together, our previous findings and results presented here indicate that long-distance transport of Mir1-CP is critical for providing enhanced resistance to insect attack in maize.

Plant defenses to insect herbivores have been studied in response to several insect behaviors on plants such as feeding, crawling, and oviposition. However, we have only scratched the surface about how insect feces induce plant defenses. In this study, we measured frass-induced plant defenses in maize, rice, cabbage, and tomato by chewing herbivores such as European corn borer (ECB), fall armyworm (FAW), cabbage looper (CL), and tomato fruit worm (TFW). We observed that caterpillar frass induced plant defenses are specific to each host-herbivore system, and they may induce herbivore or pathogen defense responses in the host plant depending on the composition of the frass deposited on the plant, the plant organ where it is deposited, and the species of insect. This study adds another layer of complexity in plant-insect interactions where analysis of frass-induced defenses has been neglected even in host-herbivore systems where naturally frass accumulates in enclosed feeding sites over extended periods of time.

Viruses are one of the most devastating plant pathogens causing severe economic losses worldwide. RNA silencing is a robust technology to knock down the expression of specific genes. This mechanism can be exploited to generate virus resistant plants through expression of the viral derived sequences. Viruses in turn have evolved to encode suppressors of RNA silencing to combat host defense. Mixed infection of plants is of common occurrence in nature and simultaneous targeting of suppressor(s) of multiple viruses offers an effective strategy. In this study, we have in silico designed siRNAs against suppressors of the two most devastating viruses of tomato, leaf curl causing tomato begomoviruses and Cucumber mosaic virus. Three different siRNA prediction programs were used to evaluate siRNAs generating capability of each sequence and common putative candidate siRNAs were selected fulfilling the stringent parameters. Our results indicated that in the case of each suppressor a particular region of 100–150 base pairs could be source of potent siRNAs referred as hotspots. Expression of these viral hot spots as a single construct in the plants would facilitate development of transgenic plants with a high degree of broad spectrum resistance against multiple viruses.