Herbarium specimens reveal herbivory patterns across the genus Cucurbita.

PREMISE: Quantifying how closely related plant species differ in susceptibility to insect herbivory is important for understanding the variation in evolutionary pressures on plant functional traits. However, empirically measuring in situ variation in herbivory spanning the geographic range of a plant-insect complex is logistically difficult. Recently, new methods have been developed using herbarium specimens to investigate patterns in plant-insect symbioses across large geographic scales. Such investigations provide insights into how accelerated anthropogenic changes may impact plant-insect interactions that are of ecological or agricultural importance. METHODS: Here, we analyze 274 pressed herbarium samples to investigate variation in herbivory damage in 13 different species of the economically important plant genus Cucurbita (Cucurbitaceae). This collection is composed of specimens of wild, undomesticated Cucurbita that were collected from across their native range, and Cucurbita cultivars collected from both within their native range and from locations where they have been introduced for agriculture in temperate North America. RESULTS: Herbivory is common on individuals of all Cucurbita species collected throughout their geographic ranges. However, estimates of herbivory varied considerably among individuals, with mesophytic species accruing more insect damage than xerophytic species, and wild specimens having more herbivory than specimens collected from human-managed habitats. CONCLUSIONS: Our study suggests that long-term evolutionary changes in habitat from xeric to mesic climates and wild to human-managed habitats may mediate the levels of herbivory pressure from coevolved herbivores. Future investigations into the potential factors that contribute to herbivory may inform the management of domesticated crop plants and their insect herbivores.

Identification of Bacterial Wilt (Erwinia tracheiphila) Resistances in USDA Melon Collection.

Bacterial wilt (BW) caused by the Gram-negative bacterium, Erwinia tracheiphila (Et.), is an important disease in melon (Cucumis melo L.). BW-resistant commercial melon varieties are not widely available. There are also no effective pathogen-based disease management strategies as BW-infected plants ultimately die. The purpose of this study is to identify BW-resistant melon accessions in the United States Department of Agriculture (USDA) collection. We tested 118 melon accessions in two inoculation trials under controlled environments. Four-week-old seedlings of test materials were mechanically inoculated with the fluorescently (GFP) labeled or unlabeled E. tracheiphila strain, Hca1-5N. We recorded the number of days to wilting of inoculated leaf (DWIL), days to wilting of whole plant (DWWP) and days to death of the plant (DDP). We identified four melon lines with high resistance to BW inoculation based on all three parameters. Fluorescent microscopy was used to visualize the host colonization dynamics of labeled bacteria from the point of inoculation into petioles, stem and roots in resistant and susceptible melon accessions, which provides an insight into possible mechanisms of BW resistance in melon. The resistant melon lines identified from this study could be valuable resistance sources for breeding of BW resistance as well as the study of cucurbit-E. tracheiphila interactions.

Biological Control and Microbial Ecology Draft Genome Sequence Data of Glutamicibacter sp. FBE-19, a Bacterium Antagonistic to the Plant Pathogen Erwinia tracheiphila.

Glutamicibacter sp. FBE-19 was isolated based on its strong antagonism to the cucurbit bacterial blight pathogen Erwinia tracheiphila on plates. Members of the Glutamicibacter genus can promote plant growth under saline conditions and antagonize fungi on plates via chitinolytic activity; however, their production of antibacterial compounds has not been examined. Here, we report the genome sequence of strain FBE-19. The genome is 3.85 Mbp with a G+C content of 60.1% and comprises 3,791 genes. Genes that may contribute to its antagonistic activity include genes for the secondary metabolites stenothricin, salinosporamide A, a second ß-lactone compound, and a carotenoid. The Glutamicibacter sp. FBE-19 genome data may be a useful resource if this strain proves to be an effective biocontrol agent against E. tracheiphila.

Discovery and Characterization of Low-Molecular Weight Inhibitors of Erwinia tracheiphila.

Plant pathogenic bacteria in the genus Erwinia cause economically important diseases, including bacterial wilt of cucurbits caused by Erwinia tracheiphila. Conventional bactericides are insufficient to control this disease. Using high-throughput screening, 464 small molecules (SMs) with either cidal or static activity at 100 µM against a cucumber strain of E. tracheiphila were identified. Among them, 20 SMs (SM1 to SM20), composed of nine distinct chemical moiety structures, were cidal to multiple E. tracheiphila strains at 100 µM. These lead SMs had low toxicity to human cells and honey bees at 100 µM. No phytotoxicity was observed on melon plants at 100 µM, except when SM12 was either mixed with Silwet L-77 and foliar sprayed or when delivered through the roots. Lead SMs did not inhibit the growth of beneficial Pseudomonas and Enterobacter species but inhibited the growth of Bacillus species. Nineteen SMs were cidal to Xanthomonas cucurbitae and showed >50% growth inhibition against Pseudomonas syringae pv. lachrymans. In addition, 19 SMs were cidal or static against Erwinia amylovora in vitro. Five SMs demonstrated potential to suppress E. tracheiphila when foliar sprayed on melon plants at 2× the minimum bactericidal concentration. Thirteen SMs reduced Et load in melon plants when delivered via roots. Temperature and light did not affect the activity of SMs. In vitro cidal activity was observed after 3 to 10 h of exposure to these five SMs. Here, we report 19 SMs that provide chemical scaffolds for future development of bactericides against plant pathogenic bacterial species.

Potential biological control of Erwinia tracheiphila by internal alimentary canal interactions in Acalymma vittatum with Pseudomonas fluorescens.

AIMS: We aim to determine if Pseudomonas fluorescens is a viable biological control for Erwinia tracheiphila within the insect vector, Acalymma vittatum. METHODS AND RESULTS: Pseudomonas fluorescens secreted fluorescein and inhibited growth of E. tracheiphila in disc diffusion assays. To determine if this antagonism was conserved within the insect vector, we performed in vivo assays by orally injecting beetles with bacterial treatments and fluorescent in situ hybridization to determine bacterial presence within the alimentary canal. CONCLUSIONS: Pseudomonas fluorescens inhibited the growth of E. tracheiphila on a nutrient-limiting medium. In situ experiments demonstrated that P. fluorescens is maintained within the alimentary canal of the beetle for at least 4 days, and co-occurred with E. tracheiphila. When beetles were first presented with Pseudomonas and then challenged with E. tracheiphila, E. tracheiphila was not recovered via FISH after 4 days. These data suggest that P. fluorescens has potential as a biological control agent to limit E. tracheiphila within the insect vector. SIGNIFICANCE AND IMPACT OF THE STUDY: This is a novel approach for controlling E. tracheiphila that has the potential to decrease reliance on insecticides, providing a safer environment for pollinators and growers.

Modified inoculation and disease assessment methods reveal host specificity in Erwinia tracheiphila-Cucurbitaceae interactions.

We conducted a greenhouse trial to determine specific compatible interactions between Erwinia tracheiphila strains and cucurbit host species. Using a modified inoculation system, E. tracheiphila strains HCa1-5N, UnisCu1-1N, and MISpSq-N were inoculated to cucumber (Cucumis sativus) cv. 'Sweet Burpless', melon (Cucumis melo) cv. 'Athena Hybrid', and squash (Cucubita pepo) cv. 'Early Summer Crookneck'. We observed symptoms and disease progression for 30 days; recorded the number of days to wilting of the inoculated leaf (DWIL), days to wilting of the whole plant (DWWP), and days to death of the plant (DDP). We found significant interactions between host cultivar and pathogen strains, which imply host specificity. Pathogen strains HCa1-5N and UnisCu1-1N isolated from Cucumis species exhibited more virulence in cucumber and melon than in squash, while the reverse was true for strain MISpSq-N, an isolate from Cucurbita spp. Our observations confirm a previous finding that E. tracheiphila strains isolated from Cucumis species were more virulent on Cucumis hosts and those from Cucubita were more virulent on Cucubita hosts. This confirmation helps in better understanding the pathosystem and provides baseline information for the subsequent development of new disease management strategies for bacterial wilt. We also demonstrated the efficiency of our modified inoculation and disease scoring methods.