Final report for ONE22-431
Project Information
Fire blight, caused by a bacterial pathogen Erwinia amylovora, is a devastating disease of apples and pears. Properly-timed application of the antibiotic streptomycin is by far the most effective and widely-used control of fire blight. The intensive use of streptomycin since the 1950’s has resulted in the development of streptomycin resistance in E. amylovora populations. Since first reported in 1972, streptomycin resistance of E. amylovora has been commonly detected in almost all apple-producing regions in the United States, including some Northeastern states such as New York. In this proposal, A region-wide survey of E. amylovora was conducted to determine streptomycin resistance status of E. amylovorapopulations in New England apple and pear orchards. 70 samples were collected from 27 farms from 6 states (CT, NH, MA, RI, VT, NY). E. amylovora was isolated from the infected plant tissue and tested on LB agar media supplemented with and without streptomycin sulfate. Our results suggest all samples tested were susceptible to streptomycin at the tested concentration (100 μg/ml). The survey results were released to growers at multiple grower meetings including the New England Fruit and Vegetable Conference (2024), Connecticut Pomological Society (2024), Maine Apple Grower Association Meeting (2023), New Hampshire Apple Grower Association Meeting (2025), as well as NE IPM center online conferences. We also promoted the proper use of antibiotics, including the use of a disease prediction model to time the antibiotic spray, as well as the use of biological controls as alternatives. A "2+2" non-antibiotic program has been presented to growers at the above meetings and is currently tested on 10 organic apple farms throughout New England. These activities will reduce the risk of inducing streptomycin resistance and enhance the sustainability of fruit production in this region.
This project seeks to understand the status of fire blight pathogen E. amylovora's susceptibility to the commonly used management materials, streptomycin. Two objectives proposed are:
Objective 1: Conduct a survey evaluating the streptomycin resistance of E. amylovora isolates in New England apple and pear orchards.
Objective 2: Disseminate the findings to the New England apple and pear growers and provide education about the causes and prevention of streptomycin resistance.
Anticipated impacts: Findings from these objectives will provide increased knowledge in tree fruit management area regarding the accurate use of antibiotic streptomycin to control fire blight. Streptomycin susceptibility in E. amylovora populations in New England will be obtained. This will provide necessary knowledge to the New England apple and pear growers about whether streptomycin is still an effective bactericide for fire blight management. In case streptomycin resistance is observed, the geographical distribution of streptomycin resistant E. amylovora populations in the New England region will be acquired. If resistant populations are at the emerging stage, eradication of orchards will help to restrict the spread of the resistant population to other parts of the region.
1.1 Fire blight is a devastating disease of apples and pears
Apples and pears represent economically important commodities in the United States and New England. According to recent U.S. census data, apples and pears are grown on 142,000 and 24,300 hectares nationally, with cash receipt values of $2.6 billion and $372 million, respectively (USDA-NASS 2014) . In New England (CT, MN, MA, NH, RI and VT), cash receipt values of apple and pears exceed $54 million and $6.1 million, respectively (USDA-ERS).
Fire blight, caused by a bacterial pathogen Erwinia amylovora, is one of the most serious diseases currently limiting apple and pear production in New England and in the United States (Malnoy 2012; Zeng 2013; Zeng 2014). Fire blight can result in both yield reduction, when infection occurs on the scion, and tree death, when infection occurs in the rootstock. Annual losses to fire blight and costs of control in the United States are estimated at over $100 million (Norelli 2003). In recent years, severe fire blight outbreaks occurred throughout the New England region.
1.2 Timely application of antibiotic streptomycin is an indispensable component of fire blight IPM
Most apple and pear varieties sought after by consumers, such as “Gala”, “Fuji”, and “Barlett”, are either susceptible or highly-susceptible to fire blight. Although multiple fire blight resistant rootstocks such as “Geneva” and “Vineland” have been developed, grafting the susceptible cultivars to the resistant rootstocks only prevent rootstock infections but does not prevent scion infections. In practice, the fire blight susceptible scions are often grafted to fire blight susceptible dwarfing root stocks for the new industry standard, high-density orchard systems, making these orchards highly vulnerable to fire blight.
The lack of effective cultural control makes chemical control an indispensable component of fire blight IPM. The most critical step of fire blight management is to limit the multiplication of E. amylovora on stigmas of apple flowers to prevent blossom blight (Malnoy 2012). With appropriate disease forecasting models (e.g. Maryblyt or CougarBlight), properly timed application of bactericide during bloom can effectively reduce bacterial number in flowers and control blossom blight. Streptomycin is by far the most effective (up to 90% control) and widely-used bactericide for fire blight control in the United States . Other control alternatives include the biological control agents Serenade MAX , BlightBan A506, and the systemic acquired resistance (SAR) inducers such as Actigard. However, these control alternatives have not provided consistently high levels of control compared to streptomycin (Sundin 2009).
1.3 Streptomycin resistance raises critical challenges to fire blight IPM
The intensive, long-term use of streptomycin led to the evolution of streptomycin resistance in E. amylovora populations. Since the first report in California in 1971 (Miller 1972), streptomycin resistance in E. amylovora has been commonly detected in almost all apple producing regions in the United States, such as Washington (1972), Michigan (1990), and has recently been detected in New York (2003) (Coyier 1975; Chiou and Jones 1993; Russo 2008). Currently, the New England is the only region in the United States where streptomycin resistance in E. amylovora has not been reported.
The impact of streptomycin resistance is dramatic, both from the economic loss point of view and the environmental impact and human health point of view. First, because streptomycin is no longer effective in regions with streptomycin resistant E. amylovora populations, thousands of acres of valuable apple and pear crops are exposed to fire blight, which has caused catastrophic losses. In 2000, the fire blight epidemic in southwest Michigan resulted in the death of 400,000 apple trees, and over $42 million in losses. In addition, the loss of streptomycin led the growers to adopt other antibiotics to control fire blight. Oxytetracycline and kasugamycin have been used in fire blight control in California and Michigan due to streptomycin resistance. Introduction of these antibiotics into agriculture raises concerns of facilitating the development of antibiotic resistance in the environment, with a potential impact to human health.
Chiou, C. S. and A. L. Jones 1995. Expression and identification of the strA-strB gene pair from streptomycin-resistant Erwinia amylovora. Gene 152: 47-51.
Coyier, D. L., and Covey, R. P. 1975. Tolerance of Erwinia amylovora to streptomycin sulfate in Oregon and Washington. Plant disease reporter 59: 849-852.
Norelli, J.L., Jones, A. and H. Aldwinckle. 2003. Fire blight management in the twenty-first century: using new technologies that enhance host resistance in apple. Plant disease 87: 756-765.
Malnoy, M., and S. Martens 2012. Fire blight: applied genomic insights of the pathogen and host." Annual review of phytopathol. 50: 475-494.
Russo, N. L., Burr, T. J., Breth, D. I.,and H. S. Aldwinckle. 2008. Isolation of streptomycin-resistant isolates of Erwinia amylovora in New York. Plant disease 92: 714-718.
Sundin, G. W. W., Nicole A.; Yoder, Keith S.; H.S. Aldwinckle. 2009. Field evaluation of biological control of fire blight in the eastern United States. Plant disease 93: 386-394.
Zeng, Q., R. R. McNally, and G. Sundin. 2013. Global small RNA chaperone Hfq and regulatory small RNAs are important virulence regulators in Erwinia amylovora. Journal of bacteriol, 195: 1706-1717.
Zeng, Q. and G. W. Sundin. 2014. Genome-wide identification of Hfq-regulated small RNAs in the fire blight pathogen Erwinia amylovora discovered small RNAs with virulence regulatory function. BMC genomics 15: 414.
Cooperators
- - Producer
Research
Objective 1. Conduct a survey evaluating the streptomycin resistance of E. amylovora isolates in New England apple and pear orchards.
1.1 Collect fire blight tissues from New England orchards and isolate E. amylovora
Freshly infected apple and pear tissues with fire blight were collected from New England orchards in 2023 and 2024. The sample collection was through a collaborative effort of extension agents in the area, including but not limited to, Mike Basedow, Anna Wallingford, Win Cowgill, Kristy Grigg-McGuffin, Dan Cooley, and Jeremy Delisle. Fresh shoot and fruitlet samples with ooze droplets were collected and put in a plastic bag with a damp paper towel and shipped to Connecticut Agricultural Experiment Station using a prepaid envelope. Special attention was given to orchards that have used streptomycin in the past, observed inconsistent control of fire blight using streptomycin, or had severe fire blight symptoms in the past years. Once the samples were received, researchers from CAES performed pathogen isolation and resistance testing. Diseased samples were surface sterilized followed by removal of the outer bark. Bacteria from infected internal plant tissue / ooze droplets will be resuspended in sterile water and plated on an E. amylovora differential medium CCT. Pale violet, mucoid E. amylovora colonies were selected and preserved as putative E. amylovora strains. The identity of E. amylovora was confirmed by polymerase chain reaction (PCR) using E. amylovora diagnostic primers that amplify a region from the ubiquitous E. amylovora plasmid pEA29 (Bereswill, Pahl et al. 1992).
1.2 Evaluate the streptomycin resistance of E. amylovora isolates
The confirmed E. amylovora isolates were tested for streptomycin resistance using the Kirby-Bauer antibiotic diffusion method. E. amylovora isolates were cultured in Luria-Bertani (LB) broth at 28°C overnight. 100µl of 108 cells/ml bacterial suspension will be evenly spread onto LB plates. The growth of bacteria were challenged by placing filter paper disks impregnated with streptomycin sulfate solutions at 0, 10, 50, or 100 μg/ml. A streptomycin resistant strain and a streptomycin susceptible strain were included as positive and negative controls. We expect that zone of inhibition would be observed on plates with streptomycin susceptible strains but not with streptomycin resistant strains. The levels of resistance would be determined by measuring the sizes of inhibition zones at 24 and 48 hours post incubation.
2022 PROGRESS
A list of collaborators have been identified, and sampling methods and collection bags have been delivered accordingly. Sampling of streptomycin resistance in E. amylovora will start in May 2023, when the infected plant materials are available.
2023 PROGRESS
In 2023, we tested 48 samples collected from 13 farms in New England states. This includes 7 farms in New Hampshire, 5 farms in Connecticut, and 2 farms from Massachusetts. Most of the samples were mailed by growers with prepaid Fedex services provided by the grant. Erwinia amylovora strains have been isolated from the infected tissue, mostly bacterial ooze droplets, or infected immature fruits and occasionally shoots. The isolates were streaked on LB agar plates supplemented with streptomycin at 100ug/ml. All E. amylovora isolates were tested negative in resisting streptomycin. Results are provided to growers / extension agents typically within 10 days.
2024 PROGRESS
In 2024, we tested 22 samples collected from 14 additional farms in New England states. This includes 3 farms in New Hampshire, 9 farms in Connecticut, and 1 farm from Massachusetts, and 1 farm from New York. Most of the samples were mailed by growers with prepaid Fedex services provided by the grant. Erwinia amylovora strains have been isolated from the infected tissue, mostly bacterial ooze droplets, or infected immature fruits and occasionally shoots. The isolates were streaked on LB agar plates supplemented with streptomycin at 100ug/ml. All E. amylovora isolates were tested negative in resisting streptomycin. Results are provided to growers / extension agents typically within 10 days.
There is currently no E. amylovora that can resist streptomycin at 100ug/ml at the tested locations in New England. Measures to prevent streptomycin resistance include restricting transporting plant cuttings, nursery materials from regions where streptomycin resistant E. amylovora is endemic. In tandem with this project, we developed and tested a "2+2" program which two Blossom Protect applications are conducted at 40-80% bloom and 2 applications of Oxidate 5.0 are conducted at full bloom and petal fall, which showed great control effect against fire blight and may help reduce the risk of streptomycin resistance.
Education & Outreach Activities and Participation Summary
Participation Summary:
The survey results of sample collection location and stretpomycin succeprtibility of E. amylovora isolates were disseminated to growers through extension presentations at regional grower meetings such as the CT Pomological Society Meeting (2024), Maine Apple Grower Association meeting (2024) at the Northeastern Tree Fruit Management Working Group meetings (2023 and 2024), the New England Vegetable and Fruit Conference (2024), Northeastern IPM center webinars (2022 and 2023) and one-on-one consulting with growers will be used to further reach fruit growers in New England.
Management of fire blight using the “2+2” non-antibiotic Program Factsheet
Actigard Paint Therapy to Manage Fire Blight Canker Fact Sheet
Learning Outcomes
- Is streptomycin still effective to control fire blight on their farm
- how to use disease prediction modeles to time their antibiotic sprays
- what alternative options of antibiotics can they adopt
Project Outcomes
The outcomes of this project include
- informed choice of disease control materials. Since streptomycin resistance has not been found in New England, it means it is safe to continue to use streptomycin to control fire blight in our region.
- raised awareness of antibiotic resistance. Growers are more aware of the risk of antibiotic resistance in fruit production, risky measures of summer sprays of antibiotics are avoided.
- raised awareness of other antibiotic alternatives. More growers are willing to adopt or test the "2+2" progrm using Blossom Protect yeast and Oxidate. In 2024, 9 farms signed up for a USDA funded control trials of the "2+2" program including growers in MA, CT, NH, NY, and RI.
A prepaid envelope format of sampling by growers and direct shipment to the testing lab (CAES) may save time for extension agents and reduce the turnaround time for the testing results. The monitoring of the streptomycin resistance should be conducted in this region on a routine basis. Some strepotmycin resistant E. amylovora have been recently described in the Hudson Valley area, which suggests they may enter New England in the near future. Benefited audience of this project is fruit growers (apple, pear) in the Northeast.