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• 3231. [Article] Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; September 1995

  Previously established field trials of MTTC-Fume continue to show that this chemical remains in Douglas-fir and southern pine poles at fungitoxic levels 5 years after treatment. In general, increasing ...

  Citation × Citation Citation Abstract Copy Title: Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; September 1995 Author: Morrell, Jeffrey J., Oregon State University. Dept. of Forest Products, Corden, M. E. (Malcolm E.) Previously established field trials of MTTC-Fume continue to show that this chemical remains in Douglas-fir and southern pine poles at fungitoxic levels 5 years after treatment. In general, increasing dosages resulted in higher chemical levels in the poles. All of the MITC-Fume treatments resulted in higher residual M1TC levels than comparable metham sodium treatments. Field trials with solid basaniid with and without copper sulfate continue to show the promise of this chemical. Basamid treated poles continue to contain MITC at levels which exceed those for comparable metham sodium treatments. These results, which are currently being confirmed in poles in service, indicate that this formulation can deliver fiingitoxic MITC levels to wood in service, without the risk of spills associated with metham sodium. We have completed trials with gelled and pelletized metham sodium formulations. Both of these formulations offered improved safety during application. Initial laboratory trials also suggested that the gelled formulation was more effective than comparable liquid metham sodium. Field trials, however, indicate that the formulations provided protection comparable to the liquid formulation in poles over the test period. The gelled formulation still offers the advantage of reduced risks of spills during application. Trials are also continuing with various solid, water diffusible treatments for arresting internal decay. Trials of fused borate rods continue to show that these treatments require more moisture for effective movement in Douglas-fir poles. Field trials have also been established with borate rods with glycol as an additive to determine if glycols can accelerate diffusion in drier wood. The poles in these trials have also been sensored to monitor internal moisture changes over time in order to better correlate boron diffusion with wood moisture content. Trials underway with a boronlfluoride rod indicate that the boron is diffusing well from these rods, while the fluoride is moving somewhat slower. Neither chemical has approached a fiungal threshold one year after treatment, but the combination of chemicals may lead to more effective fungal control. Trials with pelletized metham sodium/basamid mixtures suggests that using ratios of these two chemicals can produce an initial rapid burst ofMITC release followed by a slower MITC release with time. This combination allows for rapid control of existing fungal infestations followed by long term protection against reinvasion. These laboratory trials will be further confirmed with field trials. The trials to evaluate the effects of voids on fumigant movement initially suggested that voids have little influence on subsequent fumigant levels on either side of the void. Sampling 8 years after treatment, however, indicates that chioropicrin levels were generally higher in poles without voids. We plan further trials of actual field poles containing voids to confirm these effects. Trials to identify treatments for protecting the sapwood of western redcedar poles as well as wood exposed in field drilled bolt holes are continuing. Trials in western redcedar sapwood have identified a number of chemicals which can be remedially applied to protect this wood against fungal attack. At present, however, commercial pole spraying has largely ceased making further field trials difficult. Field trials to identify treatments for protecting untreated wood exposed during drilling for various pole attachments continue to show that diffusible boron and fluoride compounds provide excellent long term protection against flingal attack. The protective effects of one diffusible treatment, Boracol, however, has begun to decline. Further sampling will be undertaken to identify the long term effectiveness of the remaining treatments. Trials to identify enhanced patterns for through boring of Douglas-fir poles are complete. The results indicate that patterns as widely spaced as 400 mm apart longitudinally still produce a nearly completely treated pole i n the through bored zone. Pentachlorophenol levels in the through bored zone were generally above the threshold for ftrngal growth. Prior sampling of Douglas-fir poles in service suggest that even poles with small skips in the through bored zone contained no evidence of internal decay. These results suggest that the through boring pattern can be extended without adversely risking pole service life. A reduced through boring pattern would decrease treatment costs while minimizing impacts on pole strength. Trials to evaluate the application of boron to freshly peeled Douglas-fir poles as a means of preventing fungal colonization using a thermal process suggest that thermal treatment failed to produce a boron loading sufilcient to permit subsequent diffusion across the pole section after a 3 month diffusion period. Evaluations of various external groundline preservative formulations continue to indicate that replacement formulations based upon copper naphthenate, boron or fluoride perform comparably to earlier formulations employing pentachlorophenol and creosote. Pentachlorophenol levels in some treatments have fallen below the threshold for fungal growth, while those in all of the replacement treatments remain above a U protective level. These results suggest that the newer groundline preservative systems should provide a reasonable level of protection against external decay. Laboratory trials to better understand the levels of combinations of chemicals required for protection in the groundline zone are continuing. Fungal cellar evaluations of copper naphthenate treated western redcedar continue to show excellent performance at levels specified in the American Wood Preservers' Association Standards. Performance is generally better for wood which was freshly sawn prior to treatment, while wood cuts from weathered poles in service has provided slightly lower levels of protection. The weather wood apparently has higher permeability, making it more likely to lose chemical in soil contact. Further evaluations are planned. Title: Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; September 1995 Author: Morrell, Jeffrey J., Oregon State University. Dept. of Forest Products, Corden, M. E. (Malcolm E.) Previously established field trials of MTTC-Fume continue to show that this chemical remains in Douglas-fir and southern pine poles at fungitoxic levels 5 years after treatment. In general, increasing dosages resulted in higher chemical levels in the poles. All of the MITC-Fume treatments resulted in higher residual M1TC levels than comparable metham sodium treatments. Field trials with solid basaniid with and without copper sulfate continue to show the promise of this chemical. Basamid treated poles continue to contain MITC at levels which exceed those for comparable metham sodium treatments. These results, which are currently being confirmed in poles in service, indicate that this formulation can deliver fiingitoxic MITC levels to wood in service, without the risk of spills associated with metham sodium. We have completed trials with gelled and pelletized metham sodium formulations. Both of these formulations offered improved safety during application. Initial laboratory trials also suggested that the gelled formulation was more effective than comparable liquid metham sodium. Field trials, however, indicate that the formulations provided protection comparable to the liquid formulation in poles over the test period. The gelled formulation still offers the advantage of reduced risks of spills during application. Trials are also continuing with various solid, water diffusible treatments for arresting internal decay. Trials of fused borate rods continue to show that these treatments require more moisture for effective movement in Douglas-fir poles. Field trials have also been established with borate rods with glycol as an additive to determine if glycols can accelerate diffusion in drier wood. The poles in these trials have also been sensored to monitor internal moisture changes over time in order to better correlate boron diffusion with wood moisture content. Trials underway with a boronlfluoride rod indicate that the boron is diffusing well from these rods, while the fluoride is moving somewhat slower. Neither chemical has approached a fiungal threshold one year after treatment, but the combination of chemicals may lead to more effective fungal control. Trials with pelletized metham sodium/basamid mixtures suggests that using ratios of these two chemicals can produce an initial rapid burst ofMITC release followed by a slower MITC release with time. This combination allows for rapid control of existing fungal infestations followed by long term protection against reinvasion. These laboratory trials will be further confirmed with field trials. The trials to evaluate the effects of voids on fumigant movement initially suggested that voids have little influence on subsequent fumigant levels on either side of the void. Sampling 8 years after treatment, however, indicates that chioropicrin levels were generally higher in poles without voids. We plan further trials of actual field poles containing voids to confirm these effects. Trials to identify treatments for protecting the sapwood of western redcedar poles as well as wood exposed in field drilled bolt holes are continuing. Trials in western redcedar sapwood have identified a number of chemicals which can be remedially applied to protect this wood against fungal attack. At present, however, commercial pole spraying has largely ceased making further field trials difficult. Field trials to identify treatments for protecting untreated wood exposed during drilling for various pole attachments continue to show that diffusible boron and fluoride compounds provide excellent long term protection against flingal attack. The protective effects of one diffusible treatment, Boracol, however, has begun to decline. Further sampling will be undertaken to identify the long term effectiveness of the remaining treatments. Trials to identify enhanced patterns for through boring of Douglas-fir poles are complete. The results indicate that patterns as widely spaced as 400 mm apart longitudinally still produce a nearly completely treated pole i n the through bored zone. Pentachlorophenol levels in the through bored zone were generally above the threshold for ftrngal growth. Prior sampling of Douglas-fir poles in service suggest that even poles with small skips in the through bored zone contained no evidence of internal decay. These results suggest that the through boring pattern can be extended without adversely risking pole service life. A reduced through boring pattern would decrease treatment costs while minimizing impacts on pole strength. Trials to evaluate the application of boron to freshly peeled Douglas-fir poles as a means of preventing fungal colonization using a thermal process suggest that thermal treatment failed to produce a boron loading sufilcient to permit subsequent diffusion across the pole section after a 3 month diffusion period. Evaluations of various external groundline preservative formulations continue to indicate that replacement formulations based upon copper naphthenate, boron or fluoride perform comparably to earlier formulations employing pentachlorophenol and creosote. Pentachlorophenol levels in some treatments have fallen below the threshold for fungal growth, while those in all of the replacement treatments remain above a U protective level. These results suggest that the newer groundline preservative systems should provide a reasonable level of protection against external decay. Laboratory trials to better understand the levels of combinations of chemicals required for protection in the groundline zone are continuing. Fungal cellar evaluations of copper naphthenate treated western redcedar continue to show excellent performance at levels specified in the American Wood Preservers' Association Standards. Performance is generally better for wood which was freshly sawn prior to treatment, while wood cuts from weathered poles in service has provided slightly lower levels of protection. The weather wood apparently has higher permeability, making it more likely to lose chemical in soil contact. Further evaluations are planned. Close

• 3232. [Article] Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; August 1992

  Evaluation of previously established field trials of remedial internal treatments demonstrates the continued performance of chioropicrin, Vorlex, and methylisothiocyanate (MITC). While the degree of protection ...

  Citation × Citation Citation Abstract Copy Title: Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; August 1992 Author: Morrell, Jeffrey J., Oregon State University. Dept. of Forest Products, Corden, M. E. (Malcolm E.) Evaluation of previously established field trials of remedial internal treatments demonstrates the continued performance of chioropicrin, Vorlex, and methylisothiocyanate (MITC). While the degree of protection afforded by these treatments has declined with time, residual fungitoxic levels remain in many tests. Field trials of gelatin encapsulated MITC indicate that gelatin had no negative effect on fumigant performance even when no water was added at the time of treatment. Closed tube bioassays, chemical analyses, and culturing of Douglas-fir and southern pine poles treated with glass encapsulated METC (MITC-Fume) indicate that this chemical is outperforming metham sodium 3 years after application. While the glass vials lost chemical very slowly, the slow release rate did not appear to adversely affect MITC performance. Trials to evaluate the performance of fused borate rods were sampled after 1 or 2 years of exposure. Chemical analysis of cores removed from the test poles revealed that none of the treatments contained boron at levels which would be considered adequate for arresting or preventing colonization by wood decay fungi. Interestingly, boron levels in poles exposed in Hilo, Hawaii were highest above the treatment hole, suggesting that some upward diffusion of this chemical is possible. The low boron levels in these poles are reason for concern, since a number of utilities are considering the using this formulation for remedial treatment at the groundline. Evaluations of new solid fumigants are progressing. Trials with Basamid indicate that the addition of copper compounds improved the rate of decomposition to produce MITC. Simultaneous addition of copper sulfate and Basamid may be useful for accelerating the decomposition of this compound, making it practical for control of internal decay fungi. Trials have been established to evaluate the performance of gelled and pelletized metham sodium and a sodium fluoride/boron rod. These trials will be evaluated in future reports. A third field trial to evaluate the performance of a copper naphthenate/boron paste for internal treatment of Douglas-fir poles is currently be evaluated to determine chemical levels 3 years after treatment. The performance of gelled metham sodium was further evaluated under laboratory conditions to better understand the performance of this chemical. Gelled metham sodium provided improved fungal control in comparison with liquid metham sodium and appeared to produce increased MITC levels under a variety of test conditions. The improved performance of this formulation may reflect the ability of the gell to retain moisture for longer periods of time than the liquid metham sodium formulation. Further studies of this formulation are underway. Laboratory studies were also performed to evaluate the effects of various additives on the performance of Basamid. Once again, the addition of copper compounds enhanced the production of MITC. A number of other compounds shifted decomposition to the production of carbon disulfide and carbonyl sulfide, two less fungitoxic compounds. Further studies are underway to identify non-sulfur products which may provide some protection against wood decay fungi. Evaluations of the effects of artificial voids on performance of fumigants in Douglas-fir poles indicate that voids had little or no effect on fumigant distribution. As a result, fumigant treatment of solid wood around voids represents a viable strategy for improving pole service life. Evaluation of timbers treated with metham sodium indicate that detectable levels of MITC were present one year after treatment. These timbers will be evaluated in subsequent years to determine the protective period provided by fumigants in sawn material. We continue development of a fumigant movement model using data previously developed on MITC. This year, we evaluated a previously developed system, ANSYS. Results of preliminary trials are similar to data previously developed on MITC-Fume treated poles and indicate that modeling MITC movement should be possible. Further trials are underway to confirm and expand this model. The effect of wood moisture content, temperature and wood species on metham sodium decomposition was investigated under laboratory conditions. The efficiency of dcomposition to MITC varied widely, but was most affected by temperature and wood moisture content. The results suggests that there is considerable potential for improving decomposition efficiency to enhance performance of this fumigant. Further studies to characterize the relationship between chemical content of the wood species and decomposition are underway. Field trials to identify safer treatments for preventing decay of cedar sapwood and protecting field drilled bolt holes are continuing. Diffusible treatments continue to provide excellent protection for field drilled bolt holes. A study to develop estimates of the extent of decay above the groundline in Douglas-fir poles in the Pacific Northwest is underway. The data from this study will be used to develop estimates of the potential for damage and provide some insight into the extent of this problem. Studies to develop guidelines for sterilization of Douglas-fir poles following air-seasoning are continuing. Evaluations of internal temperature development during kiln-drying were completed this year and indicate that internal temperatures during typical pole drying schedules were more than adequate for acheiving sterilization. Further evaluations of the data are underway to develop reliable heating curves for this process. Evaluations of groundline preservative systems have been established at Corvallis, OR and Merced, CA. The results indicate that all of the formulations are moving well through the wood in a manner similar to that found with pentachlorophenol-based systems. Chemical levels in some treatments; however, are beginning to decline 30 months after treatment. Studies are now underway to establish thresholds for combinations of the various formulations. Copper naphthenate treated western redcedar stakelets continue to perform well in fungus cellar trials. Stakes weathered prior to treatment are degrading slightly faster, while freshly sawn stakelets continue to perform well. Field trials have also been established to evaluate the performance of copper naphthenate treated Douglas-fir utility poles in California and Oregon. The chemical levels and fungal colonization will be monitored in these poles to provide a guide to performance of this chemical in western wood species. Title: Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; August 1992 Author: Morrell, Jeffrey J., Oregon State University. Dept. of Forest Products, Corden, M. E. (Malcolm E.) Evaluation of previously established field trials of remedial internal treatments demonstrates the continued performance of chioropicrin, Vorlex, and methylisothiocyanate (MITC). While the degree of protection afforded by these treatments has declined with time, residual fungitoxic levels remain in many tests. Field trials of gelatin encapsulated MITC indicate that gelatin had no negative effect on fumigant performance even when no water was added at the time of treatment. Closed tube bioassays, chemical analyses, and culturing of Douglas-fir and southern pine poles treated with glass encapsulated METC (MITC-Fume) indicate that this chemical is outperforming metham sodium 3 years after application. While the glass vials lost chemical very slowly, the slow release rate did not appear to adversely affect MITC performance. Trials to evaluate the performance of fused borate rods were sampled after 1 or 2 years of exposure. Chemical analysis of cores removed from the test poles revealed that none of the treatments contained boron at levels which would be considered adequate for arresting or preventing colonization by wood decay fungi. Interestingly, boron levels in poles exposed in Hilo, Hawaii were highest above the treatment hole, suggesting that some upward diffusion of this chemical is possible. The low boron levels in these poles are reason for concern, since a number of utilities are considering the using this formulation for remedial treatment at the groundline. Evaluations of new solid fumigants are progressing. Trials with Basamid indicate that the addition of copper compounds improved the rate of decomposition to produce MITC. Simultaneous addition of copper sulfate and Basamid may be useful for accelerating the decomposition of this compound, making it practical for control of internal decay fungi. Trials have been established to evaluate the performance of gelled and pelletized metham sodium and a sodium fluoride/boron rod. These trials will be evaluated in future reports. A third field trial to evaluate the performance of a copper naphthenate/boron paste for internal treatment of Douglas-fir poles is currently be evaluated to determine chemical levels 3 years after treatment. The performance of gelled metham sodium was further evaluated under laboratory conditions to better understand the performance of this chemical. Gelled metham sodium provided improved fungal control in comparison with liquid metham sodium and appeared to produce increased MITC levels under a variety of test conditions. The improved performance of this formulation may reflect the ability of the gell to retain moisture for longer periods of time than the liquid metham sodium formulation. Further studies of this formulation are underway. Laboratory studies were also performed to evaluate the effects of various additives on the performance of Basamid. Once again, the addition of copper compounds enhanced the production of MITC. A number of other compounds shifted decomposition to the production of carbon disulfide and carbonyl sulfide, two less fungitoxic compounds. Further studies are underway to identify non-sulfur products which may provide some protection against wood decay fungi. Evaluations of the effects of artificial voids on performance of fumigants in Douglas-fir poles indicate that voids had little or no effect on fumigant distribution. As a result, fumigant treatment of solid wood around voids represents a viable strategy for improving pole service life. Evaluation of timbers treated with metham sodium indicate that detectable levels of MITC were present one year after treatment. These timbers will be evaluated in subsequent years to determine the protective period provided by fumigants in sawn material. We continue development of a fumigant movement model using data previously developed on MITC. This year, we evaluated a previously developed system, ANSYS. Results of preliminary trials are similar to data previously developed on MITC-Fume treated poles and indicate that modeling MITC movement should be possible. Further trials are underway to confirm and expand this model. The effect of wood moisture content, temperature and wood species on metham sodium decomposition was investigated under laboratory conditions. The efficiency of dcomposition to MITC varied widely, but was most affected by temperature and wood moisture content. The results suggests that there is considerable potential for improving decomposition efficiency to enhance performance of this fumigant. Further studies to characterize the relationship between chemical content of the wood species and decomposition are underway. Field trials to identify safer treatments for preventing decay of cedar sapwood and protecting field drilled bolt holes are continuing. Diffusible treatments continue to provide excellent protection for field drilled bolt holes. A study to develop estimates of the extent of decay above the groundline in Douglas-fir poles in the Pacific Northwest is underway. The data from this study will be used to develop estimates of the potential for damage and provide some insight into the extent of this problem. Studies to develop guidelines for sterilization of Douglas-fir poles following air-seasoning are continuing. Evaluations of internal temperature development during kiln-drying were completed this year and indicate that internal temperatures during typical pole drying schedules were more than adequate for acheiving sterilization. Further evaluations of the data are underway to develop reliable heating curves for this process. Evaluations of groundline preservative systems have been established at Corvallis, OR and Merced, CA. The results indicate that all of the formulations are moving well through the wood in a manner similar to that found with pentachlorophenol-based systems. Chemical levels in some treatments; however, are beginning to decline 30 months after treatment. Studies are now underway to establish thresholds for combinations of the various formulations. Copper naphthenate treated western redcedar stakelets continue to perform well in fungus cellar trials. Stakes weathered prior to treatment are degrading slightly faster, while freshly sawn stakelets continue to perform well. Field trials have also been established to evaluate the performance of copper naphthenate treated Douglas-fir utility poles in California and Oregon. The chemical levels and fungal colonization will be monitored in these poles to provide a guide to performance of this chemical in western wood species. Close

• 3233. [Article] Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; October 1996

  The Cooperative continues to actively address a diverse array of issues related to the effective use of wood utility poles. The trials to evaluate the effectiveness of MITC-Fume are now in their seventh ...

  Citation × Citation Citation Abstract Copy Title: Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; October 1996 Author: Morrell, Jeffrey J., Oregon State University. Dept. of Forest Products, Corden, M. E. (Malcolm E.) The Cooperative continues to actively address a diverse array of issues related to the effective use of wood utility poles. The trials to evaluate the effectiveness of MITC-Fume are now in their seventh year and continue to show that methylisothiocyanate (MITC) levels in MITC-Fume treatments remain higher than comparable metham sodium treatments. The levels of chemical are, however, declining, suggesting that this treatment may need replenishment in 3 to 5 years. Trials with the solid wood fumigant Basamid continue to show that MITC release can be enhanced by addition of small amounts of copper. Field trials suggest that these additives become less important with time. As a result, unamended Basamid may be suitable for treatment where the risk of immediate decay is not high, but the utility wishes to protect against future attack. Field trials with various water-diffusible internal treatments continue to show that these treatments move more slowly through Douglas-fir heartwood than do fumigants. Boron levels in pole sections treated with fused borate rods remain at levels that will protect against fungal attack 6 years after treatment. Similar trials with a boron/ fluoride rod indicate that neither fluoride or boron levels in the poles are adequate for wood protection 2 years after treatment. While the dosages tested were relatively low, the volumes of chemical were similar to the liquid volumes normally applied during internal remedial treatment. We will sample these poles next year to ensure that our measurements accurately reflect the chemical levels present. Trials to evaluate the effects of glycol on boron movement from fused borate rods suggest that glycol enhanced boron diffusion to only a slight extent. This effect was most pronounced at lower moisture contents. This trial was established to identify methods for improving boron movement in drier wood. In addition, moisture measurements in these poles suggest the internal wood moisture content varies widely both seasonally and positionally. While elevated moisture levels can negatively affect the movement of gaseous fumigants, excess moisture is critical for diffusion of boron or fluoride and its absence around the treatment site can markedly reduce the efficacy of rod treatments. These poles will continue to be monitored to assess both boron movement and seasonal changes in moisture content. Trials to identify safe, effective and easily used systems for protecting wood exposed during field fabrication are continuing. Boron and fluoride continue to provide excellent protection to field drilled bolt holes. These treatments are safe and easy to apply, and have provided protection in our field test for 14 years. Trials of similar formulations on simulated decking are also reported to provide additional information on the ability of boron and fluoride to protect exposed Douglas-fir heartwood. Efforts to improve the effectiveness of through boring as a method for enhancing the treatment of Douglas-fir poles are continuing. This past year, we evaluated preservative distribution around through bored holes as a means of developing optimum through boring patterns that maximized treatment while minimizing potential strength effects. These trials suggest the diamond shaped through boring zone of effect is relatively narrow. This information will be used in the coming year to construct optimum patterns for poles of various classes. The goal of this project is to develop a standard through boring pattern that would permit automation of the process. This would create the potential for cost savings on new poles. Trials to evaluate the durability of western redcedar are nearly complete. These trials were initiated because of concerns that second growth western redcedar might be less durable than poles cut from older trees. As expected, cedar varied widely in its resistance to fungal attack. This resistance, however, was not related to tree age, suggesting that there might not be a difference between so-called "old-growth" and "second growth" material. These data will be more thoroughly analyzed once the final set of trials are completed. In addition, we are evaluating more rapid methods for assessing cedar durability by measuring tropolone content. Tropolones are an important component of the extractives that make cedar heartwood so durable. Field trials of various externally applied supplemental groundline treatments are continuing at sites in Oregon, California, and New York. Trials in Corvallis, Oregon, have shown that various copper naphthenate, fluoride or boron based systems are at least as effective as the pentachlorophenol (penta) based systems that were formerly used for this purpose. Penta concentrations in one system have now declined below a protective level, while the copper based systems continue to remain at a protective level. Field trials in California are following similar trends and indicate that the alternative systems will provide comparable performance. Fungus cellar trials of copper naphthenate treated western redcedar stakes continue to show that this chemical provides excellent protection to cedar sapwood. Weathered wood that was treated with copper naphthenate continues to perform more poorly than freshly sawn wood treated to similar retention levels. Variations in permeability likely account for these differences. A new Wood Pole Maintenance Manual has been completed and is now ready for distribution. This update of the 1979 publication includes more information on initial pole procurement and closely follows the video by the same name that we produced in 1994. Title: Conserving energy by environmentally acceptable practices in maintaining and procuring transmission poles for long service ; October 1996 Author: Morrell, Jeffrey J., Oregon State University. Dept. of Forest Products, Corden, M. E. (Malcolm E.) The Cooperative continues to actively address a diverse array of issues related to the effective use of wood utility poles. The trials to evaluate the effectiveness of MITC-Fume are now in their seventh year and continue to show that methylisothiocyanate (MITC) levels in MITC-Fume treatments remain higher than comparable metham sodium treatments. The levels of chemical are, however, declining, suggesting that this treatment may need replenishment in 3 to 5 years. Trials with the solid wood fumigant Basamid continue to show that MITC release can be enhanced by addition of small amounts of copper. Field trials suggest that these additives become less important with time. As a result, unamended Basamid may be suitable for treatment where the risk of immediate decay is not high, but the utility wishes to protect against future attack. Field trials with various water-diffusible internal treatments continue to show that these treatments move more slowly through Douglas-fir heartwood than do fumigants. Boron levels in pole sections treated with fused borate rods remain at levels that will protect against fungal attack 6 years after treatment. Similar trials with a boron/ fluoride rod indicate that neither fluoride or boron levels in the poles are adequate for wood protection 2 years after treatment. While the dosages tested were relatively low, the volumes of chemical were similar to the liquid volumes normally applied during internal remedial treatment. We will sample these poles next year to ensure that our measurements accurately reflect the chemical levels present. Trials to evaluate the effects of glycol on boron movement from fused borate rods suggest that glycol enhanced boron diffusion to only a slight extent. This effect was most pronounced at lower moisture contents. This trial was established to identify methods for improving boron movement in drier wood. In addition, moisture measurements in these poles suggest the internal wood moisture content varies widely both seasonally and positionally. While elevated moisture levels can negatively affect the movement of gaseous fumigants, excess moisture is critical for diffusion of boron or fluoride and its absence around the treatment site can markedly reduce the efficacy of rod treatments. These poles will continue to be monitored to assess both boron movement and seasonal changes in moisture content. Trials to identify safe, effective and easily used systems for protecting wood exposed during field fabrication are continuing. Boron and fluoride continue to provide excellent protection to field drilled bolt holes. These treatments are safe and easy to apply, and have provided protection in our field test for 14 years. Trials of similar formulations on simulated decking are also reported to provide additional information on the ability of boron and fluoride to protect exposed Douglas-fir heartwood. Efforts to improve the effectiveness of through boring as a method for enhancing the treatment of Douglas-fir poles are continuing. This past year, we evaluated preservative distribution around through bored holes as a means of developing optimum through boring patterns that maximized treatment while minimizing potential strength effects. These trials suggest the diamond shaped through boring zone of effect is relatively narrow. This information will be used in the coming year to construct optimum patterns for poles of various classes. The goal of this project is to develop a standard through boring pattern that would permit automation of the process. This would create the potential for cost savings on new poles. Trials to evaluate the durability of western redcedar are nearly complete. These trials were initiated because of concerns that second growth western redcedar might be less durable than poles cut from older trees. As expected, cedar varied widely in its resistance to fungal attack. This resistance, however, was not related to tree age, suggesting that there might not be a difference between so-called "old-growth" and "second growth" material. These data will be more thoroughly analyzed once the final set of trials are completed. In addition, we are evaluating more rapid methods for assessing cedar durability by measuring tropolone content. Tropolones are an important component of the extractives that make cedar heartwood so durable. Field trials of various externally applied supplemental groundline treatments are continuing at sites in Oregon, California, and New York. Trials in Corvallis, Oregon, have shown that various copper naphthenate, fluoride or boron based systems are at least as effective as the pentachlorophenol (penta) based systems that were formerly used for this purpose. Penta concentrations in one system have now declined below a protective level, while the copper based systems continue to remain at a protective level. Field trials in California are following similar trends and indicate that the alternative systems will provide comparable performance. Fungus cellar trials of copper naphthenate treated western redcedar stakes continue to show that this chemical provides excellent protection to cedar sapwood. Weathered wood that was treated with copper naphthenate continues to perform more poorly than freshly sawn wood treated to similar retention levels. Variations in permeability likely account for these differences. A new Wood Pole Maintenance Manual has been completed and is now ready for distribution. This update of the 1979 publication includes more information on initial pole procurement and closely follows the video by the same name that we produced in 1994. Close

• 3234. [Article] Grape and Wine Aroma Influenced by Vine Nutrient Status, Vigor and Crop Levels in Oregon Pinot Noir

  Wine volatile composition is one of the most important constitutes of wine quality, and it is greatly influenced by number of factors. The studies herein were conducted to evaluate the grape and wine aroma ...

  Citation × Citation Citation Abstract Copy Title: Grape and Wine Aroma Influenced by Vine Nutrient Status, Vigor and Crop Levels in Oregon Pinot Noir Author: Yuan, Fang Wine volatile composition is one of the most important constitutes of wine quality, and it is greatly influenced by number of factors. The studies herein were conducted to evaluate the grape and wine aroma affected by different vine nutrient status, vigor and crop levels of Oregon Pinot noir. In the first study, aroma potentials in early and late maturity Pinot noir grapes were investigated in two consecutive vintages. The grape samples were hydrolyzed under acidic conditions and the released odorants were studied by aroma extract dilution analysis (AEDA). Forty-nine main odor-active compounds were detected in the AEDA. The odorants released with high flavor dilution values were 1-hexanal, β-damascenone, guaiacol and vanillin, together with C₆ aldehydes and alcohols, 4-vinylguaiacol, 4-vinylphenol and 1-octen-3-one. The concentrations of aroma-active compounds were further quantitated by SPME-GC-MS and SBSE-GC-MS. Comparing with early harvest grapes, late harvest grapes released more β-damascenone, vanillin, 4-vinylguaiacol and 4-vinylphenol in both years according to both AEDA and quantitation results, suggesting they were important aroma compounds that contribute to the characteristic of mature Pinot noir grapes. The second study evaluated the volatile composition of grape berries subjected to varying levels of nitrogen (N), phosphorous (P) and potassium (K) supply. Pinot noir grapevines were grown in a pot-in-pot system for three years (beginning when they were 4-years-old) and fertigated with either complete nutrition (Control) or with reduced levels of either N, P, or K supply while holding all other nutrients constant. Nitrogen was varied from 7.50 mM total N supply (Control) to 1.13 mM in five discreet treatments, while P and K supply were each varied in four discreet treatments with the lowest rate of 0 mM during fertigation events. Aroma volatiles in berries were determined in each year using SPME-GC-MS technique. Results showed that reducing N supply resulted in lower concentrations of C₆ compounds and total β-damascenone in berries across all three years. Low N supply had little impact on monoterpenes. Reducing N supply resulted in higher bound form α-terpineol in two of the three years. Compared to P and K, N supply showed a more profound influence on the volatile profiles of the grape berries, indicating the important role of N in the biosynthesis of berry volatile and volatile precursors. The volatile composition of resulting wines from the same study was also investigated. Results showed that reducing N supply increased the total phenolic content in wine across all years. N supply has a profound impact on wine volatiles with the greatest effect on yeast-derived esters and higher alcohols. In general, reducing N supply decreased straight-chain esters and many straight chain alcohols, but increased the branched-chain esters, and phenethyl ester and phenethyl alcohol in wine across all years. The effect of N supply on monoterpenes in wine was not consistent from year to year. Low N wines also had reduced levels of volatile sulfur compounds in 2 of 3 years. Compared to the Control, reducing N and K supply resulted in lower quantities of total β-damascenone across all years. Altering P supply did not have reproducible effects on wine volatiles from year to year. The third study investigated the composition of Pinot noir wines produced from vines with varying vegetative vigor levels and two crop levels over three vintages (2011, 2012, and 2013) in the cool climate viticulture region of western Oregon. Wine was produced from grapes grown with two inter-row floor management treatments (tilled or grass) and two crop levels, including full crop and half crop achieved through cluster thinning. Crop thinning treatments didn’t alter the wine volatile composition. Certain wine volatiles were affected by the vineyard floor management treatments. Wines produced from grapes grown in the Grass treatment (lower vigor) had higher levels of grape-derived compounds such as α-terpineol, β-citronellol, vitispirane and TDN compared to wines produced from the Tilled treatment. Wines from Grass treatment also had higher isoamyl alcohol, phenethyl alcohol, isoamyl acetate, ethyl isobutyrate, ethyl isovalerate, and phenethyl acetate but lower levels of linalool, 1-propanol and 1-octanol. A subproject of this study investigated the carotenoid breakdown and C₁₃-norisoprenoids synthesis in Pinot noir grapes from vines of differing vigor and crop levels (yield). Carotenoid degradation and C₁₃-norisoprenoid synthesis during berry development with different vine balance was monitored from véraison to harvest in year 2013. Our data suggests that vineyard floor management had a greater influence than did cluster thinning on carotenoids composition in grapes. Grape berries from Tilled treatment vines have higher carotenoids and C₁₃-norisoprenoids than the grapes from Grass treatment vines. Multivariance analyses also showed some interactions between the vineyard floor treatments and cluster thinning treatments on the composition of carotenoids and C₁₃-norisoprenoids. Title: Grape and Wine Aroma Influenced by Vine Nutrient Status, Vigor and Crop Levels in Oregon Pinot Noir Author: Yuan, Fang Wine volatile composition is one of the most important constitutes of wine quality, and it is greatly influenced by number of factors. The studies herein were conducted to evaluate the grape and wine aroma affected by different vine nutrient status, vigor and crop levels of Oregon Pinot noir. In the first study, aroma potentials in early and late maturity Pinot noir grapes were investigated in two consecutive vintages. The grape samples were hydrolyzed under acidic conditions and the released odorants were studied by aroma extract dilution analysis (AEDA). Forty-nine main odor-active compounds were detected in the AEDA. The odorants released with high flavor dilution values were 1-hexanal, β-damascenone, guaiacol and vanillin, together with C₆ aldehydes and alcohols, 4-vinylguaiacol, 4-vinylphenol and 1-octen-3-one. The concentrations of aroma-active compounds were further quantitated by SPME-GC-MS and SBSE-GC-MS. Comparing with early harvest grapes, late harvest grapes released more β-damascenone, vanillin, 4-vinylguaiacol and 4-vinylphenol in both years according to both AEDA and quantitation results, suggesting they were important aroma compounds that contribute to the characteristic of mature Pinot noir grapes. The second study evaluated the volatile composition of grape berries subjected to varying levels of nitrogen (N), phosphorous (P) and potassium (K) supply. Pinot noir grapevines were grown in a pot-in-pot system for three years (beginning when they were 4-years-old) and fertigated with either complete nutrition (Control) or with reduced levels of either N, P, or K supply while holding all other nutrients constant. Nitrogen was varied from 7.50 mM total N supply (Control) to 1.13 mM in five discreet treatments, while P and K supply were each varied in four discreet treatments with the lowest rate of 0 mM during fertigation events. Aroma volatiles in berries were determined in each year using SPME-GC-MS technique. Results showed that reducing N supply resulted in lower concentrations of C₆ compounds and total β-damascenone in berries across all three years. Low N supply had little impact on monoterpenes. Reducing N supply resulted in higher bound form α-terpineol in two of the three years. Compared to P and K, N supply showed a more profound influence on the volatile profiles of the grape berries, indicating the important role of N in the biosynthesis of berry volatile and volatile precursors. The volatile composition of resulting wines from the same study was also investigated. Results showed that reducing N supply increased the total phenolic content in wine across all years. N supply has a profound impact on wine volatiles with the greatest effect on yeast-derived esters and higher alcohols. In general, reducing N supply decreased straight-chain esters and many straight chain alcohols, but increased the branched-chain esters, and phenethyl ester and phenethyl alcohol in wine across all years. The effect of N supply on monoterpenes in wine was not consistent from year to year. Low N wines also had reduced levels of volatile sulfur compounds in 2 of 3 years. Compared to the Control, reducing N and K supply resulted in lower quantities of total β-damascenone across all years. Altering P supply did not have reproducible effects on wine volatiles from year to year. The third study investigated the composition of Pinot noir wines produced from vines with varying vegetative vigor levels and two crop levels over three vintages (2011, 2012, and 2013) in the cool climate viticulture region of western Oregon. Wine was produced from grapes grown with two inter-row floor management treatments (tilled or grass) and two crop levels, including full crop and half crop achieved through cluster thinning. Crop thinning treatments didn’t alter the wine volatile composition. Certain wine volatiles were affected by the vineyard floor management treatments. Wines produced from grapes grown in the Grass treatment (lower vigor) had higher levels of grape-derived compounds such as α-terpineol, β-citronellol, vitispirane and TDN compared to wines produced from the Tilled treatment. Wines from Grass treatment also had higher isoamyl alcohol, phenethyl alcohol, isoamyl acetate, ethyl isobutyrate, ethyl isovalerate, and phenethyl acetate but lower levels of linalool, 1-propanol and 1-octanol. A subproject of this study investigated the carotenoid breakdown and C₁₃-norisoprenoids synthesis in Pinot noir grapes from vines of differing vigor and crop levels (yield). Carotenoid degradation and C₁₃-norisoprenoid synthesis during berry development with different vine balance was monitored from véraison to harvest in year 2013. Our data suggests that vineyard floor management had a greater influence than did cluster thinning on carotenoids composition in grapes. Grape berries from Tilled treatment vines have higher carotenoids and C₁₃-norisoprenoids than the grapes from Grass treatment vines. Multivariance analyses also showed some interactions between the vineyard floor treatments and cluster thinning treatments on the composition of carotenoids and C₁₃-norisoprenoids. Close

• 3235. [Article] Kinetic and molecular effects of sulfate reduction on a dechlorinating culture under chemostat growth conditions

  Chlorinated ethenes are common groundwater contaminants that may be treated through in-situ bioremediation. Relationships between the reducing environment, available electron donors and acceptors, reaction ...

  Citation × Citation Citation Abstract Copy Title: Kinetic and molecular effects of sulfate reduction on a dechlorinating culture under chemostat growth conditions Author: Berggren, Dusty Rose V. Chlorinated ethenes are common groundwater contaminants that may be treated through in-situ bioremediation. Relationships between the reducing environment, available electron donors and acceptors, reaction kinetics, and microbial community composition must be further understood to successfully engineer remediation schemes in the complex subsurface environment. This thesis work investigated the effect of sulfate reduction on a dehalogenating culture grown under very controlled conditions. Two chemostats containing the Point Mugu (PM) culture were maintained using an influent containing tetrachloroethene (PCE) as an electron acceptor and lactate as a fermenting electron donor. One of these chemostats, PM-5L, was used as a control, while the influent to the PM-2L chemostat was amended with sulfate on an equal electron-equivalent basis to PCE. The effluent composition of these two chemostats was monitored over time, and periodic batch rate tests and molecular analyses were performed with cells harvested from the chemostat to elucidate the changes in performance and microbial composition within the chemostat culture. A numerical model based on Monod kinetics with competitive inhibition was developed to fit data from batch PCE-to-ethene rate tests by simultaneously solving for the k[subscript m]X parameters of each CAH dechlorination step given a standard set of Ks values. Non-linear regression of multi-equilibrium VC Monod test data provided the Monod parameters (k[subscript m]X and K[subscript s]) for VC dechlorination. These parameters were used to quantify changes in dechlorinating performance of each chemostat over time and compare the performance of the two chemostats. The effluent chemical composition of the PM-5L chemostat appeared to be steady after approximately six residence times, with 1120 μM PCE being transformed to 98% ethene and 2% VC, H₂ tensions remaining between 2-3 nM, acetate around 4.3 mM, and biomass around 23 mg protein/L. Batch rate tests during this time showed rapid rates of transformation for all CAHs, agreeing well with chemostat performance. The k[subscript m]X parameters derived from the PCE-to-ethene data and the pseudo-mixed order rate coefficient of VC dechlorination determined through multi-equilibrium VC Monod tests also remained essentially constant over the one-year period of study. Changes in the PM-2L chemostat performance following the initiation of sulfate reduction were observed. Sulfate reduction began almost immediately after its addition to the chemostat, and total sulfide concentrations rose to 100-300 μM. Chemostat performance with respect to CAH and H₂ concentrations was roughly steady over approximately 250 days, with PCE being dechlorinated to 9 μM cis-DCE, 230 μM VC, and 860 μM ethene under H₂ tensions around 4 nM. Total protein levels nearly doubled during this period, increasing from 25 to 47 mg protein/L. Sulfate reduction then rapidly increased to completion, resulting in 620-720 μM dissolved sulfide and a decrease in the H₂ concentration to 2 nM. At this time, the extent of PCE dechlorination also decreased to 280, 760, and 80 μM cis-DCE, VC, and ethene, respectively. Batch rate tests showed a decrease in all chlorinated ethene reduction rates; however, VC dechlorination was the most effected by sulfate reduction, showing a 97% reduction in rate following sulfate addition. Multi-equilibrium VC rate tests were impossible to conduct following the sharp increase in sulfate reduction in the chemostat due to lack of measurable dechlorination over a days' time. A simple chemostat model employed Monod kinetics for the series of CAH reactions to determine the steady-state extent of dechlorination in the chemostat predicted by the best-fit kinetic parameters of each PCE-to-ethene rate test. The extent of dechlorination was well-modeled for the PM-5L chemostat when a H₂ limitation factor of 0.3 was applied to the rate of each CAH. Using a dual Monod kinetic model with H₂ as the electron donor, the limitation factor corresponded with a half-velocity coefficient (K[subscript H]) of 4.6 nM. When this same K[subscript H] was used to model the PM-2L chemostat, a greater extent of dechlorination was predicted than what was observed in the chemostat, possibly suggesting other inhibitory factors of dechlorination were present in the PM-2L chemostat. DNA and RNA analyses of cells periodically harvested from the chemostat were performed by Ian P.G. Marshall at Stanford University. His work revealed shifts in the chemostats' Dehalococcoides population over time. Analysis of the PM-5L culture using the H2ase chip he developed and a clone library of hupL genes showed that the Dehalococcoides population was predominately related to strains BAV-1 or CBDB1/GT and did not undergo a significant shift over time. Clone libraries constructed for cells harvested from the PM-2L chemostat revealed two shifts in the chemostat Dehalococcoides population. A genetically homogenous strain relative of BAV-1 was eliminated following a decline in chemostat H2 tensions from around 27 nM to 2 nM and a corresponding increase in dehalogenation efficiency. In a second shift, a strain 195 relative outcompeted the CBDB1/GT relatives following enhanced sulfate reduction. A general decrease in the Dehalococcoides concentration within the chemostat culture was also suggested by qPCR analysis of Dehalococcoides 16S genes. These molecular results correlated well with the decline in VC reduction rates reported in batch kinetic tests given the characteristic co-metabolic VC reduction of the dominant strain 195 relative and overall lower concentrations of Dehalococcoides. Our work suggests that sulfate reduction in the anaerobic chemostat environment caused a shift in the dechlorinating microbial population to a strain with less efficient VC reduction. This shift was also accompanied by a decline in Dehalococcoides concentration within the culture. Both of these factors contributed to the decline in chlorinated ethene transformation rates observed through batch rate tests. Competition for H₂ was not expected to be the primary cause for the changes observed in the PM-2L chemostat. Long-term batch tests involving the control culture are proposed to elucidate whether sulfide or other factors of sulfate reduction are responsible for this shift, and to confirm the suspected role of H₂ competition between dechlorination and sulfate reduction in the chemostat-grown PM culture. Title: Kinetic and molecular effects of sulfate reduction on a dechlorinating culture under chemostat growth conditions Author: Berggren, Dusty Rose V. Chlorinated ethenes are common groundwater contaminants that may be treated through in-situ bioremediation. Relationships between the reducing environment, available electron donors and acceptors, reaction kinetics, and microbial community composition must be further understood to successfully engineer remediation schemes in the complex subsurface environment. This thesis work investigated the effect of sulfate reduction on a dehalogenating culture grown under very controlled conditions. Two chemostats containing the Point Mugu (PM) culture were maintained using an influent containing tetrachloroethene (PCE) as an electron acceptor and lactate as a fermenting electron donor. One of these chemostats, PM-5L, was used as a control, while the influent to the PM-2L chemostat was amended with sulfate on an equal electron-equivalent basis to PCE. The effluent composition of these two chemostats was monitored over time, and periodic batch rate tests and molecular analyses were performed with cells harvested from the chemostat to elucidate the changes in performance and microbial composition within the chemostat culture. A numerical model based on Monod kinetics with competitive inhibition was developed to fit data from batch PCE-to-ethene rate tests by simultaneously solving for the k[subscript m]X parameters of each CAH dechlorination step given a standard set of Ks values. Non-linear regression of multi-equilibrium VC Monod test data provided the Monod parameters (k[subscript m]X and K[subscript s]) for VC dechlorination. These parameters were used to quantify changes in dechlorinating performance of each chemostat over time and compare the performance of the two chemostats. The effluent chemical composition of the PM-5L chemostat appeared to be steady after approximately six residence times, with 1120 μM PCE being transformed to 98% ethene and 2% VC, H₂ tensions remaining between 2-3 nM, acetate around 4.3 mM, and biomass around 23 mg protein/L. Batch rate tests during this time showed rapid rates of transformation for all CAHs, agreeing well with chemostat performance. The k[subscript m]X parameters derived from the PCE-to-ethene data and the pseudo-mixed order rate coefficient of VC dechlorination determined through multi-equilibrium VC Monod tests also remained essentially constant over the one-year period of study. Changes in the PM-2L chemostat performance following the initiation of sulfate reduction were observed. Sulfate reduction began almost immediately after its addition to the chemostat, and total sulfide concentrations rose to 100-300 μM. Chemostat performance with respect to CAH and H₂ concentrations was roughly steady over approximately 250 days, with PCE being dechlorinated to 9 μM cis-DCE, 230 μM VC, and 860 μM ethene under H₂ tensions around 4 nM. Total protein levels nearly doubled during this period, increasing from 25 to 47 mg protein/L. Sulfate reduction then rapidly increased to completion, resulting in 620-720 μM dissolved sulfide and a decrease in the H₂ concentration to 2 nM. At this time, the extent of PCE dechlorination also decreased to 280, 760, and 80 μM cis-DCE, VC, and ethene, respectively. Batch rate tests showed a decrease in all chlorinated ethene reduction rates; however, VC dechlorination was the most effected by sulfate reduction, showing a 97% reduction in rate following sulfate addition. Multi-equilibrium VC rate tests were impossible to conduct following the sharp increase in sulfate reduction in the chemostat due to lack of measurable dechlorination over a days' time. A simple chemostat model employed Monod kinetics for the series of CAH reactions to determine the steady-state extent of dechlorination in the chemostat predicted by the best-fit kinetic parameters of each PCE-to-ethene rate test. The extent of dechlorination was well-modeled for the PM-5L chemostat when a H₂ limitation factor of 0.3 was applied to the rate of each CAH. Using a dual Monod kinetic model with H₂ as the electron donor, the limitation factor corresponded with a half-velocity coefficient (K[subscript H]) of 4.6 nM. When this same K[subscript H] was used to model the PM-2L chemostat, a greater extent of dechlorination was predicted than what was observed in the chemostat, possibly suggesting other inhibitory factors of dechlorination were present in the PM-2L chemostat. DNA and RNA analyses of cells periodically harvested from the chemostat were performed by Ian P.G. Marshall at Stanford University. His work revealed shifts in the chemostats' Dehalococcoides population over time. Analysis of the PM-5L culture using the H2ase chip he developed and a clone library of hupL genes showed that the Dehalococcoides population was predominately related to strains BAV-1 or CBDB1/GT and did not undergo a significant shift over time. Clone libraries constructed for cells harvested from the PM-2L chemostat revealed two shifts in the chemostat Dehalococcoides population. A genetically homogenous strain relative of BAV-1 was eliminated following a decline in chemostat H2 tensions from around 27 nM to 2 nM and a corresponding increase in dehalogenation efficiency. In a second shift, a strain 195 relative outcompeted the CBDB1/GT relatives following enhanced sulfate reduction. A general decrease in the Dehalococcoides concentration within the chemostat culture was also suggested by qPCR analysis of Dehalococcoides 16S genes. These molecular results correlated well with the decline in VC reduction rates reported in batch kinetic tests given the characteristic co-metabolic VC reduction of the dominant strain 195 relative and overall lower concentrations of Dehalococcoides. Our work suggests that sulfate reduction in the anaerobic chemostat environment caused a shift in the dechlorinating microbial population to a strain with less efficient VC reduction. This shift was also accompanied by a decline in Dehalococcoides concentration within the culture. Both of these factors contributed to the decline in chlorinated ethene transformation rates observed through batch rate tests. Competition for H₂ was not expected to be the primary cause for the changes observed in the PM-2L chemostat. Long-term batch tests involving the control culture are proposed to elucidate whether sulfide or other factors of sulfate reduction are responsible for this shift, and to confirm the suspected role of H₂ competition between dechlorination and sulfate reduction in the chemostat-grown PM culture. Close

• 3236. [Article] Science and efficacy of mild sodium hydroxide treatments in enzyme-based wheat straw-to-glucose processing

  The work described in this dissertation focused on chemistry related to the use of aqueous sodium hydroxide as a treatment in the processing of wheat straw. A major emphasis was the comprehensive evaluation ...

  Citation × Citation Citation Abstract Copy Title: Science and efficacy of mild sodium hydroxide treatments in enzyme-based wheat straw-to-glucose processing Author: Sophonputtanaphoca, Supaporn The work described in this dissertation focused on chemistry related to the use of aqueous sodium hydroxide as a treatment in the processing of wheat straw. A major emphasis was the comprehensive evaluation of straw component partitioning due to sodium hydroxide (NaOH) processing. This was evaluated over a range of NaOH concentrations (0­‐10%, w/v), all at 50°C, 5 h treatment period, and 3% solid loading. Solid and liquid phases resulting from NaOH treatments were evaluated. Total solids recovered in the NaOH­‐treated solid phase ranged from 47.4­‐88.0%. Overall carbohydrate recovery in the combined solid and liquid phases was negatively correlated with the alkali concentration of the treatment liquor. The glucan content of the NaOH‐treated solid phase ranged from 37.2­‐67.4%. Glucan recovery in the solid phase was relatively high in all cases, the minimum value being ~98%. Increasing amounts of xylan partitioned into the liquid phase as sodium hydroxide concentrations increased – it ranged from 31­‐83% of the xylan being recovered in the soluble phase. Carbohydrate analyses of the pretreated liquor revealed that the majority of carbohydrate loss from the solid fraction could be recovered in the liquid phase in form of oligomers and monomers due to alkaline degradation. The interconversion of glucose, fructose, and mannose under the alkaline condition played an important role in the presence of those sugars. Increase in NaOH concentration contributed to increase in amount of cellulose­‐derived and hemicellulose‐derived oligomers in the pretreated liquor. All oligomers except fructooligomers in NaOH pretreated liquor were higher than those found in water extraction at 50°C, 5 h. Total carbohydrate recovery from the solid and liquid fractions was as high as 99% for glucose and glucan in 5% NaOH treatment and 80‐95% for xylose and xylan in 1-­10% NaOH treatment. The presence of NaOH as extraction reagent dramatically induced lignin and ash removal from the pretreated solid with up to 63% acid insoluble lignin (AIL) and 87% ash extraction. Solid fractions resulting from NaOH pretreatments (up to 5% NaOH) were tested for their susceptibility to enzymatic saccharification using cellulase and cellulase/xylanase enzyme preparations. The cellulase/xylanase enzyme preparation was found to be more effective at cellulose saccharification than the cellulase enzyme preparation alone. Maximum glucose yield, which corresponded to the 5% NaOH treatment, was 82% over the standard 48 h saccharification period. Extended saccharifications times to 120 h showed that the conversion yield approached 90%. Sequential treatments of the straw (i.e. initial alkali treatment – first enzyme saccharification – second alkali treatment ‐ second enzyme treatment) revealed the NaOH treatment has the potential to render essentially all (~99%) of the straw glucan susceptible to enzyme saccharification. This suggests that the layered molecular arrangement of cellulose, hemicellulose, and lignin in the cell wall impacts biomass recalcitrance and glucan conversion yield. The other major focus of this dissertation research was the characterization of alkali neutralization, which occurs during the aqueous alkali processing of wheat straw. The approach taken was to evaluate the time course of alkali uptake and to determine the underlying nature of alkali uptake. The knowledge generated from this study is useful for understanding the nature of the alkali‐induced chemistry that is at the heart of alkali processing of agricultural byproducts, foods, and forest products. Alkali uptake/acid generation measurements were monitored for wheat straw suspensions at pH 11 and 30°C. The first phase of alkali uptake corresponded to the 30‐second time period over which the pH of the wheat straw suspension was adjusted from its original pH (~6.6) to pH 11. Alkali neutralization during this period was attributed to the instantaneous ionization of solvent accessible Bronstad acids. Following pH adjustment to 11.0, the time course of subsequent alkali uptake was recorded. The time course appeared biphasic. The early phase, which corresponded to the relatively rapid uptake of alkali, was evident during the first 24 hours. The later phase, which was characterized by the relatively slow uptake of alkali, was maintained for the length of the study (up to 96 hours). Alkali uptake during the early phase of the time course appears to be determined by the rate of hydrolysis of readily accessible esters – primarily acetic acid esters (acetyl groups). Alkali uptake during the later phase of the time course appears to be impacted by the rate of alkali penetration into the straw and the rate of production of alkali‐induced acid degradation products. The uptake of alkali in the pH adjustment phase was ~ 120 μEq per gram wheat straw, the uptake of alkali in the early phase of time course was ~ 1,064 μEq per gram wheat straw, and the rate of uptake in the later phase of the time course 6.10 μEq per gram wheat straw per hour. Amount of acetyl groups, ferulic acid, and p-­coumaric acid generated during 96-­h pretreatment revealed that they are major esters being hydrolyzed under the studied condition. Combined, these ester-­derived acids contributed up to ~ 28% of overall alkali uptake. In addition, alkaline degradation products quantified in this study showed additional ~ 28% contribution to the overall alkali uptake. Title: Science and efficacy of mild sodium hydroxide treatments in enzyme-based wheat straw-to-glucose processing Author: Sophonputtanaphoca, Supaporn The work described in this dissertation focused on chemistry related to the use of aqueous sodium hydroxide as a treatment in the processing of wheat straw. A major emphasis was the comprehensive evaluation of straw component partitioning due to sodium hydroxide (NaOH) processing. This was evaluated over a range of NaOH concentrations (0­‐10%, w/v), all at 50°C, 5 h treatment period, and 3% solid loading. Solid and liquid phases resulting from NaOH treatments were evaluated. Total solids recovered in the NaOH­‐treated solid phase ranged from 47.4­‐88.0%. Overall carbohydrate recovery in the combined solid and liquid phases was negatively correlated with the alkali concentration of the treatment liquor. The glucan content of the NaOH‐treated solid phase ranged from 37.2­‐67.4%. Glucan recovery in the solid phase was relatively high in all cases, the minimum value being ~98%. Increasing amounts of xylan partitioned into the liquid phase as sodium hydroxide concentrations increased – it ranged from 31­‐83% of the xylan being recovered in the soluble phase. Carbohydrate analyses of the pretreated liquor revealed that the majority of carbohydrate loss from the solid fraction could be recovered in the liquid phase in form of oligomers and monomers due to alkaline degradation. The interconversion of glucose, fructose, and mannose under the alkaline condition played an important role in the presence of those sugars. Increase in NaOH concentration contributed to increase in amount of cellulose­‐derived and hemicellulose‐derived oligomers in the pretreated liquor. All oligomers except fructooligomers in NaOH pretreated liquor were higher than those found in water extraction at 50°C, 5 h. Total carbohydrate recovery from the solid and liquid fractions was as high as 99% for glucose and glucan in 5% NaOH treatment and 80‐95% for xylose and xylan in 1-­10% NaOH treatment. The presence of NaOH as extraction reagent dramatically induced lignin and ash removal from the pretreated solid with up to 63% acid insoluble lignin (AIL) and 87% ash extraction. Solid fractions resulting from NaOH pretreatments (up to 5% NaOH) were tested for their susceptibility to enzymatic saccharification using cellulase and cellulase/xylanase enzyme preparations. The cellulase/xylanase enzyme preparation was found to be more effective at cellulose saccharification than the cellulase enzyme preparation alone. Maximum glucose yield, which corresponded to the 5% NaOH treatment, was 82% over the standard 48 h saccharification period. Extended saccharifications times to 120 h showed that the conversion yield approached 90%. Sequential treatments of the straw (i.e. initial alkali treatment – first enzyme saccharification – second alkali treatment ‐ second enzyme treatment) revealed the NaOH treatment has the potential to render essentially all (~99%) of the straw glucan susceptible to enzyme saccharification. This suggests that the layered molecular arrangement of cellulose, hemicellulose, and lignin in the cell wall impacts biomass recalcitrance and glucan conversion yield. The other major focus of this dissertation research was the characterization of alkali neutralization, which occurs during the aqueous alkali processing of wheat straw. The approach taken was to evaluate the time course of alkali uptake and to determine the underlying nature of alkali uptake. The knowledge generated from this study is useful for understanding the nature of the alkali‐induced chemistry that is at the heart of alkali processing of agricultural byproducts, foods, and forest products. Alkali uptake/acid generation measurements were monitored for wheat straw suspensions at pH 11 and 30°C. The first phase of alkali uptake corresponded to the 30‐second time period over which the pH of the wheat straw suspension was adjusted from its original pH (~6.6) to pH 11. Alkali neutralization during this period was attributed to the instantaneous ionization of solvent accessible Bronstad acids. Following pH adjustment to 11.0, the time course of subsequent alkali uptake was recorded. The time course appeared biphasic. The early phase, which corresponded to the relatively rapid uptake of alkali, was evident during the first 24 hours. The later phase, which was characterized by the relatively slow uptake of alkali, was maintained for the length of the study (up to 96 hours). Alkali uptake during the early phase of the time course appears to be determined by the rate of hydrolysis of readily accessible esters – primarily acetic acid esters (acetyl groups). Alkali uptake during the later phase of the time course appears to be impacted by the rate of alkali penetration into the straw and the rate of production of alkali‐induced acid degradation products. The uptake of alkali in the pH adjustment phase was ~ 120 μEq per gram wheat straw, the uptake of alkali in the early phase of time course was ~ 1,064 μEq per gram wheat straw, and the rate of uptake in the later phase of the time course 6.10 μEq per gram wheat straw per hour. Amount of acetyl groups, ferulic acid, and p-­coumaric acid generated during 96-­h pretreatment revealed that they are major esters being hydrolyzed under the studied condition. Combined, these ester-­derived acids contributed up to ~ 28% of overall alkali uptake. In addition, alkaline degradation products quantified in this study showed additional ~ 28% contribution to the overall alkali uptake. Close

• 3237. [Article] The effects of selected feeding programs on the reproductive performance and specific blood plasma chemistries of caged broiler breeder males and dwarf breeder females

  I. THE EFFECT OF FEEDING PROGRAMS ON REPRODUCTIVE TRAITS AND SELECTED BLOOD CHEMISTRIES OF CAGED BROILER BREEDER MALES: 1. FEED RESTRICTION LEVELS: The effects of five different feeding levels (136, 125, ...

  Citation × Citation Citation Abstract Copy Title: The effects of selected feeding programs on the reproductive performance and specific blood plasma chemistries of caged broiler breeder males and dwarf breeder females Author: Buckner, Robert E. I. THE EFFECT OF FEEDING PROGRAMS ON REPRODUCTIVE TRAITS AND SELECTED BLOOD CHEMISTRIES OF CAGED BROILER BREEDER MALES: 1. FEED RESTRICTION LEVELS: The effects of five different feeding levels (136, 125, 113, 102, and 91 g/male/day of a 13.1% crude protein (CP), 3167 kcal ME/kg feed) on the reproductive traits of adult broiler breeder males in cages were evaluated from 30 to 60 weeks of age. Individual body weights, semen volume, sperm cells per ejaculate, and fertilizing ability were measured at 30, 40, 50, and 60 weeks of age. To assist in measuring the effect of feed restriction on metabolism during these periods, average hematocrits, plasma cholesterol, total protein, and uric acid levels were analyzed. Significant (P<.05) reductions were observed in average body weights, semen volume, sperm cell numbers per ejaculate, testicular weights, and hematocrits with feed restriction at the 91 g compared to the 136 g feeding level. At 40 weeks of age the percentage of males producing semen was significantly reduced for the 102 and 91 g compared to the 136 g feeding level. Average plasma cholesterol levels were significantly increased at the 91 g compared to the 136 g feeding level, but no significant effect (P>.05) was observed in plasma total protein and uric acid levels. Correlation coefficients were negative for plasma cholesterol to body weights, sperm numbers per ejaculate, and testicular weights but positive for body weights to hematocrits and testicular weights. II. THE EFFECT OF FEEDING PROGRAMS ON REPRODUCTIVE TRAITS AND SELECTED BLOOD CHEMISTRIES OF CAGED BROILER BREEDER MALES: 2. FREE CHOICE, LOW ENERGY RATIONS OF 9, 7, AND 5 PERCENT CRUDE PROTEIN: Three isocaloric feeds consisting of 9, 7, and 5% CP each containing 2315 kcal ME/kg were fed free choice to individually caged broiler breeder males from 22 to 65 weeks of age. To obtain these protein and energy levels, sand was incorporated into the rations at levels ranging from 26.1 to 33.6% of the feed. The average daily protein intake per male was 18.7, 14.7, and 10.7 g for the 9, 7, and 5% CP, respectively, with an average caloric intake of 480 kcal ME. Individual body weights, semen characteristics, testicular weights, hematocrits, plasma cholesterol, total protein, uric acid and triglyceride levels were evaluated during each 4-week period from 24 to 44 weeks and also at 50 and 65 weeks of age. Average body weights from 24 to 32 weeks of age were significantly lower for the males fed 5% CP then those fed 9% CP rations. Semen volumes and sperm cell numbers per ejaculate were significantly reduced only at 24 weeks of age for males fed 5% CP feeds. At 65 weeks of age there were no significant differences among treatments means for body weight, sperm cells numbers per ejaculate, nor testicular weights. Plasma cholesterol levels of males fed the 5% CP diet were significantly higher at 28 and 32 weeks while levels of plasma total protein were decreased at 28 weeks of age. Plasma uric acid levels were consistently and significantly decreased as protein in take was reduced. The results suggest that within the parameters of this study broiler breeder males can be successfully maintained on low energy (2315 kcal/ kg) feeds of 7% CP from 22 to 40 weeks and then 5% CP from 40 to 65 weeks of age. III. INFLUENCES OF 16 AND 17.5 PERCENT DIETARY PROTEIN AND SUPPLEMENTAL METHIONINE ON THE REPRODUCTIVE TRAITS AND SELECTED PLASMA CHEMISTRIES OF CAGED DWARF (dw) BROILER BREEDER FEMALES: A total of 240 individually caged dwarf (dw) broiler breeder females were fed one of six isocaloric treatment diets (2740 kcal/kg) containing 16 or 17.5% CP to which d,l-methionine (MET) was supple mented at 0, .08, or .15% from 22 to 50 weeks of age. The resulting dietary MET levels ranged from .256 to .424% and total sulfur amino acids (TSAA) ranged from .527 to .718% of the rations. Treatment means were compared for body weights, mortality, total egg produc tion, egg weights, and egg fertility and hatchability resulting from artificial insemination with .05 ml broiler breeder semen. Blood plasma was analyzed at 32, 40, and 50 weeks of age for cholesterol, uric acid, total protein, and albumin as possible monitors of dietary protein utilization and of the physiological effects of protein and methionine levels on reproductive traits. There were no significant differences observed among dietary treatment means for body weights, mortality, total egg production, fertility, nor hatchability from 24 to 50 weeks of age. Significant differences were not observed among the treatment means for plasma cholesterol, total protein, and albumin at 32, 40, or 50 weeks of age. Between the two CP levels, plasma uric acid was significantly increased for the females on 17.5% compared to those on 16% CP rations. A consistent although nonsignificant trend of decreased uric acid levels with supplemental methionine was observed only in the females fed 16% CP diets and not in those on 17.5% CP. Under the conditions of this experiment, protein requirements for caged dwarf broiler breeder females from 22 to 50 weeks of age appear to be satisfied with daily dietary intakes of 20.3 g CP, 325 mg MET, and 670 mg TSAA. Title: The effects of selected feeding programs on the reproductive performance and specific blood plasma chemistries of caged broiler breeder males and dwarf breeder females Author: Buckner, Robert E. I. THE EFFECT OF FEEDING PROGRAMS ON REPRODUCTIVE TRAITS AND SELECTED BLOOD CHEMISTRIES OF CAGED BROILER BREEDER MALES: 1. FEED RESTRICTION LEVELS: The effects of five different feeding levels (136, 125, 113, 102, and 91 g/male/day of a 13.1% crude protein (CP), 3167 kcal ME/kg feed) on the reproductive traits of adult broiler breeder males in cages were evaluated from 30 to 60 weeks of age. Individual body weights, semen volume, sperm cells per ejaculate, and fertilizing ability were measured at 30, 40, 50, and 60 weeks of age. To assist in measuring the effect of feed restriction on metabolism during these periods, average hematocrits, plasma cholesterol, total protein, and uric acid levels were analyzed. Significant (P<.05) reductions were observed in average body weights, semen volume, sperm cell numbers per ejaculate, testicular weights, and hematocrits with feed restriction at the 91 g compared to the 136 g feeding level. At 40 weeks of age the percentage of males producing semen was significantly reduced for the 102 and 91 g compared to the 136 g feeding level. Average plasma cholesterol levels were significantly increased at the 91 g compared to the 136 g feeding level, but no significant effect (P>.05) was observed in plasma total protein and uric acid levels. Correlation coefficients were negative for plasma cholesterol to body weights, sperm numbers per ejaculate, and testicular weights but positive for body weights to hematocrits and testicular weights. II. THE EFFECT OF FEEDING PROGRAMS ON REPRODUCTIVE TRAITS AND SELECTED BLOOD CHEMISTRIES OF CAGED BROILER BREEDER MALES: 2. FREE CHOICE, LOW ENERGY RATIONS OF 9, 7, AND 5 PERCENT CRUDE PROTEIN: Three isocaloric feeds consisting of 9, 7, and 5% CP each containing 2315 kcal ME/kg were fed free choice to individually caged broiler breeder males from 22 to 65 weeks of age. To obtain these protein and energy levels, sand was incorporated into the rations at levels ranging from 26.1 to 33.6% of the feed. The average daily protein intake per male was 18.7, 14.7, and 10.7 g for the 9, 7, and 5% CP, respectively, with an average caloric intake of 480 kcal ME. Individual body weights, semen characteristics, testicular weights, hematocrits, plasma cholesterol, total protein, uric acid and triglyceride levels were evaluated during each 4-week period from 24 to 44 weeks and also at 50 and 65 weeks of age. Average body weights from 24 to 32 weeks of age were significantly lower for the males fed 5% CP then those fed 9% CP rations. Semen volumes and sperm cell numbers per ejaculate were significantly reduced only at 24 weeks of age for males fed 5% CP feeds. At 65 weeks of age there were no significant differences among treatments means for body weight, sperm cells numbers per ejaculate, nor testicular weights. Plasma cholesterol levels of males fed the 5% CP diet were significantly higher at 28 and 32 weeks while levels of plasma total protein were decreased at 28 weeks of age. Plasma uric acid levels were consistently and significantly decreased as protein in take was reduced. The results suggest that within the parameters of this study broiler breeder males can be successfully maintained on low energy (2315 kcal/ kg) feeds of 7% CP from 22 to 40 weeks and then 5% CP from 40 to 65 weeks of age. III. INFLUENCES OF 16 AND 17.5 PERCENT DIETARY PROTEIN AND SUPPLEMENTAL METHIONINE ON THE REPRODUCTIVE TRAITS AND SELECTED PLASMA CHEMISTRIES OF CAGED DWARF (dw) BROILER BREEDER FEMALES: A total of 240 individually caged dwarf (dw) broiler breeder females were fed one of six isocaloric treatment diets (2740 kcal/kg) containing 16 or 17.5% CP to which d,l-methionine (MET) was supple mented at 0, .08, or .15% from 22 to 50 weeks of age. The resulting dietary MET levels ranged from .256 to .424% and total sulfur amino acids (TSAA) ranged from .527 to .718% of the rations. Treatment means were compared for body weights, mortality, total egg produc tion, egg weights, and egg fertility and hatchability resulting from artificial insemination with .05 ml broiler breeder semen. Blood plasma was analyzed at 32, 40, and 50 weeks of age for cholesterol, uric acid, total protein, and albumin as possible monitors of dietary protein utilization and of the physiological effects of protein and methionine levels on reproductive traits. There were no significant differences observed among dietary treatment means for body weights, mortality, total egg production, fertility, nor hatchability from 24 to 50 weeks of age. Significant differences were not observed among the treatment means for plasma cholesterol, total protein, and albumin at 32, 40, or 50 weeks of age. Between the two CP levels, plasma uric acid was significantly increased for the females on 17.5% compared to those on 16% CP rations. A consistent although nonsignificant trend of decreased uric acid levels with supplemental methionine was observed only in the females fed 16% CP diets and not in those on 17.5% CP. Under the conditions of this experiment, protein requirements for caged dwarf broiler breeder females from 22 to 50 weeks of age appear to be satisfied with daily dietary intakes of 20.3 g CP, 325 mg MET, and 670 mg TSAA. Close

• 3238. [Article] Addressing wastewater epidemiology limitations with the use of dynamic population surrogates, complementary urinalyses and in-situ experiments

  Wastewater epidemiology is an emerging discipline that requires collaborative research involving analytical chemists, drug epidemiologists, and wastewater engineers. Wastewater epidemiology involves the ...

  Citation × Citation Citation Abstract Copy Title: Addressing wastewater epidemiology limitations with the use of dynamic population surrogates, complementary urinalyses and in-situ experiments Author: Brewer, Alex J. Wastewater epidemiology is an emerging discipline that requires collaborative research involving analytical chemists, drug epidemiologists, and wastewater engineers. Wastewater epidemiology involves the sampling and quantitative analysis of raw wastewaters from communities for illicit drugs and their metabolites. Mass loads (mass per day) and per capita (mg per day per person) are then calculated from concentrations and indicate the approximate quantity of illicit drugs used and excreted by the community. Limitations to wastewater epidemiology include that the population served by wastewater treatment plants within a day and between days is not well known. In addition, biodegradation of illicit drugs during transit in sewers may affect the concentration and mass flows that reach wastewater treatment plants. This thesis describes a series of studies conducted by an international collaboration between scientists and engineers from the United States and Switzerland to answer these two limitations. The experimental approaches for these studies used included high-frequency wastewater sampling strategies, the use of creatinine as a human urinary biomarker, as well as the use of unique locations as test sites including an open community, a prison in the state of Oregon, and a 5 km section of sewer in Zürich Switzerland. In Chapter 2, the diurnal study on the mass flows of illicit drugs or metabolites was formed over four days in a municipality with a population of approximately 55,000 people. The diurnal trends in illicit substances vary by substance. The high (g/day) mass flows of caffeine, methamphetamine, and creatinine indicate that lower-frequency sampling (approximately one sample per h) may representatively capture the use and excretion of these substances. However, lower and episodic mass flows of cocaine and its primary human metabolite, benzoylecgonine, indicate that higher-frequency is needed to accurately assess the use of the cocaine within the municipality. Normalization of illicit substances to creatinine gave between-day trends in illicit and legal substances that differed from non-normalized trends. Resident use of cocaine and methamphetamine were indicated by normalized mass flows that increased during early morning hours while commuters are largely absent from the community. Chapter 3 describes a series of experiments conducted at an Oregon state prison. The prison setting provided a unique opportunity to study a nearly-fixed population of individuals and their corresponding mass flows of illicit substances, the number of doses per person consumed, as well as an opportunity to quantify the level of agreement between numbers of individuals and the measured mass flows of creatinine. Methamphetamine use was more prevalent than cocaine/benzoylecgonine in the prison over the one month study in which single daily (24 h) composite samples of wastewater were collected. The hypothesis that the mass flows of methamphetamine and cocaine would be lower on days on which random urinalysis testing (RUA) is typically conducted by the prison (Monday-Thursday) was rejected. While the mass flows (mg/d) of methamphetamine were less than those for a nearby open community, the number of estimated doses per person was higher for the prison population. A higher number of positive RUA results were obtained for methamphetamine while none were positive for cocaine, which is consistent with the data obtained from wastewater. The hourly (diurnal) trend in methamphetamine mass loads indicated continual methamphetamine use/excretion inside the prison while cocaine and benzoylecgonine were detected in five hourly composite samples. Use of methamphetamine and cocaine by inmates could not be unambiguously distinguished from that of non-inmates (employees and visitors). The observed diurnal trends in creatinine mass loads were similar to those of an open community and are indicative of the general pattern of human wakefulness/activity. Predicted creatinine mass loads based on the total prison (inmates + non-inmates) were in good agreement with the measured mass loads, which indicates the potential use of creatinine as a quantitative population indicator. Additional research on the biodegradability of creatinine is needed because the prison setting was deliberately selected to minimize the potential for creatinine biodegradation. Chapter 4 addresses the data gap that exists on illicit drug transformation during in situ transit in sewers. The rates of in situ biodegradation have not yet been determined for conditions that are relevant to sewers, which include low to variable oxygen concentrations, the presence of a biofilm, and temperatures ≤ 20 °C. For this reason, two tracer tests were conducted in a 5 km stretch of sewer located near Zürich, Switzerland. The stable-isotope forms (deuterated) of cocaine and benzoylecgonine were injected into flowing wastewater and three locations up to 5 km downstream were sampled over time. Breakthrough curves were constructed from measurements of cocaine-d3 and benzoylecgonine-d3 concentration with time. The area under the curve (mass) was determined by integrating concentration over time. Benzoylecgonine-d3 was present in the injectate that should have only contained cocaine-d3; because the benzoylecgonine-d3 formation prior to injection is not known. The injected mass of cocaine-d3 did not decline over the 5 km distance. The observed mass of cocaine-d3 at 5 km was 10% greater than at 500 m, which indicates that the transformation of cocaine was not significant over the 1.5 h experiment. At 5 km downgradient, the apparent mass of benzoylecgonine-d3 had increased by 35% over that observed at 500 m. However, the apparent increase in benzoylecgonine-d3 mass was not accompanied by a corresponding loss of cocaine-d3. While uncertainty is apparent in the increase of both cocaine-d3 and benzoylecgonine-d3, the ratio of cocaine-d3/benzoylecgonine-d3 is subject only to analytical error because any errors associated with sampling and the integration of masses cancel out. The ratio of cocaine-d3/benzoylecgonine-d3 declined from 2.98 in the injectate to 1.66 at Location 3, which indicated a greater increase in benzoylecgonine-d3 relative to cocaine over the 5 km distance. Due to the benzoylecgonine-d3 that was present in the injectate, any biodegradation of cocaine-c3 to form benzoylecgonine-d3 could not be unambiguously distinguished. During the second tracer test in which benzoylecgonine-d3 was injected, the mass of benzoylecgonine-d3 did not significantly decline, which suggests that the apparent loss of benzoylecgonine-d3 during the cocaine-d3 test cannot be attributed to in-situ biodegradation. Overall, while uncertainty exists about the integrated masses for cocaine-d3 and benzoylecgonine-d3, the 5 km distance was too short in order to observe a significant loss of cocaine-d3 and formation of benzoylecgonine-d3. Recommendations for future research include conducting analysis on the injectate solution to ensure that only cocaine-d3 is introduced so that any formation of benzoylecgonine-d3 is readily apparent and quantifiable. In addition, the tracer tests should be repeated in a longer section of sewer to increase the residence time beyond 1.5 hr and degradation products of benzoylecgonine-d3 should be monitored including ecgonine and ecgonine methyl ester. Title: Addressing wastewater epidemiology limitations with the use of dynamic population surrogates, complementary urinalyses and in-situ experiments Author: Brewer, Alex J. Wastewater epidemiology is an emerging discipline that requires collaborative research involving analytical chemists, drug epidemiologists, and wastewater engineers. Wastewater epidemiology involves the sampling and quantitative analysis of raw wastewaters from communities for illicit drugs and their metabolites. Mass loads (mass per day) and per capita (mg per day per person) are then calculated from concentrations and indicate the approximate quantity of illicit drugs used and excreted by the community. Limitations to wastewater epidemiology include that the population served by wastewater treatment plants within a day and between days is not well known. In addition, biodegradation of illicit drugs during transit in sewers may affect the concentration and mass flows that reach wastewater treatment plants. This thesis describes a series of studies conducted by an international collaboration between scientists and engineers from the United States and Switzerland to answer these two limitations. The experimental approaches for these studies used included high-frequency wastewater sampling strategies, the use of creatinine as a human urinary biomarker, as well as the use of unique locations as test sites including an open community, a prison in the state of Oregon, and a 5 km section of sewer in Zürich Switzerland. In Chapter 2, the diurnal study on the mass flows of illicit drugs or metabolites was formed over four days in a municipality with a population of approximately 55,000 people. The diurnal trends in illicit substances vary by substance. The high (g/day) mass flows of caffeine, methamphetamine, and creatinine indicate that lower-frequency sampling (approximately one sample per h) may representatively capture the use and excretion of these substances. However, lower and episodic mass flows of cocaine and its primary human metabolite, benzoylecgonine, indicate that higher-frequency is needed to accurately assess the use of the cocaine within the municipality. Normalization of illicit substances to creatinine gave between-day trends in illicit and legal substances that differed from non-normalized trends. Resident use of cocaine and methamphetamine were indicated by normalized mass flows that increased during early morning hours while commuters are largely absent from the community. Chapter 3 describes a series of experiments conducted at an Oregon state prison. The prison setting provided a unique opportunity to study a nearly-fixed population of individuals and their corresponding mass flows of illicit substances, the number of doses per person consumed, as well as an opportunity to quantify the level of agreement between numbers of individuals and the measured mass flows of creatinine. Methamphetamine use was more prevalent than cocaine/benzoylecgonine in the prison over the one month study in which single daily (24 h) composite samples of wastewater were collected. The hypothesis that the mass flows of methamphetamine and cocaine would be lower on days on which random urinalysis testing (RUA) is typically conducted by the prison (Monday-Thursday) was rejected. While the mass flows (mg/d) of methamphetamine were less than those for a nearby open community, the number of estimated doses per person was higher for the prison population. A higher number of positive RUA results were obtained for methamphetamine while none were positive for cocaine, which is consistent with the data obtained from wastewater. The hourly (diurnal) trend in methamphetamine mass loads indicated continual methamphetamine use/excretion inside the prison while cocaine and benzoylecgonine were detected in five hourly composite samples. Use of methamphetamine and cocaine by inmates could not be unambiguously distinguished from that of non-inmates (employees and visitors). The observed diurnal trends in creatinine mass loads were similar to those of an open community and are indicative of the general pattern of human wakefulness/activity. Predicted creatinine mass loads based on the total prison (inmates + non-inmates) were in good agreement with the measured mass loads, which indicates the potential use of creatinine as a quantitative population indicator. Additional research on the biodegradability of creatinine is needed because the prison setting was deliberately selected to minimize the potential for creatinine biodegradation. Chapter 4 addresses the data gap that exists on illicit drug transformation during in situ transit in sewers. The rates of in situ biodegradation have not yet been determined for conditions that are relevant to sewers, which include low to variable oxygen concentrations, the presence of a biofilm, and temperatures ≤ 20 °C. For this reason, two tracer tests were conducted in a 5 km stretch of sewer located near Zürich, Switzerland. The stable-isotope forms (deuterated) of cocaine and benzoylecgonine were injected into flowing wastewater and three locations up to 5 km downstream were sampled over time. Breakthrough curves were constructed from measurements of cocaine-d3 and benzoylecgonine-d3 concentration with time. The area under the curve (mass) was determined by integrating concentration over time. Benzoylecgonine-d3 was present in the injectate that should have only contained cocaine-d3; because the benzoylecgonine-d3 formation prior to injection is not known. The injected mass of cocaine-d3 did not decline over the 5 km distance. The observed mass of cocaine-d3 at 5 km was 10% greater than at 500 m, which indicates that the transformation of cocaine was not significant over the 1.5 h experiment. At 5 km downgradient, the apparent mass of benzoylecgonine-d3 had increased by 35% over that observed at 500 m. However, the apparent increase in benzoylecgonine-d3 mass was not accompanied by a corresponding loss of cocaine-d3. While uncertainty is apparent in the increase of both cocaine-d3 and benzoylecgonine-d3, the ratio of cocaine-d3/benzoylecgonine-d3 is subject only to analytical error because any errors associated with sampling and the integration of masses cancel out. The ratio of cocaine-d3/benzoylecgonine-d3 declined from 2.98 in the injectate to 1.66 at Location 3, which indicated a greater increase in benzoylecgonine-d3 relative to cocaine over the 5 km distance. Due to the benzoylecgonine-d3 that was present in the injectate, any biodegradation of cocaine-c3 to form benzoylecgonine-d3 could not be unambiguously distinguished. During the second tracer test in which benzoylecgonine-d3 was injected, the mass of benzoylecgonine-d3 did not significantly decline, which suggests that the apparent loss of benzoylecgonine-d3 during the cocaine-d3 test cannot be attributed to in-situ biodegradation. Overall, while uncertainty exists about the integrated masses for cocaine-d3 and benzoylecgonine-d3, the 5 km distance was too short in order to observe a significant loss of cocaine-d3 and formation of benzoylecgonine-d3. Recommendations for future research include conducting analysis on the injectate solution to ensure that only cocaine-d3 is introduced so that any formation of benzoylecgonine-d3 is readily apparent and quantifiable. In addition, the tracer tests should be repeated in a longer section of sewer to increase the residence time beyond 1.5 hr and degradation products of benzoylecgonine-d3 should be monitored including ecgonine and ecgonine methyl ester. Close

• 3239. [Article] Coral-reef fishes : insights into larval dispersal and invasion ecology

  Because many coral-reef fishes are observable in situ, are amenable to transplantation, have small home ranges and short generation times, they provide a excellent system to investigate many topics within ...

  Citation × Citation Citation Abstract Copy Title: Coral-reef fishes : insights into larval dispersal and invasion ecology Author: Pusack, Timothy J. Because many coral-reef fishes are observable in situ, are amenable to transplantation, have small home ranges and short generation times, they provide a excellent system to investigate many topics within general ecology, fisheries biology, and conservation biology. The primary goal of this dissertation was to use the coral-reef fishes system to investigate two pressing sets of issues that face marine ecologists and managers of living marine resources. The first topic is the spatial and temporal patterns of larval dispersal and reproductive success in a marine metapopulation (Chapter 2). Because miniscule larvae are difficult to track in the vast pelagic environment, little is known about the patterns of larval dispersal. Yet, the more that is understood about the spatial and temporal variability in larval dispersal, the easier it will be to identify sites that are self-sustaining and exporting larvae to unprotect sites, a common goal of marine reserves. Incorporating this information into siting of marine reserves will improve their effectiveness. The second topic is fundamental in the ecology of biological invasions: species specific interactions between an invasive predator and native species. Specifically, I investigated the ability of an invasive predator to disrupt natural population regulation of a native prey species (Chapter 3), and the ability of a native predator to provide biotic resistance against the invasive species (Chapter 4). Because management of the lionfish invasion is needed throughout the Caribbean and Atlantic waters, it is necessary to understand not only how lionfish can change the native system, but also potential ways to moderate the negative effects. To address the first topic, we collected a total of 3,278 genetic tissue samples from bicolor damselfish (Stegastes partitus) over a four year period from reefs near four islands that encompass Exuma Sound, Bahamas (Chapter 2). Using a Bayesian parentage analysis, eight parent-offspring pairs were detected, which directly documented both connectivity between and self-recruitment on an ecological time scale. Remarkably, some larvae returned to the exact same reef where they were spawned, while others traveled to sites greater than 100 km distance. The only study island without a detected parent-offspring pair, Lee Stocking Island, was also the island that showed the most restrictive gene flow on evolutionary time scales. Additionally, variability was documented in the spatial and temporal signatures of sweepstakes reproduction and Wahlund effects. The variation we observed may be influenced by seasonal mesoscale gyres present in Exuma Sound, which play a prominent role in shaping local oceanographic patterns. Understanding how to identify pathways of larval dispersal is important to designing networks of marine reserves, because a common goal of reserves is to protect populations that are self-seeding and can export larvae. Thus, this research not only demonstrates that temporal variability is a prominent characteristic of larval dispersal, but provides an example of how to identify these populations. To address the second topic, two studies using both lab observations and manipulative field experiments were conducted to study the interaction between invasive Indo-Pacific red lionfish (Pterois volitans) and two different native species. In the first study (Chapter 3) three different data sets were used to document the effect of lionfish predation on their top prey species, the bridled goby (Coryphopterus glaucofraenum). The first data set was extracted from three previous studies to compare the change in abundance of bridled goby between patch reefs with lionfish and patch reefs that were predator-free. The second data set came from laboratory feeding trials to test for the presence of a size refuge for bridled goby caused by lionfish gape limitation. The third data set came from a manipulative field experiment using 22 nearshore reefs where the per capita mortality of bridled gobies was compared among four orthogonal predator treatments: (1) predator-free control, (2) a single native predator only -- the graysby grouper (Cephalopholis cruentatus) -- representing the pre-invasion system, (3) a single lionfish only, and (4) one native grouper and one lionfish, representing the invaded system. The combined results from these three data sets demonstrated that lionfish can consistently consume a significant amount and an extremely high proportion of bridled goby on small patch reefs. While small lionfish cannot eat bridled goby larger than 0.42 times their body size, large lionfish eat virtually the entire size range of bridled goby. These findings indicate that lionfish have the potential to extirpate local goby populations. In the second experiment, lionfish were exposed to different abundances of a native grouper, the Nassau grouper (Epinephelus striatus), on 28 nearshore patch reefs in the Bahamas. Lionfish persistence and growth was monitored over 10 weeks, as well as the abundance of small, < 5 cm total length (TL), native reef fishes. Nassau grouper did not affect lionfish persistence or growth. Yet, reefs with a lionfish and many Nassau grouper saw a significant increase in the abundance of small reef fishes compared to reefs with only lionfish or lionfish with few grouper. Thus, it appears that high densities of Nassau grouper provide some biotic resistance on Bahamian patch reefs by potentially interfering with lionfish predation on native fishes, perhaps by interrupting stalking behavior. This dissertation highlights the flexibility of the coral-reef fishes system for investigation of broadly different topics in basic and applied ecology. Coral-reef fishes are a tractable system for both large and small scale studies, as well as laboratory observations and manipulative field experiments. The findings of this dissertation advance the understanding of metapopulation dynamics and have implications for fisheries management and marine reserve design. This research also documents further evidence of the negative effects of lionfish, yet provides some promising findings that may indicate some native biotic resistance to this devastating of marine invasions. Title: Coral-reef fishes : insights into larval dispersal and invasion ecology Author: Pusack, Timothy J. Because many coral-reef fishes are observable in situ, are amenable to transplantation, have small home ranges and short generation times, they provide a excellent system to investigate many topics within general ecology, fisheries biology, and conservation biology. The primary goal of this dissertation was to use the coral-reef fishes system to investigate two pressing sets of issues that face marine ecologists and managers of living marine resources. The first topic is the spatial and temporal patterns of larval dispersal and reproductive success in a marine metapopulation (Chapter 2). Because miniscule larvae are difficult to track in the vast pelagic environment, little is known about the patterns of larval dispersal. Yet, the more that is understood about the spatial and temporal variability in larval dispersal, the easier it will be to identify sites that are self-sustaining and exporting larvae to unprotect sites, a common goal of marine reserves. Incorporating this information into siting of marine reserves will improve their effectiveness. The second topic is fundamental in the ecology of biological invasions: species specific interactions between an invasive predator and native species. Specifically, I investigated the ability of an invasive predator to disrupt natural population regulation of a native prey species (Chapter 3), and the ability of a native predator to provide biotic resistance against the invasive species (Chapter 4). Because management of the lionfish invasion is needed throughout the Caribbean and Atlantic waters, it is necessary to understand not only how lionfish can change the native system, but also potential ways to moderate the negative effects. To address the first topic, we collected a total of 3,278 genetic tissue samples from bicolor damselfish (Stegastes partitus) over a four year period from reefs near four islands that encompass Exuma Sound, Bahamas (Chapter 2). Using a Bayesian parentage analysis, eight parent-offspring pairs were detected, which directly documented both connectivity between and self-recruitment on an ecological time scale. Remarkably, some larvae returned to the exact same reef where they were spawned, while others traveled to sites greater than 100 km distance. The only study island without a detected parent-offspring pair, Lee Stocking Island, was also the island that showed the most restrictive gene flow on evolutionary time scales. Additionally, variability was documented in the spatial and temporal signatures of sweepstakes reproduction and Wahlund effects. The variation we observed may be influenced by seasonal mesoscale gyres present in Exuma Sound, which play a prominent role in shaping local oceanographic patterns. Understanding how to identify pathways of larval dispersal is important to designing networks of marine reserves, because a common goal of reserves is to protect populations that are self-seeding and can export larvae. Thus, this research not only demonstrates that temporal variability is a prominent characteristic of larval dispersal, but provides an example of how to identify these populations. To address the second topic, two studies using both lab observations and manipulative field experiments were conducted to study the interaction between invasive Indo-Pacific red lionfish (Pterois volitans) and two different native species. In the first study (Chapter 3) three different data sets were used to document the effect of lionfish predation on their top prey species, the bridled goby (Coryphopterus glaucofraenum). The first data set was extracted from three previous studies to compare the change in abundance of bridled goby between patch reefs with lionfish and patch reefs that were predator-free. The second data set came from laboratory feeding trials to test for the presence of a size refuge for bridled goby caused by lionfish gape limitation. The third data set came from a manipulative field experiment using 22 nearshore reefs where the per capita mortality of bridled gobies was compared among four orthogonal predator treatments: (1) predator-free control, (2) a single native predator only -- the graysby grouper (Cephalopholis cruentatus) -- representing the pre-invasion system, (3) a single lionfish only, and (4) one native grouper and one lionfish, representing the invaded system. The combined results from these three data sets demonstrated that lionfish can consistently consume a significant amount and an extremely high proportion of bridled goby on small patch reefs. While small lionfish cannot eat bridled goby larger than 0.42 times their body size, large lionfish eat virtually the entire size range of bridled goby. These findings indicate that lionfish have the potential to extirpate local goby populations. In the second experiment, lionfish were exposed to different abundances of a native grouper, the Nassau grouper (Epinephelus striatus), on 28 nearshore patch reefs in the Bahamas. Lionfish persistence and growth was monitored over 10 weeks, as well as the abundance of small, < 5 cm total length (TL), native reef fishes. Nassau grouper did not affect lionfish persistence or growth. Yet, reefs with a lionfish and many Nassau grouper saw a significant increase in the abundance of small reef fishes compared to reefs with only lionfish or lionfish with few grouper. Thus, it appears that high densities of Nassau grouper provide some biotic resistance on Bahamian patch reefs by potentially interfering with lionfish predation on native fishes, perhaps by interrupting stalking behavior. This dissertation highlights the flexibility of the coral-reef fishes system for investigation of broadly different topics in basic and applied ecology. Coral-reef fishes are a tractable system for both large and small scale studies, as well as laboratory observations and manipulative field experiments. The findings of this dissertation advance the understanding of metapopulation dynamics and have implications for fisheries management and marine reserve design. This research also documents further evidence of the negative effects of lionfish, yet provides some promising findings that may indicate some native biotic resistance to this devastating of marine invasions. Close

• 3240. [Article] Population Genetics of American Pika (Ochotona princeps) : Investigating Gene Flow and Genetic Diversity Across Multiple, Complex Landscapes

  American pikas (Ochotona princeps) are considered an indicator species of climate change. Adaptations for cold climates and active winters make pikas particularly sensitive to increasing temperatures. ...

  Citation × Citation Citation Abstract Copy Title: Population Genetics of American Pika (Ochotona princeps) : Investigating Gene Flow and Genetic Diversity Across Multiple, Complex Landscapes Author: Castillo, Jessica A. American pikas (Ochotona princeps) are considered an indicator species of climate change. Adaptations for cold climates and active winters make pikas particularly sensitive to increasing temperatures. This, combined with evidence that multiple historically occupied populations have been extirpated within the past century, contributed to American pikas becoming a focal species for climate change research. This dissertation is primarily the product of a collaboration among the National Parks Service and multiple academic institutions whose principal goal was to assess the vulnerability of the American pika within eight national parks representing a variety of habitat types and environmental conditions. While much of the previous climate change-related research on American pikas has focused on how climate affects occupancy, this work sought to establish how landscape characteristics and climate may interact and influence dispersal, population connectivity, and ultimately population vulnerability or resilience. I addressed these questions using population and landscape genetics approaches within nine national parks, two national wildlife refuges, and a national forest. American pikas represent one of the best described mammalian metapopulation systems. They are restricted to rocky habitats, such as talus and lava, which are frequently patchily distributed on the landscape, both at the local and regional scales. Functional connectivity, the combination of habitat configuration and an organism’s ability to move through the landscape, is essential for maintaining robust genetic populations. This is particularly true within metapopulations which, by definition, rely on dispersal of individuals among habitat patches for overall long-term stability. In Chapters 1 and 2, I used a landscape genetics approach to identify features that either promote or inhibit dispersal of American pikas. There has been significant debate over the past few years about the best statistical approaches within landscape genetics and the appropriateness of certain methods in particular. In Chapter 1, I used Crater Lake National Park as a test case to evaluate the effectiveness of Mantel and partial Mantel tests in a causal modeling framework to correctly identify underlying landscape variables (e.g., topography, aspect, water barriers) and their effect on gene flow. I demonstrated, through simulations, that this approach was able to correctly identify the landscape variables, but not precisely the magnitude of their effect on gene flow. I concluded that while results need to be interpreted with caution, this method was effective for identifying which variables are important in shaping functional connectivity. In Chapter 2, I applied the causal modeling approach from Chapter 1 to seven additional study sites. I identified a general trend that south-west facing aspects pose greater resistance to dispersal than north and east-facing aspects, suggesting that exposure to high temperatures may limit dispersal in American pikas. I also found that amount and configuration of habitat (i.e., rocky substrate) influenced the degree to which other landscape variables impact dispersal. I then applied the models of landscape resistance to investigate habitat patch connectivity within each site using a graph theoretic approach. This allowed me to assess overall patch connectivity, as well as identify specific patches and areas within study sites that are particularly important for maintaining functional connectivity, or in contrast, at risk of becoming isolated. Functional connectivity is an important component when assessing population vulnerability because it describes the ability of genetic material to flow through the landscape. A loss of gene flow can lead to isolation of population segments, reduction of effective population size, loss of genetic diversity and subsequent erosion of evolutionary potential, and ultimately metapopulation collapse. However, other factors including habitat area and quality, affect effective population size and population stability. Genetic diversity is the material upon which evolution acts. In the face of rapid environmental change, species must either shift their range in order to track their ecological niche, or adapt in situ to the new environment. The former strategy requires sufficient available habitat as well as dispersal ability, which is unlikely for American pikas. Genetic diversity, therefore, is an important indicator of long-term population viability. In Chapter 3, I quantified genetic diversity within thirteen study sites and investigated the relationship between genetic diversity and environmental variables related to climate as well as habitat configuration and quality. As expected, habitat area was important at both the local and regional scales. Temperature was also a significant predictor of genetic diversity, with hotter sites having lower genetic diversity. However, and somewhat surprisingly, precipitation was a better predictor of genetic diversity than temperature, with sites receiving moderate levels of precipitation having higher genetic diversity. I also compared population differentiation among study sites and identified sites that are particularly distinct. In the process of the above analyses, I also identified a previously undescribed contact zone between the northern and southern Rocky Mountain genetic lineages, recognized as separate subspecies, within Rocky Mountain National Park. Climate change is predicted to affect habitat quality and landscape permeability, both of which have detectable genetic consequences. In Chapter 4, I quantified genetic diversity and structure within two study sites, Yosemite and Lassen Volcanic National Parks, from two sampling periods separated by approximately a century in time. I extracted DNA from historic study skins housed in the University of California Berkeley Museum of Vertebrate Zoology (MVZ) and compared multilocus microsatellite genotypes to those generated from modern fecal and tissue samples. Additionally, the historic survey data, including specimens, field journals, and other artifacts, were used in a series of systematic resurveys and subsequent species distribution modelling by other researchers. The nature of national parks as protected areas affords the opportunity to isolate the effects of climate change to a greater degree than other areas that have also experienced significant land use changes. I found no evidence for a change in genetic diversity within these two sites, consistent with observations from other studies that occupancy has remained relatively stable. I did find some evidence suggesting increasing population differentiation, potentially as a result of eroding landscape permeability. These results provide an important baseline for comparison with other sites, as well as reference points for future genetic monitoring of these populations. In Chapter 5, I provide general synthesis and conclusions, as well as considerations for future research. Title: Population Genetics of American Pika (Ochotona princeps) : Investigating Gene Flow and Genetic Diversity Across Multiple, Complex Landscapes Author: Castillo, Jessica A. American pikas (Ochotona princeps) are considered an indicator species of climate change. Adaptations for cold climates and active winters make pikas particularly sensitive to increasing temperatures. This, combined with evidence that multiple historically occupied populations have been extirpated within the past century, contributed to American pikas becoming a focal species for climate change research. This dissertation is primarily the product of a collaboration among the National Parks Service and multiple academic institutions whose principal goal was to assess the vulnerability of the American pika within eight national parks representing a variety of habitat types and environmental conditions. While much of the previous climate change-related research on American pikas has focused on how climate affects occupancy, this work sought to establish how landscape characteristics and climate may interact and influence dispersal, population connectivity, and ultimately population vulnerability or resilience. I addressed these questions using population and landscape genetics approaches within nine national parks, two national wildlife refuges, and a national forest. American pikas represent one of the best described mammalian metapopulation systems. They are restricted to rocky habitats, such as talus and lava, which are frequently patchily distributed on the landscape, both at the local and regional scales. Functional connectivity, the combination of habitat configuration and an organism’s ability to move through the landscape, is essential for maintaining robust genetic populations. This is particularly true within metapopulations which, by definition, rely on dispersal of individuals among habitat patches for overall long-term stability. In Chapters 1 and 2, I used a landscape genetics approach to identify features that either promote or inhibit dispersal of American pikas. There has been significant debate over the past few years about the best statistical approaches within landscape genetics and the appropriateness of certain methods in particular. In Chapter 1, I used Crater Lake National Park as a test case to evaluate the effectiveness of Mantel and partial Mantel tests in a causal modeling framework to correctly identify underlying landscape variables (e.g., topography, aspect, water barriers) and their effect on gene flow. I demonstrated, through simulations, that this approach was able to correctly identify the landscape variables, but not precisely the magnitude of their effect on gene flow. I concluded that while results need to be interpreted with caution, this method was effective for identifying which variables are important in shaping functional connectivity. In Chapter 2, I applied the causal modeling approach from Chapter 1 to seven additional study sites. I identified a general trend that south-west facing aspects pose greater resistance to dispersal than north and east-facing aspects, suggesting that exposure to high temperatures may limit dispersal in American pikas. I also found that amount and configuration of habitat (i.e., rocky substrate) influenced the degree to which other landscape variables impact dispersal. I then applied the models of landscape resistance to investigate habitat patch connectivity within each site using a graph theoretic approach. This allowed me to assess overall patch connectivity, as well as identify specific patches and areas within study sites that are particularly important for maintaining functional connectivity, or in contrast, at risk of becoming isolated. Functional connectivity is an important component when assessing population vulnerability because it describes the ability of genetic material to flow through the landscape. A loss of gene flow can lead to isolation of population segments, reduction of effective population size, loss of genetic diversity and subsequent erosion of evolutionary potential, and ultimately metapopulation collapse. However, other factors including habitat area and quality, affect effective population size and population stability. Genetic diversity is the material upon which evolution acts. In the face of rapid environmental change, species must either shift their range in order to track their ecological niche, or adapt in situ to the new environment. The former strategy requires sufficient available habitat as well as dispersal ability, which is unlikely for American pikas. Genetic diversity, therefore, is an important indicator of long-term population viability. In Chapter 3, I quantified genetic diversity within thirteen study sites and investigated the relationship between genetic diversity and environmental variables related to climate as well as habitat configuration and quality. As expected, habitat area was important at both the local and regional scales. Temperature was also a significant predictor of genetic diversity, with hotter sites having lower genetic diversity. However, and somewhat surprisingly, precipitation was a better predictor of genetic diversity than temperature, with sites receiving moderate levels of precipitation having higher genetic diversity. I also compared population differentiation among study sites and identified sites that are particularly distinct. In the process of the above analyses, I also identified a previously undescribed contact zone between the northern and southern Rocky Mountain genetic lineages, recognized as separate subspecies, within Rocky Mountain National Park. Climate change is predicted to affect habitat quality and landscape permeability, both of which have detectable genetic consequences. In Chapter 4, I quantified genetic diversity and structure within two study sites, Yosemite and Lassen Volcanic National Parks, from two sampling periods separated by approximately a century in time. I extracted DNA from historic study skins housed in the University of California Berkeley Museum of Vertebrate Zoology (MVZ) and compared multilocus microsatellite genotypes to those generated from modern fecal and tissue samples. Additionally, the historic survey data, including specimens, field journals, and other artifacts, were used in a series of systematic resurveys and subsequent species distribution modelling by other researchers. The nature of national parks as protected areas affords the opportunity to isolate the effects of climate change to a greater degree than other areas that have also experienced significant land use changes. I found no evidence for a change in genetic diversity within these two sites, consistent with observations from other studies that occupancy has remained relatively stable. I did find some evidence suggesting increasing population differentiation, potentially as a result of eroding landscape permeability. These results provide an important baseline for comparison with other sites, as well as reference points for future genetic monitoring of these populations. In Chapter 5, I provide general synthesis and conclusions, as well as considerations for future research. Close