The Northeast Arugula Team (NEAT): Evaluation and Mitigation of Limitations to Profitability for Arugula and Brassica Leafy Green Production

Identification of arugula and brassica leafy green bacterial pathogens.

Bacterial disease outbreaks were recognized through NEAT scouting & collection efforts, collections made by an extension personnel, and from growers contacting us. For each plant sample, bacteria were isolated using published methods using KBBC, a semi-selective medium for Pseudomonas and general nutrient agar media. Pure cultures were stored, and they were used in hypersensitivity test on tobacco to determine if isolates are likely to be plant pathogens. Organisms inducing an HR in 24-48 hours were stored. The strains selected for long-term storage are then used in PCR experiments to identify the species of the pathogen. Once the bacteria are identified, we will conduct pathogenicity tests on the plants from which they were isolated (arugula or other brassica leafy green). To date, we have collected approximately 132 samples of arugula and brassica leafy greens. In the 2023 growing season, we collected isolates from 62 samples from five Pennsylvania arugula growing locations and none from other states. Of these 62 samples, 13 plant pathogenic bacteria were isolated, six of which have been successfully genotyped using multi locus sequence analysis. Six Pseudomonas allivorans isolates or a closely related member of the P. syringae species complex were identified. In March 2024 a Grower Network Webinar was held and we demonstrated the differences between significant bacterial disease of arugula and BLGs for growers and NEAT collaborators. This material was also shared with clinic directors from the Northeast Plant Diagnostics Network (NEPDN). In 2024, 34 samples were collected or received by NEAT members. Nine farms were scouted for bacterial disease outbreaks. Of the 34 samples collected, six induced an HR+ response in tobacco. Through the efforts of 16S PCR, four of these six strains are believed to be Pseudomonas viridiflava, an opportunistic plant pathogen. In 2025, we processed 36 samples of diseased arugula and brassica leafy green tissue, as well as over 10,000 seeds from arugula and mustard seed lots. 42 farms were scouted for bacterial disease outbreaks. In diseased mustard microgreen samples sent from New York, we successfully identified our target pathogen, Pseudomonas cannabina pv. alisalensis, the causal agent of bacterial blight in arugula and brassica leafy greens. For the remaining isolates, we have yet to identify any known plant pathogens. At the end of 2025, we determined 11 of our recently isolated strains may be plant pathogens via HR on tobacco. We will proceed with further genetic identification and pathogenicity testing in 2026.

In collaborative work with cooperator Dr. Hehe Wang, we published an article which highlights a qPCR rapid detection method for Pseudomonas cannabina pv. alisalensis, the causal agent of bacterial blight in arugula and brassica leafy greens. We now utilize this method to rapidly detect bacterial blight in arugula samples. (A qPCR assay for specific detection of Pseudomonas cannabina pv. alisalensis”, Noh et al., 2024 DOI: 10.1094/PDIS-06-24-1217-RE ). In 2025, we contributed to the determination of specificity to newly developed conventional PCR protocols for Pseudomonas alliivorans, an emerging pathogen isolated from crucifers as part of this part of this project. This protocol has been applied to over 58 strains in our collection to determine their identity and were used to confirm the identity of P. alliivorans isolated in rapini and radish in 2023.This work will be published in the coming year and was reported at a regional scientific conference. MacLellan, M. P., Ray, S., Bushong, K., Paret, M., Potnis, N., Bull, C. (Author), Jones, J., & Dutta, B. Genetic traits of Pseudomonas alliivorans strains isolated from plants in the Southeastern United States. APS Souther Division Meeting, March 9-13, 2025.

Field Germplasm Evaluation

In fall 2023, a preliminary field trial was established to determine nutrient management, irrigation techniques, and pest management strategies for our farming situation where we had not previously grown brassica leafy greens. We were able to successfully grow commercial arugula cultivars under the parameter recommended to NEAT by our PAC & cooperator Bill Boone. We grew 10 cultivars in randomized plots which we did preliminary evaluations on for bolting and flea beetle damage and scouted regularly for bacterial diseases. The cultivars used included: five Eruca sativa cultivars (‘Astro’, ‘Runaway’, ‘Esmee’, ‘Standard’, ‘Balboa’), four Diplotaxis tenuifolia cultivars (‘Bellezia’, ‘Sylvetta’, ‘Wild’, ‘Dragon’s Fire’), and one cultivar of Diplotaxis erucoides (‘Wasabi’). Arugula was planted in plots at high density (3-5 seeds per inch) and in 36 inch raised beds as recommended to us by farmers from NEAT’s PAC as industry standards. Plots were randomized throughout raised beds and each raised bed within the research field is considered a replicate block. Overhead irrigation was utilized, a 20-10-10 NPK nutrient treatment was applied, and no forms of pest control have been applied. Observations were made twice a week to record bolting and pest damage. Dates were recorded for the first signs of bolting within a plot and once >50% of the plants within a plot have bolted. However, late season plantings, cold weather, & shorter days delayed bolting in most cultivars. Consistent bolting throughout the field was only observed in the ‘Bellezia’, ‘Sylvetta’, ‘Wild’, and ‘Wasabi’ cultivars by the time frost terminated the plantings in December 2023. Additionally, no flea beetle damage was seen in any cultivar in this field. This was likely due to the colder weather and late season start. In 2024, two successful field evaluations were conducted in the early summer and fall seasons at the Rock Springs Research Station at Penn State. 26 commercially available cultivars of arugula were screened for resistance to P. cannabina pv. alisalensis. The 26 cultivars evaluated were distributed among three species of arugula: 14 Diplotaxis tenuifolia (‘Aphrodite’, ‘Apollo’, ‘Ares’, ‘Athena’, ‘Bellezia’, ‘Dragon’s Fire’, ‘Eros’, ‘Nemesis’, ‘Poseidon’, ‘Red Streaked’, ‘Sylvetta’, ‘Wild’, ‘Wildfire’, ‘Zeus’), 11 Eruca sativa, (‘Astro’, ‘Balboa’, ‘Esmee’, ‘Green Brigade’, ‘Roquette’, ‘Runaway’, ‘Slow Bolt’, ‘Speedy’, ‘Standard’, ‘Surrey’, ‘Uber’), and 1 Diplotaxis erucoides (‘Wasabi’). Arugula was planted in 5ft plots at high density (3-5 seeds per inch) and in 36-inch-wide raised beds as recommended to us by farmers from NEAT’s PAC as industry standards. Plots are randomized throughout raised beds and each raised bed within the research field is considered a replicate block. Overhead irrigation was utilized, a 20-10-10 NPK nutrient treatment was applied, and no forms of pest control were applied. There were three experimental treatments in each evaluation: a natural plot, one plot inoculated with P. cannabina pv. alisalensis (bacterial blight), and another plot inoculated with X. campestris pv. campestris (black rot). Each treatment was made of 4 replicate blocks, with each cultivar randomly grown in 5ft plots within the blocks (26 plots/block). For the experimental plots, bacteria were grown up in nutrient broth and the plants were spray inoculated at maturity. After bacteria was inoculated, disease severity ratings were recorded 7, 14, and 21 days after inoculation. For both bacteria, disease severity was moderate to high among the E. sativa cultivars, and little to no disease was observed among the Diplotaxis cultivars. Flea beetle predation was moderate to high for all cultivars in the summer trial. The fall trial was planted in September and due to the colder temperatures, no flea beetles or bolting was observed among cultivars until the experiments ended in December. For the bacterial blight experiments, similar results were observed in the fall as to the summer trial, with moderate to high disease observed across the E. sativa cultivars, and low-to severe disease symptoms observed in the Diplotaxis cultivars. For the fall Xanthomonas field experiment, there was significantly less disease post-inoculation than was observed in the summer trial. Low-no disease symptoms were observed in the Diplotaxis plots, and only one plot of the E. sativa cultivars had moderate-severe disease symptoms. The low disease spread throughout the field could have been due to a cold front that moved in two days after inoculating the Xanthomonas plots. In 2025, we replicated our Rock Springs field germplasm evaluation experiments from 2024, planting both summer and fall research plots, with the addition of one new cultivar, ‘Coyote’. For summer 2025, in the black rot inoculation plot, moderate to high disease severity was observed for all cultivars. For the bacterial blight treatment, moderate to high disease severity was observed for all E. sativa cultivars. Disease severity was low for all Diplotaxis cultivars expect for ‘Dragon’s Fire’, where moderate severity was observed, comparable to the performance of ‘Esmee’. Flea beetle predation was moderate to high among all cultivars except ‘Dragon’s Fire’ & ‘Nemesis’, which had the lowest predation. In the fall experiments, there was no flea beetle predation observed in the natural plot. As well, there were no black rot symptoms observed after inoculation of X. campestris pv. campestris in the fall trial, likely due to cold temperatures. The P. cannabina pv. alisalensis treatment saw moderate-high disease severity in the E. sativa cultivars, as well as moderate severity in the ‘Eros’ and ‘Wildfire’ D. tenuifolia cultivars. The remaining Diplotaxis cultivars saw low-no disease severity.

On October 1st, 2024, experimental plots were planted to determine whether commonly grown commercial varieties of Eruca sativa arugula differ in their susceptibility to naturally occurring Pseudomonas spp. bacterial disease in a commercial high tunnel setting at Pleasant Valley Farm in Argyle, NY. The trials were planted under organic growing conditions, with the growers abiding by the Certified Naturally Grown audit requirements. P. cannabina pv. alisalensis bacterial blight in arugula was first confirmed on a high tunnel crop in January 2021 at the farm. Using a randomized complete block design, we planted five cultivars of arugula (‘Astro’, ‘Esmee’, ‘Standard’, ‘Surrey’, and ‘Uber’) in a 35’x144’ heated high tunnel. Each variety treatment was replicated five times in 2.5’ wide by 3’ plots. Each plot contained 12 rows of arugula seeded with a push seeder (Jang JP-1 with roller size X-24; Jang Automation Co. Ltd, Seoul, Korea). Soil fertility, irrigation, high tunnel climate control, row cover application, and other growing practices followed the host farm’s typical management for fall and spring arugula crops. Prior to planting for the experiment, the bed was fertilized at the following rates (lbs/ac) using a combination of NatureSafe (10-2-8; Darling Ingredients, Irving, TX), sulfur, aragonite, azomite, and granular boron fertilizer: 641 N, 128 P2O5, 514 K2O. 68 S, 74 Ca, 1.5 Mg, and 5 B. The arugula beds were irrigated using drip irrigation with three lines per bed. The arugula was covered with floating row cover at night and on cloudy days. The high tunnel was heated when temperatures underneath the row cover fell below 27F. No pesticides were applied to the arugula during either experiment. To determine whether the varieties differed in susceptibility to bacterial disease, we conducted two disease assessments on October 24^th and December 2^nd. During each assessment, we recorded overall disease incidence (percentage of plants displaying disease symptoms) in each plot and gave plots each an overall disease severity rating based on average symptom appearance and percentage of blighted leaf surface. On both dates, we also collected data on plant density. Plant density was measured by counting the number of arugula plants in a 12” section of row in each plot in random locations. Between 24 Oct and 13 Dec 2024, the arugula was harvested for farm sales. In the fall 2024 experiment, all arugula plots showed symptoms of bacterial disease, with some varieties developing symptoms more slowly than others. Disease incidence varied significantly by arugula variety at the time of observation in October prior to the first harvest. The variety ‘Uber’ displayed the greatest percentage of plants with disease symptoms, with over 55% of plants having leaves with gray or black lesions. ‘Surrey’ plots had significantly less diseased plants than ‘Uber,’ with other varieties not differing significantly. In our October data collection, we observed no statistically significant differences in disease severity of symptoms across varieties. In December, all plots had nearly 100% of plants showing some disease symptoms, and there were no significant differences in disease across varieties. In March 2025, a replicated arugula varietal susceptibility experiment was planted at Pleasant Valley Farm, using similar methods to our fall 2024 experiment. In 2025, however, soil fertility rates were reduced per spring practices commonly used on the farm, and plots were smaller (1.5 ft in length). On April 23 and May 8, we evaluated each plot for disease severity, incidence of disease (% of plants displaying disease symptoms), and counted the density of plants per row foot. Flea beetles were observed in the plots on both dates, and flea beetle damage was assessed. Disease symptoms were present in all plots by May. Esmee, Surrey, and Uber plots had nearly 100% of plants showing disease symptoms. Yield data were collected by the host farm. The growers abandoned the planting in early May after one harvest from some of the plots; no marketable arugula was harvested from Surrey and Uber plots due to small size and disease. Flea beetle damage did not differ by variety.

In fall 2025, two demonstration plantings of arugula were sown at the Cornell Willsboro Research Farm using the same seed lots from the Pleasant Valley Farm experiments. Five varieties (‘Standard’, ‘Astro’, ‘Esmee’, ‘Uber’, and ‘Surrey’) were seeded in 3 ft and 1 ft long plots that were 2.5 ft wide, 12 rows of arugula each. The first planting was seeded using a Jang JP-1 seeder with X-24 roller on Sept.18, and the second by hand on Sept. 24 due to uneven establishment in the first planting. Gaps in rows were reseeded approximately a week later in the first planting. The goal of this demonstration was to gather evidence for seedborne disease in the seed lots used at Pleasant Valley Farm, and the plots were seeded at high density (approx. 80 seeds per foot) to stimulate disease progression. Arugula was harvested twice in October and November when at ~5 cm in length. Few disease symptoms were observed, and samples revealed no P. cannabina pv. alisalensis, lending no support to the theory that the seeds are carrying this pathogen.

Greenhouse Germplasm Evaluations

In 2023 we established a protocol for our greenhouse evaluations. Arugula seeds were planted randomly among 6-pack planting inserts with a standard Osmocote fertilizer application. An experimental rep would consist of multiple 6-packs, with each cultivar of arugula represented once in each rep. Initial experiments used the original ten cultivars used in the fall 2023 field trials, and the other cultivars were added for later experiments. Planting was staggered for Eruca & Diplotaxis arugula so that all plants would be sprayed at the same time at their maturity date based on the seed packet information. For the bacterial blight evaluations, P. cannabina pv. alisalensis was grown on KMB agar and suspended in buffer to an OD of 600nm which is approximately 10⁸ CFU/ml. For the black rot evaluations, X. campestris pv. campestris was grown in nutrient broth, pelleted, and resuspended in buffer at approximately 10⁸ CFU/ml. The bacteria were sprayed onto the tops & undersides of arugula foliage, and the arugula was placed in high humidity for 48 hours to create optimal conditions for disease infection. Disease severity was recorded at 7, 14, & 21 days after inoculation. Additional bacterial blight trials were performed in spring-summer 2024, and black rot trials were performed in fall-winter 2024. In the bacterial blight trials, results were similarly mirrored to the summer and fall field experiments, although there was more variation of disease severity among the E. sativa cultivars. In the black rot trials, disease severity was moderate-high among all arugula cultivars. In 2025, we selected 25 lines from the USDA-ARS Germplasm Resources Information Network (GRIN) to evaluate based on previous performance data. In addition, we added our 4 best-performing E. sativa commercial germplasm from 2024, and the recently obtained ‘Coyote’ cultivar. A total of 30 germplasm evaluated for resistance to bacterial blight & black rot. Across the two black rot inoculation experiments, moderate-severe disease symptoms were observed in all cultivars. In the two bacterial blight experiments, low disease severity was observed in one GRIN line, with moderate-severe disease observed in all other lines. This one GRIN line along with the four other best performers were sent to our seed breeder collaborator to be bulk increased for replicated field trials.