Progress 10/01/16 to 09/30/21
Outputs
Target Audience:The target audiences for this project have been 1) government, land managers, and academic groups combating a beetle-vectored fungal disease called Rapid Ohia Death that decimates native Hawaiian forests, 2) those involved in public outreach about this dilemma, 3) forestry and academic groups concerned with invasive bark beetles and the fungal diseases they vector, and 4) academic groups studying insect genomics, how insects select plant hosts, and how invasive insects differ from native ones. Changes/Problems:As our project progressed, we broadened our initial approach to include a broader genomic angle to both provide a resource for those developing methods to control pest bark beetles, and to understand what genes enable pest bark beetles to attack plants. This was a natural extension of our initial plan to explore the metagenomics of the microbial community within each species, because a shotgun whole-genome sequencing method would reveal both the microbiomes and the beetles' host-selection genes. What opportunities for training and professional development has the project provided?The first year of this grant funded a postdoctoral fellow, Curtis Ewing, and a technician, Kylle Roy, who both obtained training in entomology and forest pathology. Roy continues to collaborate on Rapid Ohia Projects in her role at USGS-PIERC. Subsequently, the grant has funded a PhD candidate at the University of Hawaii-Manoa, Jared Bernard, who is being trained in entomology, curating data, molecular biology analysis, and project management. We are grateful to have recently started an extension of our McIntire-Stennis funds for a further two years. How have the results been disseminated to communities of interest?Aspects and results of this work have been variously presented to legislators, land managers, the public, and academic interest groups at the following events: Nov 2016: Presentation by Gordon Bennett at the Rapid Ohia Death Summit convened by Gov. David Ige. Feb 2017: Researchers met with homeowners in Kaloko near Kailua-Kona, Hawaii. Mar 2017: Curtis Ewing presented "How does the fungus spread?" at the Rapid Ohia Death symposium at the University of Hawaii-Hilo, repeated Apr. 2017 at West Hawaii Civic Center in Kailua-Kona, Hawaii. Apr 2017: Curtis Ewing and Kylle Roy presented "Infectivity of boring dust produced by ambrosia beetles from Rapid Ohia Death positive ohia trees" at the University of Hawaii-Hilo. May 2017: Rapid Ohia Death training for Hawaii County field workers, coordination with Dept. of Public Works. Jul 2017: Kylle Roy presented "Monitoring assessment of current ROD management strategies on Hawaii Island" at the Hawaii Conservation Conference in Honolulu, Hawaii. Sep 2017: Jared Bernard presented "Learning about beetles to save Hawaii's native forests from Rapid Ohia Death" at a mini-symposium for Rep. Colleen Hanabusa (D.-HI). Sep 2017: Jared Bernard presented "Sampling methods on Kauai" at the Farm Bill 10007 Coffee Berry Borer Biological Control and IPM Planning Workshop in Kalaheo, Hawaii. Oct 2018: Kylle Roy and Curtis Ewing, alongside co-authors at USDA-PBARC, finalized the peer-review process for their paper entitled "Presence and viability of Ceratocystis lukuohia in ambrosia beetle frass from Rapid Ohia Death-affected Metrosideros polymorpha trees on Hawaii Island," which was published in the journal Forest Pathology. Nov 2018: Jared Bernard presented "Community dynamics of bark beetles on Kauai" for an Ecology, Evolution and Conservation Biology meeting at the University of Hawaii-Manoa. This presentation contrasted the seasonality and population influences of the dozens of invasive bark beetles in Hawaii, including pests of forestry and agriculture. We are still in the process of finalizing the genomes, so we have not yet disseminated this information. However, we plan to make the genomes publicly available and publish our findings on their comparative genomics as soon as possible. We plan to also share this resource with groups investigating control methods as soon it is ready. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
This project has used multiple pathways of inquiry to understand what makes invasive bark beetles successful and thus provide information to aid control methods, with special emphasis on those that transmit the fungal disease Rapid Ohia Death that kills native ohia trees, a cornerstone species in Hawaiian forests. Determine the current diversity of native and adventive Hawaiian scolytine species: We collected bark beetles in numerous ways to determine the distribution of invasive and native species, and which attack ohia trees. First, using a variety of traps to determine which species are in forests on different Hawaiian Islands. We tested 8 trap lures, finding that methanol-ethanol was most effective, as we reported to the US Forest Service, so we used this bait during our surveys of forests across the state. Next, we developed boring-dust traps to collect bark beetles directly from live trees, as described in the "Instruments or Equipment" section. Finally, we reared specimens from bolts of wood from various native Hawaiian trees that include ohia. Ultimately, we found 5 species to be associated with ohia: Xyleborinus saxesenii, Xyleborus ferrugineus, Xyleborus affinis, Xyleborus perforans, and Xyleborus simillimus. All are invasive except for X. simillimus, a native Hawaiian species whose only known host is ohia, but we seldomly collected it. Xyleborus ferrugineus was mostly in bolts we collected at elevations less than 520 meters, and X. affinis was in bolts from over 600 meters. From all our surveys across the state, we collected 27 species of invasive bark beetles and 20 species of native bark beetles during our surveys across the archipelago. Circumscribe fungal and bacterial symbiont diversity for each species and geographic localities: Following the publication in the next section (Roy et al. 2019), we are currently investigating how microbiomes enables bark beetles to use their hosts. As described below in the section labeled "Host-Selection of Bark Beetles," this part of the project has required organizing a large amount of genomic data, which will be analyzed in a new 2-year extension of the funding we have acquired from USDA-NIFA. Determine the prevalence of plant pathogenic fungi among invasive and introduced scolytine species: We detected genetic markers for the fungal pathogen responsible for Rapid Ohia Death, known as Ceratocystis lukuohia, in both beetles and their boring-dust using a molecular technique known as quantitative PCR. The results showed that 62% of the frass/dust contained Ceratocystis, most of which was produced by the invasive bark beetle Xyleborus ferrugineus. The beetles themselves, however, were mostly negative for the fungal pathogen, although we have so far tested few beetles. To assess the viability of the fungal disease, we inoculated growth media in a lab with fungal spores. By this process, which involved culturing on slices of carrots, we found that a quarter of our positive dust samples produced viable cultures. We collaborated with the plant pathology lab at USDA's PBARC in Hilo, Hawaii for this work, which was published in Forest Pathology in 2019 (doi: 10.1111/efp.12476). USGS-PIERC in Hilo, Hawaii is continuing this phase of the project. Aerial sampling of fungal pathogen: To determine whether Ceratocystis fungal spores that cause Rapid Ohia Death move through the air in native forests, we used Rotorod devices to collect dust samples and tested them for the presence of fungal spores via quantitative PCR. After initially failing to detect the spores in an outbreak area, we tested whether cutting down trees infected with Ceratocystis spores exuded aerosolized spores. Here the Rotorods detected the fungal spores downwind from tree cutting. Finally, we set up 4 Rotorods for 8 weeks at a Rapid Ohia Death outbreak in eastern Hawaii Island, finding 10% of samples to be positive for the spores. Genetic variation of invasive bark beetles: To gain a preliminary idea of the genetic variation of invasive bark beetles in Hawaii, we extracted genetic markers from 48 specimens. These represented 7 species, 3 of which we had previously identified as being associated with Rapid Ohia Death: Xyleborinus saxesenii, Xyleborus ferrugineus, and Xyleborus affinis. The results showed very low genetic variation among most populations, for example 0.05% among X. affinis individuals. This is likely the result of the semi-clonal life history of bark beetles. However, the highest variation was among X. ferrugineus individuals at 14%. We speculate this is the result of multiple founder events for this species, but further work would be needed to confirm these findings. Reference genomes of bark beetles: To aid in the development of control methods against invasive bark beetles that transmit Rapid Ohia Death, we developed 2 genomes. One is for Xyleborus affinis, an invasive species we previously found to be attacking ohia trees and a putative vector of Rapid Ohia Death. This species uses at least 240 plants as hosts. A reference genome for this species will be crucial for identifying genes that enable it to attack these species, and such genes can also be targeted by molecular control methods. To avoid targeting native Hawaiian bark beetles, however, we need to know how they differ from X. affinis. We therefore also developed a genome of a native Hawaiian species, X. molokaiensis, which specializes on just 1 or 2 native Hawaiian plants. We collected live specimens of both species on Oahu, which we flash froze and extracted high quality genetic material. We sequenced the genomes of each, and also the totality of their gene-coding transcripts (known as a transcriptome) to use as a guide to annotate the genomes. We are still finalizing our analysis of these genomes before publication, but our initial results show an interesting contrast between the 2 species. Xyleborus affinis appears to have a genome that is roughly 36% smaller than that of the native beetle, but the native beetle has over 3 times as many noncoding regions. After initial data cleaning, we detected more than 34,000 genes for X. affinis and over 28,000 genes for X. molokaiensis. More importantly, we found that X. affinis has almost 4 times as many odor-sensing genes and over 2.6 times as many toxin metabolizers than X. molokaiensis. The genome of the invasive species, especially when compared to the native species, will be a valuable resource for academic and government groups that develop control methods for beetle vectors of Rapid Ohia Death. Host-selection of bark beetles: The final phase of our project broadens the genomic comparisons between invasive and native bark beetles, specifically the genes involved in host selection. Additionally, this work will assess whether microbiomes are correlated with host selection. This work will be significant to academic and government groups developing control methods for invasive bark beetles that are pests of forestry and crops. This phase of the project relies on the 2 genomes as references, and uses 137 specimens from our archipelago-wide collections, encompassing 16 invasive species and 20 native species. During the final reporting year of this grant, we organized the material needed for this work. This included trials of 15 DNA extraction techniques. We then prepared the samples for shotgun genomic sequencing, which we did in collaboration with Scott Geib's lab at USDA-ARS. USDA granted us a 2-year extension of funding, during which time we will complete the sequencing of the 137 specimens and analyze their host-selecting genes and microbiomes for insights into what makes the invaders successful and problematic.
Publications
- Type:
Other
Status:
Other
Year Published:
2016
Citation:
Presentation by Gordon Bennett at the Rapid Ohia Death Summit convened by Gov. David Ige
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2017
Citation:
Curtis Ewing presented How does the fungus spread? at the Rapid Ohia Death symposium at the University of HawaiiHilo
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2017
Citation:
Curtis Ewing and Kylle Roy presented Infectivity of boring dust produced by ambrosia beetles from Rapid Ohia Death positive ohia trees at the University of HawaiiHilo
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2017
Citation:
Kylle Roy presented Monitoring assessment of current ROD management strategies on Hawaii Island at the Hawaii Conservation Conference in Honolulu, Hawaii
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2017
Citation:
Jared Bernard presented Sampling methods on Kauai at the Farm Bill 10007 Coffee Berry Borer Biological Control and IPM Planning Workshop in Kalaheo, Hawaii
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2018
Citation:
Jared Bernard presented Community dynamics of bark beetles on Kauai for an Ecology, Evolution and Conservation Biology meeting at the University of HawaiiManoa
- Type:
Journal Articles
Status:
Other
Year Published:
2022
Citation:
Publication on invasive bark beetle genomics still in progress, expected to be submitted by summer 2022.
- Type:
Journal Articles
Status:
Published
Year Published:
2019
Citation:
Roy, K., Ewing, C.P., Hughes, M.A., Keith, L., and Bennett, G.M. (2019). Presence and viability of Ceratocystis lukuohia in ambrosia beetle frass from Rapid Ohia Death-affected Metrosideros polymorpha trees on Hawaii Island. Forest Pathology 49: e12476. doi: 10.1111/efp.12476
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2017
Citation:
Jared Bernard presented Learning about beetles to save Hawaiis native forests from Rapid Ohia Death at a mini-symposium for Rep. Colleen Hanabusa (D.-HI)
- Type:
Journal Articles
Status:
Published
Year Published:
2019
Citation:
Gillett, C.P.D.T., Honsberger, D., and Rubinoff, D. (2019) Rediscovery of the Hawaiian endemic bark beetle Xyleborus pleiades Samuleson, 1981 on Molokai, with records of three new exotic bark beetles for the island (Coleoptera: Curculionidae: Scolytinae: Xyleborini). Journal of Natural History, 53, 14811490. doi:10.1080/00222933.2019.1657976
- Type:
Journal Articles
Status:
Published
Year Published:
2018
Citation:
Gillett, C.P.D.T., Pulakkatu-Thodi, I., and Rubinoff, D. (2018) Rediscovery of an enigmatic bark beetle endemic to the Hawaiian Islands (Coleoptera: Curculionidae: Scolytinae). The Coleopterists Bulletin, 72, 811815. doi:10.1649/0010-065X-72.4.811
- Type:
Journal Articles
Status:
Published
Year Published:
2018
Citation:
Bernard, J., Ewing, C.P., and Messing, R.H. (2018) The structure and phenology of non-native scolytine beetle communities in coffee plantations on Kauai. Insects 9: 123. doi: 10.3390/insects9040123
|
Progress 10/01/19 to 09/30/20
Outputs
Target Audience:There are two audiences that will be interested in our efforts during this reporting period. The primary audience is academics who study the biology of bark beetles, particularly how they associate with host plant species, including forests and crops. Another audience that will find this work pertinent is regional research communities who are actively confronting invasive bark beetles associated with fungal pathogens that eliminate native forests, not only in Hawaii, but across the country. Changes/Problems:The progress of the project stalled for several months because collaborating labs had reduced operations due to the COVID-19 pandemic. Although this has caused significant delays, it did not result in a change to the project. As these labs become functional again, we are optimistic about resuming our progress. What opportunities for training and professional development has the project provided?A PhD candidate at UH-Manoa, Jared Bernard, completed the third year of support from McIntire-Stennis funds during this fiscal year. These resources will allow him to continue researching the genetic basis for host selection and utilization by bark beetles, including those that are important pests of forests. How have the results been disseminated to communities of interest?The progress of the project was not anticipated to yield deliverables during the 2019-2020 reporting period. The research is ongoing and we hope to be able to disseminate more information to interested parties next year, pending results. What do you plan to do during the next reporting period to accomplish the goals?The resources provided by the McIntire-Stennis will support the ongoing research into host specificity of bark beetles being pursued by a PhD candidate, Jared Bernard. The project will provide insights about how bark beetles are able to adapt to new hosts, and why some are able to utilize hundreds of hosts while others are highly specific. As the genomes described above are drafted, and the shotgun sequencing project moves into its next stage, we will also plan to explore how the microbial community within each beetle species dictates its host usage. All of these avenues of investigation will have significance for understanding forest pests.
Impacts
What was accomplished under these goals?
This reporting period included 2 primary goals outlined below. Both relate to continuing our investigation into the genetic and microbial basis for how bark beetles use host plants, including those impacting native Hawaiian ohia trees (a dominant tree in Hawaiian forests, vital to many endangered species). This work involves contrasting the native and invasive bark beetle species. 1. ASSEMBLING REFERENCE GENOMES OF BARK BEETLES During the last reporting period, we initiated a genetic project to study the basis for host specificity of both native and non-native bark beetles. Not only will this be a significant addition to the scientific community interested in how insects associate with hosts, but this will be informative for ongoing programs by state and federal agencies that combat the Ceratocystis fungal pathogen impacting ohia trees. We are analyzing the genomes of a native bark beetle and an invasive bark beetle. The latter is Xyleborus affinis, which recent research indicates as a putative vector of this Ceratocystis pathogen. As stated in the previous interim report, beetles were collected, flash frozen, and sent to the Genomic Center at the University of California-Davis for both whole genome and transcriptome sequencing. During this reporting period, we assembled the genome of each species. Our preliminary results suggest that X. affinis has a genome that is ~36% smaller than that of the native beetle, but the native beetle has over 3 times as many noncoding regions. We also assembled the transcriptome for each species, which will enable us to begin the next part of this project: annotating the genome so we can understand which areas are associated with biological aspects of interest. 2. USING REFERENCE GENOMES TO EXPLORE BROADER TRENDS Last year, we reported that we were planning a broader genetic project that would be based on the genomes. This will involve about 134 specimens tabulated in the database mentioned above, which includes around 20 native species and 16 invasive species. This year, methods for extracting the DNA from these specimens has been undertaken, and we are now starting to prepare the DNA for shotgun sequencing. This is an approach that will permit us to scan across each beetle's genome for areas of interest, using the reference genomes being constructed as a guide. The goal is to determine whether genes that enable beetles to find host plants are more numerous in the bark beetle species that have wider host ranges. This dataset includes X. affinis mentioned above. The results of this investigation will provide valuable information about how such pests are able to attack so many forestry and crop species.
Publications
|
Progress 10/01/18 to 09/30/19
Outputs
Target Audience:There are two audiences during this reporting period falling into two groups. The first includes local and national research communities dealing with fungal pathogens associated with bark beetles that affect native trees and forest health. The second audience is academic researchers interested in the biology of bark beetles, including pest species that impact native forests and crops, and native bark beetle species. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?A PhD graduate student at UH-Manoa, Jared Bernard, completed the second year of support from McIntire-Stennis funds during this fiscal year. These resources will allow him to continue researching the genetic basis for host selection and utilization by invasive bark beetles in Hawaii, including those implicated as vectors of Rapid Ohia Death as well as other bark beetles that are important pests of forests. How have the results been disseminated to communities of interest?The ongoing results of the above projects were presented to the public, research community and land managers, as summarized below: At the start of the reporting period (October 2018), Kylle Roy and Curtis Ewing, alongside co-authors at USDA-PBARC, published a paper entitled "Presence and Viability of Ceratocystis lukuohia in Ambrosia Beetle Frass from Rapid Ohia Death-Affected Metrosideros polymorpha Trees on Hawaii Island" in the journal Forest Pathology. In November 2018, Jared Bernard gave a presentation entitled "Community Dynamics of Bark Beetles on Kauai" for an Ecology, Evolution and Conservation Biology meeting at the University of Hawaii-Manoa. This presentation contrasted the seasonality and population influences of the dozens of invasive bark beetles in Hawaii, including pests of forestry and agriculture. What do you plan to do during the next reporting period to accomplish the goals?The funds will continue to support an investigation into the basis of host utilization by non-native and native bark beetles in Hawaii, which is being done by a PhD graduate student, Jared Bernard. This work will be instrumental in understanding how pest bark beetles are able to attack certain plants, and how bark beetles can shift hosts. This research will include understanding what roles genes and microbial symbionts play in host selection and utilization, which can have important implications for forest pests in the Hawaiian Islands.
Impacts
What was accomplished under these goals?
For this reporting period, our goals were to (1) contribute research into the mode of transmission of the fungal pathogen killing the native Hawaiian ohia trees (vital to native Hawaiian forests as cornerstone species), and (2) begin to understand the basis of host utilization for pest bark beetles as compared with native bark beetles in Hawaii. Other funding sources are aiding our ability to pursue these projects. 1. PRESENCE OF FUNGAL PATHOGEN IN BEETLE BORING-DUST AND BEETLES We published a paper in October 2018 in the journal Forest Pathology, which we were finalizing during the previous fiscal year. By collecting boring dust/frass from 4 sites in native forests, the results revealed that dust/frass is positive for the fungal pathogen Ceratocystis lukuohia (i.e. Rapid Ohia Death) and was associated with the invasive bark beetle Xyleborus ferrugineus. Nevertheless, the beetles themselves were mostly negative for the fungal pathogen. Continued work on this aspect of the project is being carried out by USGS-PIERC in Hilo, Hawaii. 2. HOST-SPECIFICITY OF BARK BEETLES As a follow-up to the previously mentioned publication, we are organizing protocols to launch a project to determine how the microbiome enables bark beetles to use their hosts. As these bark beetles have a microbial symbiont in the form of ambrosia fungus, including Ceratocystis, this could have important implications for forest pathogens. Furthermore, we are initiating a genetic project to understand the genetic basis for host utilization by non-native bark beetles, compared to native species. We collected live invasive and native bark beetles in the forests of Oahu for the initial phase of this project, which involved obtaining reference genomes for two bark beetle species: one native and one invasive for comparison of genetic markers. Beetles were identified and flash frozen in liquid nitrogen before sending to the Genomic Center at the University of California-Davis. After high quality extraction and preparation, the genomes were sequenced. Assembly of these genomes will occur during the 2019-2020 fiscal year. To aid in the annotation and analysis of the genomes, the transcriptomes will also be sequenced during the next fiscal reporting year. We are currently developing the protocols to begin a larger genetic project involving over 100 specimens, which will use the aforementioned reference genomes to enable us to compare the genetics of pest and native bark beetles. This is an essential step in evaluating the pathogenicity and vectoring role of both native and invasive bark beetles.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2018
Citation:
Roy, K., Ewing, C.P., Hughes, M.A., Keith, L., and Bennett, G.M. 2018. Presence and viability of Ceratocystis lukuohia in ambrosia beetle frass from Rapid Ohia Death-affected Metrosideros polymorpha trees on Hawaii Island. Forest Pathology 2018: e12476.
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Progress 10/01/17 to 09/30/18
Outputs
Target Audience:Target audiences during this reporting period fall into two groups. First, the largest audience includes state, federal, and academic groups confronting a serious native tree pathogen that causes the forest tree disease known as Rapid Ohia Death. In this case, outreach has focused on educating these groups on the latest research concerning the potential of bark beetles to act as vectors of the fungal pathogen. This has included communicating with legislators. The second audience includes academic researchers interested in the biology of bark beetles, including non-native species that impact native forests and crops.Target audiences during this reporting period fall into two groups. First, the largest audience includes state, federal, and academic groups confronting a serious native tree pathogen that causes the forest tree disease known as Rapid Ohia Death. In this case, outreach has focused on educating these groups on the latest research concerning the potential of bark beetles to act as vectors of the fungal pathogen. This has included communicating with legislators. The second audience includes academic researchers interested in the biology of bark beetles, including non-native species that impact native forests and crops. Changes/Problems:The new PI, Dr. Dan Rubinoff, has assumed control of the project and funds in the middle of this reporting period, after Gordon Bennett relocated to University of California-Merced.The funds are now used to support an investigation into the genetic variation of non-native bark beetles in Hawaii, which is being done by a PhD graduate student, Jared Bernard. The work will trace the origins of bark beetle invasions in Hawaii, and seek evidence of multiple introductions. Additional changes to the project stem from the departure of the postdoc, Curtis Ewing, who worked on infected forests with state and federal partners; he started a new position in California in the middle of this reporting period. His assistant, Kylle Roy, also started a new position at USGS in the middle of this reporting period, although her work with Rapid Ohia Death projects will continue. What opportunities for training and professional development has the project provided?In the early months of this reporting period, a postdoctoral researcher and his assistant finalized their work completed during the first year of funding. The postdoc, Curtis Ewing, now works for the State of California as an entomologist, and the assistant, Kylle Roy, is continuing to collaborate on the Rapid Ohia Death projects listed above with her new role at USGS-PIERC in Hilo, Hawaii. A PhD graduate student at UH-Manoa, Jared Bernard, is now supported by the McIntire-Stennis funds, which will enable him to continue researching the genetic variation and tracing the invasions of bark beetles into Hawaii, including those implicated as vectors of Rapid Ohia Death as well as other bark beetles that are important pests of croplands and forests. How have the results been disseminated to communities of interest?The ongoing results of the above projects and associated outreach material Work under this project was presented to the public, state legislators, and land managers through presentations to each of these groups. Below are summary of these events: At the start of the reporting period, Jared Bernard gave a presentation at a mini-symposium for Rep. Colleen Hanabusa (D.-HI) entitled "Learning About Beetles to Save Hawaii's Native Forests from Rapid Ohia Death" at UH-Manoa. At the Hawaii Conservation Conference on July 25, 2018, Kylle Roy gave a presentation entitled "Monitoring Assessment of Current ROD Management Strategies on Hawaii Island" as part of the Rapid Ohia Death Status symposium in Honolulu, Hawaii. Kylle Roy and Curtis Ewing, alongside co-authors at USDA-PBARC, finalized the peer-review process for their paper entitled "Presence and Viability of Ceratocystis lukuohia in Ambrosia Beetle Frass from Rapid Ohia Death-Affected Metrosideros polymorpha Trees on Hawaii Island" that will be published in the journal Forest Pathology in October 2018. Curtis Ewing and Kylle Roy continued to work with other state and federal agencies to deliver the findings from these projects, particularly to the Rapid Ohia Death Working Group consortium. What do you plan to do during the next reporting period to accomplish the goals?PhD graduate student Jared Bernard will continue collecting bark beetle species that inhabit native forests and croplands across the archipelago to determine their genetic structure. From this we will address from where bark beetle invasions arrived in the rest of the world, and importantly whether there have been multiple introductions. By analyzing these trends in bark beetle invasions alongside detecting the fungal symbionts of the beetles, we intend to study whether bark beetles recruit new fungal symbionts in their invaded ranges. This could have important implications for the Rapid Ohia Death crisis, as well as other bark beetle pests in the Hawaiian Islands.
Impacts
What was accomplished under these goals?
For this reporting period, our goals were to (1) contribute research into the mode of transmission of the fungal pathogen killing the native Hawaiian ohia trees (vital to native Hawaiian forests as cornerstone species), and (2) begin to understand the genetic variation and population structure of non-native bark beetles in Hawaii. These projects were addressed with the aid of other funding sources. 1. PRESENCE OF FUNGAL PATHOGEN IN BEETLE BORING-DUST AND BEETLES We finalized our analysis of the presence of the fungal pathogen in beetle boring dust/frass, which was started in the previous reporting period. Samples came from 200 boring dust traps we established across 4 sites in native forests on Hawaii Island. We screened these using a molecular lab technique called quantitative PCR, which could detect genetic markers specific to the fungal pathogen. The results showed that 62% of the frass/dust was positive for the Ceratocystis lukuohia fungal pathogen, most of which was produced by the non-native bark beetle Xyleborus ferrugineus. This analysis, which was completed in conjunction with labs at USDA-PBARC, was finalized for publication, after completing a peer-review process, and will be published in Forest Pathology in October 2018. However, the beetles themselves gave mostly negative results for the presence of the fungal pathogen. During this reporting period, we have screened more beetle individuals to understand whether the beetles are transporting the disease as a byproduct of gallery formation, or whether the beetles have developed a mutual relationship with the fungus. We have continued to analyze our tentative results and have a partnership with a USGS lab to further investigate the relationship between the fungi and the beetles. In addition, further work has been done regarding the aerial sampling discussed in the last reporting period. In these trials, we monitored aerial sampling devices (Rotorod) in the districts with the forest pathogen to screen for aerosolized spores/dust from the disease. We also performed before-and-after control experiments to determine the amount of aerosolized spores/dust from infected trees that were felled. Samples were analyzed with quantitative molecular methods for the presence of fungal pathogens. The results of the work are still being assessed with partner agencies. 2. GENETIC VARIATION IN NON-NATIVE BARK BEETLES We isolated molecular markers from 48 bark beetle specimens collected in Kauai, Oahu, and Maui. These represent 7 species, 3 of which have purported association with the pathogen that causes Rapid Ohia Death. These are taken from over 4000 specimens collected from Kauai, as well as numerous specimens collected across the archipelago. With this small trial, we analyzed the variation of these markers within each species to investigate genetic diversity within populations. We are developing protocols to isolate genetic markers to identify fungal symbionts, and the use of genomic methods to better understand the genetic variation of the non-native bark beetles. To understand the ability of the bark beetles to impact the forests and croplands, we will additionally use these methods to determine the origin of some of the invasive bark beetle species, and whether there are multiple introductions into the Hawaiian Islands. Ideally, this will also enable us to assess the movement of beetles between Hawaiian Islands.
Publications
|
Progress 10/01/16 to 09/30/17
Outputs
Target Audience:The target audience for this reporting period was largely the local and national research communities dealing with a recent fungal pathogen affecting native trees and natural resource health (e.g., watersheds). Several researchers involved in this effort were further involved in direct public outreach. These outreach efforts were aimed at communicating the problems Hawaii is facing by beetle-vectored fungal pathogens and ways that they can either help report disease spread or mitigate its spread. Finally, outreach efforts also included several meetings and presentations with Hawaii State representatives, including the Hawaii State Governor, State Legislators, and State Land Managers. Changes/Problems:Nomajorproblems were facedover the term of this project, resulting in revision of approaches or objectives. The project wil be transferred to a new PI in the coming year. What opportunities for training and professional development has the project provided?A postdoctoral researcher worked on the project and was successful in obtaining an entomological position with the State of California. During the current and upcoming years, a PhD graduate student is/will be paid in part from these funds. This will help support his training and research on the main project objectives. Finally, a post-baccalaureate (Kylle Roy, currently employed by the USGS)research technician was hired from other funds, but was trained in part by the postdoctoral researcher. How have the results been disseminated to communities of interest?Work under this project was presented to the public, state legislators, and land managers through presentations to each of these groups. Below are summary of these events: Information has been presented at community meetings in Hilo and Kona (see outreach section below) and additional community outreach is planned for the future (May 9, 2017: In coordination with the Department of Public Works and the Mayor's Office - RapidOhia Death training for Hawaii County field workers). Gordon Bennett presented current work at the Rapid Ohia Death Summit convened by Governor David Ige. 30Nov2016, State Capitol Building, Honolulu, HI. Curtis Ewing presented at the Community Outreach ROD Symposia in Hilo and Kona: How does the fungus spread? 18March2017, University of Hawaii at Hilo and 1April2017 West Hawai'i Civic Center Kona. Curtis Ewing and Kylle Roy presented preliminary results for project #2 at the Tropical Conservation Biology and Environmental Science Research Symposium: Infectivity of boring dust produced by ambrosia beetles from ROD positive ohia trees. 7 April 20017, UH Hilo. Curtis Ewing and Kylle Roy will present at the Hawaii Conservation Conference 18-20July2017, Honolulu Hawaii. Researchers met with students from the West Hawaii Explorations Academy and homeowners in the Kaloko area of Kailua-Kona on 3-Feb-17 and 7-Mar-17. Trees were surveyed for insect activity and visible evidence of Ceratocystis infection. What do you plan to do during the next reporting period to accomplish the goals?The current PI, Dr. Gordon Bennett, will be leaving Hawaii at the end of this academic year. The project will be transitioned to a new Principal Investigator. Funds will be used to support and Ph.D. graduate student, Jared Bernard. Bernard will continue collecting bark beetle species that inhabit native forests across the archipelago to determine the relatively abundance and distributions of their populations. From this information population sizes and community structure will be estimated to better predict the potential of the tree disease introduction, establishment, and spread. Second, bark beetles will continue to be screened of fungal pathogens using established molecular techniques. Anticipated results will reveal which species carry potential pathogen(s) that cause Rapid Ohia Death, among others. Third, Bernard will analyze genomic markers and individual genes with modeling software to map the connectivity of beetle populations. These strategies will infer the corridors of movement for the bark beetle species that transport the disease.
Impacts
What was accomplished under these goals?
Several project goals were addressed in coordination with ongoing efforts by our lab and the State of Hawaii to understand and to manage the outbreak of a recent fungal pathogen killing Hawaii's dominant native tree, ohia (Metrosideros polymoprha). Ohia comprises the dominant tree in most of Hawaii's watersheds and is critically important in the health of natural resources and native environment. Bark Beetles (Scolytinae) are likely to play a major role in the spread of this disease. Our ongoing projects are addressing this question. Below is a list of project objectives the postdoctoral researcher accomplished with partial funding from the HATCH project over the past year: 1. ASSESSMENT OF BEETLE SPECIES ASSOCIATED WITH ??HI?A TREES Ohia trees were monitored for beetle emergence. Cut tree bolts were collected primarily on the windward side of the Big Island. Four species were reared in high numbers, indicating active breeding in infected and dying ohia trees. All species are adventive and include (i) Xyleborinus saxesenii, (ii) Xyleborus ferrugineus, (iii) Xyleborus affinis, and (iv) Xyleborus perforans. Xyleborinus saxesenii is the most common and widespread species associated with ohia and likely other trees in the environment. Thousands of individuals for this species were collected during the experiment. Xyleborinus saxesenii was also reared from near sea level to over 5,000 ft. elevation and from all areas of the island. Xyleborus ferrugineus was also commonly reared, but was not found in bolts collected from above 1,700 ft. elevation. These results indicate habitat and/or climate specificity for the distribution of this adventive species in Hawaii. Xyleborus perforans was reared simultaneously with X. ferrugineus, but in much smaller numbers. Xyleborus affinis was reared from bolts collected >2,000 ft. The native species Xyleborus simillimus, known to attack ohia, but was never reared from bolts, even when collected in their known habitat range. Three species were reared in very small numbers (n < 10 individuals), suggesting that they may on occasion attack ohia, but likely do not breed successfully in it: (i) Coccotrypes cyperi, (ii) Xylosandrus crassiusculus, and (iii) Xylosandrus morigerus. 2. BEETLE BORING-DUST AND BEETLE CARRIAGE OF CERATOCYSTIS 2a. Beetle Boring-Dust: Two hundred boring dust Traps (BDTs) were affixed directly to ohia trees that (i) known to be infected with the Ceratocystis pathogen and (ii) that had observable beetle infestations. BDTs were attached to the opening of active beetle galleries at four sites in the Puna district of the Big Island. Boring dust (BD) samples were screened for the presence of Ceratocystis with (i) DNA marker quantitative analysis (qPCR) and (ii) viable spore inoculum with a carrot-culturing method. A total of 160 BDT samples were analyzed. Nearly all BD collected was produced by X. ferrugineus; two X. perforans galleries/colonies were found with the traps as well. A total of 99 of 160 trap sampled BD (62%) tested positive for Ceratocystis, and 25 of 160 (16%) produced viable Ceratocystis culture. The latter indicates that at least some galleries produce inoculum that could infect other trees. We are currently preparing a publication of this work titles "Infectivity of boring dust produced by ambrosia beetles on Ceratocystis positive ohia trees on Hawaii Island" expected submission Forest Pathology, November 2017. 2b. Beetles: To date, a limited number of beetle specimens have been screened with qPCR. Of those species screened, all X. ferrugineus and Xy. saxesenii have tested negative. Of the ten X. perforans screened, two tested positive (~20%). Three of the Hawaiian endemic X. simillimus were further tested. One of the samples screened tested positive for Ceratocystis, indicating potential carriage of spores through wood consumption in this species. In summary, the limited number of beetles screened indicates that they do carry detectible Ceratocystis DNA. Whether or not these spores/fungi are viable, needs to be further tested. Potential disease carriage may vary between the different species in relation to their different methods of gallery formation, which is known to be distinct between some species and genera (wood consumption vs. non-wood consumption). Screening of more individuals is underway. 4. AERIAL SAMPLING OF CERATOCYSTIS INOCULUM In order to determine if (i) Ceratocystis spores are moved through the air column and (ii) if we have appropriate methods and technology to detect it, three rounds of Rotorod sampling have been carried out. All samples were subsequently screened for Ceratocystis DNA with the qPCR method: First, samplers were deployed for five days in Hawaiian Acres, Puna District. This site was heavily impacted by Ceratocystis disease and we have determined that beetle activity is high. Samples were taken daily, and no Ceratocystis was detected from the aerial samplers when deployed for only one week. Second, the Rotorod samples were moved to a managed site along the Wailuku River in the Piihonua area to monitor airborne inoculum before, during, and after felling of infected trees (i.e., a proposed management strategy by the State of Hawaii). Samples were taken for two days before cutting, two days after cutting, and six days afterward cutting. Positive samples for Ceratocystis were obtained downwind from the tree felling events. Tree felling likely increased the particulate matter from infected ohia, which can be detected with Rotorods. Third, a more intensive trial of Rotorod efficiency was carried out. Four traps were set: one at the CTAHR Waiakea Research Station upwind of local ohia dieback, and three were set near Stainback Hwy within the ohia dieback area. Samples were taken after 2 and 5-day intervals, and at once/week intervals thereafter for a total of eight weeks. A total of 30 samples were collected. Three positive (10%) samples were obtained for Ceratocystis DNA (two Ceratocystis sp. A and one Ceratocystis sp. B) when Rotorods were deployed for eight weeks. The latter sample collected at the "upwind" site at Waiakea station. In summary, the Rotorod samplers appear to be effective at detecting some level of Ceratocystis inoculum in the wind or environment. This is particularly the case when tree particle matter is high, as can occur during active tree felling. The Rotorods perform best when left in the environment over longer time periods with regular sampling and servicing. Rotorods should be sampled (i) at least at once/week intervals under normal environmental conditions over the course of several months, and (ii) daily under higher inoculum levels as found during cutting.
Publications
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